Lipid Peroxide-Mediated Oxidative Rearrangement of the Pyrazinone Carboxamide Core of Neutrophil Elastase Inhibitor AZD9819 in Blood Plasma Samples

  • Drug Metab Dispos. 2015 Oct;43(10):1441-9. doi: 10.1124/dmd.115.065136.
Chungang Gu  1 ,  Richard J Lewis  2 ,  Andrew S Wells  2 ,  Per H Svensson  2 ,  Vinayak P Hosagrahara  2 ,  Eskil Johnsson  2 ,  Gösta Hallström  2
Affiliations
  • 1. AstraZeneca, R&D Boston, Waltham, Massachusetts (C.G., V.P.H.); AstraZeneca, R&D Mölndal, Mölndal, Sweden (R.J.L., E.J.); AstraZeneca, R&D Charnwood, Loughborough, United Kingdom (R.J.L., A.S.W.); AstraZeneca, R&D Södertälje, Södertälje, Sweden (P.H.S); AstraZeneca, R&D Wilmington, Wilmington, Delaware (C.G.); AstraZeneca, R&D Lund, Lund, Sweden (E.J., G.H.); SP Process Development, Södertälje, Sweden (P.H.S.); and Applied Physical Chemistry, Royal Institute of Technology, Stockholm, Sweden (P.H.S.) [email protected].
  • 2. AstraZeneca, R&D Boston, Waltham, Massachusetts (C.G., V.P.H.); AstraZeneca, R&D Mölndal, Mölndal, Sweden (R.J.L., E.J.); AstraZeneca, R&D Charnwood, Loughborough, United Kingdom (R.J.L., A.S.W.); AstraZeneca, R&D Södertälje, Södertälje, Sweden (P.H.S); AstraZeneca, R&D Wilmington, Wilmington, Delaware (C.G.); AstraZeneca, R&D Lund, Lund, Sweden (E.J., G.H.); SP Process Development, Södertälje, Sweden (P.H.S.); and Applied Physical Chemistry, Royal Institute of Technology, Stockholm, Sweden (P.H.S.).
PMID: 26203069 DOI: 10.1124/dmd.115.065136
Abstract

This study focused on the mechanistic interpretation of ex vivo oxidation of a candidate drug in blood plasma samples. An unexpected lipid peroxide-mediated epoxidation followed by a dramatic rearrangement led to production of a five-membered oxazole derivative from the original six-membered pyrazinone-carboxamide core of a human neutrophil Elastase Inhibitor, 6-(1-(4-cyanophenyl)-1H-pyrazol-5-yl)-N-ethyl-5-methyl-3-oxo-4-(3-(trifluoromethyl)phenyl)-3,4-dihydropyrazine-2-carboxamide (AZD9819). The rearranged oxidation product 2-(1-(4-cyanophenyl)-1H-pyrazol-5-yl)-5-(N-ethylacetamido)-N-(3-(trifluoromethyl)phenyl)oxazole-4-carboxamide was characterized by accurate-mass tandem mass spectrometry fragmentations, by two-dimensional NMR and X-ray crystallography of an authentic standard, and by incorporation of an (18)O atom from molecular (18)O2 to the location predicted by our proposed mechanism. The lipid peroxide-mediated oxidation was demonstrated by using human low-density lipoprotein (LDL) in pH 7.4 phosphate buffer and by inhibiting the oxidation with ascorbic acid or l-glutathione, two Antioxidants effective in both plasma and the LDL incubation. A nucleophilic mechanism for the epoxidation of AZD9819 by lipid hydroperoxides explains the prevention of its ex vivo oxidation by acidification of the plasma samples. The discovery of the lipid peroxide-dependent oxidation of an analyte and the means of prevention could provide valuable information for biotransformation and bioanalysis.

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