RESUMO
The computer-designed Top7 served as a scaffold to produce immunoreactive proteins by grafting of the 2F5 HIV-1 antibody epitope (Top7-2F5) followed by biotinylation (Top7-2F5-biotin). The resulting nonimmunoglobulin affinity proteins were effective in inducing and detecting the HIV-1 antibody. However, the grafted Top7-2F5 design led to protein aggregation, as opposed to the soluble biotinylated Top7-2F5-biotin. The structure-based model predicted that the thermodynamic cooperativity of Top7 increases after grafting and biotin-labeling, reducing their intermediate state populations. In this work, the folding kinetic traps that might contribute to the aggregation propensity are investigated by the diffusion theory. Since the engineered proteins have similar sequence and structural homology, they served as protein models to study the kinetic intermediate traps that were uncovered by characterizing the position-dependent drift-velocity (v(Q)) and the diffusion (D(Q)) coefficients. These coordinate-dependent coefficients were taken into account to obtain the folding and transition path times over the free energy transition states containing the intermediate kinetic traps. This analysis may be useful to predict the aggregated kinetic traps of scaffold-epitope proteins that might compose novel diagnostic and therapeutic platforms.
Assuntos
Biotina , Dobramento de Proteína , Biotina/metabolismo , Proteínas/química , Termodinâmica , Epitopos , Proteína gp41 do Envelope de HIV , Anticorpos Anti-HIVRESUMO
Broadly neutralizing antibodies against HIV-1 are rare with the 2F5 antibody being one of the most protective. Insertion of an antibody epitope into a stable and small protein scaffold overcomes many of the obstacles found to produce antibodies. However, the design leads to grafting of epitopes that may cause protein aggregation. Here, I investigated the 2F5 epitope grafted into the Top7 as the scaffold in which the resulting immunoreactive protein precipitates along the storage time, as opposed to its completely soluble biotinylated version. Molecular dynamics showed that biotinylation eliminates the intermediate state of the scaffold-epitope Top7-2F5 by switching a noncooperative to a cooperative folding. The aggregation propensity of the Top7-designed proteins is examined in light of thermodynamic cooperativity and kinetic traps along the decreasing depth of the intermediate ensemble in the free energy landscape. This protocol may predict stable and soluble scaffold-epitopes with the purpose of composing novel therapeutic and diagnostic platforms.
Assuntos
HIV-1 , Biotinilação , Anticorpos Amplamente Neutralizantes , Epitopos , Anticorpos Anti-HIV/metabolismo , Proteína gp41 do Envelope de HIV/metabolismo , HIV-1/metabolismo , Agregados Proteicos , Proteínas/metabolismoRESUMO
PROBLEM: Pregnancy appears to favor maternal antibody production. In contrast, by damaging follicular helper T cells (TFH ), HIV-1 infection compromises protective humoural immune response. Therefore, we aimed to investigate the frequency of different TFH -like cells in HIV-infected pregnant women (PW) before and after antiretroviral (ARV) therapy. METHOD OF STUDY: Peripheral blood mononuclear cells, CD4+ T and B cells, were obtained from asymptomatic HIV-1-infected non-PW and PW just before and after ARV therapy. In some experiments, healthy HIV-1-negative PW were also tested. The frequency of different TFH -like cell subsets was determined by flow cytometry. The plasma titers of IgG anti-tetanus toxoid (TT), anti-HBsAg, and anti-gp41 were determined by ELISA. The in vitro production of total IgG, IL-21, and hormones (estrogen and progesterone) was quantified also by ELISA. RESULTS: Our results demonstrate that antiretroviral (ARV) therapy was more efficient in elevating the percentage of circulating IL-21-secreting TFH cells in HIV-1-infected pregnant women (PW) than in non-pregnant patients (nPW). Moreover, in co-culture systems, CD4+ T cells from ART-treated PW were more efficient in assisting B cells to produce IgG production. The in vivo anti-HBsAg IgG titers after ARV therapy were also significantly higher in PW, and their levels were directly associated with both IL-21+ TFH frequency and plasma concentration of estrogen. CONCLUSION: In summary, our results suggest that pregnancy favors the recovery of TFH -like cells after ARV therapy in HIV-1-infected women, which could help these mothers to protect their newborns from infectious diseases by transferring IgG across the placenta.
