Cell specific delivery of novel therapies to enhance glomerular regeneration and repair

项目摘要/摘要 该项目的目标是改变蛋白尿性肾小球疾病的治疗模式，通过联合 细胞特异性递送疗法的发展，以增强体内足细胞的修复和再生。 足细胞是一种高度特化的终末分化上皮细胞，在大多数情况下都会受到损伤 肾小球疾病。由于足细胞不能自我更新，足细胞丢失会导致肾小球瘢痕形成。一个 顶叶上皮细胞(PECs)亚群可作为足细胞干细胞(PEC前体细胞)，但其 再生潜力不足以克服与疾病相关的肾小球损伤。增强 因此，足细胞的生产性修复需要双重同步的方法：(I)将丢失的足细胞替换为 增加它们的数量，以及(Ii)限制/逆转对剩余足细胞的损害。然而，梅杰 知识差距阻碍了我们实现这些目标；其中包括我们对分子的有限知识。 刺激PEC自我更新和足细胞再生/修复的因素以及可选的方法 将这些因子传递给体内特定类型的肾脏细胞。 我们由四名专家调查人员组成的团队将利用互补的工具来弥合这些知识差距，并 产生创新的疗法。韦斯利博士将应用实验设计(DoE)方法来识别小说 增加PEC祖细胞和减少足细胞损失的分子组合；罗伯茨博士将结合 这些疗法用于将VHH(纳米体)输送给PEC祖细胞和足细胞；弗里德曼博士将 生成经过基因编辑的人类肾脏器官类化合物，以验证VHH的效果与临床数据的比较 患者；Shankland博士将使用足细胞耗竭和人类器官样体的血统追踪动物模型 移植到小鼠肾脏进行体内安全性和有效性分析。这条管道最终将测试 靶向递送PEC和足细胞特异性治疗货物可以增强足细胞的假说 体内修复和再生，将肾小球功能恢复到临床疾病阈值以下。 这项工作将通过两个具体目标来完成，每个目标都有独特的里程碑。第一个目标是 通过细胞靶向递送新型多肽和小分子药物组合促进体内肾小球再生 增强壁上皮细胞起源的足细胞前体的分子。第二个目标是增加 通过新确定的治疗方法的细胞型特异性传递对受损足细胞的生产性修复。对两个人都是 AIMS，我们将使用上述渠道来发现美国能源部的候选疗法，并与 来自人类患者的肾小球疾病签名。这些将与来自的特定细胞类型的VHH结合 高多样性重组VHH文库，选择性地将其输送到人PECs(Aim 1)或足细胞(Aim 2)。在FSGS的动物模型和移植的人类中将证实体内增强的再生 有机化合物。这一过程将建立一种治疗肾脏疾病的新范式，并产生铅 进一步的临床前开发和最终的人类临床试验的候选治疗方法。

PROJECT SUMMARY/ABSTRACT The goal of this project is to change the treatment paradigm for proteinuric glomerular diseases by combining therapeutics development with cell-specific delivery to enhance podocyte repair and regeneration in vivo. Podocytes, highly specialized terminally differentiated epithelial cells, are injured in the majority of glomerular diseases. As podocytes cannot self-renew, podocyte loss leads to glomerular scarring. A subpopulation of parietal epithelial cells (PECs) can serve as podocyte stem cells (`PEC progenitors'), but their regenerative potential is insufficient to overcome disease-associated glomerular damage. Enhancing productive repair of podocytes thus requires a dual synchronized approach: (i) replacing lost podocytes to increase their number, and (ii) limiting/reversing damage to the remaining podocytes. However, major knowledge gaps prevent us from achieving these goals; these include our limited knowledge on the molecular factors stimulating PEC self-renewal and podocyte regeneration/repair, as well as options methods for delivering these factors to specific kidney cell types in vivo. Our team of four expert investigators will wield complementary tools to close these knowledge gaps and produce innovative therapies. Dr. Wessely will apply Design of Experiment (DoE) approaches to identify novel combinations of molecules that increase PEC progenitors and reduce podocyte loss; Dr. Roberts will conjugate these therapeutics to VHHs (nanobodies) for delivery to PEC progenitors and podocytes; Dr. Freedman will generate gene-edited human kidney organoids to validate effects of VHHs compared to clinical data from patients; Dr. Shankland will use lineage tracing animal models of podocyte depletion and human organoids transplanted into mouse kidneys for in vivo safety and efficacy analysis. This pipeline will ultimately test the hypothesis that targeted delivery of PEC- and podocyte-specific therapeutic cargos can enhance podocyte repair and regeneration in vivo, and restore glomerular function to below the clinical disease threshold. The work will be accomplished through two Specific Aims, each with unique Milestones. The first Aim is to increase glomerular regeneration in vivo by cell targeted delivery of novel combinations of peptides and small molecules to augment podocyte progenitors of parietal epithelial cell origin. The second Aim is to increase productive repair of damaged podocytes by cell-type specific delivery of newly identified therapies. For both aims, we will employ the above pipeline to discover candidate therapeutics by DoE and cross-referenced with glomerular disease signatures from human patients. These will be combined with cell type-specific VHHs from high diversity recombinant VHH libraries to selectively deliver them to human PECs (Aim 1), or podocytes (Aim 2). Enhanced regeneration in vivo will be demonstrated in animal models of FSGS and transplanted human organoids. This process will establish a new paradigm for the treatment of kidney disease, and produce lead therapeutic candidates for further pre-clinical development and ultimately human clinical trials.