Small Round Cell Tumors of Bone
Meera Hameed, MD
● Context.—Primary small round cell tumors of the bone
are a heterogeneous group of malignant neoplasms pre-
senting predominantly in children and adolescents. They
include Ewing sarcoma/peripheral neuroectodermal tumor
or Ewing family tumors, lymphoma, mesenchymal chon-
drosarcoma, and small cell osteosarcoma. Even though
they share many morphological similarities, their unique
biological and genetic characteristics have provided sub-
stantial insights into the pathology of these diverse neo-
plasms.
Objective.—To provide an overview of the clinical, ra-
diologic, pathologic, and genetic characteristics of these
tumors along with a pertinent review of the literature.
EWING SARCOMA FAMILY OF TUMORS
Since the original descriptions of tumors of the Ewing
sarcoma family in 1918 by Arthur Purdy Stout1 and in
1921 by James Ewing, who called the tumor diffuse endo-
thelioma of bone,2 several nomenclatures have been assigned
to these tumors, albeit with much skepticism in the liter-
ature3 A possible relationship between Ewing sarcoma
(ES) and peripheral neuroectodermal tumor (PNET) was
established after the description of extraosseous ES by An-
gerwall and Enzinger4 and PNET of the bone by Jaffe.5 At
the present time, ES and PNET, otherwise called Ewing
family tumors (EFTs), form a single neoplastic entity
sharing common histologic and molecular features, and
differing only in their extent of cellular differentiation.
Askin’s tumor of the thoracopulmonary region is also
currently grouped under EFT. Nonrandom translocation
involving the EWS gene with one of the ETS family of
transcription factors is the hallmark of their oncogene-
sis.6–8
Incidence and Location
Ewing family tumor commonly occurs in the adolescent
age group between 10 and 20 years, but is extremely rare
in African American children.9 The annual incidence is
about 3 per million, and there is a slight male predomi-
nance.10 These neoplasms constitute the second most com-
mon pediatric malignancy occurring in the skeletal sys-
tem.10 Adult patients (older than 40 years) with EFT are
rare, but isolated cases in patients up to 81 years old have
been reported.11,12
Ewing family tumor involves both the axial and appen-
dicular skeleton, including the pelvis. Among long tubular
Accepted for publication April 19, 2006.
From the University of Medicine and Dentistry of New Jersey–New
Jersey Medical School, Newark.
The author has no relevant financial interest in the products or com-
panies described in this article.
Reprints: Meera Hameed, MD, Surgical Pathology, UMDNJ–New Jer-
sey Medical School, Newark, NJ 07103 (e-mail: hameedmr@
umdnj.edu).
192 Arch Pathol Lab Med—Vol 131, February 2007
Data Sources.—A literature search using PubMed and
Ovid MEDLINE was performed, and data were obtained
from various articles pertaining to clinicopathologic, bio-
logical, and genetic findings in these tumors. Additionally,
findings from rare cases have been included from author’s
subspecialty experience.
Conclusion.—The diagnosis of small round cell tumors
can be made accurately by applying clinicopathologic cri-
teria, as well as a panel of immunohistochemical and ge-
netic studies in appropriate cases. Molecular genetic stud-
ies may provide further insight into the biology, histogen-
esis, and prognosis of these tumors.
(Arch Pathol Lab Med. 2007;131:192–204)
bones, the femur is most commonly affected, followed by
tibia, fibula, and humerus. However, virtually any bone
can be affected, including the acral skeleton, craniofacial
bones, and clavicle.13 Neoplasms in the long bones are of-
ten diaphyseal.14,15
Clinical Presentation
Local tenderness or mass is the most common clinical
presentation. In about 10% to 15% of the cases, there is a
pathological fracture when the femur is involved.16 It is
not uncommon to encounter children with systemic symp-
toms such as fever, fatigue, leukocytosis, and raised eryth-
rocyte sedimentation rate mimicking signs and symptoms
of acute osteomyelitis.15 This is an important clinical and
radiologic differential diagnosis.
Radiographic Features
Radiographic correlation is a prerequisite to all ortho-
pedic pathology and more so in tumors such as EFT,
where clinical findings may point to benign processes
such as osteomyelitis. The radiographic features are that
of a diaphyseal or metadiaphyseal lesion in the long
bones. Typically, the lesion is intramedullary (symmetric
or eccentric) and is associated with cortical thickening and
cyclical periosteal reaction, giving rise to the characteristic
onion-skin appearance (Figure 1). The rapid growth of the
tumor produces a continuous lifting of the periosteum
leading to perpendicular striations. Within the marrow,
the lesion is poorly marginated, with a permeative pattern
of osteolysis (without a beginning and an end) (Figure 1).
Magnetic resonance imaging reveals an accompanying
soft tissue mass in about 90% of cases (Figures 2 and 3).
Computed tomographic scan will demonstrate that the le-
sion itself does not produce any matrix (bone or cartilage).
Occasionally there is prominent diffuse thickening of the
bone and cloudy opacities mimicking osteosarcoma (OS).
Radiologic differential diagnoses include osteomyelitis,
Langerhans histiocytosis, lymphoma, and OS.17 Imaging
workup of these patients involves skeletal scintigraphy
(bone scan) and computed tomography of the chest to rule
out metastatic disease at the time of presentation.
Small Round Cell Tumors of Bone—Hameed
Figure 1. Plain x-ray. Diaphyseal intramedullary lesion of radius showing cyclical periosteal reaction with periosteal elevation and onion skinning.
