RESUMO
DNA vaccines require a considerable enhancement of immunogenicity. Here, we optimized a prototype DNA vaccine against drug-resistant HIV-1 based on a weak Th2-immunogen, HIV-1 reverse transcriptase (RT). We designed expression-optimized genes encoding inactivated wild-type and drug-resistant RTs (RT-DNAs) and introduced them into mice by intradermal injections followed by electroporation. RT-DNAs were administered as single or double primes with or without cyclic-di-GMP, or as a prime followed by boost with RT-DNA mixed with a luciferase-encoding plasmid ("surrogate challenge"). Repeated primes improved cellular responses and broadened epitope specificity. Addition of cyclic-di-GMP induced a transient increase in IFN-γ production. The strongest anti-RT immune response was achieved in a prime-boost protocol with electroporation by short 100V pulses done using penetrating electrodes. The RT-specific response, dominated by CD4+ T-cells, targeted epitopes at aa 199-220 and aa 528-543. Drug-resistance mutations disrupted the epitope at aa 205-220, while the CTL epitope at aa 202-210 was not affected. Overall, multiparametric optimization of RT strengthened its Th2- performance. A rapid loss of RT/luciferase-expressing cells in the surrogate challenge experiment revealed a lytic potential of anti-RT response. Such lytic CD4+ response would be beneficial for an HIV vaccine due to its comparative insensitivity to immune escape.
Assuntos
Vacinas contra a AIDS , Farmacorresistência Viral , Infecções por HIV/terapia , Transcriptase Reversa do HIV/imunologia , Células Th2/imunologia , Vacinação/métodos , Vacinas de DNA , Vacinas contra a AIDS/administração & dosagem , Vacinas contra a AIDS/genética , Animais , Calibragem , Células Cultivadas , Códon , Sistemas de Liberação de Medicamentos , Farmacorresistência Viral/genética , Farmacorresistência Viral/imunologia , Epitopos/genética , Epitopos/imunologia , Infecções por HIV/imunologia , Transcriptase Reversa do HIV/genética , HIV-1/genética , HIV-1/imunologia , Células HeLa , Humanos , Evasão da Resposta Imune/genética , Evasão da Resposta Imune/imunologia , Imunização Secundária/métodos , Imunização Secundária/normas , Imunogenicidade da Vacina/genética , Camundongos , Camundongos Endogâmicos BALB C , Melhoria de Qualidade , Células Th2/metabolismo , Vacinação/normas , Vacinas de DNA/administração & dosagem , Vacinas de DNA/genéticaRESUMO
BACKGROUND: Genetic immunization is expected to induce the expression of antigens in a native form. The encoded peptide epitopes are presented on endogenous MHC molecules, mimicking antigen presentation during a viral infection. We have explored the potential of enfuvirtide (T20), a short HIV peptide with antiviral properties, to enhance immune response to HIV antigens. To generate an expression vector, the T20 sequence was cloned into a conventional plasmid, the novel minicircle construct, and a replicon plasmid. In addition, 3 conventional plasmids that express the envelope of HIV-1 subtypes A, B and C and contain T20 in their gp41 sequences were also tested. RESULTS: All combinations induced HIV-specific antibodies and cellular responses. The addition of T20 as a peptide and as an expression cassette in the 3 DNA vectors enhanced antibody responses. The highest anti-HIV-1 Env titers were obtained by the replicon T20 construct. This demonstrates that besides its known antiviral activity, T20 promotes immune responses. We also confirm that the combination of slightly divergent antigens improves immune responses. CONCLUSIONS: The antiretroviral T20 HIV-1 sequence can be used as an immunogen to elicit binding and neutralizing antibodies against HIV-1. These, or similarly modified gp41 genes/peptides, can be used as priming or boosting components for induction of broadly neutralizing anti-HIV antibodies. Future comparative studies will reveal the optimal mode of T20 administration.
