Investigation of a shipworm endosymbiont compound with activity against the AIDS-associated pathogens Cryptosporidium and Toxoplasma

 描述(申请人提供)：隐孢子虫属。弓形虫和弓形虫是机会性的尖端复合体寄生虫，在免疫功能低下的人中会导致显著的发病率和死亡率。治疗这些寄生虫的新疗法是医学上的当务之急。我们已经确定了一种由软体动物共生细菌产生的化合物，它可以抑制微小弧菌和弓形虫的细胞内生长，而对宿主细胞没有毒性。船虫是一种海洋双壳软体动物，它们钻进洞穴并贪婪地消耗木材。像大多数食木动物一样，船虫在共生细菌的帮助下消化木材，但在船虫中，共生体寄生在鳃细胞内，并将纤维素酶从细菌液泡、宿主细胞和盲肠腔中输出到盲肠腔中，在那里它们可以消化船虫的木质食物。尽管环境营养丰富，但船虫盲肠缺乏细菌群落，这表明存在强大的抗微生物化合物。船虫共生体已被证明在体内产生抗微生物的次生代谢产物，这些化合物可能在维持盲肠几乎无菌的环境中发挥作用。我们发现，轮虫鳃内共生菌Teredinibacter turnerae在体外产生一种化合物，在NM水平上抑制微小弧菌和弓形虫的细胞内生长，并抑制小鼠的微小弧菌感染。该化合物对相关的血寄生虫牛巴贝斯虫也有活性，表明对尖端复合体有广泛的疗效。在这项应用中，我们将测试这种化合物针对弓形虫和微小弧菌共同的基因产物或过程的假设，以及这种活性在体内有效的假设。为了验证这一假设，我们提出了以下具体目标： 1A-确定共生菌化合物针对的弓形虫和微小弧菌的感染过程。我们将评估共生菌化合物对特定寄生虫过程的影响，并测试该化合物对其他顶端复合体的作用，包括II型弓形虫速殖子和缓殖子。目的1B-确定微小弧菌和弓形虫共生化合物的活性靶标。我们将使用遗传和生化方法来鉴定这种化合物所针对的基因产物。目的1C-评价共生菌化合物对小鼠微小弧菌感染的疗效。这些实验将为未来涉及靶标验证、结构-活性关系、药代动力学-药效学评价以及其他微小弧菌和弓形虫感染动物模型的疗效测试等研究奠定基础。以前从来没有人研究过船虫共生体产生抗寄生虫化合物的潜力。因此，这些研究具有开辟抗寄生虫药物发现的新领域的巨大潜力，有望为药物开发提供新的分子靶点，并有可能广泛应用于许多难治病原体。

 DESCRIPTION (provided by applicant): Cryptosporidium spp. and Toxoplasma gondii are opportunistic apicomplexan parasites that cause significant morbidity and mortality in immunocompromised people. New therapeutics to treat these parasites are a medical imperative. We have identified a compound produced by a mollusk symbiotic bacterium that inhibits intracellular growth of C. parvum and T. gondii without toxicity to the host cell. Shipworms are marine bivalve mollusks that burrow into and voraciously consume wood. Like most xylophagous animals, shipworms digest wood with the assistance of their symbiotic bacteria but in shipworms, the symbionts are housed within the cells of the gill and export cellulolytic enzymes out of the bacterial vacuole, out of the host cell, and into the lumen of the caecum where they effect digestion of the shipworm's woody diet. Despite the nutrient rich environment, the shipworm caecum lacks a bacterial community suggesting the presence of potent anti-microbial compounds. Shipworm symbionts have been shown to produce anti-microbial secondary metabolites in vivo and it is likely these compounds play a role in maintaining the nearly sterile environment of the caecum. We found that the shipworm gill endosymbiont, Teredinibacter turnerae, produces a compound that inhibits intracellular growth of C. parvum and T. gondii at nM levels in vitro and inhibits C. parvum infection in mice. This compound is also active against the related hemoparasite, Babesia bovis, suggesting a broad efficacy against apicomplexans. In this application, we will test the hypothesis that this compound targets a gene product or process common to T. gondii and C. parvum, and that this activity is effective in vivo. To test this hypothesis, we propose the following specific aim: Aim 1A - Identify the T. gondii and C. parvum infection processes targeted by the symbiont compound. We will evaluate the effect of the symbiont compound on specific parasite processes and test the compound against other apicomplexans, including Type II T. gondii tachyzoites and bradyzoites. Aim 1B - Identify the target of the symbiont compound's activity in C. parvum and T. gondii. We will employ both genetic and biochemical approaches to identify the gene products targeted by this compound. Aim 1C - Evaluate the efficacy of the symbiont compound against murine C. parvum infection. These experiments will provide the foundation for future studies involving target verification, structure-activity relationships, pharmacokinetic-pharmacodynamic evaluation and tests of efficacy in other animal models of C. parvum and T. gondii infection. Shipworm symbionts have never before been investigated for their potential to produce anti-parasitic compounds. Thus, these studies have tremendous potential to open up a new area of anti-parasitic drug discovery, holding the promise of new molecular targets for drug development, potentially with broad application to many intractable pathogens.