Guidelines for

Acute Leukemias
Vol X

Part A

Editors
Dr. Hari Menon MD, DM
Associate Professor
Department of Medical Oncology
Tata Memorial Centre
Dr. Brijesh Arora MD, DM
Associate Professor
Department of Medical Oncology
Tata Memorial Centre
Dr. Manju Sengar MD, DM
Assistant Professor
Department of Medical Oncology
Tata Memorial Centre
Dr. Akash Nahar, MD, DM, MPH
Assistant Professor
Department of Medical Oncology
Tata Memorial Centre
Dr. Tushar Vora
Published by
Tata Memorial Centre
Mumbai
Tata Memorial Hospital
Dr. Ernesh Borges Road, Parel
Mumbai 400 012. INDIA.
Tel.: +91-22-2417 7000
Fax: +91-22-2414 6937
Email: crs@tmc.gov.in
Website: http: //tmc.gov.in

Evidence Based Management of Cancers in India Vol. X

Three Parts
Set ISBN: 978-93-80251-07-3
Guidelines for Acute Leukemia
Part A ISBN: 978-93-80251-08-0
Guidelines for Bone and Soft Tissue Tumors
Part B ISBN: 978-93-80251-09-7
Guidelines for Colorectal Cancers
Part C ISBN: 978-93-80251-10-3

Set ISBN: 978-93-80251-07-3

Part A ISBN: 978-93-80251-08-0
Published by the Tata Memorial Hospital, Mumbai
Printed at the Sundaram Art Printing Press, Mumbai
© 2011 Tata Memorial Hospital, Mumbai
All rights reserved.
 Dedicated to
all our patients at
The Tata Memorial Hospital
 Contributers

Dr. Seema Gulia

Dr. A Nahar
Dr. Amol Dongre
Dr. B Arora
Dr. Maya Prasad
Dr. Vamashi
Dr. Nandish
Dr. Manju Sengar
Dr. Hari Menon
Dr. Saurabh
Dr. Naveen Khattry
Dr. Vikas
Dr. Akash Nahar
Dr. Randeep Singh
Dr. Amit Joshi
Dr. Ravi
Dr. Jayant
Dr. M Sengar
Dr. Bahusaheb
Dr. S D Banavali
 Contents

1. Algorithm for Laboratory Diagnosis 1

of Acute Leukemias
Part-1- ALL:
2. Approach to risk-stratification of Pediatric ALL 25
3. Management of standard Risk & 47
High risk ALL in children
4. CNS prophylaxis for Pediatric ALL 70
5. Evidence Based Management of 90
relapsed Pediatric ALL Including the
role of stem cell transplantation
6. Management of ALL in adults 111
7. Guidelines for stem cell transplant in 120
Adult ALL and management of relapsed ALL
Part-2- AML:
8. Molecular Cytogenetics and 146
Risk Stratification in AML
9. Optimizing induction therapy in AML 156
10. Optimizing post induction therapy in AML 167
11. Current state of HSCT and 177
Management of relapsed refractory AML
12. Pediatric AML: differences and similarities 186
13. Current management of 219
Elderly Patients with AML
 Preface

The Centre for Evidence Based Medicine (EBM) defines EBM

as “the conscientious, explicit and judicious use of current
best evidence in making decisions about the care of
individual patients”. EBM has percolated into all fields and
levels of medical practice and this has been particularly
exemplified in current oncology practice. There is an
increasing need to update our knowledge and be guided
by EBM, especially in an era where there have been rapid
developments and innovations in oncology.
Considerable progress has been made in diagnosis,
response prediction, prognostication and management
of acute leukemias. The new WHO classification of 2008
emphasizes the importance of molecular pathways and
impact of cytogenetics. Newer molecular techniques refine
it further and help individualize treatment for patients.
Prognostication has changed considerably based on this
new understanding. The role of tyrosine kinase inhibitors
in the treatment of Philadelphia positive ALL has come to
stay with the possibility of even avoiding transplants in
such patients. The concept of risk-adapted therapy has
gone a long way in allowing for minimizing toxicity and
bringing about optimum results in pediatric leukemias.
In the internet era, information overload can be as much
of a problem as paucity of information. The busy clinican
is frequently unable to separate real data from mere hype;
the ninth annual EBM meeting and the guidelines book
on acute leukemias is planned to do precisely this. As
always, in addition to collating the best available evidence,
the meeting and book also highlight areas where strong
evidence is lacking. Exciting new research is ongoing in
both myeloid and lymphatic leukemias with ongoing
efforts in targeted therapy. Controversies in management
can only be resolved with large multi centric studies. I
hope that in addition to updating practicing oncologists,
this book and meeting serves as a stimulus for investigators
to actively participate in clinical research and further
improve treatment outcomes.

C S Pramesh
Central Research Secretariat and
DAE Clinical Trials Centre
 Algorithmic approach to the
laboratory diagnosis of
Acute Leukemia
Galani KS, Gujral S

Acute Leukemia is a neoplasm of precursor (blastic)

hematopoietic cells (myeloid, B- and T-lymphoblasts)
involving bone marrow (BM) or other tissues like lymph
node, thymus with or without peripheral blood
involvement. It is a heterogenous disease clinically,
morphologically, immunophenotypically and genetically.
The modern era for classification of acute leukemia dates
to 1976 when one of the landmark publication was
published by a cooperative group of hematologists and
hematopathologists from France, America, and Britain and
was designated the French-American-British (FAB)
classification. It provided long needed standard
terminology for the acute leukemias and was quickly
accepted by most of the multi-institutional study groups.
The major advantage of the FAB lineage-based
classification system was its ease of use. The cytologic
criteria are well defined; they do not require high
technology and can be applied in most laboratories

1
throughout the world. The classification is also applicable
to the majority of cases of acute leukemia, and it partially
defines prognostic groups. The major disadvantage is its
modest clinical relevance; as it does not adequately define
biologic and treatment groups.
Other was the WHO classification (2001) which requires
not only morphology but also immuno-phenotyping (IPT),
cytogenetics, and molecular studies for proper
classification and prognostication. WHO 2008
classification stratifies neoplasms primarily according to
lineage. In addition, it has expanded the genetic disease
subtypes of acute leukemia, providing broader criteria for
the identification of the poor-prognosis AMLs with
myelodysplasia-related changes and introduced
cytogenetic subtypes of ALL. Also, it provides more
restrictive criteria for the diagnosis of mixed phenotype
acute leukemia (table 1). However its use is restricted to
only few cancer centers in India highlighting poor
applicability of WHO 2008 classification.
Table 1: Classification of acute leukemia by WHO
classification of tumors of hematopoietic and
lymphoid tissue (4th edition) 2008

I) Acute myeloid leukemia and related

precursor neoplasms
AML with recurrent genetic abnormalities
AML with t(8;21)(q22;q22)
AML with inv(16)(p13;q22) or t(16;16)(p13;q22)
Acute Promyelocytic Leukemia with
t(15;17)(q22;q21)
AML with t(9;11) (p22;q23)
AML with t(6;9)(p23;q34)

2
 AML with inv(3)(q21;q26.2)
AML (megakaryoblastic) with t(1;22)(p13;q13)
Provisional entity: AML with mutated NPM1 and
AML with mutated CEBPA
AML with myelodysplasia-related changes
Therapy-related myeloid neoplasms
AML, NOS
AML with minimal differentiation (FAB M0)
AML without maturation (FAB M1)
AML with maturation (FAB M2)
Acute myelomonocytic leukemia (FAB M4)
Acute monblastic and monocytic leukemia (FAB M5)
Acute erythroid leukemia (FAB M6)
Acute megakaryocytic leukemia
Acute basophilic leukemia
Acute panmyelosis with myelofibrosis
Myeloid sarcoma
Myeloid proliferations related to Down Syndrome
Transient abnormal myelopoiesis
Myeloid leukemia associated with Down syndrome
Blastic plasmacytoid dendritic cell neoplasm
II) Precursor lymphoid neoplasms
B-lymphoblastic leukemia/lymphoma (B-LL), NOS
B-LL with recurrent genetic abnormalities
B-LL with t(9;22)(q34;q11.2)
B-LL with t(v;11q23); MLL rearranged
B-LL with t(12;21)(p13;q22)
B-LL with hyperdiploidy
B-LL with hypodiploidy (Hypodiploid ALL)
B-LL with t(5;14)(q31;q32)
3
 B-LL with t(1;19)(q23;p13.3)
T- lymphoblastic leukemia/lymphoma (T-LL)

III) AMBIGUOUS LINEAGE

Acute leukemias of ambiguous lineage
Acute undifferentiated leukemia
Mixed phenotype acute leukemia with
t(9;22)9q34;q11.2)
Mixed phenotype acute leukemia with t(v;11q23);
MLL rearranged
Mixed phenotype acute leukemia, B/myeloid, NOS
Mixed phenotype acute leukemia, T/myeloid, NOS
Mixed phenotype acute leukemia, NOS-rare types
Natural killer-cell lymphoblastic leukemia/
lymphoma

Evaluation of acute leukemia patients

Evaluation starts with a detailed history and clinical
examination which includes lymphdenopathy,
organomegaly (liver/spleen) and testicular enlargement.
Patients commonly present with symptoms including
weakness, fatigue, fever, bleeding, bone pains either
gradually or abruptly. In addition, history of prior cytotoxic
therapy and presence of Down syndrome would help
classify a patient in an appropriate category.
Laboratory work up starts with complete blood counts
(CBC) and peripheral blood evaluation. CBC generally
shows anemia and thrombocytopenia. However patients
may have leucopenia, normal leukocyte count or
leukocytosis. Peripheral blood examination is usually
followed by BM examination. BM aspiration is preferably
done from the posterior superior iliac bone and sample is

4
aliquotted for morphology, flow cytometry, molecular
studies (EDTA vacutainer), cytogenetics (Heparin
vacutainer) and trephine biopsy (formalin fixed). We follow
Hammersmith’s protocol for BM biopsy processing which
uses acetic acid-zinc-formalin as the fixative and provides
optimal preservation of both the morphology and the
antigens and nucleic acids for immunohistochemistry
(IHC).
Coagulation studies like PT, aPTT, fibrinogen and D-dimer
are also important to monitor patients for disseminated
intravascular coagulation (DIC), mainly in acute
promyelocytic leukemia. In addition, serum LDH, uric acid,
serum electrolytes and renal and liver function tests are
carried out.

Role of morphology in the diagnosis of

acute leukemia
Staining of peripheral blood or BM aspirate: Meticulous
examination of a well stained smear is the cornerstone
for the diagnosis of a hematolymphoid neoplasm. Various
Romanowsky stains commonly used in the laboratory
include Wright’s stain, Leishman’s stain, Giemsa stain,
Wright-Giemsa stain, Jenner-Giemsa, May-Grunwald-
Giemsa stain,etc.
Morphologic examination: Screening of peripheral blood
smear for the blasts is important and presence of
circulating blasts suggest BM examination for confirmation
and further subtyping.
Percentage of blasts: Presence of at least 20% blasts in
BM or in blood makes the diagnosis of acute leukemia. In
rare cases, the blast count is below 20%, but cytogenetic
abnormalities are present that by convention warrant a

5
diagnosis of AML (e.g. t(8;21), t(15;17) and other recurrent
genetic abnormalities). If the blasts percentage is less,
trephine biopsy evaluation is necessary to detect the cluster
of blasts alongwith CD34 expression.
Morphologic evaluation of blasts and blast equivalent cells
(mainly promeylocytes or promonocytes): Presence of Auer
rod aids in the diagnosis of myeloid lineage. Morphological
evaluation is important in the monocytic proliferations,
mainly in distinction of promonocytes from more mature
monocytes as the diagnosis of acute monocytic leukemia
(AML M5b) versus chronic or juvenile myelomonocytic
leukemia (CMML/JMML) depends on the percentage of
these cells. Other morphologic features of importance
include the bilobed nuclei with or without fine granules
which suggest acute promyelocytic leukemia (APML),
which should result in a clinical work-up for DIC as well as
rapid cytogenetic studies by fluorescent in-situ
hybridization (FISH) or molecular confirmation of t(15;17)
and variant translocations.
Many chromosomal abnormalities have been associated
primarily with morphological or immunophenotypic
features. Blast morphology (Table 2), immunophenotype
and even non-blast morphologic features like increase in
eosinophils and Eo-basophils are of importance in selecting
specific cytogenetic panel or probes for FISH analysis., e.g.,
AML M2 with eosinophilia (associated with expression of
lymphoid marker like CD19 detected by flow cytometry or
PAX5 by IHC) correlates with frequent presence of t(8;21)
while AML M4 with eosinophilia/Eo-basophilia correlates
with presence of inv (16) and t (16;16). Presence of
multilineage dysplasia warrants a diagnosis of AML with
myelodysplasia-related changes (Table 3, 4) and requires

6
conventional karyotyping by G banding. Presence of
nuclear cupping suggests myeloid lineage and correlates
with NPM1 or FLT3 mutations.
Table 2: Usual cytological features of AML and ALL
Parameter AML ALL
Blast size Large and uniform Small to medium;
variable
Chromatin Finely dispersed Rather coarse
Nucleoli 1 to 4, often Absent or 1 or 2;
prominent indistinct
Cytoplasm Granules often Granules lacking
present in nearly all cases
Auer rods 60–70% of cases Absent
Myelodysplasia Often present Absent

Role of trephine biopsy

It is especially required in cases wherein the BM aspirate
smears are acellular or dry tap, diluted with peripheral
blood, in partially treated cases where blood transfusion/
treatment by steroids and other drugs alter the
morphological picture or in cases where the sample is
degenerated during transport to a referral laboratory for
ancillary investigations. It is recommended to obtain a
trephine biopsy upfront in all new cases of
hematolymphoid malignancies as the paraffin block is an
invaluable diagnostic archival material for
immunophenotyping. This is especially important in our
country where many of these patients are empirically
treated with blood transfusions or steroids before they
are referred to a tertiary hemato-oncology center for
management. Thus IHC can be performed on paraffin

7
blocks to broadly classify these neoplasms into AML or B-
/T- ALL. Also, expression of certain antigens like Tdt can
be well demonstrated on IHC.
Table 3: Criteria for the diagnosis of AML with
myelodysplasia-related features
>= 20% blood or marrow blasts
AND
Any one of the following
 Previous history of myelodysplastic syndrome
 Myelodysplastic syndrome-related cytogenetic
abnormality
 Multilineage dysplasia
AND
Absence of both
 Prior cytotoxic therapy for an unrelated disease
 Recurring cytogentic abnormality as described in
AML with recurrent genetic abnormalities

Table 4: Cytogenetic abnormalities sufficient to

diagnose AML with myelodysplasia-related features
when >= 20% blood or marrow blasts are present

Complex karyotype*
Unbalanced abnormalities
 -7/del(7q)
 -5/del(5q)
 i(17q)/t(17p)
 -13/del(13q)
 del(11q)
 del(12p)/t(12p)
 del(9q)
 idic(X)(q13)

8
Balanced abnormalities
 t(11;16)(q23;p13.3)**

 t(3;21)(q26.2;q22.1)**
 t(1;3)(p26.3;q21.1)
 t(2;11)(p21;q23)
 t(5;12)(q33;p12)
 t(5;7)(q33;q11.2)
 t(5;17)(q33;q13)
 t(5;10)(q33;q21)
 t(3;5)(q25;q34)
*>- 3 unrelated abnormalities, none of which are included in
the AML with recurrent genetic abnormalities subgroup;

Role of cytochemistry in acute leukemia

Though immunophenotyping by flow cytometry is
mandatory, the cytochemical myeloperoxidase (MPO) has
a role in assigning the myeloid lineage even in latest WHO
2008 classification. MPO positivity in > 3% of blasts is
considered positive. Lymphoid blasts are known to be MPO
or SBB negative. Among the myeloid leukemia, AML-M0
blasts, erythroid blasts (AML-M6), megakaryoblasts (AML-
M7) and even monoblasts (AML-M5) are MPO negative.
Thus, absence of cytochemical MPO does not exclude a
myeloid lineage and here is the role of cytoplasmic MPO
either by flow cytometry or IHC.
Sudan Black B (SBB) staining parallels MPO but is less
specific and even occasional cases (<5%) of lymphoblastic
leukemia exhibit SBB positivity. Hence for lineage
specification MPO positivity either by cytochemisty or
immunophenotyping is important.

9
Non specific esterases (NSE), alpha naphthyl butyrate (ANB)
and alpha naphthyl acetate (ANA), show diffuse
cytoplasmic activity in monoblasts and monocytes (table
5). Lymphoblasts (T-LL) may have focal punctate activity
with NSE but neutrophils are usually negative.
Megakaryoblasts and erythroid blasts may have some
multifocal, punctuate ANA positivity, but it is partially
resistant to natrium fluoride (NaF) inhibition whereas
monocyte NSE is totally inhibited by NaF. Megakaryocytes
are positive internal control for NSE stain.
Table 5: Cytochemical profiles in acute leukemia
Myeloperoxidase and Nonspecific esterase
Sudan Black B
AML + + (monocytic,diffuse)
ALL - +/-
+ positive, - negative. +/- not definitive

Role of Immunophenotyping in Acute

leukemia by FCM
Multiparameter FCM (atleast 3 color, preferably 4 or more
colors) is the preferred method for immunophenotyping
(IPT) of AL due to the ability to analyze high numbers of
cells in a relatively short period of time with simultaneous
recording of information about several antigens for each
individual cell. This permits precise characterization of
leukemic cells even when they are present in low numbers.
It plays important role in diagnosis, prognosis and
monitoring the treatment as minimal residual detection
(MRD).
Leukemia cells express the lineage associated markers,
lineage specific markers and leukemia associated antigens

10
which are aberrantly expressed as unexpected antigens
(Table 6). Multicolor FCM helps in demonstration of all
such aberrant expressions.
Table 6: Commonly used immunophenotypic markers
Lineage Markers
Myeloid CD13, CD33, CD117, CD15, CD16,
Or MPO
Monocytic At least 2 of the following:
NSE, CD14, CD64, CD11c, lysozyme
B-lymphoid CD19, CD20, CD22,
cytoplasmicCD79a,
cytoplasmic CD22
T-lymphoid CD1a, CD2, CD3, CD4, CD5, CD7,
CD8, cytoplasmic CD3
Others markers CD34, Tdt, HLADR, CD41, CD61,
CD56, CD38, CD138

Various factors influence panel selection for leukemia. It

may depend upon type of laboratory and cost
effectiveness, laboratory throughput, technical expertise
available, number of antibodies used and other factors
like indication for which IPT is being done. E.g., initial
diagnosis, subtyping, follow-up studies and detection of
minimal residual disease; or the type of flow instrument,
for example, three or more colors, etc. For the optimum
results, FCM analysis requires a strict quality control with
standardization methodology, panel of antibodies and
their combinations. Of these, antibody selection is the most
critical step to get accurate diagnosis and even during
monitoring of disease.

11
Indian experience in IPT of
hematolymphoid neoplasms:
It has been observed that few laboratories in the past had
been resorting to cost cutting excercises in the form of
short, sketchy and inadequate reagent/panel of antibodies
for IPT. A group of Indian Cytometrists in 2008 highlighted
these issues by establishing the Indian ‘Guidelines for
immunophenotyping of hematolymphoid neoplasms by
flow cytometry’ to bring about uniformity and
comparibility in reporting of leukemia and lymphoma.
A small group of 6 clinical cytometry laboratory from
Mumbai started an inter laboratory comparison program
(ILCP), a kind of proficiency testing program, which gave
the realistic input about the variations in laboratory
practices starting from sample collection, processing,
analysis of data upto final dispatch of report. An initial
consensus on panel selection was made by this group.
This document was circulated to national and international
experts in hematopathologist, hematologists, medical
oncologists and cytometrists for their suggestions and
modifications. A practice-based questionnaire including
reagent and antibodies was circulated to all participants
and that was discussed in guideline meeting held in March
2008 at Tata Memorial Hospital (TMH). Thus an approach
of minimal screening panel followed by directed additional
markers was proposed. However it was further
recommended that laboratories must refrain using lesser
reagents and instead do a more comprehensive panel. It
also emphasized that laboratories must ensure those
adequate cells/sample are/is available for additional
markers to avoid re-sampling (CD45 gating was important
in all the cases of AL especially when the blast count was

12
less than 50%). These guidelines were later published in
the Indian Journal of Pathology and Microbiology.
With the advances in the field of FCM (6, 8 or 10 color
IPT) the selection of various antibodies combination,
fluorochrome conjugate, gating strategies, software
selection and analysis are becoming more and more
complex. Most laboratories all over do there to four color
IPT, however a few laboratories in India have shifted to six
color IPT and are in the process of standardization. Few
of these laboratories have started MRD evaluation also.

Panel for Acute leukemia

Cases which are unequivocally myeloid, for example,
containing Auer rods, and/or positive for cytochemical
stains like myeloperoxidase (MPO), may not be processed
for FCM (unless a protocol based treatment is planned).
All other subtypes of AL are diagnosed by FCM (Table 7)
with the suggested minimal screening panel and
cytogenetics (as and when required). In those cases where
a diagnosis is not clearly established, additional markers
should follow. In cases of aberrant expression of two or
more antigens, a biphenotypic acute leukemia should be
ruled out using lineage-specific cytoplasmic markers.
However it is strongly recommended that cytoplasmic
stains are done upfront, so as not to miss rare subtypes.
Panels are decided after doing a meticulous morphological
examination of the peripheral blood/bone marrow/fluid
smears. Recommended minimal screening panels for
immunophenotyping of acute leukemia include CD45,
CD10, CD19, CD7, CD3, CD5, CD13, CD33, CD117, CD34,
HLA-DR. The selection of antibodies in the additional
markers may be based on the results of the minimal panel.

13
Panel recommended following antibodies in the additional
panel: cCD3, cCD22/cCD79a, anti-MPO (lineage-specific
cytoplasmic markers for T, B and myeloid series cells), SIg
(K/’»), Tdt, CD41, CD61, CD56, CD38, CD138, CD64, CD2,
CD4, CD8.
Table 7: Approach to diagnosis of acute leukemia
with minimal panels.

A cu te Le u ke m ia (Blast s > =2 0% )

A u er ro d se en A u er r o d n o t see n

AM L M PO an d /o r NSE + M P O a n d/o r N SE n e ga tive

FC M

First lin e p an el
(C D 34 an d Td t)

C D 1 0 ,C D 1 9 ,H L A-D R C D 3 ,C D 4 ,C D 8 C D 1 3 , C D 3 3, C D 1 17

If lin e age is n o t e stab lish e d ,

Sec o n d lin e pa n el*

cC D 2 2 / c C D 79 a/ C c d 3 /A nt i-M P O /C D 4 1/C D 61

Fo llo w ed b y C yto ge ne tic s an d m o le cu la r a n alysis

*Though above recommendations were made in 2008,

however, laboratories are encouraged to do a more
comprehensive and an elaborate panel as a primary panel.
It is further recommended that cytoplasmic markers be
included in the primary panel itself.

If a case is morphologically suggestive of Burkitt’s

lymphoma, surface immunoglobulin (k/»), IgM and Tdt
may be added to the primary panel. In cases of pleural
effusion or mediastinal mass, cCD3, Tdt, CD4 and CD8
may be added to the primary panel. Tdt is an important
marker for all cases of T cell neoplasms. Most of T-ALLs

14
are negative for surface CD3 and instead express
cytoplasmic CD3. We have been able to diagnose more
cases of hepatosplenic gamma delta T cell lymphoma after
increasing the immunophenotypic panel.
Acute biphenotypic leukemia has been defined by the
Immunological Characterization of Leukemias (EGIL) group
and has been recommended by the WHO. Another criteria
for work-up of these neoplasms has been defined by
Campana et al. The latter define true mixed-lineage
leukemia giving more credence to cytoplasmic expression
of highly lineage-specific markers (e.g., cCD22/cCD79a,
cCD3 and anti-MPO). Both these criteria have their own
limitations. The laboratories can use any of the above two
criteria for the diagnosis of biphenotypic acute leukemia.
WHO 2008 has come out with similar recommendation.

Existing 6 color panel for acute leukemia

We are in the process of standardizing six colour IPT. For
acute leukemia, we use following 6 tubes and the
antibodies included are:
Tube 1: (B-cell) CD10/ CD19/ CD20/ CD34/ CD38/ CD45
Tube 2: (Monocytic) CD33/ CD64/ CD14/ HLA-DR/
CD71/ CD45
Tube 3: (T-cell) CD3/ CD4/ CD8/ CD7/ CD56/ CD45
Tube 4: (Myeloid) CD13/ CD117/ CD15/ CD16/ CD11b/
CD45
Tube 5: (Cytoplasmic) cCD3/ cCD79a/ anti-MPO/ CD45
Tube 6: (Cytoplasmic) cCD3/ cCD22/ anti-Tdt/ CD45

15
Role of Cytogenetics in Acute leukemia
Genetic changes associated with both AML and ALL are
most important in prognostication of disease, to decide
the plan of treatment and also to monitor the treatment
response as minimal residual disease. The best practice is
to do complete work up as per WHO 2008.
The prognostication of AML is based on the cytogenetic
abnormalities and thus needs Fluorescent-in-situ-
hybridisation ( FISH) evaluation for the predicted genetic
abnormalities. The AML with myelodysplasia related
changes carries poor prognosis and needs bone marrow
transplantation as the treatment of choice. The MDS
related cytogenetics comprises complex karyotype,
unbalanced abnormalities or balanced translocations and
conventional karyotype by G banding is superior to define
such multiple abnormalities.
Among the B-LL with recurrent genetic abnormalities,
t(9;22) has worst prognosis and needs therapy with
Imatinib for an early event-free survival. B-LL with CD10
negativity, particularly age less than 1 year of age correlates
with MLL rearrangement. MLL gene on chromosome
11q23 has many fusion partners and thus it is done with
MLL break apart probe. Diploidy (hyperdiploidy >50 or
hypodyploidy <45) status is done by conventional
karyotype by G banding and both have prognostic value
(favourable or poor, respectively).
Prognosis in T-LL is not as dependent on cytogenetics as
in AML and B-LL, though the prognosis is better in T-LL as
compared to B-LL due to the lower incidence of cytogenetic
abnormalities in T-LL.

16
 Table 9: Prognostic groups in AML based on
cytogenetics
Prognostic group Chromosome finding
Favorable Inv(16) or t(16;16)(p13;q22)
t(8;21)(q22;q22)
t(15;17)
Intermediate t(6;9),
Normal karyotype
Unfavorable del 5q, del7q
Complex abnormalities

Table 10: Prognostic group of ALL

based on cytogenetics
Prognostic group Chromosome finding
Favorable Hyperdiploidy >50
Cryptic t(12;21)(p12;q22)
Intermediate Hyperdiploidy 47–50
Normal (diploidy)
del(6q)
Unfavorable Hypodiploidy: near haploid
Near tetraploid
del 17p
t(9;22)(q34;q11)
t(11q23), usually t(4;11)

Role of molecular analysis in acute

leukemia:
The role of molecular diagnosis is adjuvant to cytogentics
in acute leukemia work up, but it is a very powerful tool
for the identification of minimal residual disease (MRD)

17
and early relapse. Molecular analysis may be used to
establish clonality or to identify molecular translocations
producing fusion gene products, especially in cryptic
translocation. The molecular studies are best done for the
known or expected fusion transcripts or mutations.
It is important to perform molecular analysis when the
presence of a fusion gene that would impact treatment is
expected and also in the monitoring of MRD status,
especially in cases of Philadelphia positive B-LL, APML, etc.
The most important role of molecular analysis is in AML
cases especially AML having ‘normal karyotype’. Mutations
of NPM1, FLT3 (FLT3-ITD, FLT3-TKD) and CEBPA have been
reported in patients with AML with a normal karyotype
where they have a prognostic significance. In case of
normal or intermediate-risk karyotype AML, CEBPA and
NPM mutations in the absence of FLT3 mutation appear
to confer a good prognosis.
The above investigations require a standardized protocol
and advance laboratory facilities which are possible in few
tertiary cancer referral centers or referral laboratories. The
newer applications in this field are fascinating and fruitful
in prognostication and monitoring of disease.

Hematolymphoid neoplasms and Indian

experience
Various centers in India have published data in
hematolymphoid neoplasms. However, there is no data
of true incidence of AL from India. Most of the paediatric
patients are brought to the tertiary care centers for
treatment while many of the old age patients may be
managed locally. Our hospital is a tertiary care cancer
referral center where common neoplasms include head &

18
neck, breast & female genitor-urinary tract tumors.
Hematolymphoid neoplasms constitute approximately 8%
of all neoplasms presented at TMH. Acute Leukemia
constitutes approximately 3% of all new cases.
We reviewed 2520 consecutive cases of AL between 2003
– 06 and the morphology, cytochemistry,
immunophenotyping, FISH, and molecular tests data were
correlated as per WHO guidelines 2001. It included 1,471
cases (58%) of acute lymphoblastic leukemia, 964 cases
(38%) of acute myeloid leukemia, 45 cases (1.8%) of
chronic myelogenous leukemia in blast crisis, 37 cases
(1.5%) of biphenotypic acute leukemia, 1 case of
Triphenotypic AL, and 2 cases of acute undifferentiated
leukemia. Common subtypes of ALL were B-cell ALL (76%),
which comprised of intermediate stage/CALLA positive
(73%), early precursor/proBALL (3%). T-cell ALL constituted
24% (351 cases) of ALL. Common subtypes of AML
included AML-M2 (27%), AML-M5 (15%), AML-M0 (12%),
AML-M1 (12%), APML (11%), and AML t(8;21) (9%). CML-
BC was commonly of myeloid blast crisis subtype (40
cases).
B-cell ALL was the commonest subtype in children and
AML in adults. Overall incidence of AML in adults was
low (53% only). The incidence has a definite referral bias
as many of the older patients might not be brought to
the hospital. CD13 was most sensitive and CD117 most
specific for determining myeloid lineage. A minimal
primary panel of nine antibodies consisting of three
myeloid markers (CD13, CD33, and CD117), B-cell
lymphoid marker (CD19), T-cell marker (CD7), with CD45,
CD10, CD34, and HLADR could assign lineage to 92% of
AL. Cytogenetics findings lead to a change in the

19
diagnostic subtype of myeloid malignancy in 38 (1.5%)
cases.

Conclusions
Role of a diagnostic laboratory includes establishing
diagnosis, prognostication (sub-classification) of the
hematolymphoid neoplasm, prediction and evaluation of
treatment effectiveness (minimal residual disease). Many
new molecularly defined subgroups have been
incorporated in the recent WHO 2008 classification.
Though it is difficult to fully adopt the WHO classification
in India as only few centers possess facilities for
immunophenotyping and cytogenetic/molecular genetic
analyses, it is necessary to identify the prognostic subgroup
of the patient in case the patient is to be treated on a
protocol basis. Only tertiary care centers try to apply WHO
2008 classification, however most of the remaining centers
still follow FAB classification. It is mandatory to perform
IPT for all the AL cases prior to initiating appropriate
therapy as IPT provides both diagnostic and prognostic
information. Furthermore, cytogenetics and molecular
analysis could be applied for risk stratification.
Most laboratories in India do a 3-4 color IPT and recently
few have started doing 6 color IPT. With the advancements
in multicolor flow cytometry (Euroflow- 8 colour and Brent
Wood et al- 10 colour) in the west, Indian scenario is also
changing. These higher end fluororchromes add to the
cost of testing but are claimed to add invaluable
information to the disease. Cost benefit ratio has to be
studied. More number of colors add to more
troubleshoots. The laboratories are encouraged to
participate in external quality assurance programs/
proficiency testing for IPT and molecular diagnosis.

