A comparison of the in vitro and in vivo activities of IgG and F(ab')2 fragments of a mixture of three monoclonal anti-Her-2 antibodies.
OBJECTIVE
We have demonstrated previously that a mixture of three anti-Her-2 monoclonal antibodies (MAbs) that bind to different epitopes on the extracellular domain of Her-2 expressed on a human breast cancer cell line has more potent antitumor activity than the individual MAbs both in vitro and in xenografted severe combined immunodeficient mice. Because the activity of Herceptin is Fc dependent, we determined whether this would also be the case when a mixture of these three anti-Her-2 MAbs was used.


METHODS
IgG and highly purified F(ab')(2) fragments of the anti-Her-2 MAbs and Herceptin were prepared and evaluated for their ability to induce cell death, inhibit vascular endothelial growth factor secretion, and mediate antibody-dependent cellular cytotoxicity and complement-mediated cytotoxicity in vitro. They were also compared for their abilities to induce regression of large BT474 tumors in severe combined immunodeficient mice.


RESULTS
All of the F(ab')(2) fragments were >95% pure and, as expected, did not mediate antibody-dependent cellular cytotoxicity or complement-dependent cytotoxicity in vitro. The in vitro antiproliferative and proapoptotic effects of the IgGs and F(ab')(2) fragments were similar. In contrast, the IgGs had significant antitumor activity in vivo, whereas their F(ab')(2) fragments were only marginally effective even at 5-fold higher doses to offset their shorter half-lives.


CONCLUSIONS
These results confirm the importance of the Fc portion of Herceptin for optimal in vivo activity and demonstrate that even a mixture of three anti-Her-2 MAbs that are highly effective at inducing cell death in vitro requires Fc-mediated effector function for optimal in vivo activity.
Spiridon
Camelia I
CI
Cancer Immunobiology Center, University of Texas Southwestern Medical School, Dallas, Texas 75390, USA.
Guinn
Sarah
S
Vitetta
Ellen S
ES
D020533:Angiogenesis Inhibitors / Q000494:pharmacology; D000818:Animals; D017304:Annexin A5 / Q000494:pharmacology; D000911:Antibodies, Monoclonal / Q000737:chemistry; D061067:Antibodies, Monoclonal, Humanized; D017209:Apoptosis; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry*; D007128:Immunoglobulin Fragments / Q000737:chemistry; D007074:Immunoglobulin G / Q000737:chemistry*; D066298:In Vitro Techniques; D020128:Inhibitory Concentration 50; D051379:Mice; D008807:Mice, Inbred BALB C; D016513:Mice, SCID; D008856:Microscopy, Fluorescence; D009368:Neoplasm Transplantation; D017434:Protein Structure, Tertiary; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000737:chemistry*; D013997:Time Factors; D000068878:Trastuzumab; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism
D020533:Angiogenesis Inhibitors; D017304:Annexin A5; D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D000939:Epitopes; D007140:Immunoglobulin Fab Fragments; D007128:Immunoglobulin Fragments; D007074:Immunoglobulin G; D018719:Erb-b2 Receptor Tyrosine Kinases; D000068878:Trastuzumab; D042461:Vascular Endothelial Growth Factor A