15N-, 13C- and 1H-NMR Spectroscopy Characterization and Growth Inhibitory Potency of a Combi-Molecule Synthesized by Acetylation of an Unstable Monoalkyltriazene

Abstract

6-(3-Methyltriaz-1-en-1-yl)-1H-benzo[de]isoquinoline-1,3(2H)-dione referred to as EG22 (8a), is an open-chain 3-alkyl-1,2,3-triazene termed “combi-molecule” designed to inhibit poly(adenosine diphosphate ribose) polymerase (PARP) and damage DNA. To delay its hydrolysis, acetylation of N3 was required. Being a monoalkyl-1,2,3-triazene, EG22 could assume two tautomers in solution or lose nitrogen during the reaction, thereby leading to several acetylated compounds. Instead, one compound was observed and to unequivocally assign its structure, we introduced isotopically labeled reagents in its preparation, with the purpose of incorporating 15N at N2 and 13C in the 3-methyl group. The results showed that the 1,2,3-triazene moiety remained intact, as confirmed by 15N-NMR, coupling patterns between the 15N-labeled N2 and the 13C-labeled methyl group. Furthermore, we undertook heteronuclear single quantum coherence (HSQC) and heteronuclear multiple bond correlation (HMBC) experiments that permitted the detection and assignment of all four nitrogens in 6-(3-acetyl-3-methyltriaz-1-en-1-yl)-1H-benzo[de]isoquinoline-1,3(2H)-dione, referred to as ZSM02 (9a), whose structure was further confirmed by X-ray crystallography. The structure showed a remarkable coplanarity between the N-acetyltriazene and the naphtalimide moiety. Thus, we unequivocally assigned 9a as the product of the reaction and compared its growth inhibitory activity with that of its precursor, EG22. ZSM02 exhibited identical growth inhibitory profile as EG22, suggesting that it may be a prodrug of EG22.

Keywords

• 15N-NMR
• 1,2,3-triazene
• temozolomide
• monoalkyltriazene
• 4-amino-1,8-naphthalimide