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Hepatitis B Virus (HBV) is a major driver of infectious disease mortality. Curative therapies are needed and ideally should induce CD8 T cell-mediated clearance of infected hepatocytes plus anti-hepatitis B surface antigen (HBsAg) antibodies (anti-HBs) to neutralize residual virus. We developed a novel therapeutic vaccine using non-replicating arenavirus vectors. Antigens were screened for genotype conservation and magnitude and genotype reactivity of T cell response, then cloned into Pichinde virus (PICV) vectors (recombinant PICV, GS-2829) and lymphocytic choriomeningitis virus (LCMV) vectors (replication-incompetent, GS-6779). Alternating immunizations with GS-2829 and GS-6779 induced high-magnitude HBV T cell responses, and high anti-HBs titers. Dose schedule optimization in macaques achieved strong polyfunctional CD8 T cell responses against core, HBsAg, and polymerase and high titer anti-HBs. In AAV-HBV mice, GS-2829 and GS-6779 were efficacious in animals with low pre-treatment serum HBsAg. Based on these results, GS-2829 and GS-6779 could become a central component of cure regimens.
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Arenavirus , Hepatitis B , Ratones , Animales , Antígenos de Superficie de la Hepatitis B , Virus de la Hepatitis B/genética , Vacunas contra Hepatitis B , Anticuerpos contra la Hepatitis B , Inmunización , Linfocitos T CD8-positivos , Genotipo , Antígenos de SuperficieRESUMEN
OBJECTIVE: Achieving HBV cure will require novel combination therapies of direct-acting antivirals and immunomodulatory agents. In this context, the toll-like receptor 8 (TLR8) agonist selgantolimod (SLGN) has been investigated in preclinical models and clinical trials for chronic hepatitis B (CHB). However, little is known regarding its action on immune effectors within the liver. Our aim was to characterise the transcriptomic changes and intercellular communication events induced by SLGN in the hepatic microenvironment. DESIGN: We identified TLR8-expressing cell types in the human liver using publicly available single-cell RNA-seq data and established a method to isolate Kupffer cells (KCs). We characterised transcriptomic and cytokine KC profiles in response to SLGN. SLGN's indirect effect was evaluated by RNA-seq in hepatocytes treated with SLGN-conditioned media (CM) and quantification of HBV parameters following infection. Pathways mediating SLGN's effect were validated using transcriptomic data from HBV-infected patients. RESULTS: Hepatic TLR8 expression takes place in the myeloid compartment. SLGN treatment of KCs upregulated monocyte markers (eg, S100A12) and downregulated genes associated with the KC identity (eg, SPIC). Treatment of hepatocytes with SLGN-CM downregulated NTCP and impaired HBV entry. Cotreatment with an interleukin 6-neutralising antibody reverted the HBV entry inhibition. CONCLUSION: Our transcriptomic characterisation of SLGN sheds light into the programmes regulating KC activation. Furthermore, in addition to its previously described effect on established HBV infection and adaptive immunity, we show that SLGN impairs HBV entry. Altogether, SLGN may contribute through KCs to remodelling the intrahepatic immune microenvironment and may thus represent an important component of future combinations to cure HBV infection.
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OBJECTIVE: Selected populations of patients with chronic hepatitis B (CHB) may benefit from a combined use of pegylated interferon-alpha (pegIFN-α) and nucleos(t)ides (NUCs). The aim of our study was to assess the immunomodulatory effect of pegIFN-α on T and natural killer (NK) cell responses in NUC-suppressed patients to identify cellular and/or serological parameters to predict better T cell-restoring effect and better control of infection in response to pegIFN-α for a tailored application of IFN-α add-on. DESIGN: 53 HBeAg-negative NUC-treated patients with CHB were randomised at a 1:1 ratio to receive pegIFN-α-2a for 48 weeks, or to continue NUC therapy and then followed up for at least 6 months maintaining NUCs. Serum hepatitis B surface antigen (HBsAg) and hepatitis B core-related antigen (HBcrAg) levels as well as peripheral blood NK cell phenotype and function and HBV-specific T cell responses upon in vitro stimulation with overlapping HBV peptides were measured longitudinally before, during and after pegIFN-α therapy. RESULTS: Two cohorts of pegIFN-α treated patients were identified according to HBsAg decline greater or less than 0.5 log at week 24 post-treatment. PegIFN-α add-on did not significantly improve HBV-specific T cell responses during therapy but elicited a significant multispecific and polyfunctional T cell improvement at week 24 post-pegIFN-α treatment compared with baseline. This improvement was maximal in patients who had a higher drop in serum HBsAg levels and a lower basal HBcrAg values. CONCLUSIONS: PegIFN-α treatment can induce greater functional T cell improvement and HBsAg decline in patients with lower baseline HBcrAg levels. Thus, HBcrAg may represent an easily and reliably applicable parameter to select patients who are more likely to achieve better response to pegIFN-α add-on to virally suppressed patients.
