Synthesis and computational studies of highly selective inhibitors of human recombinant tissue non-specific alkaline phosphatase (h-TNAP): A therapeutic target against vascular calcification

In this study, we have discovered small druglike molecules as selective inhibitors of human tissue-nonspecific alkaline phosphatase (h-TNAP), an enzyme critical for the regulation of extracellular matrix calcification. The upregulation of h-TNAP is associated with various pathologies particularly the vascular calcification (VC). Selective inhibition of h-TNAP over h-NPP1 may serve as a useful therapeutic strategy against vascular calci-fication. A series of novel triazolyl pyrazole derivatives (10a-y) in which thiol bearing triazole moiety as the zinc binding functional group was introduced to a pyrazole based pharmacophore was synthesized and evaluated as potent and selective inhibitors of h-TNAP over h-NPP1. The biological screening against h-TNAP, h-IAP, h-NPP1 and h-NPP3 showed that many of the synthesized compounds are selective inhibitors of TNAP. Particularly, the compounds 10a-h, 10j, 10m-q, 10u, 10w and 10x displayed high potency and complete selectivity towards h-TNAP over h-NPP1. Compound 10q emerged as a highly potent inhibitor (IC50 = 0.16 mu M or 160 nM) against h-TNAP with 127-fold increased inhibition compared to levamisole. On the other hand, compound 10e was found to be most selective inhibitor against the tested APs and NPPs (IC50 = 1.59 +/- 0.36 mu M). Binding sites archi-tecture analysis, molecular-docking and molecular dynamics simulations (MDS), revealed the basis for h-TNAP and h-IAP ligand selectivity as well as selectivity towards h-TNAP over h-NPP1. These newly discovered in-hibitors are believed to represent valuable lead structures to further streamline the generation of candidate compounds to target VC.