Assuntos
Vacinas contra a AIDS/imunologia , Anticorpos Neutralizantes/sangue , Reações Cruzadas , Anticorpos Anti-HIV/sangue , Proteína gp41 do Envelope de HIV/imunologia , HIV-1/imunologia , Fragmentos de Peptídeos/imunologia , Vacinas de DNA/imunologia , Vacinas contra a AIDS/administração & dosagem , Animais , Enfuvirtida , Feminino , Proteína gp41 do Envelope de HIV/genética , Camundongos Endogâmicos BALB C , Fragmentos de Peptídeos/genética , Vacinas de DNA/administração & dosagemRESUMO
BACKGROUND: In order to develop a more effective prophylactic HIV-1 vaccine it is important optimize the components, improve Envelope glycoprotein immunogenicity as well as to explore prime-boost immunization schedules. It is also valuable to include several HIV-1 subtype antigens representing the world-wide epidemic. METHODS: HIVIS-DNA plasmids which include Env genes of subtypes A, B and C together with Gag subtypes A and B and RTmut/Rev of subtype B were modified as follows: the Envelope sequences were shortened, codon optimized, provided with an FT4 sequence and an immunodominant region mutated. The reverse transcriptase (RT) gene was shortened to contain the most immunogenic N-terminal fragment and fused with an inactivated viral protease vPR gene. HIVISopt-DNA thus contains fewer plasmids but additional PR epitopes compared to the native HIVIS-DNA. DNA components were delivered intradermally to young Balb/c mice once, using a needle-free Biojector® immediately followed by dermal electroporation. Vaccinia-based MVA-CMDR boosts including Env gene E and Gag-RT genes A were delivered intramuscularly by needle, once or twice. RESULTS: Both HIVIS-DNA and HIVISopt-DNA primed humoral and cell mediated responses well. When boosted with heterologous MVA-CMDR (subtypes A and E) virus inhibitory neutralizing antibodies were obtained to HIV-1 subtypes A, B, C and AE. Both plasmid compositions boosted with MVA-CMDR generated HIV-1 specific cellular responses directed against HIV-1 Env, Gag and Pol, as measured by IFNγ ELISpot. It was shown that DNA priming augmented the vector MVA immunological boosting effects, the HIVISopt-DNA with a trend to improved (Env) neutralization, the HIVIS-DNA with a trend to better (Gag) cell mediated immune reponses. CONCLUSIONS: HIVIS-DNA was modified to obtain HIVISopt-DNA that had fewer plasmids, and additional epitopes. Even with one DNA prime followed by two MVA-CMDR boosts, humoral and cell-mediated immune responses were readily induced by priming with either DNA construct composition. Priming by HIV-DNA augmented neutralizing antibody responses revealed by boosting with the vaccinia-based heterologous sequences. Cellular and antibody responses covered selected strains representing HIV-1 subtypes A, B, C and CRF01_AE. We assume this is related to the inclusion of heterologous full genes in the vaccine schedule.
RESUMO
Germinal cell tumors of the testis were studied for the presence of several tumor-associated antigens. Antisera were produced by immunizing rabbits with the purified antigens of alpha-fetoprotein (AFP), carcinoembryonic antigen (CEA), and hepatoma ferritin. Indirect immunofluorescence on embryonal carcinoma with or without teratoma components demonstrated that their staining range was 1--60 per cent with antiserum against AFP, 0--16 per cent with anti-serum against ferritin, and 0-40% with antiserum against CEA. Ferritin-like substances have not been described previously in germinal tumors of the testis. No staining was seen with seminoma cells or benign testicular tissues. Raised serum levels of AFP and the ferritin-like substance were related both to the presence of tumor and to dissemination of the disease. CEA occurred transiently in serum. Eleven patients with primary tumors had no antigen in their sera and have all survived, but the median survival time for 8 patients with either antigen in preoperative sera was 12 months. Five patients with advanced tumor in whom neither AFP nor ferritin was detected had a much longer median survival time (58 mo) than did 13 patients with high levels of serum AFP or ferritin (12 mo). The presence of either AFP or ferritin in sera of patients with primary or advanced disease, therefore, seemed to indicate a poor prognosis. The determination of both substances in serum may be useful in the follow-up of patients with certain types of testicular tumors. The proportion of cells containing each antigen varied in the different tumors. Similarly, each antigen could occur independently in serum. This suggested that certain germ cell tumors contained subpopulations of cells, which differed in their production and release of the antigens studied.
Assuntos
Antígenos de Neoplasias , Teratoma/imunologia , Neoplasias Testiculares/imunologia , Antígenos de Neoplasias/análise , Antígeno Carcinoembrionário , Disgerminoma/imunologia , Ferritinas/imunologia , Imunofluorescência , Humanos , Masculino , Prognóstico , Teratoma/patologia , Neoplasias Testiculares/patologia , Testículo/imunologia , alfa-FetoproteínasRESUMO
Sera from patients with diseases in the pancreas, gallbladder, and bile duct were analyzed for the tumor markers CA 19-9, CA-50, and carcinoembryonic antigen. In particular CA 19-9 and CA-50 appear to be valuable in differentiating malignant from benign disease in these organs. Our sample of 72 patients with pancreatic cancer also indicates that CA 19-9 and CA-50 complement each other in 21% of the cases. They are also shown to be reliable for monitoring disease: following radical surgery for pancreatic cancer low levels of CA 19-9 and CA-50 were noted, while progressive rises of these tumor markers were related to disease progression.
