Figure 4.

First-generation optimized leads and 3D-QSAR model for the hit scaffold. A, bioactivity of analogs AVG-094 and AVG-158 compared with resynthesized GRP-156784. No detectable cytotoxicity was measured for either AVG094 (blue dashed line) or AVG-158 (red dashed line). B, reduction of progeny RSV A2-L19F virus load by AVG-094 and AVG-158 determined in dose–response activity assays. Active concentrations were determined through four-parameter variable-slope regression modeling. LoD, level of detection. Symbols in A and B represent individual biological repeats (n = 3 each); error bars, S.D. C, in vitro RSV RdRp activity assay using synthetic template RNA and purified polymerase complexes conducted in the presence of AVG-094 or AVG-158. The compounds show dose-dependent block of 3′ RNA extension elongation after back-priming (back-priming extens. elong.) and de novo RNA synthesis from the promoter, but do not prevent extension by up to three nucleotides after back-priming (back-priming 3′ extens.). L_D811A is bio-inactive due to the alanine substitution in the L catalytic site and was included as assay specificity control. D, generation and testing of a 3D-QSAR model developed based on a subset of GRP-156784 analogs synthesized for chemical optimization of the scaffold. AutoGPA embedded in the MOE package was used for model building. Predicted (x axis) and experimentally measured (y axis) pIC₅₀ value correlations are shown. Values denote predictive capacity, goodness of fit, and, for the combined training and test sets, slope through the origin. E, graphical representation of the 3D-QSAR model from D. Gray shading, allowable space component of the model. Areas of steric constraints, steric freedom, electro-activity, and hydrophobic interactions are shown. Electrostatically active regions of the compound are shown in red and blue.