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Selection of novel affinity-matured human chondroitin sulfate proteoglycan 4 antibody fragments by yeast display.

Publication ,  Journal Article
Yu, X; Qu, L; Bigner, DD; Chandramohan, V
Published in: Protein Eng Des Sel
September 1, 2017

Chondroitin sulfate proteoglycan 4 (CSPG4) is a promising target for cancer immunotherapy due to its high level of expression in a number of malignant tumors, and its essential role in tumor growth and progression. Clinical application of CSPG4-targeting immunotherapies is hampered by the lack of fully human high-affinity CSPG4 antibodies or antibody fragments. To overcome this limitation, we performed affinity maturation on a novel human CSPG4 single-chain Fv fragment (scFv) using the random mutagenesis approach and screened for improved variants from a yeast display library using a modified whole-cell panning method followed by fluorescence-activated cell sorting. After six rounds of panning and sorting, the top seven mutant scFvs were isolated and their binding affinities were characterized by flow cytometry and surface plasmon resonance. These highly specific, affinity-matured variants displayed nanomolar to picomolar binding affinities to the CSPG4 antigen. While each of the mutants harbored only two to six amino acid substitutions, they represented ~270-3000-fold improvement in affinity compared to the parental clone. Our study has generated affinity-matured scFvs for the development of antibody-based clinical therapeutics targeting CSPG4-expressing tumors.

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Published In

Protein Eng Des Sel

DOI

EISSN

1741-0134

Publication Date

September 1, 2017

Volume

30

Issue

9

Start / End Page

639 / 647

Location

England

Related Subject Headings

  • Single-Chain Antibodies
  • Saccharomyces cerevisiae
  • Recombinant Proteins
  • Protein Binding
  • Mutation
  • Membrane Proteins
  • Kinetics
  • Humans
  • HEK293 Cells
  • Gene Expression
 

Citation

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Yu, X., Qu, L., Bigner, D. D., & Chandramohan, V. (2017). Selection of novel affinity-matured human chondroitin sulfate proteoglycan 4 antibody fragments by yeast display. Protein Eng Des Sel, 30(9), 639–647. https://doi.org/10.1093/protein/gzx038
Yu, Xin, Liang Qu, Darell D. Bigner, and Vidyalakshmi Chandramohan. “Selection of novel affinity-matured human chondroitin sulfate proteoglycan 4 antibody fragments by yeast display.Protein Eng Des Sel 30, no. 9 (September 1, 2017): 639–47. https://doi.org/10.1093/protein/gzx038.
Yu X, Qu L, Bigner DD, Chandramohan V. Selection of novel affinity-matured human chondroitin sulfate proteoglycan 4 antibody fragments by yeast display. Protein Eng Des Sel. 2017 Sep 1;30(9):639–47.
Yu, Xin, et al. “Selection of novel affinity-matured human chondroitin sulfate proteoglycan 4 antibody fragments by yeast display.Protein Eng Des Sel, vol. 30, no. 9, Sept. 2017, pp. 639–47. Pubmed, doi:10.1093/protein/gzx038.
Yu X, Qu L, Bigner DD, Chandramohan V. Selection of novel affinity-matured human chondroitin sulfate proteoglycan 4 antibody fragments by yeast display. Protein Eng Des Sel. 2017 Sep 1;30(9):639–647.
Journal cover image

Published In

Protein Eng Des Sel

DOI

EISSN

1741-0134

Publication Date

September 1, 2017

Volume

30

Issue

9

Start / End Page

639 / 647

Location

England

Related Subject Headings

  • Single-Chain Antibodies
  • Saccharomyces cerevisiae
  • Recombinant Proteins
  • Protein Binding
  • Mutation
  • Membrane Proteins
  • Kinetics
  • Humans
  • HEK293 Cells
  • Gene Expression