Wen Lina, Han Zongqiang, Li Jianhui, Du Yanlin
Department of Clinical Nutrition, Beijing Shijitan Hospital, Capital Medical University, Beijing, China.
Key Laboratory of Cancer FSMP for State Market Regulation, Beijing, China.
J Gastrointest Oncol. 2022 Jun;13(3):1152-1168. doi: 10.21037/jgo-22-389.
G-quadruplexes are molecular switches regulating gene transcription. and hypoxia-inducible factor 1-alpha () play important roles in cell proliferation, apoptosis, and metabolic regulation in colon cancer. Whether berberine can regulate metabolism by interacting with and G-quadruplexes in colon cancer needs to be explored.
The binding mode of berberine with and G-quadruplexes were explored by ultraviolet and visible absorption spectroscopy and fluorescence spectroscopy. Circular dichroism (CD) spectroscopy was performed to evaluate the effects of berberine on the stability of and G-quadruplexes. After different concentrations of berberine acting on HCT116 cells for 24 h, cell proliferation and apoptosis were detected by MTT assay and flow cytometry; quantitative real-time polymerase chain reaction and western blot were performed to detect mRNA and protein expression of and ; transcriptome sequencing was used to analyze the metabolic pathways. For the effects of berberine on colon cancer mouse model with dose of 50 mg·kg for 14 days, tumor growth were monitored, hematoxylin and eosin staining and immunofluorescence staining were performed to analyze histopathology and protein expression of and , central carbon metabolism was detected in tumor tissues.
The binding ability of berberine with G-quadruplex was different to that of berberine with G-quadruplex. Both binding modes involved π-π stacking. The stoichiometric ratios were 1:1, 1:3, and 3:1 for berberine with G-quadruplex and only 1:1 for berberine with G-quadruplex. Temperature had a greater effect on the binding of berberine to G-quadruplex. Berberine could improve the thermal stability of both and G-quadruplexes. Berberine inhibited the gene transcription and protein expression of and in colon cancer HCT116 cells. , berberine delayed tumor progression and inhibited the protein expression of and . Twelve differential metabolites such as decreased adenosine triphosphate were obtained, indicating that berberine could regulate the metabolic pathways of the tricarboxylic acid (TCA) cycle and glycolysis/gluconeogenesis, among others.
Berberine may inhibit colon cancer by regulating the TCA cycle and glycolysis/gluconeogenesis based on the interaction with and G-quadruplexes.
G-四链体是调节基因转录的分子开关。缺氧诱导因子1α（HIF-1α）在结肠癌的细胞增殖、凋亡和代谢调节中起重要作用。小檗碱是否能通过与结肠癌中的HIF-1α和G-四链体相互作用来调节代谢，有待探索。
采用紫外可见吸收光谱和荧光光谱研究小檗碱与HIF-1α和G-四链体的结合模式。利用圆二色（CD）光谱评估小檗碱对HIF-1α和G-四链体稳定性的影响。不同浓度的小檗碱作用于HCT116细胞24小时后，采用MTT法和流式细胞术检测细胞增殖和凋亡情况；通过定量实时聚合酶链反应和蛋白质免疫印迹法检测HIF-1α和HIF-2α的mRNA和蛋白表达；利用转录组测序分析代谢途径。对于给予剂量为50mg·kg的小檗碱作用14天的结肠癌小鼠模型，监测肿瘤生长情况，进行苏木精-伊红染色和免疫荧光染色以分析HIF-1α和HIF-2α的组织病理学和蛋白表达，检测肿瘤组织中的中心碳代谢。
小檗碱与HIF-1α G-四链体的结合能力不同于其与HIF-2α G-四链体的结合能力。两种结合模式均涉及π-π堆积。小檗碱与HIF-1α G-四链体的化学计量比为1:1、1:3和3:1，而小檗碱与HIF-2α G-四链体的化学计量比仅为1:1。温度对小檗碱与HIF-1α G-四链体的结合影响更大。小檗碱可提高HIF-1α和HIF-2α G-四链体的热稳定性。小檗碱抑制结肠癌HCT116细胞中HIF-1α和HIF-2α的基因转录和蛋白表达。此外，小檗碱延缓肿瘤进展并抑制HIF-1α和HIF-2α的蛋白表达。获得了12种差异代谢物，如三磷酸腺苷减少，表明小檗碱可调节三羧酸（TCA）循环和糖酵解/糖异生等代谢途径。
小檗碱可能通过与HIF-1α和HIF-2α G-四链体相互作用调节TCA循环和糖酵解/糖异生，从而抑制结肠癌。
