New Human Antibodies for CNS Drug Delivery

摘要 全世界数以百万计的人患有阿尔茨海默氏症、中风和 脑癌。蛋白质/基因图谱技术和高通量药物筛选技术的研究进展 催生了许多新的候选药物。然而，血脑屏障(BBB)阻碍了 限制脑摄取这一新一代神经疗法的发展和临床应用 大多数小分子疗法，并禁止大脑摄取基于蛋白质和基因的药物。一个 有希望的无创脑递送策略利用内源性血脑屏障转运机制作为一种 意味着将毒品货物从血液输送到大脑。这种受体介导的运输系统可以是 利用抗体的精致特异性进行靶向，这些抗体又与药物有效载荷相联系，包括 小分子、蛋白质或DNA疗法。在与血侧的受体结合后，抗体- 药物结合物充当转运体的人工底物，从血液中转运，穿过 血脑屏障，进入大脑。然而，目前的方法已经产生了有限的大脑吸收，因为靶向 转运蛋白广泛表达，抗体靶向试剂具有较低的血脑屏障通透性。 因此，这项提议的重点是识别和验证一组新的抗体和 同源转运体可以调节治疗学中的血脑屏障交叉。通过挖掘鉴定出抗体。 针对包含人诱导的多能干细胞的血脑屏障模型，建立了一个大型噬菌体展示抗体库。 来源脑微血管内皮样细胞(BMECs)。这些BMEC的势垒特性很好- 适用于筛选大的抗体库，以寻找那些能够在体外通过血脑屏障的抗体。此外，在 这些细胞中的转录组水平和转运体特征与新鲜分离的人类 BMEC。最后，人类来源确保识别的抗体识别人类血脑屏障转运体。 使用这个创新的平台，我们已经确定了一组高价值的抗体，初步数据表明 能够结合人和小鼠的血脑屏障，并提供药理上相关的量 把毒品运到小鼠的大脑里。我们建议通过识别这些脑靶向抗体来进一步验证这些抗体 他们的同源传送器。接下来，我们将测定它们的药代动力学性质、生物分布和脑内情况。 区域特异性。最后，将测试抗体是否具有调节大脑吸收结合药物的能力。 以小鼠中风模型的形式运送到正常大脑和疾病大脑。那些表现出的抗体 显著和选择性的脑摄取将代表着具有潜力的新的非侵入性药物输送载体 在许多神经疾病环境中应用。

ABSTRACT Millions of people worldwide suffer from neurological diseases such as Alzheimer’s disease, stroke, and brain cancer. Advances in protein/gene profiling techniques and high throughput drug screening technologies have spawned many new drug candidates. However, the blood-brain barrier (BBB) has impeded the development and clinical realization of this new generation of neurotherapeutics by restricting the brain uptake of most small molecule therapeutics, and prohibiting brain uptake of protein- and gene-based medicines. A promising noninvasive brain delivery strategy takes advantage of endogenous BBB transport mechanisms as a means to shuttle drug cargo from the blood to the brain. Such receptor-mediated transport systems can be targeted using the exquisite specificity of antibodies that are in turn linked to a drug payload that can include small molecules, proteins, or DNA therapeutics. After binding to the receptor on the blood side, the antibody- drug conjugate acts as an artificial substrate for the transporter and is transcytosed from the blood, across the BBB, and into the brain. However, current approaches have yielded limited brain uptake because the targeted transporters are ubiquitously expressed and the antibody targeting reagents have low BBB permeability. Therefore, this proposal is focused on the identification and validation of a new panel of antibodies and cognate transporters that can mediate BBB crossing of therapeutics. The antibodies were identified by mining a large phage display antibody library against a BBB model comprising human induced pluripotent stem cell- derived brain microvascular endothelial-like cells (BMECs). The barrier characteristics of these BMECs are well- suited to screen large antibody libraries for those antibodies capable of crossing the BBB in vitro. Moreover, at the transcriptome level, the transporter profiles in these cells correlate quite well to freshly isolated human BMECs. Finally, the human sourcing ensures that identified antibodies recognize human BBB transporters. Using this innovative platform, we have identified a panel of high value antibodies that preliminary data indicate are capable of binding both the human and murine BBB and delivering pharmacologically relevant amounts of drug cargo to the murine brain. We propose to further validate these brain targeting antibodies by identifying their cognate transporters. Next, we will determine their pharmacokinetic properties, biodistribution and brain regiospecificity. Finally, the antibodies will be tested for their ability to mediate brain uptake of conjugated drug cargo to normal brain and to diseased brain in the form of a murine stroke model. Those antibodies that exhibit significant and selective brain uptake would represent new, noninvasive drug delivery vectors with potential application in many neurological disease settings.