We will use structure-based ultra-large virtual screenings using VirtualFlow 2.0 [Gorgulla 2023]. The procedure will consist of four steps.
Step 1: Protein preparation. Protein structures for the TTP domain of SETDB1 will be obtained from the Protein Data Bank (PDB codes 7CJT, 8UWP, 6AU3), with the co-crystalized ligands in the complex structures removed. The protein structures will be prepared with Maestro from Schrödinger (protonation state assignment, assignment of missing atoms/side chains, hydrogen atoms, ...).
Step 2: Hit identification. The hit identification step will consist of two virtual screening stages.
Step 2a: Primary virtual screen (stage 1). We will use structure-based ultra-large virtual screenings using physics-based docking methods (e.g. QuickVina 2). We will screen a ligand library with 69 billion molecules with VirtuaFlow 2.0, an open-source platform for ultra-large virtual screens. The library we are using is the Enamine REAL Space (version 2022q12). We will use a new adaptive screening technique that we have developed, called Adaptive Target Guided Virtual Screens (ATG-VS). Due to the large-scale computations required for this approach, we will use the AWS Cloud, which is supported by VirtualFlow 2.0. We have extensive experience using the cloud and have used over 5 million CPUs in parallel in the past [Gorgulla 2023]. The protein will be held rigid in stage 1 of the screens. The ligand library that we will be using (Enamine REAL Space) has already been prepared by us into a ready-to-dock format [Gorgulla 2023]. The ligands have been protonated, and tautomerized, the 3D conformation has been computed, and the ligands are in the ready-to-dock PDBQT format.
Step 2b: Rescoring (stage 2). We will rescreen the top 1 million compounds of stage 1 in stage 2, and will allow the protein side chains at the binding site to be flexible. GWO Vina will be used for the flexible dockings.
Step 3: Postprocessing of the results. The screened compounds of Step 2 will be ranked by their docking score. Of the top 1000 compounds, biophysical and pharmacokinetic properties will be computed, visual inspection carried out, MM/GBSA binding energy calculated, all of which will be taken into account during the selection. Compounds with unfavorable properties (e.g. too high logP or PAINS motiv) will be filtered out. To ensure the novelty of the compounds, compounds with a similar scaffold as the ligands co-crystalized with the TTD of SETDB1 (PDB codes 7CJT, 8UWP, 6AU3) will be removed.
