肉碱棕榈酰转移酶-2(CPT2)的下调已被证明与几种癌症的进展高度相关。但是对它的表达知之甚少,结直肠癌(CRC)的生物学功能和机制。
肿瘤基因组图谱(TCGA)和基因表达综合(GEO)数据集的生物信息学分析用于探索CPT2的表达,CPT2表达与临床病理特征之间的关系。以及CRC的总体生存率。采用Cox比例风险回归模型分析影响CRC预后的独立因素。体外,通过RT-qPCR分析CRC组织,IHC,IF和蛋白质印迹以验证CPT2表达。殖民地的形成,CCK-8,细胞周期,凋亡,进行了transwell和伤口愈合试验以检查CPT2在CRC中的功能。在体内,裸鼠异种移植实验用于进一步检查CPT2对肿瘤发生的影响。此外,基因集富集分析(GSEA)探索CPT2的下游通路。通过RT-qPCR和Western印迹验证了CPT2对p53通路的调控。
CPT2表达在CRC中经常下调,并与不良预后相关。低CPT2表达与年龄显著相关,淋巴结转移,远处转移和TMN分期。单变量和多变量分析表明,低CPT2表达是总生存期较差的独立预后因素。功能上,CPT2在CRC细胞中的过表达诱导的生长抑制,细胞周期停滞在G1期,细胞凋亡增强,细胞迁移和侵袭减少。相反,敲低CPT2有助于细胞增殖,移民和入侵,增加S期细胞的比例,降低G1期细胞比例,抑制细胞凋亡。机械上,我们发现CPT2过表达可以通过激活p-p53增加p53的表达,导致p21,Bax,裂解的caspase-9,裂解的caspase-3和裂解的PARP激活和Bcl2,MDM2失活,从而抑制肿瘤增殖和促进细胞凋亡。CPT2敲低产生相反的结果。
这些结果表明,CPT2可能是CRC的一种新的预后标志物,CPT2的下调可以通过p53途径促进CRC的增殖和抑制凋亡。可以开发靶向CPT2的策略作为CRC的疗法。
Downregulation of Carnitine palmitoyltransferase-2 (
CPT2) has been shown to be highly associated with the progression of several cancers, but little known about its expression, biological functions and mechanisms in colorectal cancer (CRC).
Bioinformatics analysis of The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) data sets was used to explore the expression of CPT2, the relationship between CPT2 expression and clinicopathologic features, as well as the overall survival of CRC. Cox\'s proportional hazards regression model was used to analyze independent prognostic factors of CRC. In vitro, CRC tissues were analyzed by RT-qPCR, IHC, IF and western blotting to verify CPT2 expression. Colony formation, CCK-8, cell cycle, apoptosis, transwell and wound healing assays were performed to examine the functions of CPT2 in CRC. In vivo, nude mouse xenograft experiment was used to further examine the effect of CPT2 on tumorigenesis. Furthermore, gene set enrichment analysis (GSEA) was conducted to explore the downstream pathway of CPT2. The regulation of p53 pathway by
CPT2 was verified by RT-qPCR and Western blotting.
CPT2 expression was frequently downregulated in CRC and correlated with poor prognosis. Low CPT2 expression was significantly associated with age, lymph node metastasis, distant metastasis and TMN stage. Univariate and multivariate analysis indicated that low CPT2 expression was an independent prognostic factor for poorer overall survival. Functionally, overexpression of CPT2 in CRC cells induced growth suppression, cell cycle arrest at the G1 phase, enhanced apoptosis and reduced cell migration and invasion. Conversely, knockdown of CPT2 contributed to cell proliferation, migration and invasion, increased the proportion of S phase cells, decreased the proportion of G1 phase cells and inhibited apoptosis. Mechanistically, we found that CPT2 overexpression can increase p53 expression by activating p-p53, leading to p21, Bax, cleaved caspase-9, cleaved caspase-3 and cleaved PARP activation and Bcl2, MDM2 deactivation, thereby inhibiting tumor proliferation and promoting apoptosis.
CPT2 knockdown yielded opposite results.
These findings suggest that
CPT2 may be a novel prognostic marker of CRC and downregulation of
CPT2 can promote proliferation and inhibit apoptosis through p53 pathway in CRC. Strategies targeting
CPT2 may be developed as therapies for CRC.