5 research outputs found
XNAzymes targeting the SARS-CoV-2 genome inhibit viral infection
The unprecedented emergence and spread of SARS-CoV-2, the coronavirus responsible for the COVID-19 pandemic, underscores the need for diagnostic and therapeutic technologies that can be rapidly tailored to novel threats. Here, we show that site-specific RNA endonuclease XNAzymes â artificial catalysts composed of single-stranded synthetic xeno-nucleic acid oligonucleotides (in this case 2â-deoxy-2â-fluoro--D-arabino nucleic acid) â may be designed, synthesised and screened within days, enabling the discovery of a range of enzymes targeting SARS-CoV-2 ORF1ab, ORF7b, spike- and nucleocapsid-encoding RNA. Three of these are further engineered to self-assemble into a catalytic nanostructure with enhanced biostability. This XNA nanostructure is capable of cleaving genomic SARS-CoV-2 RNA under physiological conditions, and when transfected into cells inhibits infection with authentic SARS-CoV-2 virus by RNA knockdown. These results demonstrate the potential of XNAzymes to provide a platform for the rapid generation of antiviral reagents.Collaborator Nicholas Matheson is supported by:
MRC (TSF ref. MR/T032413/1)
NHSBT (grant ref. WPA15-02)
The Addenbrookeâs Charitable Trust (grant ref. 900239)
NIHR Cambridge BRC
Induction of HIF-1α by HIV-1 Infection in CD4+ T Cells Promotes Viral Replication and Drives Extracellular Vesicle-Mediated Inflammation
Human immunodeficiency virus type 1 (HIV-1) is a very important global pathogen that preferentially targets CD4+ T cells and causes acquired immunodeficiency syndrome (AIDS) if left untreated. Although antiretroviral treatment efficiently suppresses viremia, markers of immune activation and inflammation remain higher in HIV-1-infected patients than in uninfected individuals. The hypoxia-inducible factor 1α (HIF-1α) is a transcription factor that plays a fundamental role in coordinating cellular metabolism and function. Here we show that HIV-1 infection induces HIF-1α activity and that this transcription factor upholds HIV-1 replication. Moreover, we demonstrate that HIF-1α plays a key role in HIV-1-associated inflammation by promoting the release of extracellular vesicles which, in turn, trigger the secretion of inflammatory mediators by noninfected bystander lymphocytes and macrophages. In summary, we identify that the coordinated actions of HIF-1α and extracellular vesicles promote viral replication and inflammation, thus contributing to HIV-1 pathogenesis.Chronic immune activation and inflammation are hallmarks of HIV-1 infection and a major cause of serious non-AIDS events in HIV-1-infected individuals on antiretroviral treatment (ART). Herein, we show that cytosolic double-stranded DNA (dsDNA) generated in infected CD4+ T cells during the HIV-1 replication cycle promotes the mitochondrial reactive oxygen species (ROS)-dependent stabilization of the transcription factor hypoxia-inducible factor 1α (HIF-1α), which in turn, enhances viral replication. Furthermore, we show that induction of HIF-1α promotes the release of extracellular vesicles (EVs). These EVs foster inflammation by inducing the secretion of gamma interferon by bystander CD4+ T cells and secretion of interleukin 6 (IL-6) and IL-1ÎČ by bystander macrophages through an HIF-1α-dependent pathway. Remarkably, EVs obtained from plasma samples from HIV-1-infected individuals also induced HIF-1α activity and inflammation. Overall, this study demonstrates that HIF-1α plays a crucial role in HIV-1 pathogenesis by promoting viral replication and the release of EVs that orchestrate lymphocyte- and macrophage-mediated inflammatory responses
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Rapid discovery of monoclonal antibodies by microfluidics-enabled FACS of single pathogen-specific antibody-secreting cells
Monoclonal antibodies are increasingly used to prevent and treat viral infections and are pivotal in pandemic response efforts. Antibody-secreting cells (ASCs; plasma cells and plasmablasts) are an excellent source of high-affinity antibodies with therapeutic potential. Current methods to study antigen-specific ASCs either have low throughput, require expensive and labour-intensive screening or are technically demanding and therefore not widely accessible. Here, we present a straightforward technology for the rapid discovery of monoclonal antibodies from ASCs. Our approach combines microfluidic encapsulation of single cells into an antibody capture hydrogel with antigen bait sorting by conventional flow cytometry. With our technology, we screened millions of mouse and human ASCs and obtained anti-SARS-CoV-2 monoclonal antibodies with high affinity (85% of characterised antibodies bound the target). By facilitating access to the underexplored ASC compartment, the approach enables efficient antibody discovery as well as immunological studies into the generation of protective antibodies.This work was supported by the EU Horizon 2020 programme (ERC Advanced Investigator Awards 69566, to F.H. and EU H2020 Marie SkĆodowska-Curie Individual Fellowship (MSCA-IF 750772) to T.S.K), the MRC (grant ref. MC_UU_00025/12, to J.E.D.T and TSF ref. MR/T032413/1, to N.J.M, and a scholarship to K.F.), the Medical Research Foundation (MRF-057-0002-RG-THAV-C0798, to J.E.D.T.), the NIHR Cambridge BRC (to J.E.D.T.), NHSBT (grant ref. WPA15-02, to N.J.M), the Wellcome Trust (ISSF ref. 204845/Z/16/Z, to N.J.M), Addenbrookeâs Charitable Trust (grant ref. 900239, to N.J.M), a scholarship from the Cambridge European Trust (to K.F.), a scholarship from St. Johnâs College Cambridge (to K.F.), a Postdoctoral Research fellowship by Boehringer Ingelheim (to M.I.J.R), an AstraZeneca studentship (to T.N.K), and a Gates Cambridge Scholarship (to G.L.P)
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Longitudinal analysis reveals that delayed bystander CD8+ T cell activation and early immune pathology distinguish severe COVID-19 from mild disease.
The kinetics of the immune changes in COVID-19 across severity groups have not been rigorously
assessed. Using immunophenotyping, RNA sequencing and serum cytokine analysis, we analyzed
serial samples from 207 SARS-CoV2-infected individuals with a range of disease severities over 12
weeks from symptom onset. An early robust bystander CD8+ T cell immune response, without
systemic inflammation, characterized asymptomatic or mild disease. Hospitalized individuals had
delayed bystander responses and systemic inflammation that was already evident near symptom
onset, indicating that immunopathology may be inevitable in some individuals. Viral load did not
correlate with this early pathological response, but did correlate with subsequent disease severity.
Immune recovery is complex, with profound persistent cellular abnormalities in severe disease
correlating with altered inflammatory responses, with signatures associated with increased oxidative
phosphorylation replacing those driven by cytokines tumor necrosis factor (TNF) and interleukin (IL)-
6. These late immunometabolic and immune defects may have clinical implication