Cancer modeling in colorectal organoids reveals intrinsic differences between oncogenic RAS and BRAF variants

Abstract

Colorectal cancers (CRCs) with oncogenic mutations in RAS and BRAF are associated with anti-EGFR therapy resistance. Consequently, all RAS mutant CRC patients are being excluded from this therapy. However, heterogeneity in drug response has been reported between RAS mutant CRC patients. It is poorly understood to what extent such differences are derived from different genetic backgrounds or intrinsic differences between the various RAS pathway mutations. Therefore, using CRISPR technology we generated an isogenic panel of patientderived CRC organoids with various RAS pathway mutations (i.e. KRAS G12D , BRAF V600E , KRAS G13D and NRAS G12D ). All RAS pathway mutants promote ERK activation and tumor growth. However, KRAS G12D and BRAF V600E mutations in particular conferred robust resistance to anti-EGFR therapy, both in vitro and in vivo. Moreover, untreated KRAS G13D mutants showed fastest growth in mice but remained sensitive to anti-EGFR therapy. Together, introducing mutationspecific oncogene signaling in CRC organoids resembles clinical phenotypes and improves understanding of genotype-phenotype correlations.

Rapid generation of precision preclinical cancer models using regulatable in vivo base editing

“…Organotypic cell culture models or ‘organoids’ are a powerful system to study epithelial biology. We and others have used organoids to reveal the contribution of cancer-associated nonsense and missense mutations for cell behavior and drug response 9,35,36 ; however, efficient introduction of such mutations using base editing is a substantial practical challenge in scaling up the generation of large collections of tailored model systems. To determine if cells derived from iBE mice could streamline the creation of targeted mutations ex vivo, we generated organoids from small intestine, pancreas, and basal cells from the trachea of iBE het mice.…”

“…Organotypic cell culture models or 'organoids' are a powerful system to study epithelial biology. We and others have used organoids to reveal the contribution of cancer-associated nonsense and missense mutations for cell behavior and drug response 9,35,36 ; however, efficient introduction of such mutations using base editing is a substantial practical challenge in scaling up the generation of large collections of tailored model systems.…”

“…Cancer cells often harbor thousands of single base pair substitutions 3 and understanding the impact of specific variants is critical for defining disease drivers and highlighting therapeutic vulnerabilities. While many model systems rely on gene 'knockout' or overexpression studies to interrogate the role of specific genes in disease states, ample evidence suggests that individual single nucleotide variants (SNVs), even within the same gene [4][5][6][7] or codon 8,9 , can dictate unique cancer phenotypes and response to targeted therapies.…”