Drug Sensitivity and Allele Specificity of First-Line Osimertinib Resistance EGFR Mutations.

Osimertinib, a mutant-specific third-generation EGFR tyrosine kinase inhibitor, is emerging as the preferred first-line therapy for EGFR-mutant lung cancer, yet resistance inevitably develops in patients. We modeled acquired resistance to osimertinib in transgenic mouse models of EGFRL858R -induced lung adenocarcinoma and found that it is mediated largely through secondary mutations in EGFR-either C797S or L718V/Q. Analysis of circulating free DNA data from patients revealed that L718Q/V mutations almost always occur in the context of an L858R driver mutation. Therapeutic testing in mice revealed that both erlotinib and afatinib caused regression of osimertinib-resistant C797S-containing tumors, whereas only afatinib was effective on L718Q mutant tumors. Combination first-line osimertinib plus erlotinib treatment prevented the emergence of secondary mutations in EGFR. These findings highlight how knowledge of the specific characteristics of resistance mutations is important for determining potential subsequent treatment approaches and suggest strategies to overcome or prevent osimertinib resistance in vivo. SIGNIFICANCE: This study provides insight into the biological and molecular properties of osimertinib resistance EGFR mutations and evaluates therapeutic strategies to overcome resistance. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/10/2017/F1.large.jpg.

CRISPR/Cas9 editing of the genome for cancer modeling.

The CRISPR/Cas9 revolution has democratized access to genome editing in many biological fields, including cancer research. Cancer results from the multistep accumulation of mutations that confer to the transformed cells certain biological hallmarks typical of the malignant phenotype. One of the major goals in cancer research is to characterize such mutations and assess their implication in the oncogenic process. Through CRISPR/Cas9 technology, genetic aberrations identified in a patient's tumor can now be easily recreated in experimental models, which can then be used for basic research or for more translational applications. Here we review the different CRISPR/Cas9 strategies that have been implemented to recapitulate oncogenic mutations in both in vitro and in vivo systems, including novel strategies to model tumor evolution and genetic heterogeneity.

CRISPR-Barcoding for Intratumor Genetic Heterogeneity Modeling and Functional Analysis of Oncogenic Driver Mutations.

Intratumor genetic heterogeneity underlies the ability of tumors to evolve and adapt to different environmental conditions. Using CRISPR/Cas9 technology and specific DNA barcodes, we devised a strategy to recapitulate and trace the emergence of subpopulations of cancer cells containing a mutation of interest. We used this approach to model different mechanisms of lung cancer cell resistance to EGFR inhibitors and to assess effects of combined drug therapies. By overcoming intrinsic limitations of current approaches, CRISPR-barcoding also enables investigation of most types of genetic modifications, including repair of oncogenic driver mutations. Finally, we used highly complex barcodes inserted at a specific genome location as a means of simultaneously tracing the fates of many thousands of genetically labeled cancer cells. CRISPR-barcoding is a straightforward and highly flexible method that should greatly facilitate the functional investigation of specific mutations, in a context that closely mimics the complexity of cancer.

Modeling intratumor heterogeneity through CRISPR-barcodes.

We have devised a barcoding strategy to recapitulate cancer evolution through the emergence of subclonal mutations of interest, whose effects can be monitored in a dynamic manner. This approach can be easily adapted for a variety of applications, including combined modeling of multiple mechanisms of drug resistance or repair of oncogenic driver mutations in addicted cancer cells.