Supplementary material. Quinazoline-based analogue of adenine as an antiDOTe against MLLr cells: synthesis, inhibition and docking studies

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posted on 2022-03-25, 09:41 authored by Paola Barbara Arimondo,, Corentin Bon, Magdalena Barbachowska,, Nemanja Djokovic, Dusan Ruzic, Yang Si, Laura Soresinetti,, Corinne Jallet, Ambre Tafit, Ludovic Halby, Katarina Nikolic

Table S1. Enrichment percentages

Figure S1. ROC curve (depicted in blue) for ligand-based virtual screening on the DOT1L ChEMBL dataset

Figure S2. Presentation of co-crystal ligand EPZ4777 (magenta) and best three re-docked ligands in the active pocket of DOT1L (left) and important amino acid residues included in interaction with co-crystal ligand (right)

Figure S3. General formula of compounds for ligand based virtual screening campaign. Red is the part mimicking the second substrate, blue is the sugar analogue et green the adenine analogue. Named compounds A to W in Tables S1 and S2.

Table S2. Selected HIT compounds after ligand based virtual screening campaign. Compounds are from the CHEMBL database (https://www.ebi.ac.uk/chembl/) or were designed according to the general formula in Figure S1 (A to W compounds).

Table S3. Ranked compounds after molecular docking results. Active (Ki≤6500nM) and inactive (Ki≥7800nM) compounds selected from CHEMBL database are coloured in blue and yellow respectively.

Figure S4. Left. Cellular growth of MV4-11 (MLL-AF4) followed up to 15 days upon treatment with 10 µM (dark blue curve), 3.2 µM (blue curve) and 1.0 µM (light blue curve) of EPZ5676. Viable cells were counted every 3 to 4 days in the presence of EPZ5676 or DMSO (-) (black curve) and the ratio number of cells over initial population were plotted. Right. The viability was

evaluated every 3 to 4 days (dotted curves). Every data point are the results of technical duplicates..

Figure S5. Full gels used for western blot analysis.

Figure S6. Two western blot analysis of the H3K79me2 and the H3 signal of harvested histones from MV4-11 cells after 6 days treatment at the indicated concentration of EPZ5676. 500 ng of proteins per well were analyzed.

Figure S7. High content screening on the effect of the compound 1 (in light grey) and (±)-2 (in dark grey) on the methylation of seven different histone marks in osteosarcoma cancer cells U20S. Cells were treated with the compounds at 1µM concentration for 10 days. Integrated fluorescence density was normalised to counted cells and presented as a relative ratio to DMSO treated cells showed in arbitrary units (A.U.).

Figure S8. Predicted binding mode of enantiomer (3R,5S,3S,5R) of compound 2 obtained by molecular docking in the crystal structure of DOT1L with EPZ-5676 (PBD code: 4HRA). A) Comparison of the binding modes of EPZ-5676 (green sticks) and compound 2 (3R,5S,3S,5R) (cyan sticks) in the binding pocket of human DOT1L. B) 2D representation of intermolecular interactions predicted for compound 2 (3R,5S,3S,5R).