Figure 2. Coronal magnetic resonance imaging without contrast showing the lesion occupies the entire marrow cavity encircling the entire cortex
with soft tissue extension.
Pathology
An open biopsy is usually performed to obtain adequate
material for pathological examination and ancillary stud-
ies. Multiple core biopsies may be performed in patients
with large, accessible soft tissue component of the tumors,
keeping in mind that additional studies such as cytoge-
netics will be needed. Fine-needle aspiration can be used
in an appropriate setting where adequate material can be
obtained. Grossly, the biopsied material is often hemor-
rhagic and necrotic and usually has a tan-gray appear-
ance. Touch imprints of the submitted pieces allow one
not only to examine morphology, but also to select viable
material for cytogenetic analysis. Molecular studies (re-
verse transcriptase polymerase chain reaction [RT-PCR]
and fluorescence in situ hybridization [FISH]) can be done
using frozen and paraffin-embedded tissue. If tissue is
snap-frozen for molecular studies, care should be taken to
freeze tumor material in a timely manner to avoid RNA
degradation. Whenever possible, bony spicules should be
separated from soft tissue to avoid unnecessary decalci-
fication of the soft tissue component.
Microscopic examination of classic EFT shows a highly
cellular neoplasm consisting of a monotonous population
of uniform round blue cells (diameter, about 14 ␮m) ar-
ranged in a nested, sheetlike, or solid pattern. The nuclei
are round with fine chromatin, scant clear or eosinophilic
cytoplasm, indistinct cytoplasmic membranes, and 1 to 3
small to medium-sized nucleoli. The cytoplasm contains
Arch Pathol Lab Med—Vol 131, February 2007
abundant glycogen, which can be confirmed with periodic
acid–Schiff stain. Necrosis and apoptosis are common. The
presence of apoptotic cells sometimes gives the biphasic
appearance of dark and light cells (Figure 4). Homer-
Wright rosettes and pseudorosettes can be present (Figure
5). Stromal elements are minimal, and the tumor generally
fills the medullary cavity and replaces bone marrow ele-
ments. Interspersed collagen bundles are usually seen in
the soft tissue component. A vascular component is not
prominent and is usually composed of slitlike capillaries.
Thick-walled vessels are seen in areas where there are
prominent stroma and collagen bundles. A minimal de-
gree of spindle cell change can be present. The tumor cells
permeate the cortex and elicit periosteal new bone for-
mation with reactive osteoblasts, osteoclasts, and some-
times cartilaginous metaplasia. The tumor generally does
not induce an inflammatory response.
Morphological Variants
A number of variants have been described in EFT. These
include atypical ES (large cell EFT),18 adamantinoma-like
ES,19,20 sclerosing ES, and spindle cell sarcoma–like EFT.21
The tumor cells of the atypical or large cell EFT are het-
erogeneous compared with classic EFT. Their size ranges
between 20 and 24 ␮m, and they have irregular nuclear
membranes and prominent nucleoli (Figure 6). The cyto-
plasm is vacuolated, and cytoplasmic glycogen is less
Small Round Cell Tumors of Bone—Hameed 193
Figure 3. Axial T2-weighted magnetic resonance imaging without contrast showing the lesion occupies the entire marrow cavity encircling the
entire cortex with soft tissue extension.
Figure 4. Photomicrograph of Ewing sarcoma showing diffuse monotonous round cells with dark and light pattern (hematoxylin-eosin, original
magnification ⫻400)
Figure 5. Rosettelike pattern in Ewing sarcoma (hematoxylin-eosin, original magnification ⫻400).
Figure 6. Photomicrograph of large cell Ewing family tumor with irregular nuclear contours and prominent nucleoli (hematoxylin-eosin, original
magnification ⫻400).
Figure 7. Pelvic atypical Ewing sarcoma showing nests of tumor cells in a desmoplastic stroma (hematoxylin-eosin, original magnification ⫻200).
Figure 8. Interphase fluorescence in situ hybridization section showing split signal with EWS probes (yellow signal normal and green and red
signal representing split EWS).

abundant than in conventional EFT. In some cases, the
large cells can be accompanied by spindled tumor cells.
