title: A human Fab exclusively binding to the extracellular domain of LMP2A.
abstract: In the areas of North Africa, Southeast Asia as well as South China, Nasopharyngeal carcinoma (NPC) is among the most widespread cancers. Plenty of research findings confirmed that Epstein-Barr virus (EBV) played a crucial role in NPC. EBV-encoded Latent membrane protein 2A (LMP2A) which continuously expressed in cell membrane protein induced an epithelial-mesenchymal transition and increased the number of side population stem-like cancer cells in NPC. This reveals that LMP2A could contribute to the development and recurrence in NPC. Above evidences suggest that LMP2A could be the potential target molecule in the treatment of NPC. In the current study, a novel human antibody Fab (Fab29) against the extracellular domain of LMP2A was produced with success. Through immunofluorescence experiment it was proved that human antibody Fab29 exclusively combined the surface of SUNE cells (LMP2A-positive). Then flow cytometry result exhibited that the fluorescent intensities of SUNE cells and CNE cells were distinct (96.89% and 0.02% respectively). After that, it was shown by affinity test that the Fab29 fragment had high affinity (KD (M) 1.79E-09) with LMP2A. It was also revealed by immunohistochemical analysis that the Fab29 fragment could combine with LMP2A-positive human NPC tissues in comparison with the control group. Finally, the MTT result indicated that the Fab29 fragment could inhibit the proliferation of LMP2A-positive NPC cells. The inhibiting rate to SUNE cell proliferation reached a peak by Fab29 (19.67%) compared with unrelated Fab and CNE with Fab29 at a concentration of 500 μg/L in first 24 h and in the next 24 h the inhibition rate grew to 22.54%. In brief, it was shown that Fab29, a characteristic human antibody, could recognize LMP2A protein and inhibit the proliferation of LMP2A-expressing NPC cells in vitro.
authors.affiliation: Department of Otorhinolaryngology, Su Bei People's Hospital of Yangzhou, Yangzhou 225001, China.
authors.forename: Qing
authors.initials: Q
authors.lastname: Cao
authors.affiliation: Department of Otolaryngology, The Second Affiliated Hospital of Nanjing Medical University, Nanjing 210029, China.
authors.forename: Dawei
authors.initials: D
authors.lastname: Zhang
authors.affiliation: Department of Otorhinolaryngology, Jiangsu Province Geriatric Hospital, Nanjing 210024, China.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Mao
authors.affiliation: Yangzhou University, Yangzhou 225009, China.
authors.forename: Chanfang
authors.initials: C
authors.lastname: Meng
authors.affiliation: The Key Laboratory of Antibody Technique of Ministry of Health, Nanjing Medical University, Nanjing 210029, China.
authors.forename: Jin
authors.initials: J
authors.lastname: Zhu
authors.affiliation: The Key Laboratory of Antibody Technique of Ministry of Health, Nanjing Medical University, Nanjing 210029, China.
authors.forename: Zhenqing
authors.initials: Z
authors.lastname: Feng
authors.affiliation: Department of Otolaryngology, The Second Affiliated Hospital of Nanjing Medical University, Nanjing 210029, China. Electronic address: renjiechenent@aliyun.com.
authors.forename: Renjie
authors.initials: R
authors.lastname: Chen
mesh_terms: D001665:Binding Sites; D002478:Cells, Cultured; D045604:Extracellular Fluid; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D000072417:Protein Domains; D014763:Viral Matrix Proteins / Q000737:chemistry* / Q000378:metabolism*
chemical_list: C037662:EBV-associated membrane antigen, Epstein-Barr virus; D007140:Immunoglobulin Fab Fragments; D014763:Viral Matrix Proteins
keywords: Antibody; EBV; LMP2A; NPC