Date of Award
Doctor of Philosophy (PhD)
Viruses pose a great threat to human health. Outbreaks and pandemics due to different viruses occur in all areas of the world. Two viruses, which emerged globally in the 1980s and in 2020, that are still causing ongoing pandemics in 2021 are dengue virus (DENV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), respectively. These viruses circulate widely and currently infect hundreds of millions of people. Immunity from a previous infection may confer protection against subsequent infection of a related virus. Thus, understanding the degree of cross-protection can help define correlates of protection that are necessary to determine which people are at greatest risk of infection. DENV is a mosquito-transmitted flavivirus that causes the most common arthropod-borne viral disease in the world. Infection with DENV causes a range of disease manifestations, with the most severe being hemorrhage, vascular leakage leading to organ impairment and circulatory collapse or shock, and consequently death. Historical observations have classified DENV into four serotypes due to lack of cross-immunity and protection between each group. Instead, immunity from one DENV serotype infection can enhance disease upon secondary infection with a different serotype. Therefore, establishing how immunity against one DENV impacts infection of similar and divergent DENV strains is essential to mitigate disease. As more variant DENV strains have been isolated in humans and inducing immunity in this small but expanding population over the last several years, it remains to be determined how this level of viral variation will impact global DENV immunity. In 2007, the most divergent DENV to date was isolated from a Dengue patient in Malaysia. Sequencing results determined it had a level of genetic divergence encroaching on serotype level status, and in 2015, this virus, DKE-121, was unveiled as a novel DENV-5 serotype. Immediately, questions arose of whether immunity from other DENV serotypes or tetravalent vaccines would remain protective or become enhancing in relation to DKE-121 infection. Herein, I examined the relationship of DKE-121 to its closest circulating DENV serotype, DENV-4, for cross-neutralization and cross-protection. My experiments show that DENV-4 immune sera can neutralize DENV-4 and DKE-121 similarly, whereas DKE-121 immune sera preferentially neutralized DKE-121 compared to DENV-4. This directionality in neutralization suggests that DENV-4 and DKE-121 are antigenically related, yet distinct. In passive transfer experiments, neither DENV-4 nor DKE-121 immune sera prevented infection of DKE-121 or DENV-4, respectively, suggesting that humoral immunity alone does not confer protection against the related virus. However, tetravalent vaccination in humans induced antibody responses that neutralized highly variant DKE-121 and DENV-4 vaccination was protective upon DKE-121 challenge in non-human primates. Thus, these findings highlight the complexity of DENV genetic variation and challenge the canonical classification of distinct serotypes. In 2021, in the context of the COVID-19 pandemic, another situation arose with urgent questions of how pre-existing immunity against one virus protects against newly emerging variant viruses. As therapeutics and vaccines were rapidly developed and widely distributed in hopes of ending the COVID-19 pandemic, SARS-CoV-2 variants of concern (VOC) emerged. These VOC contain mutations within the viral spike protein receptor binding domain (RBD), which SARS-CoV-2 uses to enter host cells. Since many vaccines and antibodies were developed using historical spike and/or RBD sequences to interfere with this interaction, it was unknown whether these VOC mutations would enable virus escape. Herein, I assessed therapeutic monoclonal antibodies and vaccine or infection-elicited polyclonal antibodies for cross-neutralization and cross-protection against the SARS-CoV-2 variant viruses. To understand whether monoclonal antibodies (mAbs) targeting historical spike protein would neutralize SARS-CoV-2 VOC, I tested a panel of mAbs against multiple viruses including those with spike protein point mutations corresponding to emerging variants. Most mAbs retained neutralizing potency to these variants compared to historical SARS-CoV-2. However, some mAbs showed reduced or completely abrogated neutralizing activity to viruses containing mutations at position 484 of the spike protein. Remarkably, serum polyclonal antibodies induced against historical spike from vaccination or natural infection showed reduced potency against E484K mutation-containing SARS-CoV-2 variants. This was true for mouse, hamster, and non-human primate sera from adenoviral vectored (ChAd-SARS-CoV-2-S) vaccinated animals in addition to human sera from Pfizer BNT162b2 mRNA vaccinated or previously SARS-CoV-2 infected individuals. Thus, SARS-CoV-2 variants containing a mutation at position 484 (e.g., B.1.351 (Beta) and B.1.1.28 (or P.1, Gamma) lineages, are of particular concern for antibody-mediated protection. As some therapeutic mAbs were given Emergency Use Authorization (EUA) or under development in advanced clinical trial stages, it was essential to understand whether they retain in vivo efficacy against emerging SARS-CoV-2 variants. These mAbs included monotherapy from Lilly under EUA (at the time) and combination mAb therapy from AstraZeneca, AbbVie, and Vir Biotechnology under development and from Regeneron and Lilly under EUA. When tested for neutralization, most mAbs retained potency against VOC including those from the B.1.351 and B.1.1.28 lineages. However, mAbs that had contact residues at position 484 (those from Lilly and AbbVie) showed reduced potency or complete loss of neutralizing activity against B.1.351 and B.1.1.28. When tested for therapeutic efficacy in K18-hACE2 mice, monotherapy and combination therapy with the Lilly mAbs failed to confer protection. The other combination therapy (from Regeneron) approved for EUA use, in addition to two others (AstraZeneca and Vir Biotechnology) were highly protective. While one mAb completely lost neutralizing activity in AbbVie’s cocktail against Beta and Gamma, the other retained activity and conferred protection against weight loss in mice. Thus, neutralizing sensitivity correlated with in vivo therapeutic efficacy, which has utility in screening mAbs against any emerging SARS-CoV-2 variants. Herein, I describe two viruses that have emerged globally within the last few decades or year and how pre-existing immunity may influence subsequent infection. For DENV, these interactions are complex and can be protective or enhancing between different viruses within this family. As more divergent DENV strains are found, the canonical serotype definitions become blurred, with unclear impacts on infection and immunization outcome. For SARS-CoV-2, many therapeutics and vaccines were generated using 2019 virus spike sequences and may be susceptible to escape from emerging variants. However, combination use of multiple therapeutic mAbs targeting different areas on the spike protein may help prevent additional viral escape. In addition, rapid development of COVID-19 vaccines has provided a framework, which can be quickly adapted to create boosters against the variants. Hence, the studies I describe here expand our understanding of the cross-protective immunity that exists against DENV and SARS-CoV-2 and with this knowledge, we are better equipped to end these pandemics and hopefully prevent future ones.
Chair and Committee
Michael S. Diamond
Robyn S. Klein, Adrianus C. Boon, Ali H. Ellebedy, Daved H. Fremont,
Chen, Rita Elaine, "Assessment of Cross-protective Responses to Emerging Variant Viruses" (2023). Arts & Sciences Electronic Theses and Dissertations. 2839.