Caveolae are critical cell surface structures important in coordinated cell signaling and endocytosis. One of the major proteins of caveolae is caveolin 1 (Cav-1). Cellular levels of Cav-1 are associated with cancer progression. In prostate cancer cells, levels of Cav-1 are positively correlated with tumor progression and metastasis. Cav-1 can be secreted by prostate cancer cells into the microenvironment and triggers proliferation and anti-apoptosis of the tumor and tumor endothelial cells. Clinical studies have shown increased serum Cav-1 levels in patients with poor prognosis. In tissue culture and animal model experiments, blocking secreted Cav-1 by polyclonal antibodies inhibits tumor cell growth. Cav-1 is therefore a potential therapeutic target for prostate cancer treatment. In this study, we used Cav-1 knock-out mice as hosts to produce monoclonal anti-Cav-1 antibodies. A total of 11 hybridoma cell lines were selected for their ability to produce antibodies that bound GST-Cav-1 but not GST on glutathione-coated ELISA plates. Further screening with ELISAs using GST-Cav-1 fragments on GSH-coated plates classified these antibodies into four groups: N1-31 with five antibodies binds the far N-terminus between amino acids 1 and 31; N32-80 with three antibodies binds between amino acids 32 and 80; CSD with two antibodies potentially bind the scaffolding domain (amino acids 80-101); and Cav-1-C with 1 antibody binds parts of the C-terminal half. Binding affinities (Kd) of these antibodies to soluble Cav-1 ranged from 10 -11 to 10 -8 M. Binding competition experiments revealed that these antibodies recognized a total of six different epitopes on Cav-1. Potency of these antibodies to neutralize Cav-1-mediated signaling pathways in cultured cells and in animal models will be tested. A selected monoclonal antibody will then be humanized and be further developed into a potential anti-prostate cancer therapeutic.
ASJC Scopus subject areas
- Immunology and Allergy