20
Further reading
1. World Health Organization classification of tumors.
In: Jaffe ES, Harris NL, Stein H, Vardiman JW, editors.
Pathology and genetics of tumors of haematopoietic
and lymphoid tissues. Lyons, France: IARC Press;
2008. p. 75-118.
2. Gujral S. Hematolymphoid neoplasms: World Health
Organization versus rest of the world. Leukemia.
2009 May;23(5):978
3. Naresh KN, Lampert I, Hasserjian R, Lykidis, Elderfield
K, Horncastle D, Smith N, Murray-Brown W, Stamp
GW. Optimal processing of bone marrow trephine
biopsy: the Hammersmith Protocol. J Clin Pathol
2006;59:903-11. 4. Gujral S, Subramanian PG, Patkar
N, Badrinath Y, Kumar A, Tembhare P, Vazifdar A,
Khodaiji S, Madkaikar M, Ghosh K, Yargop M,
Dasgupta A. Report of proceedings of the national
meeting on “Guidelines for Immunophenotyping of
Hematolymphoid Neoplasms by Flow Cytometry”.
Indian J Pathol Microbiol 2008;51:161-6.
5. Gujral, S., Badrinath, Y., Kumar, A., Subramanian, P.
G., Raje, G., Jain, H., Pais, A., Kadam, P. S. A., Banavali,
S. D., Arora, B., Kumar, P., Menon, V. G. H., Kurkure,
P. A., Parikh, P. M., Mahadik, S., Chogule, A. B.,
Shinde, S. C. and Nair, C. N. (2009),
Immunophenotypic profile of acute leukemia: Critical
analysis and insights gained at a tertiary care center
in India. Cytometry Part B: Clinical Cytometry, 76B:
199–205.
6. Ghosh S, Shinde SC, Kumaran GS, Sapre RS, Dhond
SR, Badrinath Y, Ansari R, Kumar A, Mahadik S,
Chougule AB, Nair CN. Haematologic and
immunophenotypic profile of acute myeloid

21
 leukemia: An experience of Tata memorial hospital.
Indian J Cancer 2003; 40: 71–76.
7. Gujral S, Polampalli S, Badrinath Y, Kumar A,
Subramanian PG, Raje G, Amare P, Arora B, Banavali
SD, Nair CN. Clinico-hematological profile in
biphenotypic acute leukemia. Indian J Cancer. 2009
Apr-Jun;46(2):160-8
8. Clinical and laboratory standards institute. Clinical
flow cytometric analysis of neoplastic
hematolymphoid cells: Approved guideline - 2 nd
ed. CLSI document H43-A2 [ISBN 1-56238-000-0].
Wayne, Pennsylvania: Clinical and Laboratory
Standards Institute; 2006. p. 19087-1898.
9. Stewart CC, Behm FG, Carey JL, Cornbleet J, Duque
RE, Hudnall SD, et al. US-Canadian consensus
recommendations on the immunophenotypic
analysis of hematologic neoplasia by flow cytometry:
Selection of antibody combinations. Cytometry
1997;30:231-5.
10. Wood BL, Arroz M, Barnett D, DiGiuseppe J, Greig B,
Kussick SJ, et al. 2006 Bethesda international
consensus recommendations on the
immunophenotypic analysis of hematolymphoid
neoplasia by flow cytometry: Optimal reagents and
reporting for the flow cytometric diagnosis of
hematopoietic neoplasia. Cytometry 2007;72:
S14-22.
11. Bene MC, Castoldi G, Knapp W, Ludwig WD, Matutes
E, Orfao A, et al. Proposals for the immunological
classification of acute leukemias: European Group
for the Immunological characterization of Leukemias
(EGIL). Leukemia 1995;9:1783-6.
12. Gujral S, Subramanian PG, Patkar N, Badrinath Y,
Kumar A, Tembhare P, Vazifdar A, Khodaiji S,
22
 Madkaikar M, Ghosh K, Yargop M, Dasgupta A.
Report of proceedings of the national meeting on
“Guidelines for Immunophenotyping of
Hematolymphoid Neoplasms by Flow Cytometry”.
Indian J Pathol Microbiol. 2008 Apr-Jun;51(2):
161-6
13 Gujral S, Subramanian PG, Dasgupta A. Diversity
amongst various guidelines for immunophenotyping.
Cytometry A. 2009 Jul;75(7):560-1
14. Campana D, Behm FG. Immunophenotyping of
leukemia. J Immunol Met 2000;243:59-75.
15. Gujral S, Dongre K, Bhindare S, Subramanian PG,
Narayan H, Mahajan A, Batura R, Hingnekar C,
Chabbria M, Nair CN. Activity-based costing
methodology as tool for costing in hematopathology
laboratory. Indian J Pathol Microbiol. 2010 Jan-
Mar;53(1):68-74
16. Gujral S, Tembhare P, Badrinath Y, Subramanian PG,
Kumar A, Sehgal K. Intracytoplasmic antigen study
by flow cytometry in hematolymphoid neoplasm.
Indian J Pathol Microbiol. 2009 Apr Jun; 52 (2) :135-
44. Review
17. Hasset J, Parker J. Laboratory practices in reporting
flow cytometery phenotyping results for leukemia/
lemphoma specimens: Results of a survey. Cytometry
1995;22:264-81.
18. Braylan RC, Orfao A, Borowitz MJ, Davis BH. Optimal
number of reagents required to evaluate
hematolymphoid neoplasias: Results of an
international consensus meeting. Cytometry
2001;46:23-7.
19. Bain BJ, Barnett D, Linch D, Matutes E, Reilly JT.
General haematology task force of the british

23
 committee for standards in haematology (BCSH),
British Society of Haematology. Revised guideline on
immunophenotyping in acute leukemia and chronic
lymphoproliferative disorders. Clin Lab Hematol
2002;24:1-13.
20. Ruiz-Arguelles A, Rivadeneyra-Espinoza L, Duque RE,
Orfao A. Report on the second Latin American
consensus conference for flow cytometric
immunophenotyping of hematological malignancies.
Cytometry 2005;70:39-44.
21. Davis BH, Holden JT, Bene MC, Borowitz MJ, Braylan
RC, Cornfield D, et al. 2006 Bethesda International
Consensus Recommendations on the flow cytometric
immunophenotypic analysis of hematolymphoid
neoplasms: Medical Indications. Cytometry
2007;72:S5-13
22. Almeida J, Vidriales B, López-Berges MC, Garc½a-
Marcos MA, Moro MJ, Corrales A, et al. Incidence of
phenotypic aberrations in a series of 467 patients
with B chronic lymphoproliferative disorders: Basis
for the design of specific four-color stainings to be
used for minimal residual disease investigation.
Leukemia 2002;16:1460-9

24
 Approach to risk-stratification of
Pediatric ALL
Dr. Seema Gulia, Dr. Brijesh Arora

There has been significant improvement in the outcome

of children with acute lymphoblastic leukemia (ALL). More
than 95% clinical remission rates and up to 80% cure
rates have been achieved with current treatment
regimens1-3. This success has been gained through the
implementation of risk-stratified therapy and stepwise
improvement using cooperative group protocols.
ALL is a heterogeneous disease comprised of
morphologically identical leukemias arising from different
biological mechanisms. Children with ALL are treated
according to the risk groups defined by clinical features
(age, sex), laboratory features (genetic abnormalities in
leukemia cells) and response to treatment. The aim of risk
stratification is to minimize the risk of relapse by identifying
subsets of patients requiring more intensive therapy, and
to minimize the risk of toxicity for those patients likely to
be cured with modest therapies. Risk adapted therapy
tailors treatment based on the predicted risk of relapse.
However, for the 15-20% of children with newly diagnosed
ALL who relapse, traditional risk assessment remains

25
inadequate. An increasing understanding of genetic
abnormalities, patterns of gene expression, and host
pharmacogenomics supplant currently applied prognostic
criteria for use in treatment planning for childhood ALL
and may further help enhance the outcome & decrease
toxicity.
Risk-based treatment assignment requires the
identification of prognostic factors that reliably predict
outcome. For children with ALL, the prognostic factors
are grouped into the following categories:
1. The host genotype ( host biology)
2. The leukemic cell genotype (tumor biology)
3. Response to therapy

1. Host biology
a) Age - The age at diagnosis correlates with clinical
outcome. In childhood ALL, infants and adolescents have
a worse prognosis than patients aged 1-10 years4. The
improved outcome in patients between 1-10 years is due
to more frequent occurrence of favorable cytogenetic
features in the leukemic blasts including hyperdiploidy, or
translocation t (12; 21) 5 (Level of evidence 2C) . Infants
with ALL have high risk of treatment failure as they have
high presenting leukocyte counts, increased frequency of
central nervous system leukemia at presentation and a
very high incidence (~80%) of rearrangement of the MLL
gene on chromosome 11q23. Amongst infants with MLL
gene rearrangements, those presenting at a young age
(< 6 months) or with extremely high leukocyte counts (>
300, 000/ml) have the worst prognosis 6 (Level of
evidence 2A). Adolescents (ages 16-21 years) with ALL
have a less favorable outcome than children aged 1-10
years, as they frequently present with T-cell

26
immunophenotype, high leukocyte counts, and a higher
incidence of the Philadelphia chromosome [t(9;22)]7.
Adolescents are also at higher risk for certain treatment-
related complications, such as osteonecrosis, pancreatitis
and deep vein thromboses, which may also impact
prognosis8.
b) Gender -The prognosis for boys with ALL is slightly worse
than girls9. Potential reason for the better prognosis for
girls is the occurrence of testicular relapses among boys.
Also, boys appear to be at increased risk of bone marrow
and CNS relapse for reasons that are not well understood.
With current treatment regimens, there is no difference
in outcome between males and females10.
2. Tumor biology
A) White blood cell (WBC) count at diagnosis- WBC count
reflects tumor burden. Although the relationship between
WBC count and prognosis is a continuous rather than a
step function, the National Cancer Institute (NCI) stratifies
patients into two subsets based on WBC counts; standard
risk (WBC count < 50, 000) or high risk (WBC count >
50, 000) (Level of evidence 2C). High WBC count is
usually associated with unfavorable chromosomal
translocations such as t (4; 11), t (9; 22) and T cell
immunophenotype. 11
B) CNS status at diagnosis - The presence of CNS disease
at diagnosis is an adverse prognostic factor inspite of
intensification of therapy. Patients are divided into three
categories based on the number of WBC/μL and the
presence/absence of blasts on cytospin:
CNS1: Cerebrospinal fluid (CSF) that is cytospin negative
for blasts regardless of WBC count.
CNS2: CSF with fewer than five WBC/μL and cytospin
positive for blasts.

27
CNS3 (CNS disease): CSF with five or more WBC/μL and
cytospin positive for blasts, or cranial nerve palsy.
Children with ALL who present with CNS disease (i.e.,
CNS3) at diagnosis are at a higher risk of treatment failure
(both within the CNS and systemically) compared to those
who are CNS negative4 (Level of evidence 2C). The
adverse prognostic significance associated with CNS2
status can be overcome by the application of more
intensive intrathecal therapy, especially during the
induction phase12. Furthermore, a traumatic lumbar
puncture (more than 10 erythrocytes/μL) that includes
blasts at diagnosis appears to be associated with increased
risk of CNS relapse and requires intensification of therapy.

C) Testicular Involvement at diagnosis:

Overt testicular involvement at the time of diagnosis occurs
in approximately 2% of males. Historically, testicular
involvement at diagnosis was identified as an adverse
prognostic factor, but with aggressive therapy it has lost
its prognostic significance 13.
D) Immunophenotype -The World Health Organization
(WHO) classifies ALL as either “B-lymphoblastic leukemia”
or “T-lineage lymphoblastic leukemia(T-ALL)” based on its
cell of origin detected by surface or cytoplasmic expression
of B or T cell antigens. Precursor B-cell ALL (B-ALL) is defined
by the expression of cytoplasmic CD79a, CD19, HLA-DR,
and other B cell-associated antigens. It accounts for 80%
to 85% of childhood ALL and has a better prognosis
compared to T-ALL. Precursor B-cell ALL patients are further
divided into immunologic subtypes, of which Pro-B ALL
(CD10 negative and no surface or Cytoplasmic Ig) is
commonly seen in young infants with a t (4; 11)
translocation and has a poor outcome 14.

28
T-cell ALL is defined by expression of the cytoplasmic CD3,
with CD7 plus CD2 or CD5 on leukemic blasts. T- ALL is
further divided into immunologic subtypes, of which early
T-progenitor (ETP)-ALL (CD1a and CD8 negative, CD5
weak, at least one stem-cell-associated or myeloid-
associated antigen) has stem-cell-like features with high
risk of induction failure or relapse16. (Level of evidence
2 A)
Myeloid antigen expression: Myeloid-associated antigen
expression is associated with specific ALL subgroups (MLL
gene and TEL-AML1 gene rearrangement). No
independent adverse prognostic significance exists for
myeloid-surface antigen expression17.

E) Cytogenetics
Blasts in ALL contain somatically acquired genetic
abnormalities that help in understanding pathogenesis
and strongly influence prognosis. This includes changes
in chromosome number (hyperdiploid / hypodiploid) and
chromosomal translocations.
Numerical abnormalities:
a) High Hyperdiploidy: It is defined as 51 to 65
chromosomes per cell or a DNA index larger than 1.16. It
occurs in 20% to 25% of cases of precursor B-cell ALL.
High hyperdiploidy generally occurs with clinically
favorable prognostic factors (patients aged 1–9 years with
a low WBC count) and is itself an independent favorable
prognostic factor18. (Level of evidence 2 A) Hyper diploid
leukemia cells are particularly susceptible to undergoing
apoptosis and accumulate higher levels of methotrexate
and its active polyglutamate metabolites which may
explain the favorable outcome commonly observed for

29
these cases19. Among specific trisomies, patients with triple
trisomies (4, 10, and 17) have been shown to have an
improved outcome20. (Level of evidence 2B)
b)Hypodiploidy: Patients with fewer than 44 chromosomes
have a worse outcome than patients with 44 or more
chromosomes in their leukemic cells .Cases with 24 to 28
chromosomes (near haploidy) have the worst outcome21.
(Level of evidence 2C)

Structural abnormailities
(Chromosomal translocations):
a) TEL-AML1, (t [12; 21]): It is seen in 20% to 25% of
cases of B-precursor ALL. The t (12; 21) occurs most
commonly in children aged 2 to 9 years22. It has good
prognosis. However, its impact may be modified by factors
such as early response to treatment, NCI risk category,
and treatment regimen. There is a higher frequency of
late relapses in patients with TEL-AML1 fusion compared
with other B-precursor ALL23.
b) Philadelphia chromosome, (t [9; 22] translocation): It
is present in 3% of children with ALL, and is more common
in older patients with precursor B-cell ALL and high WBC
count. It is associated with poor prognosis especially in
those who present with a high WBC count or have a slow
early response to initial therapy24. However, its prognosis
seems to have improved by incorporation of tyrosine kinase
inhibitors, such as imatinib, in the treatment. A COG study,
using intensive chemotherapy and concurrent imatinib
given daily, demonstrated a 3-year EFS rate of 80.5%25.
(Level of evidence 2A)
c) MLL gene rearrangements: It is seen in 5% of childhood
ALL cases. The t (4; 11) is the most common translocation

30
involving the MLL gene in children with ALL. Patients with
t (4; 11) are usually infants with high WBC counts; usually
have CNS disease and respond poorly to initial therapy.
Children with MLL rearrangement have a a better prognosis
than infants26 (Level of evidence 2A). The t (11; 19)
occurs in 1% of cases and occurs in both early B-lineage
and T-cell ALL. Outcome for infants with t (11; 19) is poor,
but outcome appears favorable in older children with T-
cell ALL2.
d) E2A-PBX1: (t[1; 19]) translocation: It occurs in 5% of
childhood ALL cases. It is associated with pre-B ALL
immunophenotype. It was associated with poor prognosis
in the context of less intensive antimetabolite-based
therapy in the past, but with most current treatment
protocols, the t(1;19) translocation has no adverse
prognostic significance except higher risk of CNS relapses.
28

3. Early Response To Therapy

The kinetics of the reduction in tumor burden in response
to treatment has been shown to be highly prognostic of
event free survival. Response to therapy is the most reliable
prognostic factor, as it reflects leukemic cell drug sensitivity,
intensity of therapy, and pharmacogenomic as well as
pharmacodynamic features of the host.

a) Peripheral blood response to steroid prophase:

Patients with a reduction in peripheral blast count to less
than 1,000/μL after a 7-day induction prophase with
prednisone and one dose of intrathecal methotrexate
(good prednisone response) have a more favorable
prognosis than patients whose peripheral blast counts
remain above 1,000/μL (a poor prednisone response)30

31
(Level of evidence 2A). German Berlin-Frankfurt-Muenster
(BFM) clinical trials group stratifies its treatment based on
early response to the 7-day prednisone prophase (Level
of evidence 2A).

b) Day 7 and day 14 bone marrow responses:

Patients who have a rapid reduction in leukemia cells to
less than 5% (M1 marrow) in their bone marrow within 7
or 14 days following initiation of multiagent chemotherapy
have a more favorable prognosis than do patients who
have slower clearance of leukemia cells from the bone
marrow31 (Level of evidence 2A).

c) Peripheral blood response to multiagent

induction therapy:
Patients with persistent circulating leukemic cells at 7 to
10 days after the initiation of multiagent chemotherapy
are at increased risk of relapse compared with patients
who have clearance of peripheral blasts within 1 week of
therapy initiation32 (Level of evidence 2A).

d) Induction failure:
5 % of patients do not achieve complete morphologic
remission by the end of induction therapy. A cut-off of 5
% blasts in the bone marrow is used to determine the
remission status. Patients at highest risk of induction failure
include T-cell phenotype and patients with B-precursor
ALL with very high presenting leukocyte counts or the
Philadelphia chromosome33. Induction failure portends a
very poor outcome34 (Level of evidence 2B).

e) Minimal residual disease (MRD)

Bone marrow morphology cannot discriminate well
between patients at high risk of relapse and patients with

32
excellent prognosis. Therefore, more sensitive techniques
have been developed for detection of submicroscopic levels
(<5%) of malignant cells during and after treatment i.e.
MRD. MRD is defined as the detection of the clones of
cells resistant to the chemotherapy given. Three types of
techniques allow detection of MRD of 10-3 to 10-6 (1
leukemic cell in 1000 to 1 million cells):
- Multiparametric flow cytometry (MPFC) for surface
phenotype of leukemia cell (sensitivity of 10-3 – 10-4)
can be used in up to 80-90% patients with ALL
- PCR for T-cell Receptor or Immunoglobulin gene
rearrangement or fusion transcripts (sensitivity 10-3
– 10-5) can be performed in 90-95% patients with
ALL
- PCR Analysis of breakpoint fusion regions of
chromosomal aberrations can be performed in 30-
45% patients
MRD is the most robust and strongest independent
predictor of outcome in children and adolescents with
ALL, which is independent of age, sex, Immunophenotype,
WBC count and treatment group. MRD discriminates
outcome even in subsets of patients defined by cytogenetic
abnormalities and other prognostic factors. Patients with
higher levels of end-induction MRD (>0.01%) have a
poorer prognosis than those with lower or undetectable
levels. Therefore, post-induction MRD is utilized as a factor
determining the intensity of post-induction treatment.
MRD levels at earlier (e.g., day 8 and day 15 of induction)
and later time points (e.g., week 12 of therapy) also predict
outcome (Level of evidence 2A)35,36. MRD can also be
used to prognosticate patients with ALL who are planned
for stem cell transplantation. Most of the MRD techniques
are expensive, cumbersome and require considerable

33
expertise and experience which precludes its use in most
laboratories and centres in developing countries. Recently,
a simplified inexpensive flow cytometric technique to
detect MRD in B-lineage ALL through detection of > 0.01%
of lymphoid progenitors (positive for CD10, CD19 with/
without CD34) at day 19 of induction therapy has shown
great promise. This simple technique may find wider
application across the globe.

4. Newer factors
A) Molecular genetic abnormalities:
IKAROS/IKZF1 deletions, JAK mutations and kinase
expression signatures have been associated with poor
prognosis in B cell acute lymphoblastic leukemia37, 38, and
39. Based on combination of gene expression profile and

flow cytometric measures of minimal residual disease

(MRD), children with high-risk B-precursor ALL can be
classified as low, intermediate, and high risk and thus allow
prospective identification of children who respond or fail
current treatment regimens.40 [Level of evidence: 3A]
Furthermore, integrated use of both MRD and IKZF1 status
allows prediction of 79% of all the relapses with 93%
specificity in MRD-medium risk group as compared to 46
and 54% of the relapses respectively predicted by use of
MRD or IKZF1 alone. The above findings signify that the
use of combined parameters enhances the risk
stratification, particularly for patients originally classified
as non-high risk41. [Level of evidence: 3A]

B) Pharmacogenetics:
It is the study of genetic variations in drug-processing
genes and individual responses to drugs which enables

34
improved identification of patients at higher risk for either
disease relapse or chemotherapy-associated side effects.
Patients with ALL who are homozygous for TMPT mutant
alleles experience severe or fatal myelotoxicity and
increased relapse because of long delays in therapy. 42
Studies from St Jude Children’s Research Hospital (SJCRH)
have shown that when patients are treated
pharmacologically according to phenotype or genotype,
carriers of variant TMPT alleles experience outcomes as
good as, or better than, those with wild-type TPMT.43 The
reduced folate carrier (RFC) is the primary transporter of
MTX into cells. RFC expression in leukemic blasts is linked
to MTX sensitivity, while defective transport associated with
reduced RFC expression is a common mechanism of
acquired methotrexate resistance. Increased copies of RFC
are present in hyperdiploid blasts and MTX-polyglutamate
accumulation in blasts correlates with better outcome in
hyperdiploid ALL 44. The null genotype of glutathione S-
transferases, enzymes that catalyze the inactivation of
many antileukemic agents, has been associated with a
reduced risk of relapse. 45

PROGNOSTIC GROUPS
1. NCI Risk-grouping - Age and WBC count at diagnosis
strongly correlates with outcome in B-Precursor ALL. The
high predictive value of age and WBC among all studies,
and the fact that these variables can be easily and reliably
measured by all investigators worldwide, make them one
of the most important prognostic factors based on which
the patients with ALL are divided into two risk groups:
Standard risk : Age 1-9.99 yrs
WBC < 50,000/cumm

35
High risk : Age > 10 yrs.
WBC > 50,000/cumm
2. Children’s Oncology Group (COG) risk stratification-
In the current COG classification system and treatment
algorithm, patients with precursor B-cell ALL are initially
assigned to a standard-risk or high-risk group based on
NCI grouping. All children with T-cell phenotype are
considered high risk regardless of age and initial WBC
count. Early treatment response, assessed by day 7 or day
14 marrow morphology along with end-induction MRD
assessment and cytogenetics is subsequently used to
determine the intensity of postinduction therapy. Patients
are classified as very high risk if they have very high risk
cytogentics with poor response or induction failure as
detailed below 46.

A) Standard Risk – Low

NCI Standard risk
Triple trisomies/ TEL-AML1 (+)
M1 marrow on Day 8 or 15 / M1 marrow on Day 29
Day 29 MRD < 0.1%
No CNS 2/3 and no testicular disease

B) Standard Risk – Average

NCI Standard risk
No Triple trisomies / No TEL-AML1
M1 marrow on Day 8 or 15 / M1 marrow on Day 29
Day 29 MRD < 0.1%

C) Standard Risk - High:

NCI Standard risk
CNS 3 or testicular disease
36
 M2/M3 marrow on Day 15
Day 29 MRD between 0.1% - 1%
MLL translocation with RER
Steroid pretreatment

D) High Risk
NCI high risk
M1 marrow on Day 8 or 15 / M1 marrow on Day 29
Day 29 MRD < 0.1%
No CNS 2/3 or testicular disease

E) High risk (for augmented therapy)

NCI high risk
Day 15 marrow M2/M3
Day 29 MRD between 0.1% - 1%
MLL translocation with RER
Steroid pretreatment

F) Very high risk

t (9;22) or BCR/ABL fusion
Hypo diploid clone
MLL translocation with a SER
M3 marrow on Day 29
MRD burden more than 1%
3. BFM GROUP: Risk stratification in BFM group is only
based on treatment response criteria. This includes
reduction in peripheral blast count to less than 1,000/μL
after a 7-day course of prednisolone and single dose of
intrathecal methotrexate. (Good response) and MRD
assessment at two time points, at the end of induction

37
(week 5) and at the end of consolidation (week 12).
Phenotypes, leukemic cell mass, CNS status at diagnosis
were also risk factors in old protocols but are not used in
the current risk stratification scheme.

Standard risk
Good response to Prednisolone prophase
MRD negative at wk -5 & wk -12

Intermediate risk
MRD positive at wk -5 & negative at wk -12

High risk
Poor response to Prednisolone prophase
MRD >1 % at wk -12
4. ST. JUDE - Risk classification in St. Jude total XV is
based mainly on MRD level which is assessed by flow
cytometry after 6 weeks of remission induction therapy.
Initial risk stratification at baseline is based on NCI risk
group, DNA index, cytogenetic features and
immunophenotyping. Finally, patients are restratified into
low, standard and high risk depending on the level of
MRD.
Low risk – NCI standard risk, DNA index >1.16 or TEL/
AML1, MRD < 0.01 %
Standard risk - NCI standard risk or high risk with MRD
(0.01- 1%), CNS leukemia, testicular leukemia,
Hypodiploidy, MLL gene rearrangement and t (1, 19) with
MRD (0.01-1%), T-ALL with MRD (0.01- 1%)
High risk - t (9, 22), T-ALL with MRD >1%, NCI standard
risk/high risk with MRD >1%,

38
5. Risk stratification in India and TMH Experience –The
outcome of pediatric ALL has improved significantly due
to improvement in treatment protocols, supportive care
and implementation of risk adapted therapy. However,
most developing countries including India still report low
survival rates as compared to developed world. This is
because of limitations in the existing health care systems
such poor infrastructure, high rate of abandonment, toxic
deaths inability to deliver treatment regimens of sufficient
intensity, co-morbidities (nutritional status, intercurrent
infections) or high risk biology of ALL. A prospective
multicentric (MCP-841) protocol to analyze the prognostic
factors and outcome in ALL in India has found that the
most significant risk factors for EFS were WBC count,
hemoglobin and lymphadenopathy47,48. However, over
time, biology of ALL as well as facilities for diagnostics
and treatment have changed and hence most clinical and
cytogenetic factors identified in western studies along with
morphological assessment of response can now be easily
used in most centres in India. A model risk-stratification
system is presented below:
Low-Risk: NCI standard-risk ,hyperdiploidy or Tel AML-1
fusion, day 7 PS –no blasts, day 14 BM – M1/M2
Standard risk: NCI standard risk, day 7 PS-no blasts, Day
14 BM –M1/M2
High risk: T-ALL, NCI high risk, CNS disease, testicular
disease, day 7 PS-no blasts, Day 14 BM-M1/M2
High risk: t(9,22), MLL rearrangement, Hypodiploidy, NCI
standard risk or high risk with day 7 PS with blasts or day
14/29 BM- M3

39
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24. Aricò M, Valsecchi MG, Camitta B, et al.: Outcome
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 MLL-ENL fusion and t(11;19)(q23;p13.3)
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28. Uckun FM, Sensel MG, Sather HN, et al.: Clinical
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acute lymphoblastic leukemia in the context of
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30. Manabe A, Ohara A, and Hasegawa D, et al.:
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35. Zhou J, Goldwasser MA, Li A, et al.: Quantitative
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36. van Dongen JJ, Seriu T, Panzer-Grümayer ER, et al.:
Prognostic value of minimal residual disease in acute
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37. Mullighan CG, Su X, Zhang J, et al. Deletion of IKZF1
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Engl J Med. 2009; 360:470-480.
38 . Mullighan CG, Zhang J, Harvey RC, et al. JAK
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9418.
39. Den Boer ML, van Slegtenhorst M, De Menezes RX,
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40. Huining Kang,1 I.-Ming Chen,1,2 Carla S. Wilson:
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41. Wanders E, van der velden VH et al. Integrated use
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IKZF1 alteration status accurately predicts 79% of
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42. Schmiegelow K, Forestier E, Kristinsson J, et al.:
Thiopurine methyltransferase activity is related to the
risk of relapse of childhood acute lymphoblastic
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Leukemia 23 (3): 557-64, 2009.
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importance of 6-mercaptopurine dose intensity in
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association of reduced folate carrier 80G>A
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lymphoblastic leukemia in India: An analysis of
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Annals of Oncology ,1999;10: 167-176.

46
 Management of Standard Risk &
High Risk Acute Lymphoblastic
Leukemia in Children
Dr. Akash Nahar, Dr. Amol Dongre

Introduction
The treatment of childhood acute lymphoblastic leukemia
(ALL) has advanced significantly over the past 3 decades,
with overall survival rates progressing from 20% to 75%
[1, 2]. The standard “backbone” of treatment for ALL has
remained unchanged for over 25 years and includes
remission induction, consolidation, treatment to prevent
overt leukemic infiltration of the central nervous system
(CNS directed therapy), and continuing (maintenance)
therapy. The steady improvement in survival of children
with ALL is a result of a number of modifications of this
treatment, the value of which have been confirmed by
randomized clinical trials.
In an effort to appropriately balance the risks and benefits
of therapy, “risk-adapted therapy” has been adopted. The
NCI criteria risk stratified in to standard and high risk based
on : age, initial white blood cell (WBC) count, and the
presence of extramedullary disease at diagnosis.[3]
Currently most cooperative groups use additional risk
47
factors that have been shown to have an impact on patient
outcomes (eg, ploidy, blast karyotype/cytogenetics, and
early morphologic response (table 1). The resulting
classification system thus incorporates the strongest
prognostic indicators predictive of outcome, and stratifies
the treatment based on their risk of relapse (table 2).
Table 1: Prognostic factors in ALL
Adverse Favourable
Clinical Age <1, >10 1-9
WBC >50,000 <50,000
Lab DNA Index <1(Hypodiploidy) >1.16(Hyperdiploidy)
Immuno- T-ALL
phenotype

Table 2: Risk stratification in ALL

Risk groups Features Percentage
Low risk 1. Hyperdiploid (DNA Index >1.16) 20%
2. TEL-AML1 fusion 20%
Inter. risk 1. Std risk age/WBC without
genetic risk features 15%
High risk 1. T-Cell ALL 6%
2. High risk age/WBC without 15%
genetic risk factors 15%
V.High risk 1. BCR-ABL fusion with high WBC 3%
2. MLL rearrangement with slow 4%
response to treatment
3. Induction failures 2%

Management of ALL
Induction Chemotherapy for ALL
Three-drug induction therapy using vincristine,
corticosteroid (prednisone or dexamethasone), and

48
L-asparaginase in conjunction with intrathecal (IT) therapy,
results in complete remission (CR) rates of greater than
95%.[4] For patients who are at standard risk or low risk
of treatment failure, four-drug induction therapy does not
appear necessary for favorable outcome provided that
adequate postremission intensification therapy is
administered.[5-7] Because of the likelihood of increased
toxicity with four-drug induction therapy, many co-
operative groups including the Children’s Oncology Group
(COG) protocols for National Cancer Institute (NCI)
standard-risk precursor B-cell acute lymphoblastic
leukemia(ALL) utilize a three-drug induction consisting of
dexamethasone, vincristine, and PEG-L-asparaginase.
For patients presenting with high-risk features, a more
intensive induction regimen (four or five agents) may result
in improved event-free survival (EFS),[4] and such patients
generally receive induction therapy that includes an
anthracycline (e.g., daunorubicin) in addition to vincristine,
prednisone/dexamethasone, plus L-asparaginase

Consolidation/Intensification Therapy
Once remission has been achieved, systemic treatment in
conjunction with central nervous system (CNS) sanctuary
therapy follows. The intensity of the postinduction
chemotherapy varies considerably depending on risk group
assignment, but all patients receive some form of
intensification following achievement of remission and
before beginning maintenance therapy. Intensification may
involve use of the following:
 Intermediate-dose or high-dose methotrexate with
leucovorin rescue or escalating-dose methotrexate
without rescue.[ 5-8]
 Drugs similar to those used to achieve remission
(reinduction or delayed intensification).[5,9]

49
 Different drug combinations with little known cross-
resistance to the induction therapy drug
combination.[10]
 L-asparaginase for an extended period of time.[8,11]
 Combinations of the above.[5,12,13]
In children with standard-risk acute lymphoblastic
leukemia (ALL), regimens utilizing a limited number of
courses of intermediate-dose or high-dose methotrexate
as consolidation followed by maintenance therapy
(without a reinduction phase) have been used with good
results.[6,7,14] Similarly favorable results for standard-risk
patients have been achieved with regimens utilizing
multiple doses of L-asparaginase (20–30 weeks) as
consolidation, without any postinduction exposure to
alkylating agents or anthracyclines.[11,15]
Postinduction consolidation for regimens using a German
Berlin-Frankfurt-Muenster “BFM-backbone,” such as those
of the Children’s Oncology Group (COG), include a delayed
intensification phase, during which patients receive a 4-
week reinduction (including anthracycline) and
reconsolidation containing cyclophosphamide, cytarabine,
and 6-thioguanine given approximately 3 months after
remission is achieved.[5,16,17] Clinical trials conducted
in the 1980s and early 1990s demonstrated that the use
of delayed intensification improved outcome for children
with standard-risk ALL, in comparison to that achieved
without an intensification phase.[16,18,19] In a Children’s
Cancer Group (CCG) study, which included a three-drug
induction and utilized prednisone as the corticosteroid
throughout all treatment phases, two blocks of delayed
intensification produced a small event-free survival (EFS)
benefit compared with one block of delayed intensification
in intermediate-risk patients.[20] The benefit of two blocks

50
of delayed intensification, however, may depend, in part,
on the type of corticosteroid used (prednisone vs.
dexamethasone). In a subsequent CCG study for standard-
risk ALL in which dexamethasone was used instead of
prednisone, two blocks of delayed intensification were not
associated with a survival benefit in patients who were
rapid early responders.[21] All patients are randomized
to receive either Capizzi methotrexate or high-dose
methotrexate with leucovorin rescue as given in BFM
protocols during the first interim maintenance phase.
In high-risk patients, a number of different approaches
have been used with comparable efficacy.[11,22,23];
[17][Level of evidence: 2D] Treatment for high-risk
patients generally is more intensive than that for standard-
risk patients, and typically includes higher cumulative doses
of multiple agents, including anthracyclines and/or
alkylating agents. The former CCG developed an
augmented BFM treatment regimen featuring repeated
courses of escalating-dose intravenous methotrexate
(without leucovorin rescue) given with vincristine and
asparaginase during interim maintenance and additional
vincristine/L-asparaginase pulses during initial
consolidation and delayed intensification. Augmented
therapy also included a second interim maintenance and
delayed intensification phase. In the CCG-1882 trial,
National Cancer Institute (NCI) high-risk patients with slow
early response (M3 marrow on day 7 of induction) were
randomly assigned to receive either standard or
augmented BFM therapy. The augmented therapy regimen
produced a significantly better EFS compared with
standard CCG modified BFM therapy.[24] In an Italian
study, investigators showed that two applications of
delayed intensification therapy (protocol II) significantly
improved outcome for patients with a poor response to a

51
prednisone prophase.[25] Of note, there is a significant
incidence of osteonecrosis of bone in teenaged patients
who receive the augmented BFM regimen.[26]
The augmented BFM regimen has also been evaluated in
children with high-risk ALL and a rapid early response to
induction therapy. For these children, augmented intensity
during consolidation, interim maintenance, and delayed
intensification resulted in a higher EFS rate than that
achieved with standard-intensity treatment. Increased
duration of intensive therapy was not beneficial, and a
single application of delayed intensification was as effective
as two applications.[27][Level of evidence: 1A]
For children with Ph+ ALL, imatinib mesylate in
conjunction with chemotherapy during postinduction
therapy has produced a 3-year EFS of 87.7% ± 10.9%.
These patients fared better than historic controls treated
with chemotherapy alone (without imatinib), and at least
as well as the other patients on the trial who underwent
allogeneic transplantation.[28] Longer follow-up is
necessary to determine if this novel treatment improves
cure rate or merely prolongs DFS
Infant ALL is uncommon, representing approximately 2%
to 4% of cases of childhood ALL.[29] Despite the inclusion
of postinduction intensification courses with high doses
of cytarabine and methotrexate.[30-32] , long-term EFS
rates remain below 50%, and for those infants with MLL
gene rearrangement, the EFS rates continue to be in the
17% to 40% range.[30,31,33-35][Level of evidence: 2A]
Factors predicting poor outcome for MLL-rearranged
infants include a very young age (<6 months), extremely
high presenting leukocyte count (e”300,000/ìL), and high
levels of MRD at the end of induction and consolidation
phases of treatment.[31]; [36][Level of evidence: 3D].

52
The role of bone marrow transplantation in infants with
MLL-rearranged ALL remains controversial.

Maintenance Therapy
(Standard risk and High risk ALL)
The backbone of maintenance therapy in most protocols
includes daily oral mercaptopurine and weekly oral
methotrexate. On many protocols, intrathecal
chemotherapy for CNS sanctuary therapy is continued
during maintenance therapy. Some patients may develop
severe hematopoietic toxicity because of an inherited
deficiency (homozygous mutant) of thiopurine S-
methyltransferase, an enzyme that inactivates
mercaptopurine. Patients who are heterozygous for this
mutant enzyme gene generally tolerate mercaptopurine
without serious toxicity, but they do require more frequent
dose reductions for hematopoietic toxicity than patients
who are homozygous for the normal allele.[37] The use
of continuous 6-thioguanine (6-TG) instead of 6-
mercaptopurine (6-MP) during the maintenance phase is
associated with an increased risk of hepatic complications,
including veno-occlusive disease and portal
hypertension.[38-41] Because of the risk of hepatic
complications, 6-TG is no longer utilized in maintenance
therapy in current protocols. It remains unknown if short
term use of 6-TG can improve outcome without excessive
toxicity.
Pulses of vincristine and corticosteroid are often added to
the standard maintenance backbone, A CCG randomized
trial demonstrated improved outcome in patients receiving
monthly vincristine/prednisone pulses,[42] and a meta-
analysis combining data from six clinical trials showed an
EFS advantage for vincristine/prednisone pulses.[43],
Dexamethasone is preferred over prednisone for younger

53
patients based on data from a CCG study, where
dexamethasone use had significantly fewer CNS relapses
and a significantly better EFS rate. [16, 44]
Maintenance chemotherapy generally continues until 2
to 3 years of continuous complete remission. On some
studies, boys are treated longer than girls;[12] on others,
there is no difference in the duration of treatment based
on gender.[11,17] Extending the duration of maintenance
therapy beyond 3 years does not improve
outcome.[42,45][Level of evidence: 1A]

Central Nervous System (CNS) Therapy

Options for CNS-directed therapy include IT chemotherapy,
high dose systemic chemotherapy, and cranial radiation.
The type of CNS-therapy that is used is based on a patient’s
risk of CNS-relapse, with higher-risk patients receiving more
intensive treatments. A major goal of current ALL clinical
trials are to provide effective CNS therapy while minimizing
neurotoxicity and other late effects. The proportion of
patients receiving cranial radiation has decreased
significantly over time, with those receiving cranial
radiation, the dose has been significantly reduced. All
therapeutic regimens for childhood ALL include IT
chemotherapy. IT chemotherapy is usually started at the
beginning of induction, intensified during consolidation
and, in certain protocols, continued throughout the
maintenance phase. IT chemotherapy typically consists of
either methotrexate alone or methotrexate with cytarabine
and hydrocortisone. [46] Unlike IT cytarabine, IT
methotrexate has a significant systemic effect, which may
contribute to prevention of marrow relapse.[47]
Systemically administered drugs, such as dexamethasone,
L-asparaginase, high-dose methotrexate with leucovorin
rescue, and high-dose cytarabine, provide some degree

54
of CNS protection. For example, in a randomized CCG study
of standard-risk patients who all received the same dose
and schedule of IT methotrexate without cranial
irradiation, oral dexamethasone was associated with a
50% decrease in the rate of CNS relapse compared with
oral prednisone.[48]
Detail regarding CNS prophylaxis has been mentioned in
next chapter.

