Fatty acid acylated peptide therapeutics: discovery of omega-n oxidation of the lipid chain as a novel metabolic pathway in preclinical species

• Published in: Journal of pharmaceutical and biomedical analysis 2023-04, Vol.227, p.115256-115256, Article 115256
• Main Authors: Esposito, Simone, Krick, Alain, Pasquier, Olivier, Bonche, Fabrice, Ingenito, Raffaele, Magotti, Paola, Bianchi, Elisabetta, Monteagudo, Edith, Gallo, Mariana, Cicero, Daniel Oscar, Orsatti, Laura, Veneziano, Maria, Caretti, Fulvia, Mele, Riccardo, Roversi, Daniela, Gennari, Nadia, Brasseur, Denis, Gauzy-Lazo, Laurence, Duclos, Olivier, Mauriac, Christine, Illiano, Stephane, Mallart, Sergio
• Format: Article
• Language: English
• Subjects: Animals; Cytochrome P-450 Enzyme System - metabolism; Dogs; Fatty Acids - metabolism; Haplorhini; LC-HRMS; Life Sciences; Metabolic Networks and Pathways; Metabolism; Microsomes, Liver - metabolism; NMR; Omega-n oxidation; Oxidation-Reduction; Peptide drugs; Rats; Relaxin; Stearic Acids; Swine; Swine, Miniature - metabolism
• ISSN: 0731-7085; 1873-264X
• DOI: 10.1016/j.jpba.2023.115256

We recently described C18 fatty acid acylated peptides as a new class of potent long-lasting single-chain RXFP1 agonists that displayed relaxin-like activities in vivo. Early pharmacokinetics and toxicological studies of these stearic acid acylated peptides revealed a relevant oxidative metabolism occurring in dog and minipig, and also seen at a lower extent in monkey and rat. Mass spectrometry combined to NMR spectroscopy studies revealed that the oxidation occurred, unexpectedly, on the stearic acid chain at ω-1, ω-2 and ω-3 positions. Structure-metabolism relationship studies on acylated analogues with different fatty acids lengths (C15-C20) showed that the extent of oxidation was higher with longer chains. The oxidized metabolites could be generated in vitro using liver microsomes and engineered bacterial CYPs. These systems were correlating poorly with in vivo metabolism observed across species; however, the results suggest that this biotransformation pathway might be catalyzed by some unknown CYP enzymes. [Display omitted] •ω-n oxidation is firstly reported as an in vivo pathway for acylated peptide drugs.•The oxidized metabolites are identified using combination of LC-HRMS and NMR.•Extent of omega-n oxidation is higher with longer fatty acid chains.•In vitro studies suggest that ω-n oxidation might be catalyzed by CYP enzymes.