N-(2-hydroxyethyl)-N-[2-(7-guaninyl)ethyl]amine, the putative major DNA adduct of cyclophosphamide in vitro and in vivo in the rat.

Abstract

The anti-cancer agent, cyclophosphamide, metabolises to the cytotoxic alkylating agent phosphoramide mustard, which can be dephosphoramidated to give nornitrogen mustard. A rat liver mitochondrial supernatant system was used to study the binding of [chloroethyl 3H]cyclophosphamide to DNA. The reacted DNA was acid-hydrolysed and one major adduct was identified using Sephadex G-10 chromatography, followed by HPLC, using reversed-phase or ion-exchange systems. Further studies, using [14C]guanine as reaction substrate for [chloroethyl 3H]cyclophosphamide, phosphoramide mustard or nornitrogen mustard, demonstrated the main adduct from each reaction had identical chromatographic properties in these systems. The radiolabelled ratio in the [3H]cyclophosphamide-[14C]guanine reaction demonstrated a monoadducted product. From this evidence and from 1H NMR data, the common adduct was putatively identified as a hydroxylated nornitrogen mustard adduct (N-(2-hydroxyethyl)-N-[2-(7-guaninyl)ethyl]amine). In in vivo studies, rats were injected intraperitoneally with 2.775 MBq [3H]cyclophosphamide. Total organ [3H] content and DNA binding levels were ascertained. Maximal levels of [3H] binding to DNA were seen between 1-4 hr with the highest binding levels observed in the bladder. The in vivo adduct was shown, using various HPLC systems, to co-chromatograph with the in vitro adduct and thus the main in vivo adduct was putatively identified as N-(2-hydroxyethyl)-N-[2-(7-guaninyl)ethyl]amine.

• Authors: A J Benson, C N Martin, R C Garner
• Journal: Biochemical pharmacology (Biochem Pharmacol) Vol. 37 Issue 15 Pg. 2979-85 (Aug 01 1988) ISSN: 0006-2952 [Print] England
• PMID: 3395373 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Guanine
• Cyclophosphamide
• DNA
• N-(2-hydroxyethyl)-N-(2-(7-guaninyl)ethyl)amine

Topics

• Alkylation
• Animals
• Chromatography, Gel
• Chromatography, High Pressure Liquid
• Cyclophosphamide (metabolism)
• DNA (metabolism)
• Guanine (analogs & derivatives, isolation & purification, metabolism)
• Hydrogen-Ion Concentration
• Injections, Intraperitoneal
• Kidney (metabolism)
• Magnetic Resonance Spectroscopy
• Rats
• Spectrophotometry, Ultraviolet
• Urinary Bladder (metabolism)