Department of Medical Bioengineering, Okayama University, 3-1-1 Tsushimanaka, Okayama 700-8530, Japan.
Preeminent Medical Photonics Education and Research Center, Hamamatsu University School of Medicine, 1-20-1 Handayama, Higashi-ku, Hamamatsu 431-3192, Japan.
Bioconjug Chem. 2020 Mar 18;31(3):916-922. doi: 10.1021/acs.bioconjchem.0c00046. Epub 2020 Feb 18.
Cell-penetrating peptides (CPPs) are widely used for the intracellular delivery of peptides and proteins, but CPP fusion peptides and proteins are often transported by endocytosis and trapped in endosomes. Photochemical internalization (PCI) is a method for the endosomal escape of the trapped peptide or protein and release into the cytosol using light and photosensitizers. In PCI, endosomal membranes are thought to be destabilized by singlet oxygen (O) photogenerated from photosensitizers localized in endosomes. We previously developed CPP-cargo-photosensitizer (PS) conjugates able to photodependently enter the cytosol via the PCI mechanism. For example, TatU1A-PS (a covalent complex of Tat [CPP], U1A RNA-binding protein [cargo], and PS) can photodependently deliver RNAs into the cytosol, and TatBim-PS (a covalent complex of Tat, Bim [cargo], and PS) can photoinduce apoptosis in mammalian cells. However, for many newly created conjugates, the induction of PCI has been insufficient. We hypothesized that the amino acid linker sequence (XX) adjacent to the photosensitizer is an important determinant of PCI efficiency. In this study, using CPP-cargo-XX-PS platforms, we examined the relationship between PCI efficiency and the linker amino acid sequence near the photosensitizer. We found that hydrophobic FF and LL linkers enhanced the PCI efficiencies of both TatBim-XX-PS and TatU1A-XX-PS. The effectiveness of the linker depended, in part, on both the cargo moiety and the photosensitizer. These results may guide the design of CPP-cargo-PS conjugates conferring broad target functions for PCI and photodynamic therapy.
细胞穿透肽 (CPP) 广泛用于肽和蛋白质的细胞内递送，但 CPP 融合肽和蛋白质通常通过内吞作用运输，并被困在内体中。光化学内化 (PCI) 是一种利用光和光敏剂使被困的肽或蛋白质从内体中逃逸并释放到细胞质中的方法。在 PCI 中，内体膜被认为是通过定位于内体中的光敏剂产生的单线态氧 (O) 光破坏的。我们之前开发了 CPP-货物-光敏剂 (PS) 缀合物，能够通过 PCI 机制光依赖性地进入细胞质。例如，TatU1A-PS（Tat [CPP]、U1A RNA 结合蛋白 [货物] 和 PS 的共价复合物）可以光依赖性地将 RNA 递送到细胞质中，而 TatBim-PS（Tat、Bim [货物] 和 PS 的共价复合物）可以光诱导哺乳动物细胞凋亡。然而，对于许多新创建的缀合物，PCI 的诱导作用还不够。我们假设与 PS 相邻的氨基酸接头序列 (XX) 是 PCI 效率的重要决定因素。在这项研究中，我们使用 CPP-货物-XX-PS 平台，研究了 PCI 效率与 PS 附近接头氨基酸序列之间的关系。我们发现疏水性 FF 和 LL 接头增强了 TatBim-XX-PS 和 TatU1A-XX-PS 的 PCI 效率。接头的有效性部分取决于货物部分和光敏剂。这些结果可能为设计赋予 PCI 和光动力治疗广泛靶向功能的 CPP-货物-PS 缀合物提供指导。
