RESUMEN
The well-characterized association between HLA-B*27:05 and protection against HIV disease progression has been linked to immunodominant HLA-B*27:05-restricted CD8+ T-cell responses toward the conserved Gag KK10 (residues 263 to 272) and polymerase (Pol) KY9 (residues 901 to 909) epitopes. We studied the impact of the 3 amino acid differences between HLA-B*27:05 and the closely related HLA-B*27:02 on the HIV-specific CD8+ T-cell response hierarchy and on immune control of HIV. Genetic epidemiological data indicate that both HLA-B*27:02 and HLA-B*27:05 are associated with slower disease progression and lower viral loads. The effect of HLA-B*27:02 appeared to be consistently stronger than that of HLA-B*27:05. In contrast to HLA-B*27:05, the immunodominant HIV-specific HLA-B*27:02-restricted CD8+ T-cell response is to a Nef epitope (residues 142 to 150 [VW9]), with Pol KY9 subdominant and Gag KK10 further subdominant. This selection was driven by structural differences in the F pocket, mediated by a polymorphism between these two HLA alleles at position 81. Analysis of autologous virus sequences showed that in HLA-B*27:02-positive subjects, all three of these CD8+ T-cell responses impose selection pressure on the virus, whereas in HLA-B*27:05-positive subjects, there is no Nef VW9-mediated selection pressure. These studies demonstrate that HLA-B*27:02 mediates protection against HIV disease progression that is at least as strong as or stronger than that mediated by HLA-B*27:05. In combination with the protective Gag KK10 and Pol KY9 CD8+ T-cell responses that dominate HIV-specific CD8+ T-cell activity in HLA-B*27:05-positive subjects, a Nef VW9-specific response is additionally present and immunodominant in HLA-B*27:02-positive subjects, mediated through a polymorphism at residue 81 in the F pocket, that contributes to selection pressure against HIV.IMPORTANCE CD8+ T cells play a central role in successful control of HIV infection and have the potential also to mediate the eradication of viral reservoirs of infection. The principal means by which protective HLA class I molecules, such as HLA-B*27:05 and HLA-B*57:01, slow HIV disease progression is believed to be via the particular HIV-specific CD8+ T cell responses restricted by those alleles. We focus here on HLA-B*27:05, one of the best-characterized protective HLA molecules, and the closely related HLA-B*27:02, which differs by only 3 amino acids and which has not been well studied in relation to control of HIV infection. We show that HLA-B*27:02 is also protective against HIV disease progression, but the CD8+ T-cell immunodominance hierarchy of HLA-B*27:02 differs strikingly from that of HLA-B*27:05. These findings indicate that the immunodominant HLA-B*27:02-restricted Nef response adds to protection mediated by the Gag and Pol specificities that dominate anti-HIV CD8+ T-cell activity in HLA-B*27:05-positive subjects.
Asunto(s)
Linfocitos T CD8-positivos/inmunología , Infecciones por VIH/inmunología , Antígeno HLA-B27/genética , Epítopos Inmunodominantes/inmunología , Productos del Gen gag del Virus de la Inmunodeficiencia Humana/genética , Productos del Gen nef del Virus de la Inmunodeficiencia Humana/genética , Genes MHC Clase I , Infecciones por VIH/virología , VIH-1 , Humanos , Carga ViralRESUMEN
Control of virus replication in HIV-1 infection is critical to delaying disease progression. While cellular immune responses are a key determinant of control, relatively little is known about the contribution of the infecting virus to this process. To gain insight into this interplay between virus and host in viral control, we conducted a detailed analysis of two heterosexual HIV-1 subtype A transmission pairs in which female recipients sharing three HLA class I alleles exhibited contrasting clinical outcomes: R880F controlled virus replication while R463F experienced high viral loads and rapid disease progression. Near full-length single genome amplification defined the infecting transmitted/founder (T/F) virus proteome and subsequent sequence evolution over the first year of infection for both acutely infected recipients. T/F virus replicative capacities were compared in vitro, while the development of the earliest cellular immune response was defined using autologous virus sequence-based peptides. The R880F T/F virus replicated significantly slower in vitro than that transmitted to R463F. While neutralizing antibody responses were similar in both subjects, during acute infection R880F mounted a broad T cell response, the most dominant components of which targeted epitopes from which escape was limited. In contrast, the primary HIV-specific T cell response in R463F was focused on just two epitopes, one of which rapidly escaped. This comprehensive study highlights both the importance of the contribution of the lower replication capacity of the transmitted/founder virus and an associated induction of a broad primary HIV-specific T cell response, which was not undermined by rapid epitope escape, to long-term viral control in HIV-1 infection. It underscores the importance of the earliest CD8 T cell response targeting regions of the virus proteome that cannot mutate without a high fitness cost, further emphasizing the need for vaccines that elicit a breadth of T cell responses to conserved viral epitopes.
