INITIATING THE ADAPTIVE IMMUNE RESPONSE TO CANCER

越来越清楚的是，NK细胞可以促进DC活化和随后免疫应答的极化。本项目将在可移植和自发性小鼠肿瘤模型中评价NK-DC相互作用对抗肿瘤CTL的产生和功能的意义，并将通过控制/操纵这些相互作用来提高对已建立的MHC+/MHC-肿瘤的抗肿瘤活性。我们假设，活化的NK（A-NK）细胞，交付到肿瘤部位，将杀死肿瘤细胞，从而释放急性炎症介质（如HSP和HMGB 1）和抗原，可以采取的DC。此外，DC和NK细胞之间的接触依赖性和烟碱依赖性串扰将进一步激活NK细胞。 这两种细胞类型，导致产生具有增强的刺激Th 1-抗肿瘤应答能力的极化DC。这将在三种动物模型中进行评估：1）模型肿瘤抗原表达系统（MO-5，卵清蛋白转染的B16黑素瘤）。在该模型中，将通过测量肿瘤、淋巴结、血液和脾脏中过继转移的卵特异性OT-I（CDS）和OT-II（CD 4）细胞的数量来评价有效的NK /DC相互作用; 2）野生型B16模型和3）自发性黑素瘤的p16 INK 4a-/-KO模型。在全身或局部递送A-NK细胞和DC后，我们将改变肿瘤微环境中的细胞因子谱。为了实现这一点，DC和NK细胞将用含有细胞因子基因的病毒载体转导以产生各种pro-NK和pro-DC细胞因子，例如，IL2、IL18、IFN-α和GM-CSF。此外，将测试多次NK/DC注射以确保连续刺激传入肢体（从而产生CTL）以及局部CTL效应子功能的能力。将通过三个具体目标来探讨这些问题：目标1。通过不同途径将不同比例的NK/DC注射到荷瘤动物体内后，确定NK细胞和DC的体内存活率以及它们向肿瘤和淋巴结的运输。目标二。证明NK细胞和DC的肿瘤内递送促进可移植和自发性肿瘤模型中CTL的产生和功能。目标3：通过对NK细胞和DC的强制性酪氨酸基因转导增强肿瘤中NK/DC的功能。

It has become increasingly clear that NK cells can promote DC activation and polarization of the subsequent immune response. This project will evaluate, in transplantable and spontaneous murine tumor models, the significance of NK-DC interactions for the generation and function of anti-tumor CTLs and will improve antitumor activity against established MHC+/MHC- tumors by controlling/manipulating these interactions. We hypothesize that activated NK (A-NK) cells, delivered to tumor sites, will kill tumor cells and thereby liberate both mediators of acute inflammation (such as HSP and HMGB1) and antigens which can be taken up by DCs. In addition, contact- and cytokine-dependent cross-talk between DCs and NK cells will further activate both cell types, leading to the generation of polarized DCs with enhanced capability of stimulating Th1-anti tumor responses. This will be evaluated in three animal models: 1) a model tumor antigen expressing system (the MO-5, ovalbumin transfected B16 melanoma). Effective NK /DC interactions will, in this model be evaluated by measuring the number of adoptively transferred ova-specific OT-I (CDS) and OT-II (CD4) cells in tumors, lymph nodes, blood and spleen; 2) The wild-type B16 model and; 3) in the p16INK4a-/-KO model of spontaneous melanoma. Following systemic or local delivery of A-NK cells and DCs, we will vary the cytokine profile in the tumor microenvironment. To achieve this, DCs and NK cells will be transduced with cytokine gene-containing viral vectors to produce various pro-NK and pro-DC cytokines, e.g., IL2, IL18, IFN-alpha and GM-CSF. In addition, the ability of multiple NK/DC injections to ensure continuous stimulation of the afferent limb (and thereby resultant CTL generation) as well as local CTL effector function, will be tested. Three Specific Aims will be conducted to explore these issues: Aim 1. Define the in vivo survival of NK cells and DCs as well as their traffic to tumors and lymph nodes following injection different NK/DC ratios into tumor-bearing animals via different routes. Aim 2. Demonstrate that intratumoral delivery of NK cells and DCs promotes CTL generation and function in transplantable and spontaneous tumor models.; Aim 3. Enhance NK/DC function in tumors by forced cytokine-gene transduction of NK cells and DCs.