Establish A New Gap-LCR-based Method For Detecting EGFR Mutations

EGFR gene mutation is closely to the treatment of non-small cell lung cancer,Lung cancer has become one of the most common malignant tumors, including non-small cell lung cancer accounts for more than 75% to 85%, because of toxic side effects of traditional chemotherapy, with the depth study of its pathogenesis in recent years, epidermal growth factor receptor (EGFR) as a new generation of molecular targeting therapy has more and more attention, such as inhibitors of tyrosine machine gefitinib, erlotinib has been the U.S. Food and Drug Administration (FDA) approved for the treatment of advanced non-small cell lung cancer.Clinical studies have shown that EGFR mutations is closely related to the treatment of gefitinib, erlotinib in NSCLC, patients carrying EGFR mutations is more sensitive to gefitinib and erlotinib, the overall survival and no Progression-free survival with the mutations in patients have a significant increase more than those without. Therefore, the detection of EGFR mutations in gefitinib and erlotinib for the treatment of non-small cell lung cancer is very necessary.This study established a new method of detecting EGFR mutations, according to the principle of Gap-LCR polymerization and connection, designed a pair of primers for EGFR each mutations, then complete the identification of alleles by the DNA polymerase and connection Reaction of DNA ligase. Finally, three types of detection mean to detect the mutations in the genome or not, these three methods were agarose gel electrophoresis, Qubit detection system and common oligonucleotide microarray.We chose the EGFR gene mutation hot spots, including the seven SNP mutations and the absence of two long fragment, we cloned all of mutant genes successfully, as a positive control in this method. This method has good detection sensitivity can be achieved by 5%, higher than traditional sequencing methods. In 10 cases of non-small cell lung cancer samples tested and found that 3 patients carrying EGFR mutations, the mutation frequency has consistent with the literature reported.This method has high sensitive, and does not require expensive equipment and special reagents, only ordinary PCR instrument can complete the experiment, especially agarose gel electrophoresis in general laboratory platform can be completed, Qubit detection system to facilitate fast, while the common oligonucleotide microarray can be completed high-throughput testing. This method is suitable for all levels of clinical testing, have very important significance to the achievement of individual drugs.