Inhibition of vacuolar ATPase subunit in tumor cells delays tumor growth by decreasing the essential macrophage population in the tumor microenvironment

Abstract

In cancer cells, vacuolar ATPase (V-ATPase), a multi-subunit enzyme, is expressed on the plasma as well as vesicular membranes and critically influences metastatic behavior. The soluble, cleaved N-terminal domain of V-ATPase a2 isoform is associated with in vitro induction of tumorigenic characteristics in macrophages. This activity led us to further investigate its in vivo role in cancer progression by inhibition of a2 isoform (a2V) in tumor cells and the concomitant effect on tumor microenvironment in the mouse 4T-1 breast cancer model. Results showed that macrophages cocultivated with a2V knockdown (sh-a2) 4T-1 cells produce lower amounts of tumorigenic factors in vitro and have reduced ability to suppress T-cell activation and proliferation compared with control 4T-1 cells. Data analysis showed a delayed mammary tumor growth in Balb/c mice inoculated with sh-a2 4T-1 cells compared with control. The purified CD11b(+) macrophages from sh-a2 tumors showed a reduced expression of mannose receptor-1 (CD206), interleukin-10, transforming growth factor-β, arginase-1, matrix metalloproteinase and vascular endothelial growth factor. Flow cytometric analysis of tumor-infiltrated macrophages showed a significantly low number of F4/80(+)CD11c(+)CD206(+) macrophages in sh-a2 tumors compared with control. In sh-a2 tumors, most of the macrophages were F4/80(+)CD11c(+) (antitumor M1 macrophages) suggesting it to be the reason behind delayed tumor growth. Additionally, tumor-infiltrating macrophages from sh-a2 tumors showed a reduced expression of CD206 compared with control whereas CD11c expression was unaffected. These findings demonstrate that in the absence of a2V in tumor cells, the resident macrophage population in the tumor microenvironment is altered which affects in vivo tumor growth. We suggest that by involving the host immune system, tumor growth can be controlled through targeting of a2V on tumor cells.

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

“…For example, a soluble cleavage fragment of V o a2 (a2NTD) has been shown to polarize macrophages toward a “tumor‐associated” or M2 macrophage phenotype 40, 69…”

Cancer derived peptide of vacuolar ATPase ‘a2’ isoform promotes neutrophil migration by autocrine secretion of IL-8

“…Important initiators that mediate the interaction between tumor cells and leukocytes are the vacuolar ATPases (V-ATPases); that are highly expressed on cancer cell surface456789. V-ATPases are novel targets for anticancer therapeutics45.…”

“…Our previous studies have shown that specifically the a2-isoform of the ‘a’ subunit of V-ATPase (a2V) plays a significant role in the inflammation associated with cancer and pregnancy82122. In cancer cells, a2V is highly expressed on the plasma membrane and the N-terminal domain of a2V (a2NTD) is secreted in microvesicles68921. a2NTD stimulates M2 polarization of the macrophages, which behave as TAM and have a positive impact on tumor progression168.…”

“…In cancer cells, a2V is highly expressed on the plasma membrane and the N-terminal domain of a2V (a2NTD) is secreted in microvesicles68921. a2NTD stimulates M2 polarization of the macrophages, which behave as TAM and have a positive impact on tumor progression168. Our recent study showed that a2NTD promotes the pro-tumoral properties of neutrophils that stimulate the tumor cell invasion and angiogenesis23.…”

Selective upregulation of TNFα expression in classically-activated human monocyte-derived macrophages (M1) through pharmacological interference with V-ATPase

“…Thus, the lysosomal adaptor protein Lamtor1, which forms an amino-acid sensing complex with V-ATPase, was recently shown to be required for M2 polarization [55]. Moreover, V-ATPase was reported to modulate macrophage polarization in tumor-bearing mice [56], and inhibition of the V-ATPase a2 isoform in murine tumor cells delayed tumor growth by decreasing M2-like TAMs in the tumor microenvironment [57].…”