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Determining divergent metabolic requirements of T cells, and the viruses and tumours they fail to combat, could provide new therapeutic checkpoints. Inhibition of acyl-CoA:cholesterol acyltransferase (ACAT) has direct anti-carcinogenic activity. Here, we show that ACAT inhibition has antiviral activity against hepatitis B (HBV), as well as boosting protective anti-HBV and anti-hepatocellular carcinoma (HCC) T cells. ACAT inhibition reduces CD8 +  T cell neutral lipid droplets and promotes lipid microdomains, enhancing TCR signalling and TCR-independent bioenergetics. Dysfunctional HBV- and HCC-specific T cells are rescued by ACAT inhibitors directly ex vivo from human liver and tumour tissue respectively, including tissue-resident responses. ACAT inhibition enhances in vitro responsiveness of HBV-specific CD8 + T cells to PD-1 blockade and increases the functional avidity of TCR-gene-modified T cells. Finally, ACAT regulates HBV particle genesis in vitro, with inhibitors reducing both virions and subviral particles. Thus, ACAT inhibition provides a paradigm of a metabolic checkpoint able to constrain tumours and viruses but rescue exhausted T cells, rendering it an attractive therapeutic target for the functional cure of HBV and HBV-related HCC. Shared metabolic pathways could allow simultaneous manipulation of T cells, viruses and tumours. Here the authors show targeting cholesterol esterification restrains hepatitis B in vitro, whilst bolstering exhausted antigen-specific T cell responses from human liver and hepatocellular carcinoma.

Memory B cells (MBCs) can provide a recall response able to supplement waning antibodies (Abs) with an affinity-matured response better able to neutralize variant viruses. We studied a cohort of elderly care home residents and younger staff (median age of 87 years and 56 years, respectively), who had survived COVID-19 outbreaks with only mild or asymptomatic infection. The cohort was selected because of its high proportion of individuals who had lost neutralizing antibodies (nAbs), thus allowing us to specifically investigate the reserve immunity from SARS-CoV-2-specific MBCs in this setting. Class-switched spike and receptor-binding domain (RBD) tetramer-binding MBCs persisted 5 months after mild or asymptomatic SARS-CoV-2 infection, irrespective of age. The majority of spike- and RBD-specific MBCs had a classical phenotype, but we found that activated MBCs, indicating possible ongoing antigenic stimulation or inflammation, were expanded in the elderly group. Spike- and RBD-specific MBCs remained detectable in the majority of individuals who had lost nAbs, although at lower frequencies and with a reduced IgG/IgA isotype ratio. Functional spike-, S1 subunit of the spike protein- (S1-), and RBD-specific recall was also detectable by enzyme-linked immune absorbent spot (ELISPOT) assay in some individuals who had lost nAbs, but was significantly impaired in the elderly. Our findings demonstrate that a reserve of SARS-CoV-2-specific MBCs persists beyond the loss of nAbs but highlight the need for careful monitoring of functional defects in spike- and RBD-specific B cell immunity in the elderly.

IntroductionFollowing the emergence of SARS-CoV-2 in 2020, care homes were disproportionately impacted by high mortality and morbidity of vulnerable elderly residents. Non-pharmaceutical interventions (NPIs) and improved infection control measures together with vaccination campaigns have since improved outcomes of infection. We studied the utility of past infection status, recent vaccination and anti-S antibody titres as possible correlates of protection against a newly emergent Omicron variant infection.MethodsProspective longitudinal surveillance of nine sentinel London care homes from April 2020 onwards found that all experienced COVID-19 outbreaks due to Omicron (BA.1) during December 2021 and January 2022, despite extensive prior SARS-CoV-2 exposure and high COVID-19 vaccination rates, including booster vaccines (>70% residents, >40% staff).ResultsDetailed investigation showed that 46% (133/288) of Omicron BA.1 infections were SARS-CoV-2 reinfections. Two and three COVID-19 vaccine doses were protective against Omicron infection within 2-9 weeks of vaccination, though protection waned from 10 weeks post-vaccination. Prior infection provided additional protection in vaccinated individuals, approximately halving the risk of SARS-CoV-2 infection.DiscussionAnti-S antibody titre showed a dose-dependent protective effect but did not fully account for the protection provided by vaccination or past infection, indicating that other mechanisms of protection are also involved.

