ACMA (9-amino-6-chloro-2-methoxy acridine) forms three complexes in the presence of DNA

Abstract

The interaction of ACMA (9-amino-6-chloro-2-methoxy acridine) (D) with DNA (P) has been studied by absorbance, fluorescence, circular dichroism, spectrophotometry, viscometry and unwinding electrophoresis. A T-jump kinetic study has also been undertaken. The experimental data show that, totally unlike other drugs, ACMA is able to form with DNA three complexes (PD_I, PD_II, PD_III) that differ from each other by the characteristics and extent of the binding process. The main features of PD_I fulfil the classical intercalation pattern and the formation/dissociation kinetics have been elucidated by T-jump techniques. PD_II and PD_III are also intercalated species but, in addition to the dye units lodged between base pairs, they also bear dye molecules externally bound, more in PD_III relative to PD_II. A reaction mechanism is put forward here. Comparison between absorbance, fluorescence and kinetic experiments has enabled us to determine the binding constants of the three complexes, namely (6.5 ± 1.1) × 10⁴ M⁻¹ (PD_I), (5.5 ± 1.5) × 10⁴ M⁻¹ (PD_II) and (5.7 ± 0.03) × 10⁴ M⁻¹ (PD_III). The Comet assay reveals that the ACMA binding to DNA brings about genotoxic properties. The mutagenic potential studied by the Ames test reveals that ACMA can produce frameshift and transversion/transition mutations. ACMA also is able to produce base-pair substitution in the presence of S9 mix. Moreover, the MTT assays have revealed cytotoxicity. The biological effects observed have been rationalized in light of these features.