title: Enediynyl peptides and iso-coumarinyl methyl sulfones as inhibitors of proprotein convertases PCSK8/SKI-1/S1P and PCSK4/PC4: Design, synthesis and biological evaluations.
abstract: The proprotein convertases PCSK8 and PCSK4 are, respectively, the 8th and 4th members of Ca(+2)-dependent serine endoprotease of Proprotein Convertase Subtilisin Kexin (PCSK) super family structurally related to the bacterial subtilisin and yeast kexin. The membrane bound PCSK8 (also called SKI-1 or S1P) is implicated in sterol regulation and lipid synthesis via its role in the maturation of human (h) SREBP-2. It also plays role in cartilage formation, bone mineralization, as well as viral pathogenesis. On the other hand, PCSK4 has been linked to mammalian fertilization and placenta growth. Owing to these findings, interest has grown to develop specific inhibitors against these enzymes for potential biochemical and therapeutic applications. In this study we developed two types of small molecule inhibitors of PCSK8 and PCSK4 and demonstrated their anti-proteolytic activities in vitro cell-free and in vitro cell culture systems. These are isocoumarinyl methyl sulfone derivatives and enediyne amino acid containing peptides. Our in vitro data suggested that one of the 7 sulfone derivatives (methyl phenyl sulfone) inhibited PCSK8 with inhibition constant Ki ∼255μM. It also blocked PCSK8-mediated processing of hSREBP-2 in HepG2 cell in a concentration-dependent manner. However all 7 iso-coumarinyl methyl sulfones inhibited htrypsin with IC50 ranging from 2 to 165μM. In contrast, all our designed enediynyl peptides inhibited PCSK8 and PCSK4 activity with Ki and IC50 in low μM or high nM ranges. All compounds exhibited competitive inhibition as indicated by their enzyme kinetic plots and observed dependence of IC50 value on substrate concentration. Our study confirmed that incorporation at the substrate cleavage site of 'Enediyne amino acid' generates potent inhibitors of PCSK8 and PCSK4. This represents a novel approach for future development of inhibitors of PCSK or other enzymes.
authors.affiliation: Interdisciplinary School of Health Sciences Unit, Faculty of Health Science, U Ottawa, 451 Smyth Road, Ottawa, ON K1H 8M5, Canada; Chronic Disease Program, Ottawa Hospital Research Institute, U Ottawa, Canada. Electronic address: abasa2@uottawa.ca.
authors.forename: Ajoy
authors.initials: A
authors.lastname: Basak
authors.affiliation: Interdisciplinary School of Health Sciences Unit, Faculty of Health Science, U Ottawa, 451 Smyth Road, Ottawa, ON K1H 8M5, Canada; Centre for Advanced Research in Environmental Genomics, Department of Biology, U Ottawa, Canada.
authors.forename: Mukunda
authors.initials: M
authors.lastname: Goswami
authors.affiliation: Interdisciplinary School of Health Sciences Unit, Faculty of Health Science, U Ottawa, 451 Smyth Road, Ottawa, ON K1H 8M5, Canada.
authors.forename: Abishankari
authors.initials: A
authors.lastname: Rajkumar
authors.affiliation: Department of Chemistry, Indian Institute of Technology, Kharagpur, W Bengal, India.
authors.forename: Tapobrata
authors.initials: T
authors.lastname: Mitra
authors.affiliation: Interdisciplinary School of Health Sciences Unit, Faculty of Health Science, U Ottawa, 451 Smyth Road, Ottawa, ON K1H 8M5, Canada; Chemistry Department, Tripura University, Suryamaninagar 799022, India.
authors.forename: Swapan
authors.initials: S
authors.lastname: Majumdar
authors.affiliation: Interdisciplinary School of Health Sciences Unit, Faculty of Health Science, U Ottawa, 451 Smyth Road, Ottawa, ON K1H 8M5, Canada.
authors.forename: Paul
authors.initials: P
authors.lastname: O'Reilly
authors.affiliation: Chemistry Department, Taibah University, Saudi Arabia.
authors.forename: Hussam M
authors.initials: HM
authors.lastname: Bdour
authors.affiliation: Centre for Advanced Research in Environmental Genomics, Department of Biology, U Ottawa, Canada.
authors.forename: Vance L
authors.initials: VL
authors.lastname: Trudeau
authors.affiliation: Department of Chemistry, Indian Institute of Technology, Kharagpur, W Bengal, India.
authors.forename: Amit
authors.initials: A
authors.lastname: Basak
mesh_terms: D003374:Coumarins / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D004121:Dimethyl Sulfoxide / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology*; D015195:Drug Design; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000138:chemical synthesis / Q000494:pharmacology; D056945:Hep G2 Cells; D006801:Humans; D020128:Inhibitory Concentration 50; D015394:Molecular Structure; D009842:Oligopeptides / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology*; D043484:Proprotein Convertases / Q000037:antagonists & inhibitors*; D013450:Sulfones / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology*
chemical_list: D003374:Coumarins; D004791:Enzyme Inhibitors; D009842:Oligopeptides; D013450:Sulfones; C025910:dimethyl sulfone; D043484:Proprotein Convertases; D004121:Dimethyl Sulfoxide
keywords: Enediynyl peptides; Fluorogenic substrate; Iso-coumarinyl methyl sulfones; Proprotein Convertase Subtilisin Kexin 4 and 8; Protease inhibitors