R01 Estrogen therapy and APOE4 risk in Alzheimer's tested in female EFAD mice

控制载脂蛋白ε-4等位基因表达的基因apoe-4是高血压的最大遗传风险因素 阿尔茨海默病(AD)，与APOE3相比，风险增加高达15倍。尽管这一风险与 淀粉样蛋白(Aβ，Aβ)积聚增强是20多年前发现的，但很少成为研究的焦点 治疗方法；更不重要的是，女性性行为和载脂蛋白4诱导的风险之间的关键联系 痴呆症和阿尔茨海默病。女性(♀)载脂蛋白4携带者患AD的终生风险更大，患病率增加 与男性(β)载脂蛋白4携带者相比，认知能力下降和A-♂加速积累的比例，数据一致 在♀和♂家族性AD(FAD)转基因小鼠中进行了观察。雌二醇(E2)似乎是这一过程的关键 脆弱性：绝经前妇女经卵巢切除术去除循环中的E2会导致以后的认知障碍 可被雌激素治疗(ET)逆转。自然绝经后ET的影响仍然存在争议， 尤其是关于表明ET仅在早期有益的“临界窗口”假说 更年期窗口。另一个令人担忧的问题是，由于 妇女健康倡议研究，要求开发更安全的ET替代品(ALT-ET)。这样做的目的是 建议确定ET和ALT-ET中和性与载脂蛋白4的负相互作用的能力 新的临床前EFAD小鼠模型(表达人APOE+FAD突变)，建立了 安全的ALT-ET治疗或预防AD的首选时机和APOE异构体选择性。EFAD鼠标 通过引入h-apoE来研究人h-apoE与AD病理的相互作用。 5xFAD小鼠的基因分型：EFAD小鼠表现出雌性比雄性和E4FAD比. E3FAD；EFAD的初步数据表明，卵巢切除(OVX)后的ET对认知障碍具有保护作用。 将要研究的ALT-ET是临床选择性雌激素受体(ER)调节剂(SERM)、雷洛昔芬(RAL)和 Bazedoxifene(Baz)；临床Baz/E2组合；以及减毒的选择性雌激素类似物(SEM) 对妇科的影响。目的1：建立ET(E_2)对去卵巢♀EFAD小鼠的治疗效果。 载脂蛋白E基因分型，行为/记忆测量，载脂蛋白E和Aβ生物标记物，以及AD病理。目标2：建立 EFAD小鼠ET与基因相关的“临界窗口”。目的3：检测雌性EFAD小鼠ALT-ET水平 变革性AD疗法。剂量将通过测量与体外KD相关的脑浓度来定义 和EC50。为了探索ER异构体的作用，来自APOE-tr小鼠的混合胶质细胞培养将使用 选择性药理内质网探针。体外生物标记物(载脂蛋白E、Aβ、ABCA1、细胞因子)将反映体内情况 生物标志物，允许与ET和ALT-ET的体内效应相关，并建立了一种体外检测 AD的Alt-ET的未来发现和优化。

The gene that controls expression of the apolipoprotein ε4 allele, APOE4, is the greatest genetic risk factor for Alzheimer's disease (AD), increasing risk up to 15-fold compared to APOE3. Although this risk, associated with enhanced amyloid-β (Aβ) accumulation, was discovered over 20 years ago, it has rarely been the focus of therapeutic approaches; even less so, the critical link between female sex and the APOE4-induced risk for dementia and AD. Female (♀) APOE4 carriers have a greater lifetime risk for developing AD, an increased rate of cognitive decline and accelerated accumulation of Aβ compared to male (♂) APOE4 carriers, data consistent with observations in ♀ and ♂ familial AD (FAD) transgenic mice. Estradiol (E2) would appear to be key to this vulnerability: ablation of circulating E2 in pre-menopausal women by oophorectomy causes later cognitive deficits that are reversed by estrogen therapy (ET). The effects of ET after natural menopause remain controversial, particularly with regard to the “critical window” hypothesis stating that ET is beneficial only in the early menopause window. A further concern is the unfavorable safety profile associated with ET resulting from the Women's Health Initiative studies, requiring the development of safer ET alternatives (alt-ET). The goal of this proposal is to determine the ability of ET and alt-ET to counteract the negative interaction of sex with APOE4 in the novel preclinical EFAD mouse model (expressing human APOE +FAD mutations), establishing both the preferred timing and APOE isoform selectivity of safe alt-ET for therapy or prophylaxis of AD. The EFAD mouse was developed to study the interaction between human h-APOE and AD pathology by introducing the h-APOE genotypes into 5xFAD mice: EFAD mice display advanced pathology in females vs. males and E4FAD vs. E3FAD; and preliminary EFAD data suggests that ET after ovariectomy (OVX) protects against cognitive deficits. Alt-ET to be studied are clinical selective estrogen receptor (ER) modulators (SERMs), raloxifene (Ral) and bazedoxifene (Baz); the clinical Baz/E2 combination; and selective estrogen mimics (SEMs) with attenuated gynecological effects. Aim 1: Establish the efficacy of ET (E2) treatment in OVX ♀EFAD mice with respect to APOE genotype, measuring behavior/memory, apoE and Aβ biomarkers, and AD pathology. Aim 2: Establish the “critical window” for ET in EFAD mice with respect to genotype. Aim 3: Test alt-ET in female EFAD mice as transformational AD therapeutics. Dosing will be defined by measuring brain concentrations related to in vitro Kd and EC50. To probe the roles of ER isoforms, mixed glial cultures from APOE-TR mice, will be studied using selective pharmacological ER-probes. In vitro biomarkers (apoE, Aβ, ABCA1, cytokines) will mirror in vivo biomarkers, allowing correlation with in vivo effects of ET and alt-ET, and development of an in vitro assay for future discovery and optimization of Alt-ET for AD.