Development of immune supression against gene product in gene therapy

治疗隐性遗传性皮肤病的基因治疗可能会引起针对引入的基因产物的不必要的免疫应答，因为患者中不存在针对这种新蛋白质的耐受性。在这项研究中，使用桥粒芯糖蛋白3基因敲除（Dsg 3-/-）小鼠作为DSG 3基因缺陷的疾病模型，我们调查了非病毒基因治疗是否引起针对Dsg 3的免疫反应，以及这种反应是否可以预防。当通过裸DNA注射将小鼠Dsg 3 cDNA引入Dsg 3-/-小鼠的皮肤中时，一些Dsg 3-/-小鼠产生了抗Dsg 3 IgG，其结合到由体内疗法表达的Dsg 3。为了克服这一问题，我们使用抗CD 40 L单克隆抗体（MR 1）来阻断CD 40-CD 40 L之间的共刺激相互作用，这在免疫应答的触发和维持中是重要的。对于该测定，我们在Dsg 3-/-小鼠上采用Dsg 3 +/+皮肤移植物来代表Dsg 3的稳定基因转移。皮肤移植后，所有受体Dsg 3-/-小鼠定期用MR 1或仓鼠IgG治疗。所有仓鼠IgG处理的小鼠在皮肤移植后2-3周产生循环抗Dsg 3 IgG。这种IgG的产生持续4周以上，并在移植物的角质形成细胞表面观察到IgG沉积。然而，当受体小鼠用MR 1处理时，这种抗Dsg 3 IgG产生被有效抑制。这些研究结果表明，这种不良的免疫反应的有效预防是需要一个成功的基因治疗隐性遗传性皮肤病和CD 40-CD 40 L相互作用的封锁可能是一个有价值的方式来实现这种预防。

Gene therapy to treat recessive genodermatoses may provoke unwanted immune response against the introduced gene product because tolerance against this novel protein is absent in patients. In this study, using desmoglein 3 knockout (Dsg3-/-) mice as a disease model for genetic defect of DSG3, we investigated whether immune response against Dsg3 is provoked by nonviral gene therapy and whether such reaction could be prevented. When mouse Dsg3 CDNA was introduced in the skin of Dsg3-/- mice by naked DNA injection, some Dsg3-/- mice developed anti-Dsg3 IgG which bound to Dsg3 expressed by the therapy in vivo. To overcome this problem, we used anti-CD40L monoclonal antibody (MR1) to block co-stimulatory interaction between CD40-CD40L, which is important in the triggering and maintenance of immune responses. For this assay, we employed the Dsg3+/+ skin graft on Dsg3-/- mice to represent stable gene transfer of Dsg3. After skin grafting, all recipient Dsg3-/- mice were treated either with MR1 or with hamster IgG regularly. All of hamster IgG treated mice developed circulating anti-Dsg3 IgG 2-3 weeks after the skin graft. This IgG production lasted: more than 4 weeks and IgG deposition was observed on the surfaces of the keratinocytes in grafts.; However, this anti-Dsg3 IgG production was efficiently suppressed when the recipient mice were treated with MR1. These findings indicate that effective prevention of such undesirable immune response is required for a successful gene therapy for recessive genodermatoses and that the blockade of CD40-CD40L interaction may be a valuable way to achieve this prevention.