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The impact of the Covid-19 pandemic called for rapid responses in face of unprecedented challenges. In this context, earning more about the causative agent SARS-CoV-2 becomes imperative. Therefore, clinical virus isolates were studied with focus on infectivity, replication kinetic, and caspase activity.
Firstly, clinical specimens collected from patients were tested for infectivity in cell culture. Combined with polymerase chain reaction results, a formula predicting infectivity in cell culture based on abundance of viral RNA was developed. Additionally, analysis of different specimen types, sources, and material, elucidate the question of infectivity. Here, infectivity was demonstrated in specimens derived from different parts of the respiratory tract, including specimens collected from deceased persons. A protocol for virus isolation on human airway epithelium in air-liquid interface culture was established.
Secondly, replication kinetics of 20 clinical isolates were compared, including a subset of seven sequenced isolates. All isolates replicated in the colon epithelial cell culture model. Within the subset, differences between isolates carrying the D614G amino acid exchange and with original spike protein were observed.
Lastly, elevated caspase activity was demonstrated in two cell culture models including human airway epithelium in air-liquid interface culture.
Subsequently, caspase inhibition by small-molecule compound Emricasan and its effects on the cytopathic effect observed in cell culture were studied. Here, increased cell survival in a colon epithelial cell line was shown with unimpaired virus replication. Elevated caspase activity was identified as early marker of infection and validated by testing across 20 clinical virus isolates.
This study offers information on infectivity that can help shape the understanding of transmission risk. As such, parts of the data collected here were used for validation of rapid antigen tests. The insights gained by studying caspase activity contributed in part to the development of a drug screening method by Bojkova et al.,41 thus aiding routine laboratory workflow. It was demonstrated that Emricasan exhibits no antiviral effect, while the finding of increased cell survival in cell culture could give rise to further research on prevention of tissue damage.
The ongoing COVID-19 pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is partly under control by vaccination. However, highly potent and safe antiviral drugs for SARS-CoV-2 are still needed to avoid development of severe COVID-19. We report the discovery of a small molecule, Z-Tyr-Ala-CHN2, which was identified in a cell-based antiviral screen. The molecule exerts sub-micromolar antiviral activity against SARS-CoV-2, SARS-CoV-1, and human coronavirus 229E. Time-of-addition studies reveal that Z-Tyr-Ala-CHN2 acts at the early phase of the infection cycle, which is in line with the observation that the molecule inhibits cathepsin L. This results in antiviral activity against SARS-CoV-2 in VeroE6, A549-hACE2, and HeLa-hACE2 cells, but not in Caco-2 cells or primary human nasal epithelial cells since the latter two cell types also permit entry via transmembrane protease serine subtype 2 (TMPRSS2). Given their cell-specific activity, cathepsin L inhibitors still need to prove their value in the clinic; nevertheless, the activity profile of Z-Tyr-Ala-CHN2 makes it an interesting tool compound for studying the biology of coronavirus entry and replication.
The ancestral SARS-CoV-2 strain that initiated the Covid-19 pandemic at the end of 2019 has rapidly mutated into multiple variants of concern with variable pathogenicity and increasing immune escape strategies. However, differences in host cellular antiviral responses upon infection with SARS-CoV-2 variants remain elusive. Leveraging whole-cell proteomics, we determined host signaling pathways that are differentially modulated upon infection with the clinical isolates of the ancestral SARS-CoV-2 B.1 and the variants of concern Delta and Omicron BA.1. Our findings illustrate alterations in the global host proteome landscape upon infection with SARS-CoV-2 variants and the resulting host immune responses. Additionally, viral proteome kinetics reveal declining levels of viral protein expression during Omicron BA.1 infection when compared to ancestral B.1 and Delta variants, consistent with its reduced replication rates. Moreover, molecular assays reveal deferral activation of specific host antiviral signaling upon Omicron BA.1 and BA.2 infections. Our study provides an overview of host proteome profile of multiple SARS-CoV-2 variants and brings forth a better understanding of the instigation of key immune signaling pathways causative for the differential pathogenicity of SARS-CoV-2 variants.
In this comprehensive review, we will dissect the impact of research on proteoglycans focusing on recent developments involved in their synthesis, degradation, and interactions, while critically assessing their usefulness in various biological processes. The emerging roles of proteoglycans in global infections, specifically the SARS-CoV-2 pandemic, and their rising functions in regenerative medicine and biomaterial science have significantly affected our current view of proteoglycans and related compounds. The roles of proteoglycans in cancer biology and their potential use as a next-generation protein-based adjuvant therapy to combat cancer is also emerging as a constructive and potentially beneficial therapeutic strategy. We will discuss the role of proteoglycans in selected and emerging areas of proteoglycan science, such as neurodegenerative diseases, autophagy, angiogenesis, cancer, infections and their impact on mammalian diseases.