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Updated: May 24, 2026
Generation of Discriminative Human Monoclonal Antibodies from Rare Antigen-specific B Cells Circulating in Blood
Published on: February 6, 2018
Generation of human single-chain antibody to the CD99 cell surface determinant specifically recognizing Ewing's
Mara Gellini, Alessandro Ascione, Michela Flego
1Department of Drug Research and Evaluation, Istituto Superiore di Sanita, Viale Regina Elena 299 00161 Rome, Italy. mara.gellini@iss.it.
Abstract:
The survival of pediatric patients with cancer entities including osteosarcoma and Ewing's sarcoma (ES), remains extremely low hence novel treatment approaches are urgently needed. Therefore, based on the concept of targeted therapy, numerous potential targets for the treatment of these cancers have been evaluated pre-clinically or in some cases even clinically during the last decade. In ES the CD99 protein is an attractive target antigen. In this respect, a new entry site for therapeutic intervention may derive from specific human antibodies against CD99. Human scFvC7 was isolated from a semi-synthetic ETH-2 antibody phage library panned on the extracellular portion of recombinant human CD99 protein. The scFvC7 was genetically sequenced, tested for CD99 recognition on an array of recombinant CD99 fragments and measured for binding affinity by ELISA. Finally, it was tested for staining CD99 antigen on a large panel of tumor and normal cells and tissues by cytofluorimetric and immunohistochemical assays. The new antibody scFvC7 recognizes the CD99 extracellular domain included between residues 50 and 74 with a binding affinity of 2.4 x 10(-8) M. In contrast with all other antibodies to CD99 so far isolated, scFvC7 shows a unique specificity in cancer cell recognition: It stained prevalently ES cells while no or weak reactivity was observed on the majority of the other tumor and normal cells and tissues. Thanks to its properties the new anti-CD99 antibody here described represents the first step towards the construction of new selective ES therapeutics.
Insights
A novel human antibody, scFvC7, specifically targets the CD99 protein on Ewing sarcoma (ES) cells. This discovery offers a promising new avenue for developing targeted therapies for this rare pediatric cancer.
Area of Science:
- Oncology
- Immunology
- Biotechnology
Background:
- Pediatric cancers like Ewing sarcoma (ES) have poor survival rates, necessitating novel treatments.
- Targeted therapy approaches are being explored, with CD99 protein identified as a key antigen in ES.
Purpose of the Study:
- To isolate and characterize a novel human antibody targeting the CD99 protein for potential therapeutic applications in ES.
Main Methods:
- Isolation of human scFvC7 antibody from a phage library.
- Genetic sequencing and binding affinity measurements (ELISA) for CD99 recognition.
- Evaluation of antibody specificity on various cancer and normal cells/tissues using flow cytometry and immunohistochemistry.
Main Results:
- The scFvC7 antibody specifically recognizes the extracellular domain of CD99 (residues 50-74) with a binding affinity of 2.4 x 10(-8) M.
- scFvC7 demonstrated unique specificity, predominantly staining ES cells while showing minimal reactivity with other tumor and normal cells.
- This selective binding profile distinguishes scFvC7 from previously isolated anti-CD99 antibodies.
Conclusions:
- The novel anti-CD99 antibody scFvC7 exhibits high specificity for Ewing sarcoma cells.
- scFvC7 represents a significant advancement and the foundational step for developing new, selective therapeutic agents for ES.