Adamantinoma-like EFT has been described in the tibia
and fibula and shares histological and immunohistochem-
ical features of adamantinoma, such as distinct nests of
moderately pleomorphic tumor cells with peripheral pal-
isading, a prominent desmoplastic host response, and pos-
itive immunohistochemistry for cytokeratins. These tu-
mors are also positive for CD99 and FLI-1 and show EWS-
FLI-1 fusion transcript.19 We have seen a tumor of the pel-
vis with similar histology which showed positive staining
for pan-cytokeratin with a variant EWS translocation, as
evidenced by positive break-apart FISH with EWS probe
(Figures 7 and 8). RT-PCR for EWS-FLI-1, ERG, and WT1
fusion transcripts were negative in this tumor. Sclerosing
EFT resembles sclerosing epithelioid fibrosarcoma or scle-
rosing rhabdomyosarcoma because of the presence of hy-
alinized eosinophilic matrix. However, there are always at
least focal areas showing typical features of classic EFT.21
Spindle cell sarcoma–like EFT shows a greater degree of
spindling than classic EFT and thus can resemble synovial
sarcoma.21
Ultrastructure
Transmission electron microscopy of EFTs demonstrates
densely packed undifferentiated tumor cells with cyto-

Table 1. Immunohistochemical Profile*

Tumor CD99 FLI-1 LCA B Cell T Cell TdT CK Chr S100
EFT ⫹ ⫹ ⫺ ⫺ ⫺ ⫺ ⫹/⫺ ⫺ ⫹/⫺
LBL/ALL ⫹ ⫹ ⫹/⫺ ⫹/⫺ ⫹/⫺ ⫹ ⫹/⫺ ⫺ ⫺
NHL–other ⫹/⫺ ⫹ ⫹ ⫹ ⫹ ⫺ ⫺ ⫺ ⫺
Mesenchymal chondrosarcoma ⫹ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫹
Small cell OS ⫹/⫺ ? ⫺ ⫺ ⫺ ⫺ ⫹/⫺ ⫺ ⫹/⫺
Rhabdomyosarcoma ⫹/⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫹/⫺
DSCRT ⫹ ⫹/⫺ ⫺ ⫺ ⫺ ⫺ ⫹ ⫹/⫺ ⫹/⫺
Neuroblastoma ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫺ ⫹ ⫹/⫺
* LCA indicates leukocyte common antigen; TdT, terminal deoxynucleotidyltransferase; CK, pan-cytokeratin; Chr, chromogranin; EFT, Ewing
family tumors; LBL/ALL, lymphoblastic lymphoma/acute lymphoblastic leukemia; NHL, non-Hodgkin lymphoma; OS, osteosarcoma; DSCRT, des-
moplastic small round cell tumor; and ?, not known.

194 Arch Pathol Lab Med—Vol 131, February 2007 Small Round Cell Tumors of Bone—Hameed
plasmic glycogen and scarce organelles. Occasionally, des-
mosome-like junctions are seen. Evidence of neural differ-
entiation such as dense core neurosecretory granules, den-
dritic processes, and microtubules can be present.22,23
Immunohistochemistry
It is advisable to use a panel of immunohistochemical
markers in small round cell tumors of the bone which in-
clude vimentin, CD99, FLI-1, leucocyte common antigen,
terminal deoxynucleotidyl transferase (TdT), B-cell, T-cell
markers, pan-cytokeratin, desmin, MYOD1/myogenin,
chromogranin, and S100 protein. These markers assist in
differentiating EFT from other tumors such as lympho-
blastic lymphoma/acute lymphoblastic leukemia, rhab-
domyosarcoma, desmoplastic small round cell tumor, and
neuroblastoma. Other differential diagnoses to consider
would be mesenchymal chondrosarcoma, non-Hodgkin
lymphoma (NHL) (other), and small cell OS. The most
useful, though nonspecific, marker in the diagnosis of EFT
is CD99, which often produces a diffuse membranous
staining pattern in these tumors (Figure 9). This product
is a 32-kd transmembrane protein whose function is not
well established; it is encoded by the MIC2 gene located
in the pseudoautosomal region of X and Y chromo-
somes.24–26 Apart from EFT, this protein is also highly ex-
pressed in T and B lymphocytes, hence also expressed by
lymphoblastic lymphoma/acute lymphoblastic leukemia.
In addition, positive staining has been observed in some
rhabdomyosarcomas, desmoplastic small round cell tu-
mors, synovial sarcomas, and mesenchymal chondrosar-
comas.27–30 Another useful immunomarker is the FLI-1
protein, which is expressed in about 84% of EFTs (Figure
10).31 This transcription factor is also expressed by lym-
phoblastic lymphomas and other NHLs.31 Other variably
expressed markers in EFT include vimentin, cytokeratin
(up to 25%), desmin (very rare), and neural markers such Figure 9. Immunohistochemical stain for CD99 showing diffuse pos-
as synaptophysin, CD57, S100, and chromogranin.21,23,32–34 itivity in typical Ewing sarcoma (original magnification ⫻400).
The presence of neural markers in an EFT indicates neu-
Figure 10. Immunohistochemical staining for FLI-1 showing intense
roectodermal differentiation (PNET). High molecular cy- nuclear staining in tumor cells.
tokeratin (34␤E12) is positive in adamantinoma-like ES
and not in classic and other variants of EFT.21 Table 1 sum-
marizes the immunohistochemical profile of these tumors. some 22q12 to the FLI-1 gene at chromosome 11q24. The
EWS-FLI-1 chimeric protein on the (der)22, contains the 5’
Cytogenetics and Molecular Genetics end of the EWS gene and the 3’ end of the FLI-1 gene.38
The pathogenetic relationship of ES and PNET was de- The second most common translocation in EFT is t(12;
fined by the discovery of the unique and specific nonran- 22)(q22;q12), seen in about 5% to 10% of the cases, re-
dom chromosomal translocation, seen in about 85% to 95% sulting in the fusion of EWS to ERG at chromosome
of the tumors, involving chromosome 11q24 and chro- 12q22.39 Other variant translocations reported in bone EFT
mosome 22q12, t(11;22)(q24;q12) (Figure 11).35–37 This accounting for less than 1% of cases include t(7;22)(p22;
leads to an in-frame fusion of the EWS gene at chromo- q12), t(17;22)(q12;q12), and t(2;22)(q23;q12), involving fu-
sion of EWS to ETV1, E1AF, and FEV genes, respectively
(Table 2).40–43
The EWS gene has 17 exons, of which the first 7 exons
Table 1. Extended
encode the N-terminal region responsible for regulating
Desmin Myogenin/MYOD1 WT1 its RNA binding capacity. It belongs to the TET family of
⫺ ⫺ ⫺ RNA binding proteins (EWS, TLS, TAFII68) whose amino
⫺ ⫺ ⫹/⫺ terminal (NTD) has the capacity to bind to DNA-binding
⫺ ⫺ ⫺ domains of various transcription factors, thus providing a
⫹/⫺ ⫹/⫺ ⫺ strong transcriptional activating domain to the resulting
⫹/⫺ ? ?
