[3H]A-778317 [1-((R)-5-tert-Butyl-indan-1-yl)-3-isoquinolin-5-yl-urea]:: a novel, stereoselective, high-affinity antagonist is a useful Radioligand for the human transient receptor potential vanilloid-1 (TRPV1) receptor

1-((R)-5-tert-Butyl-indan-1-yl)-3-isoquinolin-5-yl-urea(A-778317) is a novel, stereoselective, competitive antagonist that potently blocks transient receptor potential vanilloid-1 (TRPV1) receptor-mediated changes in intracellular calcium concentrations (pIC(50) = 8.31 +/- 0.13). The (S)-stereoisomer, 1-((S)-5-tert-butyl-indan-1-yl)-3- isoquinolin-5-yl-urea (A-778316), is 6.8-fold less potent (pIC(50)= 7.47 +/- 0.07). A-778317 also potently blocks capsaicin and acid activation of native rat TRPV1 receptors in dorsal root ganglion neurons. A-778317 was tritiated ([H-3] A-778317; 29.3 Ci/mmol) and used to study recombinant human TRPV1 (hTRPV1) receptors expressed in Chinese ovary cells (CHO) cells. [H-3] A-778317 labeled a single class of binding sites in hTRPV1-expressing CHO cell membranes with high affinity (K-D=3.4 nM; B-max=4.0 pmol/mg protein). Specific binding of 2 nM [H-3] A-778317 to hTRPV1-expressing CHO cell membranes was reversible. The rank-order potency of TRPV1 receptor antagonists to inhibit binding of 2 nM [H-3] A-778317 correlated well with their functional potencies in blocking TRPV1 receptor activation. The present data demonstrate that A-778317 blocks polymodal activation of the TRPV1 receptor by binding to a single high-affinity binding site and that [H-3] A-778317 possesses favorable binding properties to facilitate further studies of hTRPV1 receptor pharmacology.