Targeting lysosomal HSP70 induces acid sphingomyelinase-mediated disturbance of lipid metabolism and leads to cell death in T cell malignancies

Abstract

Background: 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.

Methods: 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.

Results: 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.

Conclusions: 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.

Keywords: T cell malignancies; acid sphingomyelinase; heat shock protein 70; lipid metabolism; lysosomal membrane permeabilization; lysosome.