Hsp90 Inhibitor STA9090 induced VPS35 related extracellular vesicle release and metastasis in hepatocellular carcinoma

•VPS35 involved in the EVs formation, delivery and secretion process induced by N-terminal Hsp90 inhibitor STA9090 but not C-terminal Hsp90 inhibitor NB and promoted HCC invasion.•Bclaf1 upregulated VPS35 transcription through bZIP DNA binding domain, and upregulation of VPS35 and Bclaf1 levels positively correlates with poorer prognosis and metastasis in HCC.•HSF1 positively correlated with Bclaf1 levels.•Hsp90 inhibitor STA9090 induced EVs promote invasion and metastasis by HSF1-Bclaf1-VPS35-EVs axis in hepatocellular carcinoma.•Bclaf1 down-regulation alleviates Hsp90 inhibitor STA9090 induced VPS35 related extracellular vesicle release and invasion. Heat shock protein 90 (Hsp90) has been an important therapeutic target for cancer therapy for decades. Unexpectedly, the monotherapy of N-terminal Hsp90 inhibitor STA9090 related clinical trials halted in phase III, and metastases were reported in animal models with the treatment of N-terminal Hsp90 inhibitors. Vacuolar protein sorting-associated protein 35 (VPS35) plays a vital role in endosome-derived EV (extracellular vesicle) traffic in neurodegeneration diseases, but no vps35 related EV were reported in tumors till now. Since tumor derived EVs contributes to metastasis and VPS35 is recently found to be involved in the invasion and metastasis of hepatocellular carcinoma (HCC), whether N-terminal Hsp90 inhibitor STA9090 induced EVs generation and the role of VPS35 in it were explored in this study. We found that N-terminal Hsp90 inhibitor STA9090 upregulated Bclaf1 and VPS35 levels, increased the secretion of EVs, and STA9090-induced-EVs promoted the invasion of HepG2 cells. As the clinical data suggested that the increased Bclaf1 and VPS35 levels correlated with increased metastasis and poorer prognosis in HCC, we focused on the Bclaf1-VPS35-EVs axis to further explore the mechanism of VPS35-related metastasis. The results demonstrated that Bclaf1 facilitated the transcription of VPS35 via bZIP domain, and knockdown of Bclaf1 or VPS35 alleviated pro-metastatic capability of STA9090-induced-EVs. All the results revealed the role of Bclaf1-VPS35-EVs axis on metastasis of HCC, and VPS35 knockdown decreased Hsp90 Inhibitor STA9090 induced extracellular vesicle release and metastasis, which provided a new combination therapeutic strategy to inhibit the metastasis of HCC caused by N-terminal Hsp90 inhibitor induced extracellular vesicles.