Received: 16 October 2020 Revised: 31 December 2020 Accepted: 4 January 2021

DOI: 10.1002/pbc.28909 Pediatric

Blood &
Cancer The American Society of
Pediatric Hematology/Oncology
T R A N S P L A N TAT I O N : B R I E F R E P O R T

Pretransplant myeloid and immune suppression, reduced

toxicity conditioning with posttransplant cyclophosphamide:
Initial outcomes of novel approach for matched unrelated
donor hematopoietic stem cell transplant for
hemoglobinopathies

Gaurav Kharya Atish N. Bakane Archana M. Rauthan

Center for Bone Marrow Transplant and

Cellular Therapy, Indraprastha Apollo
Hospital, New Delhi, India
Correspondence
Gaurav Kharya, Clinical Lead, Center for Bone
Marrow Transplant and Cellular Therapy,
Indraprastha Apollo Hospital, Delhi, India.
Email: gaurav.kharya@gmail.com
Abstract
Hematopoietic stem cell transplant (HSCT) is currently the only curative option for tha-
lassemia major (TM) and sickle cell disease (SCD). We report our experience of using
pretransplant immune suppression (PTIS), augmented Johns Hopkins conditioning, and
posttransplant cyclophosphamide (PTCy) as graft-versus-host disease (GvHD) prophy-
laxis for matched unrelated donor (MUD) transplant in TM/SCD. At a median follow-
up of 307.5 days (range 251-395), all patients (three TM, one SCD) are alive and dis-
ease free. MUD HSCT with PTIS, augmented Johns Hopkins conditioning, and PTCy
as GvHD prophylaxis is a promising way of treating patients with hemoglobinopathies
with low regimen-related toxicity (RRT), no risk of graft failure (GF) and minimal GvHD
rates.

KEYWORDS
hematopoietic stem cell transplant, hemoglobinopathies, matched unrelated donor, posttrans-
plant cyclophosphamide, pretransplant immune suppression

1 INTRODUCTION
Hemoglobinopathies (thalassemia major [TM] and sickle cell dis-
ease [SCD]) are among the commonest indications for hematopoi-
etic stem cell transplant (HSCT) in nonmalignant diseases.1,2 Only 25-
30% patients have suitable human leukocyte antigen (HLA)-identical
related donor.3 Alternative donor transplants either matched unre-
Abbreviations: aGvHD, acute graft-versus-host disease; ANC, absolute neutrophil count;
ATG, antithymocyte globulin; CBT, cord blood transplant; cGvHD, chronic graft-versus-host
disease; CMV, cytomegalovirus; DFS, disease-free survival; EF, peri-engraftment fever; ES,
engraftment syndrome; GCSF, granulocyte colony-stimulating factor; GF, graft failure; GvHD,
graft-versus-host disease; HLA, human leukocyte antigen; HHSCT, haploidentical HSCT;
HSCT, hematopoietic stem cell transplant; IR, immune reconstitution; iv, intravenously; MAC,
myeloablative conditioning; MC, mixed chimerism; MMF, mycophenolate mofetil; MUD,
matched unrelated donor; OS, overall survival; PTCy, posttransplant cyclophosphamide; PTIS,
pretransplant immune and myelosuppression; rBW, recipient body weight; RIC, reduced
intensity conditioning; RRT, regimen-related toxicity; RTC, reduced toxicity conditioning;
SCD, sickle cell disease; TM, thalassemia major.
lated donor (MUD), mismatched family donor (e.g. haploidentical HSCT
[HHSCT]), or cord blood transplant (CBT) expands the donor pool but
comes with risk of regimen-related toxicity (RRT), graft failure (GF),
and graft-versus-host disease (GvHD).4–11 In a review on HSCT for
SCD, Oshrine and Talano stated that matched sibling donor allogeneic
HCT after myeloablative conditioning (MAC) offers a cure for SCD
and is accepted as standard of care. They concluded that it should
be offered to all patients with severe SCD prior to developing severe
end organ dysfunction. However, they also mentioned that utilizing
an alternative donor for transplantation and/or reduced intensity con-
ditioning (RIC) for SCD is still under investigation, and clinical tri-
als should be performed to optimize the best treatment for these
patients.12 Shenoy et al examined the outcomes of MUD HSCT for chil-
dren with severe SCD in BMT CTN phase 2 trial between 2008 and
2014. They used alemtuzumab, fludarabine, and melphalan based RIC,

