The Esteblishment Of EGFR And PI3Ks High Throughput Screening

Many mutant forms of EGFR have been found in several human tumors:Brain tumors such as glioblastoma multiforme (GBM) and non-small cell lung cancers (NSCLC). The kinase domain hotspot mutation which have been revealed a second mutation T790M not only increases kinase activity, but also grants TKI acquired resistance. The study of the four subtypes of PI3Ks I type kinases and the discovery of them small molecule inhibitors are useful to alleviate the frequent occurrence acquired drug resistance mechanisms.This article used the EZ Reader drug screening platform mobility of microfluidic chip detection technology and tried to establish high-throughput screening models about the two enzymes of wild EGFR and double mutant EGFR (T790M/L858R). It can directly detect the peaks of the substrate and phosphorylated products, and quantify their conversion rates, avoid false positive. It is direct and effective. The T790M mutation makes double mutant EGFR enhanced affinity of ATP, the double mutant kinase showed better activity and more potent inhibition against small inhibitor.This article chose ADP-Glo Lipid Kinase kit to establish the high-throughput screening of PI3Ks type â…  four subtypes which have lipid reaction substrate. The reaction buffer and substrate source and concentration when designed were discrepant. We test lipid inhibitor PI-103 to verify the effectivity of the screening models. The four subtype kinase showed similar activity but the alpha subtype was more active, and also the alpha subtype has it specific cancer mutant. Relative to the other three subtypes, alpha subtype and its mutant have been more studied.