Department of Chemistry, Seoul National University, 1 Gwanak-ro, Gwanak-gu, Seoul 08826, Republic of Korea.
ERATO Hamachi Innovative Molecular Technology for Neuroscience, Graduate School of Engineering, Kyoto University Katsura, Katsura Int'tech Center #308, Nishikyo-ku, Kyoto 615-8530, Japan.
J Control Release. 2021 Feb 10;330:161-172. doi: 10.1016/j.jconrel.2020.12.020. Epub 2020 Dec 16.
A new vehicle is designed for the intracellular delivery of antibodies at nanomolar concentrations by combination of domain Z, a small affibody with strong binding affinity to Fc regions of immunoglobulin G (IgG), and the multimers of LK sequences, α-helical cell penetrating peptides (CPP) with powerful cell penetrating activities. Domain Z and multimeric LK are fused together to form LK-domain Z proteins. The LK-domain Z can bind with IgG at a specific ratio at nanomolar concentrations by simple mixing. The IgG/LK-domain Z complexes can successfully penetrate live cells at nanomolar concentration and the delivery efficiency is strongly dependent upon the concentrations of IgG/LK-domain Z complex as well as the species and subclasses of IgGs. The IgG/LK-domain Z complexes penetrate cells via ATP-dependent endocytosis pathway and the majority of delivered IgG seems to escape endosome to cytosol. Remarkably, the delivered IgGs are able to control the targeted intracellular signaling pathway as shown in the down-regulation of pro-survival genes by the delivery of anti-NF-κB using an LK-domain Z vehicle with a cathepsin B-cleavable linker between the LK sequence and domain Z. The simple but very efficient intracellular delivery method of antibodies at nanomolar concentrations is expected to facilitate profound understanding of cell mechanisms and development of new future therapeutics on the basis of intracellular antibodies.
一种新的载体通过融合域 Z（与免疫球蛋白 G（IgG）的 Fc 区域具有强结合亲和力的小亲和体）和 LK 序列的多聚体（具有强大细胞穿透活性的α螺旋细胞穿透肽（CPP）），被设计用于在纳摩尔浓度下将抗体进行细胞内传递。域 Z 和多聚体 LK 融合在一起形成 LK-域 Z 蛋白。通过简单混合，在纳摩尔浓度下，LK-域 Z 可以以特定比例与 IgG 结合。在纳摩尔浓度下，IgG/LK-域 Z 复合物能够成功穿透活细胞，并且传递效率强烈依赖于 IgG/LK-域 Z 复合物的浓度以及 IgGs 的种类和亚类。IgG/LK-域 Z 复合物通过 ATP 依赖性内吞作用途径穿透细胞，并且大部分递送的 IgG 似乎从内涵体逃逸到细胞质。值得注意的是，通过在 LK 序列和域 Z 之间使用具有组织蛋白酶 B 可切割连接子的 LK-域 Z 载体递送抗 NF-κB，递送的 IgG 能够控制靶向的细胞内信号通路，如通过递送抗-NF-κB 来下调生存基因。这种在纳摩尔浓度下简单但非常有效的抗体细胞内传递方法有望促进对细胞机制的深入理解，并基于细胞内抗体开发新的未来治疗方法。
