Azadirachta indica A. Juss (Neem) Bark Extract Restricts Pan-Coronavirus Infection and Replication

Sarkar, Lucky (2021) Azadirachta indica A. Juss (Neem) Bark Extract Restricts Pan-Coronavirus Infection and Replication. PhD thesis, Indian Institute of Science Education and Research Kolkata.

Text (PhD thesis of Lucky Sarkar (14IP043)) 14IP043_DBS.pdf - Submitted Version Restricted to Repository staff only Download (18MB)

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is one of the most contagious and pathogenic human Coronaviruses and is a constant threat to human health and safety today. SARS-CoV-2 has triggered widespread infection and causing an acute respiratory disease pandemic termed the 'coronavirus disease 2019' (COVID-19). SARS-CoV-2 is related to the severe acute respiratory syndrome coronavirus (SARS-CoV) and Middle Eastern Respiratory syndrome coronavirus (MERS-CoV), which causes acute respiratory disease and have been studied extensively. It is more distantly related to other human Coronaviruses that induce the common cold and Coronaviruses that infect various animal species. A battery of antibodies, repurposed drugs, and anti-convalescent plasma therapy has been examined. Recent advances in diagnosis and vaccine development have been leveraged to manage this pandemic. But understanding Coronaviruses' properties, including their genomic control of pathogenic properties, host cell entry, and cell-to-cell fusion, may guide new therapeutic efforts. Mechanistic studies of SARS-CoV-2 pathogenesis are complex in surviving patients, and experimental animal models of HCoVs are limited in mimicking human disease. But related prototype murine-β-Coronaviruses (m-β-CoVs) facilitate studying conserved mechanisms of COVID pathobiology. MHV-1 infection produces a SARS-CoV-like lethal disease that may explain host immune responses against CoVs. Neurotropic m-CoV-MHV-A59 or its spike protein targeted recombinant strain m-CoV-RSA59 infection into C57BL/6 mice initiates a biphasic disorder starting from acute meningoencephalomyelitis to chronic demyelination. Clinical reports highlight neurological manifestations of SARS-CoV-2; therefore, consequences of SARS-CoV-2 neuro-infection may be inferred from m-CoV-RSA59-induced neuro-COVID. The m-CoV-MHV Spike glycoprotein is a major determinant of viral entry, virus-host interaction, infection-initiation, viral antigen spread, and consecutive pathogenesis. MHV Spike protein can trigger fusion even in the absence of the CEACAM1 receptor, initiating viral entry and subsequent pathogenesis due to immune activation and replacing a single amino acid in the Spike fusion domain alters pathology. One strategy for inhibiting CoV fusion is based on the conformational transition of the six-helix bundle viral fusion core driven by hydrophobic interactions between Heptads repeats HR1 and HR2 of the S2 domain, providing a target for the designing of the mimetic peptide with the potential to disrupt fusion. Alternatively, in this study, we have employed anti-fusogenic properties of bark extract of the ethnomedicinal plant Azadirachta indica A. Juss (Neem bark extract; NBE) both in m-CoV-RSA59 and SARS-CoV-2 infection. Data demonstrated that NBE administered pre-and post-infection inhibits SARS-CoV-2 and m-CoV-RSA59 infection and replication in vitro, with reduced Envelope and Nucleocapsid gene expression. NBE blocks in vitro virus-free cell-to-cell fusion induced by cells expressing the m-CoV-MHV-A59 spike glycoprotein. NBE ameliorates neuroinflammation and hepatitis in vivo by restricting viral replication and spread. Isolated fraction of NBE enriched in Nimbin isomers shows potent inhibition of m-CoV-RSA59 infection in vitro. In silico studies revealed that NBE could target Spike and RdRp of m-CoV and SARS-CoV-2 with high affinity. With no immediate therapies available, the rapid development of novel therapeutic strategies by understanding the pathogenesis of the COVID is of prime importance. Data obtained in this thesis advances our understanding of COVID biology using NBE as a broad-spectrum pan-CoV antiviral therapy.

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