Identification of new compounds against fungal microbes

鉴定抗真菌微生物的新化合物

• 批准号: 8667984
• 负责人: Maurizio Del Poeta
• 金额: $ 19.74万
• 依托单位: STATE UNIVERSITY NEW YORK STONY BROOK
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-06-01 至 2017-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8667984
• 关键词: Acquired Immunodeficiency Syndrome; Affect; Alveolar; Antifungal Agents; Aspergillus; Blood Circulation; Brain; Breathing; Candida; Carbon Dioxide; Cause of Death; Cells; Centers for Disease Control and Prevention (U.S.); Ceramide glucosyltransferase; Cessation of life; Cryptococcus; Cryptococcus neoformans; Cryptococcus neoformans infection; Data; Development; Disease; Drug Targeting; Environment; Enzymes; Fusarium; Gene Mutation; Glucosylceramides; Goals; Growth; HIV; Health; Human; In Vitro; Infection; Injection of therapeutic agent; Laboratories; Lead; Libraries; Lung; Mammalian Cell; Microbe; Molds; Mus; Mycoses; Pathogenesis; Pathway interactions; Patients; Pentamidine; Plants; Pneumocystis; Pneumocystis Infections; Pneumocystis carinii Pneumonia; Production; Public Health; Reporting; Research Personnel; Sphingolipids; Testing; Therapeutic; Time; Transcript; Tuberculosis; Validation; Virulence; Virulence Factors; Yeasts; fungus; in vivo Model; inhibitor/antagonist; insight; mortality; mutant; novel; novel therapeutics; pathogen; prevent; public health relevance; pulmonary granuloma; screening; treatment strategy

DESCRIPTION (provided by applicant): The goal of this project is to assess the therapeutic potential of novel antifungal agents, identified by our laboratory via screening the ChemBridge library, that target the synthesis of fungal glucosylceramide (GlcCer). Recently, we reported that the fungal GlcCer is required for the pathogenic fungus Cryptococcus neoformans (Cn) to cause a lethal meningo-encephalitis1, 2. In particular, we showed that a Cn mutant strain lacking the final enzyme for the synthesis of glucosylceramide (glucosylceramide synthase 1 or Gcs1), after inhalation, was confined in the lung granuloma and, thus, did not reach the bloodstream and did not disseminate to the brain. Later, other investigators corroborated and extended our findings that mutation of genes involved in the last steps of the GlcCer pathway affect fungal pathogenesis not only of fungi infecting humans, such as Cn3, 4, Candida albicans5-7, and Aspergillus fumigatus8, but also of fungi infecting plants, such as Fusarium graminearum9. Further studies in our lab showed that the mechanism by which GlcCer promotes virulence of Cn is by allowing fungal growth in a neutral/alkaline environment, such as that present in the alveolar spaces2, 10. The importance of GlcCer to promote fungal growth in the lung is also underscored by the fact that in most dimorphic fungi, production of GlcCer is detected only in the lung infective form (yeast) and not in the environmental form (mold)11-13, suggesting that also in these fungi GlcCer may be required for lung infection. The synthesis of GlcCer seems to be important also during Pneumocystis pneumonia (PCP) as glucosylceramide synthase transcripts have been found to be abundant at the time of isolation of the fungus from a fulminate lung infection14. Taken together, these studies suggest that GlcCer is most likely a pan-fungal virulence factor required during lung infection to promote fungal growth at the neutral/alkaline environment of alveolar spaces, and as such, it is a promising novel drug target. Therefore, we looked for inhibitors of GlcCer synthesis by screening a ChemBridge library for compounds that inhibit Cn growth in an environment similar to the lung: neutral/alkaline pH, 37C and 5% CO2. We identified 2 compounds that significantly decreased the synthesis of GlcCer in Cn but not in mammalian cells. Importantly, 90% of mice treated with the lead compound, BHBM, survived a lethal intranasal injection of Cn and their lungs (and brains) were free from fungal cells. Moreover, we found that BHBM is over 50-fold more active than pentamidine against the lung pathogen Pneumocystis in vitro. Therefore, we hypothesize that targeting the fungal GlcCer pathway will be an effective novel therapeutic strategy for impeding the development of airborne fungal diseases. To test this hypothesis, we propose the following aims: 1) identify and validate the target(s) of the active compounds; and 2) study the effect of the identified compounds in in vivo models of fungal infections.

描述（由申请人提供）：该项目的目的是评估我们实验室通过筛选Chembridge库确定的新型抗真菌剂的治疗潜力，该实验室针对真菌葡萄糖基酶（GLCCCER）的合成。 Recently, we reported that the fungal GlcCer is required for the pathogenic fungus Cryptococcus neoformans (Cn) to cause a lethal meningo-encephalitis1, 2. In particular, we showed that a Cn mutant strain lacking the final enzyme for the synthesis of glucosylceramide (glucosylceramide synthase 1 or Gcs1), after inhalation, was confined in the肺肉芽肿，因此没有到达血液，也没有传播到大脑。后来，其他研究者证实并扩展了我们的发现，即GLCCER途径的最后一步的基因突变会影响真菌发病机理，不仅会影响真菌感染人类，例如CN3、4，candida albicans5-7，candida albicans5-7和fumigaTus8和曲植虫，而且对植物的曲霉，以及诸如Fungi Infectect fungi Infectect fungi fungiium fimectioum fimectectioum graminear graminear graminacar graminacrim graminain9。 我们实验室中的进一步研究表明，GLCCER促进CN的毒力的机制是通过在中性/碱性环境中允许真菌生长，例如存在于肺泡空间2、10中的。环境形式（模具）11-13，也表明 在这些真菌中，可能需要肺部感染。在肺炎肺炎（PCP）中，GLCCER的合成似乎也很重要，因为在从富芒酸肺部感染中分离真菌时，已经发现已发现葡萄糖基酶合酶转录物很丰富14。 综上所述，这些研究表明，GLCCER很可能是肺部感染过程中所需的泛膜毒力因子，以促进肺泡空间的中性/碱环境的真菌生长，因此，这是一个有前途的新型药物靶标。 因此，我们通过筛选Chembridge库的化合物来寻找GLCCER合成的抑制剂，这些化合物在类似于肺的环境中抑制CN生长：中性/碱性pH，37C和5％CO2。我们确定了2种显着降低CN中GLCCER的合成但没有在哺乳动物细胞中的合成。重要的是，用铅化合物BHBM处理的90％的小鼠在CN的致命性鼻内注射及其肺（和大脑）中幸存下来。此外，我们发现BHBM在体外对肺炎肺囊肿的活性超过50倍。 因此，我们假设针对真菌GLCCER途径将是阻碍空气传播真菌疾病发展的有效新型治疗策略。为了检验该假设，我们提出以下目的：1）识别和验证活性化合物的靶标；和2）研究所鉴定化合物在真菌感染的体内模型中的作用。