Abstract 4434: Identification of radiation susceptibility genes in prostate cancer by whole genome RNA interference screening.

Abstract

Background: Prostate cancer (PCa) is the most commonly diagnosed cancer and the second leading cause of cancer death in Western world males. PSA testing allows early detection of localized disease. Treatment options for patients with local disease include radiotherapy and surgery. However, local tumor control rates after radiotherapy are highly variable; and ∼40% of treated patients will experience disease recurrence and progression. Advanced prostate cancer has a very poor prognosis, emphasizing the need for more effective early local treatment, preventing advanced disease. Here, we set out to identify molecular targets of radiation susceptibility in PCa cells as a starting-point to increase the therapeutic index of radiotherapy. Methods: Targets were identified by high-throughput screening using a human whole genome siRNA library on the radioresistant PCa cell line PC-3 subjected to 4Gy irradiation. Combined effects of radiation and target gene silencing were quantified using a cell viability assay based on automated cell counting four days after irradiation. Candidate targets were validated using the colony formation assay (CFA) scored 8-10 days after irradiation. Results: We conducted two genome-wide siRNA screens with and without irradiation in PC-3 cells. We used a linear regression model to identify radiation susceptibility genes and selected 45 candidate targets with FDR < 10% for further research. In a conformation screen, multiple independent siRNAs silencing 17 genes induced ≥2-fold more cell death upon 4Gy irradiation compared to a negative control siRNA. Confirmed hits were validated independently using the CFA. The dose modifying factor at 80% clonogenic cell survival (DMF0.8) was calculated for each gene-specific siRNA compared to control siRNA from linear-quadratic survival curves. Sixteen candidate genes were validated with a DMF0.8 ≥ 1.3. This included the mitotic spindle assembly checkpoint component MAD2L2, which was previously reported as a radiosensitizing target in PCa cells (JCI 2011;121:2383-2390), thus validating our screens. Importantly, silencing of 9 genes sensitized PC-3 cells to irradiation more effectively than did silencing of the known radiation susceptibility gene PRKDC. Among these were genes involved in cell cycle control, ubiquitination, cell communication and G-protein coupled receptor signaling. Interestingly, silencing of two druggable genes, i.e. CPNE7 and GPR27, was particularly effective at low dose irradiation, resulting in a very high DMF0.8 of 4.6 and spectacular decreases in surviving fractions after 2Gy irradiation from 70% in controls to 37% and 23%, respectively. Conclusion: We have identified a set of novel candidate targets that could be used to potently enhance the efficacy of radiotherapy in prostate cancer. This could contribute significantly to improving overall success of prostate cancer intervention. Citation Format: Jasmina Hodzic, Ida van der Meulen, Renee de Menezes, Ilse Dingjan, Marielle JP Maas, Winald Gerritsen, Marcel Verheij, Ab A. Geldof, Baukelien van Triest, Victor W. van Beusechem. Identification of radiation susceptibility genes in prostate cancer by whole genome RNA interference screening. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4434. doi:10.1158/1538-7445.AM2013-4434