Response Based Treatment of ALL

The rapidity with which leukemia cells are eliminated
following onset of treatment is associated with long-term
outcome, as is level of residual disease at the end of
induction [49]. Because treatment response is influenced
by the drug sensitivity of leukemic cells and host
pharmacodynamics and pharmacogenomics, early
response has strong prognostic significance. Various ways
of evaluating the leukemia cell response to treatment have
been utilized, including the following:

1. Day 7 and day 14 bone marrow responses:

Patients who have a rapid reduction in leukemia cells to
less than 5% in their bone marrow within 7 or 14 days
following initiation of multiagent chemotherapy have a
more favorable prognosis than do patients who have
slower clearance of leukemia cells from the bone
marrow.[50] Morphologic response to treatment
continues to be used in some ALL trials to stratify patients
into prognostic categories for treatment assignment [51]

2. Peripheral blood response to steroid prophase:

Patients with a reduction in peripheral blast count to less
than 1,000/μL after a 7-day induction prophase with
prednisone and one dose of intrathecal methotrexate - (a

55
good prednisone response), have a more favorable
prognosis than do patients whose peripheral blast counts
remain above 1,000/μL - (a poor prednisone
response).[52,53] Treatment stratification for protocols of
the German Berlin-Frankfurt-Muenster (BFM) clinical trials
group is partially based on early response to the 7-day
prednisone prophase (administered immediately prior to
the initiation of multiagent remission induction). Patients
with no circulating blasts on day 7 have a better outcome
than those patients whose circulating blast level is between
1 and 999/μL.[54,55]

3. Peripheral blood response to multiagent

induction therapy:
Patients with persistent circulating leukemia cells at 7 to
10 days after the initiation of multiagent chemotherapy
are at increased risk of relapse compared with patients
who have clearance of peripheral blasts within 1 week of
therapy initiation. Rate of clearance of peripheral blasts
has been found to be of prognostic significance in both
T-cell and B-lineage ALL.[56]

4. Induction failure:
The vast majority of children with ALL achieve complete
morphologic remission by the end of the first month of
treatment. The presence of greater than 5% lymphoblasts
at the end of the induction phase is observed in up to 5%
of children with ALL. Patients at highest risk of induction
failure include those with T-cell phenotype (especially
without a mediastinal mass) and patients with B-precursor
ALL with very high presenting leukocyte counts and/or
the Philadelphia chromosome.[57,58] Induction failure
portends a very poor outcome. In the French FRALLE 93
study, the 5-year OS rate for patients with initial induction
failure was 30%.[58]

56
5. MRD determination:
Multiple studies have demonstrated that end-induction
MRD is an important, independent predictor of outcome
in children and adolescents with ALL.[59-61] MRD
response discriminates outcome in subsets of patients
defined by age, leukocyte count, and cytogenetic
abnormalities.[62] Patients with higher levels of end-
induction MRD have a poorer prognosis than those with
lower or undetectable levels.[58-61] End-induction MRD
is used by almost all groups as a factor determining the
intensity of post induction treatment, with patients found
to have higher levels allocated to more intensive therapies.
MRD levels at earlier (e.g., day 8 and day 15 of induction)
and later time points (e.g., week 12 of therapy) also predict
outcome.[58,60-66]
MRD measurements, in conjunction with other presenting
features, have also been used to identify subsets of patients
with an extremely low risk of relapse. The COG reported a
very favorable prognosis (5-year EFS of 97% ± 1%) for
patients with B-precursor phenotype, NCI standard risk
age/leukocyte count, and favorable cytogenetic
abnormalities (either high hyperdiploidy with favorable
trisomies or the TEL-AML1 fusion) who had less than
0.01% MRD levels at both day 8 (from peripheral blood)
and end-induction (from bone marrow).[60]

Controversies in the treatment of ALL

(A) Dexamethasone versus prednisone
Many current regimens utilize dexamethasone instead of
prednisone during remission induction and later phases
of therapy. The Children’s Cancer Group (CCG) conducted
a randomized trial comparing dexamethasone and
prednisone in standard-risk ALL patients, and reported that

57
dexamethasone was associated with a superior EFS.[48]
Results from another randomized trial conducted by the
United Kingdom Medical Research Council (MRC)
demonstrated that dexamethasone was associated with
a more favorable outcome than prednisolone in all patient
subgroups.[67]
While dexamethasone may be more effective than
prednisone, data also suggest that dexamethasone may
also be more toxic, especially in the context of more
intensive induction regimens and in adolescents. Several
reports indicate that dexamethasone may increase the
frequency and severity of infections and/or other
complications in patients receiving anthracycline-
containing induction regimens.[68,69,] The increased risk
of infection with dexamethasone during the induction
phase has not been noted with three-drug induction
regimens (vincristine, dexamethasone, and L-
asparaginase).[67] Dexamethasone appears to have a
greater suppressive effect on short-term linear growth than
prednisone,[70] and has been associated with a higher
risk of osteonecrosis, especially in adolescent patients.[71]

(B) High dose versus Capizzi Methotrexate

The optimal dose and schedule of methotrexate is a subject
of considerable debate. Capizzi I methotrexate (escalating
IV methotrexate without leucovorin rescue, followed by
asparaginase) is a key element of augmented BFM therapy
which has been efficacious in the treatment of higher risk
childhood ALL. Recent COG trials for children with higher
risk ALL (CCG-1961, POG 9906) utilized components of
augmented BFM including Capizzi I methotrexate every
10 days in conjunction with vincristine and L-asparaginase
during interim maintenance with good results.

58
Higher dose methotrexate with leucovorin rescue has
previously been shown to be a successful strategy in BFM
clinical ALL trials and in the POG 9404 trial for T-ALL. High
risk patients with a lower steady state methotrexate levels
are at increased risk of relapse. Therefore, it has been
postulated that high dose methotrexate with rescue may
be superior to Capizzi I methotrexate. However, although
Capizzi schedule is uniformly acceptable in standard risk
ALL, its superiority in terms of efficacy in high risk ALL is
still being evaluated.

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37. Biondi A, Rizzari C, Valsecchi MG, et al.: Role of
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43. Vora A, Mitchell CD, Lennard L, et al.: Toxicity and
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pulses of vincristine and prednisone prevent bone
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46. Relling MV, Dervieux T: Pharmacogenetics and cancer
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48. Bostrom BC, Sensel MR, Sather HN, et al.:
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49. Carroll WL, Bhojwani D, Min DJ, et al.: Pediatric acute
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51. Schrappe M, Reiter A, Ludwig WD, et al.: Improved
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52. Aricò M, Basso G, Mandelli F, et al.: Good steroid
response in vivo predicts a favorable outcome in
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53. Lauten M, Stanulla M, Zimmermann M, et al.: Clinical
outcome of patients with childhood acute
lymphoblastic leukaemia and an initial leukaemic
blood blast count of less than 1000 per microliter.
Klin Padiatr 213 (4): 169-74, 2001 Jul-Aug.
54. Manabe A, Ohara A, Hasegawa D, et al.: Significance
of the complete clearance of peripheral blasts after
7 days of prednisolone treatment in children with
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55. Griffin TC, Shuster JJ, Buchanan GR, et al.: Slow
disappearance of peripheral blood blasts is an adverse
prognostic factor in childhood T cell acute
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56. Silverman LB, Gelber RD, Young ML, et al.: Induction
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57. Oudot C, Auclerc MF, Levy V, et al.: Prognostic factors
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1496-503, 2008.
58. Zhou J, Goldwasser MA, Li A, et al.: Quantitative
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59. Borowitz MJ, Devidas M, Hunger SP, et al.: Clinical
significance of minimal residual disease in childhood
acute lymphoblastic leukemia and its relationship to
other prognostic factors: a Children’s Oncology
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60. Coustan-Smith E, Sancho J, Hancock ML, et al.: Use
of peripheral blood instead of bone marrow to
monitor residual disease in children with acute
lymphoblastic leukemia. Blood 100 (7): 2399-402,
2002.
61. Conter V, Bartram CR, Valsecchi MG, et al.: Molecular
response to treatment redefines all prognostic factors
in children and adolescents with B-cell precursor
acute lymphoblastic leukemia: results in 3184
patients of the AIEOP-BFM ALL 2000 study. Blood
115 (16): 3206-14, 2010.
62. Basso G, Veltroni M, Valsecchi MG, et al.: Risk of
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27 (31): 5168-74, 2009.
63. Panzer-Grümayer ER, Schneider M, Panzer S, et al.:
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64. Coustan-Smith E, Sancho J, Behm FG, et al.:
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66. Carroll WL, Bhojwani D, Min DJ, et al.: Pediatric acute
lymphoblastic leukemia. Hematology (Am Soc
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67. Mitchell CD, Richards SM, Kinsey SE, et al.: Benefit
of dexamethasone compared with prednisolone for
childhood acute lymphoblastic leukaemia: results of
the UK Medical Research Council ALL97 randomized
trial. Br J Haematol 129 (6): 734-45, 2005.
68. Hurwitz CA, Silverman LB, Schorin MA, et al.:
Substituting dexamethasone for prednisone
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69. Belgaumi AF, Al-Bakrah M, Al-Mahr M, et al.:
Dexamethasone-associated toxicity during induction
chemotherapy for childhood acute lymphoblastic
leukemia is augmented by concurrent use of
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70. Ahmed SF, Tucker P, Mushtaq T, et al.: Short-term
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71. Mattano LA Jr, Sather HN, Trigg ME, et al.:
Osteonecrosis as a complication of treating acute
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3262-72, 2000.

69
 Central Nervous System
Prophylaxis In Paediatric Acute
Lymphoblastic Leukemia

The treatment of Paediatric acute lymphoblastic leukemia

(ALL) in children is a success story in modern medicine
with 5-year survival rates approaching 90% in the West
and 50-60 % in developing countries. Survival rates for
children with ALL did not improve until the 1970s when
central nervous system (CNS)-directed therapy was
instituted in addition to effective systemic chemotherapy.
Before then, more than half of the complete remissions
induced by systemic chemotherapy ended in CNS relapse.1
Current strategies for CNS-directed therapy involve
effective systemic chemotherapy (eg, dexamethasone,
high-dose methotrexate, intensive asparaginase, high-dose
cytarabine), CNS radiation in selected high-risk cases and
early intensification and optimization of intrathecal
therapy. Recent clinical trials in ALL focus on further
reducing CNS relapse rates and improving long-term
survival rates by providing more precise individualized
therapy that avoids over- or under-treatment. With the
vastly improved long-term survival rates, efforts are being
made to minimize late complications such as secondary

70
cancers, neurocognitive defects, and multiple
endocrinopathy by eliminating or decreasing the dose of
cranial irradiation. Still, CNS relapse remains an important
cause of morbidity and mortality and can occur in up to
6% of ALL patients. 2

Definitions of Central Nervous System Disease:

The terms commonly accepted to denote the CNS status
are;
CNS1 – no detectable blast cells in CSF;
CNS2 -fewer than five leucocytes per μL with detectable
blast cells in a cytocentrifuged preparation of CSF;
CNS3 –the presence of overt CNS leukaemia (originally
defined by the Rome Workshop) i.e. five or more leucocytes
per μL with identifiable blast cells, or the presence of
cranial-nerve palsies.3
Traumatic lumbar puncture with blasts (TLP) – defined
as 10 or more erythrocytes per μL of CSF with the presence
of blast cells 2.
The relevance of this CNS risk classification has been a
subject of debate, because a number of groups did not
observe significant differences in outcome for patients
categorized as CNS1 versus CNS2. 5–9 Older studies which
did not include early CNS directed therapy found that
patients with CNS 2 status had previously a 3.2- fold
increased risk for CNS relapse compared to those with
CNS1 status 10.However, later studies with prophylactic
CNS irradiation and early CNS directed therapy identified
no difference in relapse rates between these two groups,
6,7 which has lead to the conclusion that the adverse

prognosis in these groups can be abrogated by appropriate

treatment.

71
The adverse outcome associated with a ‘traumatic lumbar
puncture with blasts’ 2 has been attributed to several
reasons: contamination of the CNS with leukaemia blast
cells, Inefficacy of subsequent intrathecal treatments by
the collapse of the thecal sac from haematoma formation
or CSF collection, or by scarring / segmentation of the
subarachnoid membrane; and also the likelihood of
masking of a CNS 2/3 status by the traumatic tap.2

Diagnosis of Central Nervous System Disease

The standard method of diagnosing CNS disease in
leukaemia is light microscopic examination of cytospin
preparations of cerebrospinal fluid (CSF) samples for the
presence of leukemic cells. Staining for terminal
deoxynucleotidyl transferase (tDT) may help identify
leukaemic blasts when the morphology is in doubt
.Immunocytology to detect leukemia-associated cell
surface antigens has also been used to diagnose CNS
disease.11 Multicolour flow cytometry using multiple
antibody panels and polymerase chain reaction (PCR)
methods have been found to have increased sensitivity in
picking up blast cells in the CSF.12 In contrast to cytology,
magnetic resonance imaging (MRI) has been reported to
be highly sensitive for the presence of meningeal pathology
but nonspecific for the disease entity.6

Risk factors for CNS relapse

Many factors associated with an increased risk of CNS
relapse have been identified for pediatric patients with
ALL.2 The most important among these is the presence of
blasts in the cerebrospinal fluid and the CNS status as
detailed above. The presence of a T-cell phenotype and a
presenting leucocyte count of more than 100×10y /L also
have a high risk of CNS relapse 13.Several high risk genetic

72
factors which have been associated with unfavorable CNS
prognosis include hypodiploidy (<45 chromosomes per
leukemia cell), the translocation t(4;11) with the MLL-AF4
fusion gene, the translocation t(1;19) with E2A-PBX1
fusion gene, and the Philadelphia chromosome, t(9; 22).
14

Gene polymorphisms. of vitamin D receptor start site in

high-risk ALL patients and the thymidylate synthase 3/3
genotype in low-risk patients are associated with
predisposition for CNS relapse 15,16 The vitamin D receptor
regulates the expression of proteins that are important in
the disposition of some anticancer agents,including
steroids. The polymorphism of thymidylate synthase gene
has been linked to increased expression of the enzyme,
one of the major targets of methotrexate (MTX). More
recently, high expression of interleukin-15 in leukemic cells
was found to be associated with CNS leukemia at diagnosis
and subsequent relapse. It has been postulated that IL-15
is involved with leukaemic cell migration into the CNS.
17,18

How to Optimise CNS Directed Treatment?

Ideally, CNS-directed therapy starts early in the clinical
course and varies depending on the patient’s risk of relapse
and the intensity of systemic treatment. Important
prophylactic strategies for CNS control include effective
systemic chemotherapy, early intensification of intrathecal
therapy and cranial radiation in selected cases.

Optimization of Systemic Chemotherapy:

1. Selection of Corticosteroids:
Corticosteroids are an essential and time tested component
of ALL treatment. Despite some debate on a truly

73
equivalent dose, compared to prednisolone,
dexamethasone appears to have a stronger antileukemic
effect in vitro and has been shown to provide better
leukemic CNS control and lower relapse rates. However,
dexamethasone was also associated with increased side-
effects including severe infectious complications. 19,20 The
differences in CNS relapse rates may be due to either the
longer half-life of dexamethasone versus prednisone (>32
hours vs 4 to 6 hours, respectively) 19,21 or the low protein
binding property of dexamethasone resulting in a higher
CNS bioavailability.22, 23

2. Dose & timing of Methotrexate (MTX):

High-dose intravenous (IV) MTX is used as a treatment for
CNS disease because of its ability to cross the blood-brain
barrier (BBB). However, a meta-analysis of eight studies
that compared cranial radiation plus CNS-directed therapy
with high-dose MTX plus intrathecal therapy 24 found that
high-dose MTX reduced hematologic relapse and
improved EFS but had only a marginal effect on the control
of CNS relapse . However, this meta-analysis included
studies using a wide range of MTX doses (0.5 g/m2 to 8
g/m2) and dose schedules. The higher MTX doses (eg, 5
g/m2) used early in the treatment course in many current
trials are likely to be more effective in securing CNS control,
provided that the rescue with leucovorin is given at an
appropriate time (usually 42 h from the start of the
methotrexate infusion) and dose. 24

3. Selection of Asparaginase:
The efficacy of L asparaginase in preventing CNS relapse
depends on the type and dose of asparaginase used. A
study 25 showed that patients treated with Erwinia derived
asparaginase experienced CNS relapse at nearly six times
the rate compared to those treated with Escherechia coli-
74
derived asparaginase , when the two preparations were
administered at the same dose (25,000 IU/m2 weekly ×
20 doses). This finding was attributed to the fact that the
Erwinia preparation had a much shorter half-life than the
E coli preparation. Thus, the optimal dose should be
carefully chosen depending on the type of asparaginase
preparation used.

4. Use of thiopurines:
Thioguanine is more potent than mercaptopurine, results
in higher concentrations of thioguanine nucleotides in cells
and achieves higher cytotoxic concentrations in CSF. In
three randomised trials comparing the efficacy of these
two drugs, Thioguanine, given at a daily dose of 40 mg/
m2 or more, produced better antileukaemia responses and
significantly lower CNS relapse rates than a standard dose
of mercaptopurine (75 mg/m2), but was associated with
significant thrombocytopenia, increased risk of death in
remission, and an very high rate (10–20%) of hepatic veno-
occlusive disease. Hence, Thioguanine is not the drug of
choice for maintenance therapy in ALL, although whether
short courses of thioguanine could decrease CNS relapses
without adding undue toxic effects remains to be tested.
26,27, 28

Optimization of Intrathecal Therapy:

All therapeutic regimens for childhood ALL include
Intrathecal (IT) chemotherapy. IT chemotherapy is typically
started at the beginning of induction and is intensified
during consolidation (four to eight doses of IT given every
1–3 weeks), and, in certain protocols, continued
throughout the maintenance phase. IT chemotherapy
typically consists of either methotrexate alone or
methotrexate with cytarabine and hydrocortisone (triple
intrathecal treatment) . 29
75
Choice of Intrathecal Therapy:
MTX has been the standard drug used for intrathecal
therapy. Unlike IT cytarabine, IT methotrexate has a
significant systemic effect, which may contribute to
prevention of marrow relapse. 30 Triple intrathecal therapy
was designed on the premise that addition of cytarabine
and a corticosteroid to intrathecal methotrexate may have
additive or synergistic benefits, and that the added
corticosteroid may reduce arachnoiditis associated with
other intrathecal drugs.
The seminal CCG 1952 clinical trial,which compared
intrathecal MTX with triple intrathecal therapy for
presymptomatic CNS treatment of childhood ALL, found
that as compared to IT MTX, triple intrathecal therapy
reduced the risk of CNS relapse but was linked to
significantly worse overall survival (6-year survival: triple
intrathecal therapy 90.3% vs intrathecal MTX 94.4%; P =
0.01). This seemingly paradoxical reduction in overall
survival was attributed to a significantly greater number
of testicular and bone marrow relapses that have lower
salvage rates than CNS relapse. Therefore, although triple
intrathecal therapy is beneficial, it is recommended that
concurrent intense systemic treatment is also provided to
prevent bone marrow and testicular relapses.

Essentials of Intrathecal Therapy:

Several factors associated with intrathecal administration
should be ensured to optimise good CNS drug distribution
which is essential for effective CNS-directed therapy.
Keeping the patient in a prone position for 1 hour
following administration of intrathecal chemotherapy
leads to 10-fold decrease in ventricular dose. 32 Use of
atraumatic noncutting spinal needles (22 gauge or lower)
for the procedure can reduce the likelihood of CSF leaks

76
and thrombosis 33. Intrathecal medication given in a large
volume (6 mL or more) attains a better distribution in the
CNS than if given in a small volume. The early removal of
stylet following lumbar puncture immediately after
passage through the epidermal and subcutaneous tissues
is associated with a significant increase in the effectiveness
of intrathecal therapy.34 It is also equally important to
minimize chances of a traumatic lumbar puncture at
diagnosis which can lead to increased risk of CNS relapse
.4,6,8 This can be accomplished through correction of
thrombocytopenia(platelet count > 100×109/L) and
coagulopathy immediately before the diagnostic lumbar
puncture, and procedure performed by most experienced
clinician under deep sedation or general anaesthesia.

Cranial Irradiation:
Cranial irradiation is considered the most effective form
of CNS-directed treatment, and has played a central role
in the successful treatment of CNS leukemia since the
1960s, but substantial rates of long-term complications
such as secondary neoplasms, endocrine diseases, growth
impairment, neurocognitive dysfunction, and neurotoxic
effects are of concern. 35,36 Hence, its role is gradually
shrinking.

Current Indications of cranial Irradiation:

In most collaborative groups, cranial irradiation is still
recommended for 2% to 20% of patients at very high risk
of CNS relapse (excepting infants), especially those with
CNS 3 status, T-cell phenotype presenting with
hyperleukocytosis, high risk genetic factors ( t(4;11),
t(1;19), t(9; 22) and hypodiploidy) and poor response to
chemotherapy (poor prednisolone response, post
induction MRD>10-3, M2/M3 marrow on day21).2,13 The

77
St. Jude Total Therapy Study XV showed that with
intensification of systemic and intrathecal
chemotherapy(eg, reinduction therapy, intensive
asparaginase, and intensive triple intrathecal therapy),
prophylactic cranial irradiation can be totally omitted
without compromising overall survival.38 The 8-year event-
free survival and CNS relapse in the study were 85.6%
and only 2.7% respectively. 39 In the dexamethasone-based
DCOG ALL-9 protocol without cranial irradiation, the
5-year event-free survival was 81% and isolated CNS
relapses occurred in 2·6% patients only. However, in
T-ALL with high WBC counts, elimination of preventive
cranial radiation has been found to cause a significant
increase in systemic recurrences. 13
In the recently published EORTC 58881 trial accruing 2025
patients, cranial irradiation was omitted in all patients.
The 8-year EFS rate in the CNS1, TLP+, CNS2 and CNS3
group was 69.7%, 68.8%, 71.3% and 68.3% respectively.
The 8-year incidence of isolated or combined CNS relapse
incidence was 7.6%, 3.5%, 10.2% and 11.7%, respectively.
This strategy of omission of cranial irradiation is based
not only on an expected low rate of CNS relapse, but also
on a high salvage rate for patients with an isolated CNS
relapse who have not received cranial irradiation as initial
CNS-directed treatment thus reserving it for salvage
treatment in ALL and sparing most patients from its toxic
eûects. Although Overall outcome of these trials was
favorable, some subgroups of patients remained at high
risk for relapse, the acute systemic and CNS toxic effects
were substantial, and the long-term neurocognitive
outcomes are unknown. Hence, longer prospective
evaluation of outcome and toxicity is needed before
making it a standard of care.

78
Dose, type and extent of cranial irradiation:
Investigators participating in the Berlin-Frankfurt-Münster
(BFM) trials reduced the dose of cranial irradiation to 12
Gy for high-risk patients without CNS leukemia at
diagnosis. However, even with the lower dose, there was
still a cumulative incidence of 1.7% of secondary
neoplasms at 15 years.37 Finally, a randomised trial showed
that hyperfractionated (twice daily) cranial irradiation failed
to reduce neurocognitive late effects and might have
compromised efficacy against leukaemia, as compared
with conventionally dosed radiation. An early trial by the
Pediatric Oncology Group suggested that craniospinal
irradiation was more effective than cranial irradiation, this
finding needs confirmation with intensive contemporary
treatment.

CNS therapy for standard-risk patients:

In children classified as standard Risk (NCI), a combination
of appropriate (and ideally individualized) systemic
chemotherapy and intrathecal chemotherapy has been
found to be effective, resulting in CNS relapse rates of less
than 5% for children with standard-risk ALL. 40 -44 The use
of cranial radiation does not appear to be a necessary
component of CNS-directed therapy for these patients.
The DFCI ALL Consortium 95-01 trial found no significant
difference in two groups of standard risk patients who
were randomly assigned to receive either 18 Gy of cranial
irradiation with IT chemotherapy or more frequently dosed
IT chemotherapy alone (without radiation.)25 [Level of
evidence 2B.]
The CCG-1952 study compared the relative efficacy and
toxicity of triple IT chemotherapy with methotrexate as
the sole IT agent in nonirradiated patients for NCI standard-
risk patients. 31 As mentioned in the preceding text,the

79
isolated CNS relapse rate was lower for the group which
received triple intrathecal therapy as compared to those
who received IT methotrexate alone (P = .04). There were,
however, more bone marrow relapses in the group that
received the triple IT therapy, leading to a worse overall
survival .There was no significant difference in toxicity
between these two groups ,either immediate or in a follow-
up study of neurocognitive functioning. 45 [Level of
evidence: 1C]
Patients with CNS 2 status and those who had a traumatic
lumbar puncture with blasts at diagnosis have been found
to be increased risk of CNS relapse ,46 although this risk
appears to be nearly fully abrogated if they receive more
intensive IT chemotherapy, especially during the induction
phase.4, 6,8 [Level of evidence 3B]

Traumatic LP
CNS therapy for high-risk patients
Patients classified as High Risk (NCI) who present with CNS
disease at diagnosis are treated in most protocols with a
combination of Intrathecal chemotherapy and cranial
irradiation (the usual dose being 18 Gy). 25,44

Patients without CNS leukemia:

Patients stratified as high risk who do not have overt CNS
disease at presentation also routinely receive cranial
radiation in most protocols. 2 However, in view of the well
recognized long-term morbidity ,there has been a decrease
in both the proportion of patients receiving radiation and
the dose of radiation.
A reduced dose of prophylactic radiation (12 Gy instead
of 18 Gy) provided effective CNS prophylaxis in high-risk
patients enrolled in the BFM 90 trial. 37 In the BFM 95

80
trial, the proportion of patients receiving cranial radiation
was drastically reduced to 20% of patients (compared with
70% on the previous trial). 44 There was no significant
difference in the overall EFS rate between the cohort of
patients who were not irradiated and the historical controls
who had received irradiation,even though the rate of
isolated CNS relapses was higher in the former group.
[Level of evidence 2B]

Patients with CNS leukemia(CNS III):

Therapy for CNS3 disease at diagnosis typically includes IT
chemotherapy and cranial radiation (usual dose is 18 Gy).
Recent studies such as the SJCRH Total XV study, the Dutch
Childhood Oncology group (DCOG) study and EORTC
58881 study omitted cranial irradiation even in patients
with CNS3 status 40,47. The 5 year EFS and CNS relapse
rates of patients with CNS disease treated with a
combination of intensified intrathecal chemotherapy and
high dose methotrexate was 43% ± 23% & 11% in the
SJCRH study, 67% & 14% in the DCOG study and 68% &
9.3% in EORTC 58881 study respectively. Though these
studies favour avoidance of cranial irradiation, more
evidence is required before a recommendation can be
made in view of the poor EFS, high incidence of CNS
relapses and need for data on long-term neurocognitive
outcomes . [Level of evidence 3B]

References:
1. Simone JV. History of the treatment of childhood ALL:
a paradigm for cancer cure. Best Pract Res Clin
Haematol 2006;19:353–359.
2. Pui CH, Howard SC. Current management and
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paediatric leukaemia. Lancet Oncol 2008;9:257–268.

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3. Mahmoud HH, Rivera GK, Hancock ML, et al. Low
leukocyte counts with blast cells in cerebrospinal fl
uid of children with newly diagnosed acute
lymphoblastic leukemia. N Engl J Med 1993; 329:
314–19.
4. Gajjar A, Harrison PL, Sandlund JT, et al. Traumatic
lumbar puncture at diagnosis adversely aff ects
outcome in childhood acute lymphoblastic leukemia.
Blood 2000; 96: 3381–84.
5. Smith M, Arthur D, Camitta B, Carroll AJ, Crist W,
Gaynon P, et al. Uniform approach to risk
classification and treatment assignment for children
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1996;14:18–24.
6. Burger B, Zimmermann M, Mann G, Kühl J, Löning
L, Riehm H, et al. Diagnostic cerebrospinal fluid
examination in children with acute lymphoblastic
leukemia: significance of low leukocyte counts with
blasts or traumatic lumbar puncture. J Clin Oncol
2003;21:184–188.
7. Gilchrist GS, Tubergen DG, Sather HN, et al. Low
numbers of CSF blasts at diagnosis do not predict
for the development of CNS leukemia in children with
intermediate-risk acute lymphoblastic leukemia: a
Childrens Cancer Group report. J Clin Oncol
1994;12:2594–2600.
8. te Loo DM, Kamps WA, van der Does-van den Berg
A, van Wering ER, de Graaf SS. Prognostic
significance of blasts in the cerebrospinal fluid
without pleiocytosis or a traumatic lumbar puncture
in children with acute lymphoblastic leukemia:
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9. van den Berg H, Vet R, den Ouden E, Behrendt H.
Significance of lymphoblasts in cerebrospinal fluid
in newly diagnosed pediatric acute lymphoblastic
malignancies with bone marrow involvement:
possible benefit of dexamethasone. Med Pediatr
Oncol 1995;25:22–27.
10. Mahmoud HH, Rivera GK, Hancock ML, Krance RA,
Kun LE, Behm FG, et al. Low leukocyte counts with
blast cells in cerebrospinal fluid of children with newly
diagnosed acute lymphoblastic leukemia. N Engl J
Med 1993;329:314–319.
11. Zeiser R, Burger JA, Bley TA, Windfuhr-Blum M,
Schulte-Monting J, Behringer DM. Clinical followup
indicates differential accuracy of magnetic resonance
imaging and immunocytology of the cerebral spinal
fluid for the diagnosis of neoplastic meningitis - a
single centre experience. Br J Haematol
2004;124:762–768.
12. Hegde U, Filie A, Little RF, Janik JE, Grant N, Steinberg
SM, et al. High incidence of occult leptomeningeal
disease detected by flow cytometry in newly
diagnosed aggressive B-cell lymphomas at risk for
central nervous system involvement: the role of flow
cytometry versus cytology. Blood 2005;105:496–502.
13. Conter V, Schrappe M, Arico M, et al. Role of cranial
radiotherapy for childhood T-cell acute lymphoblastic
leukemia with high WBC count and good response
to prednisone. J Clin Oncol 1997; 15: 2786–91.
14. Conter V, Arico M, Valsecchi MG, et al. Long-term
results of the Italian Association of Pediatric
Hematology and Oncology (AIEOP) acute
lymphoblastic leukemia studies, 1982–1995.
Leukemia 2000; 14: 2196–204.

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15. Rocha JC, Cheng C, Liu W, Kishi S, Das S, Cook EH, et
al. Pharmacogenetics of outcome in children with
acute lymphoblastic leukemia. Blood 2005;105:
4752–4758.
16. Krajinovic M, Costea I, Chiasson S. Polymorphism of
the thymidylate synthase gene and outcome of acute
lymphoblastic leukaemia. Lancet 2002;359:1033–
1034.
17. Cario G, Izraeli S, Teichert A, Rhein P, Skokowa J,
Möricke A, et al. High interleukin-15 expression
characterizes childhood acute lymphoblastic
leukemia with involvement of the CNS. J Clin Oncol
2007;25:4813–4820.
18. Wu S, Burmeister T, Baldus C, Schwartz S, Notter M,
Goekbuget N, et al. Low expression of IL-15 in adult
B-lineage acute lymphoblastic leukemia is associated
with central nervous system involvement at initial
presentation. Blood 2007:110. Abstract 2389
19. Bostrom BC, Sensel MR, Sather HN, Gaynon PS, La
MK, Johnston K, et al. Dexamethasone versus
prednisone and daily oral versus weekly intravenous
mercaptopurine for patients with standard-risk acute
lymphoblastic leukemia: a report from the Children’s
Cancer Group. Blood 2003;101:3809– 3817.
20. Mitchell CD, Richards SM, Kinsey SE, Lilleyman J, Vora
A, Eden TO. Benefit of dexamethasone compared
with prednisolone for childhood acute lymphoblastic
leukaemia: results of the UK Medical Research Council
ALL97 randomized trial. Br J Haematol
2005;129:734–745.
21. Meikle AW, Tyler FH. Potency and duration of action
of glucocorticoids. Effects of hydrocortisone,
prednisone and dexamethasone on human pituitary-
adrenal function. Am J Med 1997;63:200–207

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22. Balis FM, Lester CM, Chrousos GP, Heideman RL,
Poplack DG. Differences in cerebrospinal fluid
penetration of corticosteroids: possible relationship
to the prevention of meningeal leukemia. J Clin Oncol
1987;5:202–207.
23. Igarashi S, Manabe A, Ohara A, Kumagai M, Saito T,
Okimoto Y, et al. No advantage of dexamethasone
over prednisolone for the outcome of standard- and
intermediate-risk childhood acute lymphoblastic
leukemia in the Tokyo Children’s Cancer Study Group
L95-14 protocol. J Clin Oncol 2005;23:6489–6498.
24. Clarke M, Gaynon P, Hann I, Harrison G, Masera G,
Peto R, et al. CNS-directed therapy for childhood
acute lymphoblastic leukemia: Childhood ALL
Collaborative Group overview of 43 randomized
trials. J Clin Oncol 2003;21:1798–1809.
25. Moghrabi A, Levy DE, Asselin B, Barr R, Clavell L,
Hurwitz C, et al. Results of the Dana-Farber Cancer
Institute ALL Consortium Protocol 95-01 for children
with acute lymphoblastic leukemia. Blood
2007;109:896–904.
26. Stork LC, Sather H, Hutchinson R. Comparison of
mercaptopurine (mp) with thioguanine (tg) and it
methotrexate with it “triples” (itt) in children with
sr-all: Results of ccg1952. Blood 2002;100:156a.
Abstract 585.
27. Vora A, Mitchell CD, Lennard L, Eden TO, Kinsey SE,
Lilleyman J, et al. Toxicity and efficacy of 6-
thioguanine versus 6-mercaptopurine in childhood
lymphoblastic leukaemia: a randomised trial. Lancet
2006;368:1339–1348.
28. Jacobs SS, Stork LC, Bostrom BC, Hutchinson R,
Holcenberg J, Reaman GH, et al. Children’s Oncology

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 Group; National Cancer Institute. Substitution of oral
and intravenous thioguanine for mercaptopurine in
a treatment regimen for children with standard risk
acute lymphoblastic leukemia: a collaborative
Children’s Oncology Group/National Cancer Institute
pilot trial (CCG-1942). Pediatr Blood Cancer
2007;49:250–255.
29. Pullen J, Boyett J, Shuster J, et al.: Extended triple
intrathecal chemotherapy trial for prevention of CNS
relapse in good-risk and poor-risk patients with B-
progenitor acute lymphoblastic leukemia: a Pediatric
Oncology Group study. J Clin Oncol 11 (5): 839-49,
1993.
30. Thyss A, Suciu S, Bertrand Y, et al.: Systemic effect of
intrathecal methotrexate during the initial phase of
treatment of childhood acute lymphoblastic
leukemia. The European Organization for Research
and Treatment of Cancer Children’s Leukemia
Cooperative Group. J Clin Oncol 15 (5): 1824-30,
1997.
31. Matloub Y, Lindemulder S, Gaynon PS, Sather H, La
M, Broxson E, et al. Children’s Oncology Group.
Intrathecal triple therapy decreases central nervous
system relapse but fails to improve eventfree survival
when compared with intrathecal methotrexate:
results of the Children’s Cancer Group (CCG) 1952
study for standard-risk acute lymphoblastic leukemia,
reported by the Children’s Oncology Group. Blood
2006;108:1165–1173.
32. Blaney SM, Poplack DG, Godwin K, McCully CL,
Murphy R, Balis FM. Effect of body position on
ventricular CSF methotrexate concentration following
intralumbar administration. J Clin Oncol
1995;13:177–179.

