Inhibition of the ecto-beta subunit of F1F0-ATPase inhibits proliferation and induces apoptosis in acute myeloid leukemia cell lines

Abstract

BackgroundLeukemia, a heterogeneous clonal disorder of hematopoietic progenitor cells, presents a world-wide health problem, especially in childhood. F1F0 ATPase, an inner mitochondrial enzyme, is expressed on the plasma membrane of tumor cells, and its inhibition induces both anti-angiogenic and anti-tumorigenic activity.MethodsMonoclonal Antibody (McAb) against ATPase was produced by polyethylene glycol-mediated fusions and screened by ELISA. Proliferation, cell cycle and apoptosis of cells were analyzed when the surface ATPase of cells was blockaded with McAb.ResultsWe detected cell-membrane expression of the F1F0 ATPase β subunit on 0.1% to 56% of the 11 cell lines derived from leukemia, including acute myeloid leukemia (AML). We produced a monoclonal antibody, McAb7E10, which recognizes both the native and recombinant ATPase β subunit, with a dissociation constant (KD) of 3.26E–10. We demonstrate that McAb7E10 binds to ATPase at the cell surface, where it is able to inhibit ATP synthesis. McAb7E10 significantly inhibited proliferation of AML cell lines in vitro: the relative inhibitory rates of 50 μg/mL McAb7E10 treated MV4-11and HL-60 cells were 69.6% and 81.9% respectively. Cell cycle analysis indicated that McAb7E10 significantly induced apoptosis in MV4-11 and HL-60 cells: the relative rates of apoptosis in 5, 10 and 50ug/mL McAb7E10 treated MV4-11 cells was 3.6 ± 0.83%, 8.4 ± 1.69% and 17.3 ± 2.56% compared to 1.5% ± 0.85% in mouse IgG treated cells (p < 0.01). The relative rate of apoptosis in 5, 10 and 50ug/mL McAb7E10 treated HL-60 cells was 5.5 ± 2.37%, 11.3 ± 3.62% and 19.9 ± 3.31% compared to 1.56% ± 0.97% in mouse IgG treated cells (p < 0.01). Annexin V staining demonstrated that the relative apoptotic rates in 50 μg/mL McAb7E10 treated MV4-11 and HL-60 cells were 50.5% ± 7.04% and 32.9% ± 4.52%, respectively, significantly higher than IgG control antibody treated cells were 21.9% ± 3.11% and 15.3% ± 3.95%, p < 0.01.ConclusionsThese findings indicate that ectopic expression of ATPase β subunit may be a tumor-associated antigen in hematological malignancies. The F1F0 ATPase β subunit provides a potential target for immunotherapy in AML and hematological malignancies.

The Multifaceted ATPase Inhibitory Factor 1 (IF1) in Energy Metabolism Reprogramming and Mitochondrial Dysfunction: A New Player in Age-Associated Disorders?

“…A similar activation of proliferation by ecto-F 1 -ATPase has been observed in lung and breast carcinoma and myeloid leukemia cell lines ( 22 , 23 , 114 , 159 ). Some studies have attributed the effect of ecto-F 1 -ATPase on tumorigenesis to the production of extracellular ATP, which could accommodate the increased energy requirement ( Fig.…”

The function of Cav-1 in MDA-MB-231 breast cancer cell migration and invasion induced by ectopic ATP5B

“…In the past decade, the beta subunit of ATPase has been shown to be ectopically expressed on the outer surface of both normal and tumor cells [ 23 , 27 – 29 ] and has been identified as a potential therapeutic target in NSCLC [ 30 ] and malignant paragangliomas [ 31 ]. Ectopic ATP5B is involved in the metabolism of lipids, activation of endothelial cells [ 32 ], development of an acidic tumor microenvironment [ 33 ], and proliferation and apoptosis of cancer cells [ 34 ]. ATP5B located on the inner membrane of mitochondria is essential for the conversion of ADP to ATP and provides energy for various life processes.…”

Early B-cell factor 3 (EBF3) is a novel tumor suppressor gene with promoter hypermethylation in pediatric acute myeloid leukemia

“…Leukemia cell lines HL-60, MV4-11, U937, THP-1 and K562 were obtained from the American Type Culture Collection (ATCC). CCRF, Raji, Jurkat, 697, Daudi and SHI-1 cell lines (gifts from Professor Wang Jian-Rong, The Cyrus Tang Hematology center of Soochow University) [ 28 ], NALM-6 cell lines (gifts from Professor Tang Yong-Ming, Zhejiang University). HL-60, MV4-11, U937, THP-1 and SHI-1 are AML cell lines.…”