Rapid Diagnostics for Mucormycosis

毛霉菌病的快速诊断

• 批准号: 8832062
• 负责人: Marwan Nasralla
• 金额: $ 15.11万
• 依托单位: VITALEX BIOSCIENCES, LLC
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-02-09 至 2017-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8832062
• 关键词: Acidosis; Adrenal Cortex Hormones; Age; Amino Acids; Amphotericin B; Angioinvasion; Animal Model; Animals; Antibodies; Antifungal Agents; Antifungal Therapy; Antigens; Applications Grants; Aspergillus fumigatus; Binding; Biological; Biological Assay; Biological Markers; Blood Vessels; Blood specimen; Bronchoalveolar Lavage; Candida; Cell Line; Cell Surface Proteins; Clinical; Colony-forming units; Data; Detection; Diabetes Mellitus; Diabetic Ketoacidosis; Diagnosis; Diagnostic; Diagnostic tests; Disease; Disease Outcome; Disease Progression; Endothelial Cells; Enzyme-Linked Immunosorbent Assay; Feasibility Studies; Goals; Heat shock proteins; Hematogenous; Hematologic Neoplasms; Hematopoietic Stem Cell Transplantation; Human; Immune system; Immunocompromised Host; Incidence; Infection; Invaded; Iron; Life; Liposomes; Malignant Neoplasms; Mammalian Cell; Medical center; Methods; Modeling; Monoclonal Antibodies; Mucorales; Mucormycosis; Mus; Mycoses; Neutropenia; Operative Surgical Procedures; Organ Transplantation; Organism; Oryza; Outcome; Pathogenesis; Patients; Penetration; Pharmaceutical Preparations; Phase; Population; Predictive Value; Prevalence; Proteins; Reporting; Rhizopus; Risk; Risk Factors; Sampling; Self-Sustained Sequence Replication; Sensitivity and Specificity; Serum; Signal Transduction; Small Business Technology Transfer Research; Techniques; Testing; Thrombosis; Tissues; United States; Urine; Western Blotting; Zygomycosis; base; clinically relevant; fungus; glucose-regulated proteins; improved; mortality; mouse model; progression marker; public health relevance; rapid detection; rapid diagnosis; response; usability

 DESCRIPTION (provided by applicant): Mucormycosis, most commonly caused by Rhizopus oryzae, is a life-threatening infection that occurs in patients immunocompromised by diabetic ketoacidosis (DKA), neutropenia, corticosteroid use, and/or increased serum iron. Because of the rising prevalence of these risk factors, the incidence of mucormycosis has risen. Despite disfiguring surgery and aggressive antifungal therapy, the mortality of mucormycosis remains >40%, and approaches 100% in patients with disseminated disease and prolonged neutropenia. A key factor which contributes to these abysmal mortality rates of mucormycosis is the current lack of rapid diagnostic tests. This deficiency often results in delayed therapy. Thus, a rapid diagnostic test for mucormycosis is likely to improve the outcome of the disease. Clinical hallmarks of R. oryzae infection include its remarkable angiotropism. We recently made the important discovery that the fungal cell surface proteins encoded by CotH facilitate disease progression by allowing R. oryzae to invade mammalian cells via binding to Glucose Regulated Protein 78 (GRP78), a heat shock protein expressed on endothelial cells lining blood vessels during mucormycosis. Importantly, CotH proteins were found to be conserved among Mucorales (organisms that cause mucormycosis) with amino acid identity ranging from 55-98%. Equally important CotH proteins are unique to Mucorales since they are absent from any other known organisms. Finally, CotH proteins were found to be expressed in clinically relevant animal models of mucormycosis including the DKA mouse model. These features strongly indicate that CotH and their gene products can be utilized for rapid detection of mucormycosis. Indeed our preliminary data show PCR-related methods using specific primers to CotH can amplify signals in blood samples spiked with different Mucorales but not Aspergillus fumigatus or Candida. Moreover, anti-CotH antibodies can recognize a band similar to the predicted size of CotH proteins in serum samples collected from infected mice. We propose to build on these exciting data to further establish CotH and/or their gene products as biomarkers for diagnosis of mucormycosis, progression of the disease and response to therapy. Our goal in this Phase I feasibility study is to use our established and clinically relevant DKA and neutropenic mucormycosis mouse models to develop a PCR-based assay and/or an antigen detection test targeting CotH in biological samples collected from mice infected with Mucorales. Phase II of this STTR application will focus on establishing a PCR-based kit, sandwiched ELISA and/or dipstick assays for the rapid detection of CotH or circulating CotH antigens by testing using human clinical samples. Establishing a rapid detection test will improve mucormycosis outcome by early initiation of proper therapy and inform the response to antifungal therapy.

 描述（由申请人提供）：毛霉菌病最常见由米根霉引起，是一种危及生命的感染，发生在因糖尿病酮症酸中毒（DKA）、中性粒细胞减少症、皮质类固醇使用和/或血清铁升高而免疫功能低下的患者中。由于这些危险因素的普遍存在，毛霉菌病的发病率有所上升，尽管进行了毁容手术和积极的抗真菌治疗，但毛霉菌病的死亡率却有所上升。毛霉菌病的死亡率仍然>40%，并且在患有播散性疾病和长期中性粒细胞减少症的患者中，导致毛霉菌病死亡率极高的一个关键因素是目前缺乏快速诊断检测，因此，这种缺陷常常导致治疗延迟。毛霉菌病的快速诊断测试可能会改善米根霉的临床特征，包括其显着的血管向性。研究发现，CotH 编码的真菌细胞表面蛋白通过与葡萄糖调节蛋白 78 (GRP78) 结合，允许米根霉侵入哺乳动物细胞，从而促进疾病进展，葡萄糖调节蛋白 78 是一种在毛霉菌病期间血管内皮细胞上表达的热休克蛋白。研究发现，毛霉菌目（引起毛霉菌病的生物体）中的蛋白质具有保守性，氨基酸同一性范围为 55-98%。 CotH 蛋白是毛霉目所独有的，因为它们在任何其他已知生物体中都不存在。最后，发现 CotH 蛋白在临床相关的毛霉菌病动物模型（包括 DKA 小鼠模型）中强烈表达。这些特征表明 CotH 及其基因产物可以被表达。事实上，我们的初步数据表明，使用 CotH 特异性引物的 PCR 相关方法可以放大掺有不同毛霉菌的血液样本中的信号，但不能放大烟曲霉或曲霉菌的信号。此外，抗 CotH 抗体可以识别与从感染小鼠收集的血清样本中预测的 CotH 蛋白大小相似的条带，我们建议以这些令人兴奋的数据为基础，进一步建立 CotH 和/或其基因产物作为诊断的生物标志物。我们在这一阶段的可行性研究试验中的目标是使用我们已建立的临床相关的 DKA 和中性粒细胞减少性毛霉菌病小鼠模型来开发基于 PCR 的和/或基于毛霉菌病的模型。该 STTR 应用第二阶段针对从感染毛霉目的小鼠收集的生物样本中的 CotH 进行抗原检测测试，重点是建立基于 PCR 的试剂盒、夹心 ELISA 和/或试纸检测，通过测试快速检测 CotH 或循环 CotH 抗原。使用人类临床样本建立快速检测测试将通过及早开始适当的治疗来改善毛霉菌病的结果并告知抗真菌治疗的反应。