Date of Award
2024
Document Type
Dissertation
Degree Name
Doctor of Philosophy (PhD)
Department
Cellular, Molecular and Biomedical Sciences
First Advisor
Jason Botte
Abstract
Monoclonal antibodies (mAbs) are an integral component of modern medicine, proving invaluable for the treatment of cancers, autoimmune disorders, and infectious diseases, including SARS-CoV-2 infections, the causative agent of COVID-19. Notably, virus-specific mAbs were pivotal for both prophylaxis and therapeutic treatment of individuals at high risk for severe COVID-19 during the early stages of the pandemic. However, emerging SARS-CoV-2 variants exhibited rapid evolution in the spike (S) protein, notably the receptor binding domain (RBD), compromising the efficacy of these antiviral mAbs, necessitating withdrawal from clinical use. Previously utilized therapeutic mAbs were derived from antibody-producing cells that were identified using antigen bait approaches. While effective in quickly identifying potent mAbs, this approach restricted discovery to mAbs that recognized regions of the SARS-CoV-2 S protein with evolutionarily volatile epitope sites.
To fully explore the broad spectrum of SARS-CoV-2-specific mAbs produced during infection and identify novel therapeutic candidates, we adopted an agnostic approach to comprehensively define the acute humoral immune repertoire in COVID-19 patients. In total, we sequenced over 11,000 natively paired antibody heavy and light chains, synthesizing 229 sequences into recombinant IgG1 mAbs. Among this preliminary subset of mAbs, 17 were capable of neutralizing SARS-CoV-2, and targeted regions in the S protein and beyond. Significantly, a subset of these mAbs broadly neutralized numerous viral variants of concern and protected mice from lethal infection with SARS-CoV-2. Ultimately, our agnostic approach allowed for the successful capture of novel mAbs, delineating an antibody repertoire that is a valuable resource for applications in cellular virology research, diagnostics, and most importantly, the development of effective mAb therapeutics in the face of the continuously evolving SARS-CoV-2.
In a separate, but complementary arm of this dissertation, we aimed to enhance candidate SARS-CoV-2-specific IgG mAbs by transforming them into multivalent structures. Using 38 candidate IgG1 sequences – including several mAbs identified in our agnostic studies – we generated dimeric IgA1 and IgA2, and pentameric IgM derivatives. Strikingly, when compared to the parental IgG mAbs, the multimerized mAbs, particularly the IgMs, consistently demonstrated massively improved binding strength, virus neutralization potency, and capacity to cross-neutralize SARS-CoV-2 variants. Importantly, selected IgM candidates protected mice from lethal SARS-CoV-2 infection in vivo. Our results support multimerization of IgG molecules as a promising strategy to create superior and enduring mAb therapeutics for SARS-CoV-2 and other potential viral pathogens.
In summary, the research presented in this dissertation yielded multiple potential therapeutic mAbs for the treatment of COVID-19 and novel methodologies to develop superior antiviral mAb therapeutics.
Language
en
Number of Pages
382 p.
Recommended Citation
Whitaker, Annalis Norman, "Expanding The Spectrum Of Covid-19 Monoclonal Antibody Therapeutics: Agnostic Exploration & Multivalent Transformations Of Sars-Cov-2 Antibody Repertoires" (2024). Graduate College Dissertations and Theses. 1907.
https://scholarworks.uvm.edu/graddis/1907