Clinical and Functional Characterization of Atypical KRAS/NRAS Mutations in Metastatic Colorectal Cancer
Jonathan M Loree1, Yucai Wang2, Muddassir A Syed3
1BC Cancer, Vancouver, British Columbia, Canada.
Purpose:
Mutations in KRAS/NRAS (RAS) predict lack of anti-EGFR efficacy in metastatic colorectal cancer (mCRC). However, it is unclear if all RAS mutations have similar impact, and atypical mutations beyond those in standard guidelines exist.
Experimental Design:
We reviewed 7 tissue and 1 cell-free DNA cohorts of 9,485 patients to characterize atypical RAS variants. Using an in vitro cell-based assay (functional annotation for cancer treatment), Ba/F3 transformation, and in vivo xenograft models of transduced isogenic clones, we assessed signaling changes across mutations.
Results:
KRAS exon 2, extended RAS, and atypical RAS mutations were noted in 37.8%, 9.5%, and 1.2% of patients, respectively. Among atypical variants, KRAS L19F, Q22K, and D33E occurred at prevalence ≥0.1%, whereas no NRAS codon 117/146 and only one NRAS codon 59 mutation was noted. Atypical RAS mutations had worse overall survival than RAS/BRAF wild-type mCRC (HR, 2.90; 95% confidence interval, 1.24-6.80; P = 0.014). We functionally characterized 114 variants with the FACT assay. All KRAS exon 2 and extended RAS mutations appeared activating. Of 57 atypical RAS variants characterized, 18 (31.6%) had signaling below wild-type, 23 (40.4%) had signaling between wild-type and activating control, and 16 (28.1%) were hyperactive beyond the activating control. Ba/F3 transformation (17/18 variants) and xenograft model (7/8 variants) validation was highly concordant with FACT results, and activating atypical variants were those that occurred at highest prevalence in clinical cohorts.
Conclusions:
We provide best available evidence to guide treatment when atypical RAS variants are identified. KRAS L19F, Q22K, D33E, and T50I are more prevalent than many guideline-included RAS variants and functionally relevant.
Insights
Atypical KRAS/NRAS mutations impact survival in metastatic colorectal cancer. Some variants, like KRAS L19F, Q22K, and D33E, are common and functionally significant, guiding treatment decisions.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- RAS mutations (KRAS/NRAS) are key predictors of anti-EGFR therapy response in metastatic colorectal cancer (mCRC).
- The clinical significance of atypical RAS variants, beyond standard guideline mutations, remains incompletely understood.
Purpose of the Study:
- To characterize the prevalence and functional impact of atypical RAS variants in mCRC.
- To assess the prognostic value of these atypical mutations on patient survival.
- To provide evidence for clinical decision-making regarding treatment strategies for mCRC patients with atypical RAS variants.
Main Methods:
- Analysis of 7 tissue and 1 cell-free DNA cohorts comprising 9,485 mCRC patients.
- Functional characterization of RAS variants using an in vitro cell-based assay (FACT), Ba/F3 transformation assays, and in vivo xenograft models.
- Assessment of signaling pathway activation and correlation with clinical outcomes.
Main Results:
- Atypical RAS mutations were found in 1.2% of patients, with KRAS L19F, Q22K, and D33E being the most prevalent (≥0.1%).
- Atypical RAS mutations were associated with significantly worse overall survival compared to RAS/BRAF wild-type mCRC (HR, 2.90; P=0.014).
- Functional assays revealed that 28.1% of atypical RAS variants were hyperactive, 40.4% showed intermediate signaling, and 31.6% had reduced signaling compared to wild-type.
Conclusions:
- Atypical RAS variants, particularly KRAS L19F, Q22K, D33E, and T50I, are functionally relevant and more prevalent than some guideline-included RAS mutations.
- These findings underscore the importance of identifying atypical RAS variants for accurate prognostic assessment and personalized treatment strategies in mCRC.
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