Development of Vaccine Strategies to prevent EBV+ Lymphoma in Patients with HIV

描述(由申请人提供)：感染艾滋病毒的患者患恶性B细胞淋巴增生性疾病(LPD)的风险增加。这些LPD中的大多数是EB病毒(EBV)阳性的，具有侵袭性的临床病程，并与不良预后相关。EBV是一种致癌的人类疱疹病毒，感染全球90%以上的成年人，并与广泛的恶性肿瘤有关。HIV感染患者中EBV-LPD的发生率部分与EBV特异性T淋巴细胞反应的数量和或质量不足有关。我们假设，针对EBV裂解抗原和潜伏抗原的细胞免疫缺陷会导致EBV-LPD风险增加，疫苗增强的EBV特异性免疫将在这一高危人群中恢复保护并防止EBV-LPD的发展。我们利用了一种SCID小鼠模型，当移植来自健康EBV血清阳性捐赠者的人白细胞时，自发地产生人EBV-LPD。我们发现了几种EBV潜伏抗原和裂解抗原，当疾病在体内得到控制时，可以被人CD8(+)T细胞识别。此外，我们验证了医源性免疫抑制的EBV-LPD患者在减少免疫抑制方案后在体内成功控制疾病的同时对这些相同抗原的识别。我们将几个EBV基因克隆到腺相关和腺病毒载体中，感染了人类抗原提呈细胞，并展示了强大的自体T细胞扩增。使用负载BZLF四聚体的BZLF1免疫优势多肽，我们证明了在这些患者体内BZLF1特异性T细胞的扩张与EBV+肿瘤的消退和患者生存的改善相关。在白血病淋巴瘤协会和公司合作伙伴的支持下，我们最近完成了对BZLF1的大规模提纯，并正在为明年计划举行的Pre IND会议启动临床前毒性研究。由于一种全面的EBV疫苗可能需要多个EBV蛋白靶点，我们建议完成对另外4个靶蛋白的表征，并进行临床前分析，以确定可能进一步在临床上开发的有前途的免疫原。这项重大机遇基金将实现三个具体目标：(1)在体外表征不同人类白细胞抗原类型对建议的5个全长基因产物中每一个的人体细胞反应，以确定最佳候选基因产物(S)以纳入疫苗接种策略；(2)在我们的EBV-LPD鼠-人嵌合模型中，评估全长纯化病毒多肽制剂在刺激和维持EBV特异性记忆T细胞增殖和预防致死性人类EBV相关淋巴增殖性疾病方面的有效性，以及(3)扩大全长候选多肽的生产，用于进一步的鉴定、最佳递送和佐剂活性的评估、毒性评估、生物标记物验证和免疫原性研究。一旦我们在这笔赠款的支持下完成了上述基本探索性研究，我们将确定最佳的病毒靶点(S)，这些靶点可以在广泛的人类白细胞抗原谱中“看到”，并能够申请IND，对向感染EBV-LPD的高危HIV+患者提供特定疫苗制剂的安全性进行第一阶段试验。最终，针对这种病毒驱动的恶性肿瘤的预防方法可能会被证明是更有效、更便宜的，并有望通过进一步降低EBV-LPD的发生率和提高感染艾滋病毒患者的生活质量，造福于世界各地更多的人。 公共卫生相关性：感染人类免疫缺陷病毒(HIV)的人患恶性B细胞淋巴增生性疾病(LPD)的风险增加。这些LPD大多为EB病毒(EBV-LPD)阳性，具有侵袭性的临床病程，并与不良预后相关。健康、有免疫能力的人不会发生EBV-LPD，因为有一种高效的、适应性的细胞、抗原特异性免疫反应。HIV感染患者中EBV-LPD的发生率部分与EBV特异性T淋巴细胞反应的数量和或质量不足有关。我们的实验室计划专注于开发有效的策略来“增强”细胞免疫监测机制，以治疗和预防潜在免疫缺陷患者的EBV-LPD。我们已经开发了一个可重复的小动物模型，它非常接近于人类EBV-LPD，开发了几种实验性治疗策略来提供EBV编码的多肽靶蛋白，并与行业和非营利性组织合作，合成了预防EBV-LPD的疫苗。在这里，我们建议利用我们在过去几年中开发的工具来系统地测试选择性EBV裂解和潜伏全长多肽疫苗预防EBV-LPD的有效性。这一重大机遇奖将提供资金，以完成所有基础科学工作，以确定关键的EBV靶标，确认生物标记物来衡量对疫苗的免疫反应，并进行临床前测试，以确定有希望的疫苗候选蛋白，然后在我们合作伙伴的帮助下，转向临床开发，为HIV+患者接种疫苗，可能还包括其他免疫抑制患者群体。该项目具有巨大的潜力，可以对公共卫生、创造就业和经济增长产生影响。

DESCRIPTION (provided by applicant): Patients infected with HIV are at increased risk of developing malignant B cell lymphoproliferative diseases (LPD). The majority of these LPDs are positive for the Epstein-Barr virus (EBV), run an aggressive clinical course and are associated with a poor prognosis. EBV is an oncogenic, human herpes virus that infects more than 90% of adults worldwide and is associated with a wide spectrum of malignancies. The incidence of EBV- LPD in HIV-infected patients is, in part, related to the inadequate quantity and or quality of EBV-specific T lymphocyte response. We hypothesize that deficiency of cellular immunity against EBV lytic and latent antigens leads to increased risk of EBV-LPD, and vaccine-enhanced EBV-specific immunity will restore protection and prevent the development of EBV-LPD in this high risk population. We utilized a SCID mouse model which, when engrafted with human leukocytes from healthy EBV seropositive donors, spontaneously develops human EBV-LPD. We discovered several EBV latent and lytic antigens recognized by human CD8(+) T cells when the disease is controlled in vivo. Furthermore, we validated the recognition of these same antigens in iatrogenically immune suppressed patients with EBV-LPD while successfully controlling their disease in vivo following the reduction of their immune suppressive regimen. We cloned several EBV genes into an adeno-associated and adenoviral constructs, infected human antigen presenting cells and demonstrated robust autologous T cell expansion. Using HLA-specific tetramers loaded with immune dominant peptides derived from BZLF, we documented that In vivo expansion of BZLF1-specific T cells occurring in these patients correlated with EBV+ tumor regression and improved patient survival. With the support of Leukemia Lymphoma Society and corporate partners, we have recently completed a large scale purification