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

Abstract

Background: Leukemia, 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. Methods. Monoclonal 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. Results: We detected cell-membrane expression of the F1F0 ATPase 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 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 50g/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 3.60.83%, 8.41.69% and 17.32.56% 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 5.52.37%, 11.33.62% and 19.93.31% compared to 1.56%0.97% in mouse IgG treated cells (p<0.01). Annexin V staining demonstrated that the relative apoptotic rates in 50g/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. Conclusions: These findings indicate that ectopic expression of ATPase subunit may be a tumor-associated antigen in hematological malignancies. The F1F0 ATPase subunit provides a potential target for immunotherapy in AML and hematological malignancies. © 2012 Wen-Li et al.; licensee BioMed Central Ltd.