OLEFINIC ANALOGS OF CSA--NEW AGENTS

环孢素A是一种由11个氨基酸组成的环肽， 已证实的器官领域的选择性免疫调节活性 移植（心脏、肝脏、肺、肾脏和骨髓）和自体 免疫疾病（葡萄膜炎，1型糖尿病，类风湿性关节炎， 等等）。 尽管这种有效的和临床上令人印象深刻的免疫抑制剂 活性，其使用受到其副作用的限制：肾毒性 （主要副作用，不可逆纤维化）、肝毒性和 神经毒性 这些毒性的表达机制不是 已知的和毒性仅部分控制监测药物 血液水平。 因此，开发更安全的类似物将大大扩展 这类强力免疫抑制剂的临床效用 该提案的长期目标是开发一种类似的 具有更高效力、更高选择性和/或不同免疫抑制剂的CsA 活性和毒性降低。 具体目标是 实现这一长期目标的途径是：半合成一系列烯烃 氨基酸1（关键氨基酸）的类似物，体外评价 这些类似物的免疫抑制活性的体内评价 类似物的肾毒性和后两者的相关性 活性与类似物的三维溶液结构， 通过NMR光谱测定。 半合成的方法 CsA类似物已经开发出来， 残基1中的双带裂解，然后烯化。 的 体外免疫抑制剂活性的评价将 通过包括抑制IL-2产生的已知方法完成， 抑制胸腺细胞增殖，竞争性结合（CsA vs类似物） 亲环素（推定的CsA受体）和CsA MAB的类似物识别。 将在先前使用的大鼠中进行肾毒性评价 通过评价肾功能（GFR、BUN和肌酐水平）建立模型， 肾脏形态学（光学显微镜：细胞增殖，小管细胞 空泡化、小管萎缩和纤维化）。 这后两 特征将与所述组合物的三维结构相关联。 将通过使用1维和2维NMR测定的类似物 光谱技术和三维结构分析程序 DSPACE。 这些研究的结果将导致 结构-活性关系，这将导致发展 更安全、更强效的CsA免疫抑制剂类似物。

Cyclosporin A is an eleven amino acid, cyclic peptide with clinically proven selective immunoregulatory activity of the fields of organ transplantations (hear, liver, lung, kidney and bone marrow) and auto- immune disorders (uveitus, type 1 diabetes mellitus, rheumatoid arthritis, etc.) Despite this potent and clinically impressive immunosuppressant activity, its use has been restricts by its side effects: nephrotoxicity (major side effect, irreversible fibrosis), hepatotoxicity and neurotoxicity. The mechanisms of expression of these toxicities are not known and the toxicities are only partially controlled by monitoring drug blood levels. Thus the development of a safer analog would greatly expand the clinical utility of this powerful class of immunosuppressants. The long term objective of this proposal is the development of an analog of CsA with higher potency, more selective and/or different immunosuppressant activity and decreased toxicity. The specific aims that are directed towards this long term goal are: the semi-synthesis of a series of olefin analogs of amino acid 1 (a critical amino acid), the in vitro evaluation of the immunosuppressant activity of these analogs, the in vivo evaluation of the nephrotoxicity of the analogs and the correlation of the latter two activities with the 3 dimensional solution structure of the analogs as determined by NMR spectroscopy. The procedure for the semi-synthesis of the CsA analogs has already been developed and involves an oxidative cleavage of the double band in residue 1 followed by olefination. The evaluation of the in vitro immunosuppressant activities will be accomplished by known procedures involving inhibition of IL-2 production, inhibition of thymocyte proliferation, competitive binding (CsA vs analog) to cyclophilin (putative CsA receptor) and analog recognition by CsA MAB. The evaluation of nephrotoxicity will be done in the previously used rat model by evaluation of kidney function (GFR,BUN and creatinine levels) and kidney morphology (light microscopy: cell proliferation, tubule cell vacuolization, tubule atrophification and fibrosis). These latter two characteristics will be correlated to the 3 dimensional structure of the analogs which will be determined by the use of 1 and 2 dimensional NMR spectroscopy techniques and the 3 dimensional structure analysis program DSPACE. The results of these studies will lead to the development of structure-activity relationships which will lead to the development of safer, more powerful CsA immunosuppressant analogs.