Generation and characterization of avian single chain variable fragment against human Alpha-Enolase

Chi Hsin Lee, Chu Hsuan Tsai, Sy Jye Leu, Ko Jiunn Liu, Wei Chu Wang, Bor Yu Tsai, Liao Chun Chiang, Yan Chiao Mao, Nhlanhla Benedict Dlamini, Chen Hsin Tsai, Yi Yuan Yang

研究成果: 雜誌貢獻文章同行評審

摘要

Overexpression of human alpha-enolase (hEno1) has been reported in a wide range of cancers and is tightly associated with poor prognosis, making it a remarkable biomarker and therapeutic target. In this study, polyclonal yolk-immunoglobulin (IgY) antibodies purified from hEno1-immunized chickens showed a noticeable specific humoral response. Phage display technology was used to construct two antibody libraries of IgY gene-derived single-chain variable fragments (scFvs) containing 7.8 × 107 and 5.4 × 107 transformants, respectively. Phage-based ELISA indicated that specific anti-hEno1 clones were significantly enriched. The nucleotide sequences of scFv-expressing clones were determined and classified into seven groups either in the short linker or the long linker. Moreover, higher mutation rates were revealed in the CDR regions, especially in the CDR3. Three distinguish antigenic epitopes were identified on the hEno1 protein. The binding activities of selected anti-hEno1 scFv on hEno1-positive PE089 lung cancer cells were confirmed using Western blot, flow cytometry, and immunofluorescence assay. In particular, hEnS7 and hEnS8 scFv antibodies significantly suppressed the growth and migration of PE089 cells. Taken together, these chicken-derived anti-hEno1 IgY and scFv antibodies have great potential to develop diagnostic and therapeutic agents for the treatment of lung cancer patients with high expression levels of hEno1 protein.
原文英語
文章編號110277
期刊International Immunopharmacology
120
DOIs
出版狀態已發佈 - 7月 2023

ASJC Scopus subject areas

  • 免疫學和過敏
  • 免疫學
  • 藥理

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