Capsaicin, the pungent ingredient of red hot chili peepers, has been shown to have anti-cancer activities in several cancer cells, including prostate cancer. Several molecular mechanisms have been proposed on its chemopreventive action, including ceramide accumulation, endoplasmic reticulum stress induction and NFκB inhibition. However, the precise mechanisms by which capsaicin exerts its anti-proliferative effect in prostate cancer cells remain questionable. Herein, we have tested the involvement of autophagy on the capsaicin mechanism of action on prostate cancer LNCaP and PC-3 cells.The results showed that capsaicin induced prostate cancer cell death in a time- and concentration-dependent manner, increased the levels of microtubule-associated protein light chain 3-II (LC3-II, a marker of autophagy) and the accumulation of the cargo protein p62 suggesting an autophagy blockage. Moreover, confocal microscopy revealed that capsaicin treatment increased lysosomes which co-localized with LC3 positive vesicles in a similar extent to that produced by the lysosomal protease inhibitors E64 and pepstatin pointing to an autophagolysosomes breakdown inhibition. Furthermore, we found that capsaicin triggered ROS generation in cells, while the levels of ROS decreased with N-acetyl-cysteine (NAC), a ROS scavenger. Co-treatment of cells with NAC and capsaicin abrogated the effects of capsaicin on autophagy and cell death. Normal prostate PNT2 and RWPE-1 cells were more resistant to capsaicin-induced cytotoxicity and did not accumulate p62 protein.Taken together, these results suggest that ROS-mediated capsaicin-induced autophagy blockage contributes to antiproliferation in prostate cancer cells, which provides new insights into the anticancer molecular mechanism of capsaicin.

Background Current chemotherapy for castration-resistant prostate cancer is established on taxane-based compounds like docetaxel. However, eventually, the development of toxic side effects and resistance limits the therapeutic benefit being the major concern in the treatment of prostate cancer. Combination therapies in many cases, enhance drug efficacy and delay the appearance of undesired effects, representing an important option for the treatment of castration-resistant prostate cancer. In this study, we tested the efficacy of the combination of docetaxel and capsaicin, the pungent ingredient of hot chili peppers, on prostate cancer cells proliferation. Methods Prostate cancer LNCaP and PC3 cell lines were used in this study. Levels of total and phosphorylated forms of Akt, mTOR, S6, LKB1, AMPK and ACC were determined by Western blot. AMPK, LKB1 and Akt knock down was performed by siRNA. PTEN was overexpressed by transient transfection with plasmids. Xenograft prostate tumors were induced in nude mice and treatments (docetaxel and capsaicin) were administered intraperitoneally. Statistical analyses were performed with GraphPad software. Combination index was calculated with Compusyn software. Results Docetaxel and capsaicin synergistically inhibited the growth of LNCaP and PC3 cells, with a combination index lower than 1 for most of the combinations tested. Co-treatment with docetaxel and capsaicin notably decreased Akt and its downstream targets mTOR and S6 phosphorylation. Overexpression of PTEN phosphatase abrogated the synergistic antiproliferative effect of docetaxel and capsaicin. The combined treatment also increased the phosphorylation of AMP-activated kinase (AMPK) and the phosphorylation of its substrate ACC. In addition, pharmacological inhibition of AMPK with dorsomorphin (compound C) as well as knock down by siRNA of AMPK or its upstream kinase LKB1, abolished the synergy of docetaxel and capsaicin. Mechanistically, we showed that the synergistic anti-proliferative effect may be attributed to two independent effects: Inhibition of the PI3K/Akt/mTOR signaling pathway by one side, and AMPK activation by the other. In vivo experiments confirmed the synergistic effects of docetaxel and capsaicin in reducing the tumor growth of PC3 cells. Conclusion Combination of docetaxel and capsaicin represents a therapeutically relevant approach for the treatment of Prostate Cancer. Electronic supplementary material The online version of this article (10.1186/s12935-019-0769-2) contains supplementary material, which is available to authorized users.