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Capsid assembly mediated by hepatitis B virus (HBV) core protein (HBc) is an essential part of the HBV replication cycle, which is the target for different classes of capsid assembly modulators (CAMs). While both CAM-A ("aberrant") and CAM-E ("empty") disrupt nucleocapsid assembly and reduce extracellular HBV DNA, CAM-As can also reduce extracellular HBV surface antigen (HBsAg) by triggering apoptosis of HBV-infected cells in preclinical mouse models. However, there have not been substantial HBsAg declines in chronic hepatitis B (CHB) patients treated with CAM-As to date. To investigate this disconnect, we characterized the antiviral activity of tool CAM compounds in HBV-infected primary human hepatocytes (PHHs), as well as in HBV-infected human liver chimeric mice and mice transduced with adeno-associated virus-HBV. Mechanistic studies in HBV-infected PHH revealed that CAM-A, but not CAM-E, induced a dose-dependent aggregation of HBc in the nucleus which is negatively regulated by the ubiquitin-binding protein p62. We confirmed that CAM-A, but not CAM-E, induced HBc-positive cell death in both mouse models via induction of apoptotic and inflammatory pathways and demonstrated that the degree of HBV-positive cell loss was positively correlated with intrahepatic HBc levels. Importantly, we determined that there is a significantly lower level of HBc per hepatocyte in CHB patient liver biopsies than in either of the HBV mouse models. Taken together, these data confirm that CAM-As have a unique secondary mechanism with the potential to kill HBc-positive hepatocytes. However, this secondary mechanism appears to require higher intrahepatic HBc levels than is typically observed in CHB patients, thereby limiting the therapeutic potential.
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Virus de la Hepatitis B , Hepatitis B Crónica , Hepatocitos , Humanos , Hepatocitos/virología , Hepatocitos/efectos de los fármacos , Animales , Virus de la Hepatitis B/efectos de los fármacos , Virus de la Hepatitis B/fisiología , Ratones , Hepatitis B Crónica/tratamiento farmacológico , Hepatitis B Crónica/virología , Proteínas del Núcleo Viral/metabolismo , Antivirales/farmacología , Antivirales/uso terapéutico , Antígenos del Núcleo de la Hepatitis B/metabolismo , Cápside/metabolismo , Cápside/efectos de los fármacos , Hígado/virología , Hígado/efectos de los fármacos , Hígado/metabolismo , Antígenos de Superficie de la Hepatitis B/metabolismo , Ensamble de Virus/efectos de los fármacos , Apoptosis/efectos de los fármacos , Replicación Viral/efectos de los fármacosRESUMEN
OBJECTIVES: A major goal of curative hepatitis B virus (HBV) treatments is the reduction or inactivation of intrahepatic viral covalently closed circular DNA (cccDNA). Hence, precise cccDNA quantification is essential in preclinical and clinical studies. Southern blot (SB) permits cccDNA visualisation but lacks sensitivity and is very laborious. Quantitative PCR (qPCR) has no such limitations but inaccurate quantification due to codetection of viral replicative intermediates (RI) can occur. The use of different samples, preservation conditions, DNA extraction, nuclease digestion methods and qPCR strategies has hindered standardisation. Within the ICE-HBV consortium, available and novel protocols for cccDNA isolation and qPCR quantification in liver tissues and cell cultures were compared in six laboratories to develop evidence-based guidance for best practices. DESIGN: Reference material (HBV-infected humanised mouse livers and HepG2-NTCP cells) was exchanged for cross-validation. Each group compared different DNA extraction methods (Hirt extraction, total DNA extraction with or without proteinase K treatment (+PK/-PK)) and nuclease digestion protocols (plasmid-safe ATP-dependent DNase (PSD), T5 exonuclease, exonucleases I/III). Samples were analysed by qPCR and SB. RESULTS: Hirt and -PK extraction reduced coexisting RI forms. However, both cccDNA and the protein-free relaxed circular HBV DNA (pf-rcDNA) form were detected by qPCR. T5 and Exo I/III nucleases efficiently removed all RI forms. In contrast, PSD did not digest pf-rcDNA, but was less prone to induce cccDNA overdigestion. In stabilised tissues (eg, Allprotect), nucleases had detrimental effects on cccDNA. CONCLUSIONS: We present here a comprehensive evidence-based guidance for optimising, controlling and validating cccDNA measurements using available qPCR assays.
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ADN Circular , Virus de la Hepatitis B , Animales , Ratones , Humanos , Virus de la Hepatitis B/genética , ADN Circular/genética , Hígado , Reacción en Cadena de la Polimerasa/métodos , Células Hep G2 , ADN Viral/genéticaRESUMEN
OBJECTIVE: Exhausted hepatitis B virus (HBV)-specific CD8 T cells in chronic HBV infection are broadly heterogeneous. Characterisation of their functional impairment may allow to distinguish patients with different capacity to control infection and reconstitute antiviral function. DESIGN: HBV dextramer+CD8 T cells were analysed ex vivo for coexpression of checkpoint/differentiation markers, transcription factors and cytokines in 35 patients with HLA-A2+chronic hepatitis B (CHB) and in 29 control HBsAg negative CHB patients who seroconverted after NUC treatment or spontaneously. Cytokine production was also evaluated in HBV peptide-stimulated T cell cultures, in the presence or absence of antioxidant, polyphenolic, PD-1/PD-L1 inhibitor and TLR-8 agonist compounds and the effect on HBV-specific responses was further validated on additional 24 HLA-A2 negative CHB patients. RESULTS: Severely exhausted HBV-specific CD8 T cell subsets with high expression of inhibitory receptors, such as PD-1, TOX and CD39, were detected only in a subgroup of chronic viraemic patients. Conversely, a large predominance of functionally more efficient HBV-specific CD8 T cell subsets with lower expression of coinhibitory molecules and better response to in vitro immune modulation, typically detected after resolution of infection, was also observed in a proportion of chronic viraemic HBV patients. Importantly, the same subset of patients who responded more efficiently to in vitro immune modulation identified by HBV-specific CD8 T cell analysis were also identified by staining total CD8 T cells with PD-1, TOX, CD127 and Bcl-2. CONCLUSIONS: The possibility to distinguish patient cohorts with different capacity to respond to immune modulatory compounds in vitro by a simple analysis of the phenotypic CD8 T cell exhaustion profile deserves evaluation of its clinical applicability.
