Pronounced antitumor efficacy by extracellular activation of a doxorubicin-glucuronide prodrug after adenoviral vector-mediated expression of a human antibody-enzyme fusion protein.
Tumor-specific activation of the glucuronide prodrug of doxorubicin, N-[4-doxorubicin-N-carbonyl(oxymethyl)phenyl]-O-beta-glucuronyl carbamate (DOX-GA3), by beta-glucuronidase present in necrotic tumor areas might be improved after transduction of tumor cells to secrete a targeted form of beta-glucuronidase. To that end, we constructed an adenovirus vector, designated Ad/C28-GUSh, encoding human beta-glucuronidase fused to a human single-chain Fv (scFv) against the epithelial cell adhesion molecule (EpCAM), C28, and preceded by a signal sequence for secretion. Antibody specificity and enzyme activity were retained in the fusion protein secreted by tumor cells infected with Ad/C28-GUSh. Diffusion of fusion protein from transduced tumor cells within MCF-7 multicellular spheroids was visualized by immunohistochemistry. Treatment of spheroids with Ad/C28-GUSh and DOX-GA3 resulted in growth inhibition comparable to treatment with doxorubicin alone. Treatment of well-established FMa human ovarian cancer xenografts with intravenous injection of DOX-GA3 (500 mg/kg) resulted in a tumor volume-doubling time of 23.8 days compared to 8.0 days for phosphate-buffered saline (PBS)-treated mice. Intratumoral administration of Ad/C28-GUSh before DOX-GA3 enhanced the growth inhibition and increased the tumor volume-doubling time to 43.1 days (p < 0.01), while virus alone had no effect. Thus, we have successfully shown that an adenovirus vector encoding a secreted, targeted form of human beta-glucuronidase can further improve DOX-GA3 monotherapy.
de Graaf
Michelle
M
Department of Medical Oncology, Division of Gene Therapy, VU University Medical Center, 1007 MB Amsterdam, The Netherlands.
Pinedo
Herbert M
HM
Oosterhoff
Dinja
D
van der Meulen-Muileman
Ida H
IH
Gerritsen
Winald R
WR
Haisma
Hidde J
HJ
Boven
Epie
E
D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D000906:Antibodies / Q000276:immunology; D000951:Antigens, Neoplasm / Q000276:immunology; D000970:Antineoplastic Agents / Q000627:therapeutic use*; D015815:Cell Adhesion Molecules / Q000276:immunology; D045744:Cell Line, Tumor; D004317:Doxorubicin / Q000031:analogs & derivatives* / Q000627:therapeutic use*; D000071858:Epithelial Cell Adhesion Molecule; D005260:Female; D005434:Flow Cytometry; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D005965:Glucuronates / Q000627:therapeutic use*; D005966:Glucuronidase / Q000378:metabolism; D006801:Humans; D051379:Mice; D009369:Neoplasms / Q000188:drug therapy*; D010051:Ovarian Neoplasms / Q000188:drug therapy; D011355:Prodrugs / Q000627:therapeutic use*; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000378:metabolism; D018874:Spheroids, Cellular; D023041:Xenograft Model Antitumor Assays
C495486:Ad-C28-GUSh; D000906:Antibodies; D000951:Antigens, Neoplasm; D000970:Antineoplastic Agents; D015815:Cell Adhesion Molecules; D000071858:Epithelial Cell Adhesion Molecule; D005965:Glucuronates; C407038:N-(4-doxorubicin-N-carbonyl(oxymethyl)phenyl)-O-glucuronyl carbamate; D011355:Prodrugs; D011993:Recombinant Fusion Proteins; D004317:Doxorubicin; D005966:Glucuronidase