Exosite inhibition of A Disintegrin And Metalloproteinase with Thrombospondin motif (ADAMTS)-5 by a glycoconjugated arylsulfonamide

  1. Santamaria Salvatore
  2. Doretta Cuffaro
  3. Elisa Nuti
  4. Lidia Ciccone
  5. Tiziano Tuccinardi
  6. Francesca Liva
  7. Felicia D’Andrea
  8. Rens de Groot
  9. Armando Rossello
  10. Josefin Ahnström

Reviewed by

Read the full article See related articles

Discuss this preprint

Start a discussion What are Sciety discussions?

Listed in

Log in to save this article

Abstract

ADAMTS-5 is a major protease involved in the turnover of proteoglycans such as aggrecan and versican. Its aggrecanase activity has been directly linked to the etiology of osteoarthritis (OA), identifying ADAMTS-5 as a pharmaceutical target for OA treatment. However, most existing ADAMTS-5 inhibitors target its active site and therefore suffer from poor selectivity. Here, using a novel approach, we have designed a new class of sugar-based arylsulfonamide inhibitors, which are selective for ADAMTS-5 through binding to a previously unknown substrate-binding site (exosite). Docking calculations combined with molecular dynamics simulations demonstrated that our lead compound is a cross-domain inhibitor that targets the interface of the metalloproteinase and disintegrin-like domains. Targeted mutagenesis identified disintegrin-like domain residues K532 and K533 as an exosite which is critical for substrate recognition. Furthermore, we show that this exosite acts as major determinant for inhibitor binding and, therefore, can be targeted for development of selective ADAMTS-5 inhibitors.

Article activity feed

  1. eLife

    ###Reviewer #2:

    The focus of this paper is the zinc metalloprotease ADAMTS-5. This protein has received attention as a therapeutic target for the treatment of degenerative joint diseases such as osteoarthritis. The primary effort is devoted to the development of non-zinc chelating exosite type inhibitors. The authors have previously identified exosites in hypervariable loops that are required for or proteolysis of both aggrecan and versican. Targeting these sites with the hope of selectivity is certainly a good approach. The authors used the glycosaminoglycan (GAG) feature of substrates to build in exosite affinity. To this end the authors probed ADAMTS-5 with a small library of GAG-mimetic glycoconjugated arylsulfonamides.

    With some minimal SAR, the authors were able to achieve some selectivity of ADAMTS-5 over ADAMTS-4 and some increase in potency over other inhibitors they have developed. They report IC50 values with the most potent (molecule 4b) at 9.4±2.8 µM. Some further SAR to more fully understand exosite binding (4b did not inhibit a peptide cleavage assay) did not lead to a more potent inhibitor. Further characterization of 4b inhibitory activity was carried out looking at synergism with a known zinc chelating inhibitor and some molecular docking studies. The docking studies led to experiments mutating residues that were thought to involve inhibitor binding. The results largely supported the in silico predictions.

    Overall the reported results advance the idea that selective inhibitors of ADAMTS enzyme that are not dependent on zinc coordination are possible; however, in the absence of more detailed studies of inhibition in cells and potentially in animals it is not possible to say how important and influential molecules such as those described here will be on sorting out complicated in vivo physiology. The potency reported for 4b suggests significant optimization would be needed before in vivo significance could be assessed.

  2. eLife

    ###Reviewer #1:

    This study pursues the development of ADAMTS-5 protease inhibitors by screening compounds linking a glycan (GlcNAc) with an arylsulfonamide, using click chemistry as the tether contains a triazene. ADAMTS-5 is a metalloprotease that has been implicated as a drug target for osteoarthritis. In prior work, this lab has identified exosites in ADAMTS-5 that can contribute to substrate recognition and processing. Here they identify a hybrid compound, 4b, that can block the protease activity of ADAMTS-5 with 9 µM potency. Using docking, they implicate several Lys residues that might confer interaction and show that potency of 4b is reduced with ADAMTS-5 mutants. Overall, compound 4b may bind as predicted although no additional experimental structural studies are performed to validate binding mode. While the study is a solid but limited medicinal chemistry effort, it is not felt that this manuscript will be of broad interest.

    -The compound 4b potency is still rather weak relative to other previously published agents which show sub-µM potency. 1) Biochem J paper (2016) from this lab and BBRC (2016) from a Japanese lab reported antibodies that blocked ADAMTS-5 in the low nM range and worked in human chondrocytes. 2) Thiazolidine-diones (sub µM) were reported as cell active ADAMTS-5 inhibitors (Eur J Med Chem 2014). 3) Acylthio-semicarbazides are sub-µM ADAMTS-5 inhibitors (Eur J Med Chem 2013) although also target ADAMTS-4 more weakly, and showed selectivity against other metallohydrolases.

    -Compound 4b was not used in a cell-based let alone animal model to analyze its pharmacological effects or promise. It is thus not clear how compound 4b stacks up to earlier agents. Compound 4b is a rather large compound for advancing clinically.

    -No new insight into ADAMTS-5 biological function was gained here.

  3. eLife

    ##Preprint Review

    This preprint was reviewed using eLife’s Preprint Review service, which provides public peer reviews of manuscripts posted on bioRxiv for the benefit of the authors, readers, potential readers, and others interested in our assessment of the work. This review applies only to version 1 of the manuscript.

  4. Version published to 10.1101/2020.06.12.146324 on bioRxiv