Isolation of an escape-resistant SARS-CoV-2 neutralizing nanobody from a novel synthetic nanobody library.
| Autor: | Dormeshkin D; Laboratory of Molecular Diagnostics and Biotechnology, Institute of Bioorganic Chemistry of the National academy of Sciences of Belarus, Minsk, Belarus., Shapira M; Laboratory of Molecular Diagnostics and Biotechnology, Institute of Bioorganic Chemistry of the National academy of Sciences of Belarus, Minsk, Belarus., Dubovik S; Department of Biology, Belarusian State University, Minsk, Belarus., Kavaleuski A; Institute of Science and Technology Austria (ISTA), Klosterneuburg, Austria., Katsin M; Imunovakcina, UAB, Vilnius, Lithuania.; Immunofusion, LLC, Minsk, Belarus., Migas A; Imunovakcina, UAB, Vilnius, Lithuania.; Immunofusion, LLC, Minsk, Belarus., Meleshko A; Imunovakcina, UAB, Vilnius, Lithuania.; Immunofusion, LLC, Minsk, Belarus., Semyonov S; Laboratory of Biosafety With Pathogens Collection, Republican Research and Practical Center for Epidemiology & Microbiology, Minsk, Belarus. |
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| Jazyk: | angličtina |
| Zdroj: | Frontiers in immunology [Front Immunol] 2022 Sep 16; Vol. 13, pp. 965446. Date of Electronic Publication: 2022 Sep 16 (Print Publication: 2022). |
| DOI: | 10.3389/fimmu.2022.965446 |
| Abstrakt: | The COVID-19 pandemic not only resulted in a global crisis, but also accelerated vaccine development and antibody discovery. Herein we report a synthetic humanized VHH library development pipeline for nanomolar-range affinity VHH binders to SARS-CoV-2 variants of concern (VoC) receptor binding domains (RBD) isolation. Trinucleotide-based randomization of CDRs by Kunkel mutagenesis with the subsequent rolling-cycle amplification resulted in more than 10 11 diverse phage display library in a manageable for a single person number of electroporation reactions. We identified a number of nanomolar-range affinity VHH binders to SARS-CoV-2 variants of concern (VoC) receptor binding domains (RBD) by screening a novel synthetic humanized antibody library. In order to explore the most robust and fast method for affinity improvement, we performed affinity maturation by CDR1 and CDR2 shuffling and avidity engineering by multivalent trimeric VHH fusion protein construction. As a result, H7-Fc and G12x3-Fc binders were developed with the affinities in nM and pM range respectively. Importantly, these affinities are weakly influenced by most of SARS-CoV-2 VoC mutations and they retain moderate binding to BA.4\5. The plaque reduction neutralization test (PRNT) resulted in IC50 = 100 ng\ml and 9.6 ng\ml for H7-Fc and G12x3-Fc antibodies, respectively, for the emerging Omicron BA.1 variant. Therefore, these VHH could expand the present landscape of SARS-CoV-2 neutralization binders with the therapeutic potential for present and future SARS-CoV-2 variants. Competing Interests: Authors MK, AMi and AMe are employees of UAB Imunovakcina and LLC Immunofusion. DD is a member of the Scientific Advisory Board of Imunovakcina. DD and MS have a pending patent application for the RBD-targeted antibodies from this study. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be constructed as a potential conflict of interest. (Copyright © 2022 Dormeshkin, Shapira, Dubovik, Kavaleuski, Katsin, Migas, Meleshko and Semyonov.) |
| Databáze: | MEDLINE |
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