Identification of chemosensitivity nodes for vinblastine through small interfering RNA high-throughput screens

Abstract

Discovering chemosensitivity pathways or nodes is an attractive strategy for formulating new drug combinations for cancer. Microtubules are among the most successful anticancer drug targets. Therefore, we implemented a small interfering RNA (siRNA) synthetic lethal screen targeting 5520 unique druggable genes to identify novel chemosensitivity nodes for vinblastine, a microtubule- destabilizing agent used clinically. We transiently transfected human glioblastoma cells with siRNAs for 48 h and then treated cells with a sublethal concentration of vinblastine. Forty-eight hours later, we analyzed cell viability and, using a series of statistical methods, identified 65 gene products that, when suppressed, sensitized glioblastoma cells to vinblastine. After completion of the secondary assays, we focused on one siRNA, B-cell lymphoma extra large (BCL-xL), because of its role in the intrinsic apoptosis signaling pathway as well as the availability of pharmacological inhibitors. We found that nontoxic concentrations of 4-[4-[[2-(4-chlorophenyl)-5,5- dimethylcyclohexen-1-yl]methyl]piperazin-1-yl]-N-[4-[[(2R)-4-morpholin-4-yl-1- phenylsulfanylbutan-2-yl] amino]-3-(trifluoromethylsulfonyl)phenyl] sulfonylbenzamide (ABT-263), an inhibitor of the BCL-2 family members (BCL-2, BCL-xL, and BCL-w), sensitized glioblastoma and non-small-cell lung cancer cells to vinblastine and induced apoptosis through the intrinsic cell death pathway. These results illustrate the usefulness of unbiased siRNA screens as a method for identifying potential novel anticancer therapeutic combinations.