NEW APPROACHES TO ANTIFOLATE CHEMOTHERAPY

这个实验室一直致力于设计、合成和 经典化合物的衍生物和结构类似物的生物学评价 和非经典的抗叶酸，目的是产生新的试剂 改善的药理和治疗特性或定性的 改变了抗肿瘤活性的光谱。这项研究涉及 与多个共享 对抗叶酸药物发现的创新方法感兴趣。一位少校 这项工作的目标是找到具有更强累积能力的化合物 对甲氨蝶呤(MTX)等经典抗叶酸耐药的肿瘤细胞 由于吸食毒品的缺陷。预计交叉抗性 这类化合物和甲氨蝶呤之间的关系将是低的，它们将有 对天然或获得性肿瘤的潜在临床效用 甲氨蝶呤耐药基于转运和/或多谷氨酰化缺陷。 其次，由于甲氨蝶呤耐药性的一种重要形式已知涉及 二氢叶酸还原酶(DHFR)的结构突变导致弱 这项工作的另一个目标是结合MTX类型的经典抗叶酸 寻找比MTX更紧密地与这些DHFR结合的化合物 变种。最后，由于人们已经广泛认识到抑制 除DHFR外的叶酸途径酶，特别是胸苷合成酶 (TS)和甘氨酰胺核苷甲酰基转移酶(GART)，提供了一种强大的 选择性杀伤肿瘤细胞的另一种方法和 基于DHFR突变或增加的MTX耐药性规避 生产野生型酶，研究的第三个目标是产生 抑制这些酶的化合物，而不是DHFR，或除了DHFR之外。 在这个项目中已经发现的一个重要线索是 水溶性非经典DHFR抑制剂N-α-(4-氨基-4- Deoxypteroyl)-N-delta-hemiphthaloyl-L-ornithine(PT523，NSC633713)。这 结构独特的抗叶酸比MTX更有效地对抗广泛的 培养的人类实体肿瘤细胞范围，包括10-30倍的细胞 由于转运受损或DHFR增加而对MTX产生抵抗力 活性--通常与获得性甲氨蝶呤相关的甲氨蝶呤抵抗水平 患者的耐药性。更大的潜力和更低的增长也是 PT523对小鼠肿瘤和人肿瘤移植瘤的体内抗肿瘤作用 在裸鼠体内。因为PT523缺少谷氨酸侧链，所以它不能 形成多聚谷氨酸；因此，与经典的抗叶酸不同，它不是 前药，不依赖多聚谷氨酰胺来表现其全部影响 一碳新陈代谢。此外，与亲脂性的非经典不同 抗叶酸如曲美曲辛和匹立曲辛，PT523保持活性 对抗典型多药耐药(MDR)表型的细胞 基于高P-糖蛋白表达；因此不太可能交叉 对许多处方广泛的抗肿瘤天然产品产生抗药性 如蒽环类、长春花碱、鬼臼毒素、喜树碱、 和紫杉烷。在这些非常令人兴奋的结果的基础上，主要关注的是 下一个资助周期将是第二代类似物的合成 和靶向前药PT523用于体内外试验和SAR 分析。PT523的结构修改将以 侧链、芳基、桥和B环。靶向前药将 包括旨在释放母体药物的多肽衍生物 在体内被肿瘤靶向的Moab偶联多肽酶切割后。 还将努力发现与突变株结合良好的类似物。 对标准经典和非经典不敏感的DHFRs 抗叶酸。这项工作的重点将是合成 分子中的桥键已从6-移动到5- B形环的位置。这个项目的长期目标是发现 并开发用于治疗人类癌症的新的和新型的抗叶酸。

This laboratory has been engaged in a program of design, synthesis, and biological evaluation of derivatives and structural analogs of classical and nonclassical antifolates with the aim of generating new agents with improved pharmacological and therapeutic properties or a qualitatively altered spectrum of antitumor activity. The research involves collaborative interactions with a number of other groups sharing an interest in innovative approaches to antifolate drug discovery. A major goal of the work is to find compounds with increase ability to accumulate in tumor cells resistant to classical antifolates like methotrexate (MTX) by virtue of a defect in drug uptake. It is expected that cross resistance between such compounds and MTX will be low, and that they will have potential clinical utility against tumors with either natural or acquired MTX resistance based on defects in transport and/or polyglutamylation. Secondly, since an important form of MTX resistance is known to involve structural mutations in dihydrofolate reductase (DHFR) resulting in weak binding of classical antifolates of the MTX type, another goal of the work is to find compounds that will bind more tightly than MTX to these DHFR variants. Finally, since it has become widely recognized that inhibition of folate pathway enzymes other than DHFR, especially thymidylate synthase (TS) and glycinamide ribotide formyltransferase (GART), offers a powerful alternative approach to the selective killing of tumor cells and the circumvention of MTX resistance based on DHFR mutation or increased production of wild-type enzyme, a third goal of research is to generate compounds that inhibit these enzymes instead of, or in addition to, DHFR. An important lead already uncovered during this project has been the water-soluble nonclassical DHFR inhibitor N-alpha-(4-amino-4- deoxypteroyl)-N-delta-hemiphthaloyl-L-ornithine (PT523, NSC633713 ). This structurally unique antifolate is more potent than MTX against a broad range of cultured human solid tumor cells, including cells 10- to 30-fold resistant to MTX by virtue of either impaired transport or increased DHFR activity -- a level of MTX resistance typically associated with acquired resistance in patients. Greater potency and decreased growth has also been shown with PT523 in vivo against murine tumors and human tumor xenografts in athymic nude mice. Because PT523 lacks a glutamate side chain it cannot form polyglutamates; thus, unlike classical antifolates, it is not a prodrug and does not rely on polyglutamation to manifest its full impact on one-carbon metabolism. Moreover, unlike lipophilic nonclassical antifolates such as trimetrexate and piritrexim, PT523 retains activity against cells with the classical multidrug resistance (MDR) phenotype based on high P-glycoprotein expression; thus it is unlikely to be cross- resistant to a number of widely prescribed antineoplastic natural products such as anthracyclines, vinca alkaloids, podophyllotoxins camptothecins, and taxanes. On the basis of these very exciting results, a major focus of the next grant cycle will be the synthesis of second-generation analogs and targeted prodrugs of PT523 for in vitro/in vivo testing and SAR analysis. Structural modification of PT523 will feature changes in the side chain, aryl moiety, bridge, and B-ring. Targeted prodrugs will include peptide derivatives that are designed to release the parent drug in vivo after cleavage by tumor-targeted MoAb-conjugated peptidases. Efforts will also be made to discover analogs that bind well to mutant DHFRs that are insensitive to standard classical and nonclassical antifolates. The focus of this effort will be on the synthesis of molecules in which the bridge has been moved from the 6- to the 5- position of the B-ring. The longterm goal of this program is to discover and develop new and novel antifolates for the treatment of human cancer.