Imidacrine

Dual inhibition of topoisomerase (topo) II and FLT3 kinase, as in the case of C-1311, was shown to overcome the shortcomings of using topo II inhibitors solely. In the present study, we designed and synthesized two series of pyrido-dipyrimidine- and pseudo-pyrido-acridone-containing compounds. The two series were evaluated against topo II and FLT3 as well as the HL-60 promyelocytic leukemia cell line in vitro. Compounds 6, 7, and 20 showed higher potency against topo II than the standard amsacrine (AMSA), whereas compounds 19 and 20 were stronger FLT3 inhibitors than the standard DACA. Compounds 19 and 20 showed to be dual inhibitors of both enzymes. Compounds 6, 7, 19, and 20 were more potent inhibitors of the HL-60 cell line than the standard AMSA. The results of the in vitro DNA flow cytometry analysis assay and Annexin V-FITC apoptosis analysis showed that 19 and 20 induced cell cycle arrest at the G2/M phase, significantly higher total percentage of apoptosis, and late-stage apoptosis in HL-60 cell lines than AMSA. Furthermore, 19 and 20 upregulated several apoptosis biomarkers such as p53, TNFα, caspase 3/7 and increased the Bax/Bcl-2 ratio. These results showed that 19 and 20 deserve further evaluation of their antiproliferative activities, particularly in leukemia. Molecular docking studies were performed for selected compounds against topo II and FLT3 enzymes to investigate their binding patterns. Compound 19 exerted dual fitting inside the active site of both enzymes.

The metabolism of antitumor-active 5-diethylaminoethylamino-8-hydroxyimidazoacridinone (C-1311) has been investigated widely over the last decade but some aspects of molecular mechanisms of its metabolic transformation are still not explained. In the current work, we have reported a direct and rapid analytical tool for better prediction of C-1311 metabolism which is based on electrochemistry (EC) coupled on-line with electrospray ionization mass spectrometry (ESI-MS). Simulation of the oxidative phase I metabolism of the compound was achieved in a simple electrochemical thin-layer cell consisting of three electrodes (ROXY^™, Antec Leyden, the Netherlands). We demonstrated that the formation of the products of N-dealkylation reactions can be easily simulated using purely instrumental approach. Newly reported products of oxidative transformations like hydroxylated or oxygenated derivatives become accessible. Structures of the electrochemically generated metabolites were elucidated on the basis of accurate mass ion data and tandem mass spectrometry experiments. In silico prediction of main sites of C-1311 metabolism was performed using MetaSite software. The compound was evaluated for cytochrome P450 1A2-, 3A4-, and 2D6-mediated reactions. The results obtained by EC were also compared and correlated with those of reported earlier for conventional in vitro enzymatic studies in the presence of liver microsomes and in the model peroxidase system. The in vitro experimental approach and the in silico metabolism findings showed a quite good agreement with the data from EC/ESI-MS analysis. Thus, we conclude here that the electrochemical technique provides the promising platform for the simple evaluation of drug metabolism and the reaction mechanism studies, giving first clues to the metabolic transformation of pharmaceuticals in the human body.