Following a rational design, a series of macrocyclic ("stapled") peptidomimetics of¹⁰Panx1, the most established peptide inhibitor of Pannexin1 (Panx1) channels, were developed and synthesized. Two macrocyclic analoguesSBL-PX1-42andSBL-PX1-44outperformed the linear native peptide. Duringin vitroadenosine 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 native¹⁰Panx1 sequence. The introduction of triazole-based cross-links within the peptide backbones increased helical content and enhancedin vitroproteolytic stability in human plasma (>30-fold longer half-lives, compared to¹⁰Panx1). In adhesion assays, a "double-stapled" peptide,SBL-PX1-206inhibited 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 futurein vivoinvestigations in animal models of cardiovascular inflammatory disease.