Assuntos
Fármacos Anti-HIV/farmacologia , Infecções por HIV/imunologia , HIV-1 , Interleucinas/metabolismo , Complicações Infecciosas na Gravidez/imunologia , Células T Auxiliares Foliculares/metabolismo , Adulto , Fármacos Anti-HIV/uso terapêutico , Anticorpos Antibacterianos/sangue , Formação de Anticorpos/efeitos dos fármacos , Linfócitos B/efeitos dos fármacos , Contagem de Linfócito CD4 , Células Cultivadas , Técnicas de Cocultura , Estrogênios/sangue , Feminino , Anticorpos Anti-HIV/sangue , Proteína gp41 do Envelope de HIV/imunologia , Infecções por HIV/sangue , Infecções por HIV/tratamento farmacológico , HIV-1/imunologia , Anticorpos Anti-Hepatite B/sangue , Antígenos de Superfície da Hepatite B/imunologia , Humanos , Imunoglobulina G/sangue , Gravidez , Complicações Infecciosas na Gravidez/sangue , Progesterona/sangue , Toxoide Tetânico/imunologia , Adulto JovemRESUMO
Fusion inhibitors are antiretroviral (ARV) drugs that prevent HIV-1 entry into host cells. Enfuvirtide (ENF) is the only ARV drug marketed in this class and, like other HIV drugs, it has been associated with the emergence and selection of therapeutic-resistant HIV-1 strains. The aims of this work were to develop a computational tool capable of identifying and classifying mutations associated with resistance to Enfuvirtide and to evaluate the prevalence of these mutations among the HIV-1 sequences deposited in public databases. The HIVfird (HIV-1 fusion inhibitor resistance detector) was developed using the PHP programming language, using 30 DNA bases obtained from the HIV-1 HXB2 gp41 protein as a reference. To assess the level of resistance in HIV-1 populations, sequences were retrieved from the Los Alamos National Laboratory (LANL) database. The HIVfird is hosted at www.hivfird.ics.ufba.br, fully functional and available for public use. Twenty-five amino acid substitutions and 15 combinations were found to be associated with some level of resistance to ENF. These mutations are located at positions 36-45 of gp41, with 36, 38, 43, and 44 having the greatest diversity and frequency of variations associated with drug resistance. Resistance mutations were found in 3.16% and 4.67% of the circulating HIV-1 isolates in the world and Brazil, respectively. Subtype B showed a significantly higher ENF resistance rate (4.9%) compared to other genetic forms, while subtype C presented the lowest rate (0.9%). We present here HIVfird, an online tool that might assist in the therapeutic management of HIV-1 patients with multiple drug failure and in population-based analysis of drug resistance.
Assuntos
Análise Mutacional de DNA/métodos , DNA Viral/genética , Farmacorresistência Viral , Enfuvirtida/farmacocinética , Proteína gp41 do Envelope de HIV/genética , Inibidores da Fusão de HIV/farmacologia , Infecções por HIV/tratamento farmacológico , HIV-1/efeitos dos fármacos , Mutação , Software , Sequência de Aminoácidos , Substituição de Aminoácidos , Sequência de Bases , Bases de Dados Genéticas , Farmacorresistência Viral/genética , Enfuvirtida/uso terapêutico , Saúde Global , Proteína gp41 do Envelope de HIV/fisiologia , Inibidores da Fusão de HIV/uso terapêutico , Infecções por HIV/virologia , HIV-1/classificação , HIV-1/genética , HIV-1/isolamento & purificação , Humanos , Internet , Mutação de Sentido Incorreto , Alinhamento de Sequência , Homologia de Sequência do Ácido NucleicoRESUMO
Antiretroviral therapy has significantly evolved in the last decade, with an increasing number of new drugs and classes. Currently, even heavily experienced patients can be successfully treated with new regimens. In Brazil, the recent incorporation of some new antiretroviral drugs made it possible to suppress HIV plasma viremia in most treated patients, with significant benefits in terms of quality of life and survival. However, little has been published on outcomes of patients under new drugs-based regimens. We reviewed the safety and efficacy of antiretroviral regimens using recently introduced drugs in Bahia. Our results confirm that patients using darunavir, raltegravir, enfuvirtide, or etravirine presented with a high rate of virological suppression without significant adverse events, after one year of follow-up.
Assuntos
Síndrome da Imunodeficiência Adquirida/tratamento farmacológico , Fármacos Anti-HIV/uso terapêutico , Terapia Antirretroviral de Alta Atividade , Adulto , Brasil , Contagem de Linfócito CD4 , Darunavir/uso terapêutico , Esquema de Medicação , Enfuvirtida , Feminino , Proteína gp41 do Envelope de HIV/uso terapêutico , Humanos , Masculino , Nitrilas , Fragmentos de Peptídeos/uso terapêutico , Piridazinas/uso terapêutico , Pirimidinas , Qualidade de Vida , Raltegravir Potássico/uso terapêutico , Terapia de Salvação , Resultado do