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Hepatitis B Crónica , Hepatitis B , Humanos , Hepatitis B Crónica/tratamiento farmacológico , Virus de la Hepatitis B , Antígeno HLA-A2/metabolismo , Antígeno HLA-A2/farmacología , Antígeno HLA-A2/uso terapéutico , Receptor de Muerte Celular Programada 1/metabolismo , Linfocitos T CD8-positivosRESUMEN
BACKGROUND & AIMS: Selgantolimod (GS-9688) is a Toll-like receptor 8 (TLR8) agonist that suppresses HBV in vitro. In a phase II study, we evaluated the safety and efficacy of weekly selgantolimod treatment in virally suppressed individuals with chronic HBV taking oral antiviral treatment. METHODS: Forty-eight patients were randomized into two cohorts (hepatitis B e antigen [HBeAg]-positive and -negative [n = 24 each]) to receive oral selgantolimod 3 mg, 1.5 mg, or placebo (2:2:1) once weekly for 24 weeks while maintaining oral antivirals. The primary efficacy endpoint was the percentage of patients with a ≥1 log10 IU/ml decline in hepatitis B surface antigen (HBsAg) from baseline to week 24. Post-treatment, patients continued on oral antivirals for 24 weeks. RESULTS: The primary endpoint was reached by one participant, who was HBeAg-negative and received selgantolimod 1.5 mg. In contrast with placebo-treated patients (n = 9), only selgantolimod-treated patients (n = 39 total) had HBsAg declines greater than 0.1 log10 IU/ml at weeks 24 (18%, 7/39) and 48 (26%, 10/39), HBsAg loss (5%, 2/39 through 48 weeks), or HBeAg loss (16%, 3/19 through 48 weeks). The most common adverse events in selgantolimod-treated groups were nausea (46%), upper respiratory tract infection (23%), and vomiting (23%). Gastrointestinal disorders were mostly mild and transient. Selgantolimod induced transient dose-dependent increases in serum cytokines, including IL-12p40, IFN-γ, and IL-1RA, as well as rapid redistribution of some circulating immune cell subsets. CONCLUSION: Oral selgantolimod up to 3 mg once weekly for 24 weeks was generally safe and well tolerated and led to serologic changes associated with progression to durable cure in two individuals by week 48. GOV IDENTIFIER: NCT03491553. IMPACT AND IMPLICATIONS: The only robust criterion for stopping treatment in chronic hepatitis B is loss of hepatitis B surface antigen (known as functional cure), which is rare during nucleos(t)ide analogue therapy. It is likely that novel antiviral and immunomodulatory therapies will be needed to achieve finite functional cure. Selgantolimod is an oral Toll-like receptor 8 agonist that has shown antiviral activity in vitro as well as safety in a phase I clinical trial with weekly dosing. In this phase II study, selgantolimod therapy was associated with transient increases in serum cytokines, rapid redistribution of circulating immune cell subsets, modest reductions in HBsAg and HBeAg levels, and occasional loss of HBsAg (5%) and HBeAg (16%) among participants with chronic hepatitis B on nucleos(t)ide analogue therapy with viral suppression. Our results support continued development of selgantolimod as a component of a future hepatitis B cure regimen.
