Studies Of Inducible Nitrogen Mustards-based DNA Cross-linking Agents And Monitorings Of Drug Release In Tumor Cells By Small Molecular

Due to DNA carrying the main source of genetic information in the body, it plays a very important role in metabolism. Common cells which contain modified DNA regions such as mutation or mismatch even at very low concentrations are closely linked to disease and apoptosis. So DNA is viewed as an important molecular target for many of the drugs that are used in chemotherapy, however, is also regarded as a non-specific target of cytotoxic agents. Accordingly, great effort has been put into discovering drugs that are more selective and effective; and there is significant progress that the identification of cancer-discernible molecular agents will evolve a new generation of less toxic side effect. DNA-interactive drugs such as inducible DNA cross-linking agents provide prominent opportunities for novel therapeutics that target DNA and its associated processes. Now it is difficult to establish precise delivery of a pharmaceutically activatable prodrug to cancer cells. Toward this end, fluorescent probes that can providea ready read-out under the typical conditions of cellular analysis are of particular interest. It would be desirable to develop a drug release system that contains both an active drug for efficacy and a fluorophore for monitoring.In the dissertation, we designed and synthesized two agents using a readily available reagent for cross-linking DNA that is controlled through an inducible process initiated by fluoride ions. The DNA-DNA cross-linking abilities of the fluoride-inducible DNA cross-linking agents were investigated by denatured alkaline agarose gel electrophoresis. The results showed that compound2resulted in nearly100%DNA cross-linking at a concentration as low as50μM and was found to be approximately twice as efficient as compound1. And the cytotoxicity of compound2towards HeLa cells was tested by MTT assays. Complementary HPLC analysis proves the proposed mechanism of fluoride-inducing deprotection.In the other part we designed a new compound naphthalimide-CLB conjugate (NCC) which is composed of a potent anticancer drugchlorambucil(CLB), a disulfide linker and a fluorescent naphthalimide moiety. Fluorescence responses of NCC toward thiol amino acids and other nonthiol amino acids were detected. HPLC analysis proves the proposed mechanism of thiol-inducing reaction. Moreover, cytotoxicities of compound NCC to cells were investigated through an integrated experiments including intracellular imaging, alkaline comet assay and y-H2AX immunofluorescence in HeLa cells. It confirmed apoptosis of cells were induced by damage of NCC targeting the DNA.