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33. Stam J. Thrombosis of the cerebral veins and sinuses.
N Engl J Med 2005;352:1791–1798.
34. Baxter AL, Fisher RG, Burke BL, Goldblatt SS, Isaacman
DJ, Lawson ML. Local anesthetic and stylet styles:
factors associated with resident lumbar puncture
success. Pediatrics 2006;117:876–881
35. Pui C-H, Cheng C, Leung W, et al. Extended follow-
up of long-term survivors of childhood acute
lymphoblastic leukemia. N Engl J Med 2003; 349:
640–49.
36. Hijiya N, Hudson MM, Lensing S, et al. Cumulative
incidence of secondary neoplasms as a fi rst event
after childhood acute lymphoblastic leukemia. JAMA
2007; 297: 1207–15.
37. Schrappe M, Reiter A, Ludwig WD, Harbott J,
Zimmermann M, Hiddemann W, et al. Improved
outcome in childhood acute lymphoblastic leukemia
despite reduced use of anthracyclines and cranial
radiotherapy: results of trial ALL-BFM 90. German-
Austrian-Swiss ALL-BFM Study Group. Blood
2000;95:3310–3322. [PubMed: 10828010]
38. Pui CH, Relling MV, Sandlund JT, Downing JR,
Campana D, Evans WE. Rationale and design of Total
Therapy Study XV for newly diagnosed childhood
acute lymphoblastic leukemia. Ann Hematol
2004;83(Suppl 1):S124–S126
39. Ching-Hon Pui Improved Treatment of Childhood
Acute Lymphoblastic Leukemia Revista de
Hematología Vol. 11, Supl. 1, April-May 2010; p.
42
40. Veerman AJ, Kamps WA, van den Berg H, et al.:
Dexamethasone-based therapy for childhood acute
lymphoblastic leukaemia: results of the prospective

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 Dutch Childhood Oncology Group (DCOG) protocol
ALL-9 (1997-2004). Lancet Oncol 10 (10): 957-66,
2009.
41. Pui CH, Sandlund JT, Pei D, et al.: Improved outcome
for children with acute lymphoblastic leukemia:
results of Total Therapy Study XIIIB at St Jude
Children’s Research Hospital. Blood 104 (9): 2690-
6, 2004.
42. Tubergen DG, Gilchrist GS, O’Brien RT, et al.:
Prevention of CNS disease in intermediate-risk acute
lymphoblastic leukemia: comparison of cranial
radiation and intrathecal methotrexate and the
importance of systemic therapy: a Childrens Cancer
Group report. J Clin Oncol 11 (3): 520-6, 1993.
43. Conter V, Aricò M, Valsecchi MG, et al.: Extended
intrathecal methotrexate may replace cranial
irradiation for prevention of CNS relapse in children
with intermediate-risk acute lymphoblastic leukemia
treated with Berlin-Frankfurt-Münster-based intensive
chemotherapy. The Associazione Italiana di
Ematologia ed Oncologia Pediatrica. J Clin Oncol 13
(10): 2497-502, 1995.
44. Möricke A, Reiter A, Zimmermann M, et al.: Risk-
adjusted therapy of acute lymphoblastic leukemia
can decrease treatment burden and improve survival:
treatment results of 2169 unselected pediatric and
adolescent patients enrolled in the trial ALL-BFM 95.
Blood 111 (9): 4477-89, 2008.
45. Kadan-Lottick NS, Brouwers P, Breiger D, et al.:
Comparison of neurocognitive functioning in
children previously randomly assigned to intrathecal
methotrexate compared with triple intrathecal
therapy for the treatment of childhood acute

88
 lymphoblastic leukemia. J Clin Oncol 27 (35): 5986-
92, 2009.
46. Mahmoud HH, Rivera GK, Hancock ML, et al.: Low
leukocyte counts with blast cells in cerebrospinal fluid
of children with newly diagnosed acute
lymphoblastic leukemia. N Engl J Med 329 (5): 314-
9, 1993.
47. Pui CH, Campana D, Pei D, et al.: Treating childhood
acute lymphoblastic leukemia without cranial
irradiation. N Engl J Med 360 (26): 2730-41, 2009.
48. Sirvent N, Suciu S, Rialland X, et al. Prognostic
significance of the initial cerebro-spinal fluid (CSF)
involvement of children with acute lymphoblastic
leukaemia (ALL) treated without cranial irradiation:
Results of European Organization for Research and
Treatment of Cancer (EORTC) Children Leukemia
Group study 58881. Eur J Cancer. 2011
Jan;47(2):239-47.

89
 Biology and Evidence Based
Management of Relapsed Pediatric
Acute Lymphoblastic Leukemia
Dr. Shripad D. Banavali; Dr. Bhausaheb Bagal

Introduction:
Acute Lymphoblastic Leukemia (ALL) is the commonest
childhood cancer and despite dramatic improvement in
therapy nearly 25% of children will fail therapy making
relapsed ALL the fourth commonest childhood cancer. 1-3
In fact, relapsed ALL is the second commonest leukemia
after ALL. Although survival of children with ALL has
improved greatly in the past two decades, the outcome
of those who relapse has remained static4, 5.
Management of marrow relapse has not been as successful
as primary treatment despite intensified chemotherapy and
use of haematopoietic stem cell transplantation (HSCT),
suggesting the need for better therapeutic options. The
overall survival of relapsed ALL has remained between 30% and
60% in recent years4. Today few recurrences occur solely
in the CNS or other extra-medullary sites. However most
of these extra-medullary relapses are curable, especially
those that happen well after initial treatment6–8.
90
Origin and biology of relapse:
Retrospective comparison of gene sequences from
specimens at diagnosis, remission, and relapse reveal that
the relapsing clone is present in the initial specimen, often
beneath the level of detection.9, 10 After induction, this
clone persists in greater copy numbers than others,
probably indicating its resistance to induction therapy.9,11
Although the relapsing clone typically becomes
undetectable after post induction therapy, it eventually
reappears as minimal residual disease and subsequently
recurrence. Hence, early relapse probably results from
selection of a relatively resistant clone present at initial
diagnosis rather than generation of a new clone by
mutation (figure 1).
By contrast, late relapses may represent de novo
development of a second leukaemia from a common
premalignant clone. Studies of cord blood have established
that the ETV6–RUNX1 (TEL–AML1) translocation occurs
in utero, but is not sufficient to cause leukaemia. A second
mutation, most often a deletion of the normal ETV6 allele,
is required for leukaemogenesis.12 Each patient’s unique
translocation breakpoint sequence is a marker for the pre-
leukemic clone that develops into ALL. Comparisons of
matched initial and relapse specimens document identical
ETV6–RUNX1 breakpoint sequences and typical shifts in
immunoglobulin and T-cell receptor rearrangements, but
different mutations in the un-rearranged ETV6 allele.
Therefore, two separate episodes of leukaemia must have
arisen from metachronous independent mutations in a
common ETV6–RUNX1 pre-leukemic cell. Perhaps this
explains why the second leukaemia is commonly cured by
therapy similar to that used to treat initial disease (figure
1).12

91
Recent data has also shown that the gene expression is
different in early and late relapses13. In early relapses, there
is usually up-regulation of genes that are related to cell
proliferation, whereas cells in late relapses have more
diverse pathways.13, 14 These relapsed ALL cells develop
anti-cancer drug resistance which can be either drug
specific or have multi-drug resistance15. Gene expression
studies have shown a link between drug resistance and
the microenvironment that has previously only been
considered in context of solid tumor biology14.

Figure 1: Potential mechanisms of clinical relapse

Top: Early recurrence may arise due to selection of resistant
leukemic cells during first-line therapy. Bottom: Late relapse
may represent a second, independent transforming event
in an underlying pre-leukemic clone. (Adapted from L
Charles Bailey, Lancet Oncol 2008; 9: 873–83)
By use of the 3-[4,5-dimethyl-thiazole-2,5-diphenyl]
tetrazolium bromide (MTT) cytotoxicity assay, both case–
control and matched-set comparisons of initial and relapse
samples showed that relapse samples are more resistant
to glucocorticoids, L-asparaginase, thiopurines, and
anthracyclines in-vitro - these drugs are central
components of initial treatment. However, the samples

92
retain sensitivity to etoposide and cyclophosphamide,
which are sparingly used in first-line protocols.16 These
findings suggests that patients might benefit after relapse
from different drugs or dosing strategies, but does not
distinguish between acquired and inherent drug resistance
in leukemic blasts.

Simple Ways to prevent relapse in the

Indian Scenario:-
Though more than 75% of children with ALL are cured in
the high income countries (HIC), the results are much
inferior in low and middle income countries (LAMIC).17, 18
As opposed to the HIC, where relapses occur in a much
smaller percentage of children with ALL and mostly
because of biological reasons, relapses in LAMIC occur in
a much higher percentage of children and in many cases
because of non-biological reasons. Given the dismal
outcome of relapsed ALL, it would be appropriate to
discuss regarding ways to prevent relapse before going to
management of relapses per se.
1) If possible, use risk-adapted therapy with appropriate
protocols that best suit the biology of patients seen
at your centre. Do not keep on changing or modifying
treatment protocols at short intervals. This way you
will get acclimatized with the protocol and start
anticipating problems.
2) Induction phase is one the most important phase of
treatment, hence during Induction continue
chemotherapy irrespective of blood counts. Hold
chemotherapy only if patient is unstable.
3) During Maintenance phase titrate the dose of 6MP/
Methotrexate to keep ANC between 1000 and 2000/
cmm. It is better to give lower but continuous doses
of 6MP / MTX, than higher but intermittent doses

93
 with many breaks. 6-MP is best given at bedtime, 2
hours after dinner with water only. Maintenance is
to be given for two years in patients with B-lineage
ALL and for one and a half years for patients with T-
lineage ALL.
4) Maintain dose-intensity during the entire therapy,
during both induction and maintenance phase.
5) Keep a watch on drug interactions as they have a
potential to cause altered levels of chemotherapeutic
agents leading to un-favorable outcomes. For
example voriconazole, phenytoin sodium, 6MP and
food/milk.
6) There are many generics available in India. Use
standard chemotherapeutic drugs, especially original
molecules where ever possible.
7) Check and maintain required temperature for storage
of different drugs especially L-asparaginase,
vincristine.

Diagnosing relapse of ALL:

Recent data has shown lack of benefit of early detection
of relapse after completion of therapy for ALL and thus it
is no longer recommended to do BM Aspiration studies
to rule out relapse in patients with ALL who have achieved
remission, unless patient is symptomatic or the CBC is
abnormal. 19 Suspected patients should undergo bone
marrow aspiration with morphology, surface marker
studies and cytogenetic studies. Examination of extra-
medullary sites like CNS, testis, eyes, and skin should be
done at the time of diagnosis of relapse.

Risk stratification of relapsed ALL:

It is important to have risk stratification of relapse for
counselling and tailoring the salvage treatment.

94
1) Length of 1st CR: - Patients with very early (< 18
months from start of therapy) BM relapse have <
10% long term survival (LTS) compared to
approximately 50% LTS for patients who have late
BM relapse. The outcome is usually very poor for
patients with second or subsequent relapse.
2) Immunophenotype: - Childhood ALL patients with
T-cell ALL fare more poorly than those with CALLA+
ALL.
3) Site of relapse:-Historically, isolated marrow relapse
has had the worst prognosis; isolated CNS, testicular,
or other extra-medullary relapse carried a significantly
better prognosis, and combined marrow and extra-
medullary relapse, an intermediate prognosis. 4
Greater cure rates for combined relapse compared
with isolated marrow relapse might reflect differences
in disease biology, but might also result from
therapeutic craniospinal-irradiation.
4) The nature and intensity of previous therapy: -
Patients previously treated with lower intensity
primary therapy have a higher re-induction rate.
Similarly, patients with CNS relapses who have not
previously received Cranial Radiotherapy do better.
5) Minimal Residual Studies (MRD) Studies:- MRD is an
important tool even for patients with ALL at time of
relapse. The level of minimal residual disease after
achieving second remission or before transplant may
predict outcomes.5 Recent data has shown that
patients who are MRD negative at end of re-induction
have a relapse rate of 40% as compared to patients
who are MRD positive at end of re-induction, who
have a relapse rate of 70% 20

95
 Figure 2: Risk Stratification on UK ALL R3 Protocol4
Stratification according to immunophenotype, site of
relapse, and time to relapse. Risk groups: very early refers
to less than 18 months from first diagnosis; early refers to
18 months or more from first diagnosis and less than 6
months from stopping therapy; and late refers to 6 months
or more from stopping therapy (Adapted from Parker C,
et al. Lancet 2010; 376: 2009–17)

Treatment of marrow relapse:

Re-induction therapies after marrow relapse:
Typical treatment of first relapse involves a combination
of vincristine, a glucocorticoid (prednisone, prednisolone,
or dexamethasone), and asparaginase, plus an
anthracycline, methotrexate, or cytarabine in varying doses
and schedules. Only a few randomised trials have
investigated re-induction therapy.4, 21-23
Asparginase: Aspargine depletion is important in achieving
remission. A Paediatric Oncology Group study that showed
significantly higher re-induction rates with weekly rather
than biweekly peg-asparginase (97% vs. 82%), leading to
near universal adoption of the weekly schedule.21
Vinca-Alkaloids: Vincristine has compared to vindesine as
the re-induction vinca alkaloid in a Paediatric Oncology
Group. Outcomes were similar, but vindesine was more
96
toxic and was abandoned in the USA23 though not in BFM
group trials.
Methotrexate: Over a series of trials, the BFM group
randomised dose and duration of infusional methotrexate
in re-induction and thereafter. The group found no
difference in re-induction rate or long-term outcome
between intermediate-dose (1 g/m² over 36 h) and high
dose (5G/m² over 24h) infusions, and have adopted the
former for future trials.24, 25
Anthracycline: Institutional protocols from Philadelphia
and Seattle use idarubicin (30 mg/m² total dose) as part
of a four-drug re-induction with apparent success26 and a
trial by the Italian Paediatric Haematology Oncology
Association (AIEOP) showed efficacy in combination with
high-dose cytarabine.27 However, the Children’s Cancer
Group randomised two doses of idarubicin (30 mg/m² or
37·5 mg/m² total dose) to daunorubicin (135 mg/m² total
dose) but could not prove superiority of idarubicin.
Mitoxantrone: Unlike idarubicin, mitoxantrone is an
anthracenadione. Idarubicin is a topoisomerase IIá poison
and targets cycling cells, while mitoxantrone inhibits
topoisomerase-IIâ in addition to topoisomerase IIá and
hence has activity against quiescent cells as well. It also
has the ability to create DNA adducts, stimulate binding
of nuclear factor-êâ (Kappa-beta), and potentiate immune-
based cell kill by tagging leukaemia cells with calcireticulin.
ALL R3 protocol compared mitoxantrone to idarubicin.
Mitoxantrone conferred a significant benefit in
progression-free and overall survival 4. Two additional
interesting concepts about mitoxantrone emerged in this
study. First, study of MRD in two arms suggested that
mitoxantrone seems to have a delayed cytotoxic effect.
Second, for the few patients who were not transplanted

97
and continued on chemotherapy, haematological toxic
effects increased during the later phases in the
mitoxantrone group. Thus, mitoxantrone might in some
way affect the haematopoietic stem-cell niche, making it
a less favourable environment for the leukemic cell and
more conducive to the allograft.4
Across multiple studies, second remission is achieved in
more than 70% of early relapses and up to 96% of late
relapses.21, 22 In the absence of randomised trials and
consistent risk-stratified reporting of patient’s outcomes,
with possibly exception of mitoxantrone, no re-induction
combination is significantly superior to the others. It is
noted that especially in the HIC, most children with
relapsed ALL who show no remission to treatment
according to relapsed protocol, are still treated with
curative intent, including HSCT. However recent analysis
of BFM data has shown that this approach is not successful
and was associated with high treatment related morbidity,
mortality, and minimal survival. No response patients , may
therefore, be ideal candidates for controlled Phase I / II
trials, thus offering a chance to benefit from new drugs
and promoting drug development for cohorts with better
prognosis.28

Post-remission therapy:
Therapeutic options after CR2 include further
chemotherapy and HSCT. Most pursue HSCT options for
patients with early relapses, although outcomes remain
poor for most patients with measurable MRD after re-
induction. Similar outcomes are reported for matched
related donor and matched unrelated donor transplants.
For late marrow relapse, outcomes are similar with
chemotherapy and HSCT options. Some recommend HSCT
options for patients with late marrow relapse and an MRD-

98
positive CR2. Chemotherapy options may be pursued for
isolated extra-medullary relapse with success in most
patients.

Continuation chemotherapy:
All patients who achieve a second remission receive
additional chemotherapy, even if haematopoietic stem cell
transplantation is planned. To maintain control of disease,
higher dose intensity is used, and higher regimen-related
toxicity is tolerated than in first-line treatment.
Most reports describe single-arm studies with
combinations of vincristine, glucocorticoids, methotrexate,
cytarabine, etoposide, cyclophosphamide or ifosfamide,
and thiopurines, with or without maintenance therapy
for up to 2 years.3,4,25,29 CNS prophylaxis includes high-
dose methotrexate or cytarabine, intrathecal
chemotherapy, and in more recent BFM group trials, 1200–
1800 cGy cranial irradiation.25
The Paediatric Oncology Group study 8303 reported that
the addition of four-drug re-induction pulses did not
improve upon weekly rotation of cytarabine and teniposide
with vincristine and cyclophosphamide.30 BFM group
relapse trials randomised patients with first remission
greater than 18 months to high-dose or long-duration
infusions of intermediate dose methotrexate with blocks
of multidrug chemotherapy,25,29 with equivalent outcomes.
Aside from these randomised trials, published data do
not show one chemotherapy combination to be better
than others.

Haematopoietic stem cell transplantation

(HSCT):
The only strategy that has shown to improve outcome of
patients with relapsed ALL with high risk features is to
99
give intensified consolidation chemotherapy regimen,
followed by HSCT. Recent data has shown that outcome
of HSCT is better than chemotherapy alone even in patients
with intermediate risk relapsed ALL and HSCT is currently
being offered to these patients if they have an HLA
matched sibling donor. However, only 25–30% of potential
candidates for HSCT have an HLA matched familial donor.
Though unrelated donor registries and cord blood banks
have increased the donor pool, it is still not practical to
do these transplants in the Indian settings. There is no
role of autologus transplant in children with relapsed ALL.
Conditioning regimens: Published research on
transplantation of haematopoietic stem cells includes a
mixture of conditioning regimens. Cytoreduction that
includes whole-body irradiation achieves results superior
to chemotherapy alone. 31- 33 Delaying transplantation until
only minimal residual disease remains is beneficial.5, 34, 35
Results of Allogenic transplant: The Paediatric Oncology
Group study 8303 (POG-8303) of early marrow relapse
showed that leukaemia-free survival was 7% compared
with 26% in tracked patients selected for chemotherapy
or haematopoietic stem cell transplantation, respectively,
but the difference was not significant. An International
Bone Marrow Transplant Registry (IBMTR) and Paediatric
Oncology Group case–control comparison of patients in
first marrow relapse, including patients from POG-8303,
found leukaemia-free survival in patients receiving stem
cells from matched familial donors was significantly higher
than in those on chemotherapy alone despite higher
transplant-related mortality and regardless of first
remission duration. In more recent IBMTR–POG datasets,
leukaemia-free survival was significantly higher only in early
marrow relapse and only with conditioning including total
body irradiation (TBI).36
100
In the BFM ALL-REZ-87 trial, in patients with high-risk or
intermediate-risk marrow relapse, leukaemia-free survival
from the time of second remission was significantly higher
in the group receiving haematopoietic stem cells than in
the chemotherapy group (p=0·026). In this trial 26% of
patients with very early relapse, but only 13% of those
with early relapse had stem cell transplantation.29 Results
of autologous transplantation and chemotherapy were
the same. Matched-pair comparisons across BFM group
ALL-REZ trials showed that unrelated donor transplantation
achieved significantly better leukaemia-free survival than
chemotherapy in high-risk relapse but not in intermediate-
risk relapse.37 In the BFM group ALL-REZ- 90 trial, outcomes
for chemotherapy and trans plantation of haematopoietic
stem cells in patients with intermediate risk marrow relapse
are reported as “not different”;25 the authors conclude
that they could not assess the role of transplantation “due
to its rather uncontrolled use”.
On the basis of per protocol analysis, the Children’s
Oncology Group now recommends transplantation of
allogenic haematopoietic stem cells from best possible
donor in all patients with early relapse. For later relapse, a
current Children’s Oncology Group study is randomising
between different vincristine doses during chemotherapy,
and recommends matched familial donor haematopoietic
stem-cell transplantation as definitive therapy where
possible.
Further relapse is the greatest barrier to cure after first
relapse. Best possible allogenic transplant offers a better
chance of cure than chemotherapy for patients who have
not had haematopoietic stem cell transplantation in the
recent past.38 Although some studies report occasional
long-term third remissions in patients at intermediate and

101
standard risk after second relapse,25 cure is unlikely unless
the patients have favourable features at initial diagnosis
and first relapse.38

Central Nervous System Relapse:

Intensive treatment plans have improved the results for
patients with an isolated CNS relapse as high as 70% EFS.39
The main factors determining the success rate for patients
with CNS relapse include whether the relapse occurred
more than or less than 18 months (83% and 46% EFS
respectively) from the initial diagnosis and whether the
patient received CNS directed irradiation during the initial
treatment regimen.39, 40 Several attempts have been made
to improve on the use of single-agent IT MTX for CNS
relapse. Triple therapy consisting of simultaneously
administered IT cytarabine, hydrocortisone, and MTX has
been advocated, but this produces remission durations
similar to those achieved with MTX alone.41 The role of
craniospinal irradiation to treat CNS recurrence in a patient
who originally received cranial irradiation as part of CNS
preventive therapy is less certain. Results from the MRC
Concord and UKALL-I trials demonstrated a continuous
complete remission rate of less than 10% for these
patients. Other investigators have demonstrated
substantially better results with this approach. However,
craniospinal irradiation administered in this setting is
known to pose a significantly greater risk of delayed
neurotoxicity, which must be weighed against the risk of
progressive disease. A common approach is first to induce
a CSF remission with IT chemotherapy, reinstitute systemic
therapy, and then later administer craniospinal irradiation,
at doses of 2,400 to 3,000 cGy to the cranial vault and
1,200 to 1,800 cGy to the spinal axis.42

102
Isolated Testicular Relapse:
Isolated testicular relapse, again, is very rare with modern
aggressive protocols. It was <1% in St. Jude TXIII protocol
and 0 % on the St. Jude TXIV & TXV protocols. Again
testicular relapse is considered a systemic disease. Present
issues include whether RT is necessary (considering the
late effect on fertility) and what is the role of orchidectomy?
Since testicular RT of >24 Gy causes sterility and endocrine
problems, the recent principles of management include
orchidectomy of the clinically involved testis with biopsy
of the contra-lateral testis. If biopsy is negative, RT is
reduced to 15 Gy. A Dutch study, though included small
number of patients, had shown that one can totally avoid
RT if patients are given very HD MTX (12 G/ m2) and all
patients did well43. We have also recently shown good
outcome of patients with testicular relapse.44

Improving Therapy of relapse:

Novel therapeutic agents are needed to improve the cure
rate of patients with relapsed or refractory ALL as both
current chemotherapy and transplantation of
haematopoietic stem cells are toxic, and neither is highly
effective in curing most patients with relapse. New drugs
in development/trial are listed below.
In addition to above, one promising treatment for CD 19
+ relapsed ALLs is blinatumomab (MT103), a first antibody,
which can potentially engage all cytotoxic T cells of a
patient for tumor cell lysis.45 A perception that optimization
has been reached with available drugs has shifted focus
towards newer drugs and targeted therapy. These drugs
are prohibitively expensive for most patients. Thus, while
we wait for targeted therapies to become a reality,
conventional cytotoxics like mitoxantrone still has a role
in treatment of ALL.4

103
Summary:
Because the overall cure rates for recurrent ALL remain
poor with contemporary intensified approaches, new
treatment strategies beyond intensive chemotherapy and
HSCT are needed. These include creation of trials that
rapidly identify promising new agents and
immunotherapeutic interventions. However considering
the efforts involved and the poor outcome, it can be safely
concluded that in childhood ALL, as in all other diseases,
prevention of relapse is definitely much better than the
treatment of relapse. Systematic assessment of biologic
features at the time of diagnosis that correlate with risk
for relapse will help in this regard.46 Great emphasis is
being placed on developing a better understanding of
mechanisms of relapse and drug resistance through the
implementation of genomic and proteomic studies in an
effort to define pathways for therapeutic modulation and
developing tools to assess the activity of new treatment
strategies .

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110
 Management of Adult
Acute Lymphoblastic Leukemia
Dr Manju Sengar, Dr Jayant Gawade

The progressive improvement in outcomes of childhood

acute lymphoblastic leukemia have become a success story
in field of oncology however Adult ALL still remains a
challenge with only 25% to 50% of adults achieving long-
term disease-free survival. ALL accounts for 1% of all adult
cancers. ALL is a heterogeneous disease both in terms of
its biology and the population it affects. Definition of the
age range that encompasses the adult patient is itself
controversial, because the age range described in the
literature for this population varies from 18 to 21 years
depending on the study. Patients between 16 to 21 years
comprising of adolescents and young adults are treated
with both pediatric and adult protocols based on their
referral.

Prognostic Factors and Risk Assessment

Several variables can affect the prognosis of adult ALL
patients and suggest the need for treatment
intensification.

111
Clinical: Advancing age adversely affects the outcome.
This effect may be mediated through the adverse biology
(high incidence of Philadelphia chromosome positive ALL),
comorbidities and poor organ reserve leading to poor
treatment tolerance. White blood cell count at
presentation impacts the outcome of ALL at all ages. WBC
>30,000/cumm in case of B-cell and >100,000/cumm
for T-cell is considered as poor risk. Level of evidence III
Immunophenotype: T cell phenotype carries a better
prognosis than B cell as demonstrated by three large
international trials. Level of evidence II
CNS disease at diagnosis: It confers poor overall survival
as demonstrated in UKALLXII/ECOG 2993 study. Level of
evidence III
Cytogenetics at diagnosis: Presence of t(9;22) –
Philadelphia positive ALL, t(4;11), complex karyotype,
hypodiploidy/near triploidy confers poor prognosis. Level
of evidence I
Time to achieve complete response: Patients who
achieve complete remission (CR) at the end of induction
fare better in term of their disease free survival. Level of
evidence III
Minimal residual disease: Treatment response is being
increasingly evaluated with minimal residual disease (MRD)
assays. Level of evidence I. Being a measurement of drug
resistance in vivo and reflecting multiple cellular, host, and
treatment variables, MRD is typically an independent
prognostic factor. There is solid evidence indicating that
MRD testing is clinically useful in adult ALL. Investigators
of the GMALL studied MRD in 196 patients defined as
standard risk by the absence of MLL-AFF1 or BCR-ABL1

112
gene fusion, B-lineage ALL with leukocyte counts less than
30 x 109/L or T ALL with counts less than 100 x 109/L, age
15 to 65 years and achievement of CR after first phase of
induction therapy. They identified a low risk group (10%)
with MRD lower than 0.01% on days 11 and 24 and a 3
year relapse rate of 0%; a high risk group (23%) with MRD
0.01% or more until week 16 and a relapse rate of 94%;
and an intermediate risk group (67%) with a relapse rate
of 47%. This team also studied post consolidation samples
in 105 patients. At the time of analysis relapse had
occurred in only 5 of the 77 patients who remained MRD
–ve, compared with 17 relapses observed among 28
patients who converted to MRD positivity.
More recently, pharmacogenetics (polymorphisms of
various enzymes involved in drug metabolism) application
of microaray-based, genome-wide analysis of gene
expression and DNA copy number have been found to
have biologic and therapeutic implications.

Treatment strategies for ALL

Treatment for adult ALL is modeled on therapy developed
for childhood ALL. Multiple drugs are molded into
regimen-specific sequences of dose and time intensity with
the goal of (1) reconstituting normal hematopoiesis; (2)
preventing emergence of resistant subclones; (3) providing
adequate prophylaxis of sanctuary sites (eg, CNS, testicles);
and (4) eliminating minimal residual disease (MRD)
through postremission consolidation and maintenance.
It consists of remission induction, consolidation,
maintenance therapy, and CNS prophylaxis, using a risk-
stratified approach. As the management and outcome
results significantly vary between Philadelphia (Ph) positive
and negative ALL further discussion on treatment will focus
on these entities separately.

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Induction
Ph-ALL
The goal of induction chemotherapy is to eradicate 99%
of the initial tumor burden and to restore normal
hematopoiesis. Combination of corticosteroid, vincristine,
daunorubicin, and asparaginase is the most accepted
regimen currently.Level of evidence III. Some protocols do
include cyclophosphamide and ara-C in the later part.This
regimen results in CR rates of 85-95% with induction
treatment related mortality of 5-10%. Apart from
infections, hepatic toxicity, thrombotic events and bleeding
manifestations are common complications in this patient
group. Dexamethasone is preferred to prednisone in
pediatric population because of superior
lymphocytotoxicity, better CNS penetration and fewer
thromboembolic events, however information in Adult ALL
is limited.Level of evidence II There is no good evidence
on the use, dose and frequency of L-asparaginase in adult
patients. However with increasing age its use is
compromised due to high incidence of toxicities- hepatic,
thrombosis and hyperglycemia. Peg-asparaginase may
cause more effective asparagine depletion resulting in
better outcomes. Level of evidence II. Finally, supportive
care and management of complications is of great
importance during this period. The use of growth factors
lessens the regimen-induced myelosuppression and may
allow timely administration of treatment. In a randomized
trial, the use of G-CSF during induction was associated
with faster recovery of neutrophils and decreased hospital
stay. In the G-CSF treated group, the complete remission
rate was higher (90% vs. 81%; P = .10), and the rate of
induction deaths was lower (4% vs. 11%; P = .04).Level
of evidence II

114
Ph+ ALL
Introduction of imatinib has changed the outcome of
Ph+ALL over the years. Nearly all the studies have
documented improvement in complete response rates,
recurrence free survival and overall survival with imatinib
as compared to historical controls. It is better to consider
concurrent administration of imatinib and chemotherapy
as opposed to sequential use. As these patients are at
very high-risk of relapse allogeneic stem cell transplant
should be considered in CR1 if donor is available. Level of
evidence I
Dasatinib along with prednisolone has been shown to
effective in inducing CR rates >90% in elderly Ph+ALL.
Level III.

Consolidation
It is used with intention of clearing any drug resistant
leukemia cells that have survived induction therapy and
to eliminate MRD. Little of this therapy is evidence based
in adults. Most of the data is extrapolated from the
pediatric trials. Consolidation regimens include
methotrexate, cytarabine, cyclophosphamide, and
asparaginase. Role of allogeneic stem cell transplant as
consolidation has been discussed elsewhere in this book.
Novel strategies try to emphasize subtype-oriented and
risk-oriented approaches of consolidation programs. The
German Multicenter Study Group for Adult ALL05/93
intensified consolidation in a subtype-specific manner:
high-dose methotrexate in standard-risk B-cell lineage ALL;
cyclophosphamide and cytarabine in T-cell lineage ALL;
and high-dose methotrexate and high-dose cytarabine in
high-risk B-cell lineage ALL. Induction was intensified with
high-dose cytarabine (3 g/m2, 4 doses) and mitoxantrone.

115
The CR rate was 87% in standard-risk patients, with
median remission duration of 57 months and a 5-year
survival probability of 55%. Intensified induction and
consolidation did not improve CR rate and DFS in high-
risk patients, except in patients with pro–B-cell ALL, in
whom a continuous CR rate of 41% was achieved
compared with 19% in other high-risk patients.
In CALGB study 8811, the induction course consisting of
a five drug combination was followed by early and late
intensification courses with eight drugs. This regimen
improved the median duration of complete remission and
median survival to 29 and 36 months, respectively,
considerably better than results with earlier trials. In the
Medical Research Council Adult Acute Lymphoblastic
Leukaemia Trial UKALL XA (MRC UKALL XA), patients were
randomized to receive early intensification at 5 weeks, late
intensification at 20 weeks, both treatments, or neither.67
The early block of intensive treatment prevented relapses,
although DFS at 5 years was increased only slightly.

Maintenance
It is designed to prevent leukemia relapse. Maintenance
therapy is manadatory for patient not undergoing
allogeneic stem cell transplant. The randomized controlled
data from UKALLXII/ECOG2993 has proven the benefit of
prolonged chemotherapy over autologous stem cell
transplant. It consists of a backbone of daily 6-
mercaptopurine, weekly methotrexate, and monthly pulses
of vincristine and prednisone, generally administered for
24 months .Level of evidence II. Attempts to omit
maintenance or shorten its duration to 12 to 18 months
have led to inferior results. However, maintenance therapy
is not necessary in mature B ALL, in which a high cure rate

116
is achieved with short-term, dose-intense regimens. The
best maintenance regimen for Ph+ patients is not clear,
but should include a TK inhibitor.

Central nervous system prophylaxis

Risk factors for CNS disease include elevated WBC count
or lactate dehydrogenase at diagnosis, traumatic lumbar
puncture, and mature B cell ALL or T-lineage ALL
phenotypes. CNS involvement occurs in 5% of adults at
diagnosis and although it impacts on survival (29% vs 38%;
P= 0.03) is not clear. It is unclear how much prophylactic
CNS directed therapy is required. Intrathecal chemotherapy
and high dose methotrexate seem to be adequate to
prevent CNS relapse. Role of cranial radiation in CNS
prophylaxis is now difficult to judge due to better systemic
and intrathecal therapy. For patients proceeding to TBI
containing allograft regimens intrathecal chemotherapy
alone is sufficient. More prophylaxis may be required for
RIC regimens. For patients with CNS involvement there is
a suggestion that allografting may result in superior
outcomes. Level of evidence III

Treatment of specific subgroups

Mature B cell ALL
Owing to its high growth fraction Burkitt’s leukemia
requires short duration intensive regimens for better
outcomes. Furthermore addition of rituximab to
chemotherapy significantly improves outcome. Thomas et
al. administered rituximab in addition to the hyper-CVAD
regimen and reported a complete remission rate of 86%,
with 3-year overall survival of 89%. This was significantly
better than the 19% 3-year survival reported for hyper-
CVAD alone.

117
T cell ALL and T-Lymphoblastic lymphoma
The survival of these patients has improved significantly
using the regimens designed for ALL, with overall survival
ranging from 50% to 70%. Use of mediastinal radiation
given after chemotherapy appears to reduce mediastinal
relapse. Greater understanding of the molecular
pathogenesis of this subset of ALL has led to the recent
development of novel therapies, such as nelarabine and
forodesine, developed specifically toward neoplastic T cells,
and gamma-secretase and TK inhibitors developed
specifically toward aberrant pathways. Activity for these
new agents is being investigated in phase I and II trials.

References
Larson RA, Dodge RK, Burns CP, et al. A five-drug remission
induction regimen with intensive consolidation for adults
with acute lymphoblastic leukemia: cancer and leukemia
group B study 8811. Blood 1995;85:2025.
Kantarjian H, Thomas D, O’Brien S, et al. Long-term follow-
up results of hyperfractionated cyclophosphamide,
vincristine, doxorubicin, and dexamethasone (Hyper-
CVAD), a dose-intensive regimen, in adult acute
lymphocytic leukemia. Cancer 2004;101:2788.
Kantarjian HM, Walters RS, Keating MJ, et al. Experience
with vincristine, doxorubicin, and dexamethasone (VAD)
chemotherapy in adults with refractory acute lymphocytic
leukemia. Cancer 1989;64:16.
Gokbuget N, AR, Buechner T, et al. Intensification of
induction and consolidation improves only subgroups of
adult ALL: analysis of 1200 patients in the GMALL study
05/93 American Society of Hematology; 2001.