Individuals with potential exposure to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) do not necessarily develop PCR or antibody positivity, suggesting that some individuals may clear subclinical infection before seroconversion. T cells can contribute to the rapid clearance of SARS-CoV-2 and other coronavirus infections 1 – 3 . Here we hypothesize that pre-existing memory T cell responses, with cross-protective potential against SARS-CoV-2 (refs. 4 – 11 ), would expand in vivo to support rapid viral control, aborting infection. We measured SARS-CoV-2-reactive T cells, including those against the early transcribed replication–transcription complex (RTC) 12 , 13 , in intensively monitored healthcare workers (HCWs) who tested repeatedly negative according to PCR, antibody binding and neutralization assays (seronegative HCWs (SN-HCWs)). SN-HCWs had stronger, more multispecific memory T cells compared with a cohort of unexposed individuals from before the pandemic (prepandemic cohort), and these cells were more frequently directed against the RTC than the structural-protein-dominated responses observed after detectable infection (matched concurrent cohort). SN-HCWs with the strongest RTC-specific T cells had an increase in IFI27 , a robust early innate signature of SARS-CoV-2 (ref. 14 ), suggesting abortive infection. RNA polymerase within RTC was the largest region of high sequence conservation across human seasonal coronaviruses (HCoV) and SARS-CoV-2 clades. RNA polymerase was preferentially targeted (among the regions tested) by T cells from prepandemic cohorts and SN-HCWs. RTC-epitope-specific T cells that cross-recognized HCoV variants were identified in SN-HCWs. Enriched pre-existing RNA-polymerase-specific T cells expanded in vivo to preferentially accumulate in the memory response after putative abortive compared to overt SARS-CoV-2 infection. Our data highlight RTC-specific T cells as targets for vaccines against endemic and emerging Coronaviridae . Seronegative healthcare workers with an innate signature of infection preferentially expand pre-existing T cells targeting the conserved replication transcription complex of SARS-CoV-2 in abortive infection.

BackgroundWithdrawal of prolonged nucleos(t)ide analogue (NA) treatment results in hepatitis B surface antigen (HBsAg) loss in some subjects with chronic hepatitis B (CHB), potentially revealing immune correlates of functional cure.ObjectiveWe investigated whether baseline or longitudinal changes in humoral immunity correlated with outcome of discontinuing prolonged NA treatment.DesignGlobal memory B cells (MBC) and T follicular helper cells (Tfh) were analysed by flow cytometry. HBs (small surface)/HBc (core)-MBC were quantified by ex-vivo bait staining and function assessed by cultured ELISpots (enzyme-linked immunosorbent spots). Immune parameters assessed at end-of-treatment (EOT), 12 and 48 weeks after treatment withdrawal (and at 4–8 years in a subset) were correlated with intrahepatic and longitudinal serum viral markers and alanine transaminase (ALT).ResultsIndividuals on prolonged NA had comparable frequencies of HBc-MBC and HBs-MBC, although the latter were PD-1hi and functionally defective. Following treatment withdrawal, increases in class-switched HBc-MBC were frequently temporally linked with hepatic flares. Subjects achieving HBsAg loss had an increase in activated global MBC detectable at EOT that become more marked by week 48, accompanied by significant increases in plasmablasts. HBs-MBC in those with HBsAg loss showed significant reductions in PD-1, trends to increased activation (CD71) and function and a more robust correlation with Tfh, compared with HBsAg persistence. MBC changes were maintained 4–8 years after HBsAg seroconversion.ConclusionDifferences in global and HBs-specific B cell immunity associate with HBsAg loss, whereas HBc-MBC temporally associates with flares, following withdrawal of prolonged NA treatment. Our results underscore the need to further explore the potential of B cell targets for monitoring and enhancing HBV functional cure in larger cohorts.

