Protein Biomarkers for Invasive Aspergillosis Diagnostics

用于侵袭性曲霉病诊断的蛋白质生物标志物

• 批准号: 7788036
• 负责人: Marta L. Feldmesser
• 金额: $ 21.3万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-01-01 至 2011-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7788036
• 关键词: Adrenal Cortex Hormones; Affect; Allergic; Allergic Bronchopulmonary Aspergillosis; Allergic Disease; Animal Model; Antibody Formation; Antifungal Agents; Antigens; Arizona; Aspergillosis; Aspergillus fumigatus; Asthma; Award; Biochemistry; Biological Assay; Biological Markers; Biophysics; Carbohydrates; Clinical; Collaborations; Communicable Diseases; Contracts; Detection; Development; Diagnosis; Diagnostic; Diagnostic Procedure; Diagnostic tests; Disease; Disease Marker; Early Diagnosis; Enzyme-Linked Immunosorbent Assay; Fever; Fungal Antibodies; Fungal Proteins; Growth; Head; Hypersensitivity; Immunoassay; In Vitro; Individual; Infection; Link; Lung diseases; Mass Spectrum Analysis; Mediating; Medicine; Methods; Modeling; Molecular; Monoclonal Antibodies; Mus; Organism; Outcome; Pathogenesis; Patients; Process; Proteins; Proteome; Proteomics; Reagent; Recombinant Proteins; Research; Resources; Serum; Specimen; Syndrome; United States National Institutes of Health; Universities; assay development; base; biodefense; burden of illness; clinical application; college; disease diagnosis; flexibility; immunosuppressed; improved; innovation; mortality; novel diagnostics; point of care; prototype; response

DESCRIPTION (provided by applicant): With the rise in susceptible individuals, the burden of disease due to Aspergillus fumigatus has increased substantially. In addition, because of the difficulties in diagnosis, the actual number of affected people likely has been severely underestimated. Improved outcome in patients with invasive aspergillosis has been directly linked to earlier diagnosis, underscoring the need for more accurate diagnostic tests. Among the challenges posed for diagnostic assays include: i. the ubiquity of the organism, necessitating the ability to distinguish between contamination, infection, and disease; ii. the variety of susceptible hosts, in whom the pathogenesis of disease may vary; iii. the spectrum of disease syndromes, which range from allergy-mediated processes to aspergilloma to invasive disease. At present, the majority of effort in diagnostic development has focused on serum detection of carbohydrate antigens, PCR-based indicators of fungal burden or immunoassays for detection of anti-fungal antibody responses. Fungal proteomic analyses generally have been limited to in vitro growth conditions. This application proposes a collaboration between Dr. Marta Feldmesser (Albert Einstein College of Medicine) with Drs. John Galgiani and Vicki Wysocki (University of Arizona) to identify specific protein biomarkers associated with invasive disease in animal models of neutropenic and corticosteroid-treated mice, but that are not found in a model of allergic bronchopulmonary aspergillosis. The fungal proteome will be probed for the range of markers that will allow for more sensitive and specific markers of disease using mass spectrometry approaches to biomarker identification. Recombinant proteins then will be used to create monoclonal antibodies that can be used for mass spectrometry-based assay development. Six specific aims are proposed: 1. To identify A. fumigatus proteins present specifically during invasive pulmonary disease and not during fungal asthma; 2. to express and purify recombinant proteins of three candidate biomarkers of invasive aspergillosis; 3. to make monoclonal antibodies to fungal proteins identified in Aim 1 for use in diagnostic assays; 4. to use mass spectrometry to identify monoclonal antibodies for use in immunoassays; 5. to develop a prototype ELISA to detect candidate biomarkers in experimental invasive and allergic aspergillosis murine models and in clinical specimens from other NIH contract resources; and 6. to begin to migrate the recombinant protein and MAb reagents to multiplex and point-of-care platforms for clinical applications. The first two Aims would be accomplished during the R21 portion of the award, while the remaining four would be done during the R33 period. The accomplishment of these studies would provide novel diagnostic methods with potential for flexibility in response to particular clinical settings.

描述（由申请人提供）：随着易感个体的增加，烟曲霉引起的疾病负担大幅增加。此外，由于诊断困难，受影响的实际人数可能被严重低估。侵袭性曲菌病患者预后的改善与早期诊断直接相关，这强调了需要更准确的诊断测试。对诊断测定提出的挑战包括：生物体的普遍存在，需要区分污染、感染和疾病的能力; ii.易感宿主的多样性，其中疾病的发病机理可能不同; iii.疾病综合征的范围，从过敏介导的过程到曲霉瘤到侵袭性疾病。目前，诊断开发的大部分努力集中在碳水化合物抗原的血清检测、基于PCR的真菌负荷指标或用于检测抗真菌抗体应答的免疫测定。真菌蛋白质组学分析通常仅限于体外生长条件。该申请提出了玛尔塔费尔德梅瑟博士（阿尔伯特爱因斯坦医学院）与约翰·加尔贾尼博士和维奇·威索基博士（亚利桑那大学）之间的合作，以鉴定与在经肾上腺皮质激素和皮质类固醇治疗的小鼠的动物模型中的侵袭性疾病相关的特异性蛋白质生物标志物，但在过敏性支气管肺曲霉菌病模型中未发现。真菌蛋白质组将探测标记物的范围，这将允许使用质谱方法进行生物标记物鉴定以获得更灵敏和更特异的疾病标记物。然后，重组蛋白将用于产生可用于基于质谱的测定开发的单克隆抗体。提出了六个具体目标：1。鉴定A.烟曲霉蛋白专门存在于侵袭性肺部疾病期间，而不是真菌哮喘期间; 2.表达并纯化侵袭性曲霉病三种候选生物标志物的重组蛋白; 3.制备目标1中鉴定的真菌蛋白的单克隆抗体，用于诊断测定; 4.使用质谱法鉴定用于免疫测定的单克隆抗体; 5.开发原型ELISA以检测实验侵袭性和过敏性曲霉菌病小鼠模型和来自其他NIH合同资源的临床标本中的候选生物标志物;以及6.开始将重组蛋白和单克隆抗体试剂迁移到临床应用的多重和即时平台。前两个目标将在R21期间完成，其余四个目标将在R33期间完成。这些研究的完成将提供新的诊断方法，具有响应特定临床环境的灵活性。