Anti-vascular endothelial growth factor receptor (VEGFR) 2 autoantibody identification in glioblastoma patient using single B cell-based antibody gene cloning.

Abstract

Antibody direct cloning from single B cells is simple and efficient and has been successful in antibody identification of infectious diseases. However, although a recent whole-exome sequencing revealed abundant heterogeneic mutation accumulation in cancers, identification and synthesis of autoantibodies against specific cancer-associated antigens is still difficult in cancer patients owing to the very small number of B cells producing autoantibodies. In the present study, to identify autoantibodies targeting tumor antigens, we measured the titer of autoantibodies in high-grade glioma patients' plasma and identified two patients with elevated autoantibodies to a few transmembrane proteins. Specific B cells producing autoantibody against vascular endothelial growth factor receptor (VEGFR) 2 were immunostained with labeled protein and anti-human IgG antibody, and then collected by a single cell sorter. Finally, 22 antibody genes were successfully identified using direct IgG cloning from single B cell mRNA, and two antibody clones were found to have significant VEGFR2-specific binding affinity. The current direct human IgG gene cloning technique for identifying human antibodies derived from IgG-memory B cells avoids time-consuming procedures such as phage display-based antibody-library screening, and therefore may be applicable to identifying human autoantibodies in a variety of disorders including cancers even when antibody elevation is not detected because of a very small number of memory B cells.

• Authors: Akira Iizuka, Masaru Komiyama, Chie Oshita, Akiko Kume, Tadashi Ashizawa, Koichi Mitsuya, Nakamasa Hayashi, Yoko Nakasu, Ken Yamaguchi, Yasuto Akiyama
• Journal: Immunology letters (Immunol Lett) 2014 May-Jun Vol. 159 Issue 1-2 Pg. 15-22 ISSN: 1879-0542 [Electronic] Netherlands
• PMID: 24534640 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2014 Elsevier B.V. All rights reserved.

Chemical References

• Antigens, Neoplasm
• Autoantibodies
• Immunoglobulin G
• RNA, Messenger
• KDR protein, human
• Vascular Endothelial Growth Factor Receptor-2

Topics

• Amino Acid Sequence
• Antibody Specificity
• Antigens, Neoplasm (genetics, immunology)
• Autoantibodies (biosynthesis, genetics)
• B-Lymphocytes (immunology, pathology)
• Brain Neoplasms (genetics, immunology, pathology, therapy)
• Cloning, Molecular (methods)
• Gene Expression
• Gene Library
• Glioblastoma (genetics, immunology, pathology, therapy)
• Humans
• Immunoglobulin G (genetics, immunology)
• Immunologic Memory
• Immunotherapy (methods)
• Molecular Sequence Data
• RNA, Messenger (genetics, immunology)
• Single-Cell Analysis
• Vascular Endothelial Growth Factor Receptor-2 (antagonists & inhibitors, genetics, immunology)