摘要Objective:To explore the role of the natural compound hesperidin in glycolysis,the key ratelimiting enzyme,in colorectal cancer(CRC)cell lines.Methods:In vitro,HCT116 and SW620 were treated with different doses of hesperidin(0-500 μ mol/L),cell counting kit-8 and colone formation assays were utilized to detected inhibition effect of hesperidin on CRC cell lines.Transwell and wound healing assays were performed to detect the ability of hesperidin(0,25,50 and 75 μ mol/L)to migrate CRC cells.To confirm the apoptotic-inducing effect of hesperidin,apoptosis and cycle assays were employed.Western blot,glucose uptake,and lactate production determination measurements were applied to determine inhibitory effects of hesperidin(0,25 and 50 μ mol/L)on glycolysis.In vivo,according to the random number table method,nude mice with successful tumor loading were randomly divided into vehicle,low-dose hesperidin(20 mg/kg)and high-dose hesperidin(60 mg/kg)groups,with 6 mice in each group.The body weights and tumor volumes of mice were recorded during 4-week treatment.The expression of key glycolysis rate-limiting enzymes was determined using Western blot,and glucose uptake and lactate production were assessed.Finally,protein interactions were probed with DirectDIA Quantitative Proteomics,Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)analyses.Results:Hesperidin could inhibit CRC cell line growth(P＜0.05 or P＜0.01).Moreover,hesperidin presented an inhibitory effect on the migrating abilities of CRC cells.Hesperidin also promoted apoptosis and cell cycle alterations(P＜0.05).The immunoblotting results manifested that hesperidin decreased the levels of hexokinase 2,glucose transporter protein 1(GLUT1),GLUT3,L-lactate dehydrogenase A,6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 2(PFKFB2),PFKFB3,and pyruvate kinase isozymes M2(P＜0.01).It remarkably suppressed tumor xenograft growth in nude mice.GO and KEGG analyses showed that hesperidin treatment altered metabolic function.Conclusion:Hesperidin inhibits glycolysis and is a potential therapeutic choice for CRC treatment.