Faerman C H, Savvides S N, Strickland C, Breidenbach M A, Ponasik J A, Ganem B, Ripoll D, Krauth-Siegel R L, Karplus P A
Department of Biochemistry, Molecular and Cell Biology, Cornell University, Ithaca, NY 14853, USA.
Bioorg Med Chem. 1996 Aug;4(8):1247-53. doi: 10.1016/0968-0896(96)00120-4.
Benson et al. (Biochem. J. 1992, 286, 9) reported three novel competitive inhibitors of trypanothione reductase (TR), which were selected to complement a hydrophobic region identified on the TR structure which was not present on human glutathione reductase (hGR). Benson et al. also noted that chlorpromazine, a tricyclic antidepressant known to have trypanocidal activity, was an inhibitor of TR. Here we show that chlorpromazine is a competitive inhibitor of TRs from Crithidia fasciculata (Ki = 14 microM) and Trypanosoma cruzi (Ki = 10 microM), but the drug binds > 50-fold more weakly (Ki = 762 microM) to hGR. Analogues of chlorpromazine differing in the length of the side chain carrying the positively charged R-group are also selective TR inhibitors whereas, a tricyclic structure carrying a negatively charged side chain is a competitive inhibitor with selectivity for hGR (K(hGR)i = 165 microM vs. K(TR)i = 1400 microM). This finding suggests that simple charge characteristics, rather than differences in hydrophobicity, may account for a significant portion of the selectivity of this series of inhibitors for these two enzymes. Electrostatic analysis of the structures of TR and hGR thus provides a rationale for these results, and offers a new principle for inhibitor design. The principle gains further support from the observation that all known tricyclic competitive inhibitors of TR are positively charged. In order to investigate the in vivo relevance of our findings we have examined the effect of chlorpromazine and its negatively charged analogue on the growth of C. fasciculata parasites. Consistent with our kinetics, chlorpromazine (50 microM) inhibited the growth of parasites by 50%, while no measurable decrease in parasite growth rate was noted in the presence of the negatively charged inhibitor (400 microM). Furthermore, the highly similar inhibitory profiles of C. fasciculata TR and T. cruzi TR suggest that drug-design studies using the structurally better-studied C. fasciculata TR are also relevant to the human pathogen T. cruzi.
本森等人（《生物化学杂志》，1992年，第286卷，第9期）报道了三种新型的锥虫硫醇还原酶（TR）竞争性抑制剂，这些抑制剂是为了补充在TR结构上鉴定出的一个疏水区域而筛选出来的，该区域在人类谷胱甘肽还原酶（hGR）上不存在。本森等人还指出，氯丙嗪，一种已知具有杀锥虫活性的三环抗抑郁药，是TR的抑制剂。在此我们表明，氯丙嗪是来自克氏锥虫（Ki = 14 microM）和克鲁斯锥虫（Ki = 10 microM）的TR的竞争性抑制剂，但该药物与hGR的结合能力弱> 50倍（Ki = 762 microM）。侧链长度不同且带有带正电荷R基团的氯丙嗪类似物也是选择性TR抑制剂，而带有带负电荷侧链的三环结构是对hGR具有选择性的竞争性抑制剂（K(hGR)i = 165 microM对K(TR)i = 1400 microM）。这一发现表明，简单的电荷特征而非疏水性差异可能是这一系列抑制剂对这两种酶选择性的很大一部分原因。因此，对TR和hGR结构的静电分析为这些结果提供了理论依据，并为抑制剂设计提供了新的原则。这一原则从所有已知的TR三环竞争性抑制剂都带正电荷这一观察结果中得到了进一步支持。为了研究我们发现的体内相关性，我们研究了氯丙嗪及其带负电荷的类似物对克氏锥虫寄生虫生长的影响。与我们的动力学结果一致，氯丙嗪（50 microM）使寄生虫的生长受到50%的抑制，而在存在带负电荷抑制剂（400 microM）的情况下，未观察到寄生虫生长速率有可测量的下降。此外，克氏锥虫TR和克鲁斯锥虫TR高度相似的抑制谱表明，使用结构研究更深入的克氏锥虫TR进行的药物设计研究也与人类病原体克鲁斯锥虫相关。
