Abstract 951: Loss-of-function screening identifies therapeutic targets for triple negative breast cancer (TNBC)
Authors: Bauer Joshua A. , Lehmann Brian D. , Glick Gloria , Cortez David , Lee Ethan , Pietenpol Jennifer A. 
Published: June 2012

Triple negative breast cancers (TNBCs) are characterized by the absence of estrogen/progesterone receptors and HER2 gene amplifications. Patients with TNBC have a poor prognosis and cannot be effectively treated with current targeted therapies. Our overall goal is to identify signaling targets and pathways involved in TNBC tumor cell viability and to use our findings to inform clinical trial design to target these pathways. To accomplish this we selected a subset of 220 genes spanning diverse signaling and growth pathways from a large panel of TNBC tumors (n=386). The genes were identified as being differentially overexpressed from an integrated analysis of gene expression and CGH array datasets or were selected because they contained relevant somatic oncogenic mutations. We performed a loss-of-function RNA interference (RNAi) screen on this gene set to identify genes/pathways that modulate TNBC cell viability. We screened 25 TNBC cell lines and scored (z-score) the effect that siRNAs (four per gene) had on cell viability. For potential therapeutic benefit, three independent primary human mammary epithelial cell (HMEC) cultures were screened to compare the effects of siRNAs on ‘normal’ breast cells. siRNAs to a kinesin gene KIF11, and two genes involved in kinetochore-microtubule attachment, NDC80 and NUF2, significantly decreased cell viability (&gt; 50%, z-scores -1.28). Other siRNAs from the screen that differentially targeted TNBC cells were genes involved in proliferation (BRAF, MAPK1, AKT3, KRAS), the chromosomal passenger complex (INCENP, AURKB), kinetochore assembly (CENPE, BUB3, BRD4), spindle-assembly checkpoint (AURKA, TPX2), centromere cohesion (ESPL1, RAD21), cytokinesis (KIF14, ANLN), apoptosis (BCL2L1, BIRC5) and Wnt/beta-catenin pathway (NDP, LRP5, DVL1, TCF7). In parallel, we tested therapeutic agents that targeted several of these pathways found by our RNAi screens. Oncogene-addicted cell lines containing KRAS mutations (MDA-MB-231 and SW527) and BRAF mutations (DU4475) were more sensitive to siRNAs targeting KRAS or BRAF (sum of z-score &lt; -7) and to the MEK inhibitor AZD6244 (IC50 1 µM). Likewise, cell lines that had the highest levels of BCL2L1 mRNA (CAL120, MDA-MB-231, HCC1143, HCC1937) were the most sensitive to siRNAs and to the BCL2 inhibitor ABT-737 (IC50 &lt; 3 µM). TNBC cell lines that have a mesenchymal-like phenotype (HS578T, BT549, MDA-MB-157, and CAL51) were more sensitive to targeting the Wnt/beta-catenin pathway by siRNAs, a neutralizing antibody to the LRP5 receptor, or to a compound that activates casein kinase I, a regulator of the Wnt/beta-catenin pathway. These data demonstrate that the combination of genomic information along with functional genetic RNAi screening can be a powerful approach to identify novel targets and personalize therapeutic strategies for TNBC patients.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 951. doi:10.1158/1538-7445.AM2011-951