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-折叠细胞外沉积引起的疾病由淀粉样 A (AA) 蛋白（急性期蛋白血清的 N 末端片段）组成的片状原纤维淀粉样蛋白 A (SAA) 沉积物会破坏组织结构并最终损害器官功能。沉积部位包括肾、脾、肝和肠。在人类中，肾小球引起蛋白尿。沉积物往往是首发临床表现，随着沉积物的增加，通常会出现肾病综合征进展为终末期肾病。AA 淀粉样变性通常是慢性并发症。炎症性疾病，如类风湿性关节炎、强直性脊柱炎、炎症性肠病等遗传性自身炎症综合征（例如家族性地中海热）也与之相关。患有长期感染（骨髓炎、褥疮、支气管扩张）的退伍军人。由于慢性褥疮、尿路感染和骨髓炎而发生 AA 淀粉样变性的风险。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，AA型淀粉样变性尚无特效疗法，只能通过控制促炎刺激来减轻。在最近的一项 III 期临床试验中，肝脏 SAA 合成显示出对 AA 淀粉样蛋白进展的影响。潜在的新疗法 [eprodisate (Kiacta)] 未能显示出延缓 AA 进展的意义。由于患者的需求未得到满足，我们的初步目标是启动 AA 淀粉样变性疗法的开发我们以转甲状腺素蛋白（TTR）淀粉样变性为框架进行了转化研究。治疗策略是施用人类 SAA 特异性反义寡核苷酸 (ASO) 促进 SAA mRNA 的降解，导致血液中 SAA 水平降低，可用于的底物减少淀粉样蛋白形成，携带包含SAA1的人类SAA1基因组片段。本研究将生成并使用人类 SAA1 特异性 ASO。将由 Ionis Pharmaceuticals 提供并筛选下调人类 SAA1 表达的能力有前景的化合物将与 N-乙酰半乳糖胺 (GalNAc) 结合；赋予肝脏靶向递送能力，效力提高 6-10 倍，从而可以进一步降低剂量。在正常小鼠中进行筛选以评估耐受性，将 ASO 给予人类 SAA1 转基因小鼠以确定我们产生的人类 SAA1 转基因小鼠在不同剂量下达到的抑制水平。将与不表达小鼠 SAA1 或 SAA2 的小鼠杂交；这些小鼠将由 Drs 提供。肯塔基大学的 Frederick de Beer 和 Nancy Webb 这种杂交的后代，即小鼠。表达人类 SAA1 但不表达小鼠 SAA1 或 SAA2，将被诱导发展为 AA 淀粉样变性标准方案，并用选定的人类 SAA 特定 ASO 和对照随机 ASO 进行处理，以确定 ASO 疗法减少人类 AA 淀粉样蛋白沉积的程度。探索 SAA 翻译后修饰在淀粉样原纤维形成的体外作用。研究表明，小鼠 SAA1.1 氨基末端残基的氨甲酰化具有增强作用 SAA 的这个区域对于淀粉样蛋白的起始和增殖至关重要。原纤维，表明该区域的修饰可能在体内和体外产生重要的影响。从人类和小鼠的淀粉样沉积物中提取的 AA 淀粉样蛋白以及血清中的 SAA 将被通过串联质谱分析以确定修饰的残留物将用于测试。促进体内氨甲酰化的条件是否也会促进 AA 淀粉样蛋白生成。如果发现修饰会影响，则研究氨甲酰化对 SAA 与 HDL 关联的影响。体内淀粉样蛋白的形成与体外一样，针对此类修饰可能会提供新的治疗选择治疗这种进行性、致命的疾病。