Data for 'The development of DUAL BRD4 and CBP/p300 degraders from ISOX-DUAL' thesis by Anthony Edmonds (2022)

ESI related to AE Edmonds' thesis entitled 'The Development of DUAL BRD4 and CBP/p300 Degraders From ISOX-DUAL' (published 21.03.2022)Contains ¹H, ¹³C and other NMR spectra, HRMS, HPLC data and also scans of the original data.AbstractThe development of Heterobifunctional molecules in the field of targeted degradation is a hot topic, with various modalities appearing over the past 20 years, with examples such as; PROTACs, SNIPERs, dTAGs, HaloTags, AUTACS, ATTECs and LYTACs. In 2019 alone there were 107 PROTAC publications, an increase of 55 from the previous year. ISOX-DUAL is an inhibitor of both BRD4 (IC50 = 1.5 µM) and CBP/p300 (IC50 = 0.65 µM) bromodomains and, as such, is a useful chemical probe for research into epigenetics.2 The published and our in-house protocols toward this target molecule were poor yielding and not amenable to scale-up. Here, synthetic routes towards the title compound were re- investigated, and now achieves an overall yield of 42%, compared to the literature published 1%. Using literature co-crystal structures in the bromodomains of BRD4 and CBP/p300 of the, structurally similar, inhibitor BDOIA383, two solvent exposed exit vectors were discovered for potential linkage to E3 recruiters. ISOX-DUAL was then re-designed with the optimised synthetic route to afford two degrader precursors (3.07) and (3.27) which were designed through replacement of the N,N-dimethylpropylamine to a propyl carboxylic acid (3.07) and the replacement of the morpholine moiety to a piperazine (3.27). Degrader mimics (3.09, 3.29) were synthesised from these compounds and showed no loss in binding affinities to the bromodomains of BRD4 or CBP/p300. A small library of 20 ISOX-DUAL based degraders were synthesised, guided by predicted physiochemical properties. Select degraders (4.69-4.72) were subjected to cell-free ubiquitination assays, to which, confirmed induction of ubiquitination of the target. The 20 synthesised degraders were initially treated in HeLa cells and 4.68 caused a reduction in BRD4 (75%) and CBP (73%), the most potent degrader in this assay. The investigation goes on to highlight the importance of cell lines when assessing these compounds and describes a series of future experiments, which, should be performed.