Memory B cells (MBCs) can provide a recall response able to supplement waning antibodies (Abs) with an affinity-matured response better able to neutralize variant viruses. We studied a cohort of elderly care home residents and younger staff (median age of 87 years and 56 years, respectively), who had survived COVID-19 outbreaks with only mild or asymptomatic infection. The cohort was selected because of its high proportion of individuals who had lost neutralizing antibodies (nAbs), thus allowing us to specifically investigate the reserve immunity from SARS-CoV-2-specific MBCs in this setting. Class-switched spike and receptor-binding domain (RBD) tetramer-binding MBCs persisted 5 months after mild or asymptomatic SARS-CoV-2 infection, irrespective of age. The majority of spike- and RBD-specific MBCs had a classical phenotype, but we found that activated MBCs, indicating possible ongoing antigenic stimulation or inflammation, were expanded in the elderly group. Spike- and RBD-specific MBCs remained detectable in the majority of individuals who had lost nAbs, although at lower frequencies and with a reduced IgG/IgA isotype ratio. Functional spike-, S1 subunit of the spike protein- (S1-), and RBD-specific recall was also detectable by enzyme-linked immune absorbent spot (ELISPOT) assay in some individuals who had lost nAbs, but was significantly impaired in the elderly. Our findings demonstrate that a reserve of SARS-CoV-2-specific MBCs persists beyond the loss of nAbs but highlight the need for careful monitoring of functional defects in spike- and RBD-specific B cell immunity in the elderly.

ObjectivesAn unlinked anonymous seroprevalence study was conducted to estimate the prevalence of hepatitis C virus (HCV) infection in samples derived from antenatal clinic attendees at 2 East London Hospitals. An unexpectedly high HCV seroprevalence of 2.6% (1.2% viraemic) had been revealed during an unlinked study of the emergency department at 1 of these hospitals.Design1000 stored residual samples were tested for HCV antibody (anti-HCV) and reactive samples were further tested for HCV RNA. The study was reviewed by the East Midland NRES ethics committee project ID 181154, approval number 15/WS/0125.ResultsThe anti-HCV reactivity rate was 0.5% (5/1000) with 0.1% (1/1000) confirmed viraemic. Prevalence for the other blood-borne viruses was higher: 1% (10/1000) were hepatitis B surface antigen positive and 0.3% were HIV antigen/antibody positive (3/1000). There were no co-infections.ConclusionsMore data to establish the prevalence of HCV in the antenatal population is needed. The addition of anti-HCV testing to the well-established antenatal screening programme provides a unique opportunity to impact on the health of pregnant women, their children, partners and future pregnancies in this new era of treatment for hepatitis C.

Memory B cells (MBC) can provide a recall response able to supplement waning antibodies with an affinity-matured response better able to neutralise variant viruses. We studied a cohort of vulnerable elderly care home residents and younger staff, a high proportion of whom had lost neutralising antibodies (nAb), to investigate their reserve immunity from SARS-CoV-2-specific MBC. Class-switched spike and RBD-tetramer-binding MBC with a classical phenotype persisted five months post-mild/asymptomatic SARS-CoV-2 infection, irrespective of age. Spike/RBD-specific MBC remained detectable in the majority who had lost nAb, although at lower frequencies and with a reduced IgG/IgA isotype ratio. Functional spike/S1/RBD-specific recall was also detectable by ELISpot in some who had lost nAb, but was significantly impaired in the elderly, particularly to RBD. Our findings demonstrate persistence of SARS-CoV-2-specific MBC beyond loss of nAb, but highlight the need for careful monitoring of functional defects in RBD-specific B cell immunity in the elderly. Circulating class-switched spike and RBD-specific memory B cells can outlast detectable neutralising antibodies but are functionally constrained in the elderly.

