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PHARMACOLOGY OF HUMAN INTERFERON GAMMA INHIBITORS

人干扰素γ抑制剂的药理学

基本信息

批准号：
6181226
负责人：
MURALI RAMANATHAN
金额：
$ 9.7万
依托单位：
STATE UNIVERSITY OF NEW YORK AT BUFFALO
依托单位国家：
美国
项目类别：
财政年份：
1996
资助国家：
美国
起止时间：
1996-04-01 至 2002-03-31
项目状态：
已结题

项目摘要

The long term objective of this project is to develop potent, safe an specific pharmacological inhibitors for interferon-gamma that are useful in the treatment of septic hock and autoimmune diseases such as multiple sclerois and Type 1 diabetes. The focus of the research is an oligonucleotide based inhibitor that specifically blocks a multitude of interferon-gamma effects, including the induction of the major histocompatibility complex Class II DR and Class I and ICAM-1 proteins in several cell types. This lead oligonucleotide also blocks the significant synergy between interferon-gamma and tumor necrosis factor-aalpha in mixture. It inhibits the binding of interferon-gamma to its cell surface receptor complex and thereby blocks downstream signaling by receptor associated kinases. Because our preliminary results support the feasibility of constructing oligonucleotide-based inhibitors for interferon-gamma, we propose to identify the molecular mechanisms of this inhibitory activity and use the resulting information to engineer an even more potent inhibitor for interferon-gamma. Accordingly, the specific aims of this project are to: i) identify the amino acid residues and nucleic acid bases that interact at the site of action, ii) use the mechanistic information to synthesize more potent inhibitors for interferon-gamma, iii) characterize the potency, activity, specificity and selectivity more potent inhibitors for interferon-gamma, iii) characterize the potency, activity, specificity and selectivity profiles of the newly synthesized inhibitors, iv) determine the effects of these inhibitors on human peripheral blood derived immune cells and to test the hypothesis that these inhibitors will drive the immune system to favor a humoral or TH2-like response and, v) test the inhibitors in a generalized Shwartzman model for septic shock. We will use epitope disruption and lysine protection assays to identify the amino acids of interferon-gamma that constitute the binding site. Hydroxyl radical footprinting and dimethylsulfate protection assays will be used to identify the nucleic acid bases involved in binding. The mechanistic information will then be used to optimize the length, sequence and backbone composition of these interferon-gamma inhibitory oligonucleotides. These studies will provide critical mechanistic information on the pharmacological and molecular basis for interferon-gamma inhibitory oligonucleotide activity. More importantly, the results from this model system should be generalizable to the design of oligonucleotide inhibitors for other protein, particularly cytokines.

该项目的长期目标是开发有效，安全，干扰素-γ的特异性药理学抑制剂，治疗脓毒性跗关节和自身免疫性疾病，如多发性硬化症和1型糖尿病。研究的重点是基于寡核苷酸的抑制剂，其特异性阻断多种干扰素-γ效应，包括诱导主要的组织相容性复合物II类DR和I类和ICAM-1蛋白在几种细胞类型。这种前导寡核苷酸还阻断了显著的干扰素-γ和肿瘤坏死因子-α在混合物. 它抑制干扰素-γ与其细胞表面的结合受体复合物，从而阻断受体的下游信号传导相关激酶因为我们的初步结果支持构建基于阿替西肽的抑制剂的可行性，干扰素-γ，我们建议确定这一分子机制抑制活性，并使用由此产生的信息来设计一个甚至更有效的干扰素-γ抑制剂因此，具体目标该项目的主要目的是：i）鉴定氨基酸残基和核酸在作用位点相互作用的酸碱，ii）使用合成更有效的干扰素-γ抑制剂的信息，iii）表征效力、活性、特异性和选择性更有效干扰素-γ的抑制剂，iii）表征干扰素-γ的效力，活性，新合成的抑制剂的特异性和选择性概况， iv）确定这些抑制剂对人外周血的影响，衍生的免疫细胞，并测试假设，这些抑制剂将驱动免疫系统以有利于体液或TH 2样应答，以及，v）在感染性休克的广义Shwartzman模型中测试抑制剂。我们将使用表位破坏和赖氨酸保护测定来鉴定构成结合位点的干扰素-γ的氨基酸。羟基自由基足迹法和硫酸二甲酯保护试验将用于鉴定参与结合的核酸碱基。机械论然后，信息将用于优化长度、序列和骨架这些干扰素-γ抑制性寡核苷酸的组成。这些研究将提供关键的机械信息，干扰素-γ抑制剂的药理学和分子基础寡核苷酸活性。更重要的是，这个模型的结果该系统应可推广到寡核苷酸抑制剂的设计其他蛋白质，特别是细胞因子。