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Antivirales , Hepatitis B Crónica , Receptor Toll-Like 8 , Humanos , Antivirales/uso terapéutico , Citocinas , Antígenos e de la Hepatitis B , Antígenos de Superficie de la Hepatitis B , Hepatitis B Crónica/tratamiento farmacológico , Receptor Toll-Like 8/agonistas , Resultado del TratamientoRESUMEN
BACKGROUND & AIMS: Chronic coinfection with HBV and HDV leads to the most aggressive form of chronic viral hepatitis. Herein, we aimed to elucidate the molecular mechanisms underlying the widely reported observation that HDV interferes with HBV in most coinfected patients. METHODS: Patient liver tissues, primary human hepatocytes, HepaRG cells and human liver chimeric mice were used to analyze the effect of HDV on HBV using virological and RNA-sequencing analyses, as well as RNA synthesis, stability and association assays. RESULTS: Transcriptomic analyses in cell culture and mouse models of coinfection enabled us to define an HDV-induced signature, mainly composed of interferon (IFN)-stimulated genes (ISGs). We also provide evidence that ISGs are upregulated in chronically HDV/HBV-coinfected patients but not in cells that only express HDV antigen (HDAg). Inhibition of the hepatocyte IFN response partially rescued the levels of HBV parameters. We observed less HBV RNA synthesis upon HDV infection or HDV protein expression. Additionally, HDV infection or expression of HDAg alone specifically accelerated the decay of HBV RNA, and HDAg was associated with HBV RNAs. On the contrary, HDAg expression did not affect other viruses such as HCV or SARS-CoV-2. CONCLUSIONS: Our data indicate that HDV interferes with HBV through both IFN-dependent and IFN-independent mechanisms. Specifically, we uncover a new viral interference mechanism in which proteins of a satellite virus affect the RNA production of its helper virus. Exploiting these findings could pave the way to the development of new therapeutic strategies against HBV. IMPACT AND IMPLICATIONS: Although the molecular mechanisms remained unexplored, it has long been known that despite its dependency, HDV decreases HBV viremia in patients. Herein, using in vitro and in vivo models, we showed that HDV interferes with HBV through both IFN-dependent and IFN-independent mechanisms affecting HBV RNA metabolism, and we defined the HDV-induced modulation signature. The mechanisms we uncovered could pave the way for the development of new therapeutic strategies against HBV by mimicking and/or increasing the effect of HDAg on HBV RNA. Additionally, the HDV-induced modulation signature could potentially be correlated with responsiveness to IFN-α treatment, thereby helping to guide management of HBV/HDV-coinfected patients.
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COVID-19 , Coinfección , Hepatitis B , Hepatitis D , Humanos , Ratones , Animales , Virus de la Hepatitis Delta/fisiología , Virus de la Hepatitis B/fisiología , Interferones , Antígenos de Hepatitis delta/metabolismo , Hepatitis D/complicaciones , Hepatitis B/complicaciones , Replicación Viral/fisiología , COVID-19/complicaciones , SARS-CoV-2/genética , ARN Viral/genéticaRESUMEN
The standard of care for the treatment of chronic hepatitis B (CHB) is typically lifelong treatment with nucleos(t)ide analogs (NAs), which suppress viral replication and provide long-term clinical benefits. However, infectious virus can still be detected in patients who are virally suppressed on NA therapy, which may contribute to the failure of these agents to cure most CHB patients. Accordingly, new antiviral treatment options are being developed to enhance the suppression of hepatitis B virus (HBV) replication in combination with NAs ("antiviral intensification"). Here, we describe GS-SBA-1, a capsid assembly modulator (CAM) belonging to class CAM-E, that demonstrates potent inhibition of extracellular HBV DNA in vitro (EC50 [50% effective concentration] = 19 nM) in HBV-infected primary human hepatocytes (PHHs) as well as in vivo in an HBV-infected immunodeficient mouse model. GS-SBA-1 has comparable activities across HBV genotypes and nucleos(t)ide-resistant mutants in HBV-infected PHHs. In addition, GS-SBA-1 demonstrated in vitro additivity in combination with tenofovir alafenamide (TAF). The administration of GS-SBA-1 to PHHs at the time of infection prevents covalently closed circular DNA (cccDNA) formation and, hence, decreases HBV RNA and antigen levels (EC50 = 80 to 200 nM). Furthermore, GS-SBA-1 prevents the production of extracellular HBV RNA-containing viral particles in vitro. Collectively, these data demonstrate that GS-SBA-1 is a potent CAM that has the potential to enhance viral suppression in combination with an NA.
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Hepatitis B Crónica , Hepatitis B , Animales , Ratones , Humanos , Hepatitis B Crónica/tratamiento farmacológico , Cápside , Virus de la Hepatitis B , Antivirales/farmacología , Antivirales/uso terapéutico , Proteínas de la Cápside/genética , ARN , ADN Viral/genética , ADN Circular , Hepatitis B/tratamiento farmacológicoRESUMEN
BACKGROUND: Qualitative studies have explored why people with haemophilia (PwH) might consider or forgo gene therapy, the impact it has had on those who have received it, and what support might be needed throughout the process. No studies have yet examined what withdrawal prior to transfection might mean for PwH and their families. AIMS: To understand the experiences of PwH and their families of withdrawal from gene therapy and to understand what support mechanisms might be required. METHODS: Qualitative interviews were conducted with people with severe haemophilia who consented to take part in a gene therapy study in the UK but either withdrew or were withdrawn before transfection. RESULTS: Nine PwH and a family member were invited to this sub-study. Eight participants were recruited: six PwH (five haemophilia A and one haemophilia B) and two family members. Four participants were excluded from a study after consenting but before transfection, having failed to meet all the inclusion criteria; two withdrew after consenting but before transfection due to concerns including duration of factor expression and the time commitment involved in follow-up. The mean age of participants was 40.5 years (range 25-63 years). Two major themes emerged during the interviews: Expectation and loss. CONCLUSIONS: PwH have many expectations about the difference gene therapy can make to their lives. Studies show that these expectations may not be fully realised. For those who have either withdrawn or been withdrawn from gene therapy, any expectations they had may now be unachievable. The nature of these expectations and the loss expressed by the participants indicate that support needs to be provided to help them and their families manage it.
