Evaluating kinome-wide resistance mechanisms to 3rd generation EGFR inhibitors


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Emma Fairweather1,Patrick Eyers1
1University of Liverpool

Abstract

Background

Lung cancer has the highest cancer-related mortality worldwide and is a major cause of death in the North West of England, especially Merseyside. Mutations in the EGFR tyrosine kinase (TK) are frequently observed in ~10-15% of cases of adenocarcinomas. Moreover, drug resistance to EGFR TK inhibitors develops in as little as 10-12 months in patients. Osimertinib, a third generation covalent inhibitor, was originally designed to overcome the prevalent T790M gatekeeper mutation associated with TKI-treated patients. However, induced resistance to osimertinib is already being observed clinically, including a C797S mutation at the covalent site of drug binding in the target EGFR kinase.

Method

Using NanoString kinome mRNA profiling we analysed transcriptional signatures of drug-tolerant persister cells (DTPs), which emerge in model cultured lung cancer cells that have been challenged with both osimertinib and unrelated EGFR TKIs. Computational analysis of NanoString data by PCA analysis, hierarchical cluster profiling and pathway/gene signature association analysis was used to evaluate signaling profiles. NanoString data was validated using conventional RT-PCR and immunoblotting.

Results

Interrogation of cellular ‘rewiring’ mechanisms present in DTPs identifies differential transcriptomic profiles in both drug-persister and drug-resistant cell populations, including several novel protein kinases that are upregulated in response to drug exposure.

Conclusion

The analysis of kinome-wide signalling network dynamics has the potential to identify biomarkers and/or emerging new drug targets of potential therapeutic benefit in TK-inhibitor exposed NSCLC patients. Further validated with siRNA and CRISPR will confirm whether these kinases are necessary, sufficient, or both, for drug resistance and cell survival.