118
Rowe JM BG, Fielding A, Tallman MS, et al. In adults with
standard-risk acute lymphoblastic leukemia (ALL) the
greatest benefit is achieved from an allogeneic transplant
in first complete remission (CR) and an autologous
transplant is less effective than conventional consolidation/
maintenance chemotherapy: final results of the
international ALL trial (MRC UKALL XII/ECOG E2993).
Presented at: American Society of Hematology, San Diego,
California, 2006.
Arico M, Baruchel A, Bertrand Y, et al. The seventh
international childhood acute lymphoblastic leukemia
workshop report: Palermo, Italy, January 29-30, 2005.
Leukemia 2005;19:1145.
Thomas DA, Faderl S, O’Brien S, et al.
Chemoimmunotherapy with hyper-CVAD plus rituximab
for the treatment of adult Burkitt and Burkitt-type
lymphoma or acute lymphoblastic leukemia. Cancer
2006;106:1569
Vitale A, Guarini A, Ariola C, et al. Adult T-cell acute
lymphoblastic leukemia: biologic profile at presentation
and correlation with response to induction treatment in
patients enrolled in the GIMEMA LALA 0496 protocol.
Blood 2006;107:473.

119
 Guidelines for Stem Cell Transplant
in Adult ALL and
Management of Relapsed ALL
Dr. Ravi T / Dr. N Khattry

Introduction
Although ALL can be cured in up to 80% of pediatric
patients, adult patients have a 5-year survival rate of about
40%, even with the use of allogeneic hematopoietic cell
transplantation (allo-HCT) in high-risk patients. 1,2,3
Relapsed ALL remains a difficult challenge to treat
effectively, making success with the initial strategy critically
important .4 Improvements in the cure rate for adults have
been made through use of intensive multiagent
chemotherapy, treatment of the central nervous system
(CNS) as a potential sanctuary site, and the appropriate
use of allo-HCT. The ability to detect minimal residual
disease (MRD) during hematologic remission may identify
patients who require more intensive treatment while
sparing lower-risk patients unnecessary toxicities.

Risk factors and risk stratification

In analyses including more than 350 adult ALL patients
treated on prospective trials with intensive chemotherapy,

120
the Cancer and Leukemia Group B (CALGB) identified the
following clinical and biologic features which correlated
with favorable long-term outcome:
Age younger than 60 years, white blood cell (WBC) count
less than 30 000/mm3,the presence of a mediastinal mass,
T-cell immunophenotype (with or without myeloid
markers), and absence of the Philadelphia chromosome
(Ph+).5
Patients with no adverse features had a 91% estimated
probability of survival at 3 years [95% confidence interval
(CI) 66–98%]. Patients with one, two, or three unfavorable
features had 3-year survival rates of 64, 49, and 21%,
respectively. None of the patients with four adverse risk
factors survived more than 3 years. Survival at 3 years for
patients younger than 30 years was 66%, whereas survival
was 36% for patients between 30 and 59 years of age.
Only a few patients underwent allo-HCT in CR1 while
enrolled on these CALGB trials.5
The role of SCT in ALL will be discussed by considering the
clinical data evidence related to four main areas:
(1) Sibling allogeneic SCT in first complete remission
(CR1) in standard risk and high risk cases.
(2) Transplantation using matched, unrelated donors
(3) Autologous SCT.
(4) SCT in second complete remission (CR2) and
refractory disease,
There have been very few randomized controlled trials
(RCTs), and thus it is difficult to ascertain which patients
to undergo transplantation. A number of nonrandomized
clinical trials have attempted to answer important
questions, but multiple changes in sequential trials have
made direct comparisons difficult. Also contributing to

121
the complexity of ALL studies is the heterogeneity of the
disease itself as determined by patient age,
immunophenotype, cytogenetics, and molecular features
of the leukemia.
Recent clinical trials have evaluated allo-HCT as a
component of modern risk-adapted treatment in adult
ALL. However, the optimal postremission therapy for ALL
patients remains unclear.

Graft versus Leukemia effect (GVL) effect

in ALL
There is clear evidence of a graft-versus-leukemia effect in
this disease ; this was, in fact, initially recognized in ALL in
1979.6
The potential benefits of stem cell transplantation(SCT) in
adult ALL include the ability to use myeloablative
conditioning in an attempt to eradicate the leukemic clone
and to leverage a graft versus- leukemia (GVL) effect. There
are several observations that support evidence for a GVL
effect in ALL:
A. higher relapse rates are observed in patients who
undergo syngeneic transplantation compared with
allogeneic transplantation;
B. there is also a higher risk of relapse in recipients of T
cell–depleted grafts;
C. transplantation recipients who develop chronic graft-
versus-host disease have a reduced risk of relapse;
and
D. donor lymphocyte infusions (DLI) can induce
remissions in recipients who relapse after
transplantation.7

122
The GVL effect may be less robust in ALL compared with
myeloid malignancies. In a study comparing syngeneic to
allogeneic transplantation, a difference in relapse rates
was not detected with three-year probabilities of relapse
of 36% compared with 26%, respectively (p = 0.1). In
contrast, significant differences in relapse were observed
in patients with both acute myeloid leukemia (52% versus
16%) and chronic myeloid leukemia (40% versus 7%).8.
The design and the reduced statistical power of these
comparisons limit the strength of conclusions.
Several studies of DLI are in ALL9-13, but the patient
numbers are small, and it is difficult to draw definitive
conclusions. Though there appears to be a modest GVL
effect in ALL, it is certainly less striking than in myeloid
malignancies.

Role of minimal residual studies in

transplant
MRD studies have proved extremely useful for predicting
outcome in childhood ALL, but there are less definitive
data in adults. It is possible that a similar approach will be
useful in determining which patients should be treated
by chemotherapy alone and which patients should be
considered for high-risk treatments such as bone marrow
transplantation.
Minimal residual disease can be identified at different time
points in the disease and potentially allow the definition
of different risk groups. Ten percent of patients can be
identified who certainly do not need to be considered for
the high risk of allogeneic transplantation, and a further
23% who may well be very strong candidates. Clearly,
minimal residual disease values at any time point are
dependent on specific prior therapy given, and care must

123
be exercised in extrapolating from one protocol to another.
While this theoretically better defines the strategy for
defining transplantation candidates, it remains to be
demonstrated that this is an effective strategy, since those
with high minimal residual disease have an increased risk
of relapse and may possibly need further conventional
therapy to reduce tumor load before allogeneic
transplantation.14

Role of transplantation in Philadelphia

chromosome negative (Ph-) ALL
There are no trials in the field of HSCT in which patients
with a suitable sibling-matched donor have been
randomized to an allogeneic HSCT or chemotherapy.
“Genetic randomization” or assignment, along with intent-
to-treat analyses, have been the established methods of
comparing allogeneic hematopoietic stem cell transplant
(HSCT) with chemotherapy treatment.15
Gupta and colleagues16 at the Princess Margaret Hospital
reported a small single-institution series in which adult
patients with ALL in CR1 age 55 years or younger were
offered a myeloablative allogeneic HSCT. They found no
difference in outcome between the donor versus no-donor
groups. High risk adults in CR1 (which included Ph+
patients) who had a sibling matched-related donor (N =
41) were assigned a myeloablative allogeneic HSCT, while
the others proceeded to an autologous HSCT (N = 115)
in the GOELAMS-02 trial.17 Six-year disease free survival
(DFS) and overall survival (OS) were statistically superior
in those who underwent allogeneic HSCT, 75% versus 39%
(P = .0027)
The French LALA group has addressed this question
prospectively in their LALA-94 trial.18 They designed a trial

124
of risk-adapted postremission strategy in adult ALL.
Patients with age between 15 to 55 years with any high-
risk ALL features and an HLA-identical sibling were assigned
to allogeneic HSCT. Five-year DFS in the group with Ph–
high-risk ALL in first CR was improved in the allogeneic
HSCT (45% vs 23%, sibling donor vs those with no sibling
donor). In multivariate analysis, both age and WBC count
were found to be prognostic factors for DFS; however,
the impact on the two groups (donor vs no donor) was
not discussed.
In the largest trial reported to date (MRC-ECOG UKALLXII/
E2993), 3 1826 adult patients with ALL were given
induction therapy that resulted in a 91% CR rate. All
patients younger than 50 years old, irrespective of risk,
with a sibling matched related donor were recommended
to undergo a myeloablative HSCT using a total body
irradiation (TBI)–containing conditioning regimen. For 919
patients in CR, 389 with a donor were compared with
524 patients with no donor. Event-free survival (EFS) and
OS were statistically superior in the donor group: 50%
versus 41% (P = .009) and 53% versus 45% (P = .02). As
noted historically with most HSCT reports, in general, the
optimal allogeneic effect for EFS and OS were observed in
the standard risk rather than the high-risk group (where
high risk was defined by age and WBC count). Although
in both risk groups the relapse rates were significantly
lower in the donor group compared with the no donor
group, supporting a strong GVL effect, the TRM rates at 2
years were significantly increased in the high-risk group,
39% versus 20%. Such findings, in large part, reflect poor
tolerance to a myeloablative conditioning regimen in those
patients older than 35 years. Thus, the majority of the
data in Ph– patients favor an improved outcome in younger
patients who underwent a MRD allogeneic HSCT, an effect

125
lost when older patients are conditioned using a
myeloablative regimen.3
Table1.
MRC-ECOG UKALLXII/EC29933: Outcome after
allogeneic hematopoietic stem cell(HSCT) in Ph-
patients who had donors versus those who did not
have donors.
High Risk No.of 5 year 5 year 2 year
patients overall Relapse Non Relapse
401 Survival % Rates % Mortality%
Donor 171 40 39 39
No Donor 230 36 62 12
Standard Risk 512
Donor 218 63 27 20
No Donor 294 51 50 7
* High-risk is defined as age ≥ 35 years, WBC > 30,000/μL for
patients with B-cell disease or WBC > 100,000/ìL for patients
with T-cell disease, or time to attain CR > 4 weeks.

The number of patients with sibling donors, however, far

exceeds the usual ratio of success in finding HLA-matched
siblings (such as observed in acute myeloid leukemia HSCT
trials). Although the number of patients with a donor
appears high, it is consistent with some of the previously
published ALL HSCT studies. For example, in the LALA-94
study, for the high-risk group 100 patients had a donor
compared with 159 without a donor. 18,19
Role of transplantation in Philadelphia positive (Ph+) ALL
Although Ph+ patients have a lower remission rate and
higher relapse rate using conventional chemotherapy
compared to those with Ph– disease, allogeneic HSCT is
particularly effective in Ph+ patients.

126
The French Bone Marrow Transplantation Society 18
reported outcome of 121 patients with Ph+ ALL, including
102 adults, of whom 76 were in CR1 at time of transplant.
Two-year OS in the CR1 patients was 50% and the 2-year
relapse rate was 37%.
Dombret and co-workers 19 reported the LALA-94 trial data
for 154 Ph+ patients who proceeded to transplant (N =
103) after attainment of CR1. Sixty patients who
underwent allogeneic HSCT (n = 46 matched related
donor (MRELD) and n = 14 matched unrelated donor
(MUD) had the same 2-year TRM as 43 patients who
underwent autologous HSCT, but OS at 3 years was
significantly superior, 37% versus 12% (P = .02).
Attainment of BCR-ABL–negative status conferred a
significantly better OS as well, 54% versus 12% (P = .006).
Stirewalt and colleagues20 at the Fred Hutchinson Cancer
Research Center retrospectively analyzed 90 patients with
Ph+ ALL aged 2 to 56 years (median, 33 years) who
underwent myeloablative allogeneic HSCT. Morphologic
or cytogenetic evidence of ALL at time of transplantation
as well as autologous or MRELD use predicted for highest
risk of relapse, while development of chronic graft-versus-
host disease (GVHD) after transplantation lowered the
relapse rates. These investigators proposed that patients
without molecular evidence of disease (“low risk”) undergo
a less intense (nonmyeloablative) HSCT. They advocated a
more aggressive conditioning be used in “high-risk”
patients who have minimal residual or overt disease at
time of transplantation including consideration of post-
transplantation imatinib consolidation.
Gupta and coworkers16 reported a study in which unrelated
donor(URD) allogeneic HSCT was offered to a small
number of patients with Ph+ ALL in CR1. Three-year DFS
and OS were 46% and 57%, respectively; OS did not differ
127
when compared with those Ph– patients at their center
who underwent allogeneic HSCT. Finally, in the largest
prospective allogeneic HSCT study (conducted during the
preimatinib era), 5-year OS was superior in those who
underwent MRELD allogeneic HSCT compared with
chemotherapy or autologous HSCT.21 Early relapse, prior
to transplantation, however, was common supporting
referral for allogeneic HSCT as soon as possible after
attaining CR.These and other data suggest that allogeneic
HSCT provides significant GVL effect in CR1 patients with
Ph+ ALL.

Role of imatinib.
The addition of imatinib to the armamentarium for treating
Ph+ ALL has significantly improved the CR rate from
approximately 60% to 90% and has greatly enhanced the
access of such patients to allogeneic HSCT by reducing
early pretransplantation relapses, a large problem in the
past.
Yanada et al22 recently reported a significantly improved
OS in 49 adult patients with Ph+ ALL who were given
post-transplantation imatinib therapy compared with
historic controls, i.e., 1-year EFS and OS, 60% and 76%,
respectively. Although median follow-up was short at 1
year, both patients who underwent allogeneic HSCT
recipients and patients who did not had significantly better
outcome. Further, imatinib appeared to prevent earlier
relapse, enabling a greater percentage of patients to
proceed to allogeneic HSCT.
de Labarthe and coworkers 23 similarly showed that
imatinib in combination with conventional chemotherapy
provided results comparable with allogeneic HSCT. On the
other hand, Pfeifer, for the GMALL, recently reported the
prevalence of kinase domain mutations in newly diagnosed

128
and imatinib-naive Ph+ ALL. 24 BCR-ABL mutations
conferring high-level imatinib resistance were present in
a substantial proportion of patients with de novo Ph+
ALL and eventually gave rise to relapse.
Until there are published reports of larger studies with
longer follow-up, Imatinib therapy cannot be considered
the standard treatment, and allogeneic HSCT, in general,
remains the preferred approach in this subgroup of
patients. Imatinib’s positive effect on pretransplantation
PCR status, i.e., rendering patients tumor- negative before
transplantation, could improve outcome.
One can speculate, however, that in the post-
transplantation setting, imatinib therapy is only likely to
provide time until a GVL effect can develop to eradicate
disease. Prophylactic imatinib after HSCT can be given
safely.25

Alternative Donor Stem Cell

Transplantation:
Only 25% of patients eligible for allogenic transplant have
fully matched related donor. Therefore as allogenic
transplant remains the most curative option in adult ALL,
patients especially in high risk group should undergo
alternate donor transplantation such as unrelated donor
marrow or blood, umbilical cord blood transplantation,
haploidentical donor transplantations. Below are results
of few studies in each subgroup of alternate donor
transplantation.

Unrelated Donor Blood or Marrow

Kiehl and associates 26 reported a 12-year retrospective
multicenter experience comparing myeloablative HSCT in
adult patients with ALL according to donor source. Five

129
year DFS rates were not statistically different according to
donor source, MRELD (N = 62) versus URD (N = 32), 42%
versus 45%. Similarly, TRM did not differ, 39% versus 31%,
respectively. The vast majority of these patients were
considered high risk or very high risk. Further, these results
were significantly better than if the patients underwent
HSCT at CR2 or beyond or at time of active leukemia. Of
interest, as observed in sibling MRELD HSCT studies, Ph+
patients had no poorer outcome than did Ph– patients.
Cornelissen and associates 27 reported a multi-national,
myeloablative MUD HSCT trial in CR1 patients. Their data
again support a strong GVL effect in Ph+ patients, as the
relapses were significantly lower in these patients
compared with the t[4;11] and t[1;19] groups.
Yakoub- Agha et al,28 for the French Society of Bone
Marrow Transplantation and Cell Therapy, recently reported
the first prospective study in which outcomes for standard-
risk malignancy patients who underwent sibling MRELD
HSCT were comparable with HLA-allellically matched (10
of 10) unrelated donor HSCT. Such data enhance the case
for performing MUD HSCT in this patient population
At this time, there are insufficient data to conclude whether
a myeloablative URD should be offered even to high-risk
adult patients with ALL in CR1. Given the GVL effect that
seems to be particularly beneficial in Ph+ patients, this
patient population may be reasonable to consider for URD
transplantations in CR1.

Umbilical Cord Blood Transplantation

One single-institution trial retrospectively compared a
myeloablative HSCT regimen followed by either an
umbilical cord blood (UCB) graft (N = 100) or a related
donor blood or marrow graft (N = 71).29 UCB was safe

130
and effective compared to a related marrow or mobilized
blood graft, but only 16 adult ALL patients were included
and an ALL diagnosis carried with it a nearly 2.5-fold
increased risk for relapse.
The analyses of UCB transplantation data are hampered
by small patient numbers, heterogeneity of remission
status, variability of matching, incomplete information
about UCB cell dose (including the use of double UCB
grafts), and high engraftment failure rates. At present,
there are insufficient data to recommend use of this donor
stem cell source outside the context of a clinical trial.

Haploidentical Donor Transplantations

This specialized approach has been available at only a very
few centers. The high rejection and unacceptable GVHD
rates have been lowered by use of more intense preparative
regimens and infusion of higher numbers of purified
hematopoietic stem cells. When applied to patients not
in CR1 or CR2, haploidentical transplantations appear
inferior to other alternative donor HSCT results. The major
limitations, in addition to relapse in advance disease state,
appear to be a proclivity for severe and often-fatal
opportunistic infections. Some centers performing this
procedure in high risk CR1 (including Ph+) patients,
however, attain results equivalent to MUD allogeneic
HSCT.30 Such data need to be corroborated in multi-
institutional trials, but this strategy is a distinct option in
those patients without a MRD.

Reduced-Intensity Conditioning
Although the GVL effect has been shown to be responsible
for some of the benefit in fully ablative HSCT, TRM,
especially in the older recipient, may offset the reduction
in relapse. As donor lymphocyte infusion (DLI) has been

131
shown to decrease MRD and potentially improve survival
in those patients, there may be a role for nonmyeloablative
or reduced-intensity conditioning in ALL. Hamaki et al31
investigated reduced-intensity conditioning in 33 ALL
patients with a median age of 55 years (range, 17-68 years)
using fludarabine-based conditioning regimens followed
by MRELD or URD grafts; 9 patients died of TRM. The 1-
year progression- free survival (PFS) rate for the 19 patients
who underwent transplantation in CR1 or CR2 is 31%. Six
of 13 patients have a median DFS of 14 months (range,
4-37 months).
The City of Hope recently reported their single-center
experience in 21 patients of ALL with median age 46 years
(range, 6-68 years) given fludarabine-melphalan
conditioning followed by a MRELD (N = 7) or a MUD (N
= 14) graft.32 Ten patients were in CR1 and 11 patients
had Ph+ ALL. In this high-risk group, who were either
elderly (older than 50 years), had failed a previous HSCT,
or had a comorbid condition precluding use of a
myeloablative regimen, the 100- day mortality and 1-year
relapse rates were 11% and 8%, respectively, while 1-year
DFS and OS rates were 77% and 71%, respectively.

Autologous HSCT
This modality first was used nearly 50 years ago as
treatment for relapsed/refractory ALL. More recently, this
approach has been used in patients significantly earlier in
their disease course, to provide the potential benefit of
myeloablative therapy to those for whom a MRD is not
available, as MRD are available only for a minority of
patients. 33 While long-term DFS has been attained in some
patients, few prospective, randomized trials have analyzed
sufficiently large enough numbers of patients to determine
if this treatment is appropriate therapy.

132
A variety of preparative regimens have been used for
autologous HSCT in adult patients with ALL, but there is
no clear benefit to any one approach.34 On the other hand,
an analysis from the Center for International Blood and
Marrow Transplant Research (CIBMTR) by Marks and
associates reported in CR1 and CR2 patients with ALL
receiving sibling-matched allogeneic HSCT that Etoposide
or cyclophosphamide in combination in a TBI-containing
regimen, or use of TBI dose >1300 cGy were associated
with reduced risk of relapse and treatment failure. These
allogeneic HSCT data suggest a threshold TBI dose for
antileukemia efficacy and could have implications for
Autologous HSCT regimens; these conclusions, however,
have to be regarded as speculative based on retrospective
analysis of registry data.35
The French LALA-94 trial 18-19, group reported a prospective
multicenter trial comparing Autologous HSCT, MRELD
allogeneic HSCT, and chemotherapy in 1000 patients. Ph–
patients at high risk for relapse in CR1 who did not have
MRELD (N = 129) were randomized to an autologous
HSCT and 2-year maintenance therapy (6-MP and
methotrexate) or chemotherapy. Three-year DFS were 39%
in the autologous HSCT versus 24% in the chemotherapy-
treated group, not statistically significant.
Dhédin et al 36 updated autologous HSCT results for the
LALA-85, -87 and -94 trials. A subset analysis of 180 high-
risk CR1 patients showed no differences in either DFS or
OS at 10 years for the autologous HSCT versus the
chemotherapy treatment groups, 20% versus 12% (P =
.10) and 20% versus 13% (P = .78).
Other small series include reports of long-term survivors
in younger patients who underwent autologous HSCT, but
such studies do not include control subject comparisons.37

133
The joint MRC-ECOG UKALLXII/E2993 trial3-4 compared
446 CR1 patients randomized to undergo either an
autologous HSCT (N = 223) or 2.5 years of maintenance
therapy (N = 223). EFS was statistically superior for the
chemotherapy-treated patients, 42% versus 33% (P = .02).
While OS also was improved in the chemotherapy group
(47% versus 37%), these results were not statistically
significant (P = .06). Those assigned to an autologous
HSCT did not receive post-transplantation maintenance
therapy and continued to relapse at a higher rate. Other
groups have reported that use of maintenance
chemotherapy after Autologous HSCT can prolong
remission .These and other data do not support the routine
use of autologous HSCT in adult patients with ALL, and
patients should be referred for participation in clinical trials
addressing this concept.

Post-Transplantation Maintenance
Therapy
In vitro purging of the stem cell graft also has been used
in a number of autologous HSCT studies in ALL. Although
such an approach may reduce the number of leukemic
progenitor cells in the graft, there has been no correlation
between purging efficacy or dose of residual tumor infused
and patient outcome. Two groups have suggested that
use of maintenance chemotherapy after autologous HSCT
can reduce relapse rates. 38 .In the UK, Mehta and
colleagues39 noted that increased intensity of treatment
was more efficacious but that many patients could not
tolerate such an approach, a result similar to the Johns
Hopkins group data. Until less marrow-toxic agents are
readily available to provide a more-targeted therapy, this
postautologous HSCT maintenance strategy cannot be
recommended for widespread use.

134
Allogeneic Transplantation for Patients
with Relapsed/Refractory Disease
Two large studies in adults, one from the MRC/ECOG
group4and one from the LALA group,40 confirmed that in
most cases salvage after relapse is very unlikely. Patients
who do most poorly are those with the shortest duration
in first remission and those who are older. May be half of
those are unlikely to reach another remission, and many
will not be eligible for hematopoietic stem cell
transplantation for reasons such as performance status
and age. There is some minimal evidence that the
occasional patient with refractory leukemia can benefit,
and registry data can also show sometimes an apparently
useful retrieval of relapsed refractory patients with adult
ALL. However, it remains true that for the younger adult
patient achieving a second remission some form of
allogeneic transplantation remains the best chance for
long-term survival.4,40-41

Conclusions
Allogeneic stem cell transplantation remains the single
most potent therapy for the reduction of relapse in adult
patients with ALL. The largest ever randomized trial in this
disease, the MRC/ECOG study, which studied donor versus
no donor transplantation, firmly established the benefit
of having a sibling donor. This benefit was true of patients
at all ages in the study (between 15 and 50 years).
However, the donor versus no donor approach is in danger
of becoming outmoded, as in many studies those
previously in a “no donor” category will now undergo
matched unrelated donor transplants or haploidentical
transplants and possibly also cord blood transplants. There
is increasing evidence that the outcome from “unrelated
donor” category transplants may be not very different in

135
this disease than that of a sibling transplant. Hitherto,
transplants have been ruled out for patients over the age
of 50 or 55 because of the potential toxicity of the
procedures, although those patients have by far the worse
prognosis on conventional therapy. It is now possible that
RIC transplants may offer an allogeneic effect for this group
at a level of safety hitherto unobtainable. Finally, MRD
studies that need to be worked through considerably
further in the adult group may offer the possibility of
identifying two groups of patients, those who do not need
a transplant at all and those who have a greater
predisposition to relapse and may therefore benefit from
a transplant either immediately or at the first molecular
sign of relapse.

Recommendations
1. Allogeneic transplantation using stem cells from a
matched sibling donor is recommended for adult
patients with high and standard-risk ALL who achieve
a first remission (Grade 2A).
2. Allogeneic transplantation using stem cells from a
matched sibling or unrelated donor is recommended
for adult patients with relapsed ALL who achieve a
second remission (Grade 1C).
3. Autologous transplantation is not recommended for
adult patients with ALL (Grade 1C).
4. Imatinib is recommended in combination with
chemotherapy for patients with Philadelphia
chromosome-positive ALL (Grade 1C).
5. For younger adult patient achieving a second
remission, some form of allogeneic transplantation
remains the best chance for long-term survival (Grade
2A).

136
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Appendix

Grading of recommendation
Levels of Evidence
1++ High-quality meta analyses, systematic reviews of
randomized controlled trials (RCTs), or RCTs with a very low
risk of bias
1+ Well conducted meta analyses, systematic reviews of RCTs,
or RCTs with a low risk of bias
1– Meta analyses, systematic reviews of RCTs, or RCTs with a
high risk of bias
2++ High-quality systematic reviews of case-control or cohort
studies
High-quality case-control or cohort studies with a very low
risk of confounding, bias, or chance and a high probability
that the relationship is causal
2+ Well conducted case control or cohort studies with a low
risk of confounding, bias, or chance and a moderate
probability that the relationship is causal
2– Case control or cohort studies with a high risk of
confounding, bias, or chance and a significant risk that the
relationship is not causal
3 Non-analytic studies; eg, case reports, case series
4 Expert opinion
Grading the Strength of the Treatment Recommendation
Grades of Recommendation
A At least one meta analysis, systematic review, or randomized
controlled trial (RCT) rated as 1++, and directly applicable to
the target population;
or
A systematic review of RCTs or a body of evidence consisting
principally of studies rated as 1+, directly applicable to the
target population, and demonstrating

144
Overall consistency of results
B A body of evidence including studies rated as 2++, directly
applicable to the target population, and demonstrating overall
consistency of results;
or
Extrapolated evidence from studies rated as 1++ or 1+
C A body of evidence including studies rated as 2+, directly
applicable to the target population and demonstrating overall
consistency of results;
or
Extrapolated evidence from studies rated as 2++
D Evidence level 3 or 4; or
Extrapolated evidence from studies rated as 2+
From: Harbour and Miller. Brit Med J. 2001;323:334-6.

145
 Molecular Classification And Risk
Stratification In AML
Dr. Vamshi Krishna
Dr. Hari Menon

Introduction
Acute myeloid leukemia is a heterogeneous disease and
has biologically distinct subgroups. Until the recent past,
all patients were treated with standard 7 + 3 regimen
and were found to be having differing outcomes. This
was more pronounced in the elderly and in patients with
secondary AML who uniformly had worse outcomes as
compared to normal patients. Remission rates were
approximately 33% to 50% lower than those seen in de
novo disease, and duration of remission were generally
shorter. Thus, it was recognized that a single standard
treatment protocol applied to all patients may result in
variable treatment outcomes. Further understanding of
the biology of AML recognized distinct subsets that were
categorized based on cytogenetic and molecular makeup
into distinct groups who behaved similarly in terms of
response to therapy and long-term outcomes. This
paradigm shift in the understanding of AML has resulted
in the concept of risk-adapted therapy which allows for
146
recognition of this biologic diversity. The current treatment
regimens take into account key biological features, such
as cytogenetic and molecular markers, when formulating
treatment regimens and investigating emerging targeted
therapies based on disease characteristics.

Importance of cytogenetics
The most important prognostic factor is the biology of
the underlying disorder, as reflected by the cytogenetics.
Several large studies have reported the prognostic
importance of pretreatment cytogenetics and have helped
categorize chromosomal abnormalities into good-,
intermediate-, and poor-risk groups (Table 1).
The Medical Research Council from the United Kingdom
reported that the hierarchical system of karyotype
classification retained its predictive value across different
age groups, in different AMLs and across different
treatment modalities like chemotherapy, autologous and
allogeneic bone marrow transplantation. In the AML 10
trial, which included 1,612 patients younger than 55 years,
poor risk group had remission rates of 60% with 5-yrear
relapse rates of 80% and overall survival of 20 %. In
contrast, patients with favourable risk profile had 90%
CR rates, 5 year relapse rate of 35 % and overall survival
of 65%. The ECOG/SWOG and the CALGB have similarly
shown that division of AML into cytogenetically based
groups correlates with outcomes and is a prognostic
marker for outcomes following allogenic and autologous
transplant.
Currently, the division schema for most major oncology
groups is similar. Generally, the poor-risk or unfavorable
group includes those with complex karyotypes (> 3-5
abnormalities), chromosome 5 or 7 abnormalities, or

147
chromosome 3q abnormalities. The results for these
patients are dismal with standard chemotherapy alone
and most advocate stem cell transplantation in first
remission for such patients. More recent series suggest
that stem cell transplantation may offer patients a chance
for prolonged survival. In contrast, stem cell
transplantation in first complete remission (CR) is not
recommended in patients with good-risk or favorable
cytogenetics which Include those with t(15;17), t(8;21),
t(16;16), and inv(16) translocations. Acute promyelocytic
leukemia (APL), defined by the t(15;17) translocation, is
considered a distinct entity with a distinct biology when
considering the treatment of AML. Chemotherapy along
with differentiating agents is sufficient to give cure for
such patients.

The Core-binding factor leukaemias

The core-binding factor leukemias [t(8;21), t(16:16), and
inv(16)] are so named because the specific chromosomal
translocation results in the fusion of a gene to either the á
or â subunit of the heterodimeric core-binding factor,
resulting in the recruitment of histone deacetylase to the
promoter. Activity of this enzyme results in inhibition of
transcriptional activity from promoters of genes involved
in cellular differentiation. These leukemias are exquisitely
sensitive to high-dose cytosine arabinoside. When patients
are treated with dose-intensive regimens, overall survival
may be approximately 60%. The t(8;21) translocation is
more prevalent among AML with evidence of
differentiation on morphology. The inv(16) and t(16;16)
translocations are usually found among the
myelomonocytic leukemias. They typically have
characteristic abnormal eosinophils that contain large
basophilic granules on the peripheral smear.

148
 Table 1 Risk-based classification of AML across different groups
MRC
Good risk Inv(16)/t(16;16)/Del(16q) with or
without other abnormaities
t (15;17) with or without
other abnormalities
t (8;21) with or without
other abnormalities
Intermediate risk Normal, 11q23, +8, del(9q),
del(7q), +21, +22, all others
Poor risk Complex karyotype
(>5 abnormalities), del(5q),
-5, -7, abnl(3q)
Source – J Natl Comp Cancer Network 2008
149
CALGB SWOG/ECOG
t(8;21), inv(16)/t(16;16) Inv(16)/t(16;16)/Del(16q) with or
without other abnormaities
t (15;17) with or without other
abnormalities
t (8;21) without del(9q) or
complex karyotype
Normal, -Y, del(5q), Normal, +6, +8, -Y, del(12p)
t(6;9), t(9;11), 11q23,
+8, del(9q), loss of 7q,
+8 sole, t(6,11), -7,
+11, +13, +8 with
one other, del(20q),
del(11q), +21, t(11;19)
Complex karyotype Complex karyotype
(>3 abnormalities), (>3 abnormalities), del(5q), -5,
t(3;3)/inv 3, +8 sole, -7/del(7q), abnl(3q), 9q,11q,
+8 with one other, -7, 20q, 21q, 17p, t(9;22), t(6;9)
abn(12p), t(6;11),
+21, t(11;19)
AML with normal cytogenetics
Patients who have chromosomal abnormalities that do
not fit into other categories are considered to have
intermediate-risk disease. The largest group within this
category are patients with a normal karyotype,
representing approximately 40% to 49% of adults with
de novo AML. However, this is a heterogenous group and
these patients have a broad range of treatment outcomes,
thereby suggesting specific factors that influence the
pathogenesis and outcome for this entity.
Schlenck et al compared the mutational status of the
NPM1, FLT3, CEBPA, MLL, and NRAS genes in leukemia
cells with the clinical outcome in 872 adults younger than
60 years of age with cytogenetically normal AML for the
German-Austrian AML study group. A total of 53% of
patients had NPM1 mutations, 31% had FLT3 internal
tandem duplications (ITDs), 11% had FLT3 tyrosine kinase–
domain mutations, 13% had CEBPA mutations, 7% had
MLL partial tandem duplications (PTDs), and 13% had
NRAS mutations. The overall complete-remission rate was
77%. The genotype of mutant NPM1 without FLT3-ITD,
the mutant CEBPA genotype, and younger age were each
significantly associated with complete remission. MLL-PTD
was associated with significantly increased risk of relapse
or death. The benefit of allotransplant in this study was
limited to the subgroup of patients with the prognostically
adverse genotype FLT3-ITD or the genotype consisting of
wild-type NPM1 and CEBPA without FLT3-ITD.

FLT 3 Internal tandem duplications

One of the first reproducible recurrent genetic changes
identified was the length or internal tandem duplications
(ITD) within the juxtamembrane domain of the FMS-like

150
tyrosine kinase 3 (FLT3) receptor. This results in constitutive
activation of the enzyme through destabilization of the
negative regulatory function of the juxtamembrane
domain. This constitutive activation results in the abnormal
activation of the STAT5 and FOXO transcription factors,
up-regulation of the Pim-1 and -2 proto-oncogenes, and
down-regulation of the CEBPá transcription factor that is
involved in normal myeloid differentiation.
Since this abnormality was identified in the mid-1990s, it
has been found to be common in the normal cytogenetic
group and can be detected in approximately 28% to 33%
of these patients. The presence of an ITD has been shown
to confer a worse clinical prognosis, and therefore suitable
patients with this abnormality usually undergo allogeneic
bone marrow transplantation in first remission.
Furthermore, heterogeneity may exist within the FLT3-ITD-
positive group, because some individuals are homozygous
for the mutated allele, whereas others have one wild-type
copy of the allele. Several groups have reported the
prognostic significance of the allelic ratio (mutated-to-
wild-type), with higher values ultimately having a worse
prognosis. Kottaridis in the analysis of data from the MRC
AML 10 and other trials studied 854 patients of whom 27
% had FLT3 ITD mutation. These patients had increased
relapse risk and more adverse EFS and OS.
A second mutation in the FLT3 gene has been observed
in approximately 5% of patients with AML and involves a
point mutation at aspartate 835 within the activation loop
of the enzyme. It is analogous to point mutations seen
and characterized in multiple other receptor kinases,
including KIT, and results in shifting the activation loop of
the kinase domain to an open configuration, allowing
constitutive kinase activation. The clinical prognosis for
this particular mutation is currently a point of controversy,

151
because this mutation may not confer as negative a
prognosis as the ITD.
Table 2 Molecular mutations in AML with normal cytogenetics
Implicated gene Consequence of mutation
Good risk NPM 1 Mutated nucleoplasmin Class II
(without FLT3-ITD) gene
CEPBá Mutated CCAAT/enhancer Class II
(without FLT3-ITD) binding protein á gene
FLT3-TKD Mutated tyrosine kinase Class I
domain of FMS-related
Tyrosine kinese 3 gene
Poor risk FLT3-ITD Internal tandem duplication Class I
of FMS-related tyrosine
kinase gene
MLL-PTD Partial tandem duplication Class II
of mixed lineage leukaemia
gene
BAALC Overexpression of brain Class II
and acute leukaemia,
cytoplasmic gene
ERG Overexpression of Class I
v-ets erythroblastosis
E26 oncogene homolog
WT-1 Mutated wilms tumour-1 Class II
gene
MN-1 Overexpression of high Class II
meningioma-1 gene
Source - J Natl Comp Cancer Network 2008
Class I abnormalities - result in a proliferative and survival
advantage for immature hematopoietic cells
Class II abnormalities - alter hematopoietic differentiation,
maturation, and apoptosis Reference - Gilliland and Griffin

Mixed-lineage leukaemia (MLL)

rearrangements
Length or partial tandem duplication (PTD) involving the
mixed lineage leukemia gene (MLL) was the first adverse
152
prognostic marker identified in AML with normal
cytogenetics. In normal cytogenetics AML, MLL-PTD affects
only one allele, and expression of the second, wild-type,
allele is silenced. MLL-PTD results in an elongated protein
that retains the histone H3 lysine 4 methyltransferase
activity. This is distinct from the chimeric fusion proteins
involving 11q23 translocations that have been found to
retain the DNA binding activity but lack the histone
methyltransferase domain. The poor prognosis is due to a
global DNA methylation potentially causing silencing of
the tumor suppressor genes.
A recent CALGB study showed that in patients with normal
cytogenetics AML, the poor prognostic effect of MLL-PTD
can be potentially negated through use of intensive
consolidation chemotherapy in first CR, including an
autologous peripheral stem cell transplantation.

