Cellular uptake of N-[2-(dimethylamino)ethyl]acridine-4-carboxamide (DACA)

N-[2-(Dimethylamino)ethyl]acridine-4-carboxamide (DACA), a DNA intercalating dual topoisomerase I/II poison, has high experimental antitumour activity, is able to overcome several forms of multidrug resistance, and is undergoing clinical trial. We prepared 3H-labelled DACA and investigated its uptake using cultured Lewis lung carcinoma cells (LLTC), P388 leukaemia cells and P/DACT cells that were multidrug resistant. The kinetics of uptake and efflux were very rapid and equilibrium was obtained within seconds of drug addition. Fluorescence microscopy of LLTC cells treated with DACA showed punctate fluorescence in the cytoplasm, consistent with uptake into vesicles. To investigate the role of lipophilicity in drug uptake, a fluorimetric assay was developed to measure uptake of a more hydrophilic derivative, 9-amino-5-sulphonylmethyl-DACA (as-DACA). The calculated n-octanol-water partition coefficient for as-DACA was 20-fold lower than that for DACA. On the other hand, as determined by ethidium displacement from DNA, as-DACA bound DNA 16-fold more strongly than did DACA. Uptake and efflux of DACA and as-DACA were very rapid and the uptake ratios in LLTC cells were 550 for DACA and 54 for as-DACA. At equitoxic concentrations (corresponding to the IC50 values), LLTC cell association was estimated to be approximately 1.6 x 10(8) molecules per cell for DACA and 3.0 x 10(6) molecules per cell for as-DACA. It is argued that DACA binds predominantly to lipophilic sites such as proteins and cellular membranes, while as-DACA associates predominantly with DNA. The high affinity of DACA for membranes may contribute to the rapidity of its uptake and efflux, as well as to its ability to overcome multidrug resistance.