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BACKGROUND: Understanding how SARS-CoV-2 breakthrough infections impacts the breadth of immune responses against existing and pre-emergent SARS-CoV-2 strains is needed to develop an evidence-based long-term immunisation strategy. METHODS: We performed a randomised, controlled trial to assess the immunogenicity of homologous (BNT162b2) versus heterologous (mRNA-1273) booster vaccination in 100 BNT162b2-vaccinated infection-naïve individuals enrolled from October 2021. Post hoc analysis was performed to assess the impact of SARS-CoV-2 infection on humoral and cellular immune responses against wild-type SARS-CoV-2 and/or Omicron subvariants. FINDINGS: 93 participants completed the study at day 360. 71% (66/93) of participants reported first SARS-CoV-2 Omicron infection by the end of the study with similar proportions of infections between homologous and heterologous booster groups (72.3% [34/47] vs 69.6% [32/46]; p = 0.82). Mean wildtype SARS-CoV-2 anti-S-RBD antibody level was significantly higher in heterologous booster group compared with homologous group at day 180 (14,588 IU/mL; 95% CI, 10,186-20,893 vs 7447 IU/mL; 4646-11,912; p = 0.025). Participants who experienced breakthrough infections during the Omicron BA.1/2 wave had significantly higher anti-S-RBD antibody levels against wildtype SARS-CoV-2 and antibody neutralisation against BA.1 and pre-emergent BA.5 compared with infection-naïve participants. Regardless of hybrid immunity status, wildtype SARS-CoV-2 anti-S-RBD antibody level declined significantly after six months post-booster or post-SARS-CoV-2 infection. INTERPRETATION: Booster vaccination with mRNA-1273 was associated with significantly higher antibody levels compared with BNT162b2. Antibody responses are narrower and decline faster among uninfected, vaccinated individuals. Boosters may be more effective if administered shortly before infection outbreaks and at least six months after last infection or booster. FUNDING: Singapore NMRC, USFDA, MRC.
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Context: HNF4A-maturity-onset diabetes of the young (MODY1) is a relatively rare subtype of monogenic diabetes caused by loss of function of the HNF4A gene, which encodes the transcription factor HNF4α. HNF4α is known to form heterodimers, and the various combinations of isoforms that make up these heterodimers have been reported to result in a diversity of targeted genes. However, the function of individual HNF4α variant isoforms and the heterodimers comprising both wild-type (WT) and variant HNF4α have not yet been assessed. Objective: In this study, we analyzed the functional consequence of the HNF4A D248Y variant in vitro. Methods: We investigated the case of a 12-year-old Japanese girl who developed diabetes at age 11 years. Genetic sequencing detected a novel heterozygous missense HNF4A variant (c.742G > T, p.Asp248Tyr; referred as "D248Y") in the patient and her relatives who presented with diabetes. Results: Although the WT HNF4α isoforms (HNF4α2, HNF4α3, HNF4α8, HNF4α9) enhanced the INS gene promoter activity in HepG2 cells, the promoter activity of D248Y was consistently low across all isoforms. The presence of D248Y in homodimers and heterodimers, comprising either HNF4α8 or HNF4α3 or a combination of both isoforms, also reduced the INS promoter activity in Panc-1 cells. Conclusion: We report the clinical course of a patient with HNF4A-MODY and the functional analysis of novel HNF4A variants, with a focus on the isoforms and heterodimers they form. Our results serve to improve the understanding of the dominant-negative effects of pathogenic HNF4A variants.