⫹ ⫹ ⫹/⫺(cyto)
chimeric protein.44,45 EWS itself is ubiquitously expressed
⫹ ⫺ ⫹ in all tissues.46 The partner genes of EWS in the EFT of
⫺ ⫺ ⫺ bone, such as FLI-1, ERG, ETV1, EIAF, and FEV, belong to
the ETS family of transcription factors possessing a con-
served ETS domain recognizing a core DNA motif of
GGAA/T.47 FLI-1 expression is seen in high levels in he-
Arch Pathol Lab Med—Vol 131, February 2007 Small Round Cell Tumors of Bone—Hameed 195
Figure 11. Karyotyping of Ewing sarcoma with t(11;22)(q24;q12).
Figure 12. EWS-FLI-1 gene fusion diagram showing common fusion products with long arrows indicating the most common break points. All
fusions include exons 1 through 7 of the EWS gene (NTD) and FLI-1 exon 9 (DNA binding domain). The involved exons are shown as numbers
for each transcript of EWS-FLI-1.
Figure 13. Reverse transcriptase polymerase chain reaction showing 329 bp type I fusion product of EWS-FLI-1.
Figure 14. Metaphase fluorescence in situ hybridization with EWS dual-color break-apart probe showing translocation of green signal from
chromosome 22 to chromosome 11.

matopoietic cells, endothelial cells, and mesenchyme de- the EWS-ETS fusion are FISH and RT-PCR. Multiple chi-
rived from neural crest cells.48,49 The product of EWS-ETS meric transcripts are possible, all of which involve exons
translocation results in fusion proteins containing the C- 1 through 7 of EWS and exon 9 of the ETS gene. The
terminal region of the FLI-1 and N-terminal region of genomic break points are intronic, and occur at 1 of 4
EWS, thus bringing the ETS component under the pow- EWS introns and, in the case of EWS-FLI-1, 1 of 6 FLI-1
erful transactivating domain of EWS.50 These fusion prod- introns (Figure 12).51 Type 1 (exon 7 of EWS to exon 6 of
ucts modulate multiple target genes, resulting in malig- FLI-1) and type II (exon 7 of EWS to exon 5 of FLI-1) are
nant transformation. the most common (⬎85%) fusion transcripts.52 Commonly
The most commonly used molecular assays to identify used RT-PCR assays use EWS exon 7 forward primer and
FLI-1 exon 9 reverse primer, which amplifies all fusion
products, or EWS exon 7 forward primer and FLI-1 exon
Table 2. Translocation and Fusion Genes in Ewing 6 to detect type 1 or type II fusion products, which would
Family Tumors of Bone identify 85% of the cases (Figure 13). In addition, RT-PCR
Translocation Fusion Genes Frequency for EWS-ERG fusion product can be performed, which
t(11;22)(q24;q12) EWS-FLI-1 85%–95% would identify the 10% of cases with this fusion. A mul-
t(21;22)(q22;q12) EWS-ERG 5%–10% tiplex real-time PCR has also been used to identify EWS-
t(7;22)(p22;q12) EWS-ETV1 ⬍1% ETS fusion products.53 RT-PCR has also been applied to
t(17;22)(q12;q12) EWS-E1AF ⬍1% formalin-fixed, paraffin-embedded tissues, and a combi-
t(2;22)(q23;q12) EWS-FEV ⬍1% nation of FISH and RT-PCR may yield increased sensitiv-
196 Arch Pathol Lab Med—Vol 131, February 2007 Small Round Cell Tumors of Bone—Hameed

ity and accuracy due to limitations of formalin-fixed, par-
affin-embedded tissue material with suboptimal preser-
vation of nucleic acids.54 Fluorescence in situ hybridization
assay with the commercially available (such as Vysis)
dual-color break-apart DNA probes flanking the EWS
break point region on chromosome 22 detects the trans-
location of EWS and can be used in fresh and formalin-
fixed, paraffin-embedded tissues. In a metaphase spread,
the split signal enables one to identify the chromosome
involved in the translocation; however, the partner gene is
not identified by this methodology (Figure 14).
Additional reported secondary karyotypic changes in
EFT include gains of chromosomes 8, 12, and 18, deletion
of the short arm of chromosome 1, unbalanced translo-
cation t(1;16)(q12;q11), and other rare structural abnor-
malities.55,56
Treatment and Treatment Response
The management of EFTs includes systemic chemother-
apy followed by surgery, with or without radiation and
continuation chemotherapy.57 Induction chemotherapy be-
fore definitive surgery allows us to assess the chemores-
Table 3. Chemotherapy Response—Pediatric
Oncology Group Study
Grade Tumor Response 3-y Survival, %
I No chemotherapy effect 30
IIA 1%–10% necrosis 30
IIB 11%–90% necrosis 49
III 91%–99% necrosis 73
IV 100% necrosis 100
Arch Pathol Lab Med—Vol 131, February 2007
Figure 15. Gross photograph (A) and spec-
imen x-ray (B) of resected Ewing sarcoma of
radius after chemotherapy. The grid lines
serve as a map for histology sections.
