Oxidative ipso Substitution of 2,4-Difluoro-benzylphthalazines: Identification of a Rare Stable Quinone Methide and Subsequent GSH Conjugate

Abstract

ABSTRACT:In vitro metabolite identification and GSH trapping studies in human liver microsomes were conducted to understand the bioactivation potential of compound 1 [2-(6-(4-(4-(2,4-difluorobenzyl)phthalazin-1-yl)piperazin-1-yl)pyridin-3-yl)propan-2-ol], an inhibitor of the Hedgehog pathway. The results revealed the formation of a unique, stable quinone methide metabolite (M1) via ipso substitution of a fluorine atom and subsequent formation of a GSH adduct (M2). The stability of this metabolite arises from extensive resonance-stabilized conjugation of the substituted benzylphthalazine moiety. Cytochrome P450 (P450) phenotyping studies revealed that the formation of M1 and M2 were NADPH-dependent and primarily catalyzed by CYP3A4 among the studied P450 isoforms. In summary, an unusual and stable quinone methide metabolite of compound 1 was identified, and a mechanism was proposed for its formation via an oxidative ipso substitution.

Metabolic and Pharmaceutical Aspects of Fluorinated Compounds

“…The fluorination pattern of the hedgehog pathway inhibitor 452 was designed to reduce the electron density of the benzyl moiety and thereby lower its susceptibility to oxidative metabolism . However, metabolite profiling studies with 452 in HLM identified a metabolite, proposed to be the quinone methide 454 , that formed during oxidative defluorination of the benzyl moiety.…”

“…This arene oxide would rearrange to 454 , a compound that is stabilized by resonance delocalization of the lone pair of electrons on the piperazine N atom. An alternative proposal envisioned H atom abstraction by P450 from the benzylic site and subsequent oxidation of the delocalized radical . In addition to the quinone methide 454 , a GSH adduct of undefined regiochemistry, 455 , was identified along with a hydroxylated analogue of this metabolite .…”

“…323 The fluorination pattern of the hedgehog pathway inhibitor 452 was designed to reduce the electron density of the benzyl moiety and thereby lower its susceptibility to oxidative metabolism. 324 However, metabolite profiling studies with 452 in HLM identified a metabolite, proposed to be the quinone methide 454, that formed during oxidative defluorination of the benzyl moiety. Quinone methide 454 was sufficiently stable to be separated and characterized on a small scale by LC−MS, but this technique did not allow the quinone methide regiochemistry and olefin geometry to be determined (Scheme 57).…”

Die Erschließung von Wirkstoffmetaboliten durch übergangsmetallkatalysierte C‐H‐Oxidation: die Leber als Inspiration für die Synthese

“…Die Transformation genügte jedoch nicht, um die beschriebene Situation zu verbessern. CYP‐vermittelte ipso ‐Defluorierungen laufen leicht unter Bildung von 184 und 185 ab, wie durch LC/MS und Glutathion‐Abfangstudien bestätigt wurde . Mit diesem Beispiel soll verdeutlicht werden, dass der Ersatz eines Wasserstoffatoms durch ein Halogenatom mitnichten eine unfehlbare Strategie ist, um die Ausscheidung zu verringern oder um die Bildung reaktiver Zwischenprodukte zu vermeiden.…”

Formation of A Novel Purine Metabolite through CYP3A4 Bioactivation and Glutathione Conjugation

“…In most cases, the formation of glutathione-S-conjugates results in cellular detoxification of free radical intermediates, however, in some instance glutathione-S-conjugates can result in bioactivation leading to generation of potentially toxic metabolites [ 10 ]. Several groups have reported novel sites of glutathione conjugations in diaminothiophene [ 11 ] via stable quinone methide intermediates [ 12 ].…”