Abstract
Kallikrein‐related peptidase 6 (KLK6) is a secreted serine protease that belongs to the family of tissue kallikreins. Aberrant expression of KLK6 has been found in different cancers and neurodegenerative diseases, and KLK6 is currently studied as a potential target in these pathologies. We report a novel series of KLK6 inhibitors discovered in a high‐throughput screen within the European Lead Factory program. Structure‐guided design based on docking studies enabled rapid progression of a hit cluster to inhibitors with improved potency, selectivity and pharmacokinetic properties. In particular, inhibitors 32 ((5R)‐3‐(4‐carbamimidoylphenyl)‐N‐((S)‐1‐(naphthalen‐1‐yl)propyl)‐2‐oxooxazolidine‐5‐carboxamide) and 34 ((5R)‐3‐(6‐carbamimidoylpyridin‐3‐yl)‐N‐((1S)‐1‐(naphthalen‐1‐yl)propyl)‐2‐oxooxazolidine‐5‐carboxamide) have single‐digit nanomolar potency against KLK6, with over 25‐fold and 100‐fold selectivities against the closely related enzyme trypsin, respectively. The most potent compound, 32, effectively reduces KLK6‐dependent invasion of HCT116 cells. The high potency in combination with good solubility and low clearance of 32 make it a good chemical probe for KLK6 target validation in vitro and potentially in vivo.
Highlights
KLK6 was found to be secreted by the keratinocytes surrounding the tumor cells in response to stimuli from the tumor, and to act in a paracrine fashion to activate proteaseactivated receptors (PARs)-1 receptors, which are overexpressed on melanocytes
In an orthotopic colon cancer mouse model, mice injected with KLK6 positive HCT116 cells had significantly more metastases and worse survival than mice injected with shKLK6 HCT116 clones.[13]
After applying a threshold cutoff of 25 % inhibition, and discarding compounds with inherent high fluorescence (> 3 times the background), 1026 compounds were selected for dose-response curve analysis (Figure 2)
Summary
G. KLKs and matrix-metalloproteinases) are investigated as potential therapeutic drug targets due to their role in extracellular signaling via proteolysismediated production of small signaling molecules or proteolytic activation of membrane receptors.[10] KLK6 can activate PARs, and this signaling pathway has been found to be dysregulated in cutaneous malignant melanoma.[11] In this cancer, KLK6 was found to be secreted by the keratinocytes surrounding the tumor cells in response to stimuli from the tumor, and to act in a paracrine fashion to activate PAR-1 receptors, which are overexpressed on melanocytes. Given the growing interest in KLK6 as a drug target and the potential benefit of being able to control its enzymatic activity to validate current biological hypotheses, we set out to find a novel series of selective reversible KLK6 inhibitors
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