Combined Substrate Polymerase Inhibitors

描述（由申请人提供）：研究DNA聚合酶和逆转录酶的组合底物抑制剂作为用于治疗病毒性疾病的更特异性和毒性更小的疗法的潜在途径，肝炎、艾滋病等，也许是为了改善目前的癌症化疗，也许是为了找到暴露于生物恐怖生物（包括病毒和细菌）的一线治疗方法。这些新型抑制剂基于两种抑制剂探针，即，DNA聚合酶抑制剂2-S-4-叠氮苯酰基-硫代-2 ′-脱氧腺苷5 ′-三磷酸（1）和RT抑制剂2-S-4-叠氮苯酰基-硫代-1，N6-乙烯基-2 ′-脱氧腺苷5 ′-三磷酸（2）。这两种结构类别（与第三个探针不包括在本项目中）一起包围了腺苷碱基小沟侧周围的整个空间，当三磷酸与催化三联体配位时，将侧链放置在与模板结合位点相互作用的位置。这些抑制剂将在多种酶上进行测试，包括HIV-1、M-MLV、AMV和RAV 2逆转录酶、E. coli DNA聚合酶I、T4、T7和Taq DNA聚合酶以及RNA聚合酶和末端转移酶的选择性，并对合成和天然模板/引物以及所有dNTP的抑制机制进行了完整表征。 将生成两种抑制剂类型的文库，其中侧链的长度增加和减少，然后通过苯环取代或用环烷基环取代，将该文库中鉴定的每种酶的特异性抑制剂进一步多样化。该建议是基于这样的假设，即虽然组合的底物抑制剂性质的线索1，2和3将保持跨聚合酶，具体的抑制性质将取决于个别的聚合酶，从而允许识别非常具体的抑制模式和抑制剂结合位点。

DESCRIPTION (provided by applicant): Combined substrate inhibitors of DNA polymerases and reverse transcriptases are investigated as potential avenues to more specific and less toxic therapies for treating viral diseases, e.g., hepatitis, AIDS, etc., perhaps to improve current cancer chemotherapies, and perhaps to find frontline treatments for exposure to bio-terror organisms, including viruses and bacteria. These new classes of inhibitors are based on two inhibitor probes, i.e., the DNA polymerase inhibitors 2-S-4-azidophenacyl-thio-2'- deoxyadenosine 5'-triphosphate (1) and the RT inhibitor 2-S-4-azidophenacyl-thio-l,N6-etheno-2 '- deoxyadenosine 5'-triphosphate (2). These two structural classes (with a third probe not included in this project) together envelop the entire space surrounding the minor groove side of the adenosine base, placing the side chain in position to interact with template binding sites when the triphosphate is coordinated to the catalytic triad. These inhibitors will be tested on a large variety of enzymes, including HIV-1, M-MLV, AMV, and RAV2 reverse transcriptase, E. coli DNA polymerase I, T4, T7 and Taq DNA polymerases, and RNA polymerase and terminal transferase for selectivity, with complete characterization of inhibition mechanism versus synthetic and natural template/primers and all dNTPs. A library will be generated for the two inhibitor types where the length of the side chain is increased and decreased, and then specific inhibitors for each enzyme identified in this library will be further diversified by phenyl ring substitutions or replacement with cycloalkyl rings. This proposal is based on the hypothesis that although the combined substrate inhibitor properties of leads 1, 2, and 3 will be maintained across the polymerases, specific inhibitory properties will depend on individual polymerases, thus allowing identification of very specific inhibition patterns and inhibitor binding sites.