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Identification of novel chemosensitivity nodes using siRNA synthetic lethal screens
Author(s) -
Kitchens Carolyn A,
McDonald Peter R,
Thaker Nikhil G,
Shun Tong Ying,
Pollack Ian F,
Lazo John S
Publication year - 2010
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.24.1_supplement.964.11
Subject(s) - viability assay , small interfering rna , resazurin , transfection , cell , chemistry , biology , pharmacology , cancer research , microbiology and biotechnology , gene , biochemistry
Understanding the different modes of action of chemotherapeutic agents and how they synergize may lead to improved dosing regimens and novel combination treatments with molecularly targeted drugs. To identify chemosensitivity nodes for anticancer drugs, we implemented a short interfering RNA (siRNA) screen targeting 5,520 unique druggable genes in T98G human glioblastoma cells. In this screen we transiently transfected cells with targeting siRNAs for 48 hours, then treated with a sub‐lethal concentration of vinblastine (VBL), a specific anticancer agent, for an additional 48 hours. We analyzed cell viability using a resazurin fluorescent dye assay and identified gene products that sensitized T98G cells to VBL using two orthogonal statistical methods. The Median of the Absolute Deviations (MAD) method detected outliers using a MAD score threshold of ±3.5. Samples were ranked according to their viability ratio (cell viability with VBL and siRNA to cell viability with siRNA only) and the top 2.5 percent were selected as hits. The cell viability of samples treated with VBL and siRNA versus siRNA alone were compared using a two‐sample t ‐test and samples with a p ≤ 0.01 were considered to be statistically different and therefore synergistic VBL candidates. These analysis methods have identified 65 gene products that sensitize cells to VBL, which are currently being investigated further.