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Hemofilia A , Hemofilia B , Humanos , Adulto , Persona de Mediana Edad , Hemofilia A/genética , Hemofilia A/terapia , Motivación , Familia , Modalidades de FisioterapiaRESUMEN
INTRODUCTION: The UK National Haemophilia Database (NHD) collects data from all UK persons with haemophilia A with inhibitors (PwHA-I). It is well-placed to investigate patient selection, clinical outcomes, drug safety and other issues not addressed in clinical trials of emicizumab. AIMS: To determine safety, bleeding outcomes and early effects on joint health of emicizumab prophylaxis in a large, unselected cohort using national registry and patient reported Haemtrack (HT) data between 01 January 2018 and 30 September 2021. METHODS: Prospectively collected bleeding outcomes were analysed in people with ≥6 months emicizumab HT data and compared with previous treatment if available. Change in paired Haemophilia Joint Health Scores (HJHS) were analysed in a subgroup. Adverse events (AEs) reports were collected and adjudicated centrally. RESULTS: This analysis includes 117 PwHA-I. Mean annualised bleeding rate (ABR) was .32 (95% CI, .18; .39) over a median 42 months treatment with emicizumab. Within-person comparison (n = 74) demonstrated an 89% reduction in ABR after switching to emicizumab and an increase in zero treated bleed rate from 45 to 88% (p < .01). In a subgroup of 37 people, total HJHS improved in 36%, remained stable in 46% and deteriorated in 18%, with a median (IQR) within-person change of -2.0 (-9, 1.5) (p = .04). Three arterial thrombotic events were reported, two possibly drug related. Other AEs were generally non-severe and usually limited to early treatment, included cutaneous reactions (3.6%), headaches (1.4%), nausea (2.8%) and arthralgia (1.4%). CONCLUSIONS: Emicizumab prophylaxis is associated with sustained low bleeding rates and was generally well-tolerated in people with haemophilia A and inhibitors.
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Anticuerpos Biespecíficos , Hemofilia A , Humanos , Hemofilia A/complicaciones , Hemofilia A/tratamiento farmacológico , Estudios de Seguimiento , Anticuerpos Biespecíficos/efectos adversos , Hemorragia/complicaciones , Reino Unido , Factor VIII/uso terapéuticoRESUMEN
BACKGROUNDS AND AIMS: Toll-like receptor (TLR) agonists have been developed as adjuvants to efficiently induce antiviral immune responses. Specificity and potency of these compounds are essential requirements for clinical trial applications. In patients with hepatitis B virus (HBV) infections, sustained loss of hepatitis B surface antigen (HBsAg) is a therapeutic goal, which may be achievable by the sequential activation of follicular helper T cells (Tfh) and antibody-secreting B cells. We aimed to elucidate whether novel TLR7 agonist, GS-986, could activate immune responses involved in HBV elimination. METHODS: To clarify the impact of GS-986 on pDCs, we quantified the expression levels of surface markers and evaluated for Tfh induction in a culture model consisting of human pDCs with allogeneic naïve CD4+ T cells. In addition, we examined whether GS-986 could enhance HBs antibody production capacity using PBMC from CHB patients. RESULTS: pDCs from CHB patients had lower OX40L expression and as well as impaired capacity for Tfh induction compared with those from healthy donors. However, GS-986-stimulated pDCs from CHB patients expressed OX40L and produced IL-6 and IL-12, resulting in the induction of IL-21-producing Tfh cells (CXCR5+ PD-1+ CD4+ ) from naïve CD4+ T cells. The Tfh-inducing capacity of GS-986 was reduced in the presence of an anti-OX40L blocking antibody. Furthermore, GS-986 promoted HBsAg-specific antibody production in PBMCs from CHB patients. CONCLUSIONS: GS-986 is an adjuvant that stimulates pDCs to induce Tfh differentiation and antigen-specific B-cell production. This immune profile may be beneficial for therapeutic application as an immune modulator in CHB patients.
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Antivirales , Hepatitis B Crónica , Receptor Toll-Like 7 , Hepatitis B Crónica/tratamiento farmacológico , Humanos , Receptor Toll-Like 7/agonistas , Antivirales/uso terapéutico , Masculino , Femenino , Persona de Mediana Edad , Regulación hacia Arriba , Linfocitos T CD4-Positivos/citología , Linfocitos T CD4-Positivos/efectos de los fármacos , Diferenciación Celular , Células Dendríticas/efectos de los fármacos , Células Dendríticas/metabolismo , Células T Auxiliares Foliculares/citología , Células T Auxiliares Foliculares/efectos de los fármacos , Antígenos de Superficie de la Hepatitis B/metabolismo , Anticuerpos Antivirales/metabolismoRESUMEN
Interprofessional education (IPE) interventions aiming to promote collaborative competence and improve the delivery of health and social care processes and outcomes continue to evolve. This paper reports on a protocol for an update review that we will conduct to identify and describe how the IPE evidence base has evolved in the last 7 years. We will identify literature through a systematic search of the following electronic databases: Medline, Embase, CINAHL, Education Source, ERIC, and BEI. We will consider all IPE interventions delivered to health professions students and accredited professionals. Peer-reviewed empirical research studies published in any language from June 2014 onwards will be eligible for inclusion. The outcomes of interest are changes in the reaction, attitudes/perceptions, knowledge/skills acquisition, behaviors, organizational practice, and/or benefits to patients. We will perform each task of screening, critical appraisal, data abstraction, and synthesis using at least two members of the review team. The review will enable an update and comprehensive understanding of the IPE evidence base to inform future IPE developments, delivery and evaluation across education and clinical settings.
