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Breast cancer has become a worldwide threat, and chemotherapy remains a routine treatment. Patients are forced to receive continuous chemotherapy and suffer from severe side effects and poor prognosis. Natural alkaloids, such as piperine (PP) and piperlongumine (PL), are expected to become a new strategy against breast cancer due to their reliable anticancer potential. In the present study, cell viability, flow cytometry, and Western blot assays were performed to evaluate the suppression effect of PP and PL, alone or in combination. Data showed that PP and PL synergistically inhibited breast cancer cells proliferation at lower doses, while only weak killing effect was observed in normal breast cells, indicating a good selectivity. Furthermore, apoptosis and STAT3 signaling pathway-associated protein levels were analyzed. We demonstrated that PP and PL in combination inhibit STAT3 phosphorylation and regulate downstream molecules to induce apoptosis in breast cancer cells. Taken together, these results revealed that inactivation of STAT3 was a novel mechanism with treatment of PP and PL, suggesting that combination application of natural alkaloids may be a potential strategy for prevention and therapy of breast cancer.

Additional file 1: Fig. S1. A: DSPE-PEG2K-COOH MALDI-TOF REPORT; B: RGERPPR MALDI-TOF REPORT; C: DSPE-PEG2K-RGERPPR MALDI-TOF REPORT; D: R9dGR(RRRRRRRRR-dGR) MALDI-TOF REPORT; E: DSPE-PEG2K-R9dGR (RRRRRRRRR-dGR) MALDI-TOF REPORT. F: the visceral indexes of mice in different groups. G: Particle size of GA/PTX-NLC, RGE-GA/PTX-NLC and R9dGR-GA/PTX-NLC (by TEM images). H: R9dGR-GA/PTX-NLC. Fig. S2. The microscopy image of cellular uptake in MDA-MB-231 (A), 4T1 (B), and MCF-7 (C) cells after treatment with Cou-6-Sol, Cou-6-loaded targeted and nontargeted NLC (scale bar = 100 μm)