Asunto(s)
Aptitud Genética , Infecciones por VIH/diagnóstico , VIH-1/genética , VIH-1/inmunología , Interacciones Huésped-Patógeno/inmunología , Linfocitos T/inmunología , Adulto , Secuencia de Aminoácidos , Epítopos de Linfocito T/genética , Epítopos de Linfocito T/inmunología , Evolución Molecular , Femenino , Células HEK293 , Infecciones por VIH/genética , Infecciones por VIH/inmunología , Infecciones por VIH/transmisión , Interacciones Huésped-Patógeno/genética , Humanos , Evasión Inmune/genética , Masculino , Datos de Secuencia Molecular , Filogenia , Pronóstico , Carga Viral/genética , Replicación Viral/genéticaRESUMEN
OBJECTIVE: Developing a vaccine that is cross-reactive between HCV genotypes requires data on T cell antigenic targets that extends beyond genotype-1. We characterised T cell immune responses against HCV genotype-3, the most common infecting genotype in the UK and Asia, and assessed within genotype and between genotype cross-reactivity. DESIGN: T cell targets were identified in 140 subjects with either acute, chronic or spontaneously resolved HCV genotype-3 infection using (1) overlapping peptides and (2) putative human leucocyte antigens (HLA)-class-I wild type and variant epitopes through the prior assessment of polymorphic HCV genomic sites associated with host HLA, in IFNγ-ELISpot assays. CD4+/CD8+ T cell subsets were defined and viral variability at T cell targets was determined through population analysis and viral sequencing. T cell cross-reactivity between genotype-1 and genotype-3 variants was assessed. RESULTS: In resolved genotype-3 infection, T cells preferentially targeted non-structural proteins at a high magnitude, whereas in chronic disease T cells were absent or skewed to target structural proteins. Additional responses to wild type but not variant HLA predicted peptides were defined. Major sequence viral variability was observed within genotype-3 and between genotypes 1 and 3 HCV at T cell targets in resolved infection and at dominant epitopes, with limited T cell cross-reactivity between viral variants. Overall 41 CD4/CD8+ genotype-3 T cell targets were identified with minimal overlap with those described for HCV genotype-1. CONCLUSIONS: HCV T cell specificity is distinct between genotypes with limited T cell cross-reactivity in resolved and chronic disease. Therefore, viral regions targeted in natural HCV infection may not serve as attractive targets for a vaccine that aims to protect against multiple HCV genotypes.
Asunto(s)
Genotipo , Hepacivirus/genética , Antígenos de la Hepatitis C/inmunología , Hepatitis C/virología , Especificidad del Receptor de Antígeno de Linfocitos T/genética , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Reacciones Cruzadas , Hepacivirus/inmunología , Hepatitis C/inmunología , Antígenos de la Hepatitis C/genética , Humanos , Especificidad del Receptor de Antígeno de Linfocitos T/inmunología , Vacunas contra Hepatitis Viral , Proteínas no Estructurales Virales/genética , Proteínas no Estructurales Virales/inmunologíaRESUMEN
The rapid and extensive spread of the human immunodeficiency virus (HIV) epidemic provides a rare opportunity to witness host-pathogen co-evolution involving humans. A focal point is the interaction between genes encoding human leukocyte antigen (HLA) and those encoding HIV proteins. HLA molecules present fragments (epitopes) of HIV proteins on the surface of infected cells to enable immune recognition and killing by CD8(+) T cells; particular HLA molecules, such as HLA-B*57, HLA-B*27 and HLA-B*51, are more likely to mediate successful control of HIV infection. Mutation within these epitopes can allow viral escape from CD8(+) T-cell recognition. Here we analysed viral sequences and HLA alleles from >2,800 subjects, drawn from 9 distinct study cohorts spanning 5 continents. Initial analysis of the HLA-B*51-restricted epitope, TAFTIPSI (reverse transcriptase residues 128-135), showed a strong correlation between the frequency of the escape mutation I135X and HLA-B*51 prevalence in the 9 study cohorts (P = 0.0001). Extending these analyses to incorporate other well-defined CD8(+) T-cell epitopes, including those restricted by HLA-B*57 and HLA-B*27, showed that the frequency of these epitope variants (n = 14) was consistently correlated with the prevalence of the restricting HLA allele in the different cohorts (together, P < 0.0001), demonstrating strong evidence of HIV adaptation to HLA at a population level. This process of viral adaptation may dismantle the well-established HLA associations with control of HIV infection that are linked to the availability of key epitopes, and highlights the challenge for a vaccine to keep pace with the changing immunological landscape presented by HIV.
