Study On Antitumor Activity And Mechanism Of Bromophenol Compound LXQ-217 Targeting SHP2

Cancer is the second biggest killer threatening human health.It has the characteristics of short survival period and poor quality of life.Moreover,most types of cancer lack curable means at present,which makes people turn negative about cancer.At present,the drugs for lung cancer are mostly targeted drugs,such as gefitinib.However,due to the incomplete understanding of target function and adverse drug reactions,the application of targeted drugs is still difficult.Therefore,the development of new targeted drugs with better activity and safety for specific targets is currently the focus of anti-tumor drug research and development.SHP2(SH2 domain-containing protein Tyrosine Phosphatase-2)is a non-receptor protein tyrosine phosphatase(PTP)encoded by PTPN11 gene,which is involved in RAS-ERK and PI3K-AKT cell signal transduction.SHP2 has been shown to contribute to the progression of many cancers,including lung cancer,and has become an important target for anti-tumor drug research.However,past efforts to develop SHP2 inhibitors as drugs have been unsuccessful because the positively charged nature of the active site pocket tends to bind to negatively charged groups,which make it difficult for the compound to cross the cell membrane.Our research group used structure-based strategy to design and synthesize a series of uncharged bromophenol pyrazoline derivatives.LXQ-217 showed significant SHP2 inhibitory activity(IC50=2.01 μM)by enzyme activity screening in vitro.Therefore,the antitumor activity and mechanism of LXQ-21 7 were evaluated at molecular level,cellular level and animal level.At the molecular level,the binding ability of LXQ-217 to SHP2 protein was investigated by surface plasmon resonance(SPR)experiment,and the binding mode of LXQ-217 to SHP2 protein was predicted by computer molecular docking technology.These results indicate that LXQ-217 is a potent SHP2 catalytic site inhibitor,which competitively binds to the catalytic domain of SHP2 through multiple hydrogen bonds and has strong interaction with SHP2 protein,thus showing a significant inhibitory activity against SHP2.At the cellular level,MTT assay and SRB assay were used to detect the proliferation inhibition activity of LXQ-217 on various tumor cells and normal cells.LXQ-217 exhibited strong proliferation inhibition against various tumor cells,especially against A549(human non-small cell lung cancer cell line)(IC50=2.06 μM),but low toxicity against normal HUVEC(human umbilical vein endothelial cell line).This suggests that LXQ-217 has a good safety in vitro.Plate colony formation experiments further demonstrated that LXQ-217 inhibited the colony formation of A549 cells.Cell wound scratch assay showed that LXQ-217 inhibited the migration of A549 cells.In subsequent studies,the mechanism of LXQ-217 inhibiting the proliferation of A549 cells was explored by Western blotting,Hoechst 33258 staining and CETSA method.The results showed that LXQ-217 could bind to SHP2 in cells and down-regulation of RAS-ERK and PI3K-Akt signaling pathways resulted in cell cycle arrest and apoptosis,thereby inhibiting the growth of A549 cells.At the animal level,the antitumor activity of LXQ-217 was evaluated through the subcutaneous A549 xenograft mouse model.The results showed that LXQ-217 not only inhibited the growth of A549 xenograft tumor in mice,but also decreased the phosphorylation of AKT and ERK in tumor tissue.It was noteworthy that LXQ-217 showed no significant toxic effects on body weight,relative organ weight and organs of mice in acute and subacute oral toxicity tests,indicating that LXQ-217 has a good safety in vivo.In summary,this study clarifies the antitumor mechanism of LXQ-217 targeting SHP2 in vitro and in vivo,lays the experimental foundation for further development of LXQ-217 as a more safe and effective antitumor targeted drug,and provides new ideas for the development of novel SHP2 inhibitors.