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Educación Interprofesional , Estudiantes del Área de la Salud , Humanos , Empleos en Salud , Relaciones Interprofesionales , Cuidados Paliativos , Literatura de Revisión como AsuntoRESUMEN
OBJECTIVE: Therapeutic strategies silencing and reducing the hepatitis B virus (HBV) reservoir, the covalently closed circular DNA (cccDNA), have the potential to cure chronic HBV infection. We aimed to investigate the impact of small interferring RNA (siRNA) targeting all HBV transcripts or pegylated interferon-α (peg-IFNα) on the viral regulatory HBx protein and the structural maintenance of chromosome 5/6 complex (SMC5/6), a host factor suppressing cccDNA transcription. In particular, we assessed whether interventions lowering HBV transcripts can achieve and maintain silencing of cccDNA transcription in vivo. DESIGN: HBV-infected human liver chimeric mice were treated with siRNA or peg-IFNα. Virological and host changes were analysed at the end of treatment and during the rebound phase by qualitative PCR, ELISA, immunoblotting and chromatin immunoprecipitation. RNA in situ hybridisation was combined with immunofluorescence to detect SMC6 and HBV RNAs at single cell level. The entry inhibitor myrcludex-B was used during the rebound phase to avoid new infection events. RESULTS: Both siRNA and peg-IFNα strongly reduced all HBV markers, including HBx levels, thus enabling the reappearance of SMC5/6 in hepatocytes that achieved HBV-RNA negativisation and SMC5/6 association with the cccDNA. Only IFN reduced cccDNA loads and enhanced IFN-stimulated genes. However, the antiviral effects did not persist off treatment and SMC5/6 was again degraded. Remarkably, the blockade of viral entry that started at the end of treatment hindered renewed degradation of SMC5/6. CONCLUSION: These results reveal that therapeutics abrogating all HBV transcripts including HBx promote epigenetic suppression of the HBV minichromosome, whereas strategies protecting the human hepatocytes from reinfection are needed to maintain cccDNA silencing.
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Proteínas de Ciclo Celular/metabolismo , Virus de la Hepatitis B/fisiología , Hepatitis B Crónica/metabolismo , Hepatitis B Crónica/virología , Animales , Quimera , ADN Circular/metabolismo , Genoma Viral , Hepatitis B Crónica/prevención & control , Humanos , RatonesRESUMEN
BACKGROUND AND AIMS: GS-9688 (selgantolimod) is an oral selective small molecule agonist of toll-like receptor 8 in clinical development for the treatment of chronic hepatitis B. In this study, we evaluated the antiviral efficacy of GS-9688 in woodchucks chronically infected with woodchuck hepatitis virus (WHV), a hepadnavirus closely related to hepatitis B virus. APPROACH AND RESULTS: WHV-infected woodchucks received eight weekly oral doses of vehicle, 1 mg/kg GS-9688, or 3 mg/kg GS-9688. Vehicle and 1 mg/kg GS-9688 had no antiviral effect, whereas 3 mg/kg GS-9688 induced a >5 log10 reduction in serum viral load and reduced WHV surface antigen (WHsAg) levels to below the limit of detection in half of the treated woodchucks. In these animals, the antiviral response was maintained until the end of the study (>5 months after the end of treatment). GS-9688 treatment reduced intrahepatic WHV RNA and DNA levels by >95% in animals in which the antiviral response was sustained after treatment cessation, and these woodchucks also developed detectable anti-WHsAg antibodies. The antiviral efficacy of weekly oral dosing with 3 mg/kg GS-9688 was confirmed in a second woodchuck study. The antiviral response to GS-9688 did not correlate with systemic GS-9688 or cytokine levels but was associated with transient elevation of liver injury biomarkers and enhanced proliferative response of peripheral blood mononuclear cells to WHV peptides. Transcriptomic analysis of liver biopsies taken prior to treatment suggested that T follicular helper cells and various other immune cell subsets may play a role in the antiviral response to GS-9688. CONCLUSIONS: Finite, short-duration treatment with a clinically relevant dose of GS-9688 is well tolerated and can induce a sustained antiviral response in WHV-infected woodchucks; the identification of a baseline intrahepatic transcriptional signature associated with response to GS-9688 treatment provides insights into the immune mechanisms that mediate this antiviral effect.
