Division of Medical Genetics, Department of Pediatrics, Duke University Medical Center, Durham, NC 27710, USA.
Hum Gene Ther. 2012 May;23(5):460-72. doi: 10.1089/hum.2011.063. Epub 2012 Mar 29.
Pompe disease can be treated effectively, if immune tolerance to enzyme replacement therapy (ERT) with acid α-glucosidase (GAA) is present. An adeno-associated viral (AAV) vector carrying a liver-specific regulatory cassette to drive GAA expression (AAV-LSPhGAA) established immune tolerance in GAA knockout (KO) mice, whereas ubiquitous expression with AAV-CBhGAA provoked immune responses. Therefore, we investigated the hypothesis that immune tolerance induced by hepatic-restricted expression was dominant. AAV-LSPhGAA and AAV-CBhGAA were administered singly or in combination to groups of adult GAA-KO mice, and AAV-LSPhGAA induced immune tolerance even in combination with AAV-CBhGAA. The dual vector approach to GAA expression improved biochemical correction of GAA deficiency and glycogen accumulations at 18 weeks, and improved motor function testing including wire-hang and grip-strength testing. The greatest efficacy was demonstrated by dual vector administration, when both vectors were pseudotyped as AAV8. T cells from mice injected with AAV-LSPhGAA failed to proliferate at all after an immune challenge with GAA and adjuvant, whereas mock-treated GAA-KO mice mounted vigorous T cell proliferation. Unlike AAV-LSPhGAA, AAV-CBhGAA induced selective cytokine and chemokine expression in liver and spleen after the immune challenge. AAV-CBhGAA transduced dendritic cells and expressed high-level GAA, whereas AAV-LSPhGAA failed to express GAA in dendritic cells. The level of transduction in liver was much higher after dual AAV8 vector administration at 18 weeks, in comparison with either vector alone. Dual vector administration failed to provoke antibody formation in response to GAA expression with AAV-CBhGAA; however, hepatic-restricted expression from dual vector expression did not prevent antibody formation after a strong immune challenge with GAA and adjuvant. The relevance of immune tolerance to gene therapy in Pompe disease indicates that hepatic expression might best be combined with nonhepatic expression, achieving the benefits of ubiquitous expression in addition to evading deleterious immune responses.
庞贝病如果存在酸α-葡萄糖苷酶（GAA）酶替代治疗（ERT）的免疫耐受，可以得到有效治疗。携带肝脏特异性调控盒以驱动 GAA 表达的腺相关病毒（AAV）载体（AAV-LSPhGAA）在 GAA 敲除（KO）小鼠中建立了免疫耐受，而用 AAV-CBhGAA 进行普遍表达则会引发免疫反应。因此，我们提出假设，肝脏特异性表达诱导的免疫耐受占主导地位。AAV-LSPhGAA 和 AAV-CBhGAA 分别或联合给药于成年 GAA-KO 小鼠，AAV-LSPhGAA 诱导免疫耐受，即使与 AAV-CBhGAA 联合给药也是如此。GAA 表达的双载体方法改善了 18 周时 GAA 缺乏和糖原积累的生化纠正，并且改善了包括线悬挂和握力测试在内的运动功能测试。当两种载体均为 AAV8 假型时，双载体给药显示出最大的疗效。用 GAA 和佐剂进行免疫挑战后，用 AAV-LSPhGAA 注射的小鼠的 T 细胞完全没有增殖，而模拟处理的 GAA-KO 小鼠则产生了强烈的 T 细胞增殖。与 AAV-LSPhGAA 不同，用 GAA 和佐剂进行免疫挑战后，AAV-CBhGAA 仅在肝脏和脾脏中诱导选择性细胞因子和趋化因子表达。AAV-CBhGAA 转导树突状细胞并表达高水平的 GAA，而 AAV-LSPhGAA 未能在树突状细胞中表达 GAA。与单独使用任一载体相比，18 周时，双 AAV8 载体给药后，肝脏中的转导水平更高。双载体给药未能引起针对 AAV-CBhGAA 表达的抗体形成；然而，双载体表达的肝脏特异性表达并不能阻止用 GAA 和佐剂进行强烈免疫挑战后的抗体形成。免疫耐受对庞贝病基因治疗的相关性表明，肝脏表达最好与非肝脏表达相结合，除了避免有害的免疫反应外，还能实现普遍表达的益处。
