Abstract A2-13: Targeting genomic instability to identify molecular drivers of poor prognosis in cancer.

Abstract

Abstract Background: Cancer is a disease of the genome whereby mutational events that confer a survival advantage to cells are selectively retained. Consequently, genome instability and evasion of cell death are fundamental hallmarks of cancer. Certain cancers, including epithelial ovarian cancers (EOC), pancreatic ductal adenocarcinomas (PDAC) and a subset of sarcomas with complex karyotypes are characterized as being very genomically unstable suggesting that such tumors are driven by defects in DNA damage recognition and repair. Additionally, these tumors are also characterized by a high frequency of p53 mutation, extensive intra-tumoral heterogeneity and resistance, acquired or intrinsic, to DNA damaging chemotherapeutic agents: these tumor types consequently carry a poor prognosis. This highlights the need for detailed molecular characterization to identify new therapeutic targets and stratification biomarkers. Aims: The aim of the study is to use a bioinformatic approach to identify commonly amplified genes, that function in DNA damage response and apoptotic processes, across the three tumor types, and which confer a poorer prognosis in these patients. Methods: For target identification, datasets for EOC and PDAC (obtained from TCGA) and sarcoma (obtained from GEO, NCBI) were analyzed for common copy number aberrations (CNA) in 734 genes relating to DNA damage response and apoptosis, and confer a poor prognosis in progression-free survival (PFS) data in EOC patients. Genes with prognostic significance (p <0.05) using the log rank test were picked from the top 100 most frequently amplified loci. Next, the targets were functionally validated by siRNA-mediated knockdown, overexpression and/or pharmacological inhibition using apoptosis, proliferation and migration assays. Results: Four genes, SGK3, c19orf40, MRPS12 and ZBTB32 were highlighted as being commonly amplified across all three tumor types, of which only SGK3 and c19orf40 were statistically significant in circular binary segmentation CNA calling when examining PFS data in EOC patients. SGK3, a member of the serum/glucocorticoid regulated kinase (SGK) family appeared interesting as it has similar functions and substrates to the AKT kinase family, which we have previously shown to have key roles in tumor cell survival in response to therapy. Intra-patient paired platinum sensitive (PEA1) and resistant (PEA2) and SKOV3 ovarian cancer cell lines, and Aspc1 and Panc-1 pancreatic cancer cell lines were used for validation studies. siRNA-mediated knockdown of SGK3 did not alter induction of caspase 3/7 activity in response to chemotherapy, relative to control treatments in all cell lines. To account for any compensatory effects by the other SGK members in the presence of SGK3 knockdown, all three SGK members (SGK1-3) were knocked down by siRNA and this also did not increase induction of caspase 3/7 activity. Wound healing migration assays however, revealed that over-expression of SGK3 increases cell motility suggesting SGK3's role in prognosis is via migration/tumor spread rather than response to therapy. The remaining targets identified in the bioinfomatic analysis are currently undergoing validation and preliminary results will be summarized. Conclusion: To conclude, bioinformatic analysis highlighted four genes related to DNA damage response/apoptosis that were commonly amplified across tumors characterized by genomic instability and resistance to chemotherapy. One of these genes, SGK3, appears to have a role in poor prognosis via migration. Citation Format: Karen A. Menezes, Paula Cunnea, Phillip Lawton, Ed Curry, Hani Gabra, Harpreet Wasan, Surinder K. Sharma, Euan A. Stronach. Targeting genomic instability to identify molecular drivers of poor prognosis in cancer.. [abstract]. In: Proceedings of the AACR Special Conference on Translation of the Cancer Genome; Feb 7-9, 2015; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(22 Suppl 1):Abstract nr A2-13.