P189 Aldehyde Oxidase Dependent Metabolite Profiling Methodologies in Early Drug Discovery Projects

Lloyd M King , UCB Biopharma, Slough, United Kingdom
Jehan Claessens , UCB Biopharma, Braine-l'Alleud, Belgium
Christelle Derwa , UCB Biopharma, Braine-l'Alleud, Belgium
Liz Jones , UCB Biopharma, Slough, United Kingdom
Anne Foley , UCB Biopharma, Slough, United Kingdom
Emre M Isin , UCB Biopharma, Braine-l'Alleud, Belgium
Nenad Manevski , UCB Biopharma, Slough, United Kingdom
Harold Mackenzie , UCB Biopharma, Slough, United Kingdom
The use of aromatic heterocycles (e.g. pyridine, pyrimidine, quinazoline) in medicinal chemistry drug design are common due to their relative resistance to CYP mediated metabolism [1]. This however renders them potential substrates to aldehyde oxidase (AO) mediated oxidation via nucleophilic attack at the electron deficient carbons. AO substrates can exhibit interspecies differences in biotransformation, making human dose predictions challenging [2] as well as introducing potential for toxicological [3] and drug-drug interaction [4] liabilities. Therefore early identification of AO involvement in the metabolism of drug candidates is important to facilitate chemical design and mitigate aforementioned issues. Comparison of microsome and hepatocyte clearance rates provides early indication of non-CYP mediated metabolism, and suspected substrates of AO are submitted for the ‘Litmus test’ reaction [5] where CHF2 groups are added to electrophilic carbons, indicating a propensity for the substrates to undergo AO mediated oxidation. LC-HRMS analysis of the litmus test reaction media can detect the addition of CF2 groups, but may not allow the unambiguous identification of the site of addition. The incorporation of fluorine in to the molecule allowed us to utilise the selectivity of heteronuclear NMR so that the sites of ‘metabolism’ are identified without any further sample preparation effort, such as preparative LC. This approach has been successfully adopted as part of our strategy for the early identification of potential AO substrates where isoquinoline and pyrido-pyrimidine scaffolds have been included. The identification of AO oxidation sites facilitated further chemical design and helped improve our understanding of the chemistry and other liabilities of AO mediated metabolism. Furthermore, this has resulted in improved lead compounds with lower metabolic clearance with a reduced role of AO mediated metabolism. Data will be presented to show examples and learnings gathered from implementation of this approach.

References:

[1] Argikar UA, Potter PM, Hutzler JM, and Marathe PH. (2016) Challenges and Opportunities with Non-CYP Enzymes Aldehyde Oxidase, Carboxylesterase, and UDP-Glucuronosyltransferase: Focus on Reaction Phenotyping and Prediction of Human Clearance. AAPS J 18:1391-1405.

[2] Rashidi MR, Soltani S. (2017) An overview of aldehyde oxidase: an enzyme of emerging importance in novel drug discovery . Expert Opinion on Drug Discovery, 12:3, 305-316

[3] Diamond S, Boer J, Maduskuie TP, Falahatpisheh N, Li Y, &Yeleswaram S. (2010) Species-Specific Metabolism of SGX523 by Aldehyde Oxidase and the Toxicological Implications. Drug Metabolism and Disposition , 38 (8) 1277-1285

[4] Li Y, Xu J, Lai WG, Whitcher-Johnstone A, & Tweedie DJ. (2012) Metabolic Switching of BILR 355 in the Presence of Ritonavir. II. Uncovering Novel Contributions by Gut Bacteria and Aldehyde Oxidase. Drug Metabolism and Disposition , 40 (6) 1130-1137

[5] O'Hara F, Burns AC, Collins MR, Dalvie D, Ornelas MA, Vaz AD, Fujiwara Y, & Baran PS. (2014) A simple litmus test for aldehyde oxidase metabolism of heteroarenes. J Med Chem 57:1616-1620.