Advancing Pandemic Preparedness by Developing Antivirals for Human Coronaviruses and Mitigating Viral Infections through Inactivation and Neutralization
Loading...
Authors
Adhikari, Kabita
Issue Date
2025
Type
Dissertation
Language
en_US
Keywords
Antivirals , Coronaviruses , Inactivation , Neutralization , RdRp , SARS-CoV-2
Alternative Title
Abstract
The COVID-19 pandemic, driven by the highly transmissible SARS-CoV-2, remains a primary global health concern, supporting the critical need for effective strategies to combat the virus. This study addresses three critical areas: developing antiviral drugs effective in controlling virus replication, decontaminating the viruses from PPE for sustainability, and evaluating vaccine efficacy against emerging variants. The existing antiviral treatments primarily target the main protease; developing drugs against additional viral targets, such as the RdRp to inhibit the viral transcription and replication, could be an attractive for broad-spectrum antiviral drug development against SARS-CoV-2 and other human coronaviruses. In this study, identified the broad-spectrum antiviral activity of small molecules through a cell-based assay to determine the enzymatic activity of SARS-CoV-2 RdRp through a luciferase reporter assay. Among the tested small molecules, TK018, a potent inhibitor of SARS-CoV-2 RdRp with minimal cytotoxicity. TK018 demonstrated broad-spectrum antiviral activity against SARS-CoV-2 and other HCoVs such as HCoV-OC43, and HCoV-229E positioning as a promising candidate for therapeutic and prophylactic use. The inhibitory effect of TK018 was further evaluated on SARS-CoV-2 replication in cell-based infection assay by determining the levels of replicated and cell-free infectious virions following treatment of infected cells. The broad-spectrum antiviral activity of CADA compounds was evaluated using cell-based infection assays. Cells infected with HCoVs were treated with the compounds, and the levels of virus production were measured post-treatment. Using HCoV-229E (an alpha-coronavirus) and HCoV-OC43 (a beta-coronavirus) as representative strains, we found that TK018 effectively inhibited the replication of both viruses, confirming its potential as a broad-spectrum antiviral agent.
Developing effective, scalable, and sustainable decontamination methods is essential for safely removing viral pathogens from PPE, enabling reuse, and reducing exposure to infectious agents. O₃ has emerged as a promising, eco-friendly disinfectant due to its strong oxidative properties, fast action, and residue-free breakdown into oxygen. We evaluated a Trinion Disinfector (FATHHOME Inc.), an ozone-based sterilization device, for sustainable PPE decontamination. In this study, we investigated the effectiveness of ozone in inactivating viral pathogens. After treating with 60 ppm of ozone for 10 minutes, the Trinion Disinfector significantly reduced SARS-CoV-2, AAV, HSV-1, and HBV on PPE surfaces. Contamination from SARS-CoV-2 on face shields and N95 masks was decreased by 99.9%, while AAV infectivity was nearly eliminated. A similar level of effectiveness was observed against HSV-1 and HBV, with substantial reductions in infectious virus particles following ozone exposure.
Finally, we evaluated the neutralizing efficacy of various COVID-19 vaccines by testing pooled sera from vaccinated individuals against a comprehensive panel of SARS-CoV-2 variants and subvariants, including the ancestral strain WA1, Delta, and multiple Omicron lineages, using the gold-standard PRNT. The rapid emergence of SARS-CoV-2 variants, particularly those within the Omicron lineage, poses a significant challenge due to their extensive mutations in the spike protein, especially within the RBD. These mutations have been linked to increased transmissibility and enhanced immune escape, diminishing the effectiveness of vaccine-elicited neutralizing antibodies.
In this study, we assessed serum samples collected after both primary vaccinations and booster doses across multiple platforms. Our findings revealed that the neutralization capacity was highest against WA1 and Delta, consistent with the antigenic match to the original vaccine formulations. However, the neutralization potency declined markedly against Omicron subvariants, particularly in samples collected after single-dose regimens or heterologous booster strategies, suggesting limited cross-protective immunity.
Conversely, sera from individuals who received homologous mRNA booster doses (Moderna or Pfizer) demonstrated broader and more potent neutralizing responses against several Omicron subvariants, including BA.1, BA.2.12.1, and BA.5. While neutralization against the most recent subvariants (e.g., XBB.1.5 and JN.1) remained limited, homologous boosters still provided improved cross-variant protection compared to heterologous approaches.
These results emphasize the importance of booster vaccinations in maintaining protective immunity and highlight the relative advantage of homologous mRNA booster regimens in extending the breadth of neutralization. Furthermore, they reinforce the need for continuous surveillance of emerging variants and the adaptation of vaccine formulations to ensure effectiveness against evolving SARS-CoV-2 lineages.
In summary, the findings highlight the potential of TK018 as a lead compound for developing therapeutic and prophylactic strategies against SARS-CoV-2 and potentially future coronaviruses, offering a valuable tool to enhance pandemic preparedness and improve treatment outcomes. Additionally, optimizing ozone concentration and exposure time was shown to significantly enhance the pathogen-inactivation capability of the FATHHOME decontamination device, supporting its potential role in infection control. Finally, evaluating vaccine efficacy remains critical for understanding the breadth and durability of immune protection, informing the design of next-generation vaccines, and equipping global health systems to respond more effectively to current and future viral threats.