Abstract 131: Identification Of A Molecule To Modulate A Platelet Olfactory Receptor And Limit Thrombosis

Background and objective: Olfactory Receptors (ORs) are G protein-coupled receptors (GPCR) we recently discovered in human platelets. OR2L13 activation by an olfactory ligand Levo-Carvone (L-Carvone) limits platelet activation. Our goal was to better characterize the pharmacology of OR2L13 ligands through structural modifications of L-Carvone and by using an 8K bioactive non-olfactory ligand library and deciphering their role as anti-thrombotic. Methods and Results: With an IC 50 of 58 μM, 2-tert-Butyl-5-Methylphenol was the best odorant for inhibiting ADP-mediated platelet activation (Fig. 1). 169 molecular hits >25% threshold in an OR2L13-expressiong reporter cell line for cyclic AMP (cAMP) production were found. By counter screen (cell line without OR2L13 receptor), 12 ligands were identified for OR2L13 and 6 (CCF0054500, CCF0054432, CCF0053070, CCF0052249, CCF0051970, CCF0058399) made endogenous cAMP. Healthy platelet rich plasma (PRP) (n=7) was incubated with OR2L13 ligand for 30 minutes, then subjected to Light Transmission Aggregometry (LTA). 3/6 ligands (CCF0054500, CCF0052249 and CCF0058399) inhibited platelet aggregation through the P2Y 12 receptor and so were further validated by FACS using washed healthy platelets (n=8, attenuation of agonist-activated platelets surface P-selectin expression) for the following receptors: ADP, TRAP-6, U46619, and CRP. CCF0054500 suppressed platelet activation and aggregation through P2Y 12, PAR-1, Thromboxane and GPVI receptors. Whole blood (n=5) traversing collagen-coated capillaries reduced thrombosis when compared to vehicle under arterial sheer stress with CCF0054500. Conclusions: Like neurons, platelets have a fully functional OR signal transduction cascade. Platelet OR2L13 operates as a negative regulator of platelet reactivity and thrombosis in response to odorant and non-odorant surface agonists (biochemical activation) and under shear stress (biomechanical activation).