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Chronic infection with hepatitis B virus (HBV) is incurable, as the current therapeutics cannot eliminate its persistent genomic material, cccDNA. Screening systems for cccDNA-targeting therapeutics are unavailable, as low copies of cccDNA in vitro complicate detection. To address this, cccDNA copies were massively increased to levels detectable via automated plate readers. This was achieved via continuous infection in a contact-free co-culture of an HBV generator (clone F881), which stably produced clinically relevant amounts of HBV, and HBV acceptors selected to carry high cccDNA loads. cccDNA-targeted therapeutics were then identified via reduced cccDNA-specific fluorescence, taking differences in the cell numbers and viability into account. Amongst the drugs tested, the H1 antihistamine Bilastine, HBVCP inhibitors and, surprisingly, current HBV therapeutics downregulated the cccDNA significantly, reflecting the assay's accuracy and sensitivity in identifying drugs that induce subtle changes in cccDNA levels, which take years to manifest in vivo. Bilastine was the only therapeutic that did not reduce HBV production from F881, indicating it to be a novel direct suppressor of cccDNA levels. When further assessed, only the structurally similar antihistamines Pitolisant and Nizatidine suppressed cccDNA levels when other H1 antihistamines could not. Taken together, our rapid fluorescence cccDNA-targeted drug screen successfully identified a class of molecules with the potential to treat hepatitis B.
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Vírus da Hepatite B , Hepatite B , Humanos , Vírus da Hepatite B/genética , Replicação Viral/genética , DNA Viral/genética , Antagonistas dos Receptores Histamínicos/farmacologia , Antagonistas dos Receptores Histamínicos/uso terapêuticoRESUMO
BACKGROUND: Emergence of the SARS-CoV-2 omicron (B.1.1.529) variant with high immune evasion has led to the development and roll-out of bivalent mRNA vaccines targeting original and omicron strains. However, real-world observational data on effectiveness of bivalent vaccines are scarce. We aimed to assess the relative effectiveness of a fourth vaccine dose with the BA.1-adapted or BA.4/BA.5-adapted bivalent vaccines against medically attended symptomatic SARS-CoV-2 infection and COVID-19-related hospital admission among SARS-CoV-2-naive and previously infected individuals in Singapore. METHODS: We conducted a retrospective cohort study among Singapore residents aged 18 years and older who had received three monovalent mRNA vaccine doses and were eligible for a fourth dose. Data were collected from official databases on COVID-19 cases and vaccinations maintained by the Singapore Ministry of Health. We analysed the incidence of medically attended symptomatic SARS-CoV-2 infection and COVID-19-related hospital admission between Oct 14, 2022, and Jan 31, 2023, by previous infection status and type of fourth vaccine dose received. Inverse probability-weighted Cox regressions were used to estimate hazard ratios (HRs). FINDINGS: 2 749 819 individuals were included in the analysis. For the SARS-CoV-2-naive group, a fourth monovalent vaccine dose did not confer additional protection over three monovalent doses against symptomatic infection (HR 1·09 [95% CI 1·07-1·11]), whereas the bivalent vaccine did provide additional protection (0·18 [0·17-0·19]). Among individuals with previous infection, the HR was 0·87 (95% CI 0·84-0·91) and 0·14 (0·13-0·15) with receipt of the fourth monovalent and bivalent doses, respectively. Against COVID-19-related hospital admission, the bivalent vaccine (HR 0·12 [95% CI 0·08-0·18] in SARS-CoV-2-naive participants and 0·04 [0·01-0·15] in previously infected participants) conferred greater benefit compared with the fourth monovalent dose (0·84 [0·77-0·91] in SARS-CoV-2-naive participants and 0·85 [0·69-1·04] in previously infected participants). INTERPRETATION: A fourth dose with the bivalent vaccine was substantially more effective against medically attended symptomatic SARS-CoV-2 infection and COVID-19-related hospital admission than four monovalent doses among both SARS-CoV-2-naive and previously infected individuals. Boosters with the bivalent vaccine might be preferred in this omicron-predominant pandemic, regardless of previous infection history. FUNDING: None.
