Development of immune assays for HPV-32

描述(由申请人提供)：口腔的机会性感染困扰着50%的艾滋病毒感染者，包括口咽部念珠菌病(OPC)、口腔毛状白斑(OHL)和由粘膜萎缩性人乳头瘤病毒(HPV)引起的口腔疣。采用高效抗逆转录病毒疗法(HAART)积极治疗HIV，通过降低全身HIV病毒载量和恢复免疫功能，主要是通过增加CD4+T淋巴细胞，显著改善了HIV患者的健康和预后。这导致艾滋病毒相关机会性口腔疾病的发病率大幅下降，包括OPC和(OHL)。与之形成鲜明对比的是，据报道，自广泛应用HAART以来，口腔乳头状瘤(疣)的发病率有所增加。初步分析已确定HPV-32是在新奥尔良HIV+队列中发现的口腔疣的主要基因型别。人们认为，无症状的口腔HPV感染经常发生，但口腔尖锐湿疣很少发生，可能是由于对病毒的免疫控制。阻止无症状HPV感染进展到HPV疾病的免疫反应的关键方面尚不清楚，但以前的研究主要集中在针对主要衣壳蛋白LI的HPV基因型特异性反应。HIV+患者HPV相关口腔病理发生率的增加突出表明需要更彻底地了解口腔HPV感染的免疫反应。此外，口腔的可访问性为对HPV感染、免疫和发病机制进行严格分析提供了独特的机会。为了启动这些研究，需要开发针对口服HPV基因型别的免疫学检测方法，如HPV-32。我们假设HPV-32特异性体液和细胞免疫分析方法可以被开发并用于筛选人群中的HPV-32特异性免疫反应。我们建议建立一种用于检测血清中HPV-32特异性抗体的酶联免疫吸附试验(ELISA)，以及用于检测HPV-32特异性T细胞反应的淋巴增殖法和流式细胞术。将对这些检测进行最佳检测条件、特异性和重复性的测试。最终，这些检测将被用于研究免疫在口腔HPV感染的获得和随后的清除或进展中的作用，特别是在高度易感的HIV+患者中。 该项目的目标是开发必要的工具，以更好地了解口腔HPV感染，特别是HIV+患者的体液和细胞免疫反应。了解口腔HPV感染的免疫应答可能有助于预测HPV疾病的进展，并阐明HPV相关疾病的治疗和预防策略。

DESCRIPTION (provided by applicant): Opportunistic infections of the oral cavity afflict 50% of all HIV infected patients, and include oropharyngeal candidiasis (OPC), oral hairy leukoplakia (OHL) and oral warts caused by the mucosatropic human papillomavirus (HPV). The aggressive treatment of HIV with highly-active antiretroviral therapy (HAART) has significantly improved the HIV patient's health and prognosis by lowering systemic HIV viral loads and restoring immune function primarily through increases in CD4+ T lymphocytes. This has resulted in a substantial decrease in the incidence of HIV-associated opportunistic oral diseases, including OPC and (OHL). In stark contrast, the incidence of oral papillomas (warts) has reportedly increased since the widespread administration of HAART. Preliminary analysis has identified HPV-32 as the predominant genotype found in oral warts in the New Orleans HIV+ cohort. It is felt that asymptomatic oral HPV infection occurs frequently but oral warts occur infrequently, presumably due to immunological control of the virus. The critical aspects of the immune response that prevent the progression from asymptomatic HPV infection to HPV disease are unknown, but previous studies have focused on HPV genotype-specific response against the major capsid protein, LI. The increased rate of HPV-related oral pathology seen in HIV+ patient's highlights the need for a more thorough understanding of the immune response to oral HPV infections. Furthermore, the accessibility of the oral cavity affords a unique opportunity to conduct rigorous analysis of HPV infection, immunity and pathogenesis. To initiate these studies, the development of the immunological assays specific for oral HPV genotypes such as HPV-32 are required. We hypothesize that HPV-32 specific humoral and cellular immunoassays can be developed and utilized to screen populations for HPV-32 specific immune responses. We propose to develop an enzyme-linked immunosorbent assay (ELISA) for detection of HPV-32-specific antibodies in serum, and lymphoproliferative and flowcytometric assays for the detection of HPV-32-specific T cell responses. These assays will be tested for optimal assay conditions, specificity, and reproducibility. Ultimately these assays will be used to investigate the role of immunity in the acquisition and subsequent clearing or progression of oral HPV infections, particularly in the highly susceptible HIV+ patient. The goal of this project is to develop the tools necessary to gain a better understanding of the humoral and cellular immune response to oral HPV infection, particularly in the HIV+ individual. The understanding of immune response to oral HPV infection may help to predict HPV disease progression as well as elucidate therapeutic and preventive strategies for HPV-related disease.