Abstract
BackgroundEmerging evidences suggest that nucleolin expressed on the cell surface is implicated in growth of tumor cells and angiogenesis. Nucleolin is one of the major proteins of the nucleolus, but it is also expressed on the cell surface where is serves as a binding protein for variety of ligands implicated in cell proliferation, differentiation, adhesion, mitogenesis and angiogenesis.Methodology/Principal FindingsBy using a specific antagonist that binds the C-terminal tail of nucleolin, the HB-19 pseudopeptide, here we show that the growth of tumor cells and angiogenesis are suppressed in various in vitro and in vivo experimental models. HB-19 inhibited colony formation in soft agar of tumor cell lines, impaired migration of endothelial cells and formation of capillary-like structures in collagen gel, and reduced blood vessel branching in the chick embryo chorioallantoic membrane. In athymic nude mice, HB-19 treatment markedly suppressed the progression of established human breast tumor cell xenografts in nude mice, and in some cases eliminated measurable tumors while displaying no toxicity to normal tissue. This potent antitumoral effect is attributed to the direct inhibitory action of HB-19 on both tumor and endothelial cells by blocking and down regulating surface nucleolin, but without any apparent effect on nucleolar nucleolin.Conclusion/SignificanceOur results illustrate the dual inhibitory action of HB-19 on the tumor development and the neovascularization process, thus validating the cell-surface expressed nucleolin as a strategic target for an effective cancer drug. Consequently, the HB-19 pseudopeptide provides a unique candidate to consider for innovative cancer therapy.
Highlights
Nucleolin is an abundant RNA- and protein-binding protein ubiquitously expressed in exponentially growing eukaryotic cells
The fact that HB-19 forms a stable complex with the cell-surface expressed nucleolin, incubation of cells with biotinylated HB-19 (HB-19/Btn) followed by purification of surface/cytoplasmic extracts using avidin-agarose provides an efficient method for purification surface nucleolin and monitoring its expression [8]
By FACScan analysis, we showed that HB-19/Btn binding to human umbilical vein endothelial cells (HUVECs) is specific since it is prevented by the excess HB-19 but not by the basic 9Arg peptide or the F3 peptide (Figure 3B), which is reported to bind the acidic amino acid region located at the N-terminal part of nucleolin [6]
Summary
Nucleolin is an abundant RNA- and protein-binding protein ubiquitously expressed in exponentially growing eukaryotic cells. Surface nucleolin serves as a low affinity receptor for HIV-1 and various growth factors that interact with the RGG domain of nucleolin, such as midkine, pleiotrophin (PTN) and lactoferrin [8,9,10,11]. Binding of these ligands results in clustering of cell-surface nucleolin in lipid raft membrane microdomains before endocytosis of the ligand-. Nucleolin is one of the major proteins of the nucleolus, but it is expressed on the cell surface where is serves as a binding protein for variety of ligands implicated in cell proliferation, differentiation, adhesion, mitogenesis and angiogenesis
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