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Antivirales/uso terapéutico , Virus de la Hepatitis B de la Marmota/efectos de los fármacos , Virus de la Hepatitis B de la Marmota/genética , Hepatitis B Crónica/tratamiento farmacológico , Hexanoles/uso terapéutico , Pirimidinas/uso terapéutico , Receptor Toll-Like 8/agonistas , Animales , Antivirales/farmacología , ADN Viral/sangre , Modelos Animales de Enfermedad , Anticuerpos Antihepatitis/sangre , Antígenos de la Hepatitis/sangre , Virus de la Hepatitis B de la Marmota/inmunología , Hepatitis B Crónica/complicaciones , Hepatitis B Crónica/inmunología , Hexanoles/farmacología , Humanos , Marmota , Pirimidinas/farmacología , Replicación Viral/efectos de los fármacosRESUMEN
BACKGROUND AND AIMS: GS-9688 (selgantolimod) is a toll-like receptor 8 agonist in clinical development for the treatment of chronic hepatitis B (CHB). Antiviral activity of GS-9688 has previously been evaluated in vitro in HBV-infected hepatocytes and in vivo in the woodchuck model of CHB. Here we evaluated the potential of GS-9688 to boost responses contributing to viral control and to modulate regulatory mediators. APPROACH AND RESULTS: We characterized the effect of GS-9688 on immune cell subsets in vitro in peripheral blood mononuclear cells of healthy controls and patients with CHB. GS-9688 activated dendritic cells and mononuclear phagocytes to produce IL-12 and other immunomodulatory mediators, inducing a comparable cytokine profile in healthy controls and patients with CHB. GS-9688 increased the frequency of activated natural killer (NK) cells, mucosal-associated invariant T cells, CD4+ follicular helper T cells, and, in about 50% of patients, HBV-specific CD8+ T cells expressing interferon-γ. Moreover, in vitro stimulation with GS-9688 induced NK-cell expression of interferon-γ and TNF-α, and promoted hepatocyte lysis. We also assessed whether GS-9688 inhibited immunosuppressive cell subsets that might enhance antiviral efficacy. Stimulation with GS-9688 reduced the frequency of CD4+ regulatory T cells and monocytic myeloid-derived suppressor cells (MDSCs). Residual MDSCs expressed higher levels of negative immune regulators, galectin-9 and programmed death-ligand 1. Conversely, GS-9688 induced an expansion of immunoregulatory TNF-related apoptosis-inducing ligand+ NK cells and degranulation of arginase-I+ polymorphonuclear MDSCs. CONCLUSIONS: GS-9688 induces cytokines in human peripheral blood mononuclear cells that are able to activate antiviral effector function by multiple immune mediators (HBV-specific CD8+ T cells, CD4+ follicular helper T cells, NK cells, and mucosal-associated invariant T cells). Although reducing the frequency of some immunoregulatory subsets, it enhances the immunosuppressive potential of others, highlighting potential biomarkers and immunotherapeutic targets to optimize the antiviral efficacy of GS-9688.
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Antivirales/farmacología , Hepatitis B Crónica/tratamiento farmacológico , Hexanoles/farmacología , Pirimidinas/farmacología , Receptor Toll-Like 8/antagonistas & inhibidores , Adulto , Anciano , Animales , Antivirales/uso terapéutico , Linfocitos T CD8-positivos/efectos de los fármacos , Linfocitos T CD8-positivos/inmunología , Estudios de Cohortes , Modelos Animales de Enfermedad , Femenino , Voluntarios Sanos , Células Hep G2 , Hepatitis B Crónica/inmunología , Hepatitis B Crónica/virología , Hexanoles/uso terapéutico , Interacciones Huésped-Patógeno/efectos de los fármacos , Interacciones Huésped-Patógeno/inmunología , Humanos , Células Asesinas Naturales/efectos de los fármacos , Células Asesinas Naturales/inmunología , Leucocitos Mononucleares , Masculino , Marmota , Persona de Mediana Edad , Cultivo Primario de Células , Pirimidinas/uso terapéutico , Linfocitos T Reguladores/efectos de los fármacos , Linfocitos T Reguladores/inmunología , Receptor Toll-Like 8/metabolismo , Adulto JovenRESUMEN
Chronic hepatitis B virus infection is a leading cause of cirrhosis and liver cancer. Hepatitis B virus encodes the regulatory HBx protein whose primary role is to promote transcription of the viral genome, which persists as an extrachromosomal DNA circle in infected cells. HBx accomplishes this task by an unusual mechanism, enhancing transcription only from extrachromosomal DNA templates. Here we show that HBx achieves this by hijacking the cellular DDB1-containing E3 ubiquitin ligase to target the 'structural maintenance of chromosomes' (Smc) complex Smc5/6 for degradation. Blocking this event inhibits the stimulatory effect of HBx both on extrachromosomal reporter genes and on hepatitis B virus transcription. Conversely, silencing the Smc5/6 complex enhances extrachromosomal reporter gene transcription in the absence of HBx, restores replication of an HBx-deficient hepatitis B virus, and rescues wild-type hepatitis B virus in a DDB1-knockdown background. The Smc5/6 complex associates with extrachromosomal reporters and the hepatitis B virus genome, suggesting a direct mechanism of transcriptional inhibition. These results uncover a novel role for the Smc5/6 complex as a restriction factor selectively blocking extrachromosomal DNA transcription. By destroying this complex, HBx relieves the inhibition to allow productive hepatitis B virus gene expression.
