CHEMICAL STUDIES OF TAXOL AND MICROTUBULES

紫杉醇和微管的化学研究

• 批准号: 3199593
• 负责人: CHARLES S SWINDELL
• 金额: $ 13.45万
• 依托单位: BRYN MAWR COLLEGE
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-07-05 至 1995-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3199593
• 关键词: acylation; analog; antibody; antineoplastics; biological products; chemical binding; chemical models; chemical structure function; computer simulation; conformation; crosslink; diterpenes; drug design /synthesis /production; enzyme linked immunosorbent assay; microtubules; molecular dynamics; molecular site; nuclear magnetic resonance spectroscopy; paclitaxel; photochemistry; physical chemical interaction; stereochemistry; tubulin

Taxol, a diterpene isolated from the Pacific yew, Taxus brevifolia, has exhibited antineoplastic activity in phase I clinical trials against melanoma, adenocarcinoma, refractory ovarian carcinoma, non-small cell lung carcinoma, gastric, colon, and head and neck carcinomas, and lymphoblastic leukemias. In combination with cisplatin, activity has been observed against advanced large cell lung carcinoma, non-small cell lung carcinoma, breast, head and neck, and colon carcinomas, advanced ovarian carcinoma, and melanoma. Activity also has been displayed by taxol in phase II trials against advanced ovarian epithelial tumors in patients pretreated with radiation and chemotherapy, some of whom were considered resistant to cisplatin. Three areas associated with the development of taxol as an antitumor agent, and with the exploitation of its unique mode of action as a fundamentally new and general strategy for the management of neoplastic disease are addressed in this application. At the present time, the limited quantities of taxol available are insufficient for its broad application in cancer chemotherapy. Partial synthesis through the attachment of the critical A-ring side chain to more readily available but inactive natural substances can alleviate this problem. We will develop three new A-ring side chain attachment methodologies, two that involve 5,6-dihydro-6-keto-1,3 oxazine chemistry (including one that coalesces asymmetric side chain synthesis with attachment to the diterpenoid nucleus into a total of five steps), and one that involves ketene methodology. The success of the former strategy is suggested by preliminary results on the attachment of side chains with variable functionality. The binding site(s) of taxol on microtubules remain completely uncharacterized. This state of affairs prevents the understanding at the molecular level of taxol's unique mode of action, which, in turn, undermines its exploitation through the design of new drugs. Three areas of investigation aimed at the chemically relevant characterization of the taxol binding site will be pursued. First, we will evaluate experimentally (further structure-activity studies and the conformational analysis of taxol analogues through NOE and 13C T1 studies) and computationally (molecular mechanics modeling and dynamical calculations) the hypothesis that taxol and its biologically active analogues are preorganized for tight binding to microtubules. This notion is supported strongly by preliminary structure-activity and modeling results. Should this prove to be true, significant information on the nature of the binding site and for the optimization of drug design will be available in the short term. Second, we will engage in photoaffinity labeling of the binding site. These studies will reveal the domains and amino acid residues of microtubules responsible for the recognition of taxol. Third, we will combine the information from the first two studies with the available data on the structure of the tubulins in a computational model of the taxol binding site. The picture that will emerge will reveal the three-dimensional chemical features of the interaction between taxol and the important structural elements of tubulin at the binding site. Finally, we will pursue the preparation of taxol derivatives that will be useful at the cellular and subcellular levels for the investigation of the biology that underlies taxol's cytotoxicity. For example since the site on microtubules to which taxol binds cannot have evolved for that purpose, it is possible that taxol mimics an endogenous substance with similar biological activity. Antibodies generated to taxol will be employed in a search for such endogenous substances. Similar antibodies and fluorescent taxol analogues for the imaging of taxol bound to microtubules will be prepared.

紫杉醇是一种从太平洋紫杉短叶红豆杉中分离出来的二萜， 在I期临床试验中表现出抗肿瘤活性， 黑色素瘤，腺癌，难治性卵巢癌，非小细胞肺癌 癌、胃癌、结肠癌和头颈癌，以及淋巴母细胞癌 白血病 与顺铂联合使用时， 晚期大细胞肺癌，非小细胞肺癌， 乳腺癌、头颈癌和结肠癌，晚期卵巢癌， 和黑色素瘤 紫杉醇在II期临床试验中也显示出活性 对晚期卵巢上皮性肿瘤的治疗， 放疗和化疗，其中一些人被认为是耐药的， 顺铂 与紫杉醇作为抗肿瘤剂的发展相关的三个领域， 并利用其独特的行动模式， 新的和一般的战略管理肿瘤疾病是 在这个应用程序中解决。 目前，可用的紫杉醇数量有限， 不足以广泛应用于癌症化疗。 部分 通过将关键的A环侧链连接到更多的 容易获得但无活性的天然物质可以缓解这种情况 问题. 我们将开发三种新的A环侧链连接物 方法，两个涉及5，6-二氢-6-酮-1，3恶嗪化学 （包括将不对称侧链合成与 连接到二萜类化合物核共五个步骤），和一个 这涉及到乙烯酮的方法。 前一种策略的成功之处在于 这是由关于侧链与 可变功能 紫杉醇在微管上的结合位点完全保留 没有特征的 这种情况阻碍了对 紫杉醇独特作用模式的分子水平，反过来， 通过设计新药来破坏其开发。 三个领域 研究的目的是化学相关的表征， 将追踪紫杉醇结合位点。 首先，我们将通过实验评估 （进一步的结构活性研究和构象分析） 通过NOE和13 C T1研究的紫杉醇类似物）和计算 （分子力学建模和动力学计算）假设 紫杉醇及其生物活性类似物是预先组织的， 与微管结合。 这一观点得到了初步的有力支持。 构效关系和建模结果。 如果这是真的， 关于结合位点性质的重要信息， 药物设计的优化将在短期内实现。 第二、 我们将对结合位点进行光亲和标记。 这些 研究将揭示微管的结构域和氨基酸残基 负责识别紫杉醇。 第三，我们将联合收割机 前两项研究的信息与现有的数据， 紫杉醇结合计算模型中微管蛋白的结构 绝佳的价钱 出现的图片将揭示三维的 紫杉醇与重要代谢物相互作用的化学特征 微管蛋白的结构元件在结合位点。 最后，我们将继续制备紫杉醇衍生物， 在细胞和亚细胞水平上用于研究 紫杉醇细胞毒性的生物学基础。 例如，由于网站 与紫杉醇结合的微管不可能是为了这个目的而进化的， 紫杉醇可能模拟具有类似 生物活性 针对紫杉醇产生的抗体将用于 寻找这种内源性物质。 类似的抗体和荧光 用于结合于微管的紫杉醇成像的紫杉醇类似物将 制备