N-7-Guanine Adduct of the Active Monoepoxide of Prodrug Treosulfan: First Synthesis, Characterization, and Decomposition Profile Under Physiological Conditions

Abstract

(2S,3S)-1,2:3,4-diepoxybutane (DEB) cross-links DNA guanines by forming the intermediate epoxy-adduct ((2′S,3′S)-N-7-(3′,4′-epoxy-2′-hydroxybut-1′-yl)guanine [EHBG]). This process is presently considered a primary mechanism for the action of treosulfan (TREO), the prodrug that transforms to DEB via the monoepoxide intermediate (2S,3S)-1,2-epoxybutane-3,4-diol 4-methanesulfonate (EBDM). In this article, the N-7-guanine adduct of EBDM ((2′S,3′S)-N-7-(2′3′-dihydroxy-4′-methylsulfonyloxybut-1′-yl)guanine [HMSBG]) was synthesized for the first time, and its stability was investigated at physiological in vitro conditions. To synthesize HMSBG, EBDM, formed in-situ from TREO, was treated with guanosine in glacial acetic acid at 60°C followed by ribose cleavage in 1 M HCl at 80°C. HMSBG was stable during the synthesis, which showed that a β-hydroxy group protects the sulfonate moiety against hydrolysis in acid environment. At pH 7.2 and 37°C, HMSBG exclusively underwent first-order epoxidation to EHBG with a half-life of 5.0 h. EHBG further decomposed to trihydroxybutyl-guanine, chlorodihydroxybutyl-guanine (major products), phosphodihydroxy-guanine, and a structural isomer (minor products). The isomeric derivative was identified as guanine with a fused 7-membered ring, which provided a new insight into the EHBG stability. To conclude, the exclusive conversion of HMSBG to EHBG indicates that EBDM might contribute to DNA cross-linking independently from DEB and play a more important role in the TREO action than expected before.