TRAIL-expressing Recombinant Adenovirus Based Immunotherapy for RCC

描述（由申请人提供）：今年在美国将诊断出约40，000例新的肾细胞癌（RCC）病例，预计RCC将导致> 13，000例死亡。转移性RCC的中位生存期为8个月，约30%的RCC患者被诊断为晚期转移性疾病。局部RCC通过切除手术治疗;然而，目前没有有效的治疗方法可用于转移性疾病。对RCC的适应性免疫在肿瘤监测中起着关键作用，CD 8+和CD 4 + T细胞都有助于肿瘤消退。RCC转移的主要部位之一是肺，已知即使在没有肿瘤生长的情况下，肺也是免疫抑制的环境。因此，如果针对转移性RCC的新型免疫疗法要进入临床试验，与原发性肿瘤部位相比，必须了解肺部环境对保护性T细胞反应的产生和维持的影响。我们先前描述了编码人肿瘤坏死因子相关凋亡诱导配体（TRAIL; Ad 5-TRAIL）cDNA的重组腺病毒载体的开发。最近，我们的研究集中在使用一个类似的重组腺病毒编码全长鼠TRAIL（Ad 5-mTRAIL）的免疫活性小鼠实验肾细胞癌（Renca）肿瘤。来源于Ad 5-mTRAIL杀死的Renca细胞的抗原可以在体外交叉呈递给CD 8 + T细胞，并且单独使用Ad 5-mTRAIL最低限度地延长存活并在体内诱导低水平的CTL活性。为了增强DC效率，我们将Ad 5-mTRAIL与CpG寡核苷酸（CpG）组合，其显著增强体内抗原特异性T细胞增殖和CTL活性，以及延长荷Renca肿瘤小鼠的存活。有趣的是，治疗前去除CD 4+或CD 25+细胞进一步提高了存活率和体内CTL活性，去除CD 4+细胞的无肿瘤小鼠也拒绝了Renca细胞的后续攻击，但不是MHC匹配的前列腺肿瘤细胞，表明存在肿瘤特异性免疫记忆。虽然CD 4+细胞耗竭的影响是惊人的，但存活率未达到100%，表明存在其他免疫抑制机制。在该更新申请中，我们提出了以下具体目的来研究以下假设，即当与设计用于克服肿瘤源性免疫抑制的疗法联合使用时，局部Ad 5-TRAIL施用将在患有实验性转移性RCC的小鼠的肺中产生保护性抗肿瘤免疫：目的1）确定在原发性RCC肿瘤部位瘤内施用Ad 5-mTRAIL/CpG在多大程度上影响针对实验性肺转移的CD 8+效应T细胞应答的幅度;目的2）研究CD 4 + T细胞亚群，特别是CD 4 + CD 25 + Treg和Th 17对CD 8 + T细胞介导的抗肾癌肺转移瘤免疫应答的质和量的影响;和目的3）检查在Ad 5-mTRAIL/CpG免疫治疗期间肿瘤浸润性骨髓细胞群体对肺抗肿瘤免疫应答的影响。聚焦于这些细胞的刺激和抑制能力。目前的提案反映了我们致力于开发新的RCC免疫疗法，可以消除原发肿瘤和远端转移，从而延长晚期RCC患者的生存期。 公共卫生相关性：对于进行性转移性肾细胞癌（RCC）患者，非常需要继续开发创新的治疗策略。上一个资助期产生的数据显示，使用编码全长鼠TNF相关凋亡诱导配体cDNA序列的重组腺病毒和（TRAIL; Ad 5-mTRAIL）和免疫刺激性CpG寡核苷酸构成了当前提议背后的基本原理，其中我们将研究局部Ad 5-mTRAIL给药和随后的肿瘤细胞凋亡，与设计用于克服肿瘤源性免疫抑制的治疗相结合，将在含有实验性RCC转移瘤的小鼠的肺中导致全身保护性抗肿瘤免疫。我们将利用这些发现来更好地设计肿瘤免疫治疗的体内方法，不仅要了解Ad 5-mTRAIL对肿瘤细胞的直接作用机制，还要了解系统性抗肿瘤免疫应答调节背后的一些基本原则，以及肿瘤细胞凋亡和抗原交叉呈递如何用于刺激抗肿瘤免疫。

DESCRIPTION (provided by applicant): This year in the U.S. ~40,000 new cases of renal cell carcinoma (RCC) will be diagnosed and >13,000 deaths are expected from RCC. Metastatic RCC carries a median survival of 8 months and ~30% of RCC patients are diagnosed with advanced metastatic disease. Localized RCC is treated surgically through resection; however, there is no efficacious therapy currently available for metastatic disease. Adaptive immunity to RCC plays a critical role in tumor surveillance, with both CD8+ and CD4+ T cells contributing to tumor regression. One of the major sites of RCC metastasis is the lung, an environment that is known to be immunosuppressive even in the absence of tumor outgrowth. Therefore, if novel immunotherapies against metastatic RCC are to move forward into clinical trials, the effect of the lung environment on the generation and maintenance of protective T cell responses, compared to the primary tumor site, must be understood. We previously described the development of a recombinant adenoviral vector encoding the human TNF-related apoptosis-inducing ligand (TRAIL; Ad5-TRAIL) cDNA. More recently, our studies have focused on using a similar recombinant adenovirus encoding full-length murine TRAIL (Ad5-mTRAIL) in immunocompetent mice bearing experimental renal cell carcinoma (Renca) tumors. Antigens derived from Ad5-mTRAIL-killed Renca cells could be cross- presented to CD8+ T cells in vitro, and Ad5-mTRAIL used alone minimally prolonged survival and induced a low level of CTL activity in vivo. To enhance DC efficiency