SAR-Guided Scoring Function and Mutational Validation Reveal the Binding Mode of CGS-8216 at the α1+/γ2– Benzodiazepine Site

The structural resolution of a bound ligand-receptor complex is a key asset to efficiently drive lead optimization in drug design. However, structural resolution of many drug targets still remains a challenging endeavor. In the absence of structural knowledge, scientists resort on structure-activity relationship (SAR) to promote compound development. In this study we incorporated ligand-based knowledge to formulate a docking scoring function which evaluates binding poses for their agreement with known SAR. We showcased this protocol by identifying the binding mode of the pyrazoloquinolinone (PQ) CGS-8216 at the benzodiazepine binding site of the GABAA receptor. Further evaluation of the final pose by molecular dynamics and free energy simulations revealed a close proximity between PQ’s pending phenyl ring and γ2D56 congruent with the low potency of carboxy-phenyl analogues. Ultimately, we introduced the γ2D56A mutation and in fact observed an 10-fold potency increase in the carboxy-phenyl analogue, provid...