Good-risk mutations
Two class II mutations have been identified that confer a
good prognosis in patients with AML who do not also
have a FLT3-ITD abnormality. The first is a mutation within
exon 12 of the nucleophosmin (NPM) gene that results in
cytoplasmic as opposed to nuclear localization of the
protein. This abnormality can be seen in up to 25% of
patients with AML. The second is a mutation within the
CCAAT/enhancer binding protein á (C/EBPá). The presence
of either mutation in the background of a wild-
type FLT3 gene confers a favorable prognosis in patients
undergoing chemotherapy alone.

Other mutations
Activating mutations in the KIT kinase in the context of a
core-binding factor cytogenetic abnormality confers a poor

153
prognosis, the positive impact of the CBF abnormality
notwithstanding. Overexpression of other genes,
including the survival factor CXC R 4 , BAALC, MN1,
and ERG, as well as WT1 mutations are also known to
confer a poor prognosis.

Therapeutic implications of molecular

classification
Induction therapy is standard for all risk groups among
AML patients. The importance of molecular classification
is relevant in planning appropriate postremission therapy.
Patients in remission after induction who have a poor-risk
karyotype are to be considered for allogenic stem cell
transplantation. The good-risk group would require
standard chemotherapy in the form of induction followed
by 2-3 post-remission cycles. Patients with normal
cytogenetics with no obvious molecular abnormality
should also be offered standard therapy with the option
allogeneic stem cell transplant as post remission therapy
provided there is matched donor available.

Conclusions
Patients with normal cytogenetics previously presented a
challenge for clinicians, because outcomes vary widely and
no other prognostic data existed. The data now available
from molecular/genetic studies allows for further
stratification based on defined risk factors. Testing
for FLT3 gene mutations, BAALC overexpression, PTD
o f MLL, WT1 mutations, and overexpression
o f ERG and MN1 is important because of the negative
impact on patient outcomes, and therefore may be
incorporated into the standard workup for AML. Gene-
expression profiling represents an exciting evolving concept
to prognosticate and tailor treatment in AML.

154
Suggested reading
Keating M, Smith T, Kantarjian H, et al. Cytogenetic pattern
in acute myelogenous leukemia: a major reproducible
determinant of outcome. Leukemia 1988;2:403-412.
Grimwade D, Walker H, Oliver F, et al. The importance of
diagnostic cytogenetics on outcome in AML: analysis of
1,612 patents entered into the MRC AML 10 trial. Blood
1998;92:2322-2333.
Slovak M, Kopecky K, Cassileth P, et al. Karyotypic analysis
predicts outcome of preremission and postremission
therapy in adult acute myeloid leukemia: a Southwest
Oncology Group/Eastern Cooperative Oncology Group
study. Blood 2000;96:4075-4083.
Byrd J, Mrózek K, Ruppert A, et al. Pretreatment cytogenetic
abnormalities are predictive of induction success,
cumulative incidence of relapse, and overall survival in adult
patients with de novo acute myeloid leukemia: results from
Cancer and Leukemia Group B (CALGB 8461). Blood
2002;100:4325-4336.
Gilliland DG, Griffin JD. The roles of FLT3 in hematopoiesis
and leukemia. Blood 2002;100:1532-1542.
Schlenk R, Dohner K, Krauter J, et al. Mutations and
treatment outcome in cytogenetically normal acute
myeloid leukemia. N Eng J Med 2008;358:1909-1918.
Kottaridis P, Gale R, Frew M, et al. The presence of a FLT3
internal tandem duplication in patients with acute myeloid
leukemia (ML) adds important prognostic information to
cytogenetic risk group and response to the first cycle of
chemotherapy: analysis of 854 patients from the United
Kingdom Medical Research Council AML 10 and 12 trials.
Blood 2001;98:1752-1759.

155
 Optimization of induction therapy
in Acute Myeloid Leukemia
Dr Manju Sengar, Dr Nandish Kumar

Successful induction therapy to achieve complete response

is the key factor in management of acute myeloid leukemia.
Induction therapy aims to reduce the total body leukemia
cell population from approximately 1012 to below the
cytological detectable level of about 10 9 cells. This
reduction in leukemic burden allows normal hematopoiesis
to recover and subsequent more intense chemotherapy.
The most decisive factor for the type and intensity of
induction therapy is age. Among patients with acute
myeloid leukemia (AML), treatment regimens and
outcomes may differ between younger and older adults.
Although there is no clear dividing line between younger
and older adults when dealing with AML, in most studies,
“older adults” was defined as over age 60.
Criteria for response assessment: After conventional
induction therapy, response assessment is commonly
performed between day 21 and day 28 after start of
therapy.
Complete remission is defined as Bone marrow blasts
<5%; absence of blasts with Auer rods; absence of

156
extramedullary disease; absolute neutrophil count>1000/
microL , platelet count >100 000/microL); independence
of red cell transfusions.
Complete response with incomplete recovery (CRi) is
defined as all the complete response criteria except for
residual neutropenia (<1000/microL) or
thrombocytopenia (<100 000/microL).
Cytogenetic complete response is defined as reversion to
a normal karyotype at the time of morphologic complete
response or CRi in cases with an abnormal karyotype at
the time of diagnosis; based on the evaluation of 20
metaphase cells from bone marrow
Molecular CR has no standard definition but depends on
molecular target
Conventional induction therapy: Modern induction
therapy in AML began in the late 1960s, when both
cytarabine and anthracyclines were shown to have
significant activity as single agents. A series of classic
experiments by the Cancer and Leukemia Group B (CALGB)
some 25 years ago established what really became
standard of care for at least two decades. It included a
combination of 7 days of cytarabine 100 mg/m2/daily and
3 days of daunorubicin 45 mg/m2/day.1 The so-called “3
+ 7” regimen remained the gold standard against which
all new combinations of agents are compared. With this
regimen 60% to 80% of young adults and 40% to 60%
of older adults can achieve a CR. The results of induction
therapy are greatly influenced by prognostic factors, most
importantly the leukemia cell karyotype at presentation.
Due to relatively modest complete responses a continuous
search went on find better ways to administer induction
therapy. A large number of randomized phase III and phase

157
II studies have been done several cooperative groups across
the globe to improvise the conventional induction therapy
and thus the complete response rates. The main questions
answered or raised by these studies are following-
1. Is there a preferred anthracycline or anthraquinone?
2. What is the optimal dose of daunorubicin?
3. Is there a role for higher doses of cytarabine?
4. Is there are role for third drug?
5. Should induction be attenuated for older patients?
6. What is the role of new agents and cytokines like G-
CSF or GM-CSF?

Choice of anthracyclines :
To improve the results of induction therapy idarubicin and
mitoxantrone have been tested along with cytarabine.Four
prospective randomized trials comparing idarubicin to
daunorubicin suggest that idarubicin may have benefits,
particularly in young adults. In three of the studies,
idarubicin was associated with a significantly higher CR
rate, particularly in younger patients, was more effective
in eradicating leukemia after one course, and was
associated with a lower incidence of resistant leukemia.
In these prospective studies, idarubicin at 12 or 13 mg/
m2 was compared to daunorubicin at a dose of 45 mg/
m2. Therefore, it may be possible that a higher dose of
daunorubicin may confer the same apparent benefits
observed with idarubicin. Furthermore, it is not clear that
overall survival with idarubicin is superior to that achieved
with daunorubicin since two studies showed an advantage
and two did not. A meta-analysis by the AML Collaborative
Group reported similar early induction failure rates (20%
for idarubicin vs 18% for daunorubicin, P=0.4), but fewer

158
late (after day 40) induction failures with idarubicin (62%
vs 53%, P=0.002). Among patients achieving CR, fewer
patients assigned to idarubicin relapsed (P=0.008) but
somewhat more died in CR, resulting in a nonsignificant
benefit in disease free survival(P=0.07). Overall survival
was better with idarubicin compared with daunorubicin,
with 13% vs 9%, respectively, alive at 5 years (P=0.03). In
AML 10 trial, remission induction treatment consisted of
cytarabine 25 mg/m2 as intravenous bolus followed
immediately by 100mg/m2 given as a continuous infusion
daily for 10 days; etoposide 100 mg/m2 daily by
intravenous infusion (1 hour) on days 1 to 5; and on days
1, 3, and 5, one of the following: daunorubicin 50 mg/
m2 as a 5-minute infusion, mitoxantrone 12mg/m2 as a
30-minute infusion, or idarubicin 10 mg/m2as a 5-minute
infusion. In case of partial remission, a second remission
induction course with the same drugs was given. In case
of complete response, a single course of consolidation
therapy was administered, consisting of intermediate-dose
cytarabine (500mg/m2 every 12 hours in a 2-hour infusion
on days 1 through 6) and the same intercalator used
during induction, given on days 4 to 6. Younger patients
with a sibling donor were then assigned to undergo an
allogeneic stem cell transplant (SCT). Those without such
a donor, as well as older patients, were to receive an
unpurged autologous blood or bone marrow SCT. Results
show that all 3 regimens had equal CR rates. Survival was
not different in patients receiving allogenic transplant but
disease free survival was inferior with daunorubicin arm
in patients receiving autologous transplant. In adult
patients with AML who do not receive an allogeneic SCT,
the use of mitoxantrone or idarubicin instead of
daunorubicin enhances the long-term efûcacy of
chemotherapy.

159
Conclusion: Drugs other than daunorubicin like
idarubicin and mitoxantrone can be considered during
induction therapy if allogeneic stem cell transplant is not
feasible as postremission therapy. However the relevance
of this substitution needs to be reconsidered if higher doses
of daunomycin are being used. Level of evidence II

Dose of Daunorubicin :
Several major studies, particularly CALGB 9621 and the
French ALFA 9000 study, have demonstrated that higher
doses of DNR up to 80 or 90 mg/m2 can be administered
safely. Recently two major cooperative groups have
prospectively compared 45 mg/m2 of DNR to 90 mg/
m2.The Eastern Cooperative Oncology Group (ECOG)
studied younger patients up to age 60 and very recently
reported a significantly higher CR rate for patients receiving
90 mg/m2 72% versus 57% (P = .004). More importantly,
the overall survival was also significantly prolonged among
patients receiving the higher dose of daunorubicin,
particularly among patients with favorable or intermediate
cytogenetics . The Dutch-Belgian Hemato-Oncology
Cooperative Group (HOVON)/Swiss Group for Clinical
Cancer research (SAKK) recently completed a study in older
adults administering 45mg/m2 or 90 mg/m2. The CR rate
for patients receiving the 90 mg/m2 dose was superior to
that of patients receiving 45 mg/m2, 64% versus 54%,
respectively (P = .002), with a greater percentage of
patients achieving remission after only one course of
treatment, 52% versus 35%, respectively(P d” .001). There
were no significant safety issues in the high dose, finally
putting to rest the myth that older patients, even if fit,
should be treated in induction with attenuated doses.

160
Conclusion: Based on historic trials and the most recent
data, it appears probably fair to say that 45 mg/m2 should
no longer be considered as the standard of care. For
induction therapy, also for older patients, the dose should
be clearly higher - 90 mg/m2 for 3 days provided optimal
supportive care is being given. Level of evidence II.

Dose of Cytosine arabinoside :

When originally used in the 1960s and 1970s, ara-C was
administered at daily doses of 100 to 200 mg/m2 (i.e.,
standard-dose ara-C) by twice-daily intravenous bolus or
by continuous intravenous infusion. The latter approach
was used because of the drug’s very short half-life. When
given by continuous infusion, CR rates were higher when
the same total dose of ara-C was given over 5 days rather
than 2 days.
High dose cytarabine (HDAC) refers to cytarabine doses
of 1000 to 3000 mg/m2 administered intravenously over
one to three hours every 12 hours for 6 to 12 doses. Two
prospective randomized cooperative group trials have
evaluated the use of HDAC in combination with an
anthracycline for induction therapy in newly diagnosed
AML. Southwest Oncology Group (SWOG); 2 g/m2 every
12 hours [q12h] on days 1-6), and the Australian Leukemia
Study Group (ALSG; 3 g/m2 per q12h on days 1, 3, 5, and
7). None of the randomized trial showed a higher CR rate
with HDAC, and both demonstrated increased toxicity. In
a trial by the AML Cooperative Group (AMLCG), 1 versus
2 courses with HDAC (3 g/m2 per q12h on days 1-3) in
induction produced equal CR rates, disease-free survival
and moderate toxicity. The review of three randomized
trials showed that induction therapy with HDAraC
improved long-term disease control and overall survival
in adults age 60 ≤ years with de novo AML. It remains
161
unknown whether patients should receive HDAraC during
induction or if it is to be given during postremission
therapy. Further analyses should focus on this issue and
on the effects of HDAraC in prognostically different
subgroups of patients with AML.
Conclusion: HDAC can be included either in induction
or postremision therapy. Level of evidence II

Role of third drug:

Although 6-thioguanine often is included in the 3 + 7
regimen, a Cancer and Acute Leukemia Group B (CALGB)
study consisted of 668 patients who were randomly
assigned to three initial therapy groups. They received
either 7 days of cytarabine and 3 days of daunorubicin or
the same regimen with thioguanine. The third group
received 10 days of cytara bine and 3 days of daunorubicin.
The remission rates were 53%, 57%, and 57% for the
three groups (P =0 .6). There was no demonstrable
difference in toxicity and no statistically signiûcant
difference in remission duration & survival between
patients treated on the three induction arms. Bishop’s
study included 264 adults who were randomized to receive
either cytarabine by continuous infusion for 7 days and 3
days of daunorubicin or the same regimen with the
addition of etoposide 75 mg/ m2 per day for 7 days. The
remission rate was 56% without etoposide and 59% with
it. Survival was similar in the two arms but there was
signiûcantly improved remission duration with etoposide
(mean 18 v 12 months, P = .01) in patients younger than
55years. These two trials were too small to provide
conclusive evidence as to whether the addition of a third
drug is beneûcial, although in both studies the three-drug
regimen did slightly better.

162
A larger randomized trial by the Medical Research Council
(MRC) in Great Britain conducted with patients younger
than 55 years and older than 55 years found that
replacement of 6-thioguanine in DAT by etoposide had
no effect on CR rate, remission duration, or survival. There
was also no significant difference between the groups in
the longer-term measures of efficacy: disease-free survival
at 6 years from CR was 42% for DAT and 43% for ADE
(P= .8); relapse rate at 6 years was 50% for DAT and 49%
for ADE (P = .6); survival at 6 years was 40% for both DAT
and ADE(P =0.9).
Conclusion: The addition of third drug in induction
regimen has not been proven to provide a benefit over
conventional regimen. Level of evidence III. However if a
three drug regimen is being considered etoposide is a
better choice. Level of evidence III.

Role of gemtuzumab ozogamicin (GO):

In the Medical Research Council (MRC) AML15 study of
younger adults, 1115 patients were randomized in
induction to receive, or not, gemtuzumab ozogamicin
(GO) in addition to the induction regimen. There was also
a randomization between 3 different induction regimens,
but all patients underwent the GO randomization. The
postremission therapy was identical in both arms of the
study. This study showed a benefit in relapse free survival
and overall survival in favourable cytogenetics group but
not in all patients as a group.
Conclusion: Role of GO in induction therapy in young
and older adults still remain unproven. Level of evidence II

Role of growth factors :

Sensitization of leukemic cells with hematopoietic growth
factors, such as granulocyte colony-stimulating factor
163
(G-CSF) and granulocyte macrophage (GM)–CSF, has been
studied to increase cytotoxicity of chemotherapy. The
Dutch-Belgian Hemato-Oncology Cooperative Group
(HOVON) and Swiss Group for Clinical Cancer Research
(SAKK) showed priming with G-CSF resulted in a
significantly better DFS, and in the intermediate-risk group
also a significantly better OS. Similarly, in a study by the
Acute Leukemia French Association (ALFA) group, priming
with GM-CSF resulted in a higher CR rate and a better
EFS, in particular in the intermediate risk group, albeit
without influencing OS. In contrast, in a study by the
AMLCG, G-CSF priming did not impact OS or RFS.
Long term results of Acute Leukemia French Association
(ALFA)-9802 trial suggest that sensitization of leukemic
cells and their progenitors by GM-CSF appears as a
plausible strategy for improving the outcome of patients
with newly diagnosed AML. Patients with poor-prognosis
FLT3-ITD or MLL rearrangement might be a good target
population to further investigate priming strategies.
Conclusion: Priming with growth factors remains an active
field of clinical investigation; it cannot be recommended
in routine practice. Level of evidence II

References :
1. The AML Collaborative Group. A systematic
collaborative overview of randomized trials
comparing idarubicin with daunorubicin (or other
anthracyclines) as induction therapy for acute
myeloid leukaemia. Br J Haematol. 1998;103:100-
109.
2. Daunorubicin Versus Mitoxantrone Versus Idarubicin
As Induction and Consolidation Chemotherapy for
Adults With Acute Myeloid Leukemia: The EORTC and

164
 GIMEMA Groups Study AML-10, J Clin Oncol. 2009:
27 :5397-.5401
3. Fernandez HF, Sun Z, Yao X, et al. Anthracycline dose
intensification in acute myeloid leukemia. N Engl
JMed. 2009;361:1249-1259.
4. Löwenberg B, Ossenkoppele GJ, van Putten W, et al.
High-dose daunorubicin in older patients with acute
myeloid leukemia. N Engl J Med. 2009;361:1235-
1248.
5. Weick JK, Kopecky KJ, Appelbaum FR, et al. A
randomized investigation of high-dose versus
standard-dose cytosine arabinoside with
daunorubicin in patients with previously untreated
acute myeloid leukemia: a Southwest Oncology
Group study. Blood. 1996;88(8):2841-2851.
6. Bishop JF, Matthews JP, Young GA, et al. Randomized
study of high-dose cytarabine in induction in acute
myeloid leukemia. Blood. 1996;87(5):1710-1717.
7. Preisler H, Davis RB, Kirshner J, the Cancer and
Leukemia Group B, et al: Comparison of three
remission induction regimens and two post induction
strategies for the treatment acute non-lymphocytic
leukemia: A Cancer and Leukemia Group B study.
Blood 1987; 69:1441.
8. Bishop JF, Lowenthal RM, Joshua D, et
al: Etoposide in acute non-lymphocytic leukemia.
Australian Leukemia Study Group. Blood 1990;
75:27.
9. Hann IM, Stevens RF, Goldstone AH, et
al: Randomized comparison of DAT versus ADE as
induction chemotherapy in children and younger
adults with acute myeloid leukemia: Results of the
Medical Research Council’s 10th AML trial (MRC
AML.10). Blood 1997; 89:2311
165
10. Lo¨wenberg B, van Putten W, Theobald M, et al.Effect
of priming with granulocyte colony-stimulating
factor on the outcome of chemotherapy for acute
myeloid leukemia. N Engl J Med. 2003; 349(8):743-
752.
11. Thomas X, Raffoux E, de Botton S, et al. Effect of
priming with granulocyte-macrophage colonystimu-
lating factor in younger adults with newly diagnosed
acute myeloid leukemia: a trial by the Acute Leukemia
French Association (ALFA) Group. Leukemia.
2007;21(3):453-461.

166
 Optimising Post-induction
therapy in AML patients
Dr. Hari Menon,
Dr. Sourabh Radhakrishnan

Introduction
Acute myeloid leukaemia is a heterogeneous group of
clonal disorders involving maturation at early phases of
hemopoietic differentiation leading to accumulation of
abnormal cells in marrow and eventually causing a bone
marrow failure state. Insight into its pathogenesis and
recognition of both AML specific cytogenetic and
molecular abnormalities as prognostic markers has been
a major advance influencing response and outcomes to
therapy. Though the last few decades has witnessed little
change in induction regimen used in AML, the prognostic
value of cytogenetic and molecular profile has strongly
influenced the strategy for optimal post remission
consolidation therapy.
The choices for optimal post remission therapy are made
based on the probability of future relapse. The low risk
patient traditionally receives only cytotoxic chemotherapy
while the high risk patients are candidates for allogeneic
transplantation in first complete remission (CR1). Current
167
recommendations for post- remission therapy depends on
whether the relapse risk is high enough to merit the
potential toxicities of allogeneic transplantation, or
sufficiently low to avoid the procedure altogether. The key
in selecting the optimal post-remission therapy for each
patient has been the advances in risk stratification and
understanding of the molecular basis of AML.

Risk stratification in AML- The role of

clinical, cytogenetic and molecular profile
It is a well understood fact that advanced age, poor
performance status, leucocytosis, dyspoiesis, and a lack
of induction remission impacts treatment outcomes
adversely. However, it is the cytogenetic profile, ascertained
at the time of diagnosis that remains the most important
prognostic factor predicting outcomes after achieving a
remission post induction therapy especially in the younger
AML patient (< 60 years). Classified as good, intermediate
and poor cytogenetic risk group, the majority of the AML
patients fall into the intermediate risk group that essentially
have no cytogenetic abnormality. The 5 year survival stands
at 65% for the good risk group while dropping to less
than 20% for the poor risk.
Though cytogenetic profile is a powerful indicator of
outcomes, it remains imprecise in determining relapse.
The understanding of the heterogeneity in the molecular
markers has refined the stratification process in AML. A
number of genes are mutated, aberrantly over expressed,
or aberrantly silenced in AML. This is true across all
cytogenetic subgroups but has a particular impact in
further delineating the intermediate risk group. Internal
tandem duplication (ITD) of the fms-like tyrosine kinase
3 gene (FLT3) is particularly relevant. Their presence
adversely affects outcome, particularly when lacking a copy

168
of the wild type allele. Another mutation of the
nucleophosmin ( NPM1) gene coexists with FLT3 mutation.
Their presence is associated with a more favourable
outcome. Among patients receiving an allogeneic
transplant from a matched sibling, no benefit of
transplantation in CR1 was seen in presence of a NPM1-
mutated/ FLT3-ITD negative state. Conversely, benefit from
transplantation was seen in patients who were either FLT3-
ITD-positive or NPM1 wild type/FLT3-ITD-negative. A
number of other mutated or aberrantly over expressed
genes have been identified with a negative prognostic
value (Table-1). These include partial tandem duplications
Table -1Prognostically significant genetic aberrations in
cytogenetically normal acute myeloid leukemia
Molecular alteration Location Prognostic Impact
FLT3-ITD 13q12 OS: significantly shorter
DFS: significantly shorter
CRD: significantly shorter
ERG overexpression. 21q22.3 OS: significantly shorter
CIR: significantly shorter
BAALC overexpression 8q22.3 OS: significantly shorter
DFS: significantly shorter
EFS: significantly shorter
NPM1 mutations 5q35 OS: significantly longer
CR rate: significantly higher
CEBPA mutations 19q13.1 OS: significantly longer
DFS: significantly longer
CRD: significantly longer
MLL-PTD 11q23 OS: significantly shorter
EFS: significantly shorter
RD: significantly shorter
CR: complete remission, CIR: cumulative incidence of relapse, CRD:
complete remission duration, DFS: disease free survival, EFS: event-free
survival, OS: overall survival. Modified from Baldus CD et al. Clinical
outcome of de novo acute myeloid leukaemia patients with normal
cytogenetics is affected by molecular genetic alterations: a concise review.
Br J Haematol 2007;137:387-400.

169
of MLL (MLL-PTD), and over-expression of WT1, BAALC,
ERG, or EVI1. Other abnormalities, such as CEPBA
mutations, are associated with a more favourable
outcome.

Therapy of AML patient in CR-1 - Post

remission chemotherapy
The median disease free survival in patients not receiving
post remission therapy after induction remission is only 4
to 6 months. Hence the role of post remission
consolidation therapy for AML is undisputed. In younger
patients, there is a clear benefit from intensive post-
remission therapy, but many questions remain about the
best type, dose, and duration of treatment. There have
been several strategies adopted to identify the most
optimal and effective post remission therapy. The practice
has been to either resort to the same regimen as one would
for induction or use high dose cytosine arabinoside in
doses ranging from anywhere between 200mg/m2 to 18
and 32gms/m2. Despite years of experience in this area,
various co-operative groups, except in a few select subsets
of disease, have found it difficult to advocate one approach
as superior to another.
The only convincing concept of therapy that has emerged
in the past few years is that repeated cycles of high dose
cytarabine (HIDAC) should be administered to patients
with core binding factor (CBF) AML, a subset of AML
consisting of the good risk karyotypes t(8;21) and inv(16).
This aspect was evidenced in the CALGB study involving
596 young AML patients. Patients achieving complete
remission with standard 7+3 induction were randomized
to HIDAC (3 g/m2 every 12 hours on days 1, 3 and 5), low
dose cytarabine (100 mg/m2/day for 5 days as a
continuous intravenous infusion) or intermediate dose

170
cytarabine (400 mg/m2/day for 5 days as a continuous
intravenous infusion). The 4-year disease-free survival was
superior in patients who received HIDAC. This benefit was
particularly evident for patients with CBF AML. Repeated
two or more cycles of HIDAC consolidation therapy
produced prolonged disease-free survival in patients with
CBF AML and HIDAC had a similar impact on the normal
karyotype AML too. Those with other cytogenetic
abnormalities did not derive similar benefits. Another
concept that has emerged is the presence of C-Kit which
renders poor outcome in patients with CBF AML, their
favourable risk notwithstanding. This subset would require
alternative post remission strategies for better outcomes.
The role of multiagent chemotherapy as post remission
therapy has also been looked into in the CALGB 9222
study. This study randomized patients to receive one of
two different consolidation regimens. The first regimen
consisted of three courses of HIDAC. The second regimen
consisted of one course of HIDAC, a second course with
etoposide and cyclophosphamide, and a third course with
diaziquone and mitoxantrone. There was no DFS benefit
with either regimen. The combination only contributed
to increased toxicity.

Therapy of AML patient in

CR-1 – Hemopoietic stem cell transplant
The utility of HSCT as consolidation therapy has been
demonstrated since the 1980s and early 1990. Studies
showed lower relapse rates and improved DFS with
allogeneic HSCT in AML patients in CR1. However they
never conclusively demonstrated a survival advantage.
Several groups have published their data on the impact
of HSCT in CRI for AML which is discussed in Table-2.

171
 Table-2 - Studies on HSCT for AML in CR1
Study Result/ Inference
EORTC/GIMEMA a) Superior 4-year leukaemia free survival (LFS)
with allogeneic (55%) and autologous HSCT
(48%) compared to Intensive chemotherapy
(30%).
b) Despite this improved LFS and higher relapse
rate in patients getting intensive chemotherapy
(ICC), OS was similar in the three groups, since
majority of patients relapsing after ICC were
successfully salvaged with autologous HSCT
GOELAM a) Relapse rate after allogeneic HSCT was
unusually high (37% at 4 years) negating any
benefit with this approach.
b) Autologous HSCT showed no significant
benefit in LFS over ICC.
MRC AML10 a) Lower relapse rate and significantly better LFS
and OS following autologous HSCT
b) Better relapse rate and LFS, but not OS for
allogeneic transplant
US intergroup a) Lower relapse rates and higher treatment
related mortality (TRM) in patients randomized
to allogeneic or autologous HSCT
b) No significant difference in LFS between
patient undergoing HSCT and ICC.
HOVON-SAKK a) There was an overall survival benefit from
(Meta-analysis allogeneic transplantation in CR1 from a
of the European matched sibling donor, with myeloablative
co-operative conditioning, of 12% for patients in
group studies) intermediate or poor risk cytogenetic groups.
b) The benefit was lost in older patients (>35
years), likely due to increased transplant-
related mortality with increasing age.
c) Younger patients with an HLA- matched sibling
donor who are fit and not in the good risk
cytogenetic group should undergo allogeneic
transplantation in CR1.

172
The consistent theme of these trials for HSCT in CR-1 has
been the lower relapse rates with comparable and at times
improved LFS in patients undergoing HSCT compared to
ICC. Perhaps a higher TRM with transplantation and an
effective salvage of patient randomized to ICC with
autologous HSCT following relapse prevented an OS
benefit of transplantation from being demonstrated.
Eventually these data provide no concrete guidelines for
selecting the best consolidation therapy for an individual
patient.

Impact of of cytogenetics in defining HSCT in CR1

The role of transplant in the good risk and poor risk group
is well defined with its role limited in the former while
being a necessity in the latter for durable remissions. It is
in the intermediate group that a consensus has failed to
evolve despite several groups actively looking into this
subset. Results of EORTC/GIMEMA AML-10 trial showed
no difference between allogeneic and autologous SCT in
terms of LFS or OS for patients with good-risk or
intermediate-risk cytogenetics. However for patients with
poor-risk cytogenetics allogeneic HSCT produced
significantly superior LFS (43 vs. 18%) and OS (50 vs. 29%).
The same results came out of the US intergroup study too
which also suggested that allografting in the intermediate
risk group may not be of benefit in terms of overall survival.
This was contradicted in the MRC-10 data which however
suggested a benefit for the intermediate risk group for
allogeneic transplant. The data was however criticized for
having included certain poor risk group patients into the
intermediate group. The Dutch–Belgian haemato-
oncology co-operative group (HOVON) and Swiss group
for clinical cancer research (SAKK) trial demonstrated
superior LFS with allogeneic HSCT for both intermediate
and poor-risk groups. Currently recognition of the

173
prognostic value of additional molecular markers is
facilitating further risk stratification of patients in the
intermediate-risk cytogenetic group.

Unrelated donor transplantation for AML in CR1?

No prospective trials have looked at unrelated donor (URD)
HSCT for AML in CR1. Reports have indicated similar rates
of acute GVHD, TRM, relapse rate and OS in patients with
standard-risk undergoing HLA-identical sibling
transplantation versus HLA-allelic matched URD (10/10)
transplant. Tallman et al. analysed outcomes of URD
transplantation in AML-CR1 patients according to
cytogenetic-risk profiles in a CIBMTR/NMDP study [51].
The 5-year OS, LFS and relapse rate for patients with high-
risk cytogenetics were 31, 31 and 23%, respectively. The
31% OS of this groups of patients compares favourably
with 13–15% survival reported with autologous-HSCTand
ICC. Thus it may certainly be reasonable to consider
matched URD transplantation to AML-CR1 patients with
high-risk cytogenetics.

Autologous transplant in CR1

There is no definitive data indicating that this approach is
superior to ICC. The EORTC- GIMEMA study showed
superior LFS compared to those receiving ICC, but many
ICC patients who relapsed actually underwent late
autotransplant and there was no survival advantage. The
GOELAM trial also showed no benefit in LFS for auto-
transplant compared to ICC. Currently the use of ASCT in
CR1 may be considered unjustified owing to the
compromise it may cause to future allogeneic SCT at the
time of relapse. A possible exception might be patients
with MLL-PTD, wherein a CALGB study suggested that
autografting in CR1 can overcome the poor-prognostic
impact of this genomic abnormality.

174
Conclusions
The understanding of the heterogeneity and molecular
make up of AML patients have helped in defining what
treatment is to be given for optimal results. The
augmentation of cytogenetic risk stratification by
molecular testing in patients with normal karyotype could
be the next improvement in post remission strategies to
treat AML. The current data on post-remission therapy
for AML patients in CR1 suggest that dose modification
or intensification of presently available cytotoxic drugs is
unlikely to improve outcomes further. It is only the newer
agents targeting the pathways that may potentially
improve outcomes when incorporated into standard
therapy.

Suggested reading
1. Mrozek K, Heerema NA, Bloomfield CD. Cytogenetics
in acute leukemia. Blood Rev 2004; 18(2): 115–136
2. Cassileth PA, Harrington DP, Appelbaum FR, Lazarus
HM, Rowe JM, Paietta E, et al. Chemotherapy
compared with autologous or allogeneic bone
marrow transplantation in the management of acute
myeloid leukemia in first remission. N Engl J Med
1998; 339(23): 1649–1656.
3. British Committee for Standards in Haematology.
Milligan DW, Grimwade D, Cullis JO, Bond L, Swirsky
D, et al. Guidelines on the management of acute
myeloid leukaemia in adults. Br J Haematol 2006;
135(4): 450–474.
4. Mayer RJ, Davis RB, Schiffer CA, Berg DT, Powell BL,
Schulman P, et al. Intensive postremission
chemotherapy in adults with acute myeloid leukemia.
Cancer and Leukemia Group B. N Engl J Med 1994;
331(14): 896–903.

175
5. Cornelissen JJ, van Putten WL, Verdonck LF, Theobald
M, Jacky E, Daenen SM, et al. Myeloablative HLA-
identical sibling stem cell transplantation in first
remission acute myeloid leukemia in young and
middle aged adults: benefits for whom? results of a
HOVON/SAKK donor versus no donor analysis. Blood
2007; 109(9): 3658–3666.
6. Burnett AK, Wheatley K, Goldstone AH, Stevens RF,
Hann IM, Rees JH, et al. The value of allogeneic bone
marrow transplant in patients with acute myeloid
leukaemia at differing risk of relapse: results of the
UK MRC AML 10 trial. Br J Haematol 2002; 118(2):
385–400.
7. Whitman SP, Ruppert AS, Marcucci G, Mrozek K,
Paschka P, Langer C, et al. Long-term disease-free
survivors with cytogenetically normal acute myeloid
leukemia and MLL partial tandem duplication: a
Cancer and Leukemia Group B study. Blood
2007;109: 5164-7.
8. Harousseau JL, Cahn JY, Pignon B, Witz F, Milpied N,
Delain M, et al. Comparison of autologous bone
marrow transplantation and intensive chemotherapy
as postremission therapy in adult acute myeloid
leukemia. The Groupe Ouest Est Leucemies Aigues
Myeloblastiques (GOELAM). Blood 1997;90:2978-
86.
9. Cornelissen JJ, van Putten WL, Verdonck LF, Theobald
M, Jacky E, Daenen SM, et al. Results of a HOVON/
SAKK donor versus no-donor analysis of
myeloablative HLA-identical sibling stem cell
transplantation in first remission acute myeloid
leukemia in young and middle-aged adults: benefits
for whom? Blood 2007;109:3658-66.

