Abstract 5757: A new functional classification of lung and breast cancers based on siRNA mediated knockdown of 48 NRs and 72 CoRegulators
Authors: Ryan Carstens, Kenneth Huffman, Luc Girard, Bruce Posner, Heidi Erickson, Ignacio Wistuba, Adi Gazdar, Steven Kliewer, Suzanne Fuqua, Bert O'Malley, David Mangelsdorf, John D. Minna
Journal: Cancer Research (2012)

Abstract Introduction: Nuclear receptors (NRs) and their co-regulators (CoRegs) are master regulators of key transcriptional processes important in cancer pathogenesis and could provide molecular biomarkers for prognosis and personalizing new cancer targeted therapy. Aims and Methods: Our goal is to functionally classify a panel of 54 lung cancer, 20 breast cancer, and several immortalized human lung and breast epithelial cell strains based on their growth and survival response to siRNA mediated knockdown of 48 NRs and 72 CoRegs. Following knockdown of individual NR/CoRegs, changes in growth patterns are quantified and statistical methods are used to identify disease subgroups of similar responders. These subgroups are then integrated with our legacy datasets for clinical demographics, cancer cell oncogenotype and genome wide molecular characterization, and responses to standard and targeted chemotherapy. Results and Conclusions: A diverse, representative panel consisting of lung cancer, breast cancer, and normal epithelial cell lines was tested by siRNA transfection. After optimization to determine transfection conditions, all cell lines were screened using a focused siRNA library targeting all 48 NRs and 72 CoRegs (each gene is targeted by pools of 4 different siRNAs, Qiagen). Screening experiments were performed in triplicate, in 96 well plates, in a five-day MTS assay and assays were repeated 3 times. Correlation between replicates for the same cell line is &amp;gt;0.84 and knockdowns were validated by qRT-PCR. We found: 1) NR/CoReg-specific knockdown could have no effect, increase, or decrease tumor cell viability in a cell line dependent manner. 2) Overall ∼70% of the NRs or CoRegs reduced cell viability in at least one tumor line when their expression was knocked down (examples included NURR1, PRMT1, RXRalpha, BRCA1). 3) NR/CoReg-specific tumor cell toxicity was validated independently by siRNA knockdown combined with a liquid colony formation assay. 4) Knockdown of several NR/CoRegs selectively killed tumor but not normal epithelial cells. 5) Tumor cell lines could be stratified into functionally unique categories based on their changes in growth following NR/CoReg-specific knockdowns. We conclude that siRNA mediated knockdown of a large number of NRs and their CoRegs identifies genes that show tumor cell selective toxicity, and that tumor lines vary in their siRNA response phenotypes. These data strengthen the concept for NR/CoReg targeted therapy of lung and breast cancer and also the need to “personalize” such therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5757. doi:1538-7445.AM2012-5757