Targeted delivery of TGFbeta blockade to diseased glomeruli

描述(申请人提供)：终末期肾病的发病率和流行率正在以惊人的速度增长，迫切需要新的创新疗法来阻止慢性肾功能衰竭的进展。转化生长因子-β1的过度表达是进展性肾脏疾病最常见的分子特征。它已被确定为纤维化肾脏疾病的主要关键介质和可量化的指标。转化生长因子1似乎是治疗慢性肾脏疾病的一个有前途的靶点。然而，转化生长因子具有深刻的抗炎特性，而且转化生长因子基因缺陷的动物在出生后不久就死于压倒性炎症，这增加了全身阻断转化生长因子可能会产生严重炎症副作用的可能性。不幸的是，所有转化生长因子阻断的人类临床试验都受到其潜在的全身性副作用的阻碍。靶向向纤维化肾小球输送转化生长因子受体阻滞剂为治疗提供了新的治疗方法，而没有值得注意的全身不良反应。通过将小干扰RNA(SiRNA)与纳米颗粒技术相结合，我们最近开发了一种用于基因传递的系膜细胞特异性、配体靶向、自组装的siRNA-纳米载体系统。该纳米载体在将针对肾素受体基因的siRNA靶向输送到大鼠肾炎肾小球方面表现出良好的效率和特异性，而对其他组织的影响有限。R21探索性项目提案的目的是探索将我们的新型纳米载体系统用于以系膜细胞为靶向的纳米颗粒siRNA治疗肾小球纤维化的可行性。我们建议使用转化生长因子1-Steath siRNA来检验两个相互关联的假说：(1)自组装的纳米载体可以选择性地将siRNA运送到肾小球系膜细胞，并有效地沉默靶向转化生长因子1；(2)在肾小球系膜细胞中特异性地沉默转化生长因子1将改善实验性肾小球肾炎的基质扩张进程。我们的长期目标是开发以系膜细胞为靶点的纳米siRNA，作为治疗肾小球疾病的新的分子疗法。为了验证我们的假设，我们提出了两个特定的目标：目的1：制备和优化纳米载体，通过RNA干扰在体外和体内有效地下调转化生长因子β1基因的表达；目的2：研究纳米载体介导的RNAi对肾小球局部高水平转化生长因子1的体内治疗潜力。如果成功，我们的研究将提供概念证明，即siRNA可以通过自组装的纳米载体选择性地运送到系膜细胞。这一新的策略将对肾小球纤维化的发展和进展的研究至关重要。它还将对以肾小球为目标的抗转化生长因子1 siRNA分子的开发产生重大影响，作为一种新的辅助疗法，以克服肾小球纤维化的全身副作用，并克服可能有助于抗击当前慢性肾脏疾病流行的全身副作用。公共卫生相关性：转化生长因子1作为治疗慢性肾脏疾病的有希望的靶点，具有深刻的抗炎特性，这可能会阻碍全身转化生长因子1阻断的所有临床试验。通过将小干扰RNA(SiRNA)与纳米颗粒技术相结合，我们最近开发了一种用于基因传递的系膜细胞特异性、配体靶向、自组装的siRNA-纳米载体系统。该纳米载体在将针对肾素受体基因的siRNA靶向输送到大鼠肾炎肾小球方面表现出良好的效率和特异性，而对其他组织的影响有限。R21探索性项目提案的目的是探索将我们的新型纳米载体系统用于以系膜细胞为靶向的纳米颗粒抗转化生长因子1 siRNA治疗肾小球纤维化的可行性。我们建议(1)制备和优化纳米载体，通过RNA干扰在体外和体内有效地下调转化生长因子β1基因的表达；(2)研究纳米载体介导的RNAi在实验性肾小球肾炎中的体内治疗潜力，因为肾小球局部存在高水平的转化生长因子β。成功开展后，我们的研究将对开发一种器官靶向的抗转化生长因子1 siRNA分子作为一种治疗肾小球纤维化的新辅助疗法产生重大影响，同时克服可能有助于抗击当前慢性肾脏疾病流行的全身不良反应。

DESCRIPTION (provided by applicant): The incidence and prevalence of end-stage renal disease is increasing at an alarming rate, new innovative therapies for halting the progress of chronic renal failure are urgently demanded. Overexpression of transforming growth factor-beta 1 (TGF¿1) is the most common molecular feature of progressive renal disease. It has been identified as a primary key mediator and quantifiable measure of fibrotic renal disease. TGF¿1 appears to be a promising target for treatment of chronic renal disease. However, that TGF¿ has profound anti-inflammatory properties and that animals genetically deficient in TGF¿ die of overwhelming inflammation shortly after birth, raises the possibility that systemic TGF¿ blockade may have serious inflammatory side effects. All human clinical trials of TGF¿ blockade are unfortunately hampered by its potential systemic side effects. Targeted delivery of TGF¿ blockade to fibrotic glomeruli offers novel therapeutic approaches for treatment without noteworthy systemic adverse effects. By combining small interfering RNA (siRNA) with nanoparticle technology, we have recently developed a mesangial cell-specific, ligand-targeting, self- assembled siRNA-nanovector system for gene delivery. The nanovectors show promising efficiency and specificity in targeted delivery of siRNA against the renin receptor gene to nephritic glomeruli in rat, with limited effect on other tissues. The objective of this R21 exploratory project proposal is to explore the feasibility