176
 Current state of HSCT and
Management of relapsed/
refractory AML
Dr. Vikas Ostwal, Dr. Navin Khattry

Introduction
Cancer and Leukemia Group B (CALGB) established the
standard induction regimen for AML with which, 60% to
80% of young adults and 40% to 60% of older adults can
achieve a complete remission ( CR). Despite initial high CR
rates, the overall (OS) is unsatisfactory (1). As there are no
randomized trials to demonstrate improved overall survival
in AML relapse/ refractory patients, multiple retrospective
studies and case series form the basis of treatment for
this group of patients. Patients of age less than 55 years
with HLA matched siblings who relapse after an initial
remission in almost all cases should be considered for
allogenic transplant. Occasional relapsed patients with first
remissions of long duration and favorable cytogenetic may
be treated with further chemotherapy (2).
Results of allogeneic SCT in relapsed AML:
As the results of only chemotherapy in relapsed AML are
poor, long-term disease-free survival rates of 30–40% are
achievable in patients with AML in second CR who undergo
177
an allogenic-HSCT from a human leucocyte antigen (HLA)
identical sibling donor (3,4,10) . The European registry
reported a 35% 3-year disease-free survival rate among
459 patients transplanted from matched siblings for AML
in second remission (4). Patients who lack an HLA-identical
sibling are allografted using matched unrelated donor
(MUD) with long-term disease-free survival rates in excess
of 30 %(5). There are no randomized studies comparing
chemotherapy alone with autologous or allogeneic
transplantation. A large retrospective analysis of outcomes
of patients in first relapse by HOVON-SAKK Cooperative
group showed the results of treatment were dependent
on a prognostic index(6), (table1).
Table 1:
Prognostic Index for Adult Patients with
Acute Myeloid Leukemia in First Relapse:
Relapse-free interval (RFI) from first Points
complete remission in months (score)
>18 months 0
7-18 months 3
< 6 months 5
Cytogenetics at diagnosis
t (16;16), inv 16 0
t(8;21) 3
Others 5
Age at first relapse
</=35 0
36-45 1
>/=45 2
Stem cell transplantation before first relapse
No 0
Previous SCT(auto/allo) 2
Dimitri A. Breems, Wim L.J et al, . Clin Oncol 23:1969-1978.
© 2005 by American Society of Clinical Oncology

178
A total of 1,540 non-M3 AML patients were enrolled
during the period 1987 to 2001 onto three successive
Dutch-Belgian Hemato-Oncology Cooperative Group and
Swiss Group for Clinical Cancer Research phase III trials(6).
An analysis of cohort of 667 patients with AML in first
relapse from the above studies with a median follow-up
of 56 months were included to form the above prognostic
index. The overall survival (OS) probabilities were studied
in relation to relapse free interval (RFI) after first CR,
cytogenetics at diagnosis, age at first relapse, and whether
SCT had been undertaken before first relapse.
Table 2:
Prognostic OS at OS at Treatment No. of % OS % OS
Group 1 year 5 year Patients 1-Year 5-Year

Favorable-risk 70 % 46 % Chemotherapy 14 64 33
group A Autologous SCT 14 100 55
Score (1-6) Allogenic SCT 17 88 88
Intermediate- 49 % 18 % Chemotherapy 28 71 31
risk group B Autologous SCT 22 73 0
Score (7-9) Allogenic SCT 41 75 48
Poor-risk 16% 4% Chemotherapy 34 49 6
group C Autologous SCT 28 54 9
Score (10-14) Allogenic SCT 51 55 26
Dimitri A. Breems, Wim L.J et al, . Clin Oncol 23:1969-1978.
© 2005 by American Society of Clinical Oncology

All 3 groups showed improved 5 year OS with allogenic

stem cell transplantation see table 2(6).

Role of autologous SCT in patients with

relapsed AML
Autologous SCT produces durable second remission in
25-30% of patients with relapsed AML who achieve a
second CR but its low transplant related mortality is offset
by a high relapse rate (7). In the absence of randomized
data comparing it with allo-SCT, auto-SCT is currently

179
restricted to older patients (in whom a myeloablative
allograft is precluded) and younger patients lacking a
sibling or unrelated donor. In patients with a long first
CR, auto-SCT may produce results equivalent or superior
to allo-SCT, mainly unrelated donor.
The results of autologus SCT in patients with AML-M3
transplanted in second CR are improving particularly if
both patient and stem cell graft have no evidence of
minimal residual leukemia(8). Although based on small
numbers, autologus transplant is the preferred transplant
option in all patients with relapsed AML-M3, who have
achieved a molecular CR, provided an uncontaminated
stem-cell product is available. Allo-SCT should probably
be reserved for patients with persistent residual disease
prior to transplantation (9).

Recommendations of treatment for

relapsedAML (10, 11) :
There is no significant advantage of autologous SCT over
chemotherapy.
There is a survival advantage for allogeneic SCT vs.
chemotherapy for patients under age 55.
An HLA-matched related donor allogeneic SCT is
recommended over autologous SCT, if a matched related
donor is available.
For matched unrelated donor allogeneic SCT, there are
insufficient data to make a recommendation over
autologous SCT.
Autologous peripheral blood stem cell transplant (PBSCT)
is recommended over autologous bone marrow transplant
(BMT) due to improvements in safety and early mortality.
The impact of autologous PBSCT on overall survival is not
known.

180
For high risk disease, allogeneic PBSCT is recommended
over BMT. For low risk disease, allogeneic PBSCT and BMT
have equivalent outcomes.

Treatment for Refractory AML- Experience of

other centers across the world:
The definition of refractory AML is variable; most of the
studies have taken following points in consideration:
Patients were included if they fulfilled at least one of the
following criteria defining refractory AML(12): (1) primary
induction failure (PIF) after 2 or more cycles of
chemotherapy, (2) first early relapse after a remission
duration of fewer than 6 months, (3) relapse / refractory
to salvage combination chemotherapy containing high-
dose AraC, and (4) second or subsequent relapse.
The largest single institution and the best described series
published in manuscript form are from the City of Hope
Comprehensive Cancer Center (CHCCC) updated from their
series published a decade prior. Between 1978 and 2000,
68 patients with primary refractory AML proceed to allo-
SCT, the data reveals several relapses between the third
and fourth year, making the 4 year disease-free survival
close to 20%. An important finding in multivariate analysis
was that unfavorable karyotype and the use of an
unrelated donor predicted for a worse result. An impressive
44% of patients with intermediate risk karyotype remain
alive without disease at 3 years compared to 18% for
patients with an unfavorable karyotype. Of 12 patients,
who had an unrelated donor as the stem cell source, 11
were dead or relapsed within a year of transplant (13).
The Socie´te´ Franc¸aise de Greffe de Moelle (SFGM)
published a series of the use of allo-SCT for 379 patients
with advanced AML (14). In their series, 69 patients were

181
transplanted with primary resistant disease. The 5-year
probability of survival was 13.78%.
In 1992, the IBMTR published on a series of patients who
received HLA-identical sibling bone marrow transplants
for refractory acute leukemia(15). In all, 88 of the patients
had AML. The 3-year probability of leukemia-free survival
was 21% (range 14– 31%).
The European Group for Blood and Bone Marrow
Transplantation (EBMT) recently reported the results of the
346 patients with primary refractory disease proceeded
to matched sibling allo-SCT at a median of 136 days after
initial diagnosis. No information concerning induction
therapy, number of chemotherapies and blast percentage
was given. With a median follow-up of 22 months, the
leukemia-free survival was 18.72% at 2 years (12). Relapse
incidence was 57.76% and treatment-related mortality was
25.75% (16).
The MD Anderson Cancer Center recently published a
paper examining their results of patients who underwent
Allo-SCT for relapsed or refractory AML or myelodysplastic
syndrome (MDS) (17) . In their description of patient
characteristics of the 135 patients, 53 patients are
considered to have primary induction failure. At the time
of transplantation, 39.3% of patients had not responded
to induction therapy, 37% had not responded to first
salvage therapy, and 23.7% were beyond first salvage.
Forty-one patients (30%) received unrelated donor
progenitor cells. Eighty patients (59%) received either a
reduced-intensity or a nonmyeloablative regimen. A total
of 104 (77%) of 135 patients died, with a median survival
time of 4.9 months (95% confidence interval, 3.9-6.6
months). The median progression-free survival was 2.9
months (95% confidence interval, 2.5-4.2 months). A Cox

182
regression analysis showed that Karnofsky performance
status, peripheral blood blasts, and tacrolimus exposure
during the first 11 days after transplantation were
predictive of survival.

Recommendations for refractory AML:

The use of allogenic transplantation for patients with
refractory acute myelogenous leukemia/myelodysplastic
syndromes is recommended

References:
1. Burnett, A.K (2001) Introduction: modern
management of acute myeloid leukemia. Seminars
in Hematology 2001; 38: 381–382
2. Gale, R.P Et al. (1996) Chemotherapy versus
transplants for acute myelogenous leukemia in
second remission. Leukemia 1996;10:13–19
3. Leopold LH, Willemze R: The treatment of acute
myeloid leukemia in first relapse: A comprehensive
review of the literature. Leuk Lymphoma 43:1715-
1727, 2002
4. Reiffers, J. (2000) HLA-identical sibling
haematopoietic stem cell transplantation for acute
myeloid leukaemia. In: Clinical Bone Marrow and
Blood Stem Cell Transplantation (ed. by K. Atkinson),
pp. 433–445. Cambridge University Press,
Cambridge.
5. Sierra, J., Storer, B., et al (2000) Unrelated donor
marrow transplantation for acute myeloid leukemia:
an update of the Seattle experience. Bone Marrow
Transplantation, 26, 397–404
6. Prognostic Index for Adult Patients With Acute
Myeloid Leukemia in First Relapse, Dimitri A. Breems

183
 et al, From the Dutch-Belgian Hemato- Oncology
Cooperative Group (HOVON); Swiss Group for
Clinical Cancer Research Collaborative Group (SAKK);
J Clin Oncol 23:1969-1978. © 2005
7. Tomas (1996) Autologous or allogeneic bone marrow
transplantation for acute myeloblastic leukemia in
second complete remission. Importance of duration
of first complete remission in final outcome. Bone
Marrow Transplantation 1996; 17: 979–984
8. Meloni (1997) Autologous bone marrow
transplantation for acute promyelocytic leukemia in
second remission: prognostic relevance of
pretransplant minimal residual disease assessment
by reverse-transcription polymerase chain reaction
of the PML/RAR alpha fusion gene. Blood, 90, 1321–
1325.
9. Lo Coco (1999) Therapy of molecular relapse in acute
promyelocytic leukemia. Blood, 94, 2225–2229.
10. Oliansky D, Appelbaum F, Cassileth P, et al. The role
of cytotoxic therapy with hematopoietic stem cell
transplantation in the therapy of acute myeloid
leukemia in adults: an evidence-based review. Biol
Blood Marrow Transplant. 2007;14:135-136.
11. An evidence-based review. Biol Blood Marrow
Transplant. 2007;14:135-136.2008 American
Society for Blood and Marrow Transplantation
12. Mehta J, Powles R, Horton C et al. Bone marrow
transplantation for primary refractory acute
leukaemia. Bone Marrow Transplant 1994; 14: 415–
418
13. Fung HC, et al. A long-term follow-up report on
allogeneic stem cell transplantation for patients with
primary refractory acute myelogenous leukemia:

184
 impact of cytogenetic characteristics on
transplantation outcome. Biol Blood Marrow
Transplant 2003; 9: 766–771
14. Michallet M, et al. Long-term outcome after
allogeneic hematopoietic stem cell transplantation
for advanced stage acute myeloblastic leukemia: a
retrospective study of 379 patients reported to the
Societe Francaise de Greffe de Moelle (SFGM). Bone
Marrow Transplant 2000; 26:1157–1163
15. Biggs JC, Horowitz MM, Gale RP et al. Bone marrow
transplants may cure patients with acute leukemia
never achieving remission with chemotherapy. Blood
1992; 80:1090–1093
16. Esteve J, Labopin M, Finke J et al. Allogeneic stem-
cell transplantation for patients with de novo acute
myeloid leukemia not in complete response: results
of a survey from the European Group for Blood and
Bone Marrow Transplantation (EBMT). Blood 2004;
104: 633a
17. Wong R, Shahjahan M, Wang X et al. Prognostic
factors for outcomes of patients with refractory or
relapsed acute myelogenous leukemia or
myelodysplastic syndromes undergoing allogeneic
progenitor cell transplantation. Biol Blood Marrow
Transplant 2005; 11: 108–114

185
 PEDIATRIC AML:
Differences And Similarities
Dr. Akash Nahar, Dr. Pandeep Singh

Introduction
Acute myeloid leukemia (AML) is a heterogeneous disease
entity in children and comprises about 15 – 20% of acute
leukemia in children (1). In contrast to acute lymphoblastic
leukemia it has a poor prognosis. Therapeutic approaches
to children with AML have continued to emerge from the
largely successful, empirically developed, approaches to
therapy for childhood ALL. Despite above developments,
5-year event-free survival (EFS) of children with AML is
only 50% to 60% (2).Due to dismal results; there have been
developments in almost every aspect of this disease so as
to improve its prognosis. This review summarizes the
differences and similarities of pediatric AML with respect
to their classification, diagnosis, newer chemotherapeutics
and risk based approach to treat pediatric acute myeloid
leukemia.
(3)
Etiology & Predisposing Factors
Majority of children have de novo AML with no identiûable
predisposing factor, although a number of environmental
exposures, inherited conditions, and acquired disorders
are associated with the development of AML.
186
Enviromental: Chemotherapy (Alkylators and
topoioisomerase II inhibitors), Pesticides, Petroleum
products and Ionizing radiation.
Inherited: Down syndrome, Fanconi anemia, Severe
congenital neutropenia, Kostmann
Syndrome, Shwachman-Diamond syndrome,
Diamond-Blackfan syndrome, Dyskeratosis
congenita, Congenital amegakaryocytic
thrombocytopenia, Aaxia-telangiectasia and
Klinefelter’s syndrome.
Acquired: Aplastic anemia, Myelodysplastic syndrome
and Paroxysmal nocturnal hemoglobinuria.

Classification of AML
The WHO 2008 classification, includes morphologic
findings (FAB subtypes), genetic, immunophenotypic,
biological and clinical features to define specific disease
entities. (Table1). Though the present classification is for
adult population, it is being incorporated slowly in the
practice of pediatric oncology.
Current Classification of AML (WHO 2008) (Table I) (4)

1. AML with recurrent genetic abnormalities:

 AML with t (8; 21) (q22; q22), RUNX1-
RUNX1T1 (CBFA/ETO).
 AML with inv (16) (p13; q22) or t (16; 16) (p13;
q22), CBFB-MYH11.
 Acute promyelocytic leukemia with t (15; 17)
(q22;q11-12), PML-RARA.
 AML with t (9; 11)(p22;q23), MLLT3-MLL.
 AML with t (6; 9)(p23;q34); DEK-NUP214.
 AML (megakaryoblastic) with t (1;22)(p13;q13),
RBM15-MKL1.
187
  AML with mutated NPM1.
 AML with mutated CEBPA.
2. AML with myelodysplasia-related features.
3. Therapy-related myeloid neoplasms.
4. AML, not otherwise specified:
 AML with minimal differentiation.
 AML without maturation.
 AML with maturation.
 Acute myelomonocytic leukemia.
 Acute monoblastic and monocytic leukemia.
 Acute erythroid leukemia.
 Acute megakaryoblastic leukemia.
 Acute basophilic leukemia.
 Acute panmyelosis with myelofibrosis
5. Myeloid sarcoma
6. Myeloid proliferations related to Down syndrome:
a. Transient abnormal myelopoiesis.
b. Myeloid leukemia associated with Down
syndrome
7. Blastic plasmacytoid dendritic cell neoplasm.
Clinical Features and Diagnosis (5)
Children with AML can present with exceptionally varied
symptoms and signs. They commonly present with features
of bone marrow infiltration such as fever, fatigue, pallor,
bleeding, bone pain, and infections. Patients who have
high white blood cells counts may present with signs or
symptoms of leukostasis, most often affecting the lung
and brain. Inûltration of extramedullary sites can result in
lymphadenopathy, hepatosplenomegaly, chloromatous
tumors, leukemia cutis, orbit, epidural space, and, rarely,
testicular involvement. The diagnosis and subtype

188
classiûcation of AML is based on morphologic,
cytochemical, ûuorescent in situ hybridization analyses,
ûow cytometric immunophenotyping, and molecular
testing. The cytogentics risk group stratification varies
differently between different pediatric AML study groups.
Recent cytogenetic classification as proposed by NCCN
2010 guidelines is as follows

A. Favourable cytogenetics
The t (8; 21) and inv (16) are two of the most commonly
identified translocations in pediatric AML and are often
referred to as CBF (Core binding factor) leukemias. MRC
AML 10 trial demonstrated that 17% of the pediatric
patients had CBF leukemia (12% with t (8; 21) and 5%
with inv (16). Patients with t (8; 21) and inv (16) had
significantly better (OS) overall survival (5 yr OS, 69 & 61%
respectively) than patients with the normal karyotype (7).
AML- BFM 98 group recently confirmed a favorable
outcome in patients with t (8; 21), inv (16), and t (15;
17), with an OS of 91%, 92% and 87% respectively (8).
(Level II, Grade A).
189
B. Intermediate cytogenetics
A rearrangement of the MLL gene, located at chromosome
band 11q23 is seen in as many as 20% of cases of pediatric
AML, although the reported frequency varies (9). Among,
patients treated for AML at St. Jude, those who had t (9;
11) had a favorable prognosis (5-year EFS estimate, 65%)
(9).
In BFM-98 study, outcome of patients with t (9; 11)
(p22; q23) and t (11; 19) (q23; p13), in contrast to patients
with other MLL rearrangements, was comparable to that
of the other patients in intermediate risk group (8). (Level
II, Grade A).

C. Unfavourable cytogenetics
An abnormality of chromosome 5 and 7 has been
identified in a small proportion of pediatric patients and
is associated with poor outcome. Because of the rarity of
this abnormality, an international collaborative study was
undertaken in 258 children and adolescent to characterize
further the impact of -7 and del (7q). Patients with
monosomy 7 had lower CR rates (61% versus 89%) and
worse outcome (5-year OS 30% vs 51%) than those who
had del (7q) (10). (Level II, Grade B).

Novel Diagnsotic Molecular Markers

Significant advances in the molecular diagnostics
technique are yielding fascinating inputs for the treatment
of cytogenetically normal pediatric AML. Some of the
molecular markers studied recently in clinical trials are
described below.

A. FLT3 Mutations
(ITD-internal tandem duplication & ALM – activating loop
mutation)

190
FLT3/ITD has a prevalence of 12%–15% in pediatric
patients, 20%–25% in young adults and nearly 35% in
the elderly population (11,12). Meshinchi et al. suggested
that stem-cell transplant (SCT) might improve outcome in
children with FLT3/ITD positive AML. In their study,
multivariate analysis did not show any prognostic benefit
of SCT in first CR (11). This is in line with earlier publications
from the AML-BFM group, in which no clear benefit of
SCT in 1st CR in childhood AML is found. (Level II, Grade B).

B. c-KIT mutations
The prognostic significance of c-KIT mutations occurring
in pediatric core-binding factor leukemias remains unclear.
The Japanese Childhood AML Cooperative Study Group
found c-kit mutations (8 of 46 children) with t (8; 21),
were associated with significantly worse OS, DFS, and
relapse rates (13). (Level II, Grade B).

C. NPM1: Nucleophosphomin
NPM mutations have been reported in 30%–50% of adult
patients, it is prevalent in only 10% of children (14).
Evaluation of the prognostic significance of NPM
mutations in children with AML treated in the CCG 2961
trial failed to demonstrate any prognostic significance for
this mutation in pediatric AML (15). (Level II, Grade C).

D. CEBPA - (CCAAT/enhancer binding protein-

alpha)
The prevalence of CEBPA mutations in pediatric AML is
somewhat lower as compared to adults (< 10%) (14).
Phoenix et al evaluated prognostic signiûcance of CEBPA
mutations in 847 children with AML treated on 3
consecutive pediatric trials (16). Actuarial event free survival
at 5 years was 70% versus 38% (P= 0.015) with a
cumulative incidence of relapse from CR of 13% versus

191
44% (P= 0.007) for those with and without CEBPA
mutations. (Level II, Grade C).

Treatment
Therapeutic approaches to children with AML have
continued to emerge from the largely successful,
empirically developed approaches to therapy for childhood
ALL. Despite above developments, 5-year event-free
survival (EFS) of children with AML is only 40% to 50% (3).

Induction Therapy
The primary goal of induction therapy is to achieve a
significant reduction of leukemia burden i.e. achievement
of morphological CR (Clinical remission). The most
favorable outcomes are achieved by the use of a relatively
high cumulative dose of either anthracycline or cytarabine.
Combination studies of cytarabine and anthracyclines in
adults led to the adoption of the ‘7 + 3’ strategy of
combining cytarabine (typically 100 mg/m 2 /day)
administered by continuous infusion over 7 days) with an
anthracycline (daunorubicin 45 mg/m2 given on days 1 –
3) in many trials involving children in the US and in
Europe(17). Intensification can be achieved by the use of
high-dose chemotherapy, early administration of
subsequent courses of therapy or protracted
administration of chemotherapy. All three approaches have
been explored in Phase III trials in children. The POG
compared high dose cytarabine (1 gm/m2) to standard
dose cytarabine in 3+7 regimen. Although the tolerability
was fair to high dose, it failed to improve response rate or
event free survival (18). The Children’s Cancer Group (CCG)
reported that intensively timed induction therapy (the
second cycle delivered 10 days instead of 14 days after
the ûrst cycle) was more advantageous than standard

192
therapy (significantly higher CR & EFS rates) (19). On the
contrary intensification by protracted cytarabine
administration (over 8 to 10 days) is favored by UK-MRC
group. Certain anthracyclines (idarubucin & mitoxantrone)
are favored for their perceived greater antileukemic effect
and/or their lower cardiotoxicity, but no anthracycline
agent has been demonstrated to be superior. (3) The
examples of induction schemes from recent trials are
illustrated in Table III. (Level II, Grade B).

Post remission Therapy

Consolidation therapy with a backbone of high-dose
cytarabine is most widely practised for pediatric AML,
although the timing, dose and accompanying agents vary
considerably. Continuous delivery of multi agent low-dose
chemotherapy (thioguanine, vincristine,
cyclophosphamide, fludarabine), delivery of repeated
cycles of myelo suppressive therapy with or without stem
cell rescue and allogeneic stem cell transplantation have
all been explored alone or in sequence for pediatric AML
(17). The optimum number of postremission cycles of

myelosuppressive chemotherapy has yet to be determined

and probably depends on the therapy used for induction.
Examples of post remission therapeutics from selected
studies are shown in Table III. (Level II, Grade B).

Maintenance Therapy
The benefit of maintenance therapy in children when
tested in randomized trials has shown controversial results
(1). As remission and postremission consolidations have

intensified, maintenance therapy has been omitted or

abbreviated by most groups. In patients who have non-
APL AML, maintenance treatment showed no beneût in
two randomized studies (Leucemie Aigue Myeloblastique

193
Enfant 91 and CCG 213); these studies even suggested
that maintenance therapy may be deleterious. In contrast,
the BFM group continues to include a thioguanine based
maintenance phase with monthly pulses of cytarabine (40
mg/m2 s.c. for 4 days) (3). (Level II, Grade B).

Current Status of Stem cell Transplant

The role of allogenic SCT, particularly whether it should
be done during first CR or reserved for second remission,
remains the most controversial issue in pediatric AML. Most
groups agree that children who have APL, AML and Down
syndrome or AML with t(8;21) or inv(16) are not
candidates for SCT in first CR (21). Whereas the trend in
Europe is to reduce the use of SCT in first CR, SCT in CR 1
is supported in the United States (3). Horan et al conducted
a meta-analysis on CCG, POG and MRC AML 10 studies
and found a significant benefit in DFS and OS for
intermediate risk children treated with allogenic SCT (20).
The same dilemma, as above, also prevails for high risk
pediatric AML in CR 1. CCG 2961 suggested a benefit for
high risk patients but numbers were very small and no p
value has been provided for this subgroup. Other studies
which report subgroup analysis did not show a benefit of
allo-SCT in high risk patients (21).
Randomized studies analyzed according to intent to treat
have failed to show that autologous SCT provides a survival
advantage over intensive chemotherapy, and a meta-
analysis concluded that data were insufficient to determine
whether autologous SCT is superior to non myeloablative
chemotherapy (22). (Level II, Grade B).

Newer chemotherapeutics & Targeted Agents

Given the poor prognosis and the severe toxicity associated
with the current chemotherapeutic agents, there is an

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 Selected Phase Iii Clinical Trials For De Novo AML
Table III.

S. No. Study Induction Post remission therapy CR EFS OS

(yrs, ref) All doses in mg/m2/day Percentage (%)
1 BFM 98 1.AIE: Ara-C 100mg d1-8; 3a. “Standard consolidation” 88 42±3 52± 4
(1998- Ida 12 mg d3–5; 6 –TG 60 mg 1 – 43;
2003; 23) Eto 150 mg d 6– 8. Pred 40 mg 1 – 28;
2. HAM : VCR 1.5 mg d1, 8, 15, 22;
Ara-C 3000 mg d1 – 3; Ida 7 mg d 1, 8, 15, 22;
Mito 10mg d4 – 5. Ara-C 75mg d3–6,10–13,
24–27,17–20,31–34,38–41;
Cyclo 500 mg 29, 43;
IT Ara-C d 1, 15, 29, 43.
3b. AI (Ara-C 500mg d1-4;
Ida 7 mg d3-5 & IT Ara-C d3-5)
4. HAE :
Ara-C 3000 mg d1–3;
Eto 125 mg d2 – 5

195
 S. No. Study Induction
196
(yrs, ref) All doses in mg/m2/day
All receive CNS irradiation and allogenic SCT if family eligible donor is available
2. CCG 2961 Intensive timing
(1996- 1a) DCTER or Ida DCTER :
2002 24). continuous infusion
DNR 20 mg d0 – 3 OR
Ida 5mg d0 – 3;
followed by Ara-C 200 mg
d0– 3;Eto 100 mg
d 0 – 3;6-TG 100 mg
d0 – 3; Dex 6mg d0 – 3
1b) DCTER : days 10 – 13
Post remission therapy CR EFS OS
Percentage (%)
5) Maintenance therapy 1 year : 88 42±3 52± 4
6 TG 40 mg daily; Ara-C 40 mg s.c.
4 days monthly; IT ara-C × 4 times
2a) IdaDCTER/DCTER : As 1a
2b) Fludarabine/ara-C/
idarubicin : Ida 12 mg d 0,
1 and 2;
Flu 10.5 mg load,
then 30.5 mg for
48 h, Ara-C 390 mg-
load then 2400 mg
for 73 hrs.
3) Capizzi-II :Ara-C 3000 mg
d0–1 & 7 – 8; L-Asp 6000 units
i.m. d 1- 8
4) Maintenance : IL-2 or
observation
 S. No. Study Induction Post remission therapy CR EFS OS
(yrs, ref) All doses in mg/m2/day Percentage (%)
Patients with matched family donor receive allogeneic stem cell transplant after cycle 3.
CNS prophylaxis: Intrathecal Ara-C.
3. MRC 1a). ADE (10 + 3 + 5) 3) MACE : 92 56 66
AML 12 Or MAE (3+10+5) Amsacrine 100 mg
(1995- Ara-C 100 mg, d 1– 5; Ara-C 200 mg d 1 – 5;
2002 25) q 12 hrly, d 1– 10; Eto 100 mg d 1–5
DNR 50 mg d1, 3, 5 Good risk or no donor
1b). MAE (3 + 10 + 5) 4a). MidAC: Mito 10 mg d 1 – 5;
Ara-C 100 mg, Ara-C 1000 mg days 1 – 3. OR
q 12 hrly, d 1– 10; 4b). Capizzi-II:
Mito 12 mg d 1, 3, 5; Ara- C 3000mg d1–2 &
Eto100 mg d 1– 5 D 8–9; L-asp. 6000 U/m2
2a). ADE (8 + 3 + 5) i.m. d2 & 9
OR 2b). MAE 5). MidAC
(3 + 8 + 5) Standard or poor risk with donor
4a).Allogeneic stem cell
transplant. OR
4b). Capizzi-II followed by

197
Allogenic SCT.
 S. No. Study Induction Post remission therapy CR EFS OS

198
(yrs, ref) All doses in mg/m2/day Percentage (%)
4 NOPHO 1)ATEDox: Ara- 3) HA1M : Ara-C 1000 mg 92 48±3 65±3
AML 93 C continuous 200 mg q 12 hrly d 1 – 3;
(1993– d1–4; 6TG 100 mg d1–4; Mitoxantrone 10 mg d 3 – 5.
2001 26) Eto (continuous)100 mg 4) HdA2E : Ara-C 2000 mg
d 1–4; Dox 75 mg/m2 8 hr q 12 hrly d1 – 3;
infusion d5. Etoposide 100 mg d2–5.
2a). Patients with day 16 5) HA3 : Ara-C 3000 mg
marrow blasts d”5% q 12 hrly
await hematologic recovery, d1– 3.
then ATEDox 6) HdA 2E
2b). Patients with day 16
marrow blasts > 5%, receive
AM: Ara-C 200 mg d1 –5;
Mito 10 mg d 1–3;
Patients not in CR after
2nd course receive
3rd course induction with HdA 2 E.
Patients with matched family donor eligible for allogeneic stem cell transplant.
(CNS prophylaxis : Intrathecal methotrexate)
unmet need to develop newer novel agents for the
treatment of pediatric AML. Cytarabine and anthracycline
such as daunorubicin or idarubicin remain the gold
standard for the treatment of AML in all age groups. Many
new agents with diverse putative mechanisms of action
are currently entering clinical trials for adults (Table IV)
[27]. As the scope of this article is limited, we hereby

describe only those agents which have been tested in

clinical trials for children.
Novel agents potentially useful for acute myeloid
leukemia. (Table IV) 27.
Drug Class Agents Mechanism of Action
Immunotoxins Gemtuzumab CD33 binding, toxin
ozogamicin internalization, and double
strand DNA breaks
Monoclonal Lintuzumab CD33 binding, complement
antibodies dependent cytotoxicity, and
antibody-directed cellular
cytotoxicity
Newer Clofarabine Inhibition of ribonucleotide
nucleoside Troxacitabine reductase and DNA
analogues Sapacitabine polymerase; induction of
apoptosis
Hypomethylating Azacitidine Inhibition of DNA methylation
agents Decitabine
Farnesyltransferase Tipifarnib Inhibition of farnesyl
inhibitors Lonafarnib transferase, inhibition of
angiogenesis, and induction
of cellular adhesion
Alkylating agents Laromustine DNA alkylation and DNA
cross-linking
FLT3 inhibitors Lestaurtinib Inhibition of FLT3
Tandutinib phosphorylation, induction of
apoptosis
Multidrug- Valspodar Binding to P-glycoprotein and
resistant Zosuquidar inactivation of its
modulators efflux function

199
a. Gemtuzumab Ozagamicin (GO)
GO is a humanized anti-CD33 antibody conjugated to
cytotoxin calicheamicin. The COG evaluated the safety and
efficacy of GO (single agent) 6 mg/m 2, two doses
administered 2 weeks apart for pediatric patients with
multiple relapsed or primary refractory AML(28). Eight of
29 (28%) patients achieved overall remission. Remissions
were comparable in patients with refractory (30%) and
relapsed (26%) disease. GO was recently studied in AML-
02 trial in a dose of 3 mg/m2 in combination with low
dose of cytarabine, daunomycin and etoposide in patients
with MRD positivity of 1% or more after first induction
and as a single agent of 6 mg/m2 in patients with MRD
more than 0.1% after two inductions. They showed that
27of the 29 patients who had poor response to first
induction had a remarkable decrease in MRD (29). (Level II,
Grade C).

b. FLT3 Inhibitors
FLT3 is a transmembrane enzyme that promotes
proliferation after activation by ligand binding and plays
an important role in the survival and proliferation of AML
blasts. Phase II single-agent study of this drug in adults
with relapsed/refractory FLT3-mutant AML, showed that
it was well tolerated at doses up to 80 mg orally twice
daily, with common toxicities of mild nausea and fatigue.
Lestaurtinib is now being tested in the COG for children
with relapsed/refractory FLT3-mutant AML (30)
(Level V, Grade D).

c. Farnesyltransferase inhibitors (FTI)

Farnesylated protein products of the Ras gene family are
frequently activated in MDS and AML. Two farnesyl
transferase inhibitors, tipifarnib and lonafarnib, are

200
currently under development for the treatment of AML. A
COG pediatric Phase I trial of tipifarnib, a potent FTI, for
children with refractory leukemias has been completed,
demonstrating tolerability at a dose sufficient to inhibit
farnesyltransferase activity in leukemic blasts. Also COG is
planning to study tipifarnib in the postallogeneic SCT
setting (30). (Level V, Grade D).

e. Epigenetic agents: Histone Deacetylase

Inhibitors (HDAC) inhibitors
HDAC inhibitors are members of a new class of agents
that modulate the expression of genes by causing an
increase in histone acetylation, thereby regulating
chromatin structure and transcription. HDAC inhibitors
evaluated in clinical trials in adults with AML/MDS are
depsipeptide, vorinostat and valproic acid (31). Sodium
valproate (VPA) though long used as an antiepileptic drug,
has more recently also been identified as an antitumor
agent with HDAC inhibition as a potential mechanism of
action in myeloid malignancies. Pediatric Phase I studies
of decitabine, valproic acid, and SAHA are underway.
Chengzhi et al recently studied the synergistic effect of
valproate and cytarabine in clinically relevant pediatric AML
cell lines and diagnostic blasts from children with AML
(32).The authors noticed that t(8;21) AML blasts were

significantly more sensitive to VPA and showed far greater

sensitivities to combined cytarabine and VPA than non-
t(8;21) AML cases. (Level V, Grade C).

Minimal Residual Disease Monitoring

(MRD) Monitoring
The role of detection of disease below the level of
morphological remission in the treatment of pediatric AML
is once again following the footprints of treatment of ALL.

201
With more and more data evolving, the role of MRD
monitoring as a prognostic response to therapy is being
defined slowly. In a CCG study of 252 pediatric patients in
morphologic remission, Sievers et al studied detection of
MRD by flow cytometric technique. They demonstrated
16% of the patients in CR were identified as having occult
disease. These patients had a fivefold higher risk for relapse
than the MRD-negative patients, with a relapse-free
survival rate (RFS) from remission of 35%, compared with
65% for the MRD-negative patients. In a multivariate
analysis, flow cytometric detection of MRD showed the
strongest correlation with relapse-free survival (33). The
utility of flow-based MRD has been prospectively studied
in a multicentric study (AML-02) where risk adapted
therapy has been based upon sequential MRD assessment
(29). The outcome for patients with low levels of MRD (0·1%

to <1%) after first induction, was similar to that of patients

with undetectable MRD (<0·1%). However, the presence
of high levels of MRD (e”1%) after first induction remained
a significant adverse predictor, suggesting that novel
therapies are needed for this high-risk subset of
patients.(Level II, Grade C).

AML in Special Circumstances

1. Relapse AML
There is currently no uniform approach to treatment of
relapse, but most study groups believe that cure without
allogenic SCT is impossible. After relapse, the survival rate
is only 21% to 33% as per recent reports. Various remission
induction regimens, including fludarabine plus cytarabine
and mitoxantrone plus etoposide, seem to give similar
results (3). The targeted immunotherapy agents such as
gemtuzumab ozogamicin and clofarabine along with

202
newer chemotherapeutics as mentioned in (Table IV) have
shown some activity in clinical trials. Aladjidi et al
retrospectively analysed 96 patients who relapsed on
French LAME 89/91 protocol (34). Multiple re induction
therapies such as standard dose cytarabine, high dose
cytarabine, fludarabine, carboplatin & etoposide with
other agents led to CR2 rate of 71%. Of the 53 children
transplanted (25- autograft, sibling related -12, alternate
donor -16), 5 year OS and DFS was 33% and 45%
respectively. (Level II, Grade B).

2. AML with Down Syndrome

DS children have a 10- to 20-fold higher risk for developing
acute lymphoblastic
leukemia and acute myeloid leukemia (AML), as compared
with non-DS children. DS children with leukemia frequently
experience higher levels of treatment-related toxicity and
inferior event-free survival rates, as compared with non-
DS children. DS children also develop AML with unique
features and have a 500-fold increased risk of developing
the AML subtype, acute megakaryocytic leukemia (AMkL;
M7). Nearly 10% of DS newborns are diagnosed with a
variant of AMkL, the transient myeloproliferative disorder,
which can resolve spontaneously without treatment; event-
free survival rates for DS patients with AMkL ranges from
80% to 100%, in comparison with <30% for non-DS
children with AMkL. Somatic mutations of the GATA1 gene
have been detected in nearly all DS TMD and AMkL cases
and not in leukemia cases in non-DS children.(35). The high
EFS is related to increased in vitro sensitivity of Down
syndrome megakaryoblasts to cytarabine (ara-C) and
daunorubicin. In the POG AML 9421 study, 17.5% of
Down syndrome patients developed symptomatic
cardiomyopathies, including three who died from

203
congestive heart failure (37). In children with DS, there is a
delicate balance between the antileukemic efficacy of
intensive chemotherapy and the increased toxic morbidity
and mortality rates. COG in their 2971A trial, eliminated
etoposide and dexamethasone, as well as the 3-month
maintenance period, from the standard chemotherapy
schedule used in the 2891 AML trial. Preliminary analysis
shows outcome data (3-year probability of OS 84%) which
are comparable to the CCG 2891 standard timing arm
(38). The NOPHO reported that approximately half of the

patients included in their AML 93 protocol had dose-

reductions of 75% and 67% of anthracyclines and
cytarabine, respectively, which did not influence the relapse
rate in those patients (39).. (Level II, Grade B).