Assuntos
Antígenos de Neoplasias/análise , Doenças dos Ductos Biliares/imunologia , Antígeno Carcinoembrionário/análise , Neoplasias Pancreáticas/imunologia , Neoplasias dos Ductos Biliares/imunologia , Colangite/imunologia , Doenças da Vesícula Biliar/imunologia , Humanos , Cirrose Hepática/imunologia , Pancreatopatias/imunologia , Pancreatite/imunologiaRESUMO
The three human isozymes of alkaline phosphatases were quantitatively determined in normal testis and seminoma tissues. The highly selective assays were based on isozyme specific monoclonal antibodies. In the normal testis approximately 90% of the catalytic activity originates from the tissue unspecific alkaline phosphatase, and the remaining activity was due to trace expression of both intestinal (approximately 5%) and placental alkaline phosphatase (PLAP) or PLAP-like isozyme (approximately 5%). In homogenates of seminoma tissues, highly increased levels of all three isozymes were identified. Both the tissue unspecific alkaline phosphatase and PLAP-like enzymes displayed relative increases of 10- to 100-fold and intestinal alkaline phosphatase 2- to 10-fold compared with normal testis. This finding indicates that the entire genome coding for alkaline phosphatases may be activated in seminomas. The PLAP-like enzyme from seminoma cells comprises a heterogenous population of molecules demonstrating partial heat sensitivity and microheterogeneity upon starch gel electrophoresis in contrast to the pregnancy related PLAP. These findings have implications for the different PLAP assays used in the clinical monitoring of seminoma patients.
Assuntos
Fosfatase Alcalina/análise , Disgerminoma/análise , Isoenzimas/análise , Neoplasias Testiculares/enzimologia , Coriocarcinoma/enzimologia , Feminino , Humanos , Masculino , Placenta/enzimologia , Testículo/enzimologia , Neoplasias Uterinas/enzimologiaRESUMO
Several pyrophosphate analogues have been compared for their ability to inhibit the activities of isolated cytomegalovirus (CMV) DNA polymerase, herpes simplex virus type 1 (HSV 1) DNA polymerase and calf thymus DNA polymerase alpha. The most effective inhibitors were phosphonoformate and phosphonoacetate. Although not identical, the structural requirements for compounds inhibitory to CMV and HSV-1 DNA polymerase were specific, with two negatively charged groups in close vicinity. The CMV DNA polymerase was more susceptible to certain phosphonoacetates containing bulky hydrophobic alpha-substituents than was the HSV-1 DNA polymerase. No example of the converse preference was found. The inhibition of CMV DNA polymerase by phosphonoformate, hypophosphate, alpha-hydroxyphosphonoacetate and alpha-nonylphosphonoacetate was linear non-competitive with the deoxyribonucleoside triphosphates as variable substrates. Phosphonoformate, phosphonoacetate, and to a lesser extent alpha-hydroxyphosphonoacetate, carbonyldiphosphonate and alpha-nonylphosphonoacetate also inhibited the focus formation by CMV in cell-culture.
Assuntos
Citomegalovirus/enzimologia , Inibidores da Síntese de Ácido Nucleico , Compostos Organofosforados/farmacologia , Simplexvirus/enzimologia , Timo/enzimologia , Animais , Bovinos , Cinética , Ácido Fosfonoacéticos/farmacologia , Especificidade da EspécieRESUMO
The viral diversity of HIV-1 is likely to require a vaccine strategy that induces broad cellular and humoral anti-HIV-1 immunity. Our strategy is based on multiple HIV-1 DNA immunogens together with adjuvant recombinant granulocyte-macrophage stimulating factor. This article describes pre-clinical and clinical work preceding the initiation of clinical HIV-1 phase I/II trials.
