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Construction of small molecular CTLA4 analogs with CD80-binding affinity

Author:
Liu, Fang, Su, LiLi, Chen, Ziteng, Feng, Deming, Wei, Jing, Sun, Jian
Source:
Biochemical and biophysical research communications 2019 v.513 no.3 pp. 694-700
ISSN:
0006-291X
Subject:
binding capacity, binding sites, fibronectins, half life, humans, immunoglobulin G, monoclonal antibodies, peptides, recombinant proteins
Abstract:
A variety of CTLA4-Fc fusion proteins and anti-CTLA4 monoclonal antibody have been approved. Given the shortcomings of macromolecular antibodies, recombinant proteins derived from the tenth unit of human type III fibronectin (FN3) termed monobody were studied as CTLA4 analogs in this study. A peptide EL161 derived from CD80-binding domain (MYPPPY motifs) in the complementarity determining region (CDR) 3 of CTLA4 was found to inhibit the interaction of CTLA4 with CD80 significantly. Afterward, the peptide EL16 as well as the CDR1 of CTLA4 which is also critical for its binding to CD80 were grafted onto FN3 and obtained a novel CD80 binding monobody protein CFN13.2 CFN13 showed 80% binding affinity compared to CTLA4. In addition, to increase the half-life, CFN13 was fused to human IgG1 Fc to generate CFN13-Fc fusion protein. As expected, CFN13-Fc bound to CD80 in a dosage-dependent manner as CFN13 did, and displayed 41.0% and 31.4% inhibition on the interaction of CTLA4-Fc with CD80 at 200 μg/ml and 100 μg/ml respectively. Moreover, peptide EL16 could inhibit CFN13-Fc binding to CD80 significantly, with the inhibition ratio of 64.3% and 52.8% at 100 and 50 μg/ml respectively, indicating that the peptide EL16 and CFN13-Fc shared the similar binding sites with CD80 and the CDR3 motif of CTLA4 contributed more than CDR1 in binding to CD80. In summary, our study provides insights into small molecular analogs of CTLA4.
Agid:
6365687