Identification of new therapeutic targets for ADPKD

ADPKD 新治疗靶点的确定

• 批准号: 10298937
• 负责人: EDWARD Y SKOLNIK
• 金额: $ 50.82万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10298937
• 关键词: Affect; Affinity; Animal Model; Autosomal Dominant Polycystic Kidney; Binding; Biological Assay; Clinical Trials; Collaborations; Combined Modality Therapy; Couples; Cyclic GMP; Cyst; Data; Doxycycline; Drug Targeting; End stage renal failure; Epithelial cyst; Extracellular Matrix; Fibrosis; Focal Adhesion Kinase 1; Funding; Genes; Genetic; Genetic Transcription; Goals; Grant; Growth; Human; Kidney; Kidney Diseases; Lipids; Mass Spectrum Analysis; Mediating; Medicine; Messenger RNA; Mus; Mutation; PTK2 gene; Pathogenesis; Patients; Pharmaceutical Preparations; Pharmacology; Phosphotransferases; Play; Proprotein Convertase 1; Proprotein Convertase 2; Proteins; Renal function; Role; Signal Pathway; Signal Transduction; Techniques; Technology; Testing; Therapeutic; Time; Toxic effect; Translating; Universities; base; discoidin domain receptor 1; drug candidate; genetic approach; human disease; in vivo; inhibitor/antagonist; insight; interest; interstitial; kinase inhibitor; mouse model; new therapeutic target; novel; novel strategies; polycystic kidney disease 1 protein; preservation; tandem mass spectrometry; therapeutic target; tolvaptan

Autosomal-dominant polycystic kidney disease (ADPKD) affects more than 12 million people worldwide and is a common cause of end-stage kidney disease (ESKD). ADPKD is caused by mutations in one of two genes, PKD1 or PKD2, which encodes polycystin 1 (PC) and PC2 respectively. Loss of PC1 or PC2 results in activation of numerous kinases and downstream signaling pathways, which is central to the pathogenesis of cyst growth in ADPKD. In addition, pharmacologically inhibiting a number of different kinases up-regulated in PKD kidneys has been shown to slow cyst growth in animal models of PKD making kinase inhibitors among the most promising class of drug candidates to treat patients with ADPKD. However, while the human kinome consists of more than 500 kinases, only a fraction of these kinases have been tested to determine if they play a role in ADPKD pathogenesis. As a result, there are likely many kinases that are active in ADPKD kidneys that play prominent roles in cyst growth that are yet-to-be discovered and may be good therapeutic targets. In collaboration with James Duncan at Temple University, we have now adapted a novel approach to broadly screen PKD kidneys in an unbiased manner for kinases that are more active in lysates from PKD kidneys compared with lysates from wild type kidneys. In this assay, active kinases are affinity captured by passing lysates over multiplex inhibitor beads (MIB) containing a cocktail of kinase inhibitors. Bound kinases are then identified by LC separation followed by quantitative tandem mass spectrometry (LC- MS/MS). Using this approach, we have now identified a number of both known and unknown kinases specifically up-regulated and down-regulated in PKD kidneys. The major goal of this proposal is to assess the role of several of the kinases identified thus far whose activity is increased in PKD kidneys and determine whether inhibiting or activating any of the kinases identified slows cyst growth, inhibits interstitial fibrosis, and preserves renal function in animal models of ADPKD. We will then take both biased and unbiased approaches to identify the signaling pathways regulated by these kinases that are critical to pathogenesis with the goal of developing a more complete picture of the relevant signaling hubs and networks that are aberrantly activated in PKD kidneys. In addition, we will use this technology to broadly screen PKD kidneys from a variety of different “early” and “late” mouse models of ADPKD at different stages in cyst formation and in kidneys from human patients with ADPKD to identify in an unbiased manner additional kinases that are activated and inhibited in PKD kidneys to determine the similarities and differences between mouse models, which kinases may be most relevant to human disease, and whether distinct sets of kinases are activated early post cyst induction and function as early “drivers” of cyst growth. Ultimately, we hope this new information will identify new safe drug targets and rational approaches to combination therapies to slow cyst growth that can then be translated into clinical trials.

常染色体显性多囊肾病 (ADPKD) 影响着全世界超过 1200 万人， 终末期肾病 (ESKD) 的常见原因。 ADPKD 是由两个基因之一的突变引起的， PKD1 或 PKD2，分别编码多囊蛋白 1 (PC) 和 PC2。 PC1 或 PC2 丢失导致 许多激酶和下游信号通路的激活，这是发病机制的核心 ADPKD 中的囊肿生长。此外，在药理上抑制许多不同的激酶上调 PKD 肾脏已被证明可以减缓 PKD 动物模型中的囊肿生长，从而使激酶抑制剂 最有希望治疗 ADPKD 患者的候选药物。然而，虽然人类激酶组 由 500 多种激酶组成，只有其中一小部分经过测试以确定它们是否发挥作用 ADPKD 发病机制中的作用。因此，ADPKD 肾脏中可能有许多活跃的激酶 在囊肿生长中发挥着尚未被发现的重要作用，并且可能是良好的治疗靶点。 通过与坦普尔大学的詹姆斯·邓肯合作，我们现在采用了一种新颖的方法 以公正的方式广泛筛选 PKD 肾脏，寻找在裂解物中更活跃的激酶 PKD 肾脏与野生型肾脏裂解物的比较。在此测定中，活性激酶具有亲和力 通过将裂解物通过含有激酶抑制剂混合物的多重抑制剂珠 (MIB) 来捕获。 然后通过 LC 分离和定量串联质谱法（LC- 质谱/质谱）。使用这种方法，我们现在已经鉴定了许多已知和未知的激酶 在 PKD 肾脏中特别上调和下调。 该提案的主要目标是评估迄今为止已确定的几种激酶的作用，其活性 在 PKD 肾脏中增加，并确定抑制或激活任何已识别的激酶是否会减慢 在 ADPKD 动物模型中抑制囊肿生长、抑制间质纤维化并保留肾功能。我们随后将 采用有偏和无偏的方法来识别这些激酶调节的信号通路 对发病机制至关重要，目的是更全面地了解相关信号中枢 以及 PKD 肾脏中异常激活的网络。此外，我们还将利用这项技术来广泛 从 ADPKD 不同阶段的各种不同“早期”和“晚期”小鼠模型中筛选 PKD 肾脏 囊肿形成和 ADPKD 人类患者的肾脏中，以公正的方式识别额外的 在 PKD 肾脏中被激活和抑制的激酶，以确定两者之间的异同 小鼠模型，哪些激酶可能与人类疾病最相关，以及是否有不同的激酶组 在囊肿诱导后早期被激活，并作为囊肿生长的早期“驱动器”发挥作用。最终，我们希望这 新信息将确定新的安全药物靶点和联合疗法的合理方法，以减缓 囊肿生长，然后可以转化为临床试验。