Open AccessDual functions of a monoclonal antibody against cell surface F1F0 ATP synthase on both HUVEC and tumor cells 1
Author(s) -
ZHANG Xia,
GAO Feng,
YU Lili,
PENG Yan,
LIU Honghai,
LIU Jinying,
YIN Ming,
NI Jian
Publication year - 2008
Publication title -
acta pharmacologica sinica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.514
H-Index - 90
eISSN - 1745-7254
pISSN - 1671-4083
DOI - 10.1111/j.1745-7254.2008.00830.x
Subject(s) - microbiology and biotechnology , monoclonal antibody , flow cytometry , chemistry , cell , matrigel , cell growth , human umbilical vein endothelial cell , cytotoxicity , cell culture , mtt assay , umbilical vein , antibody , biology , biochemistry , in vitro , immunology , genetics
Aim: To generate a monoclonal antibody (McAb) against cell surface F1F0 AT P synthase (ATPase) and observe its antitumoral activity on both human umbilical vein endothelial cells (HUVEC) and tumor cells. Methods: Hybridoma cells secreting McAb against ATPase were produced by polyethylene glycol‐mediated fusions and screened by ELISA. The specificity of McAb was demonstrated by immunofluorescence and confocal imaging, as well as flow cytometry analysis. After the blockade of surface ATPase with McAb on HUVEC and human breast adenocarcinoma MDA‐MB‐231 cells, an AT P determination kit and CellTiter 96 AQueous Assay (MTS) assay were used to detect the effect of the antibody on extracellular ATP modification and cell proliferation. A cellular cytotoxicity assay in combination with doxorubicin, and a cell migration assay on MDA‐MB‐231 cells were used to determine the antitumoral activity. Finally, a HUVEC tube formation assay was used to detect the antiangiogenic effect of McAb178‐5G10. Results: A monoclonal antibody (McAb178‐5G10) against the β‐subunit of ATPase was generated, and its reactivity toward HUVEC and tumor cells was studied. We demonstrate that McAb178‐5G10 binds to ATPase at the cell surface, where it is able to inhibit AT P synthesis. This antibody also prevents the proliferation of HUVEC and MDA‐MB‐231 cells. Furthermore, McAb178‐5G10 enhances the tumoricidal effects of doxorubicin ( P <0.05), inhibits the migration of MDA‐MB‐231 in transwell assays ( P <0.01), and disrupts HUVEC tube formation on Matrigel ( P <0.01). Conclusion: McAb178‐5G10 binds preferentially to cell surface ATPase, blocks ATP synthesis, and exhibits both antiangiogenic and antitumorigenic effects.