Alkyltransferase Inhibitors for Cancer Chemotherapy

描述(申请人提供)：人类O6-烷基鸟嘌呤-DNA烷基转移酶(HAGT)是肿瘤细胞抵抗某些治疗性甲基化和氯乙基化药物杀伤的主要原因。在细胞培养中，用O6-苄基鸟嘌呤(BG)灭活HAGT可使人肿瘤对这些烷化剂增敏，并提高对裸鼠人肿瘤移植瘤的治疗指数。BG的临床试验表明，这种药物只有在有限的情况下才能成功，因为缺乏足够的效力和肿瘤特异性，而且容易产生对BG耐药的HAGT突变。因此，需要更有效、更特异的抑制剂。该提案有3个具体目的来开发这类化合物：(1)了解2-氨基-O4-苄基蝶呤衍生物对HAGT失活的机理。我们已经证明，其中一些化合物，包括O4-苄基叶酸(BF)，是HAGT的有效钝化剂。为此，将利用分子建模、结晶学、定点突变和获得信息突变的强大选择技术来了解这些化合物在BG上活性提高的基础，DNA存在干扰失活的原因，并改进这类化合物的设计。(2)探讨叶酸转运与BF灭活细胞内AGT及克服AGT介导的烷化剂抗性之间的关系。这些研究将跟进我们的初步研究，这些研究表明，在叶酸转运水平较高的肿瘤细胞系中，BF在灭活AGT方面要有效得多。(3)初步结果还表明，BG或O6-苄基-2‘-脱氧鸟苷的其他叶酸衍生物和含有多个BG残基的寡聚脱氧核苷酸也具有提供改进的AGT抑制剂的性质。这一目标的最终目标是通过对这些化合物的进一步研究和适当的修饰，可以制造出能够满足预期目标的AGT灭活剂，这些目标包括增加溶解度、增强对BG耐药突变体的效力和肿瘤特异性。抗癌药耐药性的产生是限制这些药物成功的一个主要因素。这些实验提供了一种方法来逆转众所周知的对用于此类治疗的烷化剂的耐药性来源。因此，它们将增加这些药物产生治疗反应的可能性。

DESCRIPTION (provided by applicant): Human O6-alkylguanine-DNA alkyltransferase (hAGT) is a major cause of resistance of tumor cells to killing by certain therapeutic methylating and chloroethylating agents. Inactivation of hAGT by O6-benzylguanine (BG) has been shown to sensitize human tumors to these alkylating agents in cell culture and to improve the therapeutic index against human tumor xenografts in nude mice. Clinical trials with BG indicate that this agent will be successful only in limited situations due to the lack of adequate potency and tumor specificity and the facile production of mutants of hAGT resistant to BG. Therefore, more potent and specific inhibitors are needed. The proposal has 3 specific aims to develop such compounds: (1) To understand the mechanism of the inactivation of hAGT by 2-amino- O4-benzylpteridine derivatives. We have shown that some of these compounds including O4-benzylfolic acid (BF) are potent inactivators of hAGT. In this aim, molecular modelling, crystallography, site-directed mutagenesis and powerful selection techniques to obtain informative mutants will be used to understand the basis for the improved activity of these compounds over BG, the reason that the inactivation is interfered with by the presence of DNA and to improve design of this class of compounds. (2) To investigate the relationship between folate transport and the ability of BF to inactivate cellular AGT and overcome AGT-mediated resistance to alkylating agents. These studies will follow up our preliminary studies that have shown that BF is much more effective at inactivating AGT in tumor cell lines that have high levels of folate transport. (3) Preliminary results also indicate that other folate derivatives of BG or O6-benzyl-2'-deoxyguanosine and oligodeoxynucleotides containing multiple BG residues also have properties that would provide improved AGT inhibitors. The ultimate goal of this aim is that by further study and appropriate modifications of these compounds, AGT inactivators can be made that fill the desired goals of increased solubility, increased potency against BG resistant mutants and tumor specificity. The development of resistance to anticancer agents is a major factor in limiting the success of these drugs. These experiments provide a means to reverse a well know source of resistance to alkylating agents that are used in such therapy. As such, they will increase the likelihood that these agents will produce therapeutic responses.