Immunomodulatory ligand B7-1 targets p75 neurotrophin receptor in neurodegeneration

大脑老化的特点是促炎和抗炎之间的动态平衡缓慢恶化。 兴奋性细胞因子，导致促炎状态。常驻的中枢神经系统免疫细胞通常存在于 静息状态，但随着年龄的增长，对二次侮辱表现出更高的易感性，导致 细胞表面标记物的表达和细胞因子的释放有助于促炎状态。功能界别 神经元细胞表面蛋白与免疫细胞之间相互作用的作用尚未完全发挥作用 地址。使用无偏见的蛋白质-蛋白质相互作用筛选，我们确定免疫调节 配体B7-1(CD80)与p75神经营养素受体(P75)相互作用。重要的是，B7-1：P75的相互作用是 起源于最近的进化，只存在于灵长类动物中，尽管外源应用的人类B7-1(HB7-1)结合 小鼠p75由于p75基因的广泛保守。我们绘制了负责p75的B7-1表面 参与，并证明它包括已知与CTLA-4/CD28相互作用的区域，并扩展到 其他曲面区域。鉴于这种重叠，CLTA-4/CD28直接与p75竞争结合HB7-1。 体外培养的小鼠海马神经元暴露于HB7-1可显著改变树突状细胞和棘状细胞的形态。 突触后蛋白PSD95的丢失和微管的中断以p75依赖的方式。Abatacept，一种 FDA批准的治疗蛋白(CTLA-4-FC)可抑制这些作用。小鼠体内注射HB7-1的实验研究 阿尔茨海默病中受影响的海马区下丘脑导致p75依赖的急性修剪 树枝状刺。为了研究HB7-1在完整动物和神经退行性变模型中的作用，我们 开发了一种嵌合的人源化B7-1敲入小鼠。我们的长期目标是研制出单抗 专门阻断HB7-1：p75参与预防神经退行性变，我们已经确定了两个 在10 nm浓度下阻断HB7-1：p75突触消除的单抗。目标1将进一步 评估和优先考虑这些和其他单抗，以开发更多具有广泛表位的试剂 覆盖范围。在目标2中，我们的两个铅抗体和其他优先抗体将通过海马区进行评估 神经元培养对阻断突触消除和负树突状重塑的作用。AIM 3将评估 选择性抗体预防HB7-1脊椎清除和行为障碍的体内效果 阻断WT、嵌合B7-1和AD小鼠免疫细胞表达的B7-1与神经元p75的相互作用 模特。这些研究代表了概念、机制和治疗方面的进展，通过(1)扩大我们的 了解免疫：促进神经退行性变的神经元相互作用；(2)确定 在人类中存在，但在小鼠中不存在，以克服小鼠模型的现有局限性；(3)识别临床前 老龄化人口的治疗学。

The aging brain is characterized by a slow deterioration of homeostatic balance between pro- and anti-inflam- matory cytokines, resulting in a proinflammatory state. Resident CNS immune cells are normally present in a resting state, but exhibit heightened vulnerability to secondary insults with aging, leading to a phenotypic shift in cell surface marker expression and cytokine release that contribute to the proinflammatory state. The functional contributions of interactions between cell surface proteins of neurons and immune cells have not been fully addressed. Using an unbiased protein-protein interaction screen, we determined that the immunomodulatory ligand B7-1 (CD80) interacts with the p75 neurotrophin receptor (p75). Importantly, the B7-1:p75 interaction is of recent evolutionary origin, present only in primates, although exogenously applied human B7-1 (hB7-1) binds murine p75 due to the extensive conservation of p75. We mapped the B7-1 surface responsible for p75 engagement and demonstrated that it includes regions known to interact with CTLA-4/CD28 and extends to additional surface regions. Given this overlap, CLTA-4/CD28 directly compete with p75 for binding to hB7-1. Exposure of murine hippocampal neurons in vitro to hB7-1 acutely alters dendritic and spine morphology, with loss of postsynaptic protein PSD95 and microtubule discontinuity in a p75-dependent manner. Abatacept, an FDA-approved therapeutic protein (CTLA-4-Fc), inhibits these effects. In vivo injection of hB7-1 into the murine subiculum, a hippocampal region affected in Alzheimer’s Disease, results in acute p75-dependent pruning of dendritic spines. To study the effects of hB7-1 in the intact animal, and in models of neurodegeneration, we developed a chimeric humanized B7-1 knock-in mouse. Our long-term goal is to develop monoclonal antibodies that specifically block hB7-1:p75 engagement for the prevention of neurodegeneration, and we have identified two such monoclonal antibodies that block hB7-1:p75 synapse elimination at 10nM concentration. Aim 1 will further evaluate and prioritize these and other monoclonal antibodies to develop additional reagents with broad epitope coverage. In Aim 2, our two lead antibodies, and other prioritized antibodies, will be evaluated using hippocampal neuron cultures for effects on blocking synaptic elimination and negative dendritic remodeling. Aim 3 will evaluate the in vivo effectiveness of select antibodies on preventing hB7-1 spine elimination and behavioral impairment by blocking the interaction of immune cell-expressed B7-1 with neuronal p75 in WT, chimeric B7-1 and an AD mouse model. These studies represent conceptual, mechanistic and therapeutic advances by (1) extending our understanding of immune:neuronal interactions that promote neurodegeneration; (2) identifying mechanisms that exist in humans, but not mice, to overcome existing limitations of murine models; (3) identifying preclinical therapeutics for aging populations.