Targeting granzyme B to tumor cells using a yoked human chorionic gonadotropin.

Abstract

PURPOSE:

Luteinizing hormone receptor (LHR) is found in abundance on human ovarian, breast, endometrial and prostate carcinomas but at only low levels on non-gonadal tissues. To selectively kill LHR-expressing tumors, granzyme B (GrB) was linked to a protein in which both chains of human chorionic gonadotropin were yoked together (YCG).

METHODS:

GrB-YCG was expressed and secreted from insect Sf9 cells. Its GrB enzymatic activity and binding affinity for hLHR were then characterized. The differential cytotoxicity of GrB-YCG versus GrB alone was tested in a panel of LHR-expressing tumor cells by SRB assay, and the mechanisms involved in the cell death were investigated by confocal fluorescence microscopy, flow cytometry, and western blot analysis.

RESULTS:

GrB-YCG was successfully expressed and secreted from Sf9 insect cells and purified from cell culture supernatants. The serine protease activity of GrB-YCG was equivalent to that of human recombinant GrB. An in vitro hormone binding assay revealed that the GrB-YCG molecule also retained the ability to bind to the LHR receptor with an affinity similar to that of native hCG. Upon cell binding, GrB-YCG was rapidly internalized into LHR-expressing human ovarian cancer cells and produced selective and potent tumor cell killing by inducing apoptosis through activation of caspase-3.

CONCLUSIONS:

These results validate LHR as a therapeutic target and indicate that delivery of the human pro-apoptotic enzyme GrB to tumor cells by yoked hCG has substantial selectivity and therapeutic potential for human tumors that express high levels of LHR such as ovarian carcinomas.

• Authors: Isao Kanatani, Xinjian Lin, Xiaoqin Yuan, Gerald Manorek, Xiying Shang, Lawrence H Cheung, Michael G Rosenblum, Stephen B Howell
• Journal: Cancer chemotherapy and pharmacology (Cancer Chemother Pharmacol) Vol. 68 Issue 4 Pg. 979-90 (Oct 2011) ISSN: 1432-0843 [Electronic] Germany
• PMID: 21327682 (Publication Type: Comparative Study, Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Chorionic Gonadotropin
• Receptors, LH
• Granzymes
• Caspase 3

Topics

• Animals
• Apoptosis (drug effects)
• Blotting, Western
• Caspase 3 (metabolism)
• Cell Line
• Cell Line, Tumor
• Cells, Cultured
• Chorionic Gonadotropin (chemistry, genetics, metabolism)
• Drug Delivery Systems
• Female
• Flow Cytometry
• Gene Expression Regulation, Neoplastic
• Granzymes (administration & dosage, metabolism, pharmacology)
• Humans
• Mice
• Microscopy, Confocal
• Ovarian Neoplasms (drug therapy, pathology)
• Protein Binding
• Receptors, LH (genetics, metabolism)
• Spodoptera