Pediatr Blood Cancer. 2021;68:e28909. wileyonlinelibrary.com/journal/pbc © 2021 Wiley Periodicals LLC 1 of 9

https://doi.org/10.1002/pbc.28909
2 of 9
and GvHD prophylaxis was with calcineurin inhibitor along with short-
course methotrexate and methylprednisolone. At a median follow-up
of 26 months, the overall survival (OS) and disease-free survival (DFS)
were 86% and 79%, respectively, with GF rate of 10%. They reported
day 100 incidence of grade II-IV acute GvHD (aGvHD) as 28% and
1-year incidence of chronic GvHD (cGvHD) as 62%, 38% of which
were extensive.13 In another trial, the URTH trial, Shenoy et al investi-
gated the efficacy of RIC using alemtuzumab, fludarabine, thiotepa and
melphalan based conditioning for patients with TM undergoing MUD
HSCT. GvHD prophylaxis was with calcineurin inhibitor and short-
course methotrexate. In their analysis, the OS and DFS at a median
follow-up of 58 months were 82% and 79%, respectively. The cumula-
tive incidence of grade II-IV aGvHD rates in their analysis was 24% with
bone marrow as source and that of cGvHD was 29% at 2 years.14 In
another recently published prospective multicentric trial, which stud-
ied the outcomes of HSCT for adolescents and young adults with SCD
using busulfan, fludarabine, and thymoglobulin based RIC, Krishna-
murti et al reported 1-year OS and DFS as 91% and 86%, respectively.
They reported grade II-III aGvHD rates of 18% and cGvHD rates of 27%
in their analysis.15
Transplant physicians have tried to overcome the risk of GF by
using MAC, but it increases the risk of RRT and GvHD.16,17 GvHD is
acceptable for leukemias where graft versus leukemia provides bet-
ter disease control, but for nonmalignant diseases, it impairs quality
of life and may defeat the purpose of offering HSCT.18,19 Anurathapan
et al demonstrated the use of sequential pretransplant immune and
myelosuppression (PTIS) for Lucarelli class III β-TM undergoing HLA-
identical HSCT. They demonstrated that sequential use of fludarabine
200 mg/m2 intravenously (iv) over 5 days in combination with dexam-
ethasone 25 mg/m2 /day iv over 5 days, one to two cycles 3 weeks apart
in association with hydroxyurea, followed by fludarabine and busul-
fan based reduced toxicity conditioning (RTC) is safe and effective pre-
transplant conditioning for patients with high-risk class III TM exhibit-
ing additional comorbidities decreasing the risk of GF and GvHD.20
Bhat et al used a novel pretransplant immunosuppressive prepara-
tive regimen for HHSCT and MUD HSCT in hemoglobinopathies, and
showed it to be a safe and effective approach.21
To handle the donor T cells, the posttransplant cyclophosphamide
(PTCy) platform has evolved over a period of time for T-cell replete
haploidentical transplants.10 Using either RIC or myeloablative RTC,
depending on the underlying disease and performance status of the
patient, along with use of calcineurin inhibitor or mTOR based GvHD