BACKGROUND:Neuroendocrine (NE) differentiation represents a common feature of prostate cancer and is associated with accelerated disease progression and poor clinical outcome. Nowadays, there is no treatment for this aggressive form of prostate cancer. The aim of this study was to determine the influence of the cannabinoid WIN 55,212-2 (WIN, a non-selective cannabinoid CB1 and CB2 receptor agonist) on the NE differentiation of prostate cancer cells.METHODS:NE differentiation of prostate cancer LNCaP cells was induced by serum deprivation or by incubation with interleukin-6, for 6 days. Levels of NE markers and signaling proteins were determined by western blotting. Levels of cannabinoid receptors were determined by quantitative PCR. The involvement of signaling cascades was investigated by pharmacological inhibition and small interfering RNA.RESULTS:The differentiated LNCaP cells exhibited neurite outgrowth, and increased the expression of the typical NE markers neuron-specific enolase and βIII tubulin (βIII Tub). Treatment with 3 μM WIN inhibited NK differentiation of LNCaP cells. The cannabinoid WIN downregulated the PI3K/Akt/mTOR signaling pathway, resulting in NE differentiation inhibition. In addition, an activation of AMP-activated protein kinase (AMPK) was observed in WIN-treated cells, which correlated with a decrease in the NE markers expression. Our results also show that during NE differentiation the expression of cannabinoid receptors CB1 and CB2 dramatically decreases.CONCLUSIONS:Taken together, we demonstrate that PI3K/Akt/AMPK might be an important axis modulating NE differentiation of prostate cancer that is blocked by the cannabinoid WIN, pointing to a therapeutic potential of cannabinoids against NE prostate cancer.

In this study, we investigated the antitumoral effects of combined treatment using sorafenib and capsaicin in hepatocellular carcinoma (HCC) cells. Here we showed that the combination of the two drugs had a much stronger inhibitory effect on both HepG2 and Huh-7 human HCC cells growth than either drug alone. The isobolograms demonstrated that the combinations investigated in this study produced a synergistic interaction. In the combination treatment using capsaicin and sorafenib, increased apoptosis, followed by the activation of caspase-9 and PARP, was observed. In addition, the present study demonstrated that sorafenib treatment induces activation of Akt, probably as a mechanism of resistance, whereas capsaicin inhibits Akt providing a possible pathway whereby capsaicin sensitizes to sorafenib in HCC cells. Moreover, capsaicin singly and the combination of capsaicin and sorafenib induce AMPK activation and Acetyl CoA carboxylase phosphorylation in HCC cells. Knocking down of AMPK by selective siRNA abrogates capsaicin-induced Akt inhibition, suggesting the involvement of AMPK in the antiproliferative effect. In vivo experiments further showed that that the anti-tumor effect of sorafenib was enhanced by its combination with 2.5 mg/Kg of capsaicin. Overall, these results show that combined treatment with capsaicin and sorafenib might improve sorafenib sensitivity and therefore it represents a promising and attractive strategy for the treatment of HCC.

Neuroendocrine (NE) prostate cancer (PCa) is a highly aggressive subtype of prostate cancer associated with resistance to androgen ablation therapy. In this study, we used LNCaP prostate cancer cells cultured in a serum-free medium for 6 days as a NE model of prostate cancer. Serum deprivation increased the expression of NE markers such as neuron-specific enolase (NSE) and βIII tubulin (βIII tub) and decreased the expression of the androgen receptor protein in LNCaP cells. Using cDNA microarrays, we compared gene expression profiles of NE cells and non-differentiated LNCaP cells. We identified up-regulation of 155 genes, among them LAMP2, a lysosomal membrane protein involved in lysosomal stability and autophagy. We then confirmed up-regulation of LAMP2 in NE cells by qRT-PCR, Western blot and confocal microscopy assays, showing that mRNA up-regulation correlated with increased levels of LAMP2 protein. Subsequently, we determined autophagy activity in NE cells by assessing the protein levels of SQSTM/p62 and LC3 by Western blot and LC3 and Atg5 mRNAs content by qRT-PCR. The decreased levels of SQSTM/p62 was accompanied by an enhanced expression of LC3 and ATG5, suggesting activation of autophagy in NE cells. Blockage of autophagy with 1μM AKT inhibitor IV, or by silencing Beclin 1 and Atg5, prevented NE cell differentiation, as revealed by decreased levels of the NE markers. In addition, AKT inhibitor IV as well as Beclin1 and Atg5 kwockdown attenuated LAMP2 expression in NE cells. On the other hand, LAMP2 knockdown by siRNA led to a marked blockage of autophagy, prevention of NE differentiation and decrease of cell survival. Taken together, these results suggest that LAMP2 overexpression assists NE differentiation of LNCaP cells induced by serum deprivation and facilitates autophagy activity in order to attain the NE phenotype and cell survival. LAMP2 could thus be a potential biomarker and potential target for NE prostate cancer.