Assuntos
Vacinas contra a AIDS/genética , Vacinas contra a AIDS/imunologia , Antígenos HIV/genética , Antígenos HIV/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Vacinas de DNA/imunologia , Animais , Ensaios Clínicos Fase I como Assunto , Ensaios Clínicos Fase II como Assunto , Modelos Animais de Doenças , Produtos do Gene gag/genética , Produtos do Gene gag/imunologia , Produtos do Gene nef/genética , Produtos do Gene nef/imunologia , Produtos do Gene rev/genética , Produtos do Gene rev/imunologia , Produtos do Gene tat/genética , Produtos do Gene tat/imunologia , Proteína gp160 do Envelope de HIV/genética , Proteína gp160 do Envelope de HIV/imunologia , Infecções por HIV/prevenção & controle , Infecções por HIV/terapia , Transcriptase Reversa do HIV/genética , Transcriptase Reversa do HIV/imunologia , HIV-1/genética , Humanos , Vírus da Leucemia Murina , Camundongos , Camundongos Endogâmicos C57BL , Vacinas Combinadas/genética , Vacinas Combinadas/imunologia , Vacinas de DNA/genética , Produtos do Gene nef do Vírus da Imunodeficiência Humana , Produtos do Gene rev do Vírus da Imunodeficiência Humana , Produtos do Gene tat do Vírus da Imunodeficiência HumanaRESUMO
In the past couple of years, the idea that naked DNA can be used to vaccinate against infections has been rapidly developing. In contrast to traditional protein or live attenuated vaccines, there is no risk of disease caused by DNA vaccines as only selected proteins are encoded. The ease with which DNA may be manipulated means that vaccines can be custom designed to meet many needs. In animal model systems, DNA vaccines have proved to be as effective as traditional vaccines. Additionally, this technology may also be used to control existing chronic infections. Possibilities for treating hepatitis B, herpes simplex virus-2 and HIV, as well as infections with parasites, are being explored with success.
Assuntos
Biotecnologia/tendências , Controle de Doenças Transmissíveis , Vacinas de DNA/uso terapêutico , HumanosRESUMO
The complete amino acid (aa) sequence of the hepatitis B virus (HBV) core protein (HBcAg), deduced from the genome of the HBV ayw subtype, was synthesized as decapeptides with five overlapping aas. The peptides were tested for reactivity with monoclonal antibodies (mAbs) to the beta (or HBe2) epitope of hepatitis B e antigen (HBe/b mAbs; 57/8, 78/3, 141/158 and 141/207). Cross-competition between the mAbs with a mAb to the HBe/alpha epitope (or HBe1) and an anti-HBc mAb showed that all the HBe/b mAbs specifically inhibited human anti-HBe/b binding. Screening the HBc/e peptides showed that all anti-HBe/b mAbs recognized a peptide covering the residues 126-135. Three of the mAbs, 78/3, 141/152 and 141/207, had a less restricted reactivity than the other two, suggesting the recognition of the HBe/b as a discontinuous determinant. Fine mapping of the region aa 126-135 was performed by synthesizing decapeptides with nine overlapping aas, covering residues aa 121-140. All mAbs, except 78/3, reacted with the linear sequence TPPAYR, at residues 128-133. An additional set of peptides was synthesized, where the six aas within the epitope 128-133 were substituted in turn by the other 19 possible aas. By this approach, the essential aas for mAb 57/8 were found to be the sequence of PPA at residues 129-131, and for mAb 141/158 the sequence PP-Y, at residues 129, 130 and 132, respectively. Human recognition of the linear HBe/b epitope was investigated by using a peptide covering residues 121-140 (p 33). Thirty-one sera from chronic carriers of HBsAg, of which seven were positive for HBeAg and the remaining 24 for anti-HBe, were investigated. Of the sera with HBeAg, two had low levels of anti/-HBe/b in the p 33 assay. Out of the sera with anti-HBe, eight were positive in the p 33 EIA. Thus, murine monoclonals and human sera may recognize the HBe/b epitope as a linear determinant residing around aa 130.
Assuntos
Linfócitos B/imunologia , Epitopos/imunologia , Antígenos E da Hepatite B/imunologia , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais/imunologia , Ligação Competitiva , Hepatite B/imunologia , Antígenos do Núcleo do Vírus da Hepatite B/imunologia , Humanos , Técnicas Imunoenzimáticas , Dados de Sequência Molecular , Peptídeos/síntese química , RadioimunoensaioRESUMO
A new type of immunochemical mapping of the human immunodeficiency virus type 1 (HIV-1) gag region was performed. By use of native HIV-1 viral lysates or the gag recombinant p24-15 antigen, a new set of monoclonal antibodies (Mabs) to the gag region proteins was generated. Synthetic HIV-1 peptides covering the entire gag region were used to specifically localize the continuous epitopes by direct binding to the Mabs and by blocking the Mab immunoreactivity. The identified immunogenic epitopes were localized between the gag amino acids (aa) 108-127, 203-217, 208-222, 248-282, 273-302, 288-307, 308-322, 331-354 and 408-422. These continuous epitopes formed seven immunogenic regions. One strongly p17-reactive Mab appeared to react with a discontinuous epitope, the components of which were 110 aa distant in the linear sequence: aa 23-27 and 128-132. The synthetic peptides appeared to be more congruent with the Mab-reactive sites in solution than when coated to a solid phase.
