MA11.01 Development of Novel EGFR Mutant NSCLC Mouse Models and Murine Cell Lines: New Tools for NSCLC Research

Abstract

EGFR mutant lung cancer accounts for approximately 15-30% of all non-small cell lung cancer (NSCLC) cases, with the prevalence varying in different populations. A point mutation L858R and in-frame deletions in exon 19 of the EGFR gene account for the majority of all EGFR activating mutations. EGFR tyrosine kinase inhibitors (TKIs) have led to significant advances in the treatment of EGFR mutant NSCLC; however, many patients show a partial response and all eventually develop resistance. Better preclinical models are required to design rational combination therapies. Our group has used syngeneic orthotopic mouse models of NSCLC to study interactions between cancer cells and the tumor microenvironment in the lung. These approaches are limited by the availability of murine lung cancer cell lines. To our knowledge, there are currently no EGFR mutant murine lung cancer cell lines. Therefore, we developed a novel transgenic mouse strain that conditionally expresses mutant EGFR mediated by Cre-recombinase. Administration of adeno-Cre virus into the lungs resulted in the development of EGFR-driven lung tumors. In collaboration with the Mouse Genetics Core at National Jewish Health, we developed two novel EGFR mutant mouse models (Rosa-loxP-STOP-LoxP- EGFR L860R/del19-mC3-WPRE): one with a deletion 19 and one with an L860R mutation (equivalent to the human L858R mutation) in the mouse EGFR gene. Tumors are initiated through intratracheal injections of the adeno-Cre virus. These mice were bred with p53flox mice from Jackson Laboratories to increase the diversity of tumors. Tumors were harvested and submitted to histological characterization by H&E staining or processed to create cell lines. These cell lines were characterized by in vitro growth assays, western blot analyses, and sequencing to confirm their genotype and responsiveness to EGFR TKIs. Cell lines were implanted into both the flanks and lungs of mice to determine their responsiveness to EGFR TKIs. Tumor-bearing mice were imaged by CT and treated by oral gavage with the EGFR TKIs. Intratracheal administration of adeno-Cre led to the development of multiple tumors throughout all lobes of the lung in both del19 and L860R mice. Time to tumor formation occurred within 4-12 weeks. Importantly, treatment of the mice with osimertinib resulted in tumor shrinkage and improvement in the health of mice. Moreover, we were able to successfully isolate 3 distinct cell lines (del19.1, del19.2, L860R.1) from these tumors. In vitro, these cell lines are responsive to multiple EGFR TKIs, as evidenced by a decrease in proliferation and inhibition of phospho-ERK. These cells form tumors when implanted into the flanks of nu/nu mice, del19.1-derived tumors undergo shrinkage in response to osimertinib. Orthotopically injected del19.1 cells form tumors in the left lungs of WT C57Bl/6 mice. We have developed novel mouse models as well as murine cell lines that reflect mutant EGFR-driven NSCLC which will be useful tools for studying NSCLC both in vitro and in vivo. By increasing the number of murine NSCLC cell lines, we can better recapitulate human disease to increase our understanding of NSCLC and develop novel therapeutic strategies that can be used to treat patients.