Comparative Screening of K-ras Mutations in Colorectal Cancer and Lung Cancer Patients Using a Novel Real-Time PCR with ADx-K-ras Kit and Sanger DNA Sequencing

Abstract

We determined frequency/types of K-ras mutations in colorectal/lung cancer. ADx-K-ras kit (real-time/double-loop probe PCR) was used to detect somatic tumor gene mutations compared with Sanger DNA sequencing using 583 colorectal and 244 lung cancer paraffin-embedded clinical samples. Genomic DNA was used in both methods; mutation rates at codons 12/13 and frequency of each mutation were detected and compared. The data show that 91.4% colorectal and 59.0% lung carcinoma samples were detected conclusively by DNA sequencing, whereas 100% colorectal and lung samples were detected by ADx-K-ras kit. K-ras gene mutations were detected in 32.9-27.4% colorectal samples using kit and sequencing methods, respectively. Whereas 10.6-8.3% lung cancer samples were positively detected by kit and sequencing methods, respectively. Notably, 172/677 showed mutations and 467/677 showed wild type by both methods; 38 samples showed mutations with kit but wild type with sequencing. Mutations in colorectal samples were as follows: GGT -> GAT/codon-12 (35.1%); GGC -> GAC/codon-13 (26.6%); GGT -> GTT/codon-12 (18.2%); and GGT -> GCT/codon-12 (1.6%). Mutations in lung samples were as follows: GGT > GTT/codon-12 (40.9%) and GGT>GCT/codon-12 (4.5%). In conclusion, K-ras mutations involved 32.2% colorectal and 10.6% lung samples among this cohort. ADx-K-ras real-time PCR showed higher detection rates (P < 0.05). The kit method has good clinical applicability as it is simple, fast, less prone to contamination and hence can be used effectively and reliably for clinical screening of somatic tumor gene mutations.