Wolak Daniel J, Pizzo Michelle E, Thorne Robert G
Division of Pharmaceutical Sciences, School of Pharmacy, University of Wisconsin-Madison, USA; Clinical Neuroengineering Training Program, University of Wisconsin-Madison, USA.
Division of Pharmaceutical Sciences, School of Pharmacy, University of Wisconsin-Madison, USA; Clinical Neuroengineering Training Program, University of Wisconsin-Madison, USA; Neuroscience Training Program, University of Wisconsin-Madison, USA; Cellular & Molecular Pathology Training Program, University of Wisconsin-Madison, USA; The Institute for Clinical & Translational Research, University of Wisconsin-Madison, USA.
J Control Release. 2015 Jan 10;197:78-86. doi: 10.1016/j.jconrel.2014.10.034. Epub 2014 Nov 7.
Antibody-based therapeutics exhibit great promise in the treatment of central nervous system (CNS) disorders given their unique customizable properties. Although several clinical trials have evaluated therapeutic antibodies for treatment of CNS disorders, success to date has likely been limited in part due to complex issues associated with antibody delivery to the brain and antibody distribution within the CNS compartment. Major obstacles to effective CNS delivery of full length immunoglobulin G (IgG) antibodies include transport across the blood-brain and blood-cerebrospinal fluid barriers. IgG diffusion within brain extracellular space (ECS) may also play a role in limiting central antibody distribution; however, IgG transport in brain ECS has not yet been explored using established in vivo methods. Here, we used real-time integrative optical imaging to measure the diffusion properties of fluorescently labeled, non-targeted IgG after pressure injection in both free solution and in adult rat neocortex in vivo, revealing IgG diffusion in free medium is ~10-fold greater than in brain ECS. The pronounced hindered diffusion of IgG in brain ECS is likely due to a number of general factors associated with the brain microenvironment (e.g. ECS volume fraction and geometry/width) but also molecule-specific factors such as IgG size, shape, charge and specific binding interactions with ECS components. Co-injection of labeled IgG with an excess of unlabeled Fc fragment yielded a small yet significant increase in the IgG effective diffusion coefficient in brain, suggesting that binding between the IgG Fc domain and endogenous Fc-specific receptors may contribute to the hindered mobility of IgG in brain ECS. Importantly, local IgG diffusion coefficients from integrative optical imaging were similar to those obtained from ex vivo fluorescence imaging of transport gradients across the pial brain surface following controlled intracisternal infusions in anesthetized animals. Taken together, our results confirm the importance of diffusive transport in the generation of whole brain distribution profiles after infusion into the cerebrospinal fluid, although convective transport in the perivascular spaces of cerebral blood vessels was also evident. Our quantitative in vivo diffusion measurements may allow for more accurate prediction of IgG brain distribution after intrathecal or intracerebroventricular infusion into the cerebrospinal fluid across different species, facilitating the evaluation of both new and existing strategies for CNS immunotherapy.
基于抗体的疗法因其独特的可定制特性，在中枢神经系统（CNS）疾病治疗中展现出巨大潜力。尽管已有多项临床试验评估了治疗性抗体用于治疗CNS疾病，但迄今为止的成功可能在一定程度上受到限制，这是由于与抗体递送至大脑以及抗体在CNS区域内分布相关的复杂问题。全长免疫球蛋白G（IgG）抗体有效递送至CNS的主要障碍包括跨越血脑屏障和血脑脊液屏障。IgG在脑细胞外间隙（ECS）中的扩散也可能在限制中枢抗体分布中起作用；然而，尚未使用成熟的体内方法探究IgG在脑ECS中的转运。在此，我们使用实时综合光学成像来测量在自由溶液中以及在成年大鼠体内新皮层中进行压力注射后荧光标记的非靶向IgG的扩散特性，结果显示IgG在自由介质中的扩散比在脑ECS中大约大10倍。IgG在脑ECS中明显受阻的扩散可能是由于一些与脑微环境相关的一般因素（例如ECS体积分数和几何形状/宽度），也包括分子特异性因素，如IgG的大小、形状、电荷以及与ECS成分的特异性结合相互作用。将标记的IgG与过量的未标记Fc片段共同注射，导致脑内IgG有效扩散系数有小幅但显著的增加，这表明IgG Fc结构域与内源性Fc特异性受体之间的结合可能导致IgG在脑ECS中迁移受阻。重要的是，综合光学成像得到的局部IgG扩散系数与在麻醉动物中进行控制性脑池内输注后，通过跨软脑膜脑表面的转运梯度进行离体荧光成像得到的结果相似。综上所述，我们的结果证实了扩散转运在向脑脊液中输注后全脑分布图谱形成中的重要性，尽管在脑血管周围间隙中的对流转运也很明显。我们的体内定量扩散测量可能允许更准确地预测鞘内或脑室内向脑脊液中输注后IgG在不同物种脑内的分布，有助于评估CNS免疫治疗的新策略和现有策略。
