Pathobiology of Complement C3 effects in ADPKD

描述（由申请人提供）： 由于常染色体显性遗传性多囊肾病（ADPKD）的诊断不能排除，直到40岁，许多ADPKD患者是退伍军人。本提案的目的是确定靶向C3（补体途径的轴向组分）在ADPKD中的治疗潜力。与局部C3产生的新的重要作用的发现所导致的范式转变一致，我们的研究指出，肾内C3的产生是决定肾脏囊肿形成速度的途径的调节剂。我们基于我们对PKD模型中具有快速与缓慢囊性形成的肾脏的全基因组表达分析来阐述这一假设。我们通过证明：（i）在ADPKD和常染色体隐性（AR）PKD囊肿的肾小管细胞中存在C3 mRNA和蛋白，以及这些细胞激活C3的能力来支持这一假设;（ii）在ARPKD和ADPKD及其模型中，生物活性C3裂解产物的肾脏含量增加;（iii）在PKD模型中肾C3表达与肾囊肿形成的速度之间的强相关性以及在ADPKD患者中C3低活性基因变体与疾病进展之间的强相关性;和（iv）C3缺乏对遗传性PKD模型中肾囊肿形成的显著保护作用。虽然C3可能通过不同的途径起作用，但我们发现加速的囊肿形成与补体受体CR 3调节的途径特异性相关。CR 3（或Mac- 1）是C3片段iC 3b的主要受体，其在囊性肾中高度丰富，在单核细胞/巨噬细胞的分化、附着和存活中起核心作用。虽然我们已经确定了C3，巨噬细胞标志物CD 14和C3诱导因子MCP-1作为PKD结局的候选预测因子，但其他人已经证明，巨噬细胞耗竭通过减少囊性小管的增殖来减弱ADPKD的正位模型中的囊性生成。CR 3还可诱导促囊生成性TNF释放并直接激活肾小管细胞中的c-Src。由于肾小管细胞产生和激活C3时，液体流动是缺席，我们认为，C3在PKD的影响增加，因为囊性小管扩张，形成一个恶性的囊性循环。具体来说，我们假设C3途径激活通过CR 3依赖性过程加速ADPKD囊肿形成。我们从三个相互关联的目的来阐述这一假说：1）分析C3产生对Pkd 1通路的潜在致囊效应的机制; 2）确定肾小管衍生的C3在ADPKD发病机制中的作用。（3）确定补体成分受体CR 3（或Mac 1）在Pkd 1诱导的囊肿形成中的作用。这些目标的目的将通过整合以下各项来实现：（i）探究肾囊肿发生的新型调控机制的高度创新的研究设计，（ii）产生新型最先进的试剂（例如，用于条件C3靶向）。实现所提出的目标将允许通过将已建立的和新的致囊途径与C3-CR 3联系起来来整合现有知识。拟议的研究代表了实现我们长期目标的下一步，即开发一种安全的基于C3的ADPKD和其他肾脏疾病治疗方法。

DESCRIPTION (provided by applicant): Because the diagnosis of autosomal dominant polycystic kidney disease (ADPKD) cannot be excluded until age 40, many ADPKD patients are veterans. The objective of this proposal is to define therapeutic potential of targeting C3 (the axial component of complement pathway) in ADPKD. In line with paradigm shifts resulting from discoveries of novel essential roles of local C3 production, our studies point to intra-renal production of C3 as a regulator of a pathway that dictates the pace of renal cystogenesis. We formulated this hypothesis based on our genome-wide expression analyses of kidneys with rapid vs slow pace of cystogenesis in a PKD model. We supported this hypothesis by demonstrating: (i) presence of both C3 mRNA and protein in renal tubular cells that line ADPKD and autosomal recessive (AR) PKD cysts and the ability of these cells to activate C3; (ii) increased renal content of biologically active C3 split products i ARPKD and ADPKD and their models; (iii) strong correlation between renal C3 expression and pace of renal cystogenesis in a PKD model and C3 hypoactive gene variant with the disease progression among ADPKD patients; and (iv) significant protective effect of C3 deficiency on renal cystogenesis in genetic PKD models. While C3 may act through different pathways, we have found consistent association of accelerated cystogenesis specifically with the pathway regulated by complement receptor CR3. CR3 (or Mac- 1), a major receptor for C3 fragment iC3b, which is highly abundant in cystic kidneys, plays a central role in differentiation, attachment and survival of monocytes/macrophages. While we have identified C3, macrophage marker CD14 and C3-inducible factor MCP-1 as candidate predictors of PKD outcomes, others have demonstrated that macrophage depletion attenuates cystogenesis in orthologous models of ADPKD by reducing the proliferation of cystic tubules. CR3 may also induce pro-cystogenic TNF release and directly activate c-Src in renal tubule cells. Since renal tubule cells produce and activate C3 when fluid flow is absent we suggest that the C3 effects in PKD increase as cystic tubules dilate, forming a vicious cystogenic cycle. Specifically, we hypothesize that C3 pathway activation accelerates cyst formation in ADPKD through a CR3 dependent process. We address this hypothesis in three inter-related aims: 1) Dissect mechanisms underlying cystogenic effects of C3 production on Pkd1 pathway; 2) Determine effects of renal tubule-derived C3 in the pathogenesis of ADPKD.; and 3) Determine the role of complement component receptor CR3 (or Mac1) in Pkd1-induced cystogenesis. Objectives of these Aims will be accomplished by integrating: (i) highly innovative study design of interrogating novel regulatory mechanisms of renal cystogenesis with (ii) generation of novel state of the art reagents (e.g., for conditional C3 targeting). Achieving the proposed aims will allow integration of existing knowledge by linking established and novel cystogenic pathways to the C3-CR3 nexus. The proposed studies represent the next step for attaining our long-term goal of developing a safe C3-based therapy for ADPKD and other renal disorders.