Conclusion
Inspite of remarkable advances in the field of diagnosis
and treatment of pediatric AML the survival of these
patients is not more that 50-60%. Future research is
required in the development of better cytogenetic and
molecular diagnostics, so as to improvise our risk based
approach to treatment. Evaluation of MRD has further
helped us to identify, patients at a higher risk of relapse.
Not only diagnostics, there is a necessity of testing new
chemotherapeutics and targeted agents in clinical trials
through effective international collaborations. Finally, the
similarities between AML biology in adult subgroups and
children will guide pediatric investigators in choosing the
possibilities of new therapeutic horizons in children with
AML.

Selected Abstracts:
1. Prognostic Impact of Specific Chromosomal
Aberrations in a Large Group of Pediatric Patients With

204
Acute Myeloid Leukemia Treated Uniformly According
to Trial AML-BFM 98.
Neuhoff CV, Reinhardt D, Sander A, Zimmermann M,
Bradtke J, Betts DR et al.
J Clin Oncol 2010; 28 (16): 2682-89.
Purpose: Because cytogenetic data are essential for risk
stratification of childhood acute myeloid leukemia (AML),
the impact of chromosomal aberrations is crucial.
Patients and Methods: Data of a large group of patients
younger than 18 years treated according to study AML–
Berlin-Frankfurt-Münster (BFM) 98 (n = 454), including
their cytogenetics, were analyzed.
Results The favorable outcome in the subgroups of
patients with t(8;21), inv(16), and t(15;17), with an overall
survival of 91% (SE, 4%), 92% (SE, 6%), and 87% (SE,
5%), respectively, was confirmed. Within this group, the
5-year probability of event-free survival (pEFS) of all 17
children with t(8;21) and additional aberrations apart from
del(9q) or “X/”Y was 100%. As expected, the cytogenetic
finding of a complex karyotype (n = 35; pEFS, 33%; SE,
8%) or a monosomy 7 (n = 12; pEFS, 17%; SE, 11%) was
associated with a poor outcome. Compared with
remaining patients with cytogenetic data (pEFS, 48%; SE,
2%), prognosis in patients with an MLL rearrangement (n
= 91) was inferior (pEFS, 34%; SE, 5%; P = .0005).
Particularly, children with t(9;11) and additional
aberrations (n = 13; pEFS, 31%; SE, 14%) and MLL
rearrangements other than t(9;11) and t(11;19) (n = 41;
pEFS, 24%; SE, 7%) had an unfavorable outcome. Nine
patients with aberrations in 12p showed an adverse
prognosis (pEFS, 11%; SE, 10%). The outcome of patients
with aberrations of chromosome 5 (n = 13) was better
than expected (pEFS, 50%; SE, 13%).
205
Conclusion Because the prognostic value of rare recurrent
chromosomal aberrations still has to be elucidated, these
data will contribute to future risk stratification for the
treatment of pediatric AML.
2. Successive clinical trials for childhood acute myeloid
leukemia at St Jude Children’s Research Hospital, from
1980 to 2000.
Ribeiro RC, Razzouk BI, Pounds S, N Hijiya, Pui CH, Rubnitz
JE.
Leukemia 2005; 19: 2125 -9.
Despite substantial progress in the management of
childhood acute myeloid leukemia (AML), only about 50%
of patients are cured by intensive chemotherapy. The long-
term results of clinical trials may reveal principles that can
guide the development of future therapy. From 1980 to
2000, 251 patients of 15years of age with newly
diagnosed AML were enrolled on one of the ûve
consecutive St Jude AML studies. The median age of the
128 boys and 123 girls was 6.2 years; 193 were white, 45
black, and 13 of other racial groups. With the exception
of one protocol (AML-83), outcomes improved in general
over the two decades. The estimated 5-year event-free
survival was 30.8. 75.6% for AML-80; 11.174.3% for AML-
83; 35.977.4% for AML-87; 43.576.2% for AML-91; and
45.0711.1% for AML-97. Resistant or relapsed AML caused
the great majority of treatment failures. Increasing the
intensity of chemotherapy (AML-87) did not improve
outcome, partially because of toxicity, nor did prolonging
post remission therapy by adding sequential myeloablative
(AML-80) or non myeloablative (AML-83) chemotherapy
cycles. We conclude that subtype-speciûc therapies are
needed to replace the ‘one size ûts all strategy of the past
two decades.
206
3. Long-term results in children with AML: NOPHO-AML
Study Group – report of three consecutive trials; on behalf
of the Nordic Society of Pediatric Hematology and
Oncology. (NOPHO)
Lie SO, Abrahamsson J, Clausen N, Forestier E, Hasle H,
Hovi L, et al.
Leukemia (2005) 19, 2090–2100
In all, 447 children with acute myeloid leukaemia (AML)
have been treated on three consecutive NOPHO studies
from July 1984 to December 2001. NOPHO-AML 84 was
of moderate intensity with an induction of three courses
of cytarabine, 6-thioguanine and doxorubicin followed
by four consolidation courses with high-dose cytarabine.
The 5-year event-free survival (EFS), disease free survival
(DFS) and overall survival (OS) were 29, 37 and 38%.
NOPHO-AML 88 was of high intensity with the addition
of etoposide and mitoxantrone in selected courses during
induction and consolidation. The interval between the
induction courses should be as short as possible, that is,
time intensity was introduced. The 5-year EFS, DFS and
OS were 41, 48 and 46%. In NOPHO-AML 93, the
treatment was stratiûed according to response to ûrst
induction course. The protocol utilised the same induction
blocks as NOPHO-AML 88, but after the ûrst block, children
with a hypoplastic, nonleukaemic bone marrow were
allowed to recover before the second block. Consolidation
was identical with NOPHO-AML 88. The 5-year EFS, DFS
and OS in NOPHO-AML 93 were 48, 52 and 65%. The
new NOPHO-AML protocol has been based on experiences
from previous protocols with stratiûcation of patients with
regard to in vivo response and speciûc cytogenetic
aberrations.

207
4. Treatment strategy and long-term results in paediatric
patients treated in consecutive UK AML trials.
Gibson BE, Wheatley K, Hann IM, Stevens RF, Webb D,
Hills RK, et al.
Leukemia (2005) 19, 2130–2138
Between 1988 and 2002, 758 children with acute myeloid
leukaemia (AML) were treated on Medical Research Council
(MRC) AML 10 and AML 12. MRC AML 10 tested the role
of bone marrow transplantation following four blocks of
intensive chemotherapy and found that while both
allogeneic bone marrow transplant (allo-BMT) and
autologous bone marrow transplant (A-BMT) significantly
reduced the relapse risk (RR), this did not translate into a
significant improvement in overall survival (OS). A risk
group stratification based on cytogenetics and response
to the first course of chemotherapy derived from MRC
AML 10 was used to deliver risk-directed therapy in MRC
AML 12. Allo-BMT was limited to standard and poor risk
patients and A-BMT was not employed. Instead, the benefit
of an additional block of treatment was tested by
randomising children to receive either four or five blocks
of treatment in total. While the results of MRC AML 12
remain immature, there appears to be no survival
advantage for a fifth course of treatment. The 5 year OS,
disease-free survival (DFS), event-free survival (EFS) and
RR in MRC AML 12 are 66, 61, 56 and 35%, respectively;
at present superior to MRC AML 10, which had a 5-year
OS, DFS, EFS and RR of 58, 53, 49 and 42%, respectively.
MRC AML trials employ a short course of triple intrathecal
chemotherapy alone for CNS-directed treatment and CNS
relapse is uncommon. Improvements in supportive care
have contributed to improved outcomes and the number
of deaths in remission fell between trials. Anthracycline-

208
related cardiotoxicity remains a concern and the current
MRC AML 15 trial tests the feasibility of reducing
anthracycline dosage without compromising outcome by
comparing standard MRC anthracycline-based
consolidation with high-dose ara-C. MRC studies suggest
that the role of allo-BMT is limited in 1st CR and that there
may be a ceiling of benefit from current or conventional
chemotherapy.
5. Current and emerging therapies for acute myeloid
leukemia
Tadeusz Robak T, Wierzbowska A.
Clin Ther 2009; 31:2349–2370.
Background: Acute myeloid leukemia (AML) is a clonal
disease characterized by the proliferation and
accumulation of myeloid progenitor cells in the bone
marrow, which ultimately leads to hematopoietic failure.
The incidence of AML increases with age, and older
patients typically have worse treatment outcomes than
do younger patients.
Objective: This review is focused on current and emerging
treatment strategies for nonpromyelocytic AML in patients
aged <60 years.
Methods: A literature review was conducted of the
PubMed database for articles published in English.
Publications from 1990 through March 2009 were
scrutinized, and the search was updated on August 26,
2009. The search terms used were: acute myeloid leukemia
in conjunction with treatment, chemotherapy, stem cell
transplantation, and immunotherapy. Clinical trials
including adults with AML aged e”19 years were selected
for analysis. Conference proceedings from the previous 5
years of The American Society of Hematology, The
209
European Hematology Association, and The American
Society for Blood and Marrow Transplantation were
searched manually. Additional relevant publications were
obtained by reviewing the references from the chosen
articles.
Results: Cytarabine (AraC) is the cornerstone of induction
therapy and consolidation therapy for AML. A standard
form of induction therapy consists of AraC (100–200 mg/
m2), administered by a continuous infusion for 7 days,
combined with an anthracycline, administered
intravenously for 3 days. Consolidation therapy comprises
treatment with additional courses of intensive
chemotherapy after the patient has achieved a complete
remission (CR), usually with higher doses of the same drugs
as were used during the induction period. High-dose AraC
(2–3 g/m2) is now a standard consolidation therapy for
patients aged <60 years. Despite substantial progress in
the treatment of newly diagnosed AML, 20% to 40% of
patients do not achieve remission with the standard
induction chemotherapy, and 50% to 70% of first CR
patients are expected to relapse within 3 years. The
optimum strategy at the time of relapse, or for patients
with the resistant disease, remains uncertain. Allogeneic
stem cell transplantation has been established as the most
effective form of antileukemic therapy in patients with AML
in first or subsequent remission. New drugs are being
evaluated in clinical studies, including immunotoxins,
monoclonal antibodies, nucleoside analogues,
hypomethylating agents, farnesyltransferase inhibitors,
alkylating agents, FMS-like tyrosine kinase 3 inhibitors,
and multidrug-resistant modulators. However,
determining the success of these treatment strategies
ultimately requires well-designed clinical trials, based on

210
stratification of the patient risk, knowledge of the
individual disease, and the drug’s performance status.
Conclusions: Combinations of AraC and anthracyclines
are still the mainstay of induction therapy, and use of high-
dose AraC is now a standard consolidation therapy in AML
patients aged <60 years. Although several new agents
have shown promise in treating AML, it is unlikely that
these agents will be curative when administered as
monotherapy; it is more likely that they will be used in
combination with other new agents or with conventional
therapy.
6. Minimal residual disease-directed therapy for
childhood acute myeloid leukaemia: Results of the AML
02 multicentre trial.
Rubnitz JE, Inaba H, Dahl G et al, Ribeiro RC, Bowman wP,
Taub J, et al.
Lancet Oncol 2010; 11: 543–52.
Background: We sought to improve outcome in patients
with childhood acute myeloid leukaemia (AML) by applying
risk-directed therapy that was based on genetic
abnormalities of the leukaemic cells and measurements
of minimal residual disease (MRD) done by ûow cytometry
during treatment.
Methods From Oct 13, 2002, to June 19, 2008, 232
patients with de-novo AML (n=206), therapy-related or
myelodysplasia-related AML (n=12), or mixed-lineage
leukaemia (n=14) were enrolled at eight centres. 230
patients were assigned by block, non-blinded
randomisation, stratiû ed by cytogenetics or morphological
subtype, to high-dose (18 g/m², n=113) or low-dose (2
g/m², n=117) cytarabine given with daunorubicin and

211
etoposide (ADE; induction 1). The primary aim of the study
was to compare the incidence of MRD positivity of the
high-dose group and the low dose group at day 22 of
induction 1. Inductions 2 consisted of ADE with or without
gemtuzumab ozogamicin (GO anti-CD33 monoclonal
antibody); consolidation therapy included three additional
courses of chemotherapy or haematopoietic stem-cell
transplantation (HSCT). Levels of MRD were used to
allocate GO and to determine the timing of induction 2.
Both MRD and genetic abnormalities at diagnosis were
used to determine the ûnal risk classiûcation. Low-risk
patients (n=68) received ûve courses of chemotherapy,
whereas high-risk patients (n=79), and standard-risk
patients (n=69) with matched sibling donors, were eligible
for HSCT (done for 48 high-risk and eight standard-risk
patients). All 230 randomized patients were analyzed for
the primary endpoint. Other analyses were limited to the
216 patients with AML, excluding those with mixed-
lineage leukemia. This trial is closed to accrual and is
registered with ClinicalTrials.gov, number NCT00136084.
Findings Complete remission was achieved in 80% (173
of 216 patients) after induction 1 and 94% (203 of 216)
after induction 2. Induction failures included two deaths
from toxic effects and ten cases of resistant leukemia. The
introduction of high-dose versus low-dose cytarabine did
not signiûcantly lower the rate of MRD-positivity after
induction 1 (34% vs 42%, p=0·17). The 6-month
cumulative incidence of grade 3 or higher infection was
79·3% (SE4·0) for patients in the high-dose group and
75·5% (4·2) for the low-dose group. 3-year event-free
survival and overall survival were 63·0% (SE 4·1) and 71·1%
(3·8), respectively. 80% (155 of 193) of patients achieved
MRD of less than 0·1% after induction 2, and the
cumulative incidence of relapse for this group was 17%

212
(SE 3). MRD of 1% or higher after induction 1 was the
only signiû cant independent adverse prognostic factor
for both event-free (hazard ratio 2·41, 95% CI 1·36–4·26;
p=0·003) and overall survival (2·11, 1·09–4·11; p=0·028).
Interpretation Our findings suggest that the use of
targeted chemotherapy and HSCT, in the context of a
comprehensive risk-stratiûcation strategy based on genetic
features and MRD findings, can improve outcome in
patients with childhood AML.

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218
 Management of Acute Myeloid
Leukemia in Elderly
Dr.Amit Joshi, Dr. Bharat Singh Bhosale

Acute myeloid leukemia is predominantly is a disease of

the adult age group and accounts for about 80 % of acute
leukemia diagnosed in this age group. The elderly (defined
as age group 60 years and above) are more commonly
affected and 35% of newly diagnosed AML are actually
aged above 75 with the median age at diagnosis being
67 years (1). The elderly patient in general has lower
complete remission (CR) and relapse-free survival (RFS)
rates than their younger counterparts as was demonstrated
in the Medical Research Council (MRC) AML-8 trial and
the Cancer and Leukemia group (CALGB 8525 study) .
The CR rates were 52% vs.70% for the MRC study while it
was 47% vs.73% for the CALGB study respectively. The
same inferior results were reflected in the disease free
survival too for the elderly age group.
A higher prevalence of co morbid conditions as well as
the unique biological features of elderly AML patients
account for the relatively poor response to therapy (Table-1).
Elderly patients have a multiple co morbid conditions such
as heart disease, renal insufficiency, and vascular disease.

219
 Table 1
Characteristics of AML biology in elderly versus
younger individuals
Characteristics Elderly Younger
AML AML
Population incidence, per
100,000 persons in the U.S. 17.6 1.8
Secondary AML 24%–56% 8%
TRM with induction therapy 25%–30% 5%-10%
Complete remission 38%–62% 65%–73%
FLT3 mutation 34% 20%
MDR1 expression 71% 35%
Favorable cytogenetics
t(8;21) 2% 9%
Inv 16 or t(16;16) 1%–3% 10%
t(15;17) 4% 6%–12%
Intermediate-risk cytogenetics
+8 6%–10% 4%
Normal karyotype 31% 34%–45%
Unfavorable cytogenetics
- 7, -5, inv 3, or t(3;3) 8%–9% 3%
Complex 18% 7%

Poor performance status of elderly patients at diagnosis

also contributes to poor outcomes to treatment (5,6). As
compared with the young AML patient, AML in elderly
patients have more often emerged from an antecedent
hematologic disorder (MDS) or after treatment for a
previous malignancy. They have an increased incidence of
poor-prognosis karyotypes (5q-, 7q-) coupled with
increased expression of multidrug resistant protein (MDR)
involved in intrinsic resistance to chemotherapeutic agent
220
(5,7). Various trials have shown that patients having MDR1
positive AML have lower CR and DFS rates than those with
MDR1 negative AML (8, 9). Patients with five or more
chromosomal aberrations benefited least from current
treatment and are better candidate for investigational
therapy or supportive care (11).
In a CALGB study involving 635 elderly AML patients, the
important prognostic factors were pretreatment
cytogenetics , presenting WBC count ,marrow blast
percentage, sex, and age (Table 2).
Table 2 : Prognostic cytogenetic groups in elderly
AML patients based on a study of 635 individuals
>75 years of age (11)
Cytogenetic risk group Number of Outcome (%)
patients
Complete remission
Complex karyotype 122 25
with>3 aberrations
Rare aberrations 33 30
Noncomplex > 3 or 480 56
non rare aberrations
5-Year disease-free survival
Complex karyotype with 22 0
> 5 aberrations
< 5 aberrations 285 9.1
5-Year overall survival
Complex karyotype with
>5 aberrations 94 0
CBF-AML 31 19.4
Rare aberrations 33 0
< 5 non-CBF or non rare
aberrations 477 7.5

221
TREATMENT MODALITIES FOR AML IN THE
ELDERLY
Standard Chemotherapy - Induction chemotherapy
The results of standard chemotherapy in elderly patients
are poor. Though reported CR rates are about 60%,
remissions are usually transient, with median survival being
5-10 months, and the probability of remaining in remission
at 3 years after diagnosis being less than 10% (12, 13).
The options for management of these patients are:
 Standard chemotherapy (e.g. daunorubicin +
cytosine arabinoside 3+7)
 Non-intensive (palliative) treatment
 Investigational treatment
Remission induction chemotherapy does not differ from
that for the younger patient. It includes an anthracycline
such as daunorubicin or idarubicin, (for 3days) with 7 days
of continuous infusion of cytarabine. The benefit of
intensive chemotherapy versus supportive care in the
elderly AML was demonstrated in a randomised phase III
trials. Standard chemotherapy was compared with
supportive care in patients aged > 65. Those in the
chemotherapy group received daunorubicin, vincristine,
and cytarabine as both induction and consolidation, while
those in the other study arm received supportive care. The
CR rate in patients treated with chemotherapy was 58%
and the 2-year DFS rate was 17% versus 0% in supportive
care group. The median survival time was longer (21 weeks
versus 11weeks), and days of hospitalization were similar
in the two groups (14).
In an order to improve the tolerability of induction
chemotherapy, various modifications have been made by
either reducing its intensity or substituting mitoxantrone

222
or idarubicin for daunorubicin. In the UK Medical Research
Council AML 9 study 972 patients were randomised to
receive DAT (daunorubicin, cytosine arabinoside, 6-
thioguanine) in a 1+5 or a standard 3+10 regimen:
patients receiving DAT 1+5 were less likely to achieve CR,
and required more time in hospital and more blood
product support (15). A trend towards improved survival
was also seen in another multicentre study in which
patients aged 65 and over were randomised to receive
intensive therapy or low-dose cytarabine (16). An EORTC
study randomised patients over 60 years of age to
mitoxantrone and cytarabine or daunorubicin and
cytarabine induction chemotherapy. CR rates were
improved in patients receiving mitoxantrone without an
improvement in relapse free survival or overall survival (17).
Similarly a meta-analysis of trials comparing idarubicin with
daunorubicin did not shown a survival advantage for
idarubicin. (18). In an SWOG randomised study there was
no improvement in outcome when mitoxantrone and
etoposide were compared to daunorubicin and cytosine
arabinoside (19).
The HOVON/SAKK/AMLSG published there results of
randomized phase III study of double dose of daunorubicin
in older patients. Newly diagnosed Patients of AML or high-
risk refractory anemia who were 60 to 83 years of age
(median, 67) received cytarabine, at a dose of 200 mg
per square meter by continuous infusion for 7 days, plus
daunorubicin for 3 days, either at the conventional dose
of 45 mg per square meter (411 patients) or at an escalated
dose of 90 mg per square meter (402 patients) The rates
of remission after the first cycle of induction treatment
were 52% and 35%, in escalated dose and standard dose
daunorubicin arm respectively (P<0.001). There was no
significant difference between the two groups in the

223
incidence of hematologic toxic effects, 30-day mortality
(11% versus12% in the two groups, respectively), or the
incidence of moderate, severe, or life-threatening adverse
events (P = 0.08). Survival end points too did not differ
significantly. However, patients in the escalated-treatment
group who were 60 to 65 years of age, as compared with
the patients in the same age group who received the
conventional dose, had higher rates of complete remission
(73% vs. 51%), event-free survival (29% vs. 14% p=0.002),
and overall survival (38% vs. 23% p <0.001). The HOVON/
SAKK/AMLSG concluded that daunorubicin can be dose-
intensified to 3 X 90 mg/m2 in older patients up to 65
years with more CRs and better survival, without marked
additional toxicity (20).
For patients > 70 year of age use of dose intensive
chemotherapy is not recommended except for the patient
with good risk cytogenetics and with good performance
status. In a recent review Kantarjian et al analyzed 446
patients > 70 years of age with AML (> 20% blasts)
treated with cytarabine-based intensive chemotherapy
between 1990 and 2008 to identify risk groups for high
induction (8-week) mortality. Excluding patients with
favorable karyotypes, the overall complete response rate
was 45%, 4-week mortality was 26%, and 8-week
mortality was 36%. The median survival was 4.6 months,
and the 1-year survival rate was 28%. A multivariate
analysis of prognostic factors for 8-week mortality
identified the; age > 80 years, complex karyotypes, (> 3
abnormalities), poor performance (ECOG 2-4), and
elevated creatinine > 1.3 mg/dL as independently adverse
risk factors. Patients with 0,1,2 and greater than 3 factors
had estimated 8-week mortality rates of 16%, 31%, 55%,
and 71% respectively. They concluded that the prognosis
of patients greater than 70 years of age with AML is poor

224
with intensive chemotherapy an 8-week mortality of more
than 30% and median survival of less than 6 months(21).
Currently for patients in whom intensive chemotherapy is
deemed justified (e.g. age <70 years, good performance
status, WBC <100 x 109/L, no adverse cytogenetic
abnormalities , standard induction chemotherapy with
daunorubicin (or an equivalent anthracycline) for 3 days
plus cytarabine for 7-10 days is recommended .(Grade A
recommendation, level Ib evidence). For patients >70
years of age or with adverse cytogenetic risk may be
considered for investigational therapies, or, a mild
cytoreductive therapy only. (Grade A recommendation,
level Ib evidence).

Postremission Therapy:
Postremission therapy is directed toward further reduction
in the in the residual leukemic cell. The optimal post-
remission therapy for the older adult with AML in CR
remains unclear. Standard post remission therapy includes
high dose cytotoxic chemotherapy, autologous or
allogeneic stem cell transplant.

High dose cytotoxic chemotherapy:

The use of consolidative high-dose cytarabine (HiDAC) in
the postremission setting for elderly AML patients is
controversial. These studies have generally compared
“more” versus “less” postremission therapy. In CALGB
82525 trial 596 adult AML patients, in CR were
randomized to consolidation with 4 cycles of conventional-
dose cytarabine (100 mg/m 2 per day × 5 days),
intermediate-dose cytarabine (400 mg/m2 per day × 5
days) or high-dose cytarabine (3 g/m2 total of 6 doses
over 5 days). In patients aged > 60 years, only 29% were
able to complete all four cycles of HiDAC, the 4-year DFS
225
rate was < 16% and the 4-year OS rate was only 9%.
Cerebellar toxicity occurred in > 30% of patients aged >
60 who received HiDAC. Poor tolerance of HiDAC led to
inferior outcomes observed in elderly patients treated with
HiDAC as compared with younger patients who received
such therapy. (4). In the British Medical Research Council
(MRC) AML11 study, 4 postremission courses of moderate
intensity were compared with 1 course, with equivalent
survival in the 2 arms (13). The CALGB compared 2
relatively intense cycles (cytarabine 500 mg/m2 per q12h;
mitoxantrone 5 mg/m2 per q12h, each for 6 doses) with
4 less intensive cycles (cytarabine 100 mg/m2 continuous
IV on days 1-5) and found no differences (25). In the
AMLSG AML HD98B trial, intensive consolidation
(idarubicin 12 mg/m2 on days 1 and 3; etoposide 100
mg/m2 on days 1-5) was superior to a mild 1-year oral
maintenance therapy (idarubicin 5 mg per os (p.o.) on
days 1, 4, 7, 10, and 13; etoposide 100 mg p.o. on days
1 and 13; q4 weeks) (26). From these data no clear
recommendation can be given. For patients without
adverse cytogenetics, good performance status and no
significant comorbidity, standard “3 + 7” induction
followed by repetitive cycles of modest dose consolidation
is an acceptable consolidation strategy. This view is
supported by recent results from the Swedish National
Registry suggesting that this approach is associated with
longer survival than lower doses of similar therapy (27).
(Grade B recommendation, level III evidence).

High-dose therapy followed by allogeneic or

autologous stem cell transplant is another
postremission therapeutic strategy:
Elderly AML patients are often poor candidates for SCT
because of comorbid medical conditions.

226
Nonmyeloablative or RIC regimens have been developed
to reduce TRM in older or medically less fit patients. There
is increasing evidence derived from clinical trials that
appropriately selected individuals may benefit from
allogeneic SCT following for induction remission. A
retrospective study from the Cooperative German
Transplant Study Group of 368 patients (median age, 57
years; range, 50-73) suggests that matched unrelated and
matched sibling donor allogeneic HSCT (72% had received
RIC regimens) result in comparable survival in older AML
patients(28). In one phase II trial, Nineteen older patients
(median age, 64 years; range, 60 to 70 years) with active
myeloid malignancies were treated with an RIC regimen
consisting of fludarabine, melphalan, and carmustine
followed by alloHSCT from matched unrelated (n =12) or
sibling donors (n = 7). Before transplantation, patients
had a median of 50% bone marrow blasts (range, 0% to
70%) Engraftment was successful for all 19 patients.
Seventeen assessable patients achieved complete response
(CR). Four patients experienced relapse. One -year non
relapse mortality rate was 22%. With a median follow-up
of 825 days (range, 595 to 1,028 days), 13 of 19 patients
are alive, resulting in a 1-year survival rate of 68% (95%
confidence interval, 48% to 89%). This phase II study
concluded that older patients with untreated poor-
prognosis leukemia, RIC regimen combined with allo HSCT
reduces the leukemic burden, resulting in a high CR rate.
When ATG is added, matched unrelated donor
transplantation can be performed safely in older patients
(29). (Grade C recommendation, level IV evidence)
Autologous SCT has also been attempted as a
consolidation therapy. In a phase II study involving 160
elderly patients with AML (median age, 69 years), patients
were treated with intensive induction chemotherapy and,

227
if CR was achieved, one cycle of consolidation therapy
followed by autologous SCT for selected patients >70
years of age . This measure did not prevent early relapse
and thus did not translate into longer DFS or OS (30).

Up coming therapeutic modalities for

AML in Elderly patients:
In last two decades significant progress has been made in
the molecular pathogenesis of AML which have led to
possibility of the development of molecularly targeted
therapies. Genetic alterations in AML pathogenesis
comprises:
a) Mutations in genes that activate signal transduction
cascades (eg .FLT3, KIT, and RAS).
b) Gene fusions or mutations resulting in enhanced or
repressed transcriptional activity (eg, PML-RARA,
RUNX1- RUNX1T1, CBFB-MYH11, and CEBPA).
c) Altered function of genes involved in nuclear
cytoplasmic shuttling (eg, NPM1, NUP98, NUP214).
AML phenotype usually results from multiple genetic/
epigenetic lesions affecting differentiation, proliferation,
and apoptosis. Hence targeting of a single aberrant protein
is unlikely to eradicate the leukemic clone. Various early
clinical studies have shown that most of these novel agents
will need to be used in combination with conventional
cytotoxic therapy. Only 3 modalities have reached phase
3 clinical trial development for frontline therapy of AML.
Gemtuzumab ozogamicin (GO, Mylotarg) is a
humanized anti-CD33 monoclonal antibody conjugated
to a derivative of the cytotoxic antibiotic calicheamicin.
U.S. Food and Drug Administration approved
Gemtuzumab ozogamicin in 2000 for monotherapy in

228
patients aged > 60 years with relapsed, CD33+ AML who
are not considered candidates for cytotoxic chemotherapy.
Results of three multicenter, open-label, single-arm, phase
II studies in 277 patients with CD33 + AML at first
recurrence showed that among individuals aged > 60,
the overall response rate (ORR) was 24%, the CR rate was
12%. Patients in whom a CR after initial therapy persisted
for >12 months prior to relapse experienced the best
response to GO, with an ORR of 35%. Infusion-related
events included chills, fever, nausea, hypotension, and
hypertension, all of which occurred in < 8% of cases.
Grade 3 or 4 neutropenia and thrombocytopenia
developed in 98%–99% of patients. Other side effects
included hyperbilirubinemia, transaminitis, and hepatic
veno-occlusive disease (VOD) (31). GO has also been
studied as part of first-line therapy in older patients with
AML. In a phase II trial involving 12 individuals aged > 65
with newly diagnosed AML. Patients with full recovery of
bone marrow function after induction received
consolidation therapy, followed by maintenance GO every
4 weeks. CR was attained in 25% of the patients, and
only one patient was taken off study because of
unsatisfactory bone marrow recovery (32).
Gemtuzumab ozogamicin (GO, Mylotarg) has recently
withdrawn from market by its parent company (Pfizer) in
view of updated results of SWOG Study S0106. In this
Study Mylotarg was combined with daunorubicin and
cytosine arabinoside, versus the same chemotherapy agent
combination without Mylotarg in first-line AML patients
under the age of 61 .This study was conducted to confirm
clinical benefit for Mylotarg. A total of 627 patients were
enrolled in this study. This study was stopped early based
on interim results from the study showing no evidence of
improved efficacy for patients treated with Mylotarg in

229
addition to chemotherapy for previously untreated AML.
Additionally, the fatal induction toxicity rate was
significantly higher in the daunorubicin and cytosine
arabinoside + Mylotarg arm (16/283=5.7% vs. 4/
281=1.4%, P=0.01) (33).
Multidrug Resistance Modulators: Overexpression of
the MDR1 gene product P-glycoprotein (P-gp) is the most
common mechanism of resistance in AML. In patients of
de novo AML, P-gp protein and mRNA expression is present
in the malignant cells of 20%–75% of patients (34). Two
phase III trials have used the P-gp inhibitor PSC-833 into
induction regimens for the treatment of AML in patients
aged > 60 years. In one trial, patients were randomized
to either cytarabine, daunorubicin, and etoposide or the
same regimen with PSC-833 . The study was discontinued
early because of excess early mortality in the group of
patients who received the PSC-833, and an interim analysis
revealed that the DFS and OS rates were similar in the two
arms. The incidence of hyperbilirubinemia, stomatitis,
esophagitis, and diarrhea were greater in patients who
received PSC-833 as part of their induction therapy (35).
In another trial, patients were randomized to two cycles
of cytarabine and daunorubicin induction with or without
PSC-833 followed by one cycle of consolidation with
cytarabine, mitoxantrone, and etoposide showed no
differences between the two groups with regard to the
outcome (36) . These two studies are given to conclude
that there is no firm evidence to date to support the use
of MDR-blocking agents as an adjunct to induction
chemotherapy (Grade A recommendation, level Ib
evidence).
FLT3 inhibitors: Mutations of FLT3 in AML occur in a
substantial percentage of patients with the disease.
Among elderly patients with AML, the FLT3 TKD mutation

230
is uncommon, while the FLT3 ITD is seen in approximately
35% of individuals (37). In a phase II trial oral FLT3 inhibitor
lestaurtinib (CEP701) as first-line therapy in previously
untreated older patients with AML who were not
candidates for intensive chemotherapy received
lestaurtinib at a dose of 60 mg orally twice daily initially
with dose escalation to 80 mg twice daily as tolerated.
There were no CRs or PRs, though a bone marrow response
occurred in 19% of patients, with 11% hematologic
response rates. The most common therapy-related high-
grade toxicities of nausea and diarrhea occurred in <10%
of patients (38). A randomized phase III Cancer and
Leukemia Group B trial (NCT00651261) is ongoing to
evaluate midostaurin as frontline therapy of younger adult
AML patients with activating FLT3 mutations (39).
Demethylating agents: Two demethylating agents, the
cytosine analogs azacitidine and Decitabine have been
approved for the treatment of MDS. In a phase 3
randomized trial, azacitidine prolonged OS compared with
conventional care regimens in patients with intermediate-
2 or high-risk MDS. Approximately one-third of these
patients (n = 113) were classified as having AML under
current WHO criteria (blast counts 20%-30%). Patients
were randomly assigned to receive subcutaneous
azacitidine 75 mg/m2/d or CCR (best supportive care [BSC]
only, low-dose cytarabine (LDAC), or intensive
chemotherapy [IC]). Of the 113 elderly patients (median
age, 70 years) randomly assigned to receive azacitidine (n
= 55) or CCR (n = 58; 47% BSC, 34% LDAC, 19% IC),
86% were considered unfit for IC. At a median follow-up
of 20.1 months, median OS for azacitidine-treated patients
was 24.5 months compared with 16.0 months for CCR-
treated patients (hazard ratio = 0.47; 95% CI, 0.28 to
0.79; P = .005), and 2-year OS rates were 50% and 16%,

231
respectively (P = .001). Two-year OS rates were higher
with azacitidine versus CCR in patients considered unfit
for IC (P = .0003). Azacitidine was associated with fewer
days in hospital (P < .0001) than CCR. In conclusion, for
older adult patients with low marrow blast count (20%
to 30% WHO-defined AML) azacitidine significantly
prolonged OS and significantly improved several patient
morbidity measures compared with CCR (40). Thus
azacitadine could be considered as an option for elderly
AML with low marrow blast count (20% to 30%). (Grade
A recommendation, level Ib evidence).
Conclusion: Optimal treatment of elderly AML patients
is still evolving. Patients with age < 70 year with good
cytogenetics and PS should be offered standard induction
therapy ( 3+ 7) followed by moderate dose consolidation
therapy. The patients above 70 years having adverse
cytogenetics and poor PS should be offered supportive
care or encouraged to take part in a clinical trial. G-CSF or
GM-CSF can be used to reduce the days of neutropenia
and subsequent hospitalization without impact on survival.
Multidrug Resistance Modulators such as P-gp inhibitor
PSC-833 is not of clinical benefit. Demethylating agent as
azacitadine has shown promise in Phase III study and may
be offered to elderly AML patients with low marrow blast
count (20% to 30% WHO-defined AML) not fit for
intensive chemotherapy.
With better understanding of AML pathogenesis, newer
biological agents such as FLT3 inhibitors,
farnesyltransferase inhibitors, Bcl-2 antisense
oligonucleotide may hold promise for the future.

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