Simulating the Role of RAD51 Over-Expression on DNA Repair and its Potential Impact on Tumor Prognosis via Targeted Gene T herapy

Abstract

ROS-induced DNA damage is particularly detrimental in the context of cancers as it oxidatively stresses the tumor microenvironment (TME). Cells in the TME play a vital role in cancer pathogenesis, and by understanding the therapeutic strategies, ROS-induced damage can be mitigated. ROS causes single-strand DNA breaks (SSBs) and double-strand breaks (DSBs), and the accumulation of these breaks leads to genomic instability and cancer progression. This study focuses on the pathway and effects of introducing DNA repair proteins (RAD51 in this study) via adenoviral vectors into the TME to reduce ROS-induced DNA damage. RAD51 is a repair protein involved in homologous recombination to anneal DSBs. Adenoviruses are double-stranded DNA viruses ubiquitous in common diseases; however, their DNA sequence/plasmid is commonly used in clinical gene therapies as a medium of gene insertion. This study simulates various laboratory techniques necessary for adenoviral generation.

After insertion, the effects of RAD51 are theorized in a Western Blot measuring phosphorylated H2AX (γH2AX). γH2AX can be used as a biomarker for RAD51, as it plays a vital role in accumulating DNA repair proteins at the site of the break: when there are more breaks, there will be a larger concentration of γH2AX. This study is a gene therapy methodology that simulates adenoviral generation, focusing on the engineered vector designed for therapy. If future preclinical studies validate this research, it could lead to significant benefits for patients, such as reducing their resistance to radiotherapy and decreasing overall malignancy.