Phenotypic characterization of the novel, non-hotspot oncogenic KRAS mutants E31D and E63K.

ABSTRACT: KRAS proto-oncogene, GTPase (KRAS) functions as a molecular switch at the apex of multiple signaling pathways controlling cell proliferation, differentiation, migration, and survival. Canonical KRAS mutants, such as those in codons 12 and 13, produce constitutively active oncoproteins that short-circuit epidermal growth factor receptor (EGFR)-initiated signaling, resulting in dysregulated downstream effectors associated with cellular transformation. Therefore, anti-EGFR therapy provides little to no clinical benefit to patients with activating KRAS mutations. Current genotyping procedures based on canonical mutation detection only account for ~40% of non-responders, highlighting the need to identify additional predictive biomarkers. In the present study, two novel non-hotspot KRAS mutations were functionally characterized in vitro: KRAS E31D was identified from a genetic screen of colorectal cancer specimens at the UP-National Institutes of Health. KRAS E63K is curated in the Catalogue of Somatic Mutations in Cancer database. Similar to the canonical mutants KRAS G12D and KRAS G13D, NIH3T3 cells overexpressing KRAS E31D and KRAS E63K showed altered morphology and were characteristically smaller, rounder, and highly refractile compared with their non-transformed counterparts. Filamentous actin staining also indicated cytoplasmic shrinkage, membrane ruffling, and formation of pseudopod protrusions. Further, they displayed higher proliferative rates and higher migratory rates in scratch wound assays compared with negative controls. These empirical findings suggest the activating impact of the novel KRAS mutations, which may contribute to resistance to anti-EGFR therapy. Complementary studies to elucidate the molecular mechanisms underlying the transforming effect of the rare mutants are required. In parallel, their oncogenic capacity in vivo should also be investigated.

SUBMITTER: Angeles AKJ 

PROVIDER: S-EPMC6540134 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature