Design, synthesis, and biological evaluation of DNA sequence and minor groove selective alkylating agents

Abstract

The syntheses of oligoimidazolecarboxamido analogues of distamycin wherein the N-terminus contains either a benzyl-mustard 8 or chlorambucil moiety 9-11 are reported. Based on data from an ethidium displacement assay and CD studies, these compounds, along with the N-benzoyl mustards 6 and 7, were shown to have increased acceptance of GC-rich sequences over distamycin. Compounds 8-11 which contain an electron-donating group (σ<0) para to the bischloroethylamino moiety, were found to have significantly enhanced reactivity with DNA compared to the benzoyl mustards 6 and 7. Through dialysis experiments, the benzyl and chlorambucil mustards were shown to alkylate calf thymus DNA more readily than the benzoyl mustards, presumably due to destabilization of the aziridinium intermediate by the electron-withdrawing (σ>0) carboxamido group of the benzoyl compounds. Compounds 8-11 were found to alkylate guanine-N7 in the major groove, while compounds 6 and 7 did not, suggesting that they may have different modes of DNA interaction. Mustards 8-11 were also more efficient than 6 and 7 at producing DNA interstrand cross-links in isolated DNA. In general, for these compounds, the cytotoxicity against human chronic myeloid leukemia K562 cells and the panel of human tumor cell lines of the National Cancer Institute increased with the number of imidazole moieties. The IC50 values of compounds 7 and 8 were similar, even though the latter compound was at least 100-fold more efficient at forming DNA cross-links in isolated DNA. Similarly, compounds 9-11 were less cytotoxic than 6 and 7, although they were more efficient cross-linkers in isolated DNA. A direct comparison of the three imidazole-containing benzoyl mustard 7 with the corresponding chlorambucil-containing 11 for their ability to form interstrand cross-links in cells revealed that the former compound showed no cross-linking even at doses in excess of the IC50, whereas the latter produced extensive cross-linking. This further suggests that these compounds exert their biological activity through different mechanisms. It is proposed that the aromatic moiety of compounds 8-11, which bind to the minor groove, may be able to intercalate between GC base pairs and the protruding bischloroethylamino group would be positioned to alkylate and cross-link at guanine-N7 sites in the major groove. However, the benzoyl mustards, which have a rigid amido linkage between the imidazole and aromatic-mustard moieties, do not have the flexibility to intercalate into the DNA. Thus, they are forced to produce highly cytotoxic monoalkylations in the minor groove and are not favorably positioned to form interstrand cross-links.