Abstract LB-124: RON (MST1R) is a novel prognostic marker and therapeutic target for gastroesophageal adenocarcinoma

Abstract

Abstract Background: RON (MST1R) is a member of the MET receptor tyrosine kinase family with a putative role in cancer, but its expression and function is poorly characterized in gastroesophageal cancer (GEC). Methods: We used immunohistochemistry (IHC), immunoblotting (IB) and immunofluorescence (IF) to determine RON and MET expression in human GEC tissues obtained from curative intent resection (n=94) and cell lines (n=20), and correlated tissue expression levels to overall survival using REMARK standards for prognostic markers. We assessed MST1R and MET gene copy number using quantitative polymerase chain reaction (qPCR) and confirmed findings using fluorescence in situ hybridization (FISH) and array comparative genomic hybridization in select samples. We assessed tumor tissue samples and GEC cell lines for MST1R mutation and single nucleotide polymorphisms (SNPs). We assessed the function of RON in GEC cell lines using viability, apoptosis, and invasion assays, with further evaluation using pooled small interfering RNA (siRNA) protein knockdown. The role of RON and MET coexpression and costimulation was evaluated along with downstream pathway activation of STAT3, using global and specific phophoantibodies, in order to evaluate potential synergistic signaling effects. GEC growth inhibition was evaluated using RON specific novel monoclonal blocking antibodies, small molecule tyrosine kinase inhibitors and RON siRNA/shRNA, alone or in combination with MET and STAT3 specific inhibition to evaluate for inhibitory synergism as a result of RON and MET functional reciprocity and signaling redundancy. Results: By IHC, RON was highly over-expressed in 74% of GEC samples and over-expression was prognostic of poor survival (p=0.008); MET and RON co-expression occurred in 43% of samples and was prognostic of worst survival (p=0.03). High MST1R gene copy number (GCN) was seen in 35.5% (16/45) of cases, correlated with poor survival (p=0.01), and was associated with high MET and ERBB2 GCN. A novel somatic MST1R juxtamembrane mutation R1018G was found in 11% of samples. RON was functional in cell lines, activating downstream effector STAT3, and resulting in increased viability over controls. RON and MET co-stimulation led to enhanced malignant phenotypes over either alone. Growth inhibition and apotosis was optimal using novel blocking monoclonal antibodies to both RON and MET, versus either alone. Small molecule tyrosine kinase inhibitor, SU11274, inhibited both receptors, and proved synergistic when combined with STAT3 inhibition (combination index <1). Conclusions: These preclinical studies define RON as a potential important novel prognostic marker and therapeutic target for GEC, and supports continued investigation of its role in this disease. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-124. doi:10.1158/1538-7445.AM2011-LB-124