Inhibitors of vacuolar ATPase proton pumps inhibit human prostate cancer cell invasion and prostate‐specific antigen expression and secretion

Abstract

Vacuolar ATPases (V-ATPases) comprise specialized and ubiquitously distributed pumps that acidify intracellular compartments and energize membranes. To gain new insights into the roles of V-ATPases in prostate cancer (PCa) we studied the effects of inhibiting V-ATPase pumps in androgen-dependent (LNCaP) and androgen-independent (C4-2B) cells of a human PCa progression model. Treatment with nanomolar concentrations of the V-ATPase inhibitors bafilomycin A or concanamycin A reduced the in vitro invasion in both cell types by 80%, regardless that V-ATPase was prominent at the plasma membrane of C4-2B cells and only traces were detected in the low-metastatic LNCaP parental cells. In both cell types intracellular V-ATPase was excessive and co-localized with prostate-specific antigen (PSA) in the Golgi compartment. V-ATPase inhibitors reversibly excluded PSA from the Golgi and led to the accumulation of largely dispersed PSA-loaded vesicles of lysosomal composition. Inhibition of acridine orange staining and transferrin receptor recycling suggested defective endosomal and lysosomal acidification. The inhibitors, additionally, interfered with the AR-PSA axis under conditions that reduced invasion. Bafilomycin A significantly reduced steady-state and R1881-induced PSA mRNA expression and secretion in the LNCaP cells which are androgen-dependent, but not in the C4-2B cells which are androgen ablation-resistant. In the C4-2B cells, an increased susceptibility to V-ATPase inhibitors was detected after longer treatments, as proliferation was reduced and reversibility of bafilomycin-induced responses impaired. These findings make V-ATPases attractive targets against early and advanced PCa tumors.

Deletion of Vacuolar Proton-translocating ATPase Voa Isoforms Clarifies the Role of Vacuolar pH as a Determinant of Virulence-associated Traits in Candida albicans*

“…For Western blots, vacuolar protein (80 g) was separated by SDS-PAGE and transferred to nitrocellulose overnight at 150 mA. The V 1 A subunit was visualized with a 1:000 dilution of anti-human V 1 A rabbit polyclonal antibody (30); the antibody cross-reacts with the C. albicans protein.…”

Vacuolar ATPase as a potential therapeutic target and mediator of treatment resistance in cancer

“…Increased cancer cell invasive activity is frequently associated with aberrant V‐ATPase plasma membrane localization suggesting that increased acidification of the extracellular space plays an important role 29, 33, 34, 37, 47, 48, 49, 50, 51, 52…”

“…In prostate cancer cells, V‐ATPase co‐localized with prostate specific antigen (PSA; a secreted tumor marker that is regulated by the androgen receptor [AR]) in the Golgi and V‐ATPase inhibition resulted in a relocalization of PSA to lysosome‐like intracellular vesicles 50. Interestingly, in androgen‐sensitive LNCaP prostate cancer cells, V‐ATPase inhibition also reduced PSA mRNA expression, suggesting upstream inhibition of AR's transcription activating activity 50…”

The Function of Vacuolar ATPase (V-ATPase) a Subunit Isoforms in Invasiveness of MCF10a and MCF10CA1a Human Breast Cancer Cells

“…MCF10a cells are spontaneously immortalized breast epithelial cells derived from a patient with fibrocystic disease that are unable to form tumors in immunocompromised mice (28). MCF10CA1a cells are a highly invasive cell line derived from H-Ras-transformed MCF10a cells (MCF10AT) after selection for the ability to forms tumors in mice (29,32,34). The effect of treatment of MCF10a and MCF10CA1a cells with the specific V-ATPase inhibitor concanamycin (100 nM) on invasion was determined using an in vitro Matrigel invasion assay (33).…”

“…Importantly, administration of a V-ATPase inhibitor blocks bone metastasis of B16-F10 (28). A recent report has demonstrated that inhibition of V-ATPases blocks invasion of prostate cancer cell lines as well (29).…”