Universität zu Köln

Korenkov, Michael ORCID: 0000-0002-2988-8483 (2025). Somatic hypermutation introduces bystander mutations that prepare SARS-CoV-2 antibodies for emerging variants. PhD thesis, Universität zu Köln.

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Abstract

The adaptive immune response towards pathogens relies on neutralizing antibodies induced by infection or vaccination. With the emergence of SARS-CoV-2 in late 2019, a comprehensive understanding of the human antibody response towards SARS-CoV-2 became essential for developing novel therapeutics and informing vaccination efforts. Studying the role of somatic hypermutation (SHM), especially in the context of antigenically drifted viral escape variants and immune imprinting from repeated viral exposures, is key to advancing these efforts. To address this, we initially investigated the SARS-CoV-2 serum and B cell response using ELISA, neutralization assays, and a high-throughput single-cell sequencing approach screening 4.313 B cells in convalescent individuals. We showed that a SARS-CoV-2 spike protein reactive polyclonal B cell response is readily induced in all 12 participants after infection. From those B cells, we identified 27 potent SARS-CoV-2 neutralizing monoclonal antibodies (mAbs) with a broad spectrum of V gene segments and a low degree of somatic hypermutation (SHM) that can inhibit authentic virus infection at concentrations as low as 0,04 µg/ml. We then analyzed the role of SHM in antibody functionality by reverting a broad panel of previously isolated SARS-CoV-2 neutralizing antibodies (n=92) to their V gene germline variant. Our results illustrate that most SARS-CoV-2 neutralizing antibodies (78 out of 88) depend on their mutations. However, a fraction of mutated SARS-CoV-2 neutralizing antibodies (11 out of 88), including all antibodies from the VH1-58/VK3-20 public clonotype, bound and neutralized independently of acquired mutations. For the VH1-58/VK3-20 public clonotype, only a subset of antibodies (10 out of 22) showed neutralizing activity against Omicron BA.1/BA.2 subvariants despite being isolated before the emergence of Omicron. While mutations were dispensable for Wu01, Alpha, Beta, and Delta variants, they were critical for Omicron BA.1/BA.2 neutralization. Lastly, using pseudovirus neutralization assays, we have examined the impact of emerging viral variants (BA.1, BA.1.1, BA.2, BA.2.12.1, and BA.4/5) on established humoral immunity in 50 samples comprised of vaccinated or convalescent individuals as well as in a 163 monoclonal antibody panel. Our results show that Wu01 mRNA vaccination booster elicits serum neutralization activity against diverse Omicron sublineages, underlining the importance of booster immunization in establishing a sufficient Omicron response. Additionally, the analysis of neutralization sensitivity in the 163-antibody panel revealed notable antigenic differences showcasing distinct escape patterns within the Omicron sublineages with a higher resistance to BA.4/5. Leveraging the knowledge of bystander mutations and antigenic escape, we converted the clinical antibody Tixagevimab (VH1-58/VK3-20), which is ineffective against Omicron BA.1/BA.2, into a potent neutralizer of these viruses. Our findings broaden our understanding of SHM as a mechanism that drives affinity maturation and contributes to antibody diversification through bystander mutations thus increasing the chances to neutralize viral escape variants.

Item Type:	Thesis (PhD thesis)
Creators:	Creators Email ORCID ORCID Put Code Korenkov, Michael mkorenkov52@yahoo.com https://orcid.org/0000-0002-2988-8483 UNSPECIFIED
URN:	urn:nbn:de:hbz:38-798160
Date:	2025
Language:	English
Faculty:	Faculty of Medicine
Divisions:	Faculty of Medicine > Virologie > Institut für Virologie
Subjects:	Medical sciences Medicine
Uncontrolled Keywords:	Keywords Language Somatic hypermutation UNSPECIFIED SARS-CoV-2 UNSPECIFIED Antibody UNSPECIFIED
Date of oral exam:	15 December 2025
Referee:	Name Academic Title Klein, Florian Prof. Dr. med. Lehmann, Clara Prof. Dr. med.
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/79816