Recombinant, refolded tetrameric p53 and gonadotropin-releasing hormone-p53 slow proliferation and induce apoptosis in p53-deficient cancer cells.
The p53 tumor suppressor is mutated in over 50% of human cancers. Mutations resulting in amino acid changes within p53 result in a loss of activity and consequent changes in expression of genes that regulate DNA repair and cell cycle progression. Replacement of p53 using protein therapy would restore p53 function in p53-deficient tumor cells, with a consequence of tumor cell death and tumor regression. p53 functions in a tetrameric form in vivo. Here, we refolded a wild-type, full-length p53 from inclusion bodies expressed in Escherichia coli as a stable tetramer. The tetrameric p53 binds to p53-specific DNA and, when transformed into a p53-deficient cancer cell line, induced apoptosis of the transformed cells. Next, using the same expression and refolding technology, we produced a stable tetramer of recombinant gonadotropin-releasing hormone-p53 fusion protein (GnRH-p53), which traverses the plasma membrane, slows proliferation, and induces apoptosis in p53-deficient, GnRH-receptor-expressing cancer cell lines. In addition, we showed a time-dependent binding and internalization of GnRH-p53 to a receptor-expressing cell line. We conclude that the GnRH-p53 fusion strategy may provide a basis for constructing an effective cancer therapeutic for patients with tumors in GnRH-receptor-positive tissue types.
Lafevre-Bernt
Michelle
M
GeneCopoeia, Inc., 19520 Amaranth Drive, Germantown, MD 20874, USA.
Wu
Shili
S
Lin
Xinli
X
D017209:Apoptosis; D001681:Biological Assay; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D004705:Endocytosis; D007987:Gonadotropin-Releasing Hormone / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D009369:Neoplasms / Q000473:pathology*; D009707:Nucleosomes / Q000378:metabolism; D017510:Protein Folding; D020836:Protein Structure, Quaternary; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism*; D013816:Thermodynamics; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry* / Q000172:deficiency / Q000302:isolation & purification / Q000378:metabolism*
D009707:Nucleosomes; D011993:Recombinant Fusion Proteins; D016159:Tumor Suppressor Protein p53; D007987:Gonadotropin-Releasing Hormone