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Abstract：
Uterine Corpus Endometrial Carcinoma (UCEC) is a significant health concern with a complex genetic landscape impacting disease susceptibility and progression. This study aimed to unravel the spectrum of DNA repair gene mutations in Pakistani UCEC patients through Next Generation Sequencing (NGS) and explore their potential functional consequences via downstream analyses. NGS analysis of genomic DNA from 30 UCEC patients was conducted to identify clinically significant pathogenic mutations in DNA repair genes. This analysis revealed mutations in 4 key DNA repair genes: BRCA1, BRCA2, APC, and CDH1. Kaplan-Meier (KM) analysis was employed to assess the prognostic value of these mutations on patient overall survival (OS) in UCEC. To delve into the functional impact of these mutations, we performed RT-qPCR, immunohistochemistry (IHC), and western blot analyses on the mutated UCEC samples compared to their non-mutated counterparts. These results unveiled the up-regulation in the expression of the mutated genes, suggesting a potential association between the identified mutations and enhanced gene activity. Additionally, targeted bisulfite sequencing analysis was utilized to evaluate DNA methylation patterns in the promoters of the mutated genes. Strikingly, hypomethylation in the promoters of BRCA1, BRCA2, APC, and CDH1 was observed in the mutated UCEC samples relative to the non-mutated, indicating the involvement of epigenetic mechanisms in the altered gene expression. In conclusion, this study offers insights into the genetic landscape of DNA repair gene mutations in Pakistani UCEC patients. The presence of pathogenic mutations in BRCA1, BRCA2, APC, and CDH1, coupled with their down-regulation and hypermethylation, suggests a convergence of genetic and epigenetic factors contributing to genomic instability in UCEC cells. These findings enhance our understanding of UCEC susceptibility and provide potential avenues for targeted therapeutic interventions in Pakistani UCEC patients.
摘要：
子宫内膜癌（UCEC）是一个重要的健康问题，具有影响疾病易感性和进展的复杂遗传景观。这项研究旨在通过下一代测序（NGS）揭示巴基斯坦UCEC患者的DNA修复基因突变谱，并通过下游分析探索其潜在的功能后果。对来自30名UCEC患者的基因组DNA进行NGS分析以鉴定DNA修复基因中的临床显著致病性突变。这项分析揭示了4个关键DNA修复基因的突变：BRCA1，BRCA2，APC，和CDH1。采用Kaplan-Meier（KM）分析评估这些突变对UCEC患者总生存期（OS）的预后价值。为了深入研究这些突变的功能影响，我们进行了RT-qPCR，免疫组织化学（IHC），和蛋白质印迹分析的突变的UCEC样品相比，其非突变的对应物。这些结果揭示了突变基因表达的上调，表明鉴定的突变与增强的基因活性之间存在潜在的关联。此外,靶向亚硫酸氢盐测序分析用于评估突变基因启动子中的DNA甲基化模式。引人注目的是，BRCA1，BRCA2，APC启动子中的低甲基化，和CDH1在突变的UCEC样品中观察到相对于未突变的，表明表观遗传机制参与改变的基因表达。总之,这项研究为巴基斯坦UCEC患者DNA修复基因突变的遗传前景提供了见解。BRCA1、BRCA2、APC、和CDH1，再加上它们的下调和超甲基化，表明遗传和表观遗传因素的趋同导致UCEC细胞中的基因组不稳定。这些发现增强了我们对UCEC易感性的理解，并为巴基斯坦UCEC患者的针对性治疗干预提供了潜在的途径。
