Gene therapy for cancer using intratumoral injection of dendritic cells genetically modified to express interleukin 12

树突状细胞是一种有效的专业抗原呈递细胞，在细胞免疫中起着关键作用。我们之前已经证明，肿瘤内注射经白细胞介素(IL)-12基因改造的骨髓来源的dc可以诱导特异性抗肿瘤免疫反应。在这项研究中，我们研究了OK-432刺激的DCs是否可以以同样的方式使用。体内给药重组IL-12可使NK细胞和T细胞分泌IFN-g，增强对肿瘤细胞的细胞溶解功能。然而，这些细胞因子诱导活化的巨噬细胞大量分泌一氧化氮（NO），并抑制小鼠系统的细胞免疫。OK-432已被临床用作治疗癌症患者的有效生物调节剂，已知在体外可诱导多种细胞因子，包括IFN-g和IL-12。在C57BL/6小鼠皮内肿瘤模型中，I.t.注射含有或不含OK-432的骨髓源性dc对第7天建立的肿瘤有轻微的抗肿瘤作用。然而，当DCs， OK-432和n -硝基- l -精氨酸甲酯（L-NAME）联合治疗时，观察到有效的抗肿瘤作用，其抑制诱导型一氧化氮合酶（iNOS）介导的NO生成。此外，用这种组合治疗的小鼠的脾细胞可以获得肿瘤特异性和强效的细胞毒性T淋巴细胞（ctl）。这些结果表明，巨噬细胞和树突状细胞经OK-432刺激后大量分泌NO，可强烈抑制小鼠肿瘤特异性细胞免疫。因此，如果NO的产生得到适当调节，注射dc和OK-432可能是一种很有希望的癌症免疫治疗方法。

Dendritic cells (DCs) are potent professional antigen presenting cells and play a key role in cellular immunity. We have previously demonstrated that specific antitumor immune response can be induced with intratumoral (I.t.) injection of bone marrow-derived DCs genetically engineered with interleukin (IL)-12 genes. In this study, we examined whether DCs stimulated with OK-432 could be used in the same manner. In vivo administration of recombinant IL-12 causes NK cells and T cells to secrete IFN-g and enhances the cytolytic functions against tumor cells. These cytokines, however, induce abundant secretion of nitric oxide (NO) from activated macrophages, and appear to suppress cellular immunity in murine system. OK-432 has been clinically used as a potent biological modifiers for treating cancer patients and is known to induce multiple cytokines including IFN-g and IL-12 in vitro. In intradermal tumor modeles using MCA205 into C57BL/6 mice, I.t. injection bone marrow-derived DCs with or without OK-432 showed marginal antitumor effects on day 7 established tumors. However, potent antitumor effects were observed when they are treated with the combination of DCs, OK-432, and N-nitro-L-arginine methyl ester (L-NAME), which inhibits inducible nitric oxide synthase (iNOS)-mediated NO production. Furthermore, tumor specific and potent cytotoxic T lymphocytes (CTLs) could be obtained from the splenocytes of the mice treated with this combination. These results suggested that tumor specific cellular immunity in mice could be strongly suppressed with NO, which is abundantly secreted by macrophages and DCs stimulated with OK-432. Thus, I.t. injection of DCs and OK-432 could be a promisingly cancer immunotherapy if NO production is properly regulated.