The Role of Quinols in Estrogen Neuroprotection

描述(申请人提供)：越来越多的证据表明，雌激素通过抑制神经毒性刺激来表达神经保护作用，主要是通过其直接清除自由基的活性。这项赠款申请的目的是通过关注基本化学来了解这一活动，其中该过程的分子机制以及从自由基清除反应中衍生的产物的化学性质和命运被认为是关键要素。我们的假设集中在雌激素衍生的喹酚作为特定反应产物的作用，其参与已被初步数据所牵连。通过系统地改变酚类A环化合物2-和/或4-位上的取代基，我们应该深入了解苯氧基(Aro*)稳定性、C-1I0位Aro*的电子密度以及它对羟基自由基(-0H)的反应活性的影响。Fenton反应将被用作化学模型，该反应导致0H的形成。我们假设0H暴露时会涉及羟化作用，这是从雌激素中产生非自由基产物的重要过程。在将用于研究通过系统修饰内源化合物获得的合成雌激素的命运的模型系统中，只有一种类型的(单一)羟基物种，即喹酚结构，通过两步羟基自由基清除机制被预期。预计对苯二酚的形成速度不仅与类固醇化合物的立体和电子元素有关，还与它们的神经保护作用有关。苯酚到苯二酚的途径可能增强雌激素清除自由基的能力，并在神经保护中发挥关键作用。还将研究一种将喹酚还原为苯酚的转化(因此，雌激素“循环”)，以防止体内可用神经保护雌激素的耗尽。这种再激活(就动力学和A环取代基的影响而言)将在体外无细胞和细胞系统中进行研究，并在实验动物中通过体内微透析进行研究。将寻求支持这样的假设，即对苯二酚的减少并不伴随着活性氧物种的增加形成。以谷氨酸诱导的HT22细胞氧化应激为实验范式，我们将研究雌激素酚类A环衍生物的神经保护作用。除了结构-活性关系研究外，还将寻求化合物在-OH暴露时形成其喹酚产物的倾向与其在该模型系统中用于神经保护的有效剂量之间的相关性。预计基于机制研究和体外筛选而选择的新雌激素类似物将在短暂性大脑中动脉闭塞这一脑缺血模型中显示出体内神经保护作用。因此，它们将成为新的先导化合物，用于开发效率更高的神经保护剂。

DESCRIPTION (provided by applicant): There has been mounting evidence that estrogens express neuroprotective effects by the suppression of neurotoxic stimuli largely via their direct radical-scavenging activity. The objective of this grant application is to understand this activity by focusing on the underlying chemistry in which the molecular mechanism of the process and the chemical nature and fate of the products derived from the radical-scavenging reaction are considered the key elements. Our hypotheses center on the role of estrogen-derived quinols as specific reaction products whose involvement has been implicated by preliminary data. By systematically varying substituents in the 2- and/or 4-positions of the phenolic A-ring compounds, we should gain insight into the influence of phenoxy-radical (ArO*) stability, electron density of ArO* at the C-1i0 position and, thus, its reactivity towards hydroxyl radical (-0H). The Fenton-reaction, which leads to 0H formation, will be employed as a chemical model. We hypothesize that hydroxylation will be involved upon *0H exposure as an important process that produces non-radical products from estrogens. In the model system that will be used to study the fate of the synthetic estrogens obtained by the systematic modification of the endogenous compounds, only one type of (mono)hydroxylated species, of a quinol structure, is anticipated via a two-step hydroxyl-radical scavenging mechanism. The rate of quinol formation is expected to correlate not only with the steric and electronic elements of the steroidal compounds, but also with their neuroprotective effect. The phenol to quinol pathway may augment the estrogens' free-radical scavenging efficacy, and make a pivotal contribution to neuroprotection. A reductive quinol to phenol transformation (hence, estrogen "recycling") that prevents the depletion of the available neuroprotective estrogens in vivo will also be investigated. This reactivation (in terms of kinetics and influence by the A-ring substituents) will be studied in vitro in cell-free and cellular systems, and by in vivo microdialysis in experimental animals. Support will be sought for the hypothesis that the reduction of quinols is not accompanied by an increased formation of reactive oxygen species. By using glutamate-induced oxidative stress in HT22 cells as an experimental paradigm, we will study the neuroprotective effects of the phenolic A-ring derivatives of estrogens. In addition to structure - activity relationship studies, a correlation between the propensity of the compound to form its quinol product upon -OH exposure and its effective dose in this model system for neuroprotection will be sought. It is anticipated that the new estrogen analogs selected based on mechanistic studies and in vitro screening will show in vivo neuroprotective effects in a transient middle cerebral artery occlusion, a model for cerebral ischemia. Hence, they will serve as new lead compounds for the development of neuroprotective agents with improved efficacy.