Immunotherapy with Dendritic-Allogeneic Tumor Cells

描述（由申请人提供）：树突状细胞（DC）诱导原发性T细胞应答，并作为癌症免疫治疗中抗原（Ag）递送的载体进行了广泛评价。一种策略采用通过将DC与肿瘤细胞融合产生的杂交细胞的使用。从技术上讲，使用化学融合剂如聚乙二醇（PEG）或病毒的融合一直受到效率低、毒性和重现性差的困扰。我们最近开发了一种电融合协议，异核体产生高融合率。在临床前研究中，融合细胞刺激IFN？对于免疫疗法，单次疫苗接种导致肿瘤消退。基于这些观察结果，我们最近设计了一项临床试验，通过自体DC和同种异体黑色素瘤细胞系（PMCV或CanvaxinTU）的融合疫苗接种治疗转移性黑色素瘤。在该方案中，3种选择的同种异体肿瘤细胞提供肿瘤相关Ag的来源。然而，很少有实验室分析集中在定义这种方法的免疫原性和潜在的治疗效果。由于对同种异体肿瘤细胞上的组织不相容的allo-Ag的免疫应答，对肿瘤Ag的免疫应答可能受到下调或上调。同种异体肿瘤疫苗的作用机制有待进一步研究。在本提案中，将使用两种动物肿瘤系统来开发同种异体疫苗。第一个系统将利用β-半乳糖苷酶（β-gal）作为替代肿瘤排斥Ag，该Ag已被引入B16/F10（H-2b）黑色素瘤和4 T1（H-2d）乳腺癌。第二个系统由三个小鼠黑色素瘤[B16（H-2b），K1735（H-2k）和cloudman S91（H-2d）]组成，它们具有不同的单倍型，但共享许多天然肿瘤相关的Ag。这些同种异体肿瘤将与同基因DC电融合。它们的分子特征、免疫原性和治疗效果将针对同基因肿瘤进行分析。具体目标是：1）确定通过电融合产生的同基因DC-同种异体肿瘤（allo-DC-tumor）杂合体的免疫原性和治疗功效; 2）表征和分析对肿瘤相关Ag和对同种异体组织相容性Ag的免疫应答; 3）针对晚期肿瘤的主动免疫疗法;以及4）从用allo-DC-tumor杂合体细胞免疫的小鼠产生肿瘤免疫T细胞用于过继免疫疗法。

DESCRIPTION (provided by applicant): Dendritic cells (DCs) induce primary T cell responses and are being extensively evaluated as vehicles for antigen (Ag) delivery in cancer immunotherapy. One strategy employs the use of hybrid cells generated by fusing DCs with tumor cells. Technically, fusion using chemical fusogens such as polyethylene glycol (PEG) or viruses has been plagued by low efficiency, toxicity and poor reproducibility. We recently developed an electrofusion protocol with which heterokaryons are generated with high fusion rates. In pre-clinical studies, fusion cells stimulated IFN? secretion from both CD4 and CD8 immune T cells, and for immunotherapy, a single vaccination resulted in tumor regression. Based on these observations, we have recently designed a clinical trial for the treatment of metastatic melanoma by vaccination with fusion of autologous DCs and allogeneic melanoma cell lines (PMCV or CanvaxinTU). In this protocol, 3 selected allogeneic tumor cells provide a source of tumor-associated Ags. However, little laboratory analysis has focused on defining the immunogenicity and potential therapeutic effects of this approach. Because of the immune response to histo-incompatible allo-Ags on allogeneic tumor cells, immune responses to tumor Ags may be subjected to down- or up-regulation. The mechanisms of allogeneic tumor vaccine need to be analyzed. In this proposal, two animal tumor systems will be used for the development of allogeneic vaccine. The first system will utilize beta-galactosidase (beta-gal) as a surrogate tumor rejection Ag that has been introduced into the B16/F10 (H-2b) melanoma and 4T1 (H-2d) breast carcinoma. The second system consists of three murine melanomas [B16 (H-2b), K1735 (H-2k) and cloudman S91 (H-2d)] of different haplotypes but sharing a number of natural tumor-associated Ags. These allogeneic tumors will be electrofused with syngeneic DCs. Their molecular characteristics, immunogenicities, and therapeutic effects will be analyzed against syngeneic tumors. The specific aims are: 1) To determine the immunogenicity and therapeutic efficacy of syngeneic DC-allogeneic tumor (allo-DC-tumor) hybrids generated by electrofusion; 2) To characterize and analyze immune responses to tumor-associated Ags and to allogeneic histocompatibility Ags; 3) Active immunotherapy against advanced tumors; and 4) To generate tumor-immune T cells from mice immunized with allo-DC-tumor hybrid cells for adoptive immunotherapy.