Asunto(s)
VIH-1/inmunología , Antígenos HLA-B/inmunología , Leucocitos/inmunología , Alelos , Linfocitos T CD8-positivos/inmunología , Estudios de Cohortes , Epítopos de Linfocito T/química , Epítopos de Linfocito T/genética , Epítopos de Linfocito T/inmunología , Antígenos VIH/química , Antígenos VIH/genética , Antígenos VIH/inmunología , VIH-1/genética , VIH-1/fisiología , Antígenos HLA-B/genética , Humanos , Internacionalidad , Polimorfismo Genético , Productos del Gen gag del Virus de la Inmunodeficiencia Humana/química , Productos del Gen gag del Virus de la Inmunodeficiencia Humana/genética , Productos del Gen gag del Virus de la Inmunodeficiencia Humana/inmunologíaRESUMEN
Trials of human immunodeficiency virus type 1 (HIV) pre- and postexposure prophylaxis show promise. Here, we describe a novel strategy for deciphering mechanisms of prophylaxis failure that could improve therapeutic outcomes. A healthcare worker began antiretroviral prophylaxis immediately after a high-risk needlestick injury but nonetheless became viremic 11 weeks later. Single-genome sequencing of plasma viral RNA identified 15 drug susceptible transmitted/founder HIV genomes responsible for productive infection. Sequences emanating from these genomes exhibited extremely low diversity, suggesting virus sequestration as opposed to low-level replication as the cause of breakthrough infection. Identification of transmitted/founder viruses allows for genome-wide assessment of molecular mechanisms of prophylaxis failure.
Asunto(s)
Fármacos Anti-VIH/uso terapéutico , Genoma Viral , Infecciones por VIH/tratamiento farmacológico , VIH-1/efectos de los fármacos , VIH-1/genética , Premedicación , Farmacorresistencia Viral/genética , Femenino , Infecciones por VIH/diagnóstico , Infecciones por VIH/prevención & control , VIH-1/clasificación , Humanos , Persona de Mediana Edad , Mutación , Filogenia , ARN Viral , Análisis de Secuencia de ADN , Insuficiencia del TratamientoRESUMEN
BACKGROUND: Following mucosal human immunodeficiency virus type 1 (HIV-1) transmission, type 1 interferons (IFNs) are rapidly induced at sites of initial virus replication in the mucosa and draining lymph nodes. However, the role played by IFN-stimulated antiviral activity in restricting HIV-1 replication during the initial stages of infection is not clear. We hypothesized that if type 1 IFNs exert selective pressure on HIV-1 replication in the earliest stages of infection, the founder viruses that succeed in establishing systemic infection would be more IFN-resistant than viruses replicating during chronic infection, when type 1 IFNs are produced at much lower levels. To address this hypothesis, the relative resistance of virus isolates derived from HIV-1-infected individuals during acute and chronic infection to control by type 1 IFNs was analysed. RESULTS: The replication of plasma virus isolates generated from subjects acutely infected with HIV-1 and molecularly cloned founder HIV-1 strains could be reduced but not fully suppressed by type 1 IFNs in vitro. The mean IC50 value for IFNα2 (22 U/ml) was lower than that for IFNß (346 U/ml), although at maximally-inhibitory concentrations both IFN subtypes inhibited virus replication to similar extents. Individual virus isolates exhibited differential susceptibility to inhibition by IFNα2 and IFNß, likely reflecting variation in resistance to differentially up-regulated IFN-stimulated genes. Virus isolates from subjects acutely infected with HIV-1 were significantly more resistant to in vitro control by IFNα than virus isolates generated from the same individuals during chronic, asymptomatic infection. Viral IFN resistance declined rapidly after the acute phase of infection: in five subjects, viruses derived from six-month consensus molecular clones were significantly more sensitive to the antiviral effects of IFNs than the corresponding founder viruses. CONCLUSIONS: The establishment of systemic HIV-1 infection by relatively IFNα-resistant founder viruses lends strong support to the hypothesis that IFNα plays an important role in the control of HIV-1 replication during the earliest stages of infection, prior to systemic viral spread. These findings suggest that it may be possible to harness the antiviral activity of type 1 IFNs in prophylactic and potentially also therapeutic strategies to combat HIV-1 infection.
Asunto(s)
Farmacorresistencia Viral , Infecciones por VIH/virología , VIH-1/efectos de los fármacos , VIH-1/inmunología , Interferón-alfa/inmunología , Interferón-alfa/farmacología , Replicación Viral/efectos de los fármacos , Infecciones por VIH/inmunología , VIH-1/aislamiento & purificación , Humanos , Concentración 50 Inhibidora , Pruebas de Sensibilidad MicrobianaRESUMEN
Hepatitis B virus (HBV)-specific T-cell responses are important in the natural history of HBV infection. The number of known HBV-specific T-cell epitopes is limited, and it is not clear whether viral evolution occurs in chronic HBV infection. We aimed to identify novel HBV T-cell epitopes by examining the relationship between HBV sequence variation and the human leukocyte antigen (HLA) type in a large prospective clinic-based cohort of Asian patients with chronic HBV infection recruited in Australia and China (n = 119). High-resolution 4-digit HLA class I and II typing and full-length HBV sequencing were undertaken for treatment-naïve individuals (52% with genotype B, 48% with genotype C, 63% HBV e antigen [HBeAg] positive). Statistically significant associations between HLA types and HBV sequence variation were identified (n = 49) at 41 sites in the HBV genome. Using prediction programs, we determined scores for binding between peptides containing these polymorphisms and associated HLA types. Among the regions that could be tested, HLA binding was predicted for 14/18 (78%). We identified several HLA-associated polymorphisms involving likely known anchor residues that resulted in altered predicted binding scores. Some HLA-associated polymorphisms fell within known T-cell epitopes with matching HLA restriction. Enhanced viral adaptation (defined as the presence of the relevant HLA and the escaped amino acid) was independently associated with HBeAg-negative disease (P = 0.003). Thus, HBV appears to be under immune pressure in chronic HBV infection, particularly in HBeAg-negative disease.
