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，发病率高达1/7000，包括退伍军人。 这种疾病是由参与肾胱氨酸转运的基因突变引起的，导致尿中胱氨酸水平升高。 胱氨酸与肾结石形成。随着疾病的遗传基础的定义（SLC 3A 1突变， 胱氨酸尿症A型），存在靶向分子治疗的机会。在我们之前的基础上，我们 提出了一个创新的实验设计，以证明长期的表型纠正胱氨酸尿症， 使用转座子和腺相关病毒（AAV）技术组合， 在具体的目标1中，我们将测试混合AAV-piggyBac转座子技术介导长期表达的能力。 小鼠肾脏中的转基因表达和Slc 3a 1-/-小鼠模型中的纠正胱氨酸尿症。在具体目标2中，我们 将测试新的近端小管靶向AAV在小鼠中将转基因递送至近端小管细胞的能力， vivo.在这两个目标，我们将测试交付和评估长期转基因表达在新生儿和 成年老鼠在具体目标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向人肾类器官（包括胱氨酸尿症类器官）的递送。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递送中的应用，并为临床前研究奠定基础。 为退伍军人和其他人的肾脏疾病的最终人类治疗应用基金会。

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.