title: A simple and sensitive method for detecting major mutations within the tyrosine kinase domain of the epidermal growth factor receptor gene in non-small-cell lung carcinoma.
abstract: The detection of mutations in the epidermal growth factor receptor (EGFR) gene impacts therapeutic decision-making for non-small-cell lung carcinoma (NSCLC). Although direct sequencing has been most frequently used to detect EGFR mutations, this method displays several disadvantages. We set up simple mutation-specific polymerase chain reaction (PCR) for common delE746-A750 and L858R mutations of EGFR using primers specific to these mutations. Both mutation-specific PCR and direct sequencing methods were used to investigate 62 samples of NSCLC, and the results were compared. To evaluate the sensitivity of mutation-specific PCR, DNA mixtures containing various proportions of mutant alleles were analyzed. Mutation-specific PCR revealed delE746-A750 in 8 samples and L858R in 14 samples. All samples with either delE746-A750 or L858R mutation revealed by direct sequencing also displayed positive results for mutation-specific PCR. Conversely, mutations in 3 samples revealed by L858R-specific PCR were barely detectable by direct sequencing. In DNA mixture analysis, DNA mixtures containing 2.5% of delE746-A750 allele or 0.25% of L858R allele yielded positive results with mutation-specific PCR. Our mutation-specific PCR showed satisfactory sensitivity and reliability for detecting major EGFR mutations in clinical NSCLC samples. Given the practical availability, this method could be widely applicable to the treatment of lung cancers.
authors.affiliation: Department of Laboratory Medicine, Kyorin University, Tokyo, Japan. onishi@kyorin-u.ac.jp
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Ohnishi
authors.forename: Kouki
authors.initials: K
authors.lastname: Ohtsuka
authors.forename: Akiko
authors.initials: A
authors.lastname: Ooide
authors.forename: Satsuki
authors.initials: S
authors.lastname: Matsushima
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Goya
authors.forename: Takashi
authors.initials: T
authors.lastname: Watanabe
mesh_terms: D001483:Base Sequence; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics*; D016022:Case-Control Studies; D045744:Cell Line, Tumor; D004252:DNA Mutational Analysis / Q000379:methods / Q000706:statistics & numerical data; D017931:DNA Primers / Q000235:genetics; D004273:DNA, Neoplasm / Q000235:genetics; D066246:ErbB Receptors / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics*; D009154:Mutation; D017354:Point Mutation; D016133:Polymerase Chain Reaction / Q000379:methods / Q000706:statistics & numerical data; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000235:genetics*; D012680:Sensitivity and Specificity; D017384:Sequence Deletion
chemical_list: D017931:DNA Primers; D004273:DNA, Neoplasm; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases