Transcript Comparison of genetic, antigenic and clinical features of extra-

Comparison of genetic, antigenic
and clinical features of extraosseous Ewing Sarcoma (EOEWS) and osseous Ewing sarcoma
(O-EWS)
Amanda Rivera-Begeman; Carrye Cost;
Stephen Lessnick; Richard Smith; Charles Timmons; Patrick
Leavey
Annual CTOS Meeting; Seattle, Washington, November 2007
Background

Ewing's sarcoma family of tumors (EFT)

Osseous EWS (O-EWS), Extraosseous ES
(EO-EWS), Peripheral primitive
neuroectodermal tumor (pPNET) and
Askin's tumor of chest wall
(Carvajal,R. et al; Hematol Oncol Clin North Am 2005)

Cell surface protein MIC-2 (CD99)
expressed on both O-EWS and EO-EWS
(Ambros, IM; Cancer; 1991)

FLI-1 nuclear immunostain + ve in 70% of
EWS and PNET cases (Folpe et al; Am J Surg Pathol;
2000)
Background
EWS/FLI-1 is seen in patients with
EFT (O’Sullivan,MJ et al; Hum Pathol; 2001)
 Prior reports of treatment for EOEWS demonstrated no advantage to
the addition of Doxorubicin (Raney RB et al.; J

Clin Oncol; 1997)

EO-EWS should be treated with
strategies used for O-EWS vs.
malignant mesenchymal tumors
(Castex,M.P.; J Clin Oncol; 2007)
Objective
To describe genetic, antigenic, and
clinical features of patients with EOEWS, primarily those of intraabdominal origin
 To compare genetic and antigenic
features of EO-EWS to those of
randomly chosen patients with OEWS

Patients

Eligibility criteria
EFT treated at Children’s Medical
Center Dallas (1995 – 2005; n=52)
 Availability of archival diagnostic
material and clinical data

Patients
Clinical Characteristic
EO-EWS
(n=11)
O-EWS
(n=11)
Abdominal: n=9
Pelvic: n=5
Brain: n=1
Long Bone: n=5
Nasopharynx: n=1
Rib: n=1
1035 cm3
494 cm3
Metastases at diagnosis
2
5
Median age at diagnosis
14.9
11.4
Radiation
5
7
Surgery
8
3
Died
2
5
Primary site
Mean tumor volume
Methods
AE1/AE3
CAM5.2
CK8

Cytokeratin
CK7
CEA
Carcino-embryonic antigen
Vimentin
Mesenchymal
FLI-1
Nuclear stain
CD99
MIC2
CD56
Neuro-ectodermal
Synaptophysin
NSE

Neuronal
Chromogranin
Myogenin
Desmin

All immunostains
were independently
reviewed by 2
pathologists (CT,
ARB)
Interpretation was
subjective
+ve vs. –ve
Muscle
No grading was
attempted
Fli-1
weak positive
Fli-1
strong positive
CD99 (O13)
Weak positive
CD99 (O13)
strong positive
Results – positive staining
Immunohistochemistry
EO-EWS
O-EWS
CK AE1/AE3
1
1
CAM5.2
1
0
CK7
0
0
CK8
0
1
CEA
2
2
Vimentin
8
8
FLI-1
9
9
CD99
11
11
Leu-7/CD56
4
8
Synaptophysin
4
4
NSE
11
10
Chromogranin
0
0
Myogenin
0
0
Desmin
0
0
Results – positive staining
Immunohistochemistry
EO-EWS
O-EWS
CK AE1/AE3
1
1
CAM5.2
1
0
CK7
0
0
CK8
0
1
CEA
2
2
Vimentin
8
8
FLI-1
9
9
CD99
11
11
Leu-7/CD56
4
8
Synaptophysin
4
4
NSE
11
10
Chromogranin
0
0
Myogenin
0
0
Desmin
0
0
Cytokeratin Cam 5.2
positive
Carcinoembryonic antigen (CEA)
positive
Results – positive staining
Immunohistochemistry
EO-EWS
O-EWS
CK AE1/AE3
1
1
CAM5.2
1
0
CK7
0
0
CK8
0
1
CEA
2
2
Vimentin
8
8
FLI-1
9
9
CD99
11
11
Leu-7/CD56
4
8
Synaptophysin
4
4
NSE
11
10
Chromogranin
0
0
Myogenin
0
0
Desmin
0
0
Results – positive staining
Immunohistochemistry
EO-EWS
O-EWS
CK AE1/AE3
1
1
CAM5.2
1
0
CK7
0
0
CK8
0
1
CEA
2
2
Vimentin
8
8
FLI-1
9
9
CD99
11
11
Leu-7/CD56
4
8
Synaptophysin
4
4
NSE
11
10
Chromogranin
0
0
Myogenin
0
0
Desmin
0
0
Results – positive staining
Immunohistochemistry
EO-EWS
O-EWS
CK AE1/AE3
1
1
CAM5.2
1
0
CK7
0
0
CK8
0
1
CEA
2
2
Vimentin
8
8
FLI-1
9
9
CD99
11
11
Leu-7/CD56
4
8
Synaptophysin
4
4
NSE
11
10
Chromogranin
0
0
Myogenin
0
0
Desmin
0
0
Methods

RT-PCR


RNA extracted from formalin fixed, paraffin
embedded tumor (Roche high pure RNA
paraffin kit)
PCR was performed for EWS/FLI-1
• EWS/FLI-1 fusion type identified by melt curve
analysis
• EWS-FLI-1 fusion type confirmed by agarose gel
electrophoresis

Alternate partners were examined as
necessary (ews/fev, etv1, etv4 and erg)
Results – EWS-FLI-1 type
EO- OEWS EWS
EWS-FLI-1
10
10
Type 1
7
10
Type 2
3
0
Non Type 1
or 2
1
1

Not type 1 or 2


EO-EWS: confirmed
t(11:22)(q24;q12) but
no translocation
products amplified
O-EWS: PCR
product melting
curve between types
1 and 2 but failed
sequencing
Summary
Negative FLI-1 nuclear staining does
not exclude EWS-FLI-1 translocation
positive EFT
 While CEA +ve staining can be seen in
EO-EWS it does not differentiate this
from O-EWS
 More type 2 fusions noted in patients
with EO-EWS

Conclusion

Variability may occur in
immunostaining and genotype
analysis of patients with extra-osseous
Ewing sarcoma vs. osseous Ewing
sarcoma.