Laulimalide Production Genes

劳利马内酯生产基因

• 批准号: 6584382
• 负责人: CHARLES R HUTCHINSON
• 金额: $ 16.26万
• 依托单位: KOSAN BIOSCIENCES, INC.
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-03-01 至 2004-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6584382
• 关键词: Porifera; antineoplastics; aquatic organism; artificial chromosomes; biosynthesis; biotherapeutic agent; cytochrome P450; drug design /synthesis /production; drug discovery /isolation; gene expression; genetic library; neoplasm /cancer chemotherapy; neoplasm /cancer pharmacology; nucleic acid hybridization; nucleic acid sequence; polyketide synthase; polymerase chain reaction

DESCRIPTION (provided by applicant): The long-range goal of our research is to develop economical methods for producing multigram quantities of experimental antitumor drugs from marine sources. Our approach is based on cloning the biosynthetic genes from the producing organism followed by expression of these genes in a bacterial host that can be grown in large-scale fermentations. Mechanistic and preclinical investigations of bryostatins, discodermolide, halichondrins, laulimalide and other marine cytotoxins, which are made by pathways involving polyketide synthases, have been hampered by inadequate supplies of the marine polyketides because they are available only by isolation or total synthesis at a very high cost. The goal of Phase I research is to identify the genetic locus that contains the biosynthesis genes for laulimalide by the following specific aims: (1) Obtain fresh sponge tissue from a reliable source, and show that it (or its symbionts) contains laulimalide. (2) Detect modular polyketide synthase (PKS), CYP450, "hydroxymalonate (HM)" and hydroxymethylglutaryI-Coenzyme A synthase (HMGS) gene homologs by PCR analysis of DNA from cells or cell extracts of the marine sponge (and/or their microbial symbionts) that produce laulimalide. (3) Prepare fosmid and BAC libraries using DNA isolated from the sponge/symbiont complex that produces laulimalide or from a putative microbial symbiont, and by PCR and DNA hybridization analyses obtain clones that contain the PKS, CYP450, HM and HMGS genes. (4) Sequence the region that contains the PKS, CYP450, HM and HMGS genes to the extent required to justify their involvement in laulimalide biosynthesis. Our experimental design is built on work in progress in which we are attempting to clone the discodermolide biosynthesis genes. Success will lead to development of a fermentation-based process to make large amounts of laulimalide or one of its analogs for preclinical studies as an antitumor drug.

描述（由申请人提供）： 我们研究的远程目标是开发经济的方法，用于从海洋来源生产多数量的实验抗肿瘤药物。我们的方法基于从生产生物体中克隆生物合成基因，然后在细菌宿主中表达这些基因，这些基因可以在大规模发酵中生长。对肌蛋白丁蛋白，二甲某霉素，halichondrins，Laulimalide和其他海洋细胞毒素的机械和临床前研究，这些毒素是由涉及聚酮化合物合酶的途径进行的，这些途径因仅通过隔离或总合成而被造成的海洋多酮的供应不足，因为它们在非常高昂的成本上可用。第一阶段研究的目的是通过以下特定目的确定含有Laulimalide的生物合成基因的遗传基因座：（1）从可靠的来源获得新鲜的海绵组织，并表明它（或其共生体）包含Laulimalide。 （2）检测模块化聚酮合酶（PKS），CYP450，“羟基甲酸盐（HM）”和羟基甲酰核糖核A合成酶（HMGS）基因同源物通过PCR分析DNA的细胞或细胞提取物（和或或或或或或其symbial sybial）。 （3）使用从海绵/共生体复合物中分离出的DNA来制备fosmid和BAC文库，从而产生Laulimalide或从假定的微生物共生体中，以及通过PCR和DNA杂交分析获得的克隆，可以获得包含PKS，CYP450，HM和HMGS基因的克隆。 （4）对包含PKS，CYP450，HM和HMGS基因的区域进行序列，以证明其参与Laulimalide生物合成所需的程度。我们的实验设计建立在正在进行的工作中，我们试图克隆dessodermolide生物合成基因。成功将导致发展基于发酵的过程，以制造大量的Laulimalide或其作为抗肿瘤药物的临床前研究的类似物之一。