prophylaxis, both the issues of GF and GvHD have been addressed to
a great extent using PTCy platform.22–24 However, its use in combina-
tion with PTIS for MUD HSCT still remains unexplored in nonmalignant
settings.
We developed a protocol wherein we emphasized on PTIS using flu-
darabine, cyclophosphamide, and dexamethasone to suppress recip-
ient T-cell function, followed by augmented Johns Hopkins protocol
with added thiotepa-based RTC and PTCy along with mTOR inhibitor
and mycophenolate mofetil (MMF) as GvHD prophylaxis. This adapta-
tion was based on our previous experience of using this strategy with
added plerixafor for stem cell mobilization in donors (APOLLO proto-
KHARYA ET AL .
col) to treat 25 consecutive patients undergoing haploidentical trans-
plant for severe SCD with very encouraging results.25 In the current
study, we present our experience with this novel strategy in patients
with TM and SCD undergoing MUD HSCT.
2 PATIENTS AND METHODS
2.1 Patients
Four patients (three TM and one SCD) presenting at our center for
blood and marrow transplant from September 2019 until April 2020
for MUD HSCT were enrolled in the trial. All three TM patients were on
regular transfusions and one with SCD had frequent pain crises, persis-
tent anemia, jaundice, splenic sequestration, splenectomy, and chole-
cystectomy. All four lacked suitable HLA-matched family donors (Fig-
ure 1). Ethicals approval for this retrospective analysis was granted by
our institutional ethics committee (IAH-BMR-021/07-20).
2.2 Donors
In all the cases, donors were MUD (three sourced from DATRI, an
Indian non-governmental organization stem cell registry and one
sourced from DKMS, Poland) (Figure 1).
2.3 HLA typing
All the patients and donors were high-resolution HLA-typed at the A, B,
C, DRB1, and DQB1 loci by next-generation sequencing on an Illumina
MiniSeq platform.
2.4 PTIS
Two courses of PTIS were given at three-weekly intervals. Dur-
ing each course, patients received fludarabine 150 mg/m2 (Days
1-5); cyclophosphamide 1000 mg/m2 (Day 1); dexamethasone
20 mg/m2 /per oral (PO) in two divided doses (Days 1-5). Complete
hemogram and reticulocyte counts were done on a weekly basis,
and an attempt was made to maintain absolute neutrophil count
>500/mm3 , platelet count >30 000/mm3 , and hemoglobin >10 gm/dL.
Appropriate granulocyte colony-stimulating factor (GCSF), platelet
and red cell support was given whenever required. During the entire
period, patients also received hydroxyurea 20 mg/kg/day. Target retic-
ulocyte count was kept between one and two during this period. In case
of febrile neutropenia, patients were admitted and started on broad-
spectrum antibiotics. During the two cycles of PTIS, patients received
prophylaxis with penicillin, valaciclovir, fluconazole/voriconazole,
cotrimoxazole +/− antimalarial drugs. Levetiracetam was used for
seizure prophylaxis at the start of PTIS and continued at least until Day
+90 posttransplant (Figure 2).
KHARYA ET AL . 3 of 9