of using our novel nanovector system for mesangial cell targeted nanoparticle-based siRNA therapeutics for glomerular fibrosis. We propose to use TGF¿1-Steath siRNA to test two inter-related hypotheses: (1) the self- assembled nanovectors can selectively deliver siRNA to glomerular mesangial cells and efficiently silence the target TGF¿1; (2) Specifically silencing of TGF¿1 in glomerular mesangial cells will ameliorate the progression of matrix expansion in experimental glomerulonephritis. Our long-term goal is to develop the mesangial cell- targeting nanoparticle-based siRNA as novel molecular therapy for glomerular disease. To test our hypothesis, we propose to carry out two Specific Aims: Aim 1: To prepare and optimize nanovectors for efficient knock- down of TGF¿1 gene expression by RNA interference both in vitro and in vivo; Aim 2: To investigate the in vivo therapeutic potential of nanovector-delivered RNAi in experimental glomerulonephritis where high levels of TGF¿1 occur locally in glomeruli. Successfully carried out, our study will provide proof-of-concept that siRNA can be selectively delivered to mesangial cells by the self-assembled nanovectors. This novel strategy will be critical for the studies in glomerular fibrosis development and progression. It also will have a significant impact on the development of a glomerulus-targeted anti-TGF¿1 siRNA molecule as a novel adjuvant therapy to overcome systemic adverse effects for glomerular fibrosis but overcomes systemic adverse effects that may help combat the current epidemic in chronic kidney disease. PUBLIC HEALTH RELEVANCE: TGF¿1, as a promising target for treatment of chronic renal disease, has profound anti-inflammatory properties, which may hamper all clinical trials of systemic TGF¿1 blockade. By combining small interfering RNA (siRNA) with nanoparticle technology, we have recently developed a mesangial cell-specific, ligand- targeting, self-assembled siRNA-nanovector system for gene delivery. The nanovectors show promising efficiency and specificity in targeted delivery of siRNA against the renin receptor gene to nephritic glomeruli in rat, with limited effect on other tissues. The objective of this R21 exploratory project proposal is to explore the feasibility of using our novel nanovector system for mesangial cell targeted nanoparticle-based anti-TGF¿1 siRNA therapeutics for glomerular fibrosis. We propose to (1) prepare and optimize nanovectors for efficient knockdown of TGF¿1 gene expression by RNA interference in vitro and in vivo; (2) investigate the in vivo therapeutic potential of nanovector-delivered RNAi in experimental glomerulonephritis where high levels of TGF¿1 occur locally in glomeruli. Successfully carried out, our studies will have a significant impact on the development of an organ-targeted anti-TGF¿1 siRNA molecule as a novel adjuvant therapy for glomerular fibrosis but overcomes systemic adverse effects that may help combat the current epidemic in chronic kidney disease.