Abstract
PRSS3/mesotrypsin is an atypical isoform of trypsin, the upregulation of which has been implicated in promoting tumor progression. To date there are no mesotrypsin-selective pharmacological inhibitors which could serve as tools for deciphering the pathological role of this enzyme, and could potentially form the basis for novel therapeutic strategies targeting mesotrypsin. A virtual screen of the Natural Product Database (NPD) and Food and Drug Administration (FDA) approved Drug Database was conducted by high-throughput molecular docking utilizing crystal structures of mesotrypsin. Twelve high-scoring compounds were selected for testing based on lowest free energy docking scores, interaction with key mesotrypsin active site residues, and commercial availability. Diminazene (CID22956468), along with two similar compounds presenting the bis-benzamidine substructure, was validated as a competitive inhibitor of mesotrypsin and other human trypsin isoforms. Diminazene is the most potent small molecule inhibitor of mesotrypsin reported to date with an inhibitory constant (Ki) of 3.6±0.3 μM. Diminazene was subsequently co-crystalized with mesotrypsin and the crystal structure was solved and refined to 1.25 Å resolution. This high resolution crystal structure can now offer a foundation for structure-guided efforts to develop novel and potentially more selective mesotrypsin inhibitors based on similar molecular substructures.
Highlights
PRSS3/Mesotrypsin is one of three tryptic digestive enzymes produced and secreted from the human pancreas
To identify better small molecule inhibitors, here we describe a structure guided effort utilizing virtual screening to identify compounds with predicted activity towards mesotrypsin
Small molecule inhibitors of mesotrypsin from a structure-based docking screen formulated on the “extra-precision” XP Glide software algorithm which considers factors such as: lipophilicity, displacement of water, hydrogen bonding and electrostatic interactions, and metal ion/ligand interactions as favorable interactions, while the desolvation of polar or charged groups, restriction of motion, and the entropic cost of binding adversely affect docking score [28, 29]
Summary
PRSS3/Mesotrypsin is one of three tryptic digestive enzymes produced and secreted from the human pancreas. It possesses resistance to proteinaceous trypsin inhibitors [1,2,3]. Mesotrypsin displays differential activity from the other two human trypsin isoforms, displaying reduced capability for activating pancreatic zymogens [3], reduced activation of protease activated receptors (PARs) [4, 5], and elevated proteolytic activity towards proteinaceous trypsin inhibitors [3, 6,7,8,9,10]. Research and of Basic Energy Sciences of the United States Department of Energy, and the National Center for Research Resources of the National Institutes of Health. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript
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