Non-Human Primate Model of Gluten-Sensitive Enteropathy

描述（由申请人提供）：在人类中，谷蛋白敏感性肠病（GSE）通常在遗传上易患乳糜泻（CD）的个体中发现，在恒河猴和人类中，它可以由含谷蛋白的饮食诱导。我们最近进行了一项实验，对麸质敏感的猕猴和对照猕猴分别喂食含麸质食物和无麸质食物。在GSE猕猴中实现了完全恢复-基于从他们的饮食中撤出麸质。此外，我们鉴定了2个DRB单倍型和/或4个DQ等位基因对作为恒河猴面筋敏感性免疫遗传相关的候选MHC II基因。虽然已经建立了几种有用的模型来研究CD，包括表达HLA-DQ2等位基因的转基因小鼠，但目前还没有令人满意的动物模型来满足这种自身免疫性疾病的遗传和病理标准。我们已经部分表征了恒河猴GSE模型。我们相信这样的模型将对研究CD的免疫发病机制和治疗非常有用。为了进一步开发该模型，我们计划：目的1：评估恒河猴GSE的临床、组织病理学和免疫学替代物之间的关系。将评估临床或亚临床GSE猕猴与对照组的临床症状（腹泻、体重减轻、皮肤病变等）、AGA、抗转谷氨酰胺酶2 （TG2）抗体、绒毛萎缩、IELs和产生炎症细胞因子的肠道T淋巴细胞的增加之间的关联。目的2：确认在恒河猴中鉴定的MHC II等位基因与麸质敏感性的免疫遗传学关联。从至少100只麸质敏感猕猴和100只来自印度的对照猕猴中提取的DNA将被检测与我们最近确定为MHC II候选等位基因的2个DRB单倍型和/或4个DQ等位基因对的关联。我们预测，与乳糜泻患者类似的DQ2/8关联也将在患有GSE的恒河猴中得到证实。目的3：评估谷蛋白敏感猕猴和对照猕猴之间a2-麦胶蛋白消化的差异。在对谷蛋白敏感的猕猴和对照猕猴进行的初步研究中，发现GSE动物而不是对照动物，也不是缓解动物，通过肠上皮吸收未消化的33-mer。因此，我们提出，在具有适当MHC II型的GSE猕猴中，对膳食面筋的全身体液免疫反应是由未消化的a2-麦胶蛋白通过“渗漏”上皮吸收引起的。目的4：评价口服酶治疗GSE恒河猴的效果。MHC ii -预选猕猴麸质敏感性将首先放置在一个无麸质饮食，以实现缓解。在后续的谷蛋白挑战中，对谷蛋白敏感的猕猴将增加谷蛋白水平，并每日口服固定剂量的荚膜鞘单胞菌脯氨酸内肽酶、大麦半胱氨酸内蛋白酶EP-B2，以及这两种酶一起评估它们的治疗潜力。

DESCRIPTION (provided by applicant): In humans, gluten-sensitive enteropathy (GSE) is typically found in individuals genetically predisposed to celiac disease (CD), and in rhesus monkeys as well as in humans it can be induced by a gluten-containing diet. We recently performed experiments where gluten-sensitive and control macaques were fed gluten- containing diets followed by gluten-free diets. Complete recovery was achieved in GSE macaques - based on withdrawal of gluten from their diet. Furthermore, we identified 2 DRB haplotypes and/or 4 DQ allelic pairs as candidate MHC II genes for immunogenetic association with gluten sensitivity in rhesus macaques. Although several useful models have been established to study CD, including transgenic mice expressing HLA-DQ2 allele, there is no satisfactory animal model that would fulfill both genetic and pathologic criteria of this autoimmune disease. We have partially characterized a rhesus GSE model. We believe that such a model will be extremely useful for studies of the immunopathogenesis and treatment of CD. To develop this model further, we plan to: Aim 1: Evaluate an association between clinical, histopathological and immunological surrogates of GSE in rhesus macaques. An association between clinical symptoms (diarrhea, weight loss, skin lesions, etc.), presence of AGA, anti-transglutaminase 2 (TG2) antibodies, villous atrophy, increased presence of IELs and inflammatory-cytokine producing intestinal T lymphocytes in macaques with clinical or subclinical GSE vs. controls will be evaluated. Aim 2: Confirm MHC II alleles that were identified in rhesus macaques as candidates for immunogenetic association with gluten sensitivity. DNA extracted from at least 100 gluten-sensitive and 100 control macaques of Indian origin will be examined for the association with 2 DRB haplotypes and/or 4 DQ allelic pairs that we recently identified as MHC II candidate alleles. We predict that analogous to celiac patients DQ2/8 association will also be confirmed in rhesus macaques with GSE. Aim 3: Evaluate the differences in a2-gliadin digestion between gluten sensitive and control macaques. In pilot study with gluten sensitive and control macaques, it was found that GSE animals but not controls, nor remitted animals, absorb undigested 33-mer across intestinal epithelium. Thus, it was proposed that systemic humoral immune response to dietary gluten is caused by absorption of undigested a2-gliadin across "leaky" epithelium in GSE macaques with proper MHC II type. Aim 4: Evaluate the oral enzyme treatments in rhesus macaques with GSE. MHC II-pre- selected macaques with gluten sensitivity will be first placed on a gluten-free diet to accomplish remission. In a follow-up gluten challenge, gluten sensitive macaques will be dosed with increasing levels of gluten and a fixed daily oral dose of prolyl endopeptidase from Sphingomonas capsulata, cysteine endoprotease EP-B2 from barley, and the two-enzymes together to evaluate their therapeutic potential.