Abstract 3160: Open-source RNAi genetic screen platform: discovery of drug targets in cancer cell models.

Abstract

We have developed an open-source RNAi genetic screening technology, funded in part by NIH grants and freely available to the scientific community. This platform can be used for functional discovery of drug targets associated with viability and regulation of signaling pathways in cancer cells. Here we present a novel, free public resource for genome-wide RNAi screens: DECIPHER pooled lentiviral shRNA libraries. Results are shown from two RNAi screens with DECIPHER shRNA libraries: one “drop-out” screen to identify genes essential for viability in a panel of leukemic cells, and a second “rescue” screen to identify genes required for FAS induced apoptosis. Both screens found a combination of known and novel signaling pathway and regulatory genes whose functions were confirmed to be required to produce the biological responses. From the viability screens in the panel of leukemia cell lines, subsequent validation using single shRNA-expressing constructs showed that about 80% of the shRNAs identified in each complex library screen did in fact lead to cell death when transduced in cells. Analysis of the identified essential genes for known biological interactions revealed non-random clusters of interacting proteins that provide a useful strategy for prioritization of potential targets. Analysis of the lethal combinations indicates redundant, complementary, and compensatory responses in cancer cells. In the case of the FAS-induced apoptosis, in vitro screening data also enabled us to select targets that protected mice from FAS-induced hepatic failure. These results demonstrate that complex pooled shRNA libraries provide a highly efficient, flexible, and cost-effective alternative to array-based RNAi screening methods for identifying genes regulating biological responses and possible new therapeutic targets. Citation Format: Andrei Komarov, Mikhail Makhanov, Elena Komarova, Lilya Novototzkaja, Michael Yeluashvili, Dmitry Suchkov, Kyle Bonneau, Donato Tedesco, Costa G. Frangou, Paul Diehl, Debbie Deng, Karim Hyder, Pavel Komarov, Andrei Gudkov, Alex Chenchik. Open-source RNAi genetic screen platform: discovery of drug targets in cancer cell models. [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 3160. doi:10.1158/1538-7445.AM2013-3160