Design, Synthesis, and Biological Activity of Hybrid Compounds between Uramustine and DNA Minor Groove Binder Distamycin A

The design, synthesis, characterization, DNA binding properties, and cytotoxic activity of a novel series of hybrids, namely, a molecular combination of the natural antibiotic distamycin A and the antineoplastic agent uramustine, are reported, and the structure−activity relationships are discussed. This homologous series 29 − 34 consisted of the minor groove binder distamycin A joined to uramustine (uracil mustard) by suitable aliphatic carboxylic acid moieties containing a flexible polymethylene chain that is variable in length [(CH2) n , where n = 1−6). All the hybrid compounds in this series exhibit enhanced activity compared to both distamycin A and uramustine derivatives 22 − 27 used for conjugation, giving IC50 values in the range 7.26−0.07 μM following a 1 h exposure of human leukemic K562 cells, with maximal activity shown when n = 6. The distance between the uramustine and distamycin frame is crucial for the cytotoxicity, with compounds having linker lengths of four to six being at least 20-fold more cytotoxic than liker lengths one to three. Taq polymerase stop experiments demonstrated selective covalent binding of uramustine−distamycin hybrids to A/T rich DNA sequences, which was again more efficent with compounds 32 − 34 with a longer linker length. Two consequences can be derived from our study: (a) the distamycin moiety directs binding to the minor groove of A/T rich DNA sequences and, consequently, is responsible for the alkylation regioselectivity found in footprinting studies; (b) the higher flexibility due to a longer linker between the distamycin and uracil moieties allows the formation of complexes with the mustard moiety situated more deeply in the minor groove and, hence, with better alkylating properties.