Efficient Brain Delivery of Neuroprotective TrkB Agonist Antibodies in Alzheimer’s Disease Using CD98hc Bispecific Shuttles

阿尔茨海默病(AD)是最常见的与年龄相关的神经退行性疾病。尽管有很大的 生物制品(例如，治疗性抗体)影响AD病理生理学的可能性，这些制剂通常是 无法穿透血脑屏障(BBB)，这是他们临床疗效的最大障碍。一 有希望的策略是使用双特异性抗体bbb穿梭，这涉及到将免疫球蛋白与第二个亲和配体融合。 这与脑血管内皮细胞靶点结合，并诱导跨血脑屏障的运输。虽然之前的大多数 工作重点是针对转铁蛋白受体(TFR-1)的血脑屏障穿梭，我的实验室已经开发出一种血脑屏障穿梭 这与CD98hc结合，CD98hc是大的中性氨基酸转运体(LAT-1)的重链。值得注意的是，我们的 初步数据显示通过CD98hc AS运送的IGG具有较高的脑实质递送和滞留能力 与通过TFR-1穿梭的那些相比。重要的是，这一第一代CD98hc航天飞机使大脑能够传递 通过将单链CD98hc抗体连接到两种免疫球蛋白之一的C末端，即可获得现成的免疫球蛋白 沉重的锁链。这项建议的目标有两个：i)确定CD98hc的最佳穿梭特性 (CD98hc单链亲和力、价位和表位)，最大限度地将脑实质传递和保留到 指导第二代CD98hc航天飞机的设计；以及ii)在 使用TrkB激动剂抗体诱导神经保护信号的AD小鼠模型(tau，PS19)。对于第一次 目的：我假设血脑屏障携带单价CD98hc单链抗体(如第一次使用的那样- 和降低的CD98hc单链抗体亲和力(相对于第一代航天飞机)将 通过改善抗体在脑实质中的解离，最大限度地提高脑实质中的抗体浓度 大脑内皮细胞的一侧，如TFR-1航天飞机所观察到的。对于第二个目标，我假设 将TrkB激动剂IgGS运送到tau转基因小鼠脑实质的CD98hc航天飞机将减少 神经元和突触丢失以及认知缺陷。这一假说得到了非药物的部分疗效的支持 穿梭TrkB抗体全身性高效携带TrkB配体病毒脑内注射治疗AD 老鼠模型。因此，我在目标1中建议通过以下方式开发优化的第二代CD98hc航天飞机 评估CD98hc单链抗体亲和力、效价和表位对抗体浓度的影响 在小鼠脑实质中作为时间的函数。接下来，在目标2中，我建议评估第一个- 一代CD98hc穿梭运送TrkB激动剂Ig G到脑实质，激活野生型TrkB 和tau(PS19)转基因小鼠，并在tau转基因小鼠中介导神经保护。一个关键的预期结果 是否正在开发一种优化的CD98hc航天飞机，可以用来交付各种现有的 脑实质的治疗性抗体。第二个关键的预期结果是TrkB的临床前数据 阿尔茨海默病小鼠模型中的激动症。这些发现有望使未来对单一和组合的研究成为可能 使用拮抗剂和激动剂抗体靶向机械上独特的通路的治疗。

Alzheimer’s disease (AD) is the most common age-related neurodegenerative disease. Despite the great potential of biologics (e.g., therapeutic antibodies) to affect AD pathophysiology, these agents are generally unable to penetrate the blood-brain barrier (BBB), which represents their biggest hurdle to clinical efficacy. One promising strategy is to use bispecific antibody BBB shuttles, which involve fusing IgGs to a second affinity ligand that engages a cerebrovascular endothelial target and induces transport across the BBB. While most previous work has focused on BBB shuttles that target transferrin receptor (TfR-1), my lab has developed a BBB shuttle that engages CD98hc, the heavy chain of the large neutral amino acid transporter (LAT-1). Notably, our preliminary data demonstrate superior brain parenchymal delivery and retention of IgGs shuttled via CD98hc as compared to those shuttled via TfR-1. Importantly, this first-generation CD98hc shuttle enables brain delivery of off-the-shelf IgGs simply by attaching a single-chain CD98hc antibody to the C-terminus of one of the two IgG heavy chains. The objectives of this proposal are twofold: i) to define the optimal CD98hc shuttle characteristics (CD98hc single-chain affinity, valency, and epitope) for maximal brain parenchymal delivery and retention to guide the design of second-generation CD98hc shuttles; and ii) to generate proof-of-concept efficacy data in an AD mouse model (tau, PS19) using a TrkB agonist antibody that induces neuroprotective signaling. For the first objective, I hypothesize that BBB shuttles with monovalent CD98hc single-chain antibodies (as used in the first- generation shuttle) and reduced CD98hc single-chain antibody affinity (relative to the first-generation shuttle) will maximize antibody concentrations in the brain parenchyma by improving antibody dissociation at the abluminal side of the brain endothelium, as observed for TfR-1 shuttles. For the second objective, I hypothesize that CD98hc shuttles that deliver TrkB agonist IgGs to the brain parenchyma of tau transgenic mice will reduce neuronal and synaptic loss and cognitive deficits. This hypothesis is supported by the partial efficacy of non- shuttled TrkB antibodies delivered systemically and high efficacy of viral brain delivery of the TrkB ligand in AD mouse models. Therefore, I propose in Aim 1 to develop an optimized second-generation CD98hc shuttle by evaluating the impact of CD98hc single-chain antibody affinity, valency, and epitope on antibody concentrations in the mouse brain parenchyma as a function of time. Next, in Aim 2, I propose to evaluate the ability of a first- generation CD98hc shuttle to deliver a TrkB agonist IgG to the brain parenchyma, to activate TrkB in wild-type and tau (PS19) transgenic mice, and to mediate neuroprotection in tau transgenic mice. A key expected outcome is the development of an optimized CD98hc shuttle that can be used to deliver a wide range of existing therapeutic antibodies to the brain parenchyma. A second key expected outcome is preclinical data for TrkB agonism in AD mouse models. These findings are expected to enable future studies of single and combination therapies using antagonist and agonist antibodies that target mechanistically unique pathways.