DOI: 10.5061/dryad.44j0zpcm8
title: DNDI-6174, a preclinical candidate for visceral leishmaniasis that targets the cytochrome bc1 complex – Figures 3 and S2-8 raw data

description: New drugs for visceral leishmaniasis that are safe, low cost and adapted
 to the field are urgently required. Despite concerted efforts over the
 last several years, the number of new chemical entities with novel
 mechanisms of action that are suitable for clinical development remains
 low. Here, we describe the development of DNDI-6174, an inhibitor of
 Leishmania cytochrome b that originated from a phenotypically-identified
 pyrrolopyrimidine series. This compound fulfills all Target Candidate
 Profile criteria required for progression into preclinical development. In
 addition to good metabolic stability and pharmacokinetic properties,
 DNDI-6174 demonstrates potent in vitro activity against a variety of
 Leishmania species and can reduce parasite burden in animal models of
 infection, with potential for sterile cure. No significant flags were
 identified in preliminary safety studies, including an exploratory 14-day
 toxicology study in the rat. DNDI-6174 represents the first cytochrome b
 inhibitor to enter preclinical development for visceral leishmaniasis.
description: Molecular modelling Ligand and enzyme
 preparation: The 3D model of DNDI-6174 was created using LigPrep
 (Schrödinger Release 2021-2, LigPrep, Schrödinger, LLC, New York, NY,
 2021), generating the most probable ionization state and tautomers at
 neutral pH (7.4 ± 0.2). Our previously reported homology model
 of<em> L. donovani</em> cytochrome b (8) was processed using
 the protein preparation module in the Schrödinger suite to provide
 starting points for subsequent molecular modelling studies. New structures
 were generated incorporating each of the five mutations found to drive
 drug resistance. For every mutant, the hydrogen atoms’ positions were
 optimized using the H-bond assignment/sample water orientation tool, and
 the resulting structures were subjected to a restrained minimisation
 procedure with the OPLS3e force field. This was achieved using the
 minimisation protocol of the protein preparation module of the
 Schrödinger’s suite of software. Molecular docking:
 Molecular docking was performed using the six cytochrome b structures
 (wild-type and mutated) with ubiquinone bound in the Qi site alongside one
 conserved water. Molecular docking studies were carried out using Glide
 (Schrödinger Release 2021-2, Glide, Schrödinger, LLC, New York, NY, 2021)
 in standard precision (SP). The docking energy grids for wild-type and
 mutant cytochrome b were prepared using the default value of the protein
 atom scaling factor (1.0 Å) within a cubic box centered on the ubiquinone
 (UQ2) bound in the wild-type model. The conserved water molecule
 interacting with Phe34 was also retained. After grid generation, DNDI-6174
 was docked into cytochrome b. The number of poses entered to post-docking
 minimization was set to 10. In order to assess the validity of the docking
 protocol, UQ2 was used as a reference ligand for a redocking procedure
 with all 6 enzymes (WT and 5 mutants). Molecular
 dynamics: DNDI-6174: cytochrome b complexes generated during molecular
 docking studies underwent 100 ns molecular dynamic (MD) simulations using
 Desmond<br>(Schrödinger Release 2021-2: Desmond Molecular Dynamics
 System, D. E. Shaw Research, New York, NY, 2021. Maestro-Desmond
 Interoperability Tools, Schrödinger, New York, NY, 2021). The C-terminal
 carboxylic acid of all model proteins was capped with Nmethylamine, and
 the N-terminal position acetylated. Bilayers of
 1-palmitoyl-2-oleoyl-snglycero-3-phosphocholine (POPC) membranes were
 added to the model systems to mimic the integral membrane environment of
 cytochrome b. The obtained systems were placed in orthorhombic boxes
 filled with simple point-charge (SPC) water molecules. To ensure an
 electrically neutral system for the simulation, counter ions were added to
 the system in the form of a 0.15 M salt solution. The ions were placed 10
 Å away from the ligands as a buffer zone. Prior to running simulations,
 the systems were allowed to relax. The NPγT ensemble class was chosen, so
 that fixing the surface tension ensured that the simulation box does not
 deform significantly in the plane of the membrane while the pressure is
 applied normal to the membrane surface. The analysis of all the MD
 trajectories was performed within the Schrödinger’s simulation
 interactions diagram panel. 
description: # DNDI-6174, a preclinical candidate for visceral leishmaniasis that
 targets the cytochrome bc1 complex - Figures 3 and S2-8 raw data
 [https://doi.org/10.5061/dryad.44j0zpcm8](https://doi.org/10.5061/dryad.44j0zpcm8) ## Description of the data and file structure For Fig 3, the model structure of DNDI-6174 bound in the Qi site of L. donovani cytochrome b is available as PDB file For Fig 2S, the model structures of the mutated versions of cytochrome b are available as PDB files For Fig S3-S8, the .aef files associated to the analysis of the MD trajectories are provided