Establisment of tumor vaccine using autologous dendritic cells and its application for addaptive immunotherapy

树突状细胞（DC）在各种抗原呈递细胞中具有最强的抗原呈递活性。本研究旨在将自体树突状细胞用于肿瘤疫苗的临床前应用。我们尝试用自体结肠癌细胞和树突状细胞建立自体树突状细胞疫苗，并着重研究有效诱导自体肿瘤特异性T细胞的方法和肿瘤特异性树突状细胞培养体系。我们首先从38年开始建立结肠癌细胞系CE-1。男性晚期结肠癌患者。这些肿瘤细胞显示hla - I型A31和A11。我们尝试用自体细胞和hla匹配的健康志愿者建立肿瘤特异性的腺突细胞。从100 ml肝素化的外周血和含有IL-4和GM-CSF的培养基中获得1-2 × 10^6个树突状细胞。将辐照或冻融后的肿瘤细胞加入到树突状细胞中，并在IL-4和GM-CS - F存在下进一步培养，获得肿瘤特异性树突状细胞。这些细胞IL-12和ifn - γ mRNA表达阳性，HLA-DR、CD80和ICAM-1表达阳性。将自体T细胞加入肿瘤脉冲树突状细胞中，在IL-2和IL-4存在下培养2周，获得自体肿瘤反应性CTL。因为这些T细胞对释放ifn - γ的自体肿瘤细胞具有细胞毒活性。然而，我们未能使用明胶珠大量培养树突状细胞，这是本研究的目的之一。需要进一步的研究来建立和验证有效的系统来培养自体肿瘤脉冲树突状细胞疫苗。

Dendritic cells (DC) possess the most potent antigen-prersenting activity among various antigen presenting cells. This study aims preclinical application of tumor vaccine using autologous dendritic cells. We tried to establish autologous dendritic cell vaccine using autologous colon cancer cells and dendritic cells, and focused on investigation of the methods for effective induction of autolpogpus tumor spesific T cells and culture system for tumor-spesific dendritic cells. We first established colon carcinoma cell line, CE-1, from the 38 years. male patient with advanced colon cancer. These tumor cells revealed HLA-type Class I A31, and A11. We tried to establish tumor-spesific adendritic cells using autologouis cells and HLA-matched healthy volunteer. 1-2 x 10^6 dendritic cells were obtained from 100 ml of heparinized peripheral blood and culture media containing IL-4 and GM-CSF.When irradiated or freezing-thawing tumor cells were added to the dendritic cells and furtherly cultured in the presence of IL-4 and GM-CS F, tumor-spesific dendritic cells were obtained. These cells were positive for expression of IL-12 and IFN-gamma mRNA and the expression of HLA-DR, CD80, and ICAM-1.When autologous T cells were added to the tumor-pulsed dendritic cells and cultured in the presence of IL-2 and IL-4 for subsequent 2 weeks, autologous tumor-reactive CTL were obtained. Because theses T cell possess cytotoxic activity against autologous tumor cells releasing IFN-gamma. However we failed in culture of dendritic cells in a large number using gellatin-beads, which was one of aims of this study. Further studies are needed to establish and verify effective systems for culture of autologpus-tumor pulsed dendritic cell vaccine.