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COVID-19 , SARS-CoV-2 , Humanos , COVID-19/prevenção & controle , Hospitais , Vacinas de mRNA , Estudos Retrospectivos , SARS-CoV-2/genética , Vacinas Combinadas , Adolescente , AdultoRESUMO
Allopurinol, widely used in gout treatment, is the most common cause of severe cutaneous adverse drug reactions. The risk of developing such life-threatening reactions is increased particularly for HLA-B*58:01 positive individuals. However the mechanism of action between allopurinol and HLA remains unknown. We demonstrate here that a Lamin A/C peptide KAGQVVTI which is unable to bind HLA-B*58:01 on its own, is enabled to form a stable peptide-HLA complex only in the presence of allopurinol. Crystal structure analysis reveal that allopurinol non-covalently facilitated KAGQVVTI to adopt an unusual binding conformation, whereby the C-terminal isoleucine does not engage as a PΩ that typically fit deeply in the binding F-pocket. A similar observation, though to a lesser degree was seen with oxypurinol. Presentation of unconventional peptides by HLA-B*58:01 aided by allopurinol contributes to our fundamental understanding of drug-HLA interactions. The binding of peptides from endogenously available proteins such as self-protein lamin A/C and viral protein EBNA3B suggest that aberrant loading of unconventional peptides in the presence of allopurinol or oxypurinol may be able to trigger anti-self reactions that can lead to Stevens-Johnson syndrome/toxic epidermal necrolysis (SJS/TEN) and Drug Reaction with Eosinophilia and Systemic Symptoms (DRESS).
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Alopurinol , Síndrome de Stevens-Johnson , Humanos , Alopurinol/farmacologia , Lamina Tipo A , Oxipurinol , Genótipo , Síndrome de Stevens-Johnson/etiologia , Antígenos HLA-B/genética , PeptídeosRESUMO
Waning antibody levels against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the emergence of variants of concern highlight the need for booster vaccinations. This is particularly important for the elderly population, who are at a higher risk of developing severe coronavirus disease 2019 (COVID-19) disease. While studies have shown increased antibody responses following booster vaccination, understanding the changes in T and B cell compartments induced by a third vaccine dose remains limited. We analyzed the humoral and cellular responses in subjects who received either a homologous messenger RNA(mRNA) booster vaccine (BNT162b2 + BNT162b2 + BNT162b2; ''BBB") or a heterologous mRNA booster vaccine (BNT162b2 + BNT162b2 + mRNA-1273; ''BBM") at Day 0 (prebooster), Day 7, and Day 28 (postbooster). Compared with BBB, elderly individuals (≥60 years old) who received the BBM vaccination regimen display higher levels of neutralizing antibodies against the Wuhan and Delta strains along with a higher boost in immunoglobulin G memory B cells, particularly against the Omicron variant. Circulating T helper type 1(Th1), Th2, Th17, and T follicular helper responses were also increased in elderly individuals given the BBM regimen. While mRNA vaccines increase antibody, T cell, and B cell responses against SARS-CoV-2 1 month after receiving the third dose booster, the efficacy of the booster vaccine strategies may vary depending on age group and regimen combination.
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COVID-19 , SARS-CoV-2 , Idoso , Humanos , Pessoa de Meia-Idade , SARS-CoV-2/genética , Vacina BNT162 , COVID-19/prevenção & controle , Vacinas de mRNA , Anticorpos Neutralizantes , Anticorpos Antivirais , VacinaçãoRESUMO
The emergence of new SARS-CoV-2 variants, such as the more transmissible Delta and Omicron variants, has raised concerns on efficacy of the COVID-19 vaccines. Here, we examined the waning of antibody responses against different variants following primary and booster vaccination. We found that antibody responses against variants were low following primary vaccination. The antibody response against Omicron was almost non-existent. Efficient boosting of antibody response against all variants, including Omicron, was observed following a third dose. The antibody response against the variants tested was significantly higher at one month following booster vaccination, compared with two months following primary vaccination, for all individuals, including the low antibody responders identified at two months following primary vaccination. The antibody response, for all variants tested, was significantly higher at four months post booster than at five months post primary vaccination, and the proportion of low responders remained low (6-11%). However, there was significant waning of antibody response in more than 95% of individuals at four months, compared to one month following booster. We also observed a robust memory B cell response following booster, which remained higher at four months post booster than prior to booster. However, the memory B cell responses were on the decline for 50% of individuals at four months following booster. Similarly, while the T cell response is sustained, at cohort level, at four months post booster, a substantial proportion of individuals (18.8 - 53.8%) exhibited T cell response at four months post booster that has waned to levels below their corresponding levels before booster. The findings show an efficient induction of immune response against SARS-CoV-2 variants following booster vaccination. However, the induced immunity by the third BNT162b2 vaccine dose was transient. The findings suggest that elderly individuals may require a fourth dose to provide protection against SARS-CoV-2.
