Piperine in combination with TRAIL suppresses the cell growth in both TRAIL-sensitive and TRAIL-resistant human TNBC cells.
We screened 55 compounds derived from natural products with or without TRAIL on the cell growth in human triple-negative breast cancer (TNBC) cells, MDA-MB-231 and MDA-MB-468 cell lines, each of which are known as TRAIL-sensitive and TRAIL-resistant cells, respectively (6). After the screening, alisol A, curcumine, and piperine showed the synergistic suppression on the cell growth in both TNBC cells (Synergy Index<0.8, Table 1). Contrary to TRAIL, piperine alone could suppress the growth in MDA-MB-468 but not in MDA-MB-231; therefore, we decided to focus on the piperine in this study.
To confirm the screening results, both TNBC cells were treated various concentration of piperine with or without TRAIL (Fig. 2a). Consistent with the screening results, piperine suppressed the cell growth in MDA-MB-468 cells but not in MDA-MB-231 cells, and further piperine and TRAIL combination synergistically inhibited the cell growth in both cell lines compared with either treatment alone. We next determined whether or not the growth suppression of piperine and TRAIL combination is due to the induction of typical apoptotic pathway by using annexin-V and 7-AAD staining. Piperine with TRAIL significantly increased the apoptotic cells compared with either treatment alone in both cell lines (Fig. 2b). Similarly, the cleavage of caspase-3 and PARP, both of which are known as apoptotic markers, were clearly induced by piperine and TRAIL combination in both cell lines (Fig. 2c). Unlike the growth suppression, piperine treatment did not induce the apoptosis in MDA-MB-468 cells. In regards to the cell cycle status, piperine treatment lead MDA-MB-468 cells but not MDA-MB-231 cells to cell cycle arrest at G2/M phase (Fig. 2d). Collectively, the growth suppression of piperine and TRAIL combination was mediated by typical apoptotic pathway in both MDA-MB-468 and MDA-MB-231 cells while the cell-cycle arrest was specifically observed in MDA-MB-468 cells in piperine mono-therapy.
Piperine enhances the TRAIL-responsiveness through the suppression of survivin and p65-phosphorylation in TNBC cells.
Given piperine and TRAIL combination induced apoptosis not only in TRAIL-sensitive MDA-MB-231 cells but also in TRAIL-resistant MDA-MB-468 cells, we next investigated the expression levels of Mcl-1, survivin, XIAP, BCL-xL and p65-phospohorylation that are known to correlate with TRAIL-sensitivity in cancer cells (6-11). Amongst them, piperine suppressed the expression of survivin and p65-phosphorylation in both cell lines. In addition to human TNBC cells, piperine suppressed the survivin expression and the phosphorylation of p65 in murine 4T1 TNBC cells, which is known as TRAIL-sensitive (Fig. 3). These results suggest that piperine likely induce the TRAIL-sensitivity in both TRAIL-sensitive and TRAIL-resistant cells through the suppression of survivin expression...