Asunto(s)
Antígenos HLA/genética , Antígenos HLA/inmunología , Antígenos e de la Hepatitis B/sangre , Virus de la Hepatitis B/fisiología , Hepatitis B Crónica/inmunología , Hepatitis B Crónica/virología , Adaptación Biológica , Secuencia de Aminoácidos , Australia , China , Epítopos de Linfocito T/genética , Epítopos de Linfocito T/inmunología , Femenino , Antígenos e de la Hepatitis B/genética , Antígenos e de la Hepatitis B/inmunología , Virus de la Hepatitis B/clasificación , Virus de la Hepatitis B/genética , Virus de la Hepatitis B/inmunología , Hepatitis B Crónica/genética , Interacciones Huésped-Patógeno , Humanos , Masculino , Datos de Secuencia Molecular , Filogenia , Polimorfismo Genético , Estudios ProspectivosRESUMEN
Human immunodeficiency virus (HIV)-1 amino acid sequence polymorphisms associated with expression of specific human histocompatibility leukocyte antigen (HLA) class I alleles suggest sites of cytotoxic T lymphocyte (CTL)-mediated selection pressure and immune escape. The associations most frequently observed are between expression of an HLA class I molecule and variation from the consensus sequence. However, a substantial number of sites have been identified in which particular HLA class I allele expression is associated with preservation of the consensus sequence. The mechanism behind this is so far unexplained. The current studies, focusing on two examples of "negatively associated" or apparently preserved epitopes, suggest an explanation for this phenomenon: negative associations can arise as a result of positive selection of an escape mutation, which is stable on transmission and therefore accumulates in the population to the point at which it defines the consensus sequence. Such negative associations may only be in evidence transiently, because the statistical power to detect them diminishes as the mutations accumulate. If an escape variant reaches fixation in the population, the epitope will be lost as a potential target to the immune system. These data help to explain how HIV is evolving at a population level. Understanding the direction of HIV evolution has important implications for vaccine development.
Asunto(s)
Epítopos de Linfocito T/inmunología , Infecciones por VIH/transmisión , VIH-1/inmunología , Antígenos HLA/inmunología , Polimorfismo Genético/inmunología , Linfocitos T Citotóxicos/inmunología , Vacunas contra el SIDA , Adulto , Alelos , Secuencia de Aminoácidos , Niño , Preescolar , Secuencia de Consenso/genética , Secuencia de Consenso/inmunología , Epítopos de Linfocito T/genética , Evolución Molecular , Femenino , Regulación de la Expresión Génica/genética , Regulación de la Expresión Génica/inmunología , Infecciones por VIH/genética , Infecciones por VIH/inmunología , VIH-1/genética , Antígenos HLA/genética , Humanos , Masculino , Datos de Secuencia Molecular , Mutación/genética , Mutación/inmunología , Polimorfismo Genético/genéticaRESUMEN
Mutations within cytotoxic T lymphocyte (CTL) epitopes impair T cell recognition, but escape mutations arising in flanking regions that alter antigen processing have not been defined in natural human infections. In human histocompatibility leukocyte antigen (HLA)-B57+ HIV-infected persons, immune selection pressure leads to a mutation from alanine to proline at Gag residue 146 immediately preceding the NH2 terminus of a dominant HLA-B57-restricted epitope, ISPRTLNAW. Although N-extended wild-type or mutant peptides remained well-recognized, mutant virus-infected CD4 T cells failed to be recognized by the same CTL clones. The A146P mutation prevented NH2-terminal trimming of the optimal epitope by the endoplasmic reticulum aminopeptidase I. These results demonstrate that allele-associated sequence variation within the flanking region of CTL epitopes can alter antigen processing. Identifying such mutations is of major relevance in the construction of vaccine sequences.
