GENE THERAPY FOR PHENYLKETONURIA

苯丙酮尿症（PKU）是影响北美和欧洲儿童最常见的遗传性疾病之一，发病率约为万分之一。苯丙氨酸羟化酶（PAH）的突变是PKU的主要原因。多环芳烃缺乏导致血清苯丙氨酸（Phe）水平升高，导致大脑发育异常和智力迟钝。PKU的频繁发生，加上早期饮食调整的能力，大大减少了与该疾病相关的严重神经系统问题，导致了所有州的新生儿测试项目。不幸的是，人们普遍认为控制饮食可以“治愈”这种疾病。实际上，严格遵守饮食是非常困难的，失误会导致严重的长期神经发育后遗症，特别是在pah缺乏妇女的后代中。鉴于已知的单基因缺陷，基因治疗PKU在技术上是可行的，在治疗上也是可取的。近年来基因治疗的重要进展来自于利用基于aav的载体提供有效、稳定和安全的代谢校正。假设AAV载体在PKU中是有用的。根据这一假设，我们将检验AAV载体是否能够：1)有效地挽救Pah/enu2小鼠的Phe代谢缺陷。血清Phe水平将与载体基因组和转染细胞的数量以及PKU症状的减轻程度有关。2)将在成年和新生PAH /enu2小鼠中进行载体序列非种系传播的PAH肝安全传递试验。还将检查对病媒的肝毒性和免疫反应的可能性。3)通过使用替代控制元件和组织特异性靶向优化PAH表达和肝细胞递送。这些实验为检验aav驱动基因治疗真正治愈PKU的潜力提供了一种合理的方法。

Phenylketonuria (PKU) is one of the most common genetic disorders affecting children in North America and Europe, with an incidence of about 1 in 10,000 births. Mutations in the enzyme phenylalanine hydroxylase (PAH) are the major cause of PKU. PAH deficiency results in elevated serum phenylalanine (Phe) levels, leading to abnormal brain development and mental retardation. PKU's frequency, coupled with the ability of early dietary modification to greatly reduce the severe neurological problems associated with the disorder, has led to neonatal testing programs in all states. Unfortunately, there is a widespread impression that dietary control effects a "cure" for this disease. In reality, strict adherence to the diet is exceedingly difficult and lapses can cause serious long-term neuro-development sequelae, particularly in offspring of PAH-deficient women. Given the well-understood single gene defect, gene therapy for PKU is both technically feasible and therapeutically desirable. Recent important advances in gene therapy have come from the use of AAV-based vectors to provide effective, stable, and safe correction of metabolic. It is hypothesized that AAV vectors will be useful in PKU. To this hypothesis, the ability of AAV vectors to: 1) Provide effective rescue of defective Phe metabolism in the Pah/enu2 mouse will be examined. Serum Phe levels will be related to the number of vector genomes and transfected cells as well as to the degree of reduction of PKU symptoms. 2) Give safe PAH delivery to liver with non germ-line transmission of vector sequences in either adult and neonatal Pah/enu2 mice will be tested. The potential for liver toxicity and immune responses to vector will also be examined. 3) Optimize PAH expression and hepatocyte delivery by using alternative control elements and tissue-specific targeting. These experiments provide a rational approach to examining the potential of AAV-driven gene therapy to truly cure PKU.