Assuntos
Produtos do Gene gag/imunologia , Antígenos HIV/imunologia , HIV-1/imunologia , Sequência de Aminoácidos , Animais , Anticorpos Monoclonais , Ligação Competitiva , Western Blotting , Ensaio de Imunoadsorção Enzimática , Epitopos , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Fragmentos de Peptídeos/síntese química , Fragmentos de Peptídeos/imunologia , Proteínas Recombinantes/imunologiaRESUMO
The nucleotide sequence of a murine monoclonal antibody (CB-mab-p24/13-5) against p24 core protein of the human immunodeficiency virus (HIV-1) was determined for variable regions of the heavy and light chain, respectively. Genetic elements encoding the VDJH- and VJL-regions of the antibody were generated from RNA by the polymerase chain reaction, cloned into the vector pICEM 19R and sequenced. Synthetic peptides, 10 amino acids overlapping served for the localization of the epitope. The residues 152-156 within the p24 sequence contain the epitope.
Assuntos
Anticorpos Monoclonais/genética , Proteína do Núcleo p24 do HIV/genética , Região Variável de Imunoglobulina/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Hibridomas , Cadeias Pesadas de Imunoglobulinas/genética , Cadeias Leves de Imunoglobulina/genética , Camundongos , Dados de Sequência Molecular , Reação em Cadeia da PolimeraseRESUMO
The EPIICAL (Early-treated Perinatally HIV-infected Individuals: Improving Children's Actual Life with Novel Immunotherapeutic Strategies) project arises from the firm belief that perinatally infected children treated with suppressive antiretroviral therapy (ART) from early infancy represent the optimal population model in which to study novel immunotherapeutic strategies aimed at achieving ART-free remission. This is because HIV-infected infants treated within 2-3 months of life have a much reduced viral reservoir size, and rarely show HIV-specific immunity but preserve normal immune development. The goal of EPIICAL is the establishment of an international collaboration to develop a predictive platform using this model to select promising HIV therapeutic vaccine candidates, leading to prioritisation or deprioritisation of novel immunotherapeutic strategies. To establish this platform, the EPIICAL Consortium aims to: develop predictive models of virological and immunological dynamics associated with response to early ART and to treatment interruption using available data from existing cohorts/studies of early-treated perinatally HIV-infected children; optimise methodologies to better characterise immunological, virological and genomic correlates/profiles associated with viral control; test novel immunotherapeutic strategies using in vivo proof-of-concept (PoC) studies with the aim of inducing virological, immunological and transcriptomic correlates/profiles equivalent to those defined by the predictive model. This approach will strengthen the capacity for discovery, development and initial testing of new therapeutic vaccine strategies through the integrated efforts of leading international scientific groups, with the aim of improving the health of HIV-infected individuals.
RESUMO
BACKGROUND: Long-term non-progression in HIV-1-infected patients has been reported to be associated with a 32 base-pair deletion (delta32) in one CCR-5 allele. The normal gene product acts as a coreceptor for HIV cell entry and is essential for infection of cells by non-syncytium-inducing and MT-2-negative HIV-1 strains. METHODS: Forty individuals were studied, all of whom had been HIV-1-seropositive for a mean of 8 years. RESULTS: Eight (20%) were heterozygous for the CCR-5 allele delta32 deletion. Six of these eight patients harboured MT-2-negative HIV-1 strains. Of these six, three were long-term non-progressors with a positive CD4 cell slope, not receiving antiretroviral treatment, whereas the other three were progressors (mean CD4 cell decline, 3.8 x 10(6)/l per month) receiving antiretroviral combination therapy. Two of the eight patients with the delta32 deletion had MT-2-positive HIV-1 strains. Both had very rapid CD4 cell decline (6.7 and 7.6 x 10(6)/l per month, respectively), despite triple antiretroviral therapy including a protease inhibitor. One of the patients with an MT-2-positive virus strain has suffered from Pneumocystis carinii bronchitis and the other from cytomegalovirus colitis. CONCLUSIONS: Disease progression may also occur in individuals with the coreceptor deficiency, especially in association with MT-2-positive HIV-1 strains. It is suggested that MT-2-positive HIV-1 enters cells through the CXC chemokine receptor-4 fusin coreceptor, thus circumventing the defective CC chemokine receptor-5 coreceptor. Various levels of expression of the wild-type CCR-5 gene and the gene with the delta32 deletion might explain variations in the disease progression in heterozygous patients with MT-2-negative HIV-1 strains.