Figure 16. Plain x-ray of proximal femur
showing a destructive moth-eaten lesion with
mixed lytic and sclerotic areas involving me-
taphysis and diaphysis. Periostitis is seen ad-
jacent to lateral cortex.
ponse in resected specimens, which has some prognostic
significance and has a role in selection of agents for con-
tinuation chemotherapy. A sagittal section of the resected
bone, with mapping and examining the entire central face
of the tumor and adjacent tissue after careful decalcifica-
tion, allows one to assess the chemotherapeutic response
(Figure 15).58 The grading system used to assess chemo-
response is similar to that used in OS (Table 3).58,59
Prognosis and Outcome
Up to one third of patients with EFT have metastatic
disease at the time of presentation, and their outcome re-
mains poor in spite of aggressive chemotherapy.60 The
most common sites of metastases are lung, bone, and bone
marrow. Patients who present only with lung metastases
have better survival compared with those who present
with bone metastases.61 One third of the patients with lo-
calized disease are also prone to distant relapse, especially
if there are residual viable tumor cells at definitive resec-
tion.61,62 Multiple biological factors, such as type of fusion
transcript, mutations in INK4a gene, mutations in p53, de-
letion of p16, telomerase, IGF-1, and aneuploidy, have been
implicated to play a role in the prognosis of EFT.63,64 In
patients with localized disease, EWS-FLI-1 type 1 fusion
transcript has been reported as being associated with im-
proved outcome compared to other fusion transcript
types, and it appears that type 1 fusion transcript encodes
a less active chimeric protein.65,66 Reports on the detection
of minimal residual disease by RT-PCR for the chimeric
proteins in peripheral blood and bone marrow remain
controversial with respect to disease progression.67–70 Cur-
rent chemotherapeutic regimens with selective surgery
Small Round Cell Tumors of Bone—Hameed 197
have significantly improved the overall survival up to 60%
to 80% in nonmetastatic EFT.71 Radiation-induced sarco-
mas and chemotherapy-related acute leukemias are the
most frequent secondary malignancies.13
Conclusion
Even though the cell of origin is still unknown in EFT,
considerable strides have been made since the original de-
scription by Drs Stout and Ewing. The combination of
morphology, immunohistochemistry, and molecular ge-
netics has opened many doors, including some novel ap-
proaches to therapeutics, such as the use of CD99 as a
target,72 inhibition of IGF-1 receptor with antisense oligo-
nucleotides,73 and small interfering RNA (siRNA)74 all of
which have shown promising results in cell lines and an-
imal models.
NON-HODGKIN LYMPHOMA
Primary NHL of bone was described as a series, first by
Jackson and Parker in 1939.75 Before this, Oberling in 1928
had reported a case of reticuloendothelial sarcoma histo-
logically indistinguishable from ES.76 The description of
this rare entity is restricted to lymphoma involving a sin-
gle skeletal site with or without regional lymph node in-
volvement or multiple skeletal sites without visceral or
lymph node involvement.77
Incidence and Location
Approximately 3% to 7% of extranodal lymphomas pre-
sent as primary bone neoplasms,78–81 and lymphomas con-
stitute about 7% of all bone malignancies.77 The disease
tends to involve older adults (older than 40 years), and
there is a male predominance, with male-female ratio
ranging from 1.2:1 to 1.6:1.82 The femur, pelvis, vertebra,
and humerus are the most common sites. In the long
bones, the preferred site is the metadiaphyseal region.83,84
Involvement of the small bones of the hand and feet is
extremely unusual. It is not uncommon to encounter mul-
tifocality, and soft tissue extension may be present.77,83
Clinical Presentation
Most patients present with pain, and depending on lo-
cation, such as spine, neurological symptoms may be pres-
ent. Systemic symptoms such as fever (as seen in ES) and
B symptoms of nodal lymphomas are unusual.83 Occa-
sionally patients can present with hypercalcemia, lethargy,
constipation, etc.85 For a neoplasm to be classified as a
primary bone lymphoma, an interval of 4 to 6 months
between the skeletal manifestation and extraskeletal dis-
ease is required.86 Thus appropriate staging procedures,
such as skeletal imaging, computed tomographic scans,
bone scan/positron emission tomography, and bone mar-
row biopsy, are an integral part of the patient workup.87
Radiographic Features
On conventional x-rays, primary lymphomas are lytic
radiolucent lesions with a moth-eaten or permeative de-
structive pattern. This may be associated with sclerotic ar-
eas (Figure 16).84,88 These changes are nonspecific and
overlap with other round cell tumors. Cortical erosion and
destruction with soft tissue extension is frequently seen
(Figure 17). Periosteal reaction (interrupted or solid single-
layer) can be present, but less so than in ES. Generally the
disease tends to involve the long bones extensively, some-
times occupying the entire length. In some instances, the
198 Arch Pathol Lab Med—Vol 131, February 2007
tumor may evoke a sclerotic response and present as a
blastic lesion, which can sometimes be mistaken for Paget
disease. Computed tomography and magnetic resonance
imaging are useful in assessing extent of disease, and pos-
itron emission tomography and bone scans are used for
staging and assessment of remission after therapy. Of im-
portance is the fact that occasionally plain radiographs can
be normal, and persistence of clinical symptoms would
warrant further investigation such as magnetic resonance
imaging, which will demonstrate the signal abnormalities
of the marrow. Pathologic fracture can be present in 22%
of cases.89 Due to the nonspecificity of the radiographic
findings, the differential diagnoses include OS, eosino-
philic granuloma, metastases of solid tumors, round cell
tumors (ES, neuroblastoma, rhabdomyosrcaoma), and
chronic osteomyelitis.
