Immunotherapy of B-Cell Lymphoma with an Engineered Bispecific Antibody Targeting CD19 and CD5

Sandra Lüttgau,Hossein Panjideh,Frank Breitling,Antonio Pezzutto,Oliver Schmetzer,Martin Lipp,Regina Fertig,Gerhard Moldenhauer,Dorothée Deppe,Saskia Meyer

doi:10.3390/antib2020338

Sandra Lüttgau, Hossein Panjideh + Show 8 more

Open Access

https://doi.org/10.3390/antib2020338

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Abstract

Using genetic engineering a humanized Fab fragment with specificity for CD19 was fused to a disulfide-stabilized single-chain antibody (dsFv) recognizing CD5. This format should show reduced immunogenicity and improved tissue penetration. The specificity of bsAb FabCD19xdsFvCD5 binding to target cells was verified by flow cytometry on B and T lymphoma cell lines. Binding affinities of both arms were compared with the bivalent parental antibodies against CD19 and CD5 by binding competition assay. Redirected lysis of B lymphoma cells by preactivated PBMC from healthy donors was demonstrated in a chromium-release assay. A clear dose-response relationship could be established in the range from 1 ng/mL to 10 mg/mL bsAb. To evaluate the in vivo efficacy of bsAb FabCD19xdsFvCD5, NOD/SCID mice were intravenously injected with luciferase transfected Raji lymphoma cells together with pre-activated PBMC. Mice received five injections of therapeutic bsAb or control antibodies. While in the control groups all mice died within 40 to 50 days, 40% of bsAb treated animals survived longer than 60 days.

Highlights

Non-Hodgkin’s lymphoma (NHL) is the most common lymphoma with an estimated 70,130 new cases and 18,940 deaths in 2012 as registered in the United States [1]
An antibody format was chosen in which the Fab portion of the CD19-specific chimeric antibody chiBCE19 was fused to the disulfide-stabilized Fv fragment of the CD5-directed antibody T5.16
It has to be noted that a remarkable variation of cytotoxicity among individual blood donors was observed. These results provided unambiguous evidence that the Bispecific antibodies (bsAb) is able to redirect activated T lymphocytes versus CD19-expressing target cells

Summary

Introduction

Non-Hodgkin’s lymphoma (NHL) is the most common lymphoma with an estimated 70,130 new cases and 18,940 deaths in 2012 as registered in the United States [1]. Its absence in childhood and acute lymphoblastic leukemia (B-ALL) has limited the use of CD20 antibodies. Another point of concern is the development of resistance to anti-CD20 therapy caused by a variety of mechanisms as for instance CD20 antigen down regulation, decreased expression of pro apoptotic genes of the Bcl-2 family and polymorphism of Fc-gamma receptors [4]. Bispecific antibodies (bsAb) represent a new class of therapeutics that holds big promise for the treatment of NHL. They are man-made molecules, which carry two different antigen binding sites. They are excellent activators of the immune system and have been generated against different combinations of tumor antigens and trigger molecules on effector cells [5]

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