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Pathogenesis and Treatment of AA Amyloidosis

AA型淀粉样变性的发病机制和治疗

基本信息

批准号：
10292421
负责人：
MERRILL D BENSON
金额：
--
依托单位：
RLR VA MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-07-01 至 2022-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10292421
关键词：
Acute-Phase Proteins Affect Affinity Amyloid Amyloid Fibrils Amyloid Protein AA Amyloid Proteins Amyloid beta-Protein Precursor Amyloid deposition Amyloidosis Ankylosing spondylitis Antisense Oligonucleotide Therapy Antisense Oligonucleotides Asialoglycoprotein Receptor Beer Biological Blood Bronchiectasis Cell Culture Techniques Chemical Structure Chemicals Chronic Clinical Complication Crossbreeding Deposition Development Disease Dose End stage renal failure Familial Mediterranean Fever Future Galactosamine Genomic Segment Glycosaminoglycans Hepatic Hepatocyte High Density Lipoproteins Human In Vitro Individual Infection Inflammation Inflammatory Inflammatory Bowel Diseases Inherited Interleukin-6 Intestines Kentucky Kidney Kidney Failure Ligands Liver Messenger RNA Modeling Modification Mus N-terminal Nephrotic Syndrome Organ Osteomyelitis Paraplegia Pathogenesis Patients Pharmaceutical Preparations Pharmacologic Substance Phase III Clinical Trials Post-Translational Protein Processing Prealbumin Production Proteins Proteinuria Protocols documentation Regulatory Element Rheumatoid Arthritis Risk Risk Factors Role Serum Serum amyloid A protein Site Sodium Spleen Stimulus Structure Syndrome TNF gene Testing Therapeutic Tissues Transgenic Mice Universities Urinary tract infection Veterans Work amyloid fibril formation amyloid formation amyloidogenesis autoinflammatory base beta pleated sheet decubitus ulcer design experience extracellular human model improved in vivo male mimetics mouse model novel therapeutics offspring phase III trial pre-clinical prevent promoter screening small molecule tandem mass spectrometry targeted delivery translational study

项目摘要

AA (secondary, reactive) amyloidosis is a disease caused by extracellular deposition of insoluble β-pleated sheet fibrils composed of amyloid A (AA) protein, an N-terminal fragment of the acute phase protein serum amyloid A (SAA). The deposits disrupt tissue structure and eventually compromise organ function. Common sites of deposition include kidney, spleen, liver, and intestine. In humans proteinuria due to glomerular deposits is often the first clinical manifestation, and with increasing deposition, the nephrotic syndrome usually progresses to end-stage renal disease. AA amyloidosis usually develops as a complication of chronic inflammatory conditions such as rheumatoid arthritis, ankylosing spondylitis, inflammatory bowel disease, and the hereditary auto-inflammatory syndromes (e.g., familial Mediterranean fever). It also occurs in association with long-standing infections (osteomyelitis, decubitus ulcers, bronchiectasis). Veterans with paraplegia are at risk of developing AA amyloidosis due to chronic decubitus ulcers, urinary tract infections and osteomyelitis. In addition, ankylosing spondylitis is common in our male veterans which also increases risk for AA amyloidosis. At present, there is no specific therapy for AA amyloidosis. Only control of pro-inflammatory stimuli to reduce hepatic SAA synthesis has shown an effect on AA amyloid progression. In a recent Phase III clinical trial, a potential new therapy [eprodisate (Kiacta)] failed to show significance in retarding AA progression. Prompted by the unmet needs of patients, our initial aim will be to initiate development of a therapy for AA amyloidosis using as framework translational studies we have conducted on transthyretin (TTR) amyloidosis. The therapeutic strategy will be administration of human SAA-specific antisense oligonucleotides (ASO) that facilitate degradation of SAA mRNA resulting in lower SAA levels in blood and less substrate available for amyloid formation. A transgenic mouse model carrying a human SAA1 genomic segment encompassing SAA1 promoter and regulatory elements will be generated and employed for this study. Human SAA1-specific ASOs will be provided by Ionis Pharmaceuticals and screened for ability to downregulate SAA1 expression in human cell cultures. Promising compounds will be conjugated with N-acetyl galactosamine (GalNAc); this modification confers targeted delivery to the liver and increases potency 6-10-fold, allowing for lower dosing. After further screening in normal mice to evaluate tolerability, ASOs will be given to human SAA1 transgenic mice to determine the level of suppression achieved at various doses. The human SAA1 transgenic mice we generate will be crossed with mice that do not express mouse SAA1 or SAA2; these mice will be provided by Drs. Frederick de Beer and Nancy Webb, University of Kentucky. The offspring of this cross-breeding, i.e., mice expressing human SAA1 but not mouse SAA1 or SAA2, will be induced to develop AA amyloidosis using standard protocols and also treated with selected human SAA-specific ASOs and control random ASOs to determine the extent to which ASO therapy reduces deposition of human AA amyloid. In a second aim we will explore the role of post-translational modification of SAA in the genesis of amyloid fibril formation. Our in vitro studies show that carbamylation of residues in the amino-terminal portion of mouse SAA1.1 has a potentiating effect on amyloid formation. This region of SAA is most crucial to the initiation and propagation of amyloid fibrils, suggesting that modifications in this region may have important consequences in vivo as well as in vitro. AA amyloid protein extracted from amyloid deposits of humans and mice, as well as SAA in serum, will be analyzed by tandem mass spectrometry to identify modified residues. Mouse models will be used to test whether conditions that promote in carbamylation in vivo also promote AA amyloidogenesis. Other studies will investigate the effects of carbamylation on SAA association with HDL. If modifications are found to impact amyloid formation in vivo as they do in vitro, targeting such modifications may offer new therapeutic options to treat this progressive, fatal disease.

