Binding and functional properties of a mouse-human chimeric monoclonal antibody of the human IgG1 subclass with specificity for human carcinomas

Liao, Shuen‐Kuei; Horton, Linda W.; Flahart, Robert E.; O'Rear, Lynda; Crumpacker, Dina B.; Imbaratto, John W.; Yannelli, John R.; Robinson, Randy R.; Oldham, Robert K.

doi:10.3233/hab-1990-1201

Cited by 8 publications

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“…Both antibody conjugates expressed enzyme activity and bound to antigen. The percentage of conjugate that bound to antigen was 75-85%, and the K d values are in agreement with literature indicating that the antigen combining region of the antibody was not adversely effected by the conjugation process (30,46,47).…”

Section: Conjugation Of Mutant Hcpa1-t268g To An Antibody-tosupporting

confidence: 76%

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Toward Antibody-directed Enzyme Prodrug Therapy with the T268G Mutant of Human Carboxypeptidase A1 and Novel in VivoStable Prodrugs of Methotrexate

Smith

Banks

Blumenkopf³

et al. 1997

Journal of Biological Chemistry

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Antibody-directed enzyme prodrug therapy (ADEPT) has the potential of greatly enhancing antitumor selectivity of cancer therapy by synthesizing chemotherapeutic agents selectively at tumor sites. This therapy is based upon targeting a prodrug-activating enzyme to a tumor by attaching the enzyme to a tumor-selective antibody and dosing the enzyme-antibody conjugate systemically. After the enzyme-antibody conjugate is localized to the tumor, the prodrug is then also dosed systemically, and the previously targeted enzyme converts it to the active drug selectively at the tumor. Unfortunately, most enzymes capable of this specific, tumor site generation of drugs are foreign to the human body and as such are expected to raise an immune response when injected, which will limit their repeated administration. We reasoned that with the power of crystallography, molecular modeling and site-directed mutagenesis, this problem could be addressed through the development of a human enzyme that is capable of catalyzing a reaction that is otherwise not carried out in the human body. This would then allow use of prodrugs that are otherwise stable in vivo but that are substrates for a tumor-targeted mutant human enzyme. We report here the first test of this concept using the human enzyme carboxypeptidase A1 (hCPA1) and prodrugs of methotrexate (MTX). Based upon a computer model of the human enzyme built from the well known crystal structure of bovine carboxypeptidase A, we have designed and synthesized novel bulky phenylalanine-and tyrosine-based prodrugs of MTX that are metabolically stable in vivo and are not substrates for wild type human carboxypeptidases A. Two of these analogs are MTX-␣-3-cyclobutylphenylalanine and MTX-␣-3-cyclopentyltyrosine. Also based upon the computer model, we have designed and produced a mutant of human carboxypeptidase A1, changed at position 268 from the wild type threonine to a glycine (hCPA1-T268G). This novel enzyme is capable of using the in vivo stable prodrugs, which are not substrates for the wild type hCPA1, as efficiently as the wild type hCPA1 uses its best sub-

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Section: Conjugation Of Mutant Hcpa1-t268g To An Antibody-tosupporting

confidence: 76%

“…ING-1 binds to Epcam, a molecule expressed on a variety of epithelial cell tumors (46). Campath-1H binds to CDw52, which is expressed on a variety of T and B cells but not on most epithelial cells or tumors (30,47).…”

Section: Conjugation Of Mutant Hcpa1-t268g To An Antibody-tomentioning

confidence: 99%

Toward Antibody-directed Enzyme Prodrug Therapy with the T268G Mutant of Human Carboxypeptidase A1 and Novel in VivoStable Prodrugs of Methotrexate

Smith

Banks

Blumenkopf³

et al. 1997

Journal of Biological Chemistry

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Designing Antibodies for Human Therapy

Mayforth¹

1993

Designing Antibodies

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In Vitro and In Vivo Pharmacology and Pharmacokinetics of a Human Engineered™ Monoclonal Antibody to Epithelial Cell Adhesion Molecule

et al. 2003

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ING-1(heMAb), a Human Engineered monoclonal antibody to epithelial cell adhesion molecule (Ep-CAM), was evaluated for its in vitro and in vivo activity. The dissociation constant of ING-1(heMAb) for binding to Ep-CAM on HT-29 human colon tumor cells was 2 to 5 nM, similar to chimeric ING-1. In antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity assays, ING-1(heMAb) caused a concentration-dependent lysis of BT-20 breast, MCF-7 breast, HT-29 colon, and CACO-2 colon tumor cells, with maximum cytolysis at approximately 1 microg/ml. After an intravenous injection in rats, plasma ING-1(heMAb) levels declined with an alpha half-life of 8 to 11 hours, and a beta half-life of 20 days, typical of an IgG in a species without the target for ING-1. In nude mice with human HT-29 colon tumors, plasma ING-1(heMAb) levels declined more rapidly than in non-tumor-bearing mice, suggesting an enhanced clearance via the tumor-associated human Ep-CAM. In nude mice, intravenous treatments with ING-1(heMAb) twice a week for 3 weeks significantly suppressed the growth of human HT-29 colon and PC-3 prostate tumors in a dose-dependent manner, with 1.0 mg/kg providing the greatest benefit. These results indicate that Human Engineered ING-1(heMAb) is a high-affinity antibody with potent in vitro activity that targets and suppresses the growth of human tumors in vivo.

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Binding and functional properties of a mouse-human chimeric monoclonal antibody of the human IgG1 subclass with specificity for human carcinomas

Cited by 8 publications

References 0 publications

Toward Antibody-directed Enzyme Prodrug Therapy with the T268G Mutant of Human Carboxypeptidase A1 and Novel in VivoStable Prodrugs of Methotrexate

Toward Antibody-directed Enzyme Prodrug Therapy with the T268G Mutant of Human Carboxypeptidase A1 and Novel in VivoStable Prodrugs of Methotrexate

Designing Antibodies for Human Therapy

In Vitro and In Vivo Pharmacology and Pharmacokinetics of a Human Engineered™ Monoclonal Antibody to Epithelial Cell Adhesion Molecule

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