Generation of lung cancer cell lines harboring EGFR T790M mutation by CRISPR/Cas9-mediated genome editing

Area of Science:

• Oncology
• Molecular Biology
• Genetics

Background:

• Tyrosine kinase inhibitors (TKIs) like gefitinib are effective for EGFR-mutated lung adenocarcinomas.
• Acquired resistance, often due to the EGFR T790M mutation, limits TKI efficacy.
• Novel drug development requires robust models to study resistance mechanisms.

Purpose of the Study:

• To generate a lung cancer cell line with the specific EGFR T790M mutation using CRISPR/Cas9 genome editing.
• To compare the characteristics of CRISPR/Cas9-generated EGFR T790M mutations with those developed through drug exposure.
• To evaluate the utility of these engineered cell lines in studying drug resistance and efficacy.

Main Methods:

• Utilized CRISPR/Cas9 genome editing to introduce the EGFR T790M mutation into the PC9 lung cancer cell line.
• Employed EGFR pyrosequencing and peptide nucleic acid clamping for mutation rate analysis.
• Assessed gefitinib resistance and sensitivity to the third-generation TKI AZD9291.

Main Results:

• CRISPR/Cas9 generated PC9 cells exhibited a higher EGFR T790M mutation rate compared to cells adapted through long-term gefitinib exposure (PC9-G).
• Engineered PC9 cells demonstrated increased resistance to gefitinib compared to PC9-G cells.
• The generated cell lines showed significant sensitivity to AZD9291, a third-generation EGFR TKI.

Conclusions:

• CRISPR/Cas9 is a powerful tool for generating specific cancer cell line mutations to study drug resistance.
• Engineered cell lines provide valuable models for investigating resistance mechanisms and evaluating novel therapeutic agents like AZD9291.
• This approach facilitates research into overcoming TKI resistance in EGFR-mutated lung adenocarcinoma.