AA（继发性，反应性）淀粉样变性是一种由不溶性β-折叠的细胞外沉积引起的疾病，由淀粉样蛋白A（AA）蛋白组成的片状原纤维，是急性期蛋白血清的N-末端片段淀粉样蛋白A（SAA）。这些沉积物破坏了组织结构，最终损害了器官功能。共同沉积部位包括肾、脾、肝和肠。在人类中，肾小球疾病引起的蛋白尿沉积物往往是第一个临床表现，随着沉积物的增加，肾病综合征通常进展为终末期肾病。AA淀粉样变性通常发展为慢性炎症的并发症。炎性病症，如类风湿性关节炎、强直性脊柱炎、炎性肠病，和遗传性自身炎症综合征（例如，地中海热（Mediterranean Fever）它也发生在联合长期感染（骨髓炎、褥疮、支气管扩张）。截瘫的退伍军人在由于慢性褥疮溃疡、尿路感染和骨髓炎，有发生AA淀粉样变性的风险。在此外，强直性脊柱炎在我们的男性退伍军人中很常见，这也增加了AA淀粉样变性的风险。目前，对AA淀粉样变性尚无特异性治疗方法。只有控制促炎刺激，肝脏SAA合成对AA淀粉样蛋白进展有影响。在最近的一项III期临床试验中，潜在的新疗法[依普罗地酯（Kiacta）]未能显示出延缓AA进展的显著性。促使由于患者的需求未得到满足，我们最初的目标将是开始开发AA淀粉样变性的治疗方法。使用我们已经进行的甲状腺素运载蛋白（TTR）淀粉样变性的框架翻译研究。的治疗策略是施用人SAA特异性反义寡核苷酸（阿索），促进SAA mRNA的降解，导致血液中SAA水平降低，淀粉样蛋白形成。携带包含SAA 1的人SAA 1基因组区段的转基因小鼠模型启动子和调节元件将被产生并用于本研究。人SAA 1特异性ASO 将由Ionis Pharmaceuticals提供，并筛选下调人SAA 1表达的能力。细胞培养有前途的化合物将与N-乙酰基半乳糖胺（GalNAc）缀合;这种修饰赋予靶向递送至肝脏并将效力增加6-10倍，从而允许更低的剂量。经过进一步在正常小鼠中筛选以评估耐受性，将AS 0给予人SAA 1转基因小鼠，确定在不同剂量下达到的抑制水平。我们产生的人SAA 1转基因小鼠将与不表达小鼠SAA 1或SAA 2的小鼠杂交;这些小鼠将由Dr. 弗雷德里克·德·比尔和南希·韦伯，肯塔基州大学。这种杂交的后代，即，小鼠表达人SAA 1但不表达小鼠SAA 1或SAA 2的小鼠，将被诱导发展AA淀粉样变性，并且还用选择的人SAA特异性ASO和对照随机ASO治疗，确定阿索治疗减少人AA淀粉样蛋白沉积的程度。在第二个目标中，探讨SAA翻译后修饰在淀粉样纤维形成中的作用。我们的体外研究表明，小鼠SAA 1.1的氨基末端部分残基的氨甲酰化具有增强作用，对淀粉样蛋白形成的影响。SAA的这一区域对淀粉样蛋白的起始和增殖至关重要原纤维，这表明在这一地区的修改可能有重要的后果，在体内以及在体外。从人类和小鼠的淀粉样蛋白沉积物中提取的AA淀粉样蛋白，以及血清中的SAA，通过串联质谱分析以鉴定修饰的残基。小鼠模型将用于测试促进体内氨甲酰化的条件是否也促进AA淀粉样蛋白生成。其他研究将研究氨甲酰化对SAA与HDL结合的影响。如果发现修改影响在体内淀粉样蛋白的形成，因为他们在体外，靶向这样的修饰可能提供新的治疗选择，治疗这种渐进的致命疾病