A genetic engineering strategy for the improved production of the anti-helminthic

提高抗蠕虫药物生产的基因工程策略

• 批准号: 8393663
• 负责人: Jeffrey David Kittendorf
• 金额: $ 17.05万
• 依托单位: ALLUVIUM BIOSCIENCES, INC.
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-01 至 2014-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8393663
• 关键词: Agreement; Anabolism; Animals; Antibiotics; Antifungal Agents; Area; Biological Factors; Biotechnology; Chemicals; Clinical; DNA Sequence; Data; Development; Disease; Domestic Animals; Engineering; Evaluation; Fermentation; Fungal Chromosomes; Fungal Genome; Gene Amplification; Gene Cluster; Gene Deletion; Genes; Genetic; Genetic Engineering; Genetic Recombination; Genome; Genomics; Harvest; Health; Helminths; Human; Infection; Knock-in Mouse; Knock-out; Letters; Livestock; Malignant Neoplasms; Manufacturer Name; Marketing; Mediation; Metabolic Biotransformation; Metabolism; Methods; Michigan; Motivation; Mutagenesis; Output; Parasitic nematode; Pathway interactions; Pharmaceutical Preparations; Phase; Phenotype; Population; Production; Regulation; Regulator Genes; Relative (related person); Research Activity; Risk; Rural; Safety; Site; Streptomyces; Sum; System; Technology; Testing; Therapeutic; Work; analog; antimicrobial; base; chemotherapeutic agent; combat; commercial application; cost; cost effective; drug candidate; drug development; drug discovery; effective therapy; genome sequencing; improved; interest; manufacturing process; meetings; mutant; paraherquamide; paraherquamide A; phase 2 study; programs; research study; success; synthetic biology; tool; willingness

DESCRIPTION (provided by applicant): Helminth infections are responsible for severe health problems worldwide, in both human and animal populations. It is estimated that 2-3 billion people, mainly from rural and impoverished regions of the world suffer from disease due to parasitic nematodes; however, drug development and discovery efforts lack commitment to combat human infection due to the poor commercial market. In contrast, there are robust animal health discovery efforts aimed at eradicating infection in valuable domesticated livestock herds. Thus, animal health programs are often relied on to provide drug candidates for human therapeutics. Extensively tested in animals, the cost and development risk of human drugs originating from animal health are reduced. While human drug candidates still need to surpass stringent safety standards, an equal challenge is minimizing cost of goods to enable the infected population access to potent, low cost therapeutics. Accordingly, paraherquamide represents a new class of anti-helminth antibiotics that possesses broad-spectrum anti-nematodal activity and is a precursor to a recently launched animal health anti-parasitic agent 2- desoxoparaherquamide (Derquantel(tm)). Currently paraherquamide is manufactured through fermentation of the fungal strain P. simplicissimum. To maximize yields, the drug manufacturer has embarked on strain improvement programs to randomly evolve high producing mutants; however, this program has failed to achieve the desired output. Here, Alluvium proposes a modernized, genomics-driven strain improvement strategy aimed at maximizing paraherquamide production in fermentation. Specifically, Alluvium's approach is guided by the DNA sequence of the fungal genome in which key regulatory and/or biosynthetic genes responsible for paraherquamide production have been identified. Rational genetic engineering strategies, such as gene deletion or gene amplification, will be applied to key genes in efforts to rationally generate a high producing mutant phenotype. In this Phase I proposal, work is focused on rationally manipulating three genes that are predicted to regulate, both positively and negatively paraherquamide biosynthesis. In addition, Alluvium proposes to replicate the entire paraherquamide biosynthetic gene cluster within the fungal chromosome in order to boost production of anti-helminthic natural product. Following success in these initial studies, Phase II work will focus on engineering further efficiencies in paraherquamide production, including developing biocatalytic methods capable of converting paraherquamide to 2-desoxoparaherquamide as well as biotransformation tools that can be employed to generate synthetically challenging structural analogs. In sum, the strain engineering technology under development at Alluvium will result in an optimized paraherquamide manufacturing process that will lower the cost of goods required to access this commercially important anti- helminthic therapeutic, thereby enabling the potential development of a much-needed human therapeutic. PUBLIC HEALTH RELEVANCE: Helminths, especially parasitic nematodes, cause severe health problems in both humans and domesticated animals. Approximately 2-3 billion people in rural and impoverished regions are infected; however, affordable and effective treatment options are scarce. The proposed work seeks to improve upon the production of the promising anti-helminthic animal health chemotherapeutic agent, paraherquamide, so it can be generated in a more cost-effective and efficient manner, thereby stimulating further development activities for potential human treatment.

描述（由申请人提供）：在人类和动物种群中，蠕虫感染负责全球严重的健康问题。据估计，有2-30亿人，主要来自世界上农村和贫困地区，由于寄生线虫而遭受疾病。但是，由于商业市场不佳，药物开发和发现工作缺乏打击人类感染的承诺。相比之下，有强大的动物健康发现工作旨在消除宝贵的驯养牲畜群中的感染。因此，通常依靠动物健康计划为人类治疗剂提供候选毒品。在动物中进行了广泛的测试，源自动物健康的人类药物的成本和发育风险减少了。尽管人类候选者仍然需要超过严格的安全标准，但同等的挑战是最大程度地减少商品成本，以使受感染的人口获得有效的低成本治疗方法。因此，副喹啉酰胺代表了一类新的抗螺旋抗生素，具有广谱抗新肿瘤活性，并且是最近发射的动物健康抗寄生虫2-脱氧化甲帕拉胺（Desoxoparaherquamide）（Decopare剂（TM））的前体。目前，副喹啉酰胺是通过发酵真菌菌株P来制造的。简单。为了最大程度地提高产量，药物制造商已开始进行应变改善计划，以随机进化高产生突变体。但是，该程序未能实现所需的输出。在这里，冲积物提出了一种现代化的基因组驱动的菌株改善策略，旨在最大化发酵中的副喹啉酰胺。具体而言，冲积物的方法是由真菌基因组的DNA序列指导的，在该基因组的DNA序列中，已经确定了负责副喹啉酰胺产生的关键调节和/或生物合成基因。理性基因工程策略（例如基因缺失或基因扩增）将应用于关键基因，以努力 理性地产生高产生的突变表型。在这一阶段的提案中，工作的重点是合理地操纵三种基因，这些基因被预测，这些基因可以正常和负面寄生虫生物合成。此外，冲积物建议在真菌染色体内复制整个副喹啉生物合成基因簇，以增强抗固有天然产物的产生。在这些初步研究中成功后，第二阶段 工作将集中在旁氨酰胺生产中的工程效率上，包括开发能够将旁夸雷胺转化为2-氧甲莫克胺以及可用于生成合成具有挑战性结构类似物的生物转化工具。总而言之，在冲积公司开发的应变工程技术将导致优化的副氨基胺制造过程，该过程将降低访问这种商业上重要的抗死式治疗所需的商品成本，从而使备受挑战的人类治疗的潜在发展。 公共卫生相关性：蠕虫，尤其是寄生线虫，在人类和驯养的动物中引起严重的健康问题。在农村和贫困地区大约有2-3亿人被感染；但是，负担得起的有效治疗选择很少。拟议的工作旨在改善有希望的抗幽灵动物健康化学治疗剂Paraherquamide的生产，因此可以以更具成本效益和有效的方式生成，从而刺激了潜在人类治疗的进一步发展活动。