Asunto(s)
Presentación de Antígeno , Antígenos VIH/genética , Infecciones por VIH/inmunología , Infecciones por VIH/virología , VIH-1/genética , VIH-1/inmunología , Linfocitos T Citotóxicos/inmunología , Alelos , Secuencia de Aminoácidos , Secuencia de Bases , Células Clonales , ADN Viral/genética , Epítopos/genética , Productos del Gen gag/genética , Productos del Gen gag/inmunología , Variación Genética , Infecciones por VIH/genética , Antígenos HLA-B/genética , Humanos , Datos de Secuencia Molecular , Mutación , Homología de Secuencia de AminoácidoRESUMEN
Hepatitis C virus subtype 3a is a highly prevalent and globally distributed strain that is often associated with infection via injection drug use. This subtype exhibits particular phenotypic characteristics. In spite of this, detailed genetic analysis of this subtype has rarely been performed. We performed full-length viral sequence analysis in 18 patients with chronic HCV subtype 3a infection and assessed genomic viral variability in comparison to other HCV subtypes. Two novel regions of intragenotypic hypervariability within the envelope protein E2, of HCV genotype 3a, were identified. We named these regions HVR495 and HVR575. They consisted of flanking conserved hydrophobic amino acids and central variable residues. A 5-amino-acid insertion found only in genotype 3a and a putative glycosylation site is contained within HVR575. Evolutionary analysis of E2 showed that positively selected sites within genotype 3a infection were largely restricted to HVR1, HVR495, and HVR575. Further analysis of clonal viral populations within single hosts showed that viral variation within HVR495 and HVR575 were subject to intrahost positive selecting forces. Longitudinal analysis of four patients with acute HCV subtype 3a infection sampled at multiple time points showed that positively selected mutations within HVR495 and HVR575 arose early during primary infection. HVR495 and HVR575 were not present in HCV subtypes 1a, 1b, 2a, or 6a. Some variability that was not subject to positive selection was present in subtype 4a HVR575. Further defining the functional significance of these regions may have important implications for genotype 3a E2 virus-receptor interactions and for vaccine studies that aim to induce cross-reactive anti-E2 antibodies.
Asunto(s)
Hepacivirus/genética , Hepatitis C/virología , Secuencia de Aminoácidos , Variación Genética/genética , Genoma Viral/genética , Genotipo , Humanos , Datos de Secuencia Molecular , Filogenia , Selección GenéticaRESUMEN
UNLABELLED: Many hepatitis C virus (HCV) infections worldwide are with the genotype 1 and 3 strains of the virus. Cellular immune responses are known to be important in the containment of HCV genotype 1 infection, and many genotype 1 T cell targets (epitopes) that are presented by host human leukocyte antigens (HLAs) have been identified. In contrast, there is almost no information known about the equivalent responses to genotype 3. Immune escape mechanisms used by HCV include the evolution of viral polymorphisms (adaptations) that abrogate this host-viral interaction. Evidence of HCV adaptation to HLA-restricted immune pressure on HCV can be observed at the population level as viral polymorphisms associated with specific HLA types. To evaluate the escape patterns of HCV genotypes 1 and 3, we assessed the associations between viral polymorphisms and specific HLA types from 187 individuals with genotype 1a and 136 individuals with genotype 3a infection. We identified 51 HLA-associated viral polymorphisms (32 for genotype 1a and 19 for genotype 3a). Of these putative viral adaptation sites, six fell within previously published epitopes. Only two HLA-associated viral polymorphisms were common to both genotypes. In the remaining sites with HLA-associated polymorphisms, there was either complete conservation or no significant HLA association with viral polymorphism in the alternative genotype. This study also highlights the diverse mechanisms by which viral evasion of immune responses may be achieved and the role of genotype variation in these processes. CONCLUSION: There is little overlap in HLA-associated polymorphisms in the nonstructural proteins of HCV for the two genotypes, implying differences in the cellular immune pressures acting on these viruses and different escape profiles. These findings have implications for future therapeutic strategies to combat HCV infection, including vaccine design.
Asunto(s)
Adaptación Fisiológica/genética , Adaptación Fisiológica/inmunología , Antígenos HLA/inmunología , Hepacivirus/genética , Hepacivirus/inmunología , Adulto , Alelos , Australia , Estudios de Cohortes , ADN Viral/genética , Epítopos/genética , Femenino , Genotipo , Hepatitis C Crónica/inmunología , Humanos , Masculino , Persona de Mediana Edad , Polimorfismo Genético/genética , ARN Viral/genética , Suiza , Reino UnidoRESUMEN
