pHLIP(Var7)-P1AP suppresses tumor cell proliferation in MDA-MB-231 triple-negative breast cancer by targeting protease activated receptor 1

PURPOSE: Protease-activated receptor 1 (PAR1) is a signaling protein ubiquitously present on the surface of tumor cells, and its homologous protein fragment, PAR1-activating peptide (P1AP), can inhibit protein signal transduction of PAR1/G in tumor cells. pH (Low) insertion peptide (pHLIP) can target the acidic tumor microenvironment (TME) and can be used as an excellent carrier to deliver P1AP to tumor cells for therapeutic purposes. METHODS: PAR1 expression on the surface of MDA-MB-231 cells and human MCF10A mammary epithelial cells was observed. The binding between fluorescent-labeled pHLIP(Var7)-P1AP and MDA-MB-231 cells under different pH values was analyzed. The effect of pHLIP(Var7)-P1AP on the proliferation of MDA-MB-231 cells was analyzed under the conditions of pH 7.4 and 6.0. RESULTS: PAR1 was highly expressed on the surface of MDA-MB-231 cells. In an acidic environment (pH 6.0 and 5.0), fluorescent-labeled pHLIP(Var7)-P1AP and MDA-MB-231 cells had a high binding ability, and the binding ability increased with the decrease in pH. In an acidic environment (pH 6.0), pHLIP(Var7)-P1AP significantly inhibited MDA-MB-231 cell proliferation. With 0.5 μg, 1 μg, 2 μg, 4 μg, and 8 μg of pHLIP(Var7)-P1AP, the cell proliferation inhibition rates were 3.39%, 5.27%, 14.29%, 22.14%, and 35.69%, respectively. CONCLUSION: PAR1 was highly expressed on the surface of MDA-MB-231 cells. pHLIP(Var7)-P1AP can effectively target MDA-MB-231 cells in an acidic environment and inhibit the growth of MDA-MB-231 cells by inhibiting the signal transduction of PAR1/G protein.