Assuntos
Deleção de Genes , Infecções por HIV/genética , HIV-1 , Receptores CCR5/genética , Adulto , Alelos , Proteína gp160 do Envelope de HIV/imunologia , Infecções por HIV/fisiopatologia , Heterozigoto , Humanos , Imunização , Masculino , Pessoa de Meia-Idade , Prognóstico , Sobreviventes , Carga ViralRESUMO
BACKGROUND: The beta-chemokine receptor CCR-5 is the coreceptor for cellular entry by non-syncytium-inducing (NSI) HIV-1 strains that dominate early in infection. A 32 base-pair deletion (delta32) in the CCR-5 gene renders this coreceptor non-functional. Heterozygosity for this deletion [delta32/wild-type (wt)] is associated with slow disease progression. The purpose of this study was to document the combined impact on HIV-1 disease progression of the CCR-5 genotype and the biological phenotype of HIV-1. METHODS: In a cross-sectional study of 258 HIV-1-infected Swedish individuals, the CCR-5 genotype (wt/wt or delta32/wt) was determined by polymerase chain reaction and the biological phenotype [NSI or syncytium-inducing (SI)] of virus isolates was determined in the MT-2 cell assay. Clinical status, HIV-1 RNA levels in plasma, CD4+ lymphocyte counts, and rate of CD4+ lymphocyte decline, based on retrospective analysis of CD4+ lymphocyte counts, were also recorded. None of the individuals were treated with protease inhibitors. RESULTS: The prevalence of the delta32/wt genotype was 23%. Subjects with the delta32/wt CCR-5 genotype more often carried SI virus than subjects with the wt/wt genotype (49 versus 35%; P=0.067), but there were no differences between the two groups in prevalence of AIDS, viral load, CD4+ lymphocyte count or CD4+ slope. NSI virus isolates were found in 159 (62%) out of 258 individuals. Individuals with NSI had lower prevalence of AIDS (39 versus 19%; P < 0.01), higher CD4+ lymphocyte counts (289+/-188 x 10(6)/l versus 153+/-162 x 10(6)/l; P=0.001), lower viral loads (median, 4.45 log10 versus 4.91 log10 copies/ml; P < 0.01) and a lower prevalence of the delta32/wt genotype (19 versus 29%; P=0.067) compared with individuals with SI virus. When the material was further subdivided, subjects with the delta32/wt genotype and SI virus had the highest prevalence of AIDS (P < 0.001), lowest CD4+ lymphocyte count (P=0.0001) and highest viral load (P=0.023) whereas the opposite was true for subjects with the delta32/wt genotype and NSI virus. A significantly higher proportion of subjects with NSI virus with delta32/wt and wt/wt CCR-5 genotype had been immunized with recombinant gp160. CONCLUSION: In summary, the delta32/wt CCR-5 genotype has a protective effect against HIV-1 disease progression that appears to be limited to individuals carrying HIV-1 variants with NSI phenotype. Immunization with recombinant gp160 tended to reduce the frequency of SI phenotypes.
Assuntos
Infecções por HIV/fisiopatologia , HIV-1 , Receptores CCR5/genética , Vacinas contra a AIDS/imunologia , Adulto , Idoso , Contagem de Linfócito CD4 , Linhagem Celular Transformada , Estudos de Coortes , Estudos Transversais , Progressão da Doença , Seguimentos , Genótipo , Proteína gp160 do Envelope de HIV/imunologia , Infecções por HIV/mortalidade , Infecções por HIV/prevenção & controle , Infecções por HIV/virologia , HIV-1/genética , HIV-1/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Fenótipo , Proteínas Recombinantes/imunologia , Deleção de Sequência , Vacinas Sintéticas/imunologia , Carga ViralRESUMO
BACKGROUND: The efficacy of highly active antiretroviral treatment (HAART) in HIV-1 disease may vary between nucleoside-naive and experienced patients as well as between patients with different viral phenotypes and in different stages of disease. OBJECTIVE: To investigate variables of importance for successful long-term viral suppression by analysing virological, clinical and immunological characteristics at initiation of protease inhibitor treatment on suppression of HIV RNA over 1 year. DESIGN: An open, non-randomized, observational clinical study. SETTING: Venhälsan, Department of Dermatovenereology, Söder Hospital, Stockholm, Sweden. PATIENTS: A total of 147 unselected advanced patients with known HIV-1 infection for a mean of 7 years, of whom 37% had AIDS and who started treatment with a protease inhibitor during 1996. INTERVENTIONS: All patients received HAART with at least two nucleoside analogues in combination with either indinavir (81%) or ritonavir (19%). The majority (77%) had been previously treated with nucleoside analogues for a mean of 39 months. MEASUREMENTS: CD4+ lymphocyte count, plasma HIV-1 RNA, viral phenotype and HIV-1 coreceptor CCR-5 genotype at baseline. Viral load and CD4+ lymphocyte count were determined every 3 months. RESULTS: Patients were analysed on an intention-to-treat basis. The mean CD4+ lymphocyte count at baseline was 170 x 10(6)/l and the median viral load was 68 600 copies/ml. Heterozygosity for the delta32 deletion of the CCR-5 gene (delta32/wt) was found in 27%. MT-2 positive virus (syncytium-inducing) was isolated in 46%. Logistic regression revealed that nucleoside analogue experience and baseline log10 HIV-1 RNA were the only factors independently related to plasma HIV-1 RNA levels below 500 copies/ml after 1 year of treatment, which was found in 69%. CONCLUSION: The virological outcome after 1 year of HAART was strongly correlated to prior treatment history and baseline viral load, whereas CD4+ lymphocyte count, CCR-5 genotype and viral biological phenotype had less influence. The long-term antiviral efficacy of HAART was lowest in individuals with previous nucleoside analogue treatment and a high baseline viral load. In these individuals an even more aggressive treatment should be considered.