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Proteínas de Ciclo Celular/metabolismo , Virus de la Hepatitis B/fisiología , Especificidad del Huésped , Transactivadores/metabolismo , Animales , Línea Celular Tumoral , Proteínas Cromosómicas no Histona , ADN Viral/genética , ADN Viral/metabolismo , Genes Reporteros , Genoma Viral/genética , Hepatitis B/virología , Virus de la Hepatitis B/genética , Hepatocitos/virología , Humanos , Hígado/metabolismo , Hígado/virología , Masculino , Ratones , Plásmidos/genética , Plásmidos/metabolismo , Unión Proteica , Proteolisis , Transcripción Genética , Ubiquitina/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Proteínas Reguladoras y Accesorias Virales , Replicación ViralRESUMEN
BACKGROUND: People with haemophilia A and inhibitors (PwHi) suffer more orthopaedic complications, bleeding and pain than those without inhibitors. The advent of emicizumab as a prophylactic treatment has led to a reduction in bleed frequency and a significant improvement in overall quality of life. No research to date has examined the nature of this improvement on treated individuals and their families. AIMS: The Emi & Me study aims to capture the real-life experience of using emicizumab for PwHi and their families. METHODS: Participants were recruited through treatment centres, social media and by word of mouth. Each participant and a family member, if available, took part in a semistructured qualitative interview. All interviews were recorded, transcribed verbatim and analysed thematically. All elements of the study were reviewed by local statutory authorities and informed consent was sought from all participants. RESULTS: Fifteen PwHi, mean age 27.2 years (range 8-63 years), most with a family member, participated in a single qualitative interview online (n = 13), by telephone (n = 1) or in person (n = 1). Mean time on emicizumab was 2.26 years (range 1-5 years). Six major themes emerged: bleeds; pain; treatment burden; control; freedom (for both PwHi and family members) and missed potential. Emicizumab prophylaxis has delivered significant improvements in the lives of the participants. Despite these improvements, some participants felt that their pre-existing physical disabilities and the lack of physiotherapy provision had prevented them achieving similar improvements in their functional ability. CONCLUSION: This study shows that in reducing bleeds, pain and treatment burden, emicizumab had given PwHi greater control over their condition, allowing a sense of freedom they had not experienced with factor VIII or bypassing agent prophylaxis. However, for emicizumab to be truly effective, there is a need to ensure the continued availability and accessibility of robust multidisciplinary support services. Without this, it is unlikely that PwHi will realize the life-changing potential offered either by emicizumab or any other novel treatment approach. PATIENT OR PUBLIC CONTRIBUTION: A patient participant (who did not wish to be included as an author of the paper) was involved in the design of the study protocol and interview guide.
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Hemofilia A , Actividades Cotidianas , Adolescente , Adulto , Niño , Familia , Hemofilia A/complicaciones , Hemofilia A/tratamiento farmacológico , Hemorragia/complicaciones , Hemorragia/prevención & control , Humanos , Masculino , Persona de Mediana Edad , Calidad de Vida , Adulto JovenRESUMEN
INTRODUCTION: For many people with haemophilia (PwH) gene therapy offers a potential functional cure. However, some have stated that they do not wish to have gene therapy either now or in the future. AIM: This sub-study, part of the larger Exigency programme, assesses the attitudes, views and understanding of those who do not wish to undergo gene therapy. METHODS: Participants were approached via social media and word of mouth referral and invited to participate in a focus group or individual interview to discuss their views. Interviews were recorded, transcribed verbatim and analysed thematically. RESULTS: Ten adult men with severe haemophilia (eight haemophilia A and two haemophilia B), mean age 34.3 years, participated in a 1-h focus group (n = 9) or interview (n = 1). All were on prophylaxis. None reported significant treatment burden, and all had annual bleeding rates of less than five in the previous 12 months. Four major themes emerged: self-identity and its loss, lack of long-term safety and efficacy data, ongoing concerns about past viral infection, and lack of current treatment burden. CONCLUSION: There are many concerns about gene therapy, including eligibility, effectiveness and safety, which may result in individuals declining it as a therapy. These concerns may recede as more data are published. This study reveals a psychological dynamic around self-identity and belonging for PwH. The nature of this dynamic is poorly understood and needs exploration to facilitate support for those making decisions about gene therapy.
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Hemofilia A , Hemofilia B , Adulto , Toma de Decisiones , Terapia Genética , Hemofilia A/genética , Hemofilia A/terapia , Hemofilia B/genética , Hemofilia B/terapia , Hemorragia , Humanos , MasculinoRESUMEN
INTRODUCTION: Gene therapy is used in life-limiting conditions of childhood. While not a current therapeutic option for children with haemophilia, it may be considered in the future especially for those where access to treatment is limited. AIM: To assess the attitudes and opinions of parents of children with haemophilia about gene therapy as a potential future treatment, by understanding their awareness about gene therapy and what they need to know now and in the future; gauging levels of interest in gene therapy for their children; and exploring perceived current motivations and barriers. METHODS: A mixed methods study with an online questionnaire and in-depth qualitative interviews in focus groups which were analysed using thematic analysis. RESULTS: One hundred and fifty-eight participants commenced the online survey; 63 were fully completed (39%). 60 had heard of gene therapy but few (17/60 [28.3%]) felt they had a good understanding. 38/60 (63.3%) respondents did not know that gene therapy is not available for children. However, most held positive views: 53/60 (88.3%) saying they would consider it for their child. In the interviews, participants (N = 10, all mothers) discussed their awareness and understanding of gene therapy and opinions about it for children, including how this should be communicated to the child and parents. CONCLUSION: A coherent, community-wide strategy for communicating information and news about gene therapy should now be provided for children and families living with haemophilia. This should come primarily from trusted haemophilia nursing teams, who can give tailored, age-appropriate, factual advice.