UNLABELLED: The efficacy of specifically targeted anti-viral therapy for hepatitis C virus (HCV) (STAT-C), including HCV protease and polymerase inhibitors, is limited by the presence of drug-specific viral resistance mutations within the targeted proteins. Genetic diversity within these viral proteins also evolves under selective pressures provided by host human leukocyte antigen (HLA)-restricted immune responses, which may therefore influence STAT-C treatment response. Here, the prevalence of drug resistance mutations relevant to 27 developmental STAT-C drugs, and the potential for drug and immune selective pressures to intersect at sites along the HCV genome, is explored. HCV nonstructural (NS) 3 protease or NS5B polymerase sequences and HLA assignment were obtained from study populations from Australia, Switzerland, and the United Kingdom. Four hundred five treatment-naïve individuals with chronic HCV infection were considered (259 genotype 1, 146 genotype 3), of which 38.5% were coinfected with human immunodeficiency virus (HIV). We identified preexisting STAT-C drug resistance mutations in sequences from this large cohort. The frequency of the variations varied according to individual STAT-C drug and HCV genotype/subtype. Of individuals infected with subtype 1a, 21.5% exhibited genetic variation at a known drug resistance site. Furthermore, we identified areas in HCV protease and polymerase that are under both potential HLA-driven pressure and therapy selection and identified six HLA-associated polymorphisms (P Asunto(s)
Farmacorresistencia Viral/genética
, Hepacivirus/genética
, Hepatitis C Crónica/virología
, Proteínas no Estructurales Virales/genética
, Adaptación Biológica
, Aminoácidos/genética
, Antivirales/uso terapéutico
, Estudios de Cohortes
, Análisis Mutacional de ADN
, Variación Genética
, Genoma Viral
, Genotipo
, Infecciones por VIH/complicaciones
, Antígenos HLA/genética
, Hepatitis C Crónica/complicaciones
, Hepatitis C Crónica/tratamiento farmacológico
, Hepatitis C Crónica/inmunología
, Humanos
, Datos de Secuencia Molecular
, Selección Genética
RESUMEN
The extreme polymorphism in the human leukocyte antigen (HLA) class I region of the human genome is suggested to provide an advantage in pathogen defence mediated by CD8+ T cells. HLA class I molecules present pathogen-derived peptides on the surface of infected cells for recognition by CD8+ T cells. However, the relative contributions of HLA-A and -B alleles have not been evaluated. We performed a comprehensive analysis of the class I restricted CD8+ T-cell responses against human immunodeficiency virus (HIV-1), immune control of which is dependent upon virus-specific CD8+ T-cell activity. In 375 HIV-1-infected study subjects from southern Africa, a significantly greater number of CD8+ T-cell responses are HLA-B-restricted, compared to HLA-A (2.5-fold; P = 0.0033). Here we show that variation in viral set-point, in absolute CD4 count and, by inference, in rate of disease progression in the cohort, is strongly associated with particular HLA-B but not HLA-A allele expression (P < 0.0001 and P = 0.91, respectively). Moreover, substantially greater selection pressure is imposed on HIV-1 by HLA-B alleles than by HLA-A (4.4-fold, P = 0.0003). These data indicate that the principal focus of HIV-specific activity is at the HLA-B locus. Furthermore, HLA-B gene frequencies in the population are those likely to be most influenced by HIV disease, consistent with the observation that B alleles evolve more rapidly than A alleles. The dominant involvement of HLA-B in influencing HIV disease outcome is of specific relevance to the direction of HIV research and to vaccine design.
Asunto(s)
Evolución Biológica , Infecciones por VIH/inmunología , Infecciones por VIH/virología , VIH-1/inmunología , VIH-1/fisiología , Antígenos HLA-B/inmunología , África Austral , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/inmunología , Epítopos de Linfocito T/inmunología , Femenino , Frecuencia de los Genes , Productos del Gen nef/química , VIH-1/genética , Antígenos HLA-A/genética , Antígenos HLA-A/inmunología , Antígenos HLA-B/genética , Humanos , Lactante , Masculino , Polimorfismo Genético/genética , Carga Viral , Productos del Gen nef del Virus de la Inmunodeficiencia HumanaRESUMEN
Hepatitis C virus (HCV)-specific T cell responses are critical for immune control of infection. Viral adaptation to these responses, via mutations within regions of the virus targeted by CD8+ T cells, is associated with viral persistence. However, identifying viral adaptation to HCV-specific CD4+ T cell responses has been difficult although key to understanding anti-HCV immunity. In this context, HCV sequence and host genotype from a single source HCV genotype 1B cohort (n = 63) were analyzed to identify viral changes associated with specific human leucocyte antigen (HLA) class II alleles, as these variable host molecules determine the set of viral peptides presented to CD4+ T cells. Eight sites across the HCV genome were associated with HLA class II alleles implicated in infection outcome in this cohort (p ≤ 0.01; Fisher's exact test). We extended this analysis to chronic HCV infection (n = 351) for the common genotypes 1A and 3A. Variation at 38 sites across the HCV genome were associated with specific HLA class II alleles with no overlap between genotypes, suggestive of genotype-specific T cell targets, which has important implications for vaccine design. Here we show evidence of HCV adaptation to HLA class II-restricted CD4+ T cell pressure across the HCV genome in chronic HCV infection without a priori knowledge of CD4+ T cell epitopes.
Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Genoma Viral , Hepacivirus/genética , Hepatitis C Crónica/genética , Interacciones Huésped-Patógeno/genética , Proteínas no Estructurales Virales/genética , Alelos , Linfocitos T CD4-Positivos/virología , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/virología , Estudios de Cohortes , Epítopos de Linfocito T/genética , Epítopos de Linfocito T/inmunología , Regulación de la Expresión Génica , Genotipo , Cadenas beta de HLA-DQ/genética , Cadenas beta de HLA-DQ/inmunología , Cadenas HLA-DRB1/genética , Cadenas HLA-DRB1/inmunología , Hepacivirus/inmunología , Hepatitis C Crónica/inmunología , Hepatitis C Crónica/virología , Interacciones Huésped-Patógeno/inmunología , Humanos , Mutación , Proteínas no Estructurales Virales/inmunologíaRESUMEN
Host hepatitis C virus (HCV)-specific T cell responses and the ability of the virus to escape this response are important correlates of infection outcome. Understanding this host-viral interplay has been difficult given the often asymptomatic nature of acute HCV infection. We studied a recent transmission case to determine whether adapted viral strains can be transmitted and influence the recipient's anti-HCV T cell response. The diversity of viral populations was examined using next-generation sequencing, and HCV-specific T cell interferon (IFN)-γ responses were assessed using a peptide panel representing the autologous viruses. HCV-specific T cell responses in the source were directed against peptides that did not match the dominant autologous virus but rather low-frequency variants, implying existing viral adaptation in the source strain. Most HCV T cell epitopes that elicited an IFN-γ response in the source did not in the recipient, despite the pair sharing human leukocyte antigen alleles that govern antigen presentation and similar autologous viruses. Intrahost HCV variation in the recipient fell within predicted T cell epitopes, suggesting alternative targets of the immune response. These data suggest that transmission of adapted viral species can direct the host's HCV-specific immune response profile during acute infection.
Asunto(s)
Epítopos de Linfocito T/inmunología , Hepacivirus/inmunología , Hepatitis C Crónica/inmunología , Adaptación Fisiológica/inmunología , Australia , Ensayo de Immunospot Ligado a Enzimas , Epítopos de Linfocito T/análisis , Epítopos de Linfocito T/química , Evolución Molecular , Variación Genética/inmunología , Hepacivirus/genética , Hepatitis C/inmunología , Hepatitis C/virología , Hepatitis C Crónica/transmisión , Hepatitis C Crónica/virología , Secuenciación de Nucleótidos de Alto Rendimiento , Interacciones Huésped-Patógeno/inmunología , Humanos , Compartición de AgujasRESUMEN
Characterisation of Hepatitis C virus (HCV)-specific CD8+ T-cell responses in the context of multiple HCV exposures is critical to identify broadly protective immune responses necessary for an effective HCV vaccine against the different HCV genotypes. However, host and viral genetic diversity complicates vaccine development. To compensate for the observed variation in circulating autologous viruses and host molecules that restrict antigen presentation (human leucocyte antigens; HLA), this study used a reverse genomics approach that identified sites of viral adaptation to HLA-restricted T-cell immune pressure to predict genotype-specific HCV CD8+ T-cell targets. Peptides representing these putative HCV CD8+ T-cell targets, and their adapted form, were used in individualised IFN-γ ELISpot assays to screen for HCV-specific T-cell responses in 133 HCV-seropositive subjects with high-risk of multiple HCV exposures. The data obtained from this study i) confirmed that genetic studies of viral evolution is an effective approach to detect novel in vivo HCV T-cell targets, ii) showed that HCV-specific T-cell epitopes can be recognised in their adapted form and would not have been detected using wild-type peptides and iii) showed that HCV-specific T-cell (but not antibody) responses against alternate genotypes in chronic HCV-infected subjects are readily found, implying clearance of previous alternate genotype infection. In summary, HCV adaptation to HLA Class I-restricted T-cell responses plays a central role in anti-HCV immunity and multiple HCV genotype exposure is highly prevalent in at-risk exposure populations, which are important considerations for future vaccine design.
Asunto(s)
Linfocitos T CD8-positivos/inmunología , Hepacivirus , Hepatitis C Crónica/inmunología , Hepatitis C/inmunología , Alelos , Linfocitos T CD8-positivos/virología , Estudios de Cohortes , Ensayo de Immunospot Ligado a Enzimas , Epítopos de Linfocito T/química , Femenino , Genómica , Genotipo , Antígenos HLA/inmunología , Hepatitis C/virología , Hepatitis C Crónica/virología , Humanos , Masculino , Péptidos/químicaRESUMEN
Cellular immune responses during acute Hepatitis C virus (HCV) and HIV infection are a known correlate of infection outcome. Viral adaptation to these responses via mutation(s) within CD8+ T-cell epitopes allows these viruses to subvert host immune control. This study examined HCV evolution in 21 HCV genotype 1-infected subjects to characterise the level of viral adaptation during acute and early HCV infection. Of the total mutations observed 25% were within described CD8+ T-cell epitopes or at viral adaptation sites. Most mutations were maintained into the chronic phase of HCV infection (75%). The lack of reversion of adaptations and high proportion of silent substitutions suggests that HCV has structural and functional limitations that constrain evolution. These results were compared to the pattern of viral evolution observed in 98 subjects during a similar phase in HIV infection from a previous study. In contrast to HCV, evolution during acute HIV infection is marked by high levels of amino acid change relative to silent substitutions, including a higher proportion of adaptations, likely reflecting strong and continued CD8+ T-cell pressure combined with greater plasticity of the virus. Understanding viral escape dynamics for these two viruses is important for effective T cell vaccine design.
