For donor saturating assays, Hela cells are seeded on 12-well plates and transfected with 200 ng/well of plasmid pRluc-BimL coding for BRET donor and an increasing quantity of BRET acceptor plasmids peYFP-Mcl-1 (or with pCMV-Mcl-1 as a control). Twenty-four hours after transfection, cells are trypsinized, reseeded into white flat bottom 96-well plates, and incubated for another day before measurements. A single donor/acceptor ratio (200/800) is used to carry out the drug treatment assay. After reseeding, cells are then subject to a 16 h Pyridoclax treatment. Light emission at 485 and 530 nm is measured consecutively using the Mithras fluorescence-luminescence detector LB 940 after adding the luciferase substrate, coelenterazine H, at a final concentration of 5 μM. BRET ratios are calculated^[1].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

• [1]. Gloaguen C, et al. First evidence that oligopyridines, α-helix foldamers, inhibit Mcl-1 and sensitize ovarian carcinoma cells to Bcl-xL-targeting strategies. J Med Chem. 2015 Feb 26;58(4):1644-68.

  [2]. Groo AC, et al. Comparison of 2 strategies to enhance Pyridoclax solubility: Nanoemulsion delivery system versus salt synthesis. Eur J Pharm Sci. 2017 Jan 15;97:218-226.

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