Abstract 919: Design and preclinical pharmacological evaluation of a cleavable succinate ester solubilizing group for isoindolinone MDM2-p53 protein-protein interaction inhibitors
Authors: Yan Zhao, Huw Thomas, Junfeng Liu, Timothy J. Blackburn, Sarah Cully, Anna F. Watson, Ian R. Hardcastle, Bernard T. Golding, Roger J. Griffin, John Lunec, David R. Newell
Journal: Cancer Research (2012)

Abstract The potent and selective MDM2-p53 antagonist 4-chloro-3-(4-chlorophenyl)-3-((1-(hydroxymethyl)cyclopropyl)methoxy)-2-(4-nitrobenzyl)isoindolin-1-one 1 displays promising activity in cellular tumour models; however, poor solubility precludes in vivo investigations.(Bioorg. Med. Chem. Letters 2011, 21, 5916-5919) A water-soluble succinate ester of 1 was designed as a pro-drug, based on an understanding of the interaction of the parent compound 1 with MDM2, and the cellular pharmacology and in vivo pharmacokinetics of the ester 2 were defined. Compound 2 was synthesised and evaluated as an antagonist of MDM2- and MDMX-p53 using ELISA assays. Cell growth inhibition induced by 2 and 1 were compared in paired p53 functional and p53 non-functional HCT116, A2780 and SJSA-1 cell lines. Conversion of 2 into 1 was studied in human and mouse plasma in vitro, as was uptake into tumour cells, by HPLC analysis. The pharmacokinetics of 2 was studied in vivo in tumour-bearing mice. Unexpectedly, the succinate ester 2 was 7-fold more potent than the parent compound 1 as an MDM2-p53 binding antagonist in cell-free assays (IC50 values 20 ± 13 nM versus 95 ± 96 nM, respectively) and maintained &amp;gt;100-fold selectivity for MDM2 over MDMX antagonism. In cell growth inhibition assays (GI50) 2 was 2.3-, 6.1- and 8.6-fold more potent in paired p53 functional versus non-functional HCT116, A2780 and SJSA-1 cell lines, respectively. As anticipated, the solubility of 2 (65 µM) was greatly improved over 1 (6.5 µM), and the succinate ester was converted to the parent compound in mouse plasma (&amp;gt;90% in 60 min), although interestingly not in human plasma. Following incubation of tumour cell lines with 2, both the pro-drug and parent compound could be detected. Administration of 2 to A2780 tumour-bearing mice i.v. at 10 mg/kg generated plasma concentrations approaching those required for cell growth inhibition (≥ 3 μM), and oral administration at 100 mg/kg resulted in 2 bio-availability of 7%, and an 1 AUC that was estimated to be 21% of the equivalent i.v. dose of 2. However, a major unidentified metabolite was observed which compromised the in vivo utility of the succinate ester 2. Protein-protein interaction inhibitors as exemplified by MDM2-p53 binding antagonists are a major new class of antitumour agent. However, targeting hydrophobic protein-protein interactions can result in molecules with poor physicochemical properties. A cleavable succinate ester approach can be applied to address this problem without compromising target interaction or cellular activity, as in the current study. Although additional metabolite pathways may limit the utility of this approach, the improved potency gained by addition of a solubilising group provides valuable information for further lead optimisation. (This work was supported by the CR-UK) Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 919. doi:1538-7445.AM2012-919