Validation of the Point-Exacct Method in Non-Small Cell Lung Carcinomas (Molecular Diagnostics and Genetics‪)‬

Point mutations in the Kirsten ras (K-ras) oncogene are one of the most common genetic alterations involved in various types of human cancer (1, 2). In lung cancer, K-ras mutations occur predominantly in codon 12 (3-7). The frequency of those alterations varies within different histological subtypes. K-ras point mutations are found in approximately 15-56% of the adenocarcinomas and to a lesser extent in other types of non-small cell lung carcinomas (NSCLCs) (3, 5, 7-14). The timing of K-ras activation in the process of lung cancer development is still unclear. Both human and animal studies have initially reported that K-ras mutations arise early (15-17), i.e., just before the occurrence of visible tumors. Compared with far more early chromosomal deletions demonstrated in heavy smokers, the K-ras mutation seems to be a late event (18, 19). Several methods have been utilized for the detection of K-ras point mutations in lung cancer. First reports on K-ras point mutation analysis used relatively insensitive methods such as allele-specific oligonucleotide hybridization or direct sequencing of amplification products (3,4,8,10,20-25). At the moment, a number of methods displaying a considerable increase in sensitivity of point mutation detection have been described. Those methods include amplification refractory mutation system (26), allele-specific amplification (27), mismatch amplification mutation assay (28), oligonucleotide ligation assay (29), ligation chain reaction (30), restriction fragment length polymorphism (31), enriched PCR (32), PCR-primer-introduced-restriction analysis with enrichment of mutant alleles (33), PCR-based cloning and hybridization (34), and point mutation detection using exonuclease amplification coupled capture technique (Point-EXACCT) (35, 36). Interestingly, in those studies, the percentage of cases positive for K-ras seemed to be dependent on the sensitivity of the different methods used for point mutation detection.