Pathology
Grossly, bone lymphomas tend to be fleshy gray-white
or hemorrhagic, and the gross appearance is also related
to the response of the host bone, leading to sometimes
very sclerotic and bony biopsy specimens requiring de-
calcification. Histologically, the most common finding is a
diffuse pattern of growth with tumor cells permeating be-
tween bony trabeculae and invading Haversian channels
of the cortex. There is often a sclerotic or collagenized
stroma, and the tumor cells can invade in cords, nests, or
solid sheets. As is often evidenced in the x-ray, the host
bony trabeculae can be thickened and irregular with ex-
tensive sclerosis. The most frequent type (92%)82 of pri-
mary NHL of bone is diffuse large B-cell lymphoma
(DLBCL). Unlike nodal lymphomas, DLBCL of bone char-
acteristically shows a polymorphous appearance (Figures
18 and 19). The tumor cells themselves show marked var-
iation in size and shape, often with multilobation, and are
accompanied by a mixture of small and medium-sized
lymphocytes that are reactive B and T cells. The large tu-
mor cells can have prominent nucleoli, and the cytoplasm
is not usually abundant. Occasionally, the fibrotic stromal
reaction elicited by the tumor cells can result in tumor cell
spindling with storiform features, mimicking a sarcoma.
The polymorphous nature of the infiltrate can lead to a
mistaken diagnosis of osteomyelitis, which is also a radio-
logic differential diagnosis. Crush artifact is fairly com-
mon due to the delicate nature of the tumor cells and can
be a vexing problem in pathologic diagnosis. Apart from
DLBCL, other NHLs, which can occasionally present as
primary neoplasms of bone, include anaplastic large cell
lymphoma, lymphoblastic lymphoma, Burkitt lymphoma,
and T-cell lymphomas.82,90,91 Of these, lymphoblastic lym-
phoma can be a diagnostic challenge due to morphological
and immunohistochemical overlap with ES.90 Follicular
lymphomas and small lymphocytic lymphomas such as
chronic lymphocytic lymphoma generally do not present
as primary destructive bone lesions.77
Immunohistochemistry and Genetics
In a suspected case of NHL, a panel of markers is used
to delineate the lineage and type of the neoplasm. They
include leucocyte common antigen, B-cell markers (CD20),
and T-cell markers (CD2, CD3, CD5, CD7), and additional
markers such as CD30, epithelial membrane antigen, and
Alk-1 to rule out anaplastic large cell lymphoma. In a case
with a monomorphic morphology such as lymphoblastic
Small Round Cell Tumors of Bone—Hameed
Figure 17. Computed tomographic scan showing sclerosis in medullary canal with anterior cortical destruction and soft tissue extension.
Figure 18. Photomicrograph of diffuse large B-cell lymphoma showing a diffuse polymorphous infiltrate composed of small, medium, and large
cells (hematoxylin-eosin, original magnification ⫻200).
Figure 19. Immunohistochemical stain showing diffuse positivity for CD20, B-cell marker (original magnification ⫻400).
Figure 20. Photomicrograph of atypical Burkitt lymphoma with large lymphoid cells with prominent nucleoli and mitotic figures (hematoxylin-
eosin, original magnification ⫻400).

lymphoma, the panel should include all of the lymphoma
and other round cell tumor markers (CD45, CD20, CD2,
CD3, CD5, CD7, TdT, vimentin, CD99, FLI-1, keratin, des-
min, chromogranin). In addition, other lymphoid antigens
such as CD79a and CD43 may be required, as lympho-
blastic lymphomas are often negative for CD20 and posi-
tive for CD99 and FLI-1. They can also be negative for
leucocyte common antigen, and in some instances show
focal cytoplasmic keratin positivity, a major pitfall that can
lead to an erroneous diagnosis of ES.90 In pediatric cases
of small round cell tumors, TdT appears to be a sensitive
and specific marker for precursor B or T lymphoblastic
lymphomas and should be included in the immunopanel
(Table 1). Other special stains such as ␬ and ␭ can be used
to ensure monoclonality, but the results are often variable
due to technical limitations. Primary T-cell lymphomas of
the bone are extremely rare, and the diagnosis is estab-
lished based on morphology of the polymorphous popu-
lation with large tumor cells and loss of pan–T-cell anti-
gens such as CD7. Two cases of primary adult T-cell leu-
kemia of the bone have been reported.91 Additional studies
Arch Pathol Lab Med—Vol 131, February 2007
such as flow cytometry can be used in appropriate cases.
B-cell and T-cell rearrangement studies can be performed
in fresh and paraffin-embedded tissues to confirm clon-
ality in doubtful cases. Specific cytogenetic studies of pri-
mary bone lymphomas are sparse. There is a single report
of a pediatric DLBCL of the spine with t(3;22)(q27;q11).92
Recently we encountered a case of an atypical Burkitt lym-
phoma arising in the calcaneus of an elderly male patient
histologically mimicking DLBCL (Figure 20); however, cy-
togenetic analysis showed t(8;14)(q24.1;q32), characteristic
of a Burkitt lymphoma.
