Study On The Continuous Synthesis Method Of PROTAC Molecular Library Targeting The Degradation Of Src Homologous 2 Domain Protein Tyrosine Phosphatase(SHP2)

SHP2 is a non-receptor protein tyrosine phosphatase containing Src homology 2（SH2）encoded by PTPN11.It plays a vital role in cell signaling and participates in the process of cell proliferation,differentiation,apoptosis.Although the mechanism of action is not fully understood,SHP2 has been shown to play a role in many carcinogenic signaling cascades,including RAS-ERK,PI3K-AKT,and JAK-STAT.In recent years,many SHP2 inhibitors have been discovered,but due to the high homology of PTP family proteins and the generally positively charged characteristics of the catalytic center,the development of SHP2 inhibitors is limited by cell permeability and bioavailability.In 2016,Novartis discovered the small molecule allosteric inhibitor SHP099 of wild-type SHP2,which inhibits the activity of SHP2through the binding of allosteric sites outside the active pocket,providing new ideas for the development of SHP2 inhibitors.TNO155 is currently available,RMC-4630,JAB-3068,three SHP2 allosteric inhibitors targeting non-small cell lung cancer（NSCLC）,esophageal squamous cell carcinoma（ESCC）,head and neck squamous cell carcinoma（HNSCC）and other solid tumors entered clinical trials.However,malignant diseases such as leukemia,which are closely related to SHP2 mutations,still lack effective mutant SHP2 selective inhibitors.Protein degradation targeting chimera technology（PROTACs）is a kind of hybrid bifunctional small molecule compound（degrading agent）to draw the target protein and E3 ubiquitin ligase in the cell,and use ubiquitin-proteasome New technologies for drug development that specifically degrade target proteins by degradation pathways have unique advantages such as high efficacy and targeting traditional"non-drugable"targets.Using PROTAC technology to develop mutant SHP2 degraders can provide a research tool for elucidating the mechanism of the signaling pathway mediated by mutant SHP2.We used the bis-aromatic amide derivative inhibitor found in PTP1B as the starting point to replace one of the benzene rings with a heterocyclic ring,and designed and synthesized 11novel heterocyclic bis-aryl amide derivatives.Test it for biological activity.The activity results showed that the compound SMI-6b inhibited SHP2 with an IC₅₀ value of 2.63±0.08μM.In terms of selectivity,the compound SMI-6b was 4 times more selective for SHP2 than TCPTP,and had no inhibitory activity against PTP1B and SHP1,with good selectivity.In addition,in view of the ubiquitous amide bond in PROTAC technology,and to meet the requirements of green development of innovative drug research,we have developed the application of BEP/Et₃N system,bio-based green solventγ-valerolactone（GVL）as a solvent,The compound library continuous synthesis method for the structure-activity relationship study has continuously synthesized 30 amide compounds,providing an efficient synthesis strategy for PROTAC molecules.Then we designed and synthesized 10 degradants（P-01～P-10）with SMI-6b and SHP099as the degrader warheads,and used the last step of amide bond coupling in the synthesis process of P-01～P-10 As an opportunity,through the continuous synthesis scheme we developed for the efficient synthesis of 10 degradants,the efficiency and yield were significantly higher than conventional methods.The activity test results at the cellular level show that when SHP099 and SMI-6b are connected to Linker,the inhibitory activity against SHP2 will be reduced;in terms of PROTACs,as the length of Linker increases,the activity of the compound also increases,and thalidomide It is the best E3 ubiquitin ligase ligand and the most active compound P-04 in MV4-11 cells has an IC₅₀ of 0.83±0.06μM for SHP2 inhibition.Unfortunately,in the protein immunoblot experiment,the most active compounds P-04 and P-10 did not degrade the SHP2protein,which provided a reference for the subsequent design of PROTACs.