Expression of EGFR, ERRB2, and KIT in Adult Soft Tissue Sarcomas: A Clinicopathological Study of 281 Cases Takuro Wada, O Sato, A Kawai, T Hasegawa Sapporo Medial University, Sapporo Japan National Cancer Center, Tokyo Japan

Purpose To analyze expression patterns of EGFR, ERBB2, and KIT on 281 patients with common adult STS by standard immunohistochemical technique on formalin-fixed, paraffin-embedded sections To Verify whether a significant association with prognosis is present

Patients 281 adult patients with common soft tissue sarcomas in the extremities and trunk diagnosed and treated at the NCC, Tokyo, Japan Histologic subtype Fibrosarcoma (FS) 11 Leiomyosarcoma (LMS) 11 Myxoid liposarcoma （ LSM) 52 Well differentiated liposarcoma （ LSW) 50 MPNST 18 Myxofibrosarcoma (MFS) 53 Pleomorphic MFH (PMFH) 44 Synovial sarcoma (SS) 42

Exclusion Tumor histology (379 )  Rhabdomyosarcoma, Ewing sarcoma  ASPS, angiosarcoma, epithelioid sarcoma,….. Location (42)  Retroperitoneal, head and neck, visceral, intrathoracic Stage IV (43)

Characteristic of 281 patients Gender:152 females and 129 males Age:10 to 85 years (median 50 yrs) Treatment:  ResectionAll patients  Radiotherapy25  Chemotherapy37  Chemo + Radi23

Pathology reviewing, grade and Staging Tumor typing  WHO Classification of Soft Tissue Tumors Histologic grade  Differentiation, necrosis, MIB-1 index TNM staging classification

Primary Antibodies AntibodySourceDilutionPositive control EGFR, mouse monoclonal DakoCytomation, Clone 2-18C9 PredilutedBreast ca ERBB2, rabbit polyclonal DakoCytomation1:500Breast ca KIT, rabbit polyclonal DakoCytomation1:50Mast cell

０ １＋ or ≤ 10% = Negative ２＋ ３＋ and >10% = Positive StrengthDistribution Evaluation of staining

Results

Immunohistochemical Pattern Histologic typeNo.EGFR Fibrosarcoma114 (36%) Leiomyosarcoma118 (73%) Myxoid liposarcoma523 (6%) Well-D liposarcoma5019 (38%) Myxofibrosarcoma5316 (89%) MPNST1847 (89%) PMFH4439 (89%) Synovial sarcoma4232 (76%) Total (60%)

Immunohistochemical Pattern Histologic typeNo.EGFRERBB2KIT Fibrosarcoma114 (36%)00 Leiomyosarcoma118 (73%)00 Myxoid liposarcoma523 (6%)00 Well-D liposarcoma5019 (38%)00 Myxofibrosarcoma5316 (89%)00 MPNST1847 (89%)00 PMFH4439 (89%)00 Synovial sarcoma4232 (76%)32 Total (60%)32

Diffuse EGFR (2+) in leiomyosarcoma Diffuse EGFR (3+) in MPNST

Diffuse EGFR (3+) in spindle cells of biphasic synovial sarcoma Focal ERBB2 (2+) in epithelioid cells of biphasic synovial sarcoma

Focal KIT (2+) in spindle cells of monophasic synovial sarcoma

EGFR (+) EGFR (-) 79 ％ 64 ％

Variables associated with patient survival: Univariate analysis Stage Histological grade Depth Size Positive EGFR staining

Variables associated with patient survival: Multivariate analysis Stage Histological grade Depth Size Positive EGFR staining

Kaplan-Meier survival curves stratified by EGFR status and by tumor size Size 5-10cm (n=135) Size>10cm (n=77) EGFR- EGFR+ p=0.03 p=0.06

Kaplan-Meier survival curves stratified by EGFR status and by histological grade Grade 2 (n=74) Grade 3 (n=111) EGFR- EGFR+ p=0.08 p=0.7

Discussion

Expression of EGFR in STSs AuthorsCasesPositiveSubtype Gusterson et al. (1985)3518 (51%)MFH, SS, Epi Perosio et al. (1989)4020 (50%) Duda et al. (1993)4321(49%) Barbashina (2002)1913 (68%)SS Nielsen (2002)4218 (52%)SS This study (60%) MFH, SS, MPNST

Expression of ERBB2 and KIT Merimsky et al. (2002)  No overexpression of ERBB2 in 230 STSs Hornick et al. (2002)  Very limited expression of KIT in 365 STSs This study  Limited expression of ERBB2 and KIT

EGFR (+) EGFR (-) 79 ％ 64 ％

IMC-C225 ABX-EGF Tyrosine kinase inhibitor Irresa Target-specific agents and approaches for anticancer therapy

Conclusion We analyzed the expression of EGFR, ERBB2, and KIT in 281 patients with STS Positive staining of EGFR: 168/281 (60%) Positive staining of ERBB2, KIT was limited EGFR overexpression was a negative prognostic factor associated with histological grade

Immunohistochemical Pattern

Use of Immunohistochemistry (IHC) to Detect EGFR Expression in Tumors Detects expression of EGFR protein on surface of tumor cells, with direct visualization of the target Reagents and equipment commonly available Widely used in many clinical studies Standardized methods are being developed

Risk of Invasion/ Tumor TypePrognosisSurvivalMetastasesReferences ColorectalPoorMayer (1993) Poor Increased Hemming (1992) Head and NeckDecreased DFSGrandis (1998) PoorDecreased OSMaurizi (1996) PancreaticPoorDong (1998) Decreased OSYamanaka (1993) NSCLCDecreased OSVolm (1998) Decreased OSVeale (1993) PoorDecreased OSOhsaki (2000) IncreasedPavelic (1993) Abbreviations: DFS, disease-free survival; OS, overall survival; NSCLC, non-small cell lung cancer. EGFR Expression Correlated With:

Summary EGFR overexpression was associated with poor prognosis in soft tissue sarcomas. Our study demonstrated frequent overexpression of EGFR in adult-type soft tissue sarcomas, and associated with histologic grade. We also confirmed very limited expression of KIT and HER2 in these tumors.

Target-specific agents and approaches for anticancer therapy