Treatment and Prognosis
The primary modality of therapy for primary bone lym-
phomas is chemotherapy and radiotherapy, depending on
the stage of disease and the grade and type of the lym-
phoma.93 The prognosis is dependent upon the stage of
the disease. Patients with stages I and II have a very good
prognosis compared with patients with stages III and IV.94
The 5-year overall survival ranges between 56% and
61%.82,95
Small Round Cell Tumors of Bone—Hameed 199

Figure 21. Plain x-ray of proximal femur showing a permeative pat-
tern in the intertrochanteric and subtrochanteric region with endosteal
scalloping across the distalmost aspect of the neoplasm. There are
questionable foci of matrix calcification proximally. (Courtesy of Dr
Joseph Benevenia, Department of Orthopedics, University of Medicine
and Dentistry of New Jersey, Newark.)
Conclusion
Primary NHL of bone is a rare but well-defined entity
with radiologic and morphologic heterogeneity. The his-
tologic and immunohistochemical similarities, ranging
from benign conditions such as osteomyelitis to monoto-
nous round cell sarcomas, should be borne in mind when
assessing these neoplasms to avoid misdiagnosis and in-
appropriate treatment.
MESENCHYMAL CHONDROSARCOMA
This rare neoplasm was first described by Lichtenstein
and Bernstein96 in 1959 as an unusual variant of chondro-
sarcoma affecting bone and soft tissues. This was followed
by recognition of 9 more cases from the files of the Mayo
Clinic by Dahlin and Henderson.97 Subsequently it has
been established as a separate entity distinct from conven-
tional and dedifferentiated chondrosarcoma.
Incidence and Location
Mesenchymal chondrosarcomas constitute less than 2%
of all chondrosarcomas.98,99 The tumor most commonly af-
200 Arch Pathol Lab Med—Vol 131, February 2007
flicts young adults and teenagers, with a peak incidence
during the third decade.100 Males and females are equally
affected.98 Craniofacial bones, especially the jaw, are most
frequently affected.101 Other commonly involved sites in-
clude vertebrae, ribs, pelvis, and humerus. About one
third of these tumors are extraskeletal and arise in soft
tissues, and meninges are a favored extraskeletal location
for these tumors.98,99 Skeletal neoplasms can be multifo-
cal.99
Clinical Presentation
Pain and swelling are the most common clinical fea-
tures, usually several days to months in duration,102 and
oncogenic osteomalacia (tumor-induced osteomalacia) has
been reported.103
Radiographic Features
Plain x-ray reveals a radiolucent lesion, usually eccen-
tric with stippled calcifications denoting its chondroid na-
ture (Figure 21). The lesion is often destructive, with ex-
pansion of bone, cortical destruction, and extraosseous ex-
tension (Figure 22). Sometimes the calcifications can be
large and present as discrete opacities or heavily calcified
masses. Rarely, a sharply demarcated sclerotic margin is
present. Magnetic resonance imaging appearance can be
variable and can include a heterogeneous, low attenuation
on T1-weighted images (Figure 23) and high signal inten-
sity on T2-weighted images.104,105
Pathology
Biopsy of the lesion generally shows a grossly grey to
pink tissue with foci of mineralization. Microscopically,
the tumor shows a biphasic pattern made up of solid areas
of round or spindle mesenchymal cells interspersed with
islands of well-differentiated cartilage (Figure 24). The
mesenchymal component often exhibits a hemangioperi-
cytic pattern with multiple vascular spaces. The cartilage
component is variable and can show endochondral ossi-
fication with bone formation, coarse calcifications, or
loosely arranged mature hyaline cartilage. There can be
distinct demarcation or gradual blending between the
mesenchymal component and the chondroid component
(Figure 25).98,99 The mesenchymal cells when they are
round can simulate ES or small cell OS in biopsy speci-
mens. On the other hand, the cartilage component alone
in a biopsy can lead to the erroneous diagnosis of a benign
cartilaginous neoplasm.
Ultrastructure
The primitive mesenchymal cells show round to oval
cells with little intercellular matrix, and the cartilaginous
foci reveal mature cells containing glycogen, rough en-
doplasmic reticulum, and numerous mitochondria.106
Immunohistochemistry
The chondroid foci stain for S100 protein, and the prim-
itive mesenchymal cells are positive for CD99 (Figure
26).107 Both components are vimentin positive. It has been
proposed that immunohistochemical positivity for colla-
gen II and IIA, which are considered to be markers of
chondroprogenitor cells, could be used to differentiate
mesenchymal chondrosarcoma from other small round
cell malignancies, such as small cell OS and ES.108 Another
study109 has shown that the transcription factor Sox9, a
master regulator of chondrogenesis,110,111 differentiates
Small Round Cell Tumors of Bone—Hameed
Figure 22. Computed tomographic scan. Mesenchymal chondrosarcoma of the rib with cortical destruction and soft tissue mass with calcification.
(Courtesy of Dr John Reith, Department of Pathology and Orthopedics, University of Florida, Gainesville.)
Figure 23. T1-weighted magnetic resonance imaging of mesenchymal chondrosarcoma showing lobulated heterogeneous soft tissue mass of low
signal intensity. (Courtesy of Dr John Reith, Department of Pathology and Orthopedics, University of Florida, Gainesville.)

mesenchymal chondrosarcoma from various small cell SMALL CELL OSTEOSARCOMA

malignancies, including small cell OS, lymphoma, and ES,
by immunohistochemical analysis using a rabbit polyclon-
al antibody.
