Design,Synthesis And Biological Activity Evaluation Of USP7 Inhibitors

Ubiquitin-specific proteinase 7(USP7,also known as HAUSP)is a de-ubiquitinating enzyme that is abundant in the body.USP7 can reverse ubiquitination and prevent substrate proteins from being degraded,for example,USP7 can stabilize expression of MDM2.Experimental studies have shown that USP7 can form the USP7-MDM2-p53 complex in cells.When USP7 is inhibited,its downstream protein MDM2 is degraded by ubiquitin,and the tumor suppressor p53 will increase accordingly and inhibite the proliferation of cancer cells.It is reported that USP7 is highly expressed in various cancers,such as multiple myeloma,lung cancer,cervical cancer,and prostate cancer.USP7 small molecule inhibitors have become one of the hot topics in antitumor small molecule inhibitors research.Based on literature research,this paper designed and synthesized a novel USP7 inhibitor using the USP7 inhibitor(compound SM-1)reported in the literature as a template backbone.1.Design of USP7 inhibitorsUsing compound 30 as a template compound,it was found that the following types of compounds showed good potential inhibitory activity of USP7 by means of computer simulation.(1)The 4-hydroxypiperidine ring in the template compound was replaced by 2-hydroxypropyl,and series I compounds were designed;(2)The 4-hydroxypiperidine ring in the template compound was replaced by carbonyl piperidine or carbonylpiperazine,and series II compounds were designed;(3)The 4-hydroxypiperidine ring in the template compound was replaced by 3-piperidine ring,and series Ⅲcompounds were designed.2.Synthesis and activity evaluation of USP7 inhibitors and positive compoundsAccording to the results of computational simulation,26 new compounds were designed and synthesized.Based on the need of biological activity evaluation,positive compound 30 was synthesized.The evaluation of USP7 inhibitory activity and structure-activity relationship analysis showed that after the 4-piperidine ring in the template compound was replaced respectively by 2-hydroxypropyl chain,carbonyl piperidine or carbonylpiperazine,and 3-piperidine structure,and most of the compounds showed significantly reduced inhibitory activity against USP7 and showed no or very low inhibitory activity within the measured concentration range.The best active compound Ⅳ-5 was only 40.99%at the inhibition rate of 50 μM.The above results indicate that the 4-hydroxypiperidine ring plays a key role in this class of USP7 inhibitors.3.Design,synthesis and activity evaluation of fluorescent probesAlthough no effective inhibitor was obtained,in order to further study the binding mode of compound 30 and USP7,fluorescent probe molecules were designed.In addition,it also provides a molecular basis for cell imaging detection.Positive control compound 30 was used as the ligand of USP7,and FITC was selected as the fluorophore,and a chain structure of appropriate length was selected as the linker.Finally,two fluorescent probe was synthesized and characterized its structure and the optical properties of the preliminary.Unfortunately,probe molecule have not shown USP7 inhibitory activity.In summary,twenty-eight new target compounds and two probe molecules were synthesized by computer simulation design based on template molecules.According to preliminary evaluation of USP7 inhibitory activity and structure-activity relationship analysis,it was found that 4-hydroxypiperidine ring plays a key role in this class of USP7 inhibitors.After the 4-piperidine ring in the template compound was replaced respectively by 2-hydroxypropyl chain,carbonyl piperidine or carbonylpiperazine,and 3-piperidine structure,and most of the compounds showed significantly reduced inhibitory activity against USP7 and showed no or very low inhibitory activity within the measured concentration range.The synthetic probe molecules showed good optical properties,but did not show USP7 inhibitory activity.