43 Calcium channel blockers for treatment of triple-negative breast cancer

Abstract

Objectives/Goals: Triple-negative breast cancer (TNBC) is a highly aggressive form of breast cancer (BC) with limited treatment options. Mortality rate is especially high in African American (AA) women of reproductive age. High levels of intracellular calcium (Ca2+) have been shown in TNBC cells. This study is to investigate Ca2+ channel blockers (CCBs) as therapy for TNBC. Methods/Study Population: Two human TNBC cell lines obtained from ATCC – HCC1806, and MDA-MB-453 are treated with CCBs, Cilnidipine (Cil), and Mibefradil (Mib), in a concentration- and time-dependent manner. Cell proliferation assays are performed by the MTS cell viability assay. Intracellular Ca2+ levels are measured using the fluorescent dye: Fluro 4-AM. Apoptosis is determined by flow cytometry using Annexin V staining and mitochondrial permeability will be assessed by the Mito JC-1 assay. Expression of Ca2+ signaling genes will be quantitated by real-time polymerase chain reaction (RT-PCR). Potential pathways of CCB efficacy will be identified by ingenuity pathway analysis (IPA). Results/Anticipated Results: Our findings show both CCBs decrease cell proliferation in a concentration- and time-dependent manner to a maximum of 80% vs. control in both TNBC cells. Flow cytometry findings on both TNBC cells treated with both drugs at 20 µM for 24 hours depicts late apoptosis. Interestingly, Mib did not change the intracellular Ca2+ level in HCC1806 cells yet decreased in MDA-MB-453 cells by fivefold, while Cil increased the intracellular Ca2+ level in both cells almost twofold. It is anticipated that Mito JC-1 assay depict decreased mitochondrial potential in both cells. For reverse transcription polymerase chain reaction, it is anticipated that CCB treatment will increase transient receptor potential Ca2+ channels and decrease voltage-gated Ca2+ channels in both cells. IPA analysis is expected to show apoptotic pathways are involved in TNBC via CCB treatment. Discussion/Significance of Impact: TNBC lacks the estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2. Treatment options for TNBC remain severely limited. Our findings that both Cil and Mib can inhibit proliferation of human breast cancer cell lines indicate repurposing CCBs as treatment for TNBC warrants further investigation.