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Vacinas contra COVID-19 , COVID-19 , Idoso , Humanos , Vacina BNT162 , SARS-CoV-2 , COVID-19/prevenção & controle , AnticorposRESUMO
The Asian seabass is of importance both as a farmed and wild animal. With the emergence of infectious diseases, there is a need to understand and characterize the immune system. In humans, the highly polymorphic MHC class I (MHC-I) molecules play an important role in antigen presentation for the adaptive immune system. In the present study, we characterized a single MHC-I gene in Asian seabass (Lates calcarifer) by amplifying and sequencing the MHC-I alpha 1 and alpha 2 domains, followed by multi-sequence alignment analyses. The results indicated that the Asian seabass MHC-I α1 and α2 domain sequences showed an overall similarity within Asian seabass and retained the majority of the conserved binding residues of human leukocyte antigen-A2 (HLA-A2). Phylogenetic tree analysis revealed that the sequences belonged to the U lineage. Mapping the conserved binding residue positions on human HLA-A2 and grass carp crystal structure showed a high degree of similarity. In conclusion, the availability of MHC-I α1 and α2 sequences enhances the quality of MHC class I genetic information in Asian seabass, providing new tools to analyze fish immune responses to pathogen infections, and will be applicable in the study of the phylogeny and the evolution of antigen-specific receptors.
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Bass , Perciformes , Animais , Bass/genética , Peixes , Antígeno HLA-A2/genética , Humanos , Perciformes/genética , FilogeniaRESUMO
Objective: Despite the high vaccine efficacy of mRNA COVID-19 vaccines, there are individuals who developed excessive reactogenic and/or allergic responses after the first mRNA dose and were considered ineligible for further mRNA doses. CoronaVac, an inactivated SARS-CoV-2 vaccine, is recommended in Singapore as an alternative. Methods: Individuals, ineligible for further mRNA vaccines (BNT162b2 or mRNA-1273) because of excessive reactive responses to prime mRNA vaccination, were recruited and offered two doses of CoronaVac as booster vaccination 38-224 days post their mRNA vaccine dose. Individuals who did not develop any excessive reactive responses after the prime mRNA vaccination were also recruited and given another mRNA vaccine as booster vaccination. Blood samples were collected at days 0, 21 and 90 post first CoronaVac dose and mRNA dose, respectively, for analysis. Results: We showed that two CoronaVac booster doses induced specific immunity in these mRNA vaccine-primed individuals. Although the spike-specific antibody response was lower, their memory B cell response against the receptor-binding domain (RBD) of the spike protein was similar, compared with individuals who received two BNT162b2 injections. The spike-specific memory T cell response also increased following CoronaVac booster doses. However, specific immunity against the Omicron variant was low, similar to individuals with two BNT162b2 doses. Conclusion: Our findings showed that while mRNA vaccine-primed individuals can opt for two subsequent doses of CoronaVac, an additional dose may be necessary to achieve protection, especially against newly emerging immune escape variants such as Omicron.
