Oxidative stress mediated by pyrrolidine SS13 is involved in autophagic cell death induction in colorectal cancer cells

Abstract

Abstract Pyrrolidines, nitrogenous organic compounds, are among the most intensively studied substances because of their antibacterial, antiviral, neurological, and promising antitumor effects. This study aimed to evaluate the pro-oxidative properties of SS13 on human colorectal carcinoma cell lines (HCT116 and Caco-2) using flow cytometry, western blot, fluorescence microscopy and qRT-PCR. Pyrrolidine SS13 induced the accumulation of reactive oxygen and nitrogen species in both cell lines. N-acetyl cysteine and Trolox, known antioxidants, reduced the increased levels of free radicals mediated by SS13 treatment. Moreover, modulation of both superoxide dismutase isoenzymes (SOD1, SOD2) was confirmed by western blot analysis and qRT-PCR. Oxidative stress was also associated with DNA damage response system activation and modulation of stress/survival pathways. We demonstrated for the first time that pyrrolidine SS13 is involved in the induction of autophagy by increasing the levels of autophagic markers (p-AMPK, p-ULK, LC3I/II, and ATG7). In addition, a significant decreasing effect on p62 protein levels was observed in both cell lines. Finally, chloroquine, an autophagy inhibitor, increased cell survival and suppressed the cytotoxic effect of SS13 in HCT116 and Caco-2 cells, indicating the involvement of SS13 in autophagy-mediated cell death. Taken together, our results suggest that oxidative stress and autophagy contribute to the antiproliferative effect of pyrrolidine SS13 on colorectal cancer cells.