Abstract 1409: The BP1 homeobox gene is dysregulated in triple negative breast cancer

Background: Triple negative breast cancers (TNBC) lack the expression of ER, PR and HER2 receptors, making targeted treatments against these receptors ineffective. We are studying BETA PROTEIN 1 (BP1), a member of homeobox gene family, which we cloned and showed is activated in 80% of invasive breast cancers; its expression correlates with breast cancer progression and invasion. The BP1 gene is located on 17q21, a region that we showed is altered in 30% of breast tumors. BP1 protein has been shown to regulate genes including BCL2, BRCA1, VEGF and c-MYC. Also BP1 mRNA and protein are highly expressed in ER and PR negative and clinically aggressive tumors. Thus we hypothesize that BP1 may play a role in TNBC. Methods: To delineate pathways in TNBC cells that are dysregulated by BP1 expression, we studied the 5 TNBC parental cell lines, 2 non-TNBC cell lines and a stably overexpressed BP1 TNBC cell line. Microarray analysis was performed in all these cell lines and the data obtained was verified using qPCR and ELISA. A BP1 shRNA plasmid was stably transfected into MDA-MB-231 cell lines to generate sh-MDA-MB-231 cell lines with about 80% knockdown of BP1 protein. ELISA was performed to verify the effects of BP1 expression on target genes. Formalin-Fixed, Paraffin-Embedded (FFPE) tissues of TNBC were analyzed by array-CGH and confirmed by Taqman copy number (using a BP1-TAM specific probe) and FISH (using our constructed BP1-BAC probe) assays. Results: All of the TNBC cell lines had statistically more BP1 protein (pBP1) than non-TNBC cell lines, suggesting that TNBC tumors may have higher levels of pBP1 than non-TNBC tumors. Gene expression microarrays in the TNBC cell lines revealed up/down regulation of several genes in comparison to the normal breast cell lines, including the IL-6 and IL-8 genes, which were significantly upregulated in TNBC cells overexpressing BP1. ELISA assays confirmed a direct correlation between elevated levels of IL-6 in conditioned media and overexpression of BP1. Knockdown of BP1 led to a decrease in secreted IL-6. Studies are underway to establish the mechanism by which these pathways are regulated by BP1. Increased BP1 copy number was observed in 37% (25/67) of the clinical cases and all (8/8) of TNBC cell lines by array-CGH. Confirmation of these changes by FISH and TaqMan copy number assays revealed increased copy number in 22% (18/82) and 23% (9/38) of the clinical cases, respectively. Protein expression analysis by IHC is curre