Genome engineering therapeutics for cystinuria and its metabolic consequences.

摘要 胱氨酸尿症是一种遗传性人类肾脏疾病，发病率很高，每7000人中就有一人受到影响，其中包括退伍军人。 这种疾病是由于参与肾脏胱氨酸转运的基因突变导致尿液升高所致。 胱氨酸与肾结石形成。确定了疾病的遗传基础(SLC3A1中的突变， A型半胱氨酸尿症)，存在靶向分子治疗的机会。在我们之前拨款的基础上，我们 提出了一种创新的实验设计，以证明在一个长期的表型纠正胱氨酸尿症 利用转座子和腺相关病毒(AAV)技术相结合的完整动物。 在具体目标1中，我们将测试混合AAV-PiggyBac转座子技术对长期 转基因在小鼠肾脏中的表达和纠正SLC3a1-/-小鼠模型中的胱氨酸尿症。在具体目标2中，我们 将测试新型近端小管靶向AAV向小鼠近端肾小管细胞运送转基因的能力 活着。在这两个目标中，我们将测试分娩并评估新生儿和 成年小鼠。在具体目标3中，我们建议通过以下方式将我们的基础临床前研究与人类应用联系起来 测试AAV对人肾有机类物质的传递，包括胱氨酸尿症类有机物质。我们的创新但可行 研究将扩大AAV和转座子技术在肾脏基因传递中的应用，并为临床前奠定基础 为退伍军人和其他人的肾脏疾病最终人类治疗应用奠定基础。

ABSTRACT Cystinuria is an inherited human kidney disease with significant morbidity affecting 1 in 7000, including veterans. The disease is caused by mutation of genes involved in renal cystine transport resulting in elevated urinary cystine with kidney stone formation. With the genetic basis of the disorder defined (mutation in SLC3A1, cystinuria type A), opportunities for targeted molecular therapies exist. Building upon our previous grant, we propose an innovative experimental design to demonstrate long-term phenotypic correction of cystinuria in an intact animal using a combination of transposon and adeno-associated virus (AAV) technologies. In specific aim 1, we will test the ability of hybrid AAV-piggyBac transposon technology to mediate long-term transgene expression in mouse kidney and correct cystinuria in a Slc3a1-/- mouse model. In specific aim 2, we will test the ability of novel proximal tubule targeted AAV to deliver transgenes to proximal tubule cells in mice in vivo. In both aims, we will test delivery to and evaluate for long-term transgene expression in both neonatal and adult mice. In specific aim 3, we propose to bridge our foundational pre-clinical studies to human application by testing AAV delivery to human kidney organoids including cystinuria organoids. Our innovative but feasible studies will expand use of AAV and transposon technology for kidney gene delivery and lay a pre-clinical foundation for ultimate human therapeutic application for kidney disease in veterans and others.