of BZLF1 and are initiating preclinical toxicity studies for a planned pre IND meeting over the next year. Because a comprehensive EBV vaccine will likely require multiple EBV protein targets, we propose to complete a characterization of 4 additional target proteins and perform preclinical analysis to identify promising immunogens that may be further developed clinically. This Grand Opportunities grant will accomplish three specific aims: (1) characterize the human cellular response across various HLA types in response to each of the 5 proposed full length gene products in vitro in order to identify the optimal candidate gene product(s) to include in the vaccination strategy; (2) assess the efficacy of full length purified viral polypeptide preparations in stimulating and maintaining EBV specific memory T cell expansion and preventing fatal human EBV- associated lymphoproliferative disorder in our chimeric mouse-human model of EBV-LPD and (3) scale up production of full length candidate polypeptides for further characterization, evaluation of optimal delivery and adjuvant activity, toxicity evaluation, biomarker validation and immunogenicity studies. Once we have completion the above basic exploratory studies with the support of this grant, we will have identified the optimal viral target(s) that are "seen" across a broad HLA spectrum and be in a position to apply for an IND to conduct Phase I trials for safety of delivering specific vaccine preparations to HIV+ patients who are at high risk of developing EBV-LPD. Ultimately, a preventive approach for such viral-driven malignancies will likely prove to be more effective, less expensive, and hold great promise for benefitting more people worldwide by further reducing the incidence EBV-LPD and improving the quality of life of patients infected with HIV. PUBLIC HEALTH RELEVANCE: People infected with the Human Immune Deficiency virus (HIV) are at increased risk of developing malignant B cell lymphoproliferative diseases (LPD). Most of these LPDs are positive for Epstein-Barr virus (EBV-LPD), run an aggressive clinical course and are associated with a poor prognosis. Healthy, immune competent people do not develop EBV-LPD because of a highly efficient, adaptive cellular, antigen-specific immune response. The incidence of EBV-LPD in HIV-infected patients is, in part, related to the inadequate quantity and or quality of EBV-specific T lymphocyte response. Our laboratory program has focused on developing efficient strategies to "boost" cellular immune surveillance mechanisms to treat and prevent EBV-LPD in patients with underlying immune deficiency. We have developed a reproducible small animal model that closely mimics human EBV-LPD, several experimental therapeutic strategies to deliver EBV-encoded polypeptide target proteins, and partnerships with industry and not-for profit organizations to synthesize a vaccine to prevent EBV-LPD. Here we propose to utilize tools that we have developed over the past several years to systematically test the efficacy of vaccinating with selective EBV lytic and latent full length polypeptides to prevent EBV-LPD. This Grand Opportunities Award will provide funding to complete all basic scientific work to identify the key EBV targets, confirm biomarkers to measure immune response to the vaccine, and perform preclinical testing to identify promising vaccine candidate proteins that can then, with the help of our partners, move on towards clinical development to vaccinate HIV+ patients and, perhaps other immune suppressed patient groups. The project has great potential to make an impact on public health, job creation and economic growth.