A macrocyclic kinase inhibitor overcomes triple resistant mutations in EGFR-positive lung cancer.

Brigatinib-based therapy was effective against osimertinib-resistant EGFR C797S mutants and is undergoing clinical studies. However, tumor relapse suggests additional resistance mutations might emerge. Here, we first demonstrated the binding mode of brigatinib to the EGFR-T790M/C797S mutant by crystal structure analysis and predicted brigatinib-resistant mutations through a cell-based assay including N-ethyl-N-nitrosourea (ENU) mutagenesis. We found that clinically reported L718 and G796 compound mutations appeared, consistent with their proximity to the binding site of brigatinib, and brigatinib-resistant quadruple mutants such as EGFR-activating mutation/T790M/C797S/L718M were resistant to all the clinically available EGFR-TKIs. BI-4020, a fourth-generation EGFR inhibitor with a macrocyclic structure, overcomes the quadruple and major EGFR-activating mutants but not the minor mutants, such as L747P or S768I. Molecular dynamics simulation revealed the binding mode and affinity between BI-4020 and EGFR mutants. This study identified potential therapeutic strategies using the new-generation macrocyclic EGFR inhibitor to overcome the emerging ultimate resistance mutants.

Human-Derived Corneal Epithelial Cells Expressing Cell Cycle Regulators as a New Resource for in vitro Ocular Toxicity Testing.

The Draize test has been used on rabbits since the 1960s to evaluate the irritation caused by commercial chemicals in products such as cosmetics or hairdressing products. However, since 2003, such tests, including the Draize test for cosmetics, have been prohibited in European countries because they are considered problematic to animal welfare. For this reason, replacement of in vivo methods with the alternative in vitro methods has become an important goal. In this study, we established a corneal epithelial cell line co-expressing a mutant cyclin-dependent kinase 4 (CDK4), Cyclin D1, and telomerase reverse transcriptase (TERT). The established cell line maintained its original morphology and had an enhanced proliferation rate. Furthermore, the cells showed a significant, dose-dependent decrease in viability in an irritation test using glycolic acid and Benzalkonium chloride. These cells can now be shared with toxicology scientists and should contribute to increasing the reproducibility of chemical testing in vitro.