Structure-Based Design and Synthesis of Stapled 10Panx1 Analogues for Use in Cardiovascular Inflammatory Diseases.

Following a rational design, a series of macrocyclic ("stapled") peptidomimetics of 10Panx1, the most established peptide inhibitor of Pannexin1 (Panx1) channels, were developed and synthesized. Two macrocyclic analogues SBL-PX1-42 and SBL-PX1-44 outperformed the linear native peptide. During in vitro adenosine triphosphate (ATP) release and Yo-Pro-1 uptake assays in a Panx1-expressing tumor cell line, both compounds were revealed to be promising bidirectional inhibitors of Panx1 channel function, able to induce a two-fold inhibition, as compared to the native 10Panx1 sequence. The introduction of triazole-based cross-links within the peptide backbones increased helical content and enhanced in vitro proteolytic stability in human plasma (>30-fold longer half-lives, compared to 10Panx1). In adhesion assays, a "double-stapled" peptide, SBL-PX1-206 inhibited ATP release from endothelial cells, thereby efficiently reducing THP-1 monocyte adhesion to a TNF-α-activated endothelial monolayer and making it a promising candidate for future in vivo investigations in animal models of cardiovascular inflammatory disease.

Pannexin1 channels-a potential therapeutic target in inflammation.

An exaggerated inflammatory response is the hallmark of a plethora of disorders. ATP is a central signaling molecule that orchestrates the initiation and resolution of the inflammatory response by enhancing activation of the inflammasome, leukocyte recruitment and activation of T cells. ATP can be released from cells through pannexin (Panx) channels, a family of glycoproteins consisting of three members, Panx1, Panx2, and Panx3. Panx1 is ubiquitously expressed and forms heptameric channels in the plasma membrane mediating paracrine and autocrine signaling. Besides their involvement in the inflammatory response, Panx1 channels have been shown to contribute to different modes of cell death (i.e., pyroptosis, necrosis and apoptosis). Both genetic ablation and pharmacological inhibition of Panx1 channels decrease inflammation in vivo and contribute to a better outcome in several animal models of inflammatory disease involving various organs, including the brain, lung, kidney and heart. Up to date, several molecules have been identified to inhibit Panx1 channels, for instance probenecid (Pbn), mefloquine (Mfq), flufenamic acid (FFA), carbenoxolone (Cbx) or mimetic peptides like 10Panx1. Unfortunately, the vast majority of these compounds lack specificity and/or serum stability, which limits their application. The recent availability of detailed structural information on the Panx1 channel from cryo-electron microscopy studies may open up innovative approaches to acquire new classes of synthetic Panx1 channel blockers with high target specificity. Selective inhibition of Panx1 channels may not only limit acute inflammatory responses but may also prove useful in chronic inflammatory diseases, thereby improving human health. Here, we reviewed the current knowledge on the role of Panx1 in the initiation and resolution of the inflammatory response, we summarized the effects of Panx1 inhibition in inflammatory pathologies and recapitulate current Panx1 channel pharmacology with an outlook towards future approaches.