CRISPR/Cas9 Knockout of EGFR as an Approach to Overcome Bladder Cancer Chemoresistance

Abstract

RTK EGFR regulates cell growth, proliferation, and survival by activating the PI3K/AKT and MAPK signaling pathways. Cells with EGFR overexpression uncontrollably proliferate, evade apoptosis, and undergo angiogenesis, which are typical hallmarks of cancer. Based on the world statistics, more than 200,000 patients diagnosed with bladder cancer die yearly (Kawahara et al., 2024). Patient samples with urothelial carcinoma show significantly upregulated levels of EGFR and HER2. Moreover, there is a correlation between high EGFR expression levels and poor response to treatment and increased chemoresistance. In addition, EGFR activation drives epithelial-mesenchymal transition (EMT), enhancing cancer cells’ resistance to drug treatment and promoting cell migration. Consequently, the chemoresistance of bladder cancer is a major obstacle affecting the treatment. Thus, we hypothesized that gene knockouts of the EGFR family member could act as anticancer agent that can reduce chemoresistance in bladder cancer. This thesis aimed to develop efficient knockout of the EGFR RTK in the T24 bladder cancer cell line using two sgRNAs in CRISPR/Cas9 system. This approach allows to evaluate the role of EGFR in cancer cell growth. Furthermore, this thesis aimed to determine the ability of specific gene knockouts to re-establish sensitivity to chemotherapy in bladder cancer cells.