Design, Synthesis And Biological Evaluation Of Peptidvl Epoxyketone And Substituted Benzamide Boronic Acid Derivatives As Proteasome Inhibitors

Malignant tumor has become the number-one cause of death of our country dwellers.So,it is of important practical meanings to develop effective,targeted anti-cancer agents with fewer side effects to cope with the increasingly grim cancer threat.Accumulated evidence illustrates that various regulatory proteins associated with cell growth,differentiation,and apoptosis in cancer cells are controlled by the proteasome.In addition,inhibition of proteasome leads to a dysregulation of some critical substrate proteins and results in cell cycle arrest and apoptosis.Herein,proteasome is closely related with tumorigenesis and progression,and proteasome inhibitors could be used in cancer treatment by causing cell apoptosis.To date,various proteasome inhibitors have been identified and developed,and a few of them are extensively evaluated either approved or in clinical trials with possibilities for clinical applications.Enlightened by the structural features of proteasome inhibitors and their interaction mode with proteasome,retaining the epoxyketone fragment of carfilzomib and introducing a six-membered ring in the peptidyl skeleton to improve their stability against various peptidase,47 di-and tripeptidyl target compounds were designed and synthesized.The proteasome inhibitory activities indicated that tripeptides were much more potent than dipeptides,and 14 tripeptides with IC50 values less than 10nM.Further anti-proliferation activities against MM cell lines and proteasome inhibitory activities in blood cells suggested that 6 compounds 1-218,1-223,1-236～1-237 and 1-239～1-240 had strong inhibitory activities comparable to that of carfilzomib,and all the 6 compounds potently inhibit the blood and tissue proteasome in ICR and NOD/SCID mice.Based on the above mentioned in vitro and in vivo evaluations,compounds 1-236 and 1-239 were selected and evaluated in MM RPMI 8226 xenograft in nude mice,and both compounds,especially compound 1-236(65.61%),could significantly inhibit the tumor growth,which has the potential in anti-cancer treatment.In addition,the PK studies indicated that compound 1-236 has a longer half life than carfilzomib,which might be helpful for its long term effect.In order to obtain more kinds of epoxyketone peptidyl proteasome inhibitors,we tried to make further modifications in peptidyl skeleton.It is reported that adding an extra backbone carbon by introducing α β-amino acid building block could improve the biological activity and enzymatic stability of the compound,and many natural products have β-amino acid building block.57 di-and tripeptides constructed from β-amino acid were designed and synthesized based on the previous studies.Preliminary SAR study revealed that tripeptides are much more potent than dipeptides,and the location of theβ-amino acid is critical for their activities.Compounds with β-amino acid-Phe-Leu skeleton are the most potent ones among all the compounds.Selected compounds were screened for their anti-proliferation activities against two MM cell lines,and the results were consistent with the proteasome inhibitory activities.Among them,compound 2-104 was the most potent one,and the western blot analysis of this compound suggested that proteasome inhibition caused an accumulation of poly-ubiquitin and cleavage of PARP,which resulted in cell apoptosis,thus explaining the cell growth inhibitory effects.All the biological evaluations indicated that compound 2-104 is a promising proteasome inhibitor for cancer treatment.Besides,according to the binding modes of compounds bortezomib and PI-083 with proteasome,a new series of compounds with retained boronic acid substituted leucine group of bortezomib and non-peptide skeleton of PI-083 were designed and synthesized using rational drug design strategies,which combine the structure features of this two compounds.The proteasome inhibitory activities indicated that compounds 3-47 and 3-59 were the most potent compounds,with IC50 values of 161.90±29.46 nM and 394.68±80.62 nM,respectively.However,both compounds exhibited weak anti-proliferation activities,which need to be further investigated.