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Endostatin: yeast production, mutants, and antitumor effect in renal cell carcinoma. Cancer Res 1999 Jan 01;59(1):189-97

Date

01/19/1999

Pubmed ID

9892206

Abstract

Endostatin is a Mr 20,000 COOH-terminal fragment of collagen XVIII that inhibits the growth of several primary tumors. We report here the cloning and expression of mouse endostatin in both prokaryotic and eukaryotic expression systems. Soluble recombinant protein expressed in yeast (15-20 mg/L) inhibited the proliferation and migration of endothelial cells in response to stimulation by basic fibroblast growth factor. A rabbit polyclonal antibody was raised that showed positive immunoreactivity to the recombinant protein expressed from both systems. Importantly, the biological activity of the mouse recombinant protein could be neutralized by this antiserum in both endothelial proliferation and chorioallantoic membrane assays. Systemic administration of endostatin at 10 mg/kg suppressed the growth of renal cell cancer in a nude mouse model. The inhibition of tumor growth with soluble yeast-produced protein was comparable to that obtained with non-refolded precipitated protein expressed from bacteria. In addition, two closely related COOH-terminal deletion mutants of endostatin were also tested and showed strikingly differing activity. Collectively, these findings demonstrate the expression of a biologically active form of mouse endostatin in yeast, define a role for the molecule in inhibiting endothelial cell migration, extend its antitumor effects to renal cell carcinoma, and provide a formal proof (via the neutralizing antiserum experiments and the mutant data) that endostatin (and not a possible contaminant) acts as an antiangiogenic agent. Finally, the high level expression of mouse endostatin in yeast serves as an endotoxin free, soluble source of protein for fundamental studies on the mechanisms of tumor growth suppression by angiogenesis inhibitors.

Author List

Dhanabal M, Ramchandran R, Volk R, Stillman IE, Lombardo M, Iruela-Arispe ML, Simons M, Sukhatme VP

Author

Ramani Ramchandran PhD Professor in the Pediatrics department at Medical College of Wisconsin




MESH terms used to index this publication - Major topics in bold

Animals
Antineoplastic Agents
Carcinoma, Renal Cell
Cattle
Collagen
Collagen Type XVIII
Endostatins
Kidney Neoplasms
Mice
Mutation
Neoplasms, Experimental
Peptide Fragments
Rabbits
Recombinant Proteins
Saccharomyces cerevisiae
Tumor Cells, Cultured
jenkins-FCD Prod-484 8aa07fc50b7f6d102f3dda2f4c7056ff84294d1d