we combined Ad5-mTRAIL with CpG oligonucleotides (CpG), which significantly augmented in vivo antigen-specific T cell proliferation and CTL activity, as well as prolonged survival of Renca tumor-bearing mice. Interestingly, CD4+ or CD25+ cell depletion prior to therapy further enhanced survival and in vivo CTL activity, and CD4+ cell-depleted, tumor-free mice also rejected a subsequent challenge of Renca cells, but not MHC-matched prostate tumor cells, showing the existence of tumor-specific immunologic memory. While the effects of CD4+ cell depletion were striking, survival did not reach 100%, suggesting the presence of additional immunosuppressive mechanisms. In this renewal application, we propose the following specific aims to investigate the hypothesis that localized Ad5-TRAIL administration, when used in conjunction with therapies designed to overcome tumor- derived immunosuppression, will lead to protective antitumor immunity in the lungs of mice with experimental metastatic RCC: Aim 1) Ascertain the extent to which intratumoral Ad5-mTRAIL/CpG administration at the primary RCC tumor site impacts the magnitude of CD8+ effector T cell responses against experimental lung metastases; Aim 2) Examine the effect of CD4+ T cell subsets, specifically CD4+CD25+ Treg and Th17, on the quality and quantity of the CD8+ T cell-mediated antitumor response against RCC lung metastases; and Aim 3) Examine the impact of tumor-infiltrating myeloid cell populations on the lung antitumor immune response during Ad5-mTRAIL/CpG immunotherapy - focusing on the stimulatory versus suppressive capacities of these cells. The current proposal reflects our commitment to develop novel immunotherapies for RCC that can eliminate both primary tumors and distal metastases, thereby prolonging the survival of patients with advanced RCC. PUBLIC HEALTH RELEVANCE: There is great need to continue to develop innovative treatment strategies for patients with progressive metastatic renal cell carcinoma (RCC). Data generated during the previous funding period showing that tumor- specific systemic immunity can be induced using the combination of a recombinant adenovirus encoding the cDNA sequence for full-length murine TNF-related apoptosis-inducing ligand (TRAIL; Ad5-mTRAIL) and immunostimulatory CpG oligonucleotides constitutes the rationale behind the current proposal, where we will investigate the hypothesis that that localized Ad5-mTRAIL administration and the ensuing tumor cell apoptosis, combined with therapies designed to overcome tumor-derived immunosuppression, will lead to systemic protective antitumor immunity in the lungs of mice containing experimental RCC metastases. We will utilize these findings to better devise in vivo approaches for tumor immunotherapy and to understand not only the mechanism of direct action of Ad5-mTRAIL on tumor cells, but to understand some of the fundamental tenets behind the regulation of systemic antitumor immune responses and how tumor cell apoptosis and antigen cross-presentation can be used to stimulate antitumor immunity.