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Understanding the impact of age on vaccinations is essential for the design and delivery of vaccines against SARS-CoV-2. Here, we present findings from a comprehensive analysis of multiple compartments of the memory immune response in 312 individuals vaccinated with the BNT162b2 SARS-CoV-2 mRNA vaccine. Two vaccine doses induce high antibody and T cell responses in most individuals. However, antibody recognition of the Spike protein of the Delta and Omicron variants is less efficient than that of the ancestral Wuhan strain. Age-stratified analyses identify a group of low antibody responders where individuals ≥60 years are overrepresented. Waning of the antibody and cellular responses is observed in 30% of the vaccinees after 6 months. However, age does not influence the waning of these responses. Taken together, while individuals ≥60 years old take longer to acquire vaccine-induced immunity, they develop more sustained acquired immunity at 6 months post-vaccination. A third dose strongly boosts the low antibody responses in the older individuals against the ancestral Wuhan strain, Delta and Omicron variants.
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COVID-19 , Vacinas Virais , Idoso , Anticorpos Antivirais , Formação de Anticorpos , Vacina BNT162 , COVID-19/prevenção & controle , Vacinas contra COVID-19 , Humanos , Pessoa de Meia-Idade , SARS-CoV-2 , Vacinação , Vacinas Sintéticas , Vacinas de mRNARESUMO
BACKGROUND: Over 2021, COVID-19 vaccination programs worldwide focused on raising population immunity through the primary COVID-19 vaccine series. In Singapore, two mRNA vaccines (BNT162b2 and mRNA-1273) and the inactivated vaccine CoronaVac are currently authorized under the National Vaccination Programme for use as the primary vaccination series. More than 90% of the Singapore population has received at least one dose of a COVID-19 vaccine as of December 2021. With the demonstration that vaccine effectiveness wanes in the months after vaccination, and the emergence of Omicron which evades host immunity from prior infection and/or vaccination, attention in many countries has shifted to how best to maintain immunity through booster vaccinations. METHODS: The objectives of this phase 3, randomized, subject-blinded, controlled clinical trial are to assess the safety and immunogenicity of heterologous boost COVID-19 vaccine regimens (intervention groups 1-4) compared with a homologous boost regimen (control arm) in up to 600 adult volunteers. As non-mRNA vaccine candidates may enter the study at different time points depending on vaccine availability and local regulatory approval, participants will be randomized at equal probability to the available intervention arms at the time of randomization. Eligible participants will have received two doses of a homologous mRNA vaccine series with BNT162b2 or mRNA-1273 at least 6 months prior to enrolment. Participants will be excluded if they have a history of confirmed SARS or SARS-CoV-2 infection, are immunocompromised, or are pregnant. Participants will be monitored for adverse events and serious adverse events by physical examinations, laboratory tests and self-reporting. Blood samples will be collected at serial time points [pre-vaccination/screening (day - 14 to day 0), day 7, day 28, day 180, day 360 post-vaccination] for assessment of antibody and cellular immune parameters. Primary endpoint is the level of anti-SARS-CoV-2 spike immunoglobulins at day 28 post-booster and will be measured against wildtype SARS-CoV-2 and variants of concern. Comprehensive immune profiling of the humoral and cellular immune response to vaccination will be performed. DISCUSSION: This study will provide necessary data to understand the quantity, quality, and persistence of the immune response to a homologous and heterologous third booster dose of COVID-19 vaccines. This is an important step in developing COVID-19 vaccination programs beyond the primary series. TRIAL REGISTRATION: ClinicalTrials.gov NCT05142319 . Registered on 2 Dec 2021.