FIGURE 1 Donor selection algorithm

F I G U R E 2 Treatment schema. Cy, cyclophosphamide; Dex, dexamethasone; flu, fludarabine; GCSF, granulocyte colony-stimulating factor;
HSCT, hematopoietic stem cell transplant; HYD, hydroxyurea; HT, hypertransfusion; MMF, mycophenolate mofetil; PBSC, peripheral blood stem
cells; PTIS, pre-transplant immune suppression; TBI, total body irradiation; Thymo, rabbit antithymocyte globulin (Thymoglobulin); TT, thiotepa
4 of 9
2.5 Conditioning regimen
Patients were admitted on Day −8 and underwent central line inser-
tion. Conditioning included rabbit antithymocyte globulin (ATG; Sanofi-
Genzyme) 1.5 mg/kg/day (Days −7 to −5), thiotepa 10 mg/kg (Day
−7) in two divided doses, fludarabine 30 mg/m2 /day (Days −7 to −3),
cyclophosphamide 14.5 mg/kg/d (Days −3, −2), total body irradiation
2 Gy (Day −1) (Figure 2).
2.6 GvHD prophylaxis
GvHD prophylaxis was with cyclophosphamide 50 mg/kg/day (Days
+3, +4), mTOR inhibitor (sirolimus) 2 mg/m2 /day orally once daily
starting from Day +5 and continued until 9 months, with gradual
tapering over 12 weeks if there was no evidence of GvHD, MMF
10 mg/kg/dose thrice daily orally starting from Day +5 until Day +35,
halved for next 7 days and then discontinued (Figure 2).
2.7 Graft
In all the cases, donors were MUD (three sourced from DATRI, an
Indian stem cell registry and one sourced from DKMS, Poland). All
four patients received GCSF-mobilized peripheral blood stem cells
(PBSC) after 5 days of GCSF 10 mcg/kg given subcutaneous once daily.
Peripherally harvested stem cells were collected with a target CD34
dose of 10 million cells/kg recipient body weight. All the MUD products
were cryopreserved before initiating the conditioning chemotherapy.
2.8 Supportive care
All patients received supportive care as per institutional protocol
detailed in our previous publication.25
2.9 Statistical analysis
All continuous variables were presented as medians and range, and cat-
egorical variables as counts and percentages, using Fisher’s exact test
for comparisons. Median follow-up was determined using the reverse
Kaplan-Meier method. The center reports all the transplant outcomes
to the European Society for Blood and Marrow Transplantation with
CIC number 464.
3 RESULTS
3.1 Patients
Four patients aged 5-15 years were enrolled in the trial. Three had
transfusion-dependent β-TM and one had severe SCD. Three were
female; one was male. All patients presented to our center for alter-
KHARYA ET AL .
native donor transplant in the absence of HLA-matched sibling donor.
Baseline characteristics of patients and donors are detailed in Table 1.
3.2 Complications experienced during PTIS
All four patients received PTIS along with hydroxyurea, as detailed
above. Four patients received eight courses of PTIS. All courses of PTIS
were well tolerated, and no patient experienced febrile neutropenia
needing admission. Patient 1 did experience neutropenia without fever,
which responded to short course of GCSF. There was no episode of
cytomegalovirus (CMV) reactivation post PTIS, and none experienced
fungal pneumonia.
3.3 Donor
Median age of the donors was 30.5 years (range 21-45). Two
donor/recipient pairs had major blood group mismatch and one had
minor. For all the ABO mismatch transplants, antibody titers were done
and a cutoff of >1:32 was used for neutralization using fresh frozen
plasma. PBSC were harvested after 5 days of GCSF mobilization with
a target dose of 10 million cells/kg recipient body weight (rBW). All the
products were received at the transplant center before the start of con-
ditioning and were cryopreserved.
3.4 Graft characteristics
CD34+ and CD3+ cell counts were done for all the patients (except
patient 1 where CD3+ could not be performed) from the final product
before cryopreservation. Infusion cutoff of 10 million CD34+ cells/kg
rBW was decided with no capping of CD3+ cells. Median CD34+ cells
infused was 10.73 million cells/kg (range 7.67-12.6). Median CD3+
cells infused were 52.0 × 107 /kg rBW (range 23.9-78.7). Stem cell prod-
uct was infused over 2 days in patients 3 and 4 to avoid fluid overload
and to minimize dimethylsulfoxide-related toxicity.
3.5 Engraftment characteristics
Median time to neutrophil engraftment (first day of absolute neu-
trophil count [ANC] >500/mm3 ) was 15 days (range 12-17) and median
time to platelet engraftment (PC >20 000/mm3 unsupported for more
than 5 days) was 23.5 days (range 12-26).
3.6 Complications experienced post HSCT
(Table 2)
∙ Mucositis: Grade III/IV mucositis was not seen in any patient. Enteral
nutrition was maintained in all the patients. None required par-
enteral nutrition (partial or total) in view of decreased oral intake.
TA B L E 1 Baseline characteristics of recipients and donors

Recipient Donor Cell dose

KHARYA ET AL .

