Oral tolerance in enzyme replacement therapy of Morquio A disease

描述（由申请方提供）：Morquio A病（粘多糖沉积症IVA; MPS IVA）是一种常染色体隐性溶酶体贮积症（LSD），由N-乙酰半乳糖胺-6-硫酸酯硫酸酯酶（GALNS）缺乏引起。未降解的硫酸角蛋白（KS）会积聚，尤其是在骨中，导致全身性骨骼发育不良。自从PI和合作研究者在20多年前开始Morquio A研究以来，GALNS的纯化、人GALNS cDNA的克隆、人基因组基因的克隆、常见突变的鉴定、小鼠基因的克隆、GALNS的三级结构、Morquio A小鼠模型的建立、ELISA的KS测定、Morquio的教育CD、CHO中产生的重组人GALNS、串联质谱的KS测定、建立了国际Morquio登记系统，首次在Morquio A基因敲除小鼠上进行了ERT试验，并获得了Morquio A患者的生长曲线。我们已经发表了50多篇关于Morquio A的同行评审文章。申请和/或发布了7项与治疗和诊断相关的专利。我们正在开发新的创新酶替代疗法（ERT），以针对Morquio A的骨。同时，重复给予酶可能会诱导不良的免疫反应。50 - 90%的LSD患者静脉注射后产生了抗输注酶的抗体。已广泛报道LSD患者对ERT的免疫应答，并将其作为治疗的主要并发症之一。目标：直接的载体目标是开发"口服耐受性"，用于对LSD的ERT的免疫应答，特别是Morquio A病。长期运营商的目标是优化目前的ERT方案，并以更大的可行性改变ERT对LSD的疗效。我们的目标是：1）在Morquio A小鼠中建立对人重组GALNS的口服耐受方案; 2）在耐受化和非耐受化的Morquio A小鼠之间比较ERT的功效。研究计划：为了抑制对ERT的免疫应答，将通过口服施用合成的GALNS肽来设计新的口服耐受方案。将在Morquio A基因敲除小鼠中通过多次低剂量喂养方案研究用合成GALNS肽诱导口服耐受性。在诱导对GALNS的口服耐受后，我们将对Morquio A小鼠进行ERT以评估病理学病变的改善。实验计划是1）制备合成GALNS肽，2）在Morquio A小鼠中通过合成GALNS肽的多次低剂量喂养方案诱导口服耐受性，3）通过流式细胞术通过调节性T细胞群的增加来测量炎症反应的抑制，4）测量针对GALNS的抗体的存在或不存在，5）Morquio A小鼠将通过ERT治疗，以及6）我们通过测量抗体和评估病理学改善来评估免疫耐受性。为了实现所提出的目标，将解决当前ERT中未满足的挑战之一，并可能显著改变治疗的疗效。 公共卫生相关性：Morquio A病是一种罕见的疾病，由一种溶酶体酶缺乏引起，导致主要在骨骼中的糖链积聚，并表现出全身性骨骼疾病，包括身材矮小，胸部突出，脊髓压迫，knock-knee和关节松动。最近的先进技术导致了所谓的“酶替代疗法”的发展，以弥补缺乏的酶。为了避免对输注酶的免疫反应和由此产生的不希望的副作用，迫切需要开发新的免疫耐受方法，通过口服给药酶的合成肽。

DESCRIPTION (provided by applicant): Morquio A disease (mucopolysaccharidosis IVA; MPS IVA) is an autosomal recessive lysosomal storage disorder (LSD) caused by deficiency of N-acetyl-galactosamine-6-sulfate sulfatase (GALNS). Undegraded keratin sulfate (KS) will be accumulated especially in bone, leading to systemic skeletal dysplasia. Since the PI and co-investigators started Morquio A research over 20 years ago, purification of GALNS, cloning of human GALNS cDNA, cloning of human genomic gene, identification of common mutations, cloning of mouse gene, tertiary structure of GALNS, establishment of Morquio A murine models, KS assay by ELISA, Educational CD for Morquio, recombinant human GALNS produced in CHO, KS assay by tandem mass spectrometry, development of International Morquio Registry, the first attempt of ERT on Morquio A knock-out mouse, and growth chart for Morquio A patients were achieved. We have published over 50 peer-review articles on Morquio A. Seven patents related to therapy and diagnosis were applied and/or issued. We are developing new innovative enzyme replacement therapy (ERT) to target bone for Morquio A. Meanwhile, repeated administration of the enzyme could induce an undesirable immune response. 50-90% patients with LSDs administered intravenously raised antibody against infused enzyme. Immune response to ERT in LSDs has been widely reported and presented as one of the major complications of treatment. Goals: The immediate carrier goal is to develop "oral tolerance" for immune response to ERT of LSDs, in particular, Morquio A disease. The long-term carrier goal is to optimize current ERT regimen and to change efficacy of ERT for LSDs with more feasibility. We will aim: 1) to establish an oral tolerance protocol to the human recombinant GALNS in Morquio A mice; 2) to compare the efficacy of ERT between the tolerized and non-tolerized Morquio A mice. Research Plan: For suppressing the immune response to ERT, a novel oral tolerance protocol will be designed by administering synthetic GALNS peptides orally. Induction of oral tolerance with synthetic GALNS peptides will be investigated by a multiple low-dose feeding regimen in a Morquio A knock-out mouse. After induction of oral tolerance against GALNS, we will perform ERT on Morquio A mouse to assess improvement of pathological lesions. The experimental plan is 1) synthetic GALNS peptides will be made, 2) oral tolerance is induced by a multiple low-dose feeding regimen of synthetic GALNS peptides in Morquio A mouse, 3) the inhibition of an inflammatory response will be measured by the increment of regulatory T cell populations by flow cytometry, 4) the presence or absence of antibodies against GALNS will be measured, 5) Morquio A mice will be treated by ERT, and 6) we assess the immunological tolerance by measuring antibodies and assessing pathological improvement..To accomplish the proposed aims will resolve one of unmet challenges in current ERT and could change efficacy of the therapy dramatically. PUBLIC HEALTH RELEVANCE: Morquio A disease is a rare disorder caused by a deficiency of one of lysosomal enzymes, leading to accumulation of a sugar of chain mainly in bone, and showing systemic skeletal disease with short stature, prominent chest, spinal cord compression, knock-knee, and loose joints. Recent advanced technology has led to development of so-called "enzyme replacement therapy" to make up for the deficient enzyme. To avoid immunological reaction against infused enzyme and resultant unwanted side effect is urgently demanded, therefore, we will develop novel immunological tolerance method to the infused enzyme by oral administration of synthetic peptide for the enzyme.