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KRAS Gene Mutation

The KRAS oncogene is the cellular homolog of the Kirsten rat sarcoma virus gene. The KRAS gene codes for a guanosine triphosphate (GTP) binding protein involved in downstream cell signaling pathways such as MAPK, JAK-STAT and PI3K. These pathways play key roles in regulating cell proliferation, differentiation and apoptosis. KRAS protein also acts as a key signal transducer for the Epidermal Growth Factor receptor (EGFR).

The EGF cell-signaling pathway has become a major target for treatment of colon, lung and head and neck tumors with both anti-EGFR monoclonal antibody and small-molecule tyrosine kinase inhibitors. Monoclonal antibodies targeted against the EGFR have been shown to be effective treatments in patients with metastatic colon cancer when used alone or added to standard chemotherapeutic regimens.

The mutation frequency for the KRAS gene in metastatic colon cancers has been reported to be in the range of 30–55%. For positive samples, approximately 80% of the mutations occur in codon 12, with 35G>A (G12D) and 35G>T (G12V) being the most common variants; approximately 15–20% occur in codon 13 with 38G>A (G13D) being the most common variant. Less than 5% of colon cancers have mutations in other codons such as 61 and 146. The presence of a KRAS mutation is a negative predictor of response to anti-EGFR monoclonal antibodies panitumumab (Vectibix™) and cetuximab (Erbitux™) in metastatic colon cancers.

Given the strength of the data from these studies, in 2009 the American Society of Clinical Oncology (ASCO) and the National Comprehensive Cancer Network (NCCN) recommended that colorectal tumor samples be evaluated for KRAS mutations before a monoclonal antibody based therapy against EGFR be considered for a given patient. It is now standard laboratory practice to assess formalin-fixed paraffin-embedded tumor tissues from patients with metastatic colon cancer for KRAS mutation status.

KRAS mutations are detected in 10 to 22% of patients with lung cancer. Mutation frequency is dependent on cancer type (adenocarcinoma vs squamous cell carcinoma), smoking history and presence of other genetic lesions, such as EGFR mutations and ALK gene fusions. Approximately 97% of mutations are found in codons 12 and 13, with the remainder in codons 61 and 146. The presence of a KRAS mutation is associated with resistance to the small-molecule inhibitors erlotinib (Tarceva™) and gefitinib (Iressa™).

Three of the commonly used methods for KRAS mutation detection include nucleic acid sequencing, real-time PCR with melt–curve analysis and allele-specific PCR with various modes used to distinguish mutant from wild-type sequences. PCR methods have limit of detection (LOD) of 1 to 5% for detecting mutations within a background of wild-type DNA.

Specimen requirement is formalin-fixed, paraffin-embedded (FFPE) tissue block or four unstained slides and one matching H&E-stained slide. Samples with >4 mm² and ?50% tumor content are preferred. A copy of the pathology report must accompany the specimen.

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