Mechanism of Action of CpG-ODN in Generating Tumor Immunity in the Aged

描述（由申请人提供）：我们先前已经鉴定了与HLA-A2.1/Kb转基因小鼠（A2 xneu）杂交的Her-2/neu转基因小鼠对Her-2/neu/A2. 1免疫显性表位具有免疫耐受性。我们还证明了用肽或Her-2/neu蛋白免疫不能提供针对A2 xneu小鼠中肿瘤的有效抗肿瘤应答。Her-2/neu小鼠与HLA-A2.1/Kb小鼠杂交的结果之一是，在这些动物中，当动物为20-22个月大时，自发性肿瘤出现。因此，A2 xneu小鼠模型提供了一个独特的机会，以评估抗自身抗原的抗肿瘤免疫应答，其中老化和耐受性同时存在。我们已经证明，衰老大大改变了老年小鼠的免疫系统。因此，关键是要确定和优化疫苗接种策略，有效地刺激免疫反应，导致肿瘤排斥的年轻和年老的耐受动物。有大量证据表明，用不同类型的佐剂靶向APC导致诱导抗肿瘤免疫应答。我们比较了不同TLR配体（Poly I：C、LPS、鞭毛蛋白、咪喹莫特、CpG-ODN）在年轻和老年A2 xneu小鼠中的抗肿瘤作用。结果表明，只有瘤内（i.t.）注射CpG-ODN在年轻和老年A2 xneu小鼠中诱导肿瘤排斥。尽管年轻和年老的A2 xneu小鼠在i.t注射CpG-ODN治疗后可以排斥原发肿瘤，但这些动物不能产生保护性记忆反应。t注射CpG-ODN后的肿瘤微环境分析表明：1）APC的活化; 2）促炎性Th 1型应答; 3）CD 4、CD 8 T细胞和NK细胞的活化;和4）T-T细胞的数量急剧减少。综上所述，这些结果表明，i.t. CpG-ODN的注射影响肿瘤微环境，有利于抗肿瘤反应。然而，我们还没有完全了解CpG-ODN注射如何诱导肿瘤排斥反应。该建议的目的是确定CpG-ODN在诱导抗肿瘤反应中的作用机制，并开发在老年耐受宿主中产生记忆反应的策略。在这些研究中，我们将：1）评估和表征CpG-ODN对老年小鼠免疫应答的诱导作用; 2）评估CpG-ODN与T-IFN之间的关系，以及T-IFN在调节老年小鼠免疫应答的库和激活中的关系;以及3）优化使用抗-neu-CpG-ODN以将CpG-ODN靶向于肿瘤部位用于治疗老年小鼠肿瘤。相关性：我们的研究结果首次表明，靶向肿瘤部位的CpG-ODN恢复了老年小鼠的免疫应答。研究CpG-ODN对免疫系统的影响将最终优化CpG-ODN的治疗。此外，随着抗-neu-CpG-ODN分子的产生，我们可以靶向体内任何地方的CpG-ODN，使得有可能开发用于治疗癌症的通用疫苗策略。总的来说，从这些研究中获得的信息将揭示控制和操纵免疫系统的策略，以在年轻和老年耐受宿主中开发更有效的免疫疗法。

DESCRIPTION (provided by applicant): We have previously identified that Her-2/neu transgenic mice crossed with the HLA-A2.1/Kb transgenic mice (A2xneu) are immunotolerant against Her-2/neu/A2.1 immunodominant epitopes. We have also demonstrated that immunization with peptides or Her-2/neu proteins does not provide an effective antitumor response against tumors in A2xneu mice. One of the consequences of crossing the Her-2/neu mice with the HLA-A2.1/Kb mice is that in these animals spontaneous tumors appear when animals are 20-22 months old. Therefore, the A2xneu mouse model provides a unique opportunity to evaluate antitumor immune responses against a self antigen where aging and tolerance are present at the same time. We have demonstrated that aging drastically altered the immune system of old mice. Therefore, it is critical to identify and optimize a vaccination strategy that effectively stimulates an immune response resulting in tumor rejection in young and old tolerant animals. There is plenty of evidence indicating that targeting APCs with different types of adjuvants results in the induction of an antitumor immune response. We compared the antitumor effect of different TLR-ligands (Poly I:C, LPS, flagellin, imiquimod, CpG-ODN) in young and old A2xneu mice. The result indicated that only intratumoral (i.t.) injections of CpG-ODN induced the rejection of tumors in young and old A2xneu mice. Although young and old A2xneu mice could reject the primary tumor after treatment with i.t injections of CpG-ODN, these animals could not develop a protective memory response. Analysis of the tumor microenvironment following i.t injections of CpG-ODN indicated: 1) activation of APCs; 2) a pro-inflammatory Th1 type response; 3) activation of CD4, CDS T cells and NK cells; and 4) the numbers of T-regs were drastically reduced. Taken together, these results indicate that i.t. injections of CpG-ODN influence the tumor microenvironment that favors the antitumor response. However, we do not yet completely understand how CpG-ODN injections induce the rejection of tumors. The goal of this proposal is to determine the mechanism of action of CpG-ODN in the induction of antitumor responses and, to develop a strategy to generate memory responses in old tolerant hosts. In these studies we will: 1) evaluate and characterize the effect of CpG-ODN on the induction of immune responses in old mice; 2) evaluate the relationship between CpG-ODN and T-regs, and the relationship of T-regs in regulating the repertoire and activation of immune responses in old mice; and 3) optimize the use of anti-neu-CpG-ODN to target the CpG-ODN at the tumor site for the treatment of tumors in old mice. Relevance: For the first time our results show that targeting CpG-ODN at the tumor site restores the immune response in old mice. Characterizing the effect of CpG-ODN on the immune system will ultimately lead to the optimization of this therapy. Furthermore, with the generation of the anti-neu-CpG-ODN molecule we can target the CpG-ODN anywhere in the body making it possible to develop a universal vaccine strategy for the treatment of cancer. Overall, the information gained from these studies will reveal strategies for controlling and manipulating the immune system to develop more effective immunotherapies in young and old tolerant hosts.