Pathological loss-of-function mutations in cyclin-dependent kinase-like 5 ( CDKL5 ) cause CDKL5 deficiency disorder (CDD), a rare and severe neurodevelopmental disorder associated with severe and medically refractory early-life epilepsy, motor, cognitive, visual, and autonomic disturbances in the absence of any structural brain pathology. Analysis of genetic variants in CDD has indicated that CDKL5 kinase function is central to disease pathology. CDKL5 encodes a serine-threonine kinase with significant homology to GSK3β, which has also been linked to synaptic function. Further, Cdkl5 knock-out rodents have increased GSK3β activity and often increased long-term potentiation (LTP). Thus, development of a specific CDKL5 inhibitor must be careful to exclude cross-talk with GSK3β activity. We synthesized and characterized specific, high-affinity inhibitors of CDKL5 that do not have detectable activity for GSK3β. These compounds are very soluble in water but blood–brain barrier penetration is low. In rat hippocampal brain slices, acute inhibition of CDKL5 selectively reduces postsynaptic function of AMPA-type glutamate receptors in a dose-dependent manner. Acute inhibition of CDKL5 reduces hippocampal LTP. These studies provide new tools and insights into the role of CDKL5 as a newly appreciated key kinase necessary for synaptic plasticity. Comparisons to rodent knock-out studies suggest that compensatory changes have limited the understanding of the roles of CDKL5 in synaptic physiology, plasticity, and human neuropathology.

Pathological loss-of-function mutations in cyclin-dependent kinase-like 5 ( CDKL5 ) cause CDKL5 deficiency disorder (CDD), a rare and severe neurodevelopmental disorder associated with severe and medically refractory early-life epilepsy, motor, cognitive, visual and autonomic disturbances in the absence of any structural brain pathology. Analysis of genetic variants in CDD have indicated that CDKL5 kinase function is central to disease pathology. CDKL5 encodes a serine-threonine kinase with significant homology to GSK3b, which has also been linked to synaptic function. Further, Cdkl5 knock-out rodents have increased GSK3b activity and often increased long-term potentiation (LTP). Thus, development of a specific CDKL5 inhibitor must be careful to exclude cross-talk with GSK3b activity. We synthesized and characterized specific, high-affinity inhibitors of CDKL5 that do not have detectable activity for GSK3b. These compounds are very soluble in water but blood-brain barrier penetration is low. In rat hippocampal brain slices, acute inhibition of CDKL5 selectively reduces post-synaptic function of AMPA-type glutamate receptors in a dose-dependent manner. Acute inhibition of CDKL5 reduces hippocampal LTP. These studies provide new tools and insights into the role of CDKL5 as a newly appreciated, key kinase necessary for synaptic plasticity. Comparisons to rodent knock-out studies suggest that compensatory changes have limited the understanding of the roles of CDKL5 in synaptic physiology, plasticity and human neuropathology.Pathological loss-of-function mutations in cyclin-dependent kinase-like 5 ( CDKL5 ) cause CDKL5 deficiency disorder (CDD), a rare and severe neurodevelopmental disorder associated with severe and medically refractory early-life epilepsy, motor, cognitive, visual and autonomic disturbances in the absence of any structural brain pathology. Analysis of genetic variants in CDD have indicated that CDKL5 kinase function is central to disease pathology. CDKL5 encodes a serine-threonine kinase with significant homology to GSK3b, which has also been linked to synaptic function. Further, Cdkl5 knock-out rodents have increased GSK3b activity and often increased long-term potentiation (LTP). Thus, development of a specific CDKL5 inhibitor must be careful to exclude cross-talk with GSK3b activity. We synthesized and characterized specific, high-affinity inhibitors of CDKL5 that do not have detectable activity for GSK3b. These compounds are very soluble in water but blood-brain barrier penetration is low. In rat hippocampal brain slices, acute inhibition of CDKL5 selectively reduces post-synaptic function of AMPA-type glutamate receptors in a dose-dependent manner. Acute inhibition of CDKL5 reduces hippocampal LTP. These studies provide new tools and insights into the role of CDKL5 as a newly appreciated, key kinase necessary for synaptic plasticity. Comparisons to rodent knock-out studies suggest that compensatory changes have limited the understanding of the roles of CDKL5 in synaptic physiology, plasticity and human neuropathology.