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Figure 3
Self-deconvoluting library screen. (A) The screen of plate #30328 is shown as a representative example where 80 drugs (A2-H11) were analyzed in 18 pools (A-H, 2–11). 4 pools were lethal to the embryos (A, H, 4, 7). Three pools showed a significant reduction in blood vessel numbers, but only two (G, 9) exceeded the threshold of 40% reduction in HV numbers. (B)Individually re-tested compounds from toxic pools or of pool hits from plate #30328 identified G9 as promising candidate for further testing with >40% PHV reduction. (C) Representative images from DMSO control or 10 μM G9 treated zebrafish lenses showing the GFP-positive hyaloid vessels. (D) Three independent dose curves (test1–3) of G9 drug treatments showed strong toxicity at concentrations of >5 μM and lack of reproducible antiangiogenic activity. (E) The chemical structure of and chemical name (4-(2,4-Dichlorophenoxy)-N,N-diethyl-1-butanamine) of G9. (F) Individual re-testing of all 80 compounds from plate #30325 that were toxic in pools. No single compound reduced HV >40% and 2 were lethal at 10 μM (10G, 11B). (G) Summary of library screen. Pooled drug screens were conducted for 22 plates containing 1,760 different compounds. In 63% of cases, combinations of 8 or 10 pooled drugs did not reduce HV numbers >40%. 22% of pools caused significant developmental defects or were lethal. Of the 147 compounds identified in 18 pool hits, 80% had no significant effect when tested individually, two were toxic and eight were considered hits that significantly reduce hyaloid vessel number. Data in (A), (B), (D), and (F) are means ± SD. Statistical significance calculated by t test and Mann Whitney test (*p < 0.05, **p < 0.01, ***p < 0.001).