Blood
Age Age group CMV IgG HLA ABO CD34 CD3
Pt. No (years) Sex Disease Disease burden (years) Sex Relation (R/D) (R/D) matching incompatibility (106 /kg) (107 /kg)
1 7 F TM Class III, ferritin 1986 ng/mL, 23 M MUD B+/O+ +/+ 10/10 Minor 10 DNA
liver 2-3 cm BCM
2 15 F SCD Frequent VOCs needing 21 F MUD O+/A+ +/+ 10/10 Major 7.67 23.9
hospitalization; frequent
infections, persistent anemia,
and jaundice; frequent blood
transfusions
3 5 M TM Class III, ferritin 3274 ng/mL, 38 M MUD O+/O+ +/+ 10/10 None 12.6 52.04
liver 3 cm BCM
4 6 F TM Class III, ferritin 3147 ng/mL, 45 F MUD O+/B+ +/+ 10/10 Major 11.46 78.75
liver 3 cm BCM

Abbreviations: CMV, cytomegalovirus; HLA, human leukocyte antigen; MUD, matched unrelated donor; R/D, recipient/donor; SCD, sickle cell disease; TM, thalassemia major/transfusion-dependent thalassemia;
VOC, vaso-occlusive crisis.

TA B L E 2 Engraftment, chimerism, and posttransplant details in recipients

Day of Day Chimerism (% donor) Days Status at
neu- of Viral post Other last
Pt. No trophil platelet ES Day Day Day aGvHD reactivation cGvHD IS HSCT complication follow-up
engraftment engraftment +21 +60 +100
1 12 12 No 99.87 99.69 100 Gut Gr1 No No Ongoing 192 None Alive/DF
2 14 22 No 98.37 98.64 100 No No No Ongoing 154 None Alive/DF
3 16 25 No 99.48 98.83 - No Yes (? BK No Ongoing 55 None Alive/DF
virus)
4 17 26 No 99.4 99.16 - No No No Ongoing 48 None Alive/DF

Abbreviations: aGVHD, acute graft versus host disease; cGVHD, chronic graft versus host disease; DF, disease-free; ES, engraftment syndrome; IS, immune suppression.
5 of 9
6 of 9 KHARYA ET AL .

∙ Peri-engraftment fever (EF)/engraftment syndrome (ES): EF was noticed

in patients 1, 3, and 4. Along with clinical parameters, we also per-

CD56
formed interleukin 6 levels in symptomatic patients close to engraft-

N/A
202

139

145
100
IgA

12
53

34
ment as a part of our ongoing trial on interleukin 6 and cytokine-
release syndrome to assess the clinical utility of early administra-

CD19
tion of tocilizumab. The values were 202 ng/mL in patient 1 and

Immune reconstitution at Day 100

111
309

225
244
27.37 ng/mL in patient 4 at day 14 and 11, respectively; it could not

Immune reconstitution at Day 100

be done on patient 3 due to COVID-mediated lockdown. None of

IgM
N/A
32

29
the patients fulfilled the criteria of ES. GCSF was discontinued in

1023
2983
CD8
948
535
patients 1 and 3, and short course of steroids was given with opti-
mal response. Patient 4 received single-dose tocilizumab at 4 mg/kg
subsequent to which fever settled.

CD4
282
959

618
391
∙ Bacterial/fungal infection: Culture-positive bacterial infection was

1267
N/A

428
380
IgG
not seen in any patient.
∙ CMV reactivation was not seen in any of the patients.

1251
1592

1637
3433
CD3
∙ BK polyoma virus and Epstein Barr virus reactivation: Hemorrhagic cys-
titis was seen in patient 3 at day 30. Urine culture was negative; urine
BK viral load could not be done. He responded to two doses of cido-

CD56

236

213
fovir along with supportive care. Epstein Barr virus reactivation was

<33

30

32
217
IgA
74

44
not seen in any of our patients.
∙ aGvHD: Grade III/IV aGvHD was not seen in any of our patients.

