Discovery of an enzyme and substrate selective inhibitor of ADAM10 using an exosite-binding glycosylated substrate

Abstract

ADAM10 and ADAM17 have been shown to contribute to the acquired drug resistance of HER2-positive breast cancer in response to trastuzumab. The majority of ADAM10 and ADAM17 inhibitor development has been focused on the discovery of compounds that bind the active site zinc, however, in recent years, there has been a shift from active site to secondary substrate binding site (exosite) inhibitor discovery in order to identify non-zinc-binding molecules. In the present work a glycosylated, exosite-binding substrate of ADAM10 and ADAM17 was utilized to screen 370,276 compounds from the MLPCN collection. As a result of this uHTS effort, a selective, time-dependent, non-zinc-binding inhibitor of ADAM10 with Ki = 883 nM was discovered. This compound exhibited low cell toxicity and was able to selectively inhibit shedding of known ADAM10 substrates in several cell-based models. We hypothesize that differential glycosylation of these cognate substrates is the source of selectivity of our novel inhibitor. The data indicate that this novel inhibitor can be used as an in vitro and, potentially, in vivo, probe of ADAM10 activity. Additionally, results of the present and prior studies strongly suggest that glycosylated substrate are applicable as screening agents for discovery of selective ADAM probes and therapeutics.

Sequential modes of crosslinking tune viscoelasticity of cell-instructive hydrogels

“…MSCs on TCP upregulated monocyte chemoattractant protein-1 ( CCL2 ), galectin-9 ( GAL9 ), and TNFα-stimulated gene 6 ( TNFAIP6 ) after an initial pulse stimulation (Figure 5d and Figure S13), which was consistent with previous studies on MSC polarization to an immunomodulatory phenotype. [31] The gene expression levels of PTGS2 and TNFAIP6 were further upregulated as the stiffness (G’) was increased in the Alg hydrogels (Figure 5c–d). The levels were also plotted versus loss angle at each G’, which showed a viscoelastic-dependent increase in expression at 0.5 and 2.5 kPa G’.…”

Germination of Seeds and Seedling Growth of Amaranthus retroflexus L. Following Sublethal Exposure of Parent Plants to Herbicides

“…Here, we aim to study the carry-over effects of sublethal herbicides on the F1 generation of A. retroflexus L. as an invasive plant using parent plants in the vegetative or reproductive periods treated by atrazine or tribenuron-methyl, which are two commonly used herbicides in Chinese arable cereal crops 33 and also commonly used to control A. retroflexus in China 34, 35 . Our objectives were: (i) to determine whether there was a carry-over effect of atrazine and tribenuron-methyl on the F1 generation of A. retroflexus , (ii) to compare the germination and growth of the F1 generation of A. retroflexus treated with herbicides during different growth periods (vegetative and reproductive), and (iii) to determine if an increased sublethal dose to the parent plant increases the toxic effect on the F1 generation.…”

Clinical Implications of Compounds Designed to Inhibit ECM‐Modifying Metalloproteinases

“…In recent years there has been a shift from active site to a secondary substrate binding site (exosite) inhibitor discovery . We described enzyme‐ and substrate‐selective non‐Zn 2+ binding inhibitors of ADAM17 and ADAM10 (Figure ) which are currently being tested in several pre‐clinical disease models. Both ADAM10 (CID 3117694) and ADAM17 (compound 19 in) leads were discovered as a result of screening efforts using a glycosylated, exosite‐binding substrate based on the TNFα sequence .…”

“…We described enzyme‐ and substrate‐selective non‐Zn 2+ binding inhibitors of ADAM17 and ADAM10 (Figure ) which are currently being tested in several pre‐clinical disease models. Both ADAM10 (CID 3117694) and ADAM17 (compound 19 in) leads were discovered as a result of screening efforts using a glycosylated, exosite‐binding substrate based on the TNFα sequence . Mechanistic studies showed that both inhibitors acted via a non‐Zn 2+ binding mechanism, which explained their selectivity toward ADAM10 and ADAM17 (Table ).…”