Design, Synthesis And Biological Activity Of Novel VCP/p97 Small Molecule Inhibitors

The valine-containing protein(VCP/p97)is a member of the adenosine triphosphate family(AAA)and is involved in a variety of cellular activities.It can not only maintain protein homeostasis in the body,but also mediate the degradation of misfolded peptides through the ubiquitin-proteasome system(UPS).In this paper,a series of novel p97 small molecule inhibitors with pyrimidine as the core structure were designed and synthesized,and their biological evaluation was carried out.Based on the results of the enzymatic reaction,the structure-activity relationship of the synthesized compounds was discussed in detail.Among the reported small molecule inhibitors of p97,CB-5083 is the compound with the best activity and the highest specificity so far,but due to its high toxicity,clinical trials have been stopped.In this thesis,CB-5083 was used as the lead compound,a boronic acid group was introduced at the ortho position of the benzene ring,and a series of small molecule inhibitors of p97 with the pyrimidine ring as the core structure were designed and synthesized.The specific research results and content are as follows:(1)Starting from 3-nitroaniline,ethyl 2-oxocyclopentane-1-carboxylate,ethyl 2-oxocyclohexane-1-carboxylate,and hydroxybenzamide,through condensation,reduction,ring formation,chlorination,substitution,coupling and other reaction methods to synthesize the 16a-16e parent structure.Then use EDCI as the condensing agent,HOBT as the catalyst or DCC as the catalyst to carry out the amide condensation reaction of different carboxylic acid compounds.The substitution reaction of triethylamine and different acid chloride compounds finally obtains 22 new structure target compounds.And carry out HRMS?¹³C-NMR and ¹H-NMR structure identification.(2)Use the enzyme inhibition method to test the enzyme inhibitory activity of the obtained compound.The test results show that most of the enzymatic activity data of the target compounds are at the level of 200n M,of which the enzyme inhibitory activities of compounds 17,20,26,28,37 are below 50n M,and their IC₅₀ is close to CB-5083.Subsequently,we will further determine the cytotoxicity of non-small cell lung cancer(NSCLC)A549 and multiple myeloma(MM)RPMI8226 for compounds with enzyme activity data less than 200 n M.Among the inhibitors screened,compound 17(IC50,54.7 n M)showed good enzymatic activity.Studies on the cell activity of non-small cell lung cancer A549 and multiple myeloma(MM)RPMI8226 further confirmed the activity of 17,with IC₅₀ values of 2.80?M and 0.86?M,respectively.Compound17 is currently being used as a candidate compound to be further developed.Finally,on the basis of molecular docking,we discussed the possible binding mode of p97 inhibitor and p97 protein and the structural features that play a key role in the stability of the complex from the molecular level,and further explained the experimental results.