Cytogenetics and Molecular Genetics
There are a few reported cytogenetic alterations in mes-
enchymal chondrosarcoma, and so far no specific or re-
current translocations have been identified. Case reports
of near-tetraploid and other diverse structural abnormal-
ities,112 translocation der(13;21)(q10;q10) with loss of chro-
mosomes 8 and 20 material and gain of chromosome 12
material in 2 cases,113 and 1 case with t(11;22)(q24;q12)
similar to ES have been reported.114 Molecular genetic
analyses for mutational alteration of p53 and p16 tumor
suppressor genes have shown a low incidence (⬍25%) in
these tumors.115,116 Biologic pathways involving PKC-␣,
PDGFR-␣, and Bcl-2 expression seem to play a role in
pathogenesis.117
Treatment and Prognosis
Mesenchymal chondrosarcomas are primarily treated
with surgery. They are aggressive lesions with a high pro-
pensity for metastases; however, some patients can have a
protracted clinical course, so the behavior can be unpre-
dictable.118 The overall 10-year survival rate is approxi-
mately 25%.99 Tumors of the jawbones tend to have a more
indolent course.119 Negative correlation between survival
times and proliferation rates using Ki-67 in 10 patients has
been recently reported.120 Distant metastases which can
occur after many years can involve other bones, lung,
etc.99,118 Hence long-term follow-up of these patients is ad-
vocated.
Arch Pathol Lab Med—Vol 131, February 2007
This tumor is discussed here in the context of the dif-
ferential diagnosis of the above-mentioned small round
cell tumors of the bone. This is a distinctive microscopic
variant of OS, originally described in 1979, as a tumor
simulating ES.121 This rare tumor accounts for about 1%
to 1.5% of OSs and has similar age, sex, and skeletal dis-
tribution as conventional OS.122 Radiologically the tumor
can resemble ES or conventional OS. The majority of the
lesions show a mixed lytic and blastic pattern, but they
can be purely lytic or permeative; they usually involve
metaphysis and may extend into epiphysis. A soft tissue
component is present in most of the cases.123–126 Three his-
tologic patterns have been described: ES-like, lymphoma-
like, and small spindle cell.123 The cells can be round and
uniform, similar to ES, or show variation in size. Nucleoli
can be inconspicuous to prominent. The chromatin can be
fine to hyperchromatic. The nuclei can be very small to
medium (6.7–15 ␮m), thus showing a spectrum of sizes
between ES and large cell lymphoma.125 Hemangioperi-
cytic pattern, myxoid change, cordlike arrangement, and
epithelioid features can be seen. The presence of malig-
nant cartilage in some tumors can simulate mesenchymal
chondrosarcoma.125 The presence of osteiod is a prereq-
uisite for diagnosis. The new osteoid matrix is usually
lacelike and can be difficult to find in small biopsies (Fig-
ure 27). The presence of mineralized matrix in imaging
studies supports the diagnosis of OS and is helpful in
differentiating other small blue cell tumors of bone. ES
may show a blastic component in the medullary cavity,
but production of matrix outside the bone is seen only in
OS.123,127,128 Cytoplasmic glycogen can be seen in both neo-
plasms.123 Immunohistochemical features do not distin-
guish small cell OS from sarcomas, as CD99 can be pos-
Small Round Cell Tumors of Bone—Hameed 201
Figure 24. Microscopic photograph of mesenchymal chondrosarcoma showing a biphasic pattern of chondroid and round and spindle cells
(hematoxylin-eosin, original magnification ⫻200).
Figure 25. High-power view showing malignant cartilage and primitive round cell components (hematoxylin-eosin, original magnification ⫻400).
Figure 26. Immunohistochemical stain for CD99 showing positivity in tumor cells (original magnification ⫻400).
Figure 27. Photomicrograph of small cell osteosarcoma showing small spindle cells with a diffuse growth pattern (hematoxylin-eosin, original
magnification ⫻400).

itive in small cell OS.129 Ewing family tumor–specific SUMMARY

translocations such as t(11;22) have not been reported in
small cell OS. Lymphomas can be distinguished using
lymphoma-specific markers.130 The treatment of small cell
OS is similar to conventional OS, which includes neoad-
juvant chemotherapy followed by surgery, usually a limb
salvage procedure. According to some authors, when there
is questionable matrix formation in a small round cell tu-
mor, it is best to err on the diagnosis of ES rather than
small cell OS.125
OTHER TUMORS
Rhabdomyosarcoma and desmoplastic round cell tu-
mors are primarily soft tissue neoplasms; rare primary
bone tumors are cited in the literature as case reports.131,132
Neuroblastomas can present as metastases in the bone and
can be distinguished from other primary round cell tu-
mors of bone with immunohistochemical positivity for
neuron-specific enolase, synaptophysin, and chromogran-
in and absent staining for CD99,133 lymphoid markers,
desmin, myogenin, and MyoD1.
Small round cell tumors of bone are a heterogeneous
group of neoplasms with overlapping clinical, radiologic,
morphologic, and immunohistochemical features. Even
though questions remain regarding cell of origin and his-
togenesis, cytogenetic and molecular genetic studies have
enhanced our understanding in bringing together some of
these tumors into single entities. In addition, cDNA mi-
croarray analysis and the availability of techniques to un-
ravel the various signaling pathways will open many
doors toward targeted chemotherapy in these aggressive,
highly lethal tumors.
The author thanks Seena Aisner, MD, Department of Pathology,
UMDNJ-Newark, for critical review of the manuscript.
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Small Round Cell Tumors of Bone—Hameed