KRAS Codon 12 Mutations and Detection

The KRAS gene is one of the major oncogenes whose mutations can lead to different types of cancer. KRAS, along with other RAS genes, NRAS and HRAS, is a RAS gene family member in the RAS/MAPK signal pathway that passes the signal from receptor tyrosine kinases (RTKs) to regulate cell proliferation, differentiation and programmed cell death (apoptosis). RAS mutations are found in 30% of all human cancers.

KRAS Gene Mutations

The K-Ras protein performs its regulatory role in signal transduction through its GTPase activity. Important KRAS mutations abolishes the normal KRAS regulatory role in response to signal transduction and lead to constitutively active KRAS regardless of cell signals. These KRAS mutations with clinical significance include mutations at different codons, including 12 (80%), 13 (17%), 61 and 146 (1 to 4%). These mutations are found in different types of cancers including lung cancer (non-small cell lung cancer, NSCLC accounts for 85% of lung cancer), colorectal cancer (CRC, 96% of CRCs are adenocarcinoma), pancreatic cancer (94% of pancreatic cancers are exocrine tumors) and others. Among these codons, codon 12 is the most important codon that can change from glycine to several different amino acids.

Here is the KRAS G12 mutation rates in NSCLC or CRC according to my cancer genome.

• G12A: 2.39% in NSCLC and 2.12% in CRC
• G12C: 11.77% in NSCLC and 3.22% in CRC
• G12D: 4.09% in NSCLC and 12.83% in CRC
• G12R: 0.38% in NSCLC and 0.43% in CRC
• G12S: 0.5% in NSCLC and 1.84% in CRC
• G12V: 5.32% in NSCLC and 8.92% in CRC

Drug Development Targeting KRAS Mutations

Targeted therapy based on oncogene primary and secondary mutations has motivated pharmaceutical companies to develop targeted drugs such as different generations of tyrosine kinase inhibitors for different EGFR mutations. However, KRAS was thought as undruggable until recently when Amgen has developed the first KRAS drug AMG 510targeting KRAS G12C. Mirati Therapeutics is also developing a KRAS G12C drug.

Detection of KRAS Mutations

Although multiple vendors have developed commercial kits for KRAS mutation detection, mainly based on qPCR techniques. The analytical sensitivity varies dramatically from 1 to 10% VAF (Variant Allele Frequency) between the different mutations or detection kits. It is critical to detect these mutations with highly sensitive assays for early cancer detection.

We have developed the KRAS assay by adding the novel wildtype sequence of XNA(xenonucleic acids) into the qPCR assay to increase the assay sensitivity (clinical samples at 0.1 to 0.5% VAF) for detection of 1 to 3 copies of mutant allele. Our assay covers six KRAS codons 12, 13, 59, 61, 117 and 146 and can be performed by commonly used qPCR instruments (validated for Thermo Fisher QuantStudio 5, Bio-Rad CFX384 and Roche LC-480II). The assay uses either 10 ng of FFPE DNA or plasma cfDNA samples for detection of each codon mutation. We can either accommodate your needs for one codon mutation or all the six codon mutations in the whole KRAS kit.