Enzymatic activation of a new antitumour drug, 5-diethylaminoethylamino-8-hydroxyimidazoacridinone, C-1311, observed after its intercalation into DNA

Abstract

The imidazoacridinone derivative, C-1311, is a new antitumour agent that exhibits strong antitumour activity against experimental colorectal cancer and has been selected for entry into clinical trial. The compound has previously been shown to have DNA non-covalent binding properties in vitro and to bind irreversibly to DNA of tumour cells. The latter effect has also been observed in a cell-free system, but only in the presence of activated enzymes. The present studies were aimed at finding out whether and in what way the enzymatic activation of C-1311 and its non-covalent binding to DNA influence or depend on each other. Enzymatic activation was performed with a model system containing horseradish peroxidase (HRP) and hydrogen peroxide (H 2O 2) and was followed by UV-VIS spectroscopy and by HPLC with UV-VIS and electrospray ionisation mass spectrometry detection. DNA non-covalent binding was studied in the cell-free system by means of an unwinding assay and UV-VIS spectroscopy. It was shown that C-1311 was oxidised by the HRP/H 2O 2 system in a manner dependent on the drug:H 2O 2 ratio. In the case of ratios of 1:3 and 1:5, the reaction gave highly reactive species that were quickly transformed into the further products p2 and p3 that were unable to intercalate into DNA. In the presence of DNA, C-1311 first intercalated into DNA and the intercalated compound was then oxidised. This oxidation was directed to only one product. Therefore, DNA seems to play the role of a “scavenger” of the reactive oxidation product(s) yielded from the intercalated drug and prevents its further deactivation. We conclude that, under the conditions studied, intercalation of C-1311 into DNA is followed by its HRP-mediated activation, giving rise to the intercalated species that might irreversibly bind to DNA. Since peroxidase-type enzymes are present in the cell nucleus, the proposed sequence of events may also be expected to take place in the cellular environment in vivo.