Nosalova Natalia, Majirska Monika, Keselakova Alexandra, Martinkova Miroslava, Fabianova Dominika, Mirossay Andrej, Pilatova Martina Bago, Kello Martin
Department of Pharmacology, Faculty of Medicine, P.J. Šafárik University, Košice, Slovakia.
Institute of Chemical Sciences, Department of Organic Chemistry, Faculty of Science, P.J. Šafárik University, Košice, Slovakia.
Eur J Pharm Sci. 2025 Feb 1;205:106982. doi: 10.1016/j.ejps.2024.106982. Epub 2024 Dec 5.
Pyrrolidines, nitrogenous organic compounds, are among the most intensively studied agents because of their antibacterial, antiviral, neurological, and promising antitumor effects. Moreover, many medicinal drugs contain pyrrolidine moiety such as sunitinib (anticancer drug), telaprevir and ombitasvir (antiviral drugs) or ramipril (antihypertensive drug).
Based on the pro-apoptotic effect of pyrrolidine SS13, this study focuses on the pro-oxidative properties of the tested pyrrolidine SS13 on colorectal cancer cells to deepen the understanding of its mechanisms of action.
We hypothesize that SS13 induces oxidative stress and autophagy activation in HCT116 and Caco-2 cell lines, thus contributing to antiproliferative effects.
Flow cytometry, western blot, fluorescence microscopy and qRT-PCR were used to evaluate the effect of pyrrolidine SS13.
Pyrrolidine SS13 induced oxidative stress through the accumulation of reactive oxygen and nitrogen species in both cell lines and the modulation of both superoxide dismutase isoenzymes (SOD1, SOD2). Oxidative stress was also associated with the activation of DNA damage response system and modulation of stress/survival pathways. We demonstrated for the first time that pyrrolidine SS13 is involved in the induction of autophagy accompanied by increased levels of autophagic markers (p-AMPK, p-ULK, LC3I/II and ATG7) and a significant decrease in p62 protein levels in both cell lines. Finally, chloroquine, an inhibitor of autophagy, enhanced cell survival and suppressed the cytotoxic effect of SS13 in HCT116 and Caco-2 cells, indicating that SS13 contributes to autophagy-mediated cell death. Taken together, our results suggest that oxidative stress and autophagy participate in the antiproliferative effect of pyrrolidine SS13 on colorectal cancer cells. Further research using primary cell cultures obtained from different animal tissues as well as performing in vivo experiments is needed to understand these processes in detail and to investigate the potential therapeutic application of new pyrrolidine derivatives.
吡咯烷是含氮有机化合物，因其具有抗菌、抗病毒、神经学及有前景的抗肿瘤作用，成为研究最为深入的一类物质。此外，许多药物都含有吡咯烷部分，如舒尼替尼（抗癌药）、特拉匹韦和奥比他韦（抗病毒药）或雷米普利（抗高血压药）。
基于吡咯烷SS13的促凋亡作用，本研究聚焦于所测试的吡咯烷SS13对结肠癌细胞的促氧化特性，以加深对其作用机制的理解。
我们假设SS13在HCT116和Caco - 2细胞系中诱导氧化应激和自噬激活，从而产生抗增殖作用。
采用流式细胞术、蛋白质印迹法、荧光显微镜和定量逆转录聚合酶链反应来评估吡咯烷SS13的作用。
吡咯烷SS13通过在两种细胞系中积累活性氧和氮物种以及调节两种超氧化物歧化酶同工酶（SOD1、SOD2）来诱导氧化应激。氧化应激还与DNA损伤反应系统的激活以及应激/存活途径的调节有关。我们首次证明吡咯烷SS13参与自噬的诱导，伴随着两种细胞系中自噬标志物（p - AMPK、p - ULK、LC3I/II和ATG7）水平的升高以及p62蛋白水平的显著降低。最后，自噬抑制剂氯喹增强了细胞存活并抑制了SS13在HCT116和Caco - 2细胞中的细胞毒性作用，表明SS13促成自噬介导的细胞死亡。综上所述，我们的结果表明氧化应激和自噬参与了吡咯烷SS13对结肠癌细胞的抗增殖作用。需要使用从不同动物组织获得的原代细胞培养物进行进一步研究，并开展体内实验，以详细了解这些过程，并研究新型吡咯烷衍生物的潜在治疗应用。
