沉香中2-(2-苯乙基)色酮类成分及其抗KRAS突变NSCLC活性

2-(2-Phenylethyl)chromones from agarwood of Aquilaria agallocha and their inhibitory activity against KRAS mutant NSCLC

研究瑞香科植物沉香中2-(2-苯乙基)色酮类成分及其抗KRAS突变非小细胞肺癌(non-small cell lung cancer,NSCLC)活性。综合利用硅胶、RP-18反相硅胶、MCIgel CH20P、Diol和半制备HPLC等柱色谱技术,以及1D、2DNMR和ESI-MS等方法,对沉香中2-(2-苯乙基)色酮类化合物进行分离、纯化和鉴定;采用CCK-8试剂盒(cell countingkit 8)在3种KRAS突变NSCLC细胞上对分离得到的单体化合物进行抗肿瘤活性筛选。进一步对活性较好的化合物8进行细胞增殖、克隆形成、黏附能力及周期阻滞活性分析;采用分子对接、Westernblot实验开展机制研究;通过斑马鱼异种移植瘤(cellderivedxenograft,CDX)模型评价化合物8体内抗肿瘤活性。从沉香乙酸乙酯萃取部位中分离得到19个已知的2-(2-苯乙基)色酮;在A549(KRASG12S)细胞上,化合物8、10、19抑制活性较好,在H23(KRASG12C)细胞上,化合物7、8、19抑制活性较好,在H358(KRASG12C)细胞上,化合物8、10、16抑制活性较好,化合物8在三株细胞中抑制活性均较好,且能够有效抑制细胞增殖、克隆形成和黏附能力,并将H23细胞周期阻滞于G2/M期;化合物8能够抑制c-Met及其下游信号通路的表达,能够抑制斑马鱼CDX模型体内肿瘤的增殖。综上所述,19个已知的2-(2-苯乙基)色酮化合物中,化合物8抗KRAS突变NSCLC活性最好,主要通过将细胞阻滞于G2/M期抑制KRAS突变NSCLC的增殖。

The 2-(2-phenylethyl)chromones were separated from agarwood of Aquilaria agallocha Roxb.and their anti-KRAS mutant non-small cell lung cancer(NSCLC)activities were evaluated.2-(2-Phenyethyl)chromones in agarwood were separated and purified by silica gel,RP-18 reverse phase silica gel,MCI gel CH20P,Diol,and semi preparative HPLC chromatography techniques,while the structures of the compounds were identified by extensive spectroscopic analysis,such as 1D and 2D NMR and ESI-MS.The cell counting kit 8(CCK-8)assay was used to screen the anti-tumor activity of the isolated monomeric compounds on three KRAS-mutant NSCLC cells.The cell proliferation,cloning formation,adhesion ability,and cell cycle arrest activity of compound 8 were analyzed.Molecular docking and Western blot experiments were used to study the mechanism of compound 8.The in vivo anti-tumor activity of the compound 8 was evaluated by zebrafish cell derived xenograft(CDX)model.Nineteen known 2-(2-phenylethyl)chromones were isolated from the ethyl acetate extract of agarwood.On A549(KRAS G12S)cells,compounds 8,10,and 19 showed good inhibitory activity,on H23(KRAS G12C)cells,compounds 7,8,and 19 showed good inhibitory activity,and on H358(KRAS G12C)cells,compounds 8,10,and 16 showed good inhibitory activity.Compound 8 had the best inhibitory activity in all three cell lines.It effectively inhibited cell proliferation,clone formation,adhesion ability,and arrested the H23 cell cycle at G2/M phase.Compound 8 could also inhibit the expression of c-Met and its downstream signaling pathways,effectively inhibiting tumor growth in zebrafish CDX model.In conclusion,among the nineteen known 2-(2-phenylethyl)chromones,compound 8 had the best activity and significantly inhibited the proliferation of KRAS-mutant NSCLC cells by arresting the cells in the G2/M phase.