Assuntos
Fármacos Anti-HIV/uso terapêutico , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , Carga Viral , Adulto , Fármacos Anti-HIV/efeitos adversos , Contagem de Linfócito CD4 , Progressão da Doença , Quimioterapia Combinada , Feminino , Seguimentos , Genótipo , Infecções por HIV/imunologia , HIV-1/genética , HIV-1/isolamento & purificação , Humanos , Estudos Longitudinais , Masculino , Inibidores de Proteases/uso terapêutico , RNA Viral/sangue , Análise de Regressão , Estatísticas não Paramétricas , TropismoRESUMO
OBJECTIVE: To investigate whether the specificity of antibody responses to the gp120 V3 domain in HIV-1-infected individuals is related to the variability of this region. METHODS: Sera from a cohort of 22 HIV-1-infected Ugandans were tested against peptides derived from each individual's autologous proviral V3 apex sequence. Autologous peptide reactivity was compared with reactivity to peptides derived from two Ugandan consensus sequences and previously isolated US/European and African viruses. Peptides from individuals with heterogeneous V3 apex sequences, representing different HIV-1 variants, were obtained and tested against the corresponding sera. RESULTS: A notable cross-reactivity to different V3 apex peptides was observed. However, in the majority of sera, antibody reactivity to the autologous peptides was found to exceed reactivity to any of the other peptides tested. V3 proviral sequences from the Ugandan cohort studied have been shown to be closely related to the HIV-1MN isolate and thus, their sera gave better reactivity to V3MN and related peptides than to peptides representing other African HIV-1 isolates. In individuals with heterogeneous V3 proviral sequences, we could distinguish divergent antibody responses to the genomic variants differing by single amino acids. CONCLUSION: Analysis of seroreactivity to peptides might constitute a relevant tool for investigating the variability of the HIV-1 gp120 V3 domain within infected populations and single individuals.
Assuntos
Variação Antigênica , Proteína gp120 do Envelope de HIV/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Fragmentos de Peptídeos/imunologia , Sequência de Aminoácidos , Especificidade de Anticorpos , Reações Cruzadas , Ensaio de Imunoadsorção Enzimática , Anticorpos Anti-HIV/imunologia , Humanos , Dados de Sequência Molecular , UgandaRESUMO
OBJECTIVE: To analyse the pattern of immunoglobulin (Ig) G subclasses specific for HIV-1 envelope peptides in sera from HIV-1-infected mothers and their newborns. We sought to determine whether there was a selective transfer of antibodies from mother to child, and to establish diagnostic or prognostic properties by analysing HIV-1 peptide-specific IgG isotypes. DESIGN AND METHODS: Parallel sera from 12 HIV-1-infected mothers and their newborn children were analysed for IgG subclass responses to six HIV-1 envelope peptides by enzyme-linked immunosorbent assay. Levels of IgG were compared with levels of IgG1 and IgG3 to an immunodominant gp41 peptide in children's sera obtained during the first months of life. In a longitudinal study of 16 children born to HIV-1-seropositive mothers, of whom 11 were infected, the IgG1 and IgG3 responses to peptides representing an immunodominant epitope of gp41 and the principal neutralizing determinant of the gp120 V3 region were analysed. RESULTS: IgG1 and IgG3 were found to constitute the predominant peptide-specific antibody responses. A parallel distribution of IgG subclass reactivity was seen in maternal and paediatric sera. There was no evidence of selective antibody transfer. Comparable levels of IgG and IgG1 to the immunodominant peptide were seen in infected and uninfected children, while the IgG3 had disappeared in the majority of uninfected children. A decrease in peptide-specific IgG1 and IgG3 levels was observed in sequential sera from uninfected children, although the kinetic profile varied. Sera from infected children showed de novo synthesis of IgG1 and/or IgG3 binding to the selected HIV-1 peptides. Most children with rapid disease progression failed to produce IgG1 and/or IgG3 to the V3 peptide. CONCLUSION: Analysis of IgG subclass kinetics to selected HIV-1 peptides might be a useful additional diagnostic and prognostic tool in evaluating HIV-1 infection in children born to seropositive mothers.