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Vacinas contra COVID-19 , COVID-19 , Adulto , Anticorpos Antivirais , Vacina BNT162 , COVID-19/prevenção & controle , Vacinas contra COVID-19/efeitos adversos , Ensaios Clínicos Fase III como Assunto , Humanos , Ensaios Clínicos Controlados Aleatórios como Assunto , SARS-CoV-2 , Vacinas Sintéticas , Vacinas de mRNARESUMO
BACKGROUND: Waning antibody levels post-vaccination and the emergence of variants of concern (VOCs) capable of evading protective immunity have raised the need for booster vaccinations. However, which combination of coronavirus disease 2019 (COVID-19) vaccines offers the strongest immune response against the Omicron variant is unknown. METHODS: This randomized, participant-blinded, controlled trial assessed the reactogenicity and immunogenicity of different COVID-19 vaccine booster combinations. A total of 100 BNT162b2-vaccinated individuals were enrolled and randomized 1:1 to either homologous (BNT162b2 + BNT162b2 + BNT162b2; "BBB") or heterologous messenger RNA (mRNA) (BNT162b2 + BNT162b2 + mRNA-1273; "BBM") booster vaccine. The primary end point was the level of neutralizing antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) wild-type and VOCs at day 28. RESULTS: A total of 51 participants were allocated to BBB and 49 to BBM; 50 and 48, respectively, were analyzed for safety and immunogenicity outcomes. At day 28 post-boost, mean SARS-CoV-2 spike antibody titers were lower with BBB (22 382â IU/mL; 95% confidence interval [CI], 18 210 to 27 517) vs BBM (29 751â IU/mL; 95% CI, 25 281 to 35 011; P = .034) as was the median level of neutralizing antibodies: BBB 99.0% (interquartile range [IQR], 97.9% to 99.3%) vs BBM 99.3% (IQR, 98.8% to 99.5%; P = .021). On subgroup analysis, significant higher mean spike antibody titer, median surrogate neutralizing antibody level against all VOCs, and live Omicron neutralization titer were observed only in older adults receiving BBM. Both vaccines were well tolerated. CONCLUSIONS: Heterologous mRNA-1273 booster vaccination compared with homologous BNT123b2 induced a stronger neutralizing response against the Omicron variant in older individuals. CLINICAL TRIALS REGISTRATION: NCT05142319.
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Vacina BNT162 , COVID-19 , Humanos , Idoso , SARS-CoV-2 , Formação de Anticorpos , Vacina de mRNA-1273 contra 2019-nCoV , Vacinação , Anticorpos Neutralizantes , Anticorpos AntiviraisRESUMO
Infection with the hepatitis B virus (HBV) is still a major global health threat as 250 million people worldwide continue to be chronically infected with the virus. While patients may be treated with nucleoside/nucleotide analogues, this only suppresses HBV titre to sub-detection levels without eliminating the persistent HBV covalently closed circular DNA (cccDNA) genome. As a result, HBV infection cannot be cured, and the virus reactivates when conditions are favorable. Interferons (IFNs) are cytokines known to induce powerful antiviral mechanisms that clear viruses from infected cells. They have been shown to induce cccDNA clearance, but their use in the treatment of HBV infection is limited as HBV-targeting immune cells are exhausted and HBV has evolved multiple mechanisms to evade and suppress IFN signalling. Thus, to fully utilize IFN-mediated intracellular mechanisms to effectively eliminate HBV, instead of direct IFN administration, novel strategies to sustain IFN-mediated anti-cccDNA and antiviral mechanisms need to be developed. This review will consolidate what is known about how IFNs act to achieve its intracellular antiviral effects and highlight the critical interferon-stimulated gene targets and effector mechanisms with potent anti-cccDNA functions. These include cccDNA degradation by APOBECs and cccDNA silencing and transcription repression by epigenetic modifications. In addition, the mechanisms that HBV employs to disrupt IFN signalling will be discussed. Drugs that have been developed or are in the pipeline for components of the IFN signalling pathway and HBV targets that detract IFN signalling mechanisms will also be identified and discussed for utility in the treatment of HBV infections. Together, these will provide useful insights into design strategies that specifically target cccDNA for the eradication of HBV.
