Abstract 1409: The BP1 homeobox gene is dysregulated in triple negative breast cancer
Authors: Kirolikar Saurabh , Gill Mandeep , Pereira Silma , Ghinbouschi Svetlana , Cavalli Luciane , Berg Patricia 
Published: May 2015

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 currently being conducted to determine if BP1 increased DNA copy number may be one of the mechanisms leading to pBP1 overexpression.

Conclusions: Both BP1 copy number and protein expression are dysregulated in TNBC. Microarray analysis in TNBC cell lines have revealed important cancer related genes and oncogenic pathways that may be upregulated by BP1, suggesting that BP1 could be a valuable molecular marker and therapeutic target in this aggressive form of breast cancer.

Citation Format: Saurabh Kirolikar, Mandeep Gill, Silma Pereira, Svetlana Ghinbouschi, Luciane Cavalli, Patricia Berg. The BP1 homeobox gene is dysregulated in triple negative breast cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1409. doi:10.1158/1538-7445.AM2014-1409