CD19
Patient 1 developed grade I acute gut GvHD, which resolved with

282
189
17

2
Immune reconstitution at Day 60
short course of steroids.
∙ cGvHD: cGvHD (limited or extensive) was not seen in any of our

Immune reconstitution at Day 60

<20.6
<20

<21
patients until last follow-up. IgM

1790
31

CD8
973
268

884
3.7 Chimerism analysis

CD4
138
469
214
929
Sixteen STR loci and one ameloginin locus were used to identify infor-
1214
560

652
431
IgG

mative loci between donor and recipient. The amplicon was resolved by

1116

2043
1811
CD3

746
capillary electrophoresis on Dx 3500 platform. Chimerism was sent at
the time of engraftment and then at Days +30, +60, +100, +180, +270,
and +365 using peripheral blood. At a median follow-up of 307.5 days
CD56

on December 29, 2020 (range 251-395), all four patients had ≥95%
626
130
45

84
118
294

119
IgA

donor chimerism in whole blood (Table 2).

24
Immune reconstitution at Days 30, 60, and 100

CD19

130

3.8 Immune reconstitution (IR)

3
4

6
Immune reconstitution at Day 30

Immune reconstitution at Day 30

IR is always a challenge with alternative donor transplants. Delayed

IgM

316

1936

2798
1048
CD8
21
41

223
7

IR leads to increased risk of secondary infection. For all our patients,

we assessed immunoglobulin levels (G, M, A) and IR panels (CD3, 4, 8,
19 and 56) at Days +30, +60, +100, +180, +270, and +365. Median
2075
CD4
434
116
262

IgG level at Day 100 was 428 mg/dL (range 380-1267). The pattern
of IR is shown in Table 3. Median CD4+ count at Day +100 was
1581
964

645
881
IgG

504.5 cells/mm3 (range 282-959); CD8+ was 985.5 cells/mm3 (range

2382

3098
3108

535-2983); CD19+ was 234.5 cells/mm3 (range 111-309), and CD56+

CD3

336

was 119.5 cells/mm3 (range 34-145).

TA B L E 3

Patient

Patient
No.

