A recombinant bispecific single-chain antibody, CD19 × CD3, induces rapid and high lymphoma-directed cytotoxicity by unstimulated T lymphocytes
Abstract
Although bispecific antibodies directed against malignant lymphoma have been shown to be effective in vitro and in vivo, extended clinical trials so far have been hampered by the fact that conventional approaches to produce these antibodies suffer from low yields, ill-defined byproducts, or laborious purification procedures. To overcome this problem, we have generated a small, recombinant, lymphoma-directed, bispecific single-chain (bsc) antibody according to a novel technique recently described. The antibody consists of 2 different single-chain Fv fragments joined by a glycine-serine linker. One specificity is directed against the CD3 antigen of human T cells, and the other antigen-binding site engages the pan–B-cell marker CD19, uniformly expressed on the vast majority of B-cell malignancies. The construct was expressed in Chinese hamster ovary cells and purified by its C-terminal histioline tag. Specific binding to CD19 and CD3 was demonstrated by fluorescence-activated cell sorter analysis. By redirecting unstimulated primary human T cells derived from the peripheral blood against CD19-positive lymphoma cells, the bscCD19 × CD3 antibody showed significant cytotoxic activity at very low concentrations of 10 to 100 pg/mL and at effector to target cell ratios as low as 2:1. Moreover, strong lymphoma-directed cytotoxicity at low antibody concentrations was rapidly induced during 4 hours even in experiments without any T-cell prestimulation. Thus, this particular antibody proves to be much more efficacious than the bispecific antibodies described until now. Therefore, the described bscCD19 × CD3 molecule should be a suitable candidate to prove the therapeutic benefit of bispecific antibodies in the treatment of non-Hodgkin lymphoma.
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This molecule demonstrated efficient cytotoxicity induced by extremely low concentrations (10-50 pg.mL-1, subpicomolar), at low effector:target ratio (2:1), in only four hours, without need for pre-stimulation of T cells or co-signaling.
“This molecule demonstrated outstanding in vitro properties, including efficient cytotoxicity induced by extremely low concentrations of bsAb (10-50 pg.mL -1 , i.e., subpicomolar), at low effector:target ratio (2:1), and in only four hours, without the need for pre-stimulation of T cells or, most surprisingly, the need of co-signaling (e.g., through CD28).”
From Bispecific antibodies for cancer therapy: The light at the end of the tunnel? (2009) · cites this paper as [45]
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- Authors
- Löffler, A., Kufer, P., Lutterbüse, R., Zettl, F., Daniel, P. T., Schwenkenbecher, J. M., Riethmüller, G., Dörken, B., & Bargou, R. C.
- Affiliations
- From the Max Delbrück Center for Molecular Medicine, Berlin-Buch, Germany; Department of Internal Medicine, Robert-Rössle-Klinik, University Medical Center Charité, Humboldt University of Berlin, Berlin, Germany; and Institute of Immunology, University of Munich, Munich, Germany.
- Max Delbrück Center (DE)
- ORCID
- Bargou, Ralf C.
- Venue
- Blood
- Publisher
- American Society of Hematology
- Published
- 2000-03-15
- Volume
- 95
- Issue
- 6
- Pages
- 2098-2103
- ISSN
- 0006-4971 (ISSN-L)
- 1528-0020
- OpenAlex
- W1840766718
- Cited by
- 445
- References
- 44
- Citations by year
- 2026: 12 · 2025: 23 · 2024: 24 · 2023: 18 · 2022: 28 · 2021: 31 · 2020: 28 · 2019: 39 · 2018: 18 · 2017: 27 · 2016: 20 · 2015: 27 · 2014: 16 · 2013: 17 · 2012: 21
- FWCI
- 6.60
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Blood (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Health Sciences › Medicine › Radiology, Nuclear Medicine and Imaging › Monoclonal and Polyclonal Antibodies Research
- Health Sciences › Medicine › Pathology and Forensic Medicine › Lymphoma Diagnosis and Treatment
- Health Sciences › Medicine › Oncology › CAR-T cell therapy research
- Keywords
- bispecific single-chain antibody, CD19, CD3, recombinant antibodies, T cell redirection, B-cell malignancies, non-Hodgkin lymphoma, single-chain Fv fragments
- MeSH terms
- Antibody Specificity, Cell Division, Cytotoxicity, Immunologic, Dose-Response Relationship, Drug, Flow Cytometry, Humans, Immunoglobulin Variable Region, Immunotherapy, Leukocytes, Mononuclear, T-Lymphocytes, Tumor Cells, Cultured, Lymphocyte Subsets, Lymphoma, B-Cell, Cell Death, CD3 Complex, Antibodies, Bispecific, Coculture Techniques, Antigens, CD19
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