Large-scale phenotypic drug screen identifies neuroprotectants in zebrafish and mouse models of retinitis pigmentosa

(a) Statistical summary of Mtz titration assay in rho:YFP-NTR zebrafish larvae - Figure 1B. Survival effects (normalized YFP, %), 95% confidence intervals, p-values, and sample sizes (N) for each condition at 7 dpf. Student’s t-test was used to calculate p-values for each condition relative non-ablated controls (0 mM Mtz). Bonferroni correction for multiple comparisons resulted in an adjusted alpha level of 0.01 (α=0.01). Two experimental repeats were performed for each condition and data pooled across replicates (Figure 1—source data 1). (b) Paired compound p-values relative to all control conditions in Figure 7—figure supplement 1. Student’s t-test was used to calculate p-values for each paired condition relative to ablated controls (+Mtz), BMN alone control and NEC alone control for all paired conditions. Bonferroni correction for multiple comparisons resulted in an adjusted significance level of 0.003 (α=0.003). Survival effects as shown in Figure 7—figure supplement 1 are provided for context (Figure 7—figure supplement 1—source data 1). Inhibitor abbreviations: BMN, talazoparib; NEC, necrostatin-1. Other abbreviations: CI, confidence interval; Mtz, Metronidazole; NA, not applicable. (c) Paired compound p-values relative to all control conditions in Figure 10. Student’s t-test was used to calculate p-values for each paired condition relative to ablated controls (+Mtz) and relevant individual compound controls (Cmpd A, top paired compound; Cmpd B, bottom paired compound). Bonferroni correction for multiple comparisons to +Mtz control resulted in an adjusted significance level of 0.002 (α=0.002); significance level for comparisons to individual compound controls was 0.05 (α=0.05). Survival effects as shown in Figure 10 are provided for context (Figure 10—source data 1). Lead compound abbreviations: WAR, Warfarin; CLO, Cloxyquin; CPO, Ciclopirox olamine; MIC, Miconazole; ZPT, Zinc pyrithione; DHA, Dihydroartemisinin; CHL, Chloroxine; CAL, Calcimycin; SUL, Sulindac; ART, Artemesinin; COR, Cortexolone; POS, positive control. Other abbreviations: CI, confidence interval; Mtz, Metronidazole.

(a) Compounds tested as positive controls. List of 17 compounds previously reported as neuroprotectants in RP models tested for survival effects in rho:YFP-NTR zebrafish larvae using the primary screening protocol. (b) List of eliminated compounds. Compounds that were autofluorescent (precluding YFP signal detection) or lethal at the concentrations tested (10 mM to 0.625 mM). (c) List of 113 hit compounds. Hit compounds producing a SSMD score ≥1 in the primary screen ordered according to SSMD score. Drug names, concentrations producing SSMD ≥1, SSMD scores, SSMD effect types, and whether a dose-dependent trend was observed or not are shown. Yellow highlighted drugs were selected for confirmation testing. ‘'●” denotes confirmed lead compounds (source data). (d) On-label MOA for 113 hit compounds. Implicated MOA categories and subcategories are listed in order from most common to least common. The number of compounds per each category/subcategory are provided in the parentheses and compound names are listed.

Abbreviations: parp1, poly (ADP-ribose) polymerase 1; ripk1l, receptor (TNFRSF)-interacting serine-threonine kinase 1, like; casp3a: caspase 3, apoptosis-related cysteine peptidase a; casp3b: caspase 3, apoptosis-related cysteine peptidase b; tdp1, tyrosyl-DNA phosphodiesterase 1; actb1: actin, beta 1; rplp0: ribosomal protein, large, P0.