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Hepatite B Crônica , Hepatite B , Antivirais/farmacologia , Antivirais/uso terapêutico , DNA Circular , DNA Viral/genética , DNA Viral/uso terapêutico , Vírus da Hepatite B/genética , Hepatite B Crônica/tratamento farmacológico , Humanos , Interferons/uso terapêutico , Replicação ViralRESUMO
The human cell line activation test (h-CLAT) is an OECD approved (Test No. 442E) assay to identify novel skin sensitizers. h-CLAT simulates dendritic cell activation in the skin sensitization pathway and is based on the measurement of CD54 and CD86 overexpression on monocytic, leukemic THP-1 cells. However, the current h-CLAT markers show inconsistent results with moderate and weak sensitizers. Moreover, these markers have accessory roles in cell adhesion and signaling rather than a direct role in cellular inflammation. Therefore, we have explored other inflammation-related markers in this study. PBMCs comprises a mixture of cells that resemble the complex immunological milieu in adults and were primarily used to identify markers. PBMCs (n = 10) and THP-1 cells were treated with 1-chloro-2,4-dinitrobenzene (DNCB, strong) and NiCl2 (Ni, moderate) sensitizers or DMSO (control) and incubated for 24 h. The samples were subjected to RNA sequencing to obtain log2fold change in gene expression. DNCB and NiCl2 significantly upregulated 80 genes in both cell types. Of these, CD109, CD181, CD183, CLEC5A, CLEC8A & CD354 were experimentally validated. DNCB and Ni but not isopropyl alcohol (non-sensitizer) significantly induced the expression of all novel markers except CLEC8A. Moreover, the percentage induction of all novel markers except CLEC8A satisfied the OECD acceptance criteria. In summary, we identified five novel markers that may supplement the current repertoire of h-CLAT markers.
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Alérgenos/toxicidade , Haptenos/toxicidade , Antígenos CD/genética , Biomarcadores , Sobrevivência Celular/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Lectinas Tipo C/genética , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Receptores de Superfície Celular/genética , Testes Cutâneos , Células THP-1RESUMO
The HLA-B*15:02 allele is associated with an increased risk of developing carbamazepine (CBZ)-induced Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN). Many studies, however, have demonstrated that a large majority of HLA-B*15:02 individuals are unlikely to develop the adverse drug reaction while on CBZ. This phenomenon suggests that other factors that modulate the allergic immune response, such as regulatory T cells (Tregs), might contribute to an uncontrolled immune response in SJS/TEN. Peripheral blood mononuclear cells (PBMCs) from 15 healthy HLA-B*15:02 carriers were isolated to investigate the role of Tregs in controlling the immune response towards CBZ. Recognition of CBZ was assessed using enzyme linked immunosorbent spot (ELISPOT) assay for IFN-γ, and the donor T-cell profiles were quantified by flow cytometry to differentiate CBZ responders from non-responders. As CD39 expression on Tregs promotes immune tolerance, we investigated the mechanisms of Treg suppression using inhibitors targeting the CD39/adenosinergic pathway. PBMCs from seven donors (responders) produced high levels of IFN-γ when re-exposed to CBZ, while eight donors (non-responders) did not. Flow cytometric analysis revealed that non-responders produced significantly higher frequencies of CD4+CD25+CD127loCD39+FoxP3+ Tregs compared to responders. CD39 inhibition using POM-1 inhibitor converted five of the eight non-responders into responders (P < 0.05). Higher frequencies of CD4+CD25+CD127loCD39+FoxP3+ Tregs was correlated with lower production of IFN-γ (P < 0.01). Our data suggest that CD4+CD25+CD127loCD39+FoxP3+ Tregs may play a role in promoting CBZ tolerance in HLA-B*15:02 carriers. The CD39/adenosinergic axis can be a potential target to alleviate the uncontrolled immune response during this adverse drug event.
