Design,Synthesis And Biological Evaluation Of Multi-targeting Antitumor Agents And Proteolysis Targeting Chimera

Cancer has become one of the common and frequently-occurring diseases that seriously threaten human health.The occurrence and development of cancer involves dysregulation of multiple signaling pathways and the existence of compensating signaling pathways,which leads to a high incidence and mortality.Therefore,it is urgent to develop a new generation of antitumor drugs to improve the antitumor efficacy.Histone deacetylase(HDAC)is an epigenetic target associated with a variety of signaling pathways,such as p53-murine double minute 2(MDM2)interaction and bromine domain protein(BRD).Gene regulation,signal transduction and many biological processes are involved in the protein-protein interactions.and several protein-protein interactions have been confirmed as drug targets.Currently,a variety of HDAC inhibitors have been successfully marketed,achieving good clinical effects in the treatment of skin T-cell lymphoma.However,poor efficacy in solid tumors has limited clinical application of these HDAC inhibitors.HDAC inhibitors show synergistic antitumor effects when used in combination with a variety of other antitumor agents.Nevertheless,drug combination often leads to many drawbacks,such as complex dose settings,uncontrollable pharmacokinetic properties and drug-drug interactions.In contrast,multi-targeting drugs have advantages of high efficacy,more reasonable pharmacokinetic properties and low toxicity.Therefore,designing multi-targeting molecules based on HDAC can effectively solve the above disadvantages.Chemotherapy and radiotherapy are important means in cancer therapy,but the selectivity of chemotherapy and radiotherapy is poor,which can cause great toxicity and side effects.It is urgent to develop new antitumor therapy in clinic.Proteolysis targeting chimera(PROTAC)is a new technology for the research of antitumor agents.With its unique advantages,PROTAC has become a hotspot in the field of antitumor drug discovery.This thesis focuses on design,synthesis and biological activity of multi-targeting antitumor agents and PROTAC on the basis of the epigenetic targets and protein-protein interactions.1.Design,synthesis and antitumor activities evaluation of p53-MDM2/HDAC dual inhibitors.p53-MDM2 and HDAC are important targets in the development of antitumor drugs.Inspired by the synergistic effects between MDM2 and HDAC inhibitors,the first p53-MDM2/HDAC multi-target inhibitors were designed and synthetized.Most of the target compounds showed good inhibitory activity toward two targets and excellent anti-proliferation activities against a variety of cancer cells.Among them,compound B15d showed excellent and balanced inhibitory activity toward MDM2(Ki= 110 nM)and HDAC6(IC50=17.5 nM),and showed the best inhibitory activity on A549 cells.Mechanism studies had shown that compound B15d can effectively act on the two targets at the cellular level,causing apoptosis of A549 cells and blocking the G1 phase of A549 cells.The A549 xenograft model in vivo showed that the tumor growth inhibition(TGI)of compound B15d at an oral dose of 100 mg/kg was 65.4%,which was superior to that of positive agents Nutlin-3(44%)and SAHA(57.3%)at the same dose.Compound B15d had a definite dose-effect relationship.When the dosage of Compound B15d was increased to 150 mg/kg orally,the TGI reached 74.5%,and the body weight of nude mice showed no significant change.This study provides a promising lead compound for the development of novel antitumor agents.Also,this proof-of-concept study offers a novel and efficient strategy for multi-target antitumor drug discovery.2.Design,synthesis and biological evaluation of BET/HDAC dual inhibitors.HDAC and BET bromine domain protein play a key role in the regulation of histone acetylation homeostasis.Studies have shown that inhibiting HDAC and BET,thereby affecting the transcriptional of c-Myc and other genes,is a new strategy for the treatment of pancreatic cancer.Based on the synergistic effect between BET and HDAC inhibitors,novel BET/HDAC dual inhibitors were designed and synthesized by means of pharmacophore fusion.After systematic enzyme activity assay,it was found that compound C17a showed excellent balanced inhibitory activity against BRD4(1/2)and HDAC1.The subtype selectivity results showed that compound C17a was a broad-spectrum HDAC inhibitor and a highly selective BRD4(BD1)inhibitor(IC50=10.5 nM).Compound C17a had the best activity against Capan-1 cells(IC50=0.15 M).Mechanism studies confirmed that compound C17a could effectively act on two targets at the cellular level and cause down-regulation of downstream protein expression,inducing apoptosis of tumor cells.In the Capan-1 xenograft tumor model,the TGI of compound D17a(60.8%)was higher than that of(+)-JQ1(42.6%),SAHA(44.7%)and their combination(59.8%)at a dose of 15 mg/kg by intraperitoneal injection.When the dosage was increased to 20 mg/kg,the TGI of C17a reached 87.1%,and the body weight of nude mice showed no significant change.This study provides a promising lead compound for the development of novel multi-target antitumor agents and a new research area for the development of pancreatic cancer therapeutic drugs.3.Design,synthesis and biological evaluation of MDM2-based homo-PROTAC.PROTAC has become a hotspot in the field of antitumor agents,considering its various advantages in overcoming drug resistance,expanding drug targeting space and so on.Using MDM2 as the E3 ubiquitin ligase,a series of MDM2-based homo-PROTAC molecules were synthesized by rationale drug design strategy.All the target compounds showed moderate to excellent p53-MDM2 inhibitory activity.Among them,compound D10b showed the best inhibitory activity(Ki=226 nM).The results of antitumor activity in vitro showed that compound D10b had the best inhibitory activity on A549 cell line(IC50＝1.01 μM)and could effectively induce apoptosis of A549 cells.Mechanism studies confirmed that compound D10b degraded MDM2 protein in a concentration-dependent manner in A549 cells,then up-regulated the expression of p53.This study provides a foundation for the design and development of MDM2-based PROTAC drugs.