Assuntos
Proteína gp120 do Envelope de HIV/imunologia , Proteína gp41 do Envelope de HIV/imunologia , Infecções por HIV/transmissão , HIV-1/imunologia , Imunoglobulina G/classificação , Recém-Nascido/imunologia , Troca Materno-Fetal/imunologia , Adulto , Sequência de Aminoácidos , Especificidade de Anticorpos , Reações Antígeno-Anticorpo , Feminino , Seguimentos , Infecções por HIV/imunologia , Humanos , Imunoglobulina G/química , Lactente , Dados de Sequência Molecular , GravidezRESUMO
We analysed HIV-specific immunoglobulin G (IgG) responses to gag and env peptides in infants born to HIV-positive mothers. Questions of interest were whether there are early specific markers for prognosis, and whether any specific IgG is related to the prevention of vertical transmission of infection. Fifty-three children, 0-24 months old and born to HIV-1-infected mothers, were retrospectively divided into two groups based on HIV seroreactivity or non-reactivity at 15 months of age. Their sera were used to find reactivities important in diagnosis and/or prediction of the putative HIV disease. Three important findings emerged. First, a low IgG titer against the very immunodominant penv9 in newborns was found to be associated with rapid progression to AIDS. This difference was clearly reflected in the reactivity to a small peptide representing amino acid (aa) 598-606. The second interesting finding was the putative hypervariable loop on gp120 (especially aa 324-338), reactivity to which was found only in the uninfected group, and was seen in six out of 19 uninfected children under 6 months of age. This specific response was not caused by a generally high total anti-HIV reactivity, and may indicate a role of protective antibodies against vertical transmission. The response to this region in the infected group, on the other hand, was directed to the amino terminal half of the putative loop, in particular peptide 53, aa 304-318. Finally, response to a part of the amino terminal end of P17 was seen in seven out of eight infected children over 6 months of age.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Síndrome da Imunodeficiência Adquirida/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Imunoglobulina G/imunologia , Sorodiagnóstico da AIDS , Síndrome da Imunodeficiência Adquirida/diagnóstico , Síndrome da Imunodeficiência Adquirida/transmissão , Feminino , Produtos do Gene gag/imunologia , Proteína gp120 do Envelope de HIV/imunologia , Proteína gp41 do Envelope de HIV/imunologia , Infecções por HIV/diagnóstico , Infecções por HIV/transmissão , Humanos , Lactente , Recém-Nascido , Troca Materno-Fetal/imunologia , Fragmentos de Peptídeos/imunologia , Gravidez , Complicações Infecciosas na Gravidez/sangue , Complicações Infecciosas na Gravidez/imunologia , Prognóstico , Proteínas Recombinantes/imunologia , Estudos RetrospectivosRESUMO
Amino acid sequences inducing neutralizing antibodies to HIV-1 were sought. Murine monoclonal antibodies (MAbs) were characterized by their reactivity with the envelope precursor gp160 or the Escherichia coli recombinant DNA products pB1 and pE3 representing the carboxy- and amino-terminal halves of mature envelope gp120. Fine mapping of the MAb determinants was performed using defined 15-mer synthetic peptides spanning the entire envelope gp120 region of HIV-1. One group of MAbs recognizes epitopes (amino acids 304-323) occurring in a small region with variable and conserved amino acid sequences of gp120. These MAbs mediate neutralization of the HIV-1 strain HTLV-IIIB (HIV-1IIIB) which was used for immunization. Nine out of 11 primary HIV-1 isolates were neutralized well or moderately well. In addition, prominent serological reactivity was noted with peptide sequences of strains of various European or American origins, but not with two HIV-1 strains of African origin. The cross-reactivity contrasts with previously described type-specific reactions to other sequences of this region. The reactivity to the short conserved site GPGR with its flanking amino acids may explain the broad sequence cross-reactivity seen with our neutralizing MAbs. Two other MAbs recognize conserved epitopes (amino acids 79-103) situated in the amino-terminal region of gp120. These MAbs did not neutralize HIV-1IIIB.