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Hipersensibilidade a Drogas/imunologia , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/imunologia , Antígeno HLA-B15/genética , Linfócitos T Reguladores/imunologia , Adenosina/metabolismo , Alérgenos/imunologia , Antígenos CD/metabolismo , Apirase/metabolismo , Carbamazepina/imunologia , Estudos de Casos e Controles , Células Cultivadas , ELISPOT , Fatores de Transcrição Forkhead/metabolismo , Estudos de Associação Genética , Predisposição Genética para Doença , Antígeno HLA-B15/metabolismo , Humanos , Tolerância Imunológica/genética , Imunidade Celular , Transdução de SinaisRESUMO
Peptides presented by Human leukocyte antigen (HLA) class-I molecules are generally 8-10 amino acids in length. However, the predominant pool of peptide fragments generated by proteasomes is less than 8 amino acids in length. Using the Epstein - Barr virus (EBV) Rta-epitope (ATIGTAMYK, residues 134-142) restricted by HLA-A*11:01 which generates a strong immunodominant response, we investigated the minimum length of a viral peptide that can constitute a viral epitope recognition by CD8 T cells. The results showed that Peripheral blood mononuclear cells (PBMCs) from healthy donors can be stimulated by a viral peptide fragment as short as 4-mer (AMYK), together with a 5-mer (ATIGT) to recapitulate the full length EBV Rta epitope. This was confirmed by generating crystals of the tetra-complex (2 peptides, HLA and ß2-microglobulin). The solved crystal structure of HLA-A*11:01 in complex with these two short peptides revealed that they can bind in the same orientation similar to parental peptide (9-mer) and the free ends of two short peptides acquires a bulged conformation that is directed towards the T cell receptor. Our data shows that suboptimal length of 4-mer and 5-mer peptides can complement each other to form a stable peptide-MHC (pMHC) complex.
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Linfócitos T CD8-Positivos/imunologia , Infecções por Vírus Epstein-Barr/imunologia , Antígenos HLA-A/química , Herpesvirus Humano 4/imunologia , Proteínas Imediatamente Precoces/química , Leucócitos Mononucleares/imunologia , Fragmentos de Peptídeos/química , Transativadores/química , Epitopos de Linfócito T/química , Epitopos de Linfócito T/imunologia , Infecções por Vírus Epstein-Barr/virologia , Antígenos HLA-A/imunologia , Humanos , Proteínas Imediatamente Precoces/imunologia , Leucócitos Mononucleares/virologia , Fragmentos de Peptídeos/imunologia , Conformação Proteica , Linfócitos T Citotóxicos/imunologia , Transativadores/imunologiaRESUMO
INTRODUCTION: Psoriasis is a systemic, chronic inflammatory disease that not only afflicts the skin but is also associated with cardiovascular disease and metabolic syndrome. The strongest susceptibility loci for the disease is within the human leukocyte antigen (HLA) complex, though specific HLA allelic associations vary between populations. OBJECTIVE: Our objective was to investigate HLA associations with clinical phenotypes of psoriasis and metabolic syndrome in Chinese psoriasis cases. METHODS: We conducted an observational case-control study in Singapore with a cohort of psoriasis cases consecutively recruited from an outpatient specialist dermatological center (n = 120) compared with 130 healthy controls. RESULTS: Significant HLA associations with psoriasis were observed with HLA-A*02:07, B*46:01, C*01:02, and C*06:02. The three-locus haplotype of A*02:07-C*01:02-B*46:01 was also significant (odds ratio [OR] 3.07; p = 9.47 × 10-5). We also observed an association between nail psoriasis and HLA-A*02:07 carriers (OR 4.50; p = 0.002), whereas C*06:02 carriers were less prone to have nail involvement (OR 0.16; p = 0.004). HLA-A*02:07 was also identified as a possible risk allele for hypertension (OR 2.90; p < 0.05), and C*01:02 was a possible risk allele for dyslipidemia (OR 3.36; p < 0.05), both known to be common comorbidities in patients with psoriasis. CONCLUSION: Our results demonstrate the growing importance of discerning population-specific clinical phenotypes and their association with certain HLA alleles in psoriasis.