The nucleotide-sugar–activated P2Y₁₄ receptor (P2Y₁₄-R) is highly expressed in hematopoietic cells. Although the physiologic functions of this receptor remain undefined, it has been strongly implicated recently in immune and inflammatory responses. Lack of availability of receptor-selective high-affinity antagonists has impeded progress in studies of this and most of the eight nucleotide-activated P2Y receptors. A series of molecules recently were identified by Gauthier et al. that exhibited antagonist activity at the P2Y₁₄-R. We synthesized one of these molecules, a 4,7-disubstituted 2-naphthoic acid derivative (PPTN), and studied its pharmacological properties in detail. The concentration-effect curve of UDP-glucose for promoting inhibition of adenylyl cyclase in C6 glioma cells stably expressing the P2Y₁₄-R was shifted to the right in a concentration-dependent manner by PPTN. Schild analyses revealed that PPTN-mediated inhibition followed competitive kinetics, with a K_B of 434 pM observed. In contrast, 1 μM PPTN exhibited no agonist or antagonist effect at the P2Y₁, P2Y₂, P2Y₄, P2Y₆, P2Y₁₁, P2Y₁₂, or P2Y₁₃ receptors. UDP-glucose–promoted chemotaxis of differentiated HL-60 human promyelocytic leukemia cells was blocked by PPTN with a concentration dependence consistent with the K_B determined with recombinant P2Y₁₄-R. In contrast, the chemotactic response evoked by the chemoattractant peptide fMetLeuPhe was unaffected by PPTN. UDP-glucose–promoted chemotaxis of freshly isolated human neutrophils also was blocked by PPTN. In summary, this work establishes PPTN as a highly selective high-affinity antagonist of the P2Y₁₄-R that is useful for interrogating the action of this receptor in physiologic systems.