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Adaptación Fisiológica/genética , Evolución Molecular , Hepacivirus/genética , Hepacivirus/fisiología , Hepatitis C/virología , Enfermedad Aguda , Alelos , Epítopos/inmunología , Genotipo , Antígenos HLA/inmunología , Hepacivirus/patogenicidad , Hepatitis C/diagnóstico , Hepatitis C/inmunología , Hepatitis C Crónica/diagnóstico , Hepatitis C Crónica/inmunología , Hepatitis C Crónica/virología , Humanos , Mutación , Pronóstico , Linfocitos T/inmunología , Linfocitos T/virologíaRESUMEN
The enzyme linked immunospot (ELISpot) assay is a fundamental tool in cellular immunology, providing both quantitative and qualitative information on cellular cytokine responses to defined antigens. It enables the comprehensive screening of patient derived peripheral blood mononuclear cells to reveal the antigenic restriction of T-cell responses and is an emerging technique in clinical laboratory investigation of certain infectious diseases. As with all cellular-based assays, the final results of the assay are dependent on a number of technical variables that may impact precision if not highly standardised between operators. When studies that are large scale or using multiple antigens are set up manually, these assays may be labour intensive, have many manual handling steps, are subject to data and sample integrity failure and may show large inter-operator variability. Here we describe the successful automated performance of the interferon (IFN)-gamma ELISpot assay from cell counting through to electronic capture of cytokine quantitation and present the results of a comparison between automated and manual performance of the ELISpot assay. The mean number of spot forming units enumerated by both methods for limiting dilutions of CMV, EBV and influenza (CEF)-derived peptides in six healthy individuals were highly correlated (r>0.83, p<0.05). The precision results from the automated system compared favourably with the manual ELISpot and further ensured electronic tracking, increased through-put and reduced turnaround time.
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Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Ensayo de Inmunoadsorción Enzimática/métodos , Interferón gamma/inmunología , Robótica/métodos , Antígenos Virales/inmunología , Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD8-positivos/metabolismo , Humanos , Interferón gamma/biosíntesis , Péptidos/inmunologíaRESUMEN
The relative contributions of HLA alleles and T-cell receptors (TCRs) to the prevention of mutational viral escape are unclear. Here, we examined human immunodeficiency virus type 1 (HIV-1)-specific CD8(+) T-cell responses restricted by two closely related HLA class I alleles, B*5701 and B*5703, that differ by two amino acids but are both associated with a dominant response to the same HIV-1 Gag epitope KF11 (KAFSPEVIPMF). When this epitope is presented by HLA-B*5701, it induces a TCR repertoire that is highly conserved among individuals, cross-recognizes viral epitope variants, and is rarely associated with mutational escape. In contrast, KF11 presented by HLA-B*5703 induces an entirely different, more heterogeneous TCR beta-chain repertoire that fails to recognize specific KF11 escape variants which frequently arise in clade C-infected HLA-B*5703(+) individuals. These data show the influence of HLA allele subtypes on TCR selection and indicate that extensive TCR diversity is not a prerequisite to prevention of allowable viral mutations.
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Infecciones por VIH/inmunología , VIH-1/genética , VIH-1/inmunología , Antígenos de Histocompatibilidad Clase I/genética , Receptores de Antígenos de Linfocitos T/genética , Alelos , Sustitución de Aminoácidos , Linfocitos T CD8-positivos/inmunología , Epítopos de Linfocito T/genética , Productos del Gen gag/inmunología , Antígenos de Histocompatibilidad Clase I/inmunología , Humanos , Epítopos Inmunodominantes/genética , Mutación , Especificidad de la EspecieRESUMEN
Cellular immune responses are an important correlate of hepatitis C virus (HCV) infection outcome. These responses are governed by the host's human leukocyte antigen (HLA) type, and HLA-restricted viral escape mutants are a critical aspect of this host-virus interaction. We examined the driving forces of HCV evolution by characterizing the in vivo selective pressure(s) exerted on single amino acid residues within nonstructural protein 3 (NS3) by the HLA types present in two host populations. Associations between polymorphisms within NS3 and HLA class I alleles were assessed in 118 individuals from Western Australia and Switzerland with chronic hepatitis C infection, of whom 82 (69%) were coinfected with human immunodeficiency virus. The levels and locations of amino acid polymorphisms exhibited within NS3 were remarkably similar between the two cohorts and revealed regions under functional constraint and selective pressures. We identified specific HCV mutations within and flanking published epitopes with the correct HLA restriction and predicted escaped amino acid. Additional HLA-restricted mutations were identified that mark putative epitopes targeted by cell-mediated immune responses. This analysis of host-virus interaction reveals evidence of HCV adaptation to HLA class I-restricted immune pressure and identifies in vivo targets of cellular immune responses at the population level.