Fig. 3.

Identification of top hits from primary drug screen. A, Scatter plot of average peak amplitude of NAADP-evoked Ca²⁺ release in the presence of drug (25μM) from LOPAC^®1280 (left, numbered 1–1280) or Selleck (GPCR) compound library (right, numbered 1281–1534). B, results from both libraries were combined and compounds were ranked by amplitude of response from greatest inhibition (rank #1, left) to potentiation (rank #1534, right). The majority of compounds were in a range ±25% of control response (shaded box). Compounds that exhibited >80% inhibition of NAADP-evoked Ca²⁺ release were prioritized (red box) and selected for further characterization (Table 1). C, raw data from the primary screen for selected compounds – PPADS, PPNDS, thapsigargin, A23187, thio-NADP and vehicle control (DMSO). D, enlargement of red box from ‘B’ showing ranking of top eighteen hits after pruning, which displayed >80% inhibition of NAADP-evoked Ca²⁺ release. Pruned compounds were thio-NADP and A23187 (4^th and 5^th top hits), shown in grey. E, traces of NAADP-evoked Ca²⁺ release in the presence of the two top ranked candidates (#1, PF-543; #2, SKF96365; coloured lines, 25μM).