State Key Laboratory of Natural Medicines, Jiangsu Key Laboratory of Carcinogenesis and Intervention, China Pharmaceutical University, Nanjing, China.
School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, China.
Clin Transl Med. 2023 Mar;13(3):e1229. doi: 10.1002/ctm2.1229.
T cell malignancies proliferate vigorously, are highly dependent on lysosomal function, with limited therapeutic options. Deregulation of lysosomal structure and function has been confirmed to be a key role in the treatment of hematologic malignant disease.
Cell counting kit 8 and Annexin V/PI staining were used to assess the cell viability and apoptosis rate. Flow cytometry, liquid chromatography mass spectrometry, immunofluorescence and western blot were performed to detect the effect on lysosomes. Drug affinity responsive target stability, molecular docking and cellular thermal shift assay were employed to confirm the target protein of V8 on lysosomes. A xenograft model was constructed in NOD/SCID mice to assess the effect and mechanism.
V8, a new lysosomotropic compound, could be rapidly trapped by lysosomes and accumulation in lysosomes, contributing to lysosomal-dependent cell death by evoking lysosomal membrane permeabilization (LMP), accompanied with disrupted lysosome and autophagic flux. Mechanistically, heat shock protein 70 (HSP70) was identified as the binding target of V8 in lysosome. As a downstream effect of targeting HSP70, enzymatic activity of acid sphingomyelinase (ASM) was inhibited, which induced disturbance of lipid metabolism, instability of lysosomal membrane, and leakage of cathepsin B and D, leading to LMP-mediated cell death. In vivo study showed V8 well controlled the growth of the tumour and confirmed lysosomal cell death induced by V8.
Collectively, this study suggests targeting lysosomal HSP70-ASM axis by V8 illustrates the great value of drug therapy for T cell malignancies and the unlimited potential of lysosomal targeting for cancer therapy.
T 细胞恶性肿瘤增殖活跃，高度依赖溶酶体功能，治疗选择有限。溶酶体结构和功能的失调已被证实是治疗血液恶性疾病的关键作用。
使用细胞计数试剂盒 8 和 Annexin V/PI 染色来评估细胞活力和细胞凋亡率。流式细胞术、液相色谱-质谱联用、免疫荧光和 Western blot 用于检测对溶酶体的影响。药物亲和反应靶标稳定性、分子对接和细胞热转移分析用于确认 V8 在溶酶体上的靶蛋白。在 NOD/SCID 小鼠中构建异种移植模型以评估效果和机制。
V8，一种新的溶酶体趋化化合物，可迅速被溶酶体捕获并在溶酶体中积累，通过引发溶酶体膜通透性（LMP）导致溶酶体依赖性细胞死亡，同时伴有溶酶体和自噬流的破坏。在机制上，热休克蛋白 70（HSP70）被鉴定为 V8 在溶酶体中的结合靶标。作为靶向 HSP70 的下游效应，酸性鞘磷脂酶（ASM）的酶活性被抑制，导致脂质代谢紊乱、溶酶体膜不稳定以及组织蛋白酶 B 和 D 的漏出，从而导致 LMP 介导的细胞死亡。体内研究表明，V8 很好地控制了肿瘤的生长，并证实了 V8 诱导的溶酶体细胞死亡。
总之，这项研究表明，V8 通过靶向溶酶体 HSP70-ASM 轴为 T 细胞恶性肿瘤的药物治疗提供了重要价值，并为溶酶体靶向治疗癌症提供了无限潜力。
