The Function Of KIF5B-RET Fusion Gene And Related Drug Interference

Purpose:Lung cancer is still the leading cause of cancer-related mortality worldwide. In recent years, targeted therapy for lung cancer has become a hot spot of research. More and more research and development of targeted molecular inhibitor has brought Long-term survival for lung cancer patients with hope. However, only specific genotypes of respective patients can benefit from targeted drug therapy. In order to enable more patients to benefit from targeted therapy for NSCLC (non-small cell lung cancer), distinguishing patient population for molecular subtypes is needed. In 2012, one study found thatthere is a new KIF5B-RET fusion gene in lung cancer. Positive patients exist mostly in little or never smoking adenocarcinoma patients. It is mutually exclusive with other known genetic changes such as EGFR, K-Ras, EML4-ALK. This is suggesting that KIF5B-RET is a new oncogenic driver mutation, and it may become a new molecular target for individualized diagnosis and treatment KIF5B-RET fusion gene functional studies is not yet in-depth, and its gene function needs to be further explored. This study applied lentivirus infection method to build a stable KIF5B-RET fusion gene-positive cell lines, and to explore KIF5B-RET fusion gene function in proliferation, migration.Methods:The KIF5B and RET gene fragments were amplified by PCR and then connected to achieve the full access of KIF5B-RET gene. Transfecting BEAS2B lung epithelial cells with lentivirus to get the stably expressing KIF5B-RET fusion gene cell line. Using western blot、CCK8、colony formation assay、transwell cell assay、Annexin V-FITC/PI double staining to explore cell proliferation migration and the apoptosis function. Detecting the protein activation of downstream signaling pathway and inhibition by Apatinib with western blot.Results:RT-PCR analysis showed that after transfecting cells by lentivirus of KIF5B-RET-PCDH (KV2 subtype) and KIF5B-RET4-PCDH (KV4 subtype)to BEAS2B cell,and the mRNA expression of two type is obvious. Western blot detection showed that the two KIF5B-RET fusion protein significantly expressed indicating BEAS2B-KIF5B-RET tool cells successfully constructed. Two experimental groups under normal culture conditions have no effect on cell proliferation, but proliferation case in less (5%) serum concentrations circumstances are different. The migration capability between experimental group and the control group shows the difference, less serum and adding cisplatin induce resistance to apoptosis. Western blot shows that multi-target tyrosine kinase inhibitor Apatinib can inhibit KIF5B-RET protein and the activation of downstream signaling pathways.Conclusion:1. KIF5B-RET fusion gene shows the function of resistance to apoptosis.2. KIF5B-RET fusion gene could promote cell migration.3. Apatinib can inhibit the activation of KIF5B-RET protein and the downstream singaling protein.