No.
1
2
3
4

1
2
3
4
KHARYA ET AL .
4 DISCUSSION
HLA-identical related donor transplant has been standard of care for
transfusion-dependent TM or severe SCD with very good outcomes;
however, this option is available only to 25-30% of the patients in
need.1–3 Alternative donor transplantation methods such as MUD,
CBT, or haploidentical donor have increased the donor pool, making
transplant an option for all transplant-eligible patients but RRT, GF,
and GvHD remain risks.4–11 Bernaudin et al reported a GF rate of
22.6% in SCD patients who received busulfan with cyclophosphamide
as compared to 2.9% in patients who received additional ATG.26 Out-
comes of MUD transplants have improved over the years primarily
involving MAC in combination to serotherapy.27 Various groups have
tried to reduce the intensity of chemo-conditioning, but experienced
mixed chimerism and GF. Shenoy et al reported GF in 10% of chil-
dren undergoing MUD HSCT for severe SCD in the BMT CTN phase
2 trial between 2008 and 2014. The OS and DFS were 86% and 79%,
respectively, at a median follow-up of 26 months. They reported Day
100 incidence of grade II-IV aGvHD as 28% and 1-year incidence of
cGvHD as 62%, 38% of which were extensive.13 In the URTH trial,
which studied outcomes of MUD BMT for patients of TM using RIC,
the OS and DFS at a median follow-up of 58 months were 82% and
79%, respectively. The cumulative incidence of grade II-IV aGvHD rates
in their analysis was 24% and that of cGvHD was 29% at 2 years.14
In another multicentric trial, which studied the outcomes of BMT for
adolescents and young adults with SCD using RIC, Krishnamurti et al
reported 1-year OS and DFS as 91% and 86%, respectively. They
reported grade II-III aGvHD rates of 18% and cGvHD rates of 27%
in their analysis.15 Oshrine et al reported mixed chimerism of 66.7%
and GF of 22.2% using RIC with alemtuzumab.28 Other groups also
raised similar concerns with RIC. Subsequently, the emphasis moved to
pretransplant preparative strategies. Therapies including hydroxyurea
and azathioprine along with hypertransfusion to suppress hyperactive
marrow and suppress endogenous erythropoiesis have achieved some
success.27,29,30
Initial studies reported DFS of 50-80% post HSCT in high-risk class
III TM.6,9,31 Anurathapan et al reported 5-year OS and DFS of 89%
in high-risk class 3 TM patients undergoing HLA-identical transplant
using a combination of fludarabine and dexamethasone based PTIS, fol-
lowed by fludarabine and busulfan based conditioning and calcineurin
inhibitor in combination with MMF as GvHD prophylaxis. The con-
cept was based on a nucleoside analog-alkylating agent platform that
aimed at achieving initial immune suppression by giving nucleoside
analog and subsequently stable engraftment by giving alkylating agent-
based conditioning chemotherapy.32 They reported mild reversible
veno-occlusive disease in 16% patients, CMV reactivation in 16%, and
other viral reactivation in another 16% of patients. Thirty-three per-
cent (6/17) of patients in their study developed grade II-IV aGvHD,
and limited cGvHD was seen in 28% (5/18) patients.20 Our group used
a novel strategy of adding cyclophosphamide to PTIS with 25% dose
reduction in fludarabine with added plerixafor for stem cell mobiliza-
tion in donors and post-PTCy platform for GvHD prophylaxis (APOLLO
protocol) to treat 25 consecutive patients undergoing haploidentical
7 of 9
transplant for severe SCD with very encouraging results.25 In the cur-
rent study, we used a combination of PTIS and PTCy to reduce the risk
of both GF and GvHD in MUD HSCT setting. PTIS in the current study
was different as the dose of fludarabine was reduced to 150 mg/m2 ,
thus making a cumulative dose of fludarabine 300 mg/m2 in compari-
son to Anurathapan et al, where a dose of 400 mg/m2 was used over
two courses. We also added cyclophosphamide at 1000 mg/m2 during
each course of PTIS. Our conditioning platform was a reduced toxicity
augmented Johns Hopkins protocol with added thiotepa. Thiotepa is
known to be an excellent immune- and myelosuppressive agent, with a
gradually increasing role in HSCT.23
The PTCy platform championed by the Johns Hopkins group has
been found to be effective GvHD control strategy in haploidentical
donor setting; however, it remains less explored in HLA-identical
donor settings. By the virtue of causing immune tolerance, it is known
to reduce the risk of aGvHD and cGvHD more effectively than the
conventional GvHD strategies alone.10 Persistence of recipient
antigen-presenting cells in the post-HSCT period is known to increase
the risk of GvHD by reacting to alloreactive donor T cells. Our PTIS
and PTCy platform ameliorated this risk in two ways. Sequential PTIS
helped to suppress recipient T-cell function, whereas PTCy in combina-
tion with mTOR inhibitor and MMF addressed the alloreactive donor
T cells and achieved better immune tolerance, thus minimizing the risk
of GvHD.33 PTIS-mediated recipient T-cell suppression also helped
reduce the risk of GF. Addition of cyclophosphamide in PTIS was a
novel approach that helped reduce the cumulative dose of fludarabine
to 300 mg/m2 . This also helped reduce the RRT by using RTC, which
is known to be better tolerated. None of our patients experienced
grade III/IV mucositis, and we could maintain enteral nutrition during
the entire transplant period. None of our patients developed CMV
reactivation, and only one had hemorrhagic cystitis possibly due to BK
virus (BK virus estimation not done), which responded to supportive
care and two courses of cidofovir.
IR post HSCT has been less studied and reported. Due to fewer
alternative donor transplants, the data on IR post alternative-donor
HSCT are sparse. We previously described our experience of IR post
HHSCT for SCD in 25 consecutive patients treated on APOLLO
protocol.25 In comparison to HHSCT for SCD, CD4+, CD8+, and
CD19+ recovery was better at Day +100 following MUD HSCT; how-
ever, no significant difference was noticed in CD56+ cells. Due to small
sample size, it is difficult to draw any statistically significant conclusion,
and we propose that IR patterns should be studied in larger cohorts
assessing various parameters such as type of donor, source of stem cell,
conditioning chemotherapy (MAC/RIC), choice and dose of serother-
apy, and posttransplant complications warranting need of steroids for
>7-10 days.
We feel addition of PTCy in the current study helped minimize
the risk of grade III/IV aGvHD and also cGvHD, which were not seen
in any of our patients at the last follow-up, although the number
of follow-up days is small to assess the risk of cGvHD. The current
protocol was adopted from our haploidentical transplant protocol for
SCD, where the OS and DFS were 88%.25 However, unlikethe APOLLO
protocol, upfront plerixafor was not used for mobilization of healthy
8 of 9
donors, which possibly led to slightly delayed neutrophil and platelet
engraftment in the current study.
To summarize, our study suggests, based on a small number of
patients, that MUD transplant can safely be offered to patients suf-
fering from high-risk TM or severe SCD with no risk of GF and
minimal risk of GvHD. The use of fludarabine-, cyclophosphamide-,
dexamethasone-based PTIS for recipients and PTCy in combination
with mTOR inhibitor-based GvHD prophylaxis helps to minimize risk of
GF and GvHD. It also helps to modify the conditioning from myeloab-
lative to RTC, thus decreasing RRT. The good outcomes make this
approach feasible and may be preferred over routine management for
patients with TM and SCD. Previous investigators have either focused
on PTIS to target recipient T-cell suppression, or in rare occasions on
PTCy in cases where GvHD was a major concern in MUD settings. We
believe this is the first attempt to combine both approaches for MUD
HSCT for TM or SCD. If substantiated in larger groups of patients, we
believe this approach can be further extended to high-risk TM and SCD
patients and can also be used in HLA-identical related donor settings.
Although the results of the current approach are encouraging, the
study has a few limitations, small sample size being one of them. In view
of the short follow-up, relatively longer follow-up will be required to
better assess the risk of cGvHD with the current approach. Feasibility
also needs to be explored in highest risk TM, which is expected to have
even higher complications with the standard approach while undergo-
ing MUD HSCT.
ACKNOWLEDGMENTS
Authors would like to acknowledge Ms Manju Joseph and the entire
nursing team for the excellent execution of the protocol and state of
art clinical care given to the patients, and to Ms Himshikha Yadav who
fulfilled the job of BMT coordinator. Ms Bharti Sharma, senior data ana-
lyst helped in maintenance and compilation of the entire data and also
in statistical analysis.
CONFLICT OF INTEREST
The authors declare that there is no conflict of interest.
AUTHOR CONTRIBUTIONS
Gaurav Kharya conceptualized the protocol and drafted the paper.
Atish N. Bakane and Archana M. Rauthan compiled the data and did
the statistical analysis (in association with the senior data analyst). All
the authors contributed, read, and approved the final version of the
manuscript.
ORCID
Gaurav Kharya https://orcid.org/0000-0003-2181-1825
Atish N. Bakane https://orcid.org/0000-0001-6403-2309
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How to cite this article: Kharya G, Bakane AN, Rauthan AM.
Pretransplant myeloid and immune suppression, reduced
toxicity conditioning with posttransplant cyclophosphamide:
Initial outcomes of novel approach for matched unrelated
donor hematopoietic stem cell transplant for
hemoglobinopathies. Pediatr Blood Cancer. 2021;68:e28909.
https://doi.org/10.1002/pbc.28909