Sakurai Fuminori, Inoue Shunsuke, Kaminade Tadataka, Hotani Takuma, Katayama Yuki, Hosoyamada Eri, Terasawa Yuichi, Tachibana Masashi, Mizuguchi Hiroyuki
Laboratory of Biochemistry and Molecular Biology, Graduate School of Pharmaceutical Sciences, Osaka University, Japan; Laboratory of Regulatory Sciences for Oligonucleotide Therapeutics, Clinical Drug Development Unit, Graduate School of Pharmaceutical Sciences, Osaka University, Osaka, Japan.
Laboratory of Biochemistry and Molecular Biology, Graduate School of Pharmaceutical Sciences, Osaka University, Japan.
Int J Pharm. 2017 May 30;524(1-2):238-247. doi: 10.1016/j.ijpharm.2017.04.006. Epub 2017 Apr 4.
Reovirus induces tumor cell death efficiently and specifically, and thus is currently undergoing clinical testing as an anticancer agent. In the intracellular trafficking of reovirus, proteolytic disassembly of reovirus capsid-proteins and subsequent penetration of viral particles into the cytosol are crucial steps. Cathepsins B and L are largely responsible for the proteolytic disassembly of reovirus. Reovirus efficiently lyses tumor cells exhibiting relatively high activities of cathepsins B and L, while tumor cells with low activities of cathepsins B and L are often refractory to reovirus, probably due to inefficient endo/lysosomal escape. In this study, in order to enhance the tumor cell-killing efficiencies of reovirus by promoting endo/lysosomal escape, especially in reovirus-resistant tumor cells, reovirus was complexed with a cationic liposome transfection reagent. Reovirus alone and reovirus-cationic liposome complex (reoplex) exhibited similar levels of tumor cell-killing efficiencies in reovirus-susceptible tumor cells, while reoplex mediated more than 30% higher levels of tumor cell-killing activities in reovirus-resistant tumor cells than reovirus alone. Reoplex-mediated tumor cell death was efficiently induced in the tumor cells pretreated with cathepsin inhibitors. The mRNA levels of interferon (IFN)-β and apoptotic genes were significantly elevated following reoplex treatment. These results suggest that cationic liposomes efficiently promoted delivery of reovirus to the cytosol, leading to induction of apoptosis.
呼肠孤病毒能高效且特异性地诱导肿瘤细胞死亡，因此目前正作为一种抗癌药物进行临床试验。在呼肠孤病毒的细胞内运输过程中，病毒衣壳蛋白的蛋白水解性解离以及随后病毒颗粒进入细胞质是关键步骤。组织蛋白酶B和L在很大程度上负责呼肠孤病毒的蛋白水解性解离。呼肠孤病毒能有效裂解组织蛋白酶B和L活性相对较高的肿瘤细胞，而组织蛋白酶B和L活性较低的肿瘤细胞通常对呼肠孤病毒具有抗性，这可能是由于内吞/溶酶体逃逸效率低下所致。在本研究中，为了通过促进内吞/溶酶体逃逸来提高呼肠孤病毒对肿瘤细胞的杀伤效率，尤其是在对呼肠孤病毒耐药的肿瘤细胞中，将呼肠孤病毒与阳离子脂质体转染试剂复合。单独的呼肠孤病毒和呼肠孤病毒-阳离子脂质体复合物（复配物）在对呼肠孤病毒敏感的肿瘤细胞中表现出相似水平的肿瘤细胞杀伤效率，而在对呼肠孤病毒耐药的肿瘤细胞中，复配物介导的肿瘤细胞杀伤活性比单独的呼肠孤病毒高30%以上。在用组织蛋白酶抑制剂预处理的肿瘤细胞中，复配物介导的肿瘤细胞死亡被有效诱导。复配物处理后，干扰素（IFN）-β和凋亡基因的mRNA水平显著升高。这些结果表明，阳离子脂质体有效地促进了呼肠孤病毒向细胞质的递送，从而导致凋亡的诱导。
