The Simon Flavell Leukaemia Research Laboratory, Southampton General Hospital, Southampton SO16 6YD, UK.
Biomedical Imaging Unit, University of Southampton Faculty of Medicine, Southampton General Hospital, Southampton SO16 6YD, UK.
Int J Mol Sci. 2020 Nov 19;21(22):8734. doi: 10.3390/ijms21228734.
Cholesterol seems to play a central role in the augmentation of saporin-based immunotoxin (IT) cytotoxicity by triterpenoid saponins. Endolysosomal escape has been proposed as one mechanism for the saponin-mediated enhancement of targeted toxins. We investigated the effects of lipid depletion followed by repletion on (SA)-induced endolysosomal escape of Alexa Fluor labelled saporin and the saporin-based immunotoxin OKT10-SAP, directed against CD38, in Daudi lymphoma cells. Lipid deprived cells showed reduced SA-induced endolysosomal escape at two concentrations of SA, as determined by a flow cytometric method. The repletion of membrane cholesterol by low density lipoprotein (LDL) restored SA-induced endolysosomal escape at a concentration of 5 µg/mL SA but not at 1 µg/mL SA. When LDL was used to restore the cholesterol levels in lipid deprived cells, the SA augmentation of OKT10-SAP cytotoxicity was partially restored at 1 µg/mL SA and fully restored at 5 µg/mL SA. These results suggest that different mechanisms of action might be involved for the two different concentrations of SA and that endosomal escape may not be the main mechanism for the augmentation of saporin IT cytotoxicity by SA at the sub-lytic concentration of 1 µg/mL SA.
胆固醇似乎在三萜皂苷增强基于相思子毒素的免疫毒素（IT）细胞毒性方面发挥核心作用。内体逃逸已被提议为皂苷介导靶向毒素增强的一种机制。我们研究了脂质耗竭后再补充对（SA）诱导的阿霉素淋巴瘤细胞中 Alexa Fluor 标记的相思子毒素和针对 CD38 的免疫毒素 OKT10-SAP 的内体逃逸的影响。用流式细胞术方法测定，在两种浓度的 SA 下，脂质剥夺细胞显示 SA 诱导的内体逃逸减少。通过低密度脂蛋白（LDL）补充膜胆固醇可在 5 µg/mL SA 的浓度下恢复 SA 诱导的内体逃逸，但在 1 µg/mL SA 的浓度下则不能。当用 LDL 恢复脂质剥夺细胞中的胆固醇水平时，SA 增强 OKT10-SAP 细胞毒性在 1 µg/mL SA 时部分恢复，在 5 µg/mL SA 时完全恢复。这些结果表明，对于两种不同浓度的 SA，可能涉及不同的作用机制，并且内体逃逸可能不是 SA 在亚溶血性浓度 1 µg/mL SA 增强相思子毒素 IT 细胞毒性的主要机制。
