PO.CL07.02 · 临床研究
Inavolisib在浆液性子宫内膜癌临床前模型中显示抗肿瘤作用
Inavolisib demonstrates anti-tumorigenic effects in pre-clinical models of serous endometrial cancer
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
目的：Inavolisib是一种口服选择性PI3K抑制剂，通过促进突变型p110alpha（PIK3CA编码的催化亚基）的降解，已在多种癌症临床前模型以及乳腺癌临床试验中显示出有前景的抗肿瘤活性。磷脂酰肌醇3-激酶（PI3K）信号级联在子宫浆液性癌（USC）——子宫内膜癌（EC）最具侵袭性的亚型之一——的肿瘤发生和进展中起关键作用。鉴于亟需为这一高致死性EC亚型开发更有效的治疗干预措施，且PI3Ka改变在USC中常见，我们旨在研究inavolisib在浆液性EC细胞系中的抗肿瘤和抗侵袭活性。
方法：本研究使用人浆液性EC细胞系ARK1和ARK2，并用inavolisib（获自Genentech）处理。通过MTT试验评估细胞增殖。通过Cellometer检测细胞周期进程。通过DCFH-DA试验评估细胞应激，通过JC-1试验测量线粒体膜电位的变化。使用切割型caspase-3试验评估凋亡。使用层粘连蛋白-1（laminin-1）试验评估细胞黏附，通过划痕愈合试验评估细胞迁移。使用蛋白质免疫印迹检测与细胞周期进程、细胞应激和凋亡相关的下游蛋白表达。
结果：用inavolisib处理72小时后，ARK1和ARK2细胞的增殖以剂量依赖方式受到抑制（IC50：ARK1 674 nM，ARK2 2969 nM）。用inavolisib处理两株细胞系均显著将细胞周期阻滞于G1期，升高细胞内活性氧水平，并降低线粒体膜电位（与未处理细胞相比，p<0.05）。Inavolisib还显著增加切割型caspase-3的活性，并降低两株细胞系中BcL-2和MCL-1的表达。此外，inavolisib在100和250 nM剂量下有效降低ARK1和ARK2细胞的黏附和迁移（p<0.05）。蛋白质免疫印迹结果表明，用inavolisib处理下调了PIK3CA/mTOR通路下游靶点（磷酸化[p]-AKT、p-S6）以及细胞周期蛋白CDK4和cyclin D1的表达，并上调了两株细胞中促凋亡蛋白Bax和PDI的表达。
结论：Inavolisib在浆液性EC细胞系中表现出强效的抗肿瘤和抗侵袭作用，提示这一新型药物是一种有前景的治疗选择，值得在侵袭性浆液性EC治疗中进一步探索。
查看英文原文 English abstract
Objectives: Inavolisib is an oral selective PI3K inhibitor that has shown promising anti-tumorigenic activity in multiple pre-clinical models of cancer as well as breast cancer clinical trials, via promoting the degradation of mutated p110alpha, the catalytic subunit encoded by PIK3CA. The phosphatidylinositol 3-kinase (PI3K) signaling cascade is critically implicated in the tumorigenesis and progression of uterine serous carcinoma (USC), one of the most aggressive subtypes of endometrial cancer (EC). Given the urgent need to develop more effective treatment interventions for this highly lethal subtype of EC and that PI3Ka alterations are common in USC, we aimed to investigate the anti-tumorigenic and anti-invasive activities of inavolisib in serous EC cell lines.
Methods: The human serous EC cell lines, ARK1 and ARK2, were used in this study and treated with inavolisib (obtained from Genentech). Cell proliferation was evaluated by MTT assay. Cell cycle progression was examined by Cellometer. Cellular stress was evaluated by DCFH-DA assay and change in mitochondrial membrane potential was measured by JC-1 assay. Apoptosis was evaluated using a cleaved caspase-3 assay. Cell adhesion was evaluated using a laminin-1 assay, and cell migration was assessed by wound healing assay. Western immunoblotting was used to measure downstream protein expression related to cell cycle progression, cellular stress, and apoptosis.
Results: After 72 hours of treatment with inavolisib, the proliferation of ARK1 and ARK2 cells was inhibited in a dose-dependent manner (IC50: ARK1 674 nM, ARK2 2969 nM). Treatment of both cell lines with inavolisib significantly arrested the cell cycle at the G1 phase, increased intracellular reactive oxygen species levels, and decreased mitochondrial membrane potential compared with untreated cells (p<0.05). Inavolisib also significantly increased the activity of cleaved caspase-3 and reduced the expression of BcL-2 and MCL-1 in both cell lines. Moreover, inavolisib effectively decreased cell adhesion and migration at doses of 100 and 250 nM in the ARK1 and ARK2 cells (p<0.05). Western immunoblotting results demonstrated that treatment with inavolisib downregulated the expression of downstream targets of the PIK3CA/mTOR pathway (phosphorylated [p]-AKT, p-S6) and the cell cycle proteins CDK4 and cyclin D1 and upregulated the expression of the pro-apoptotic Bax and PDI proteins in both cells.
Conclusions: Inavolisib exhibits potent anti-tumorigenic and anti-invasive effects in serous EC cell lines, suggesting that this novel agent is a promising therapeutic option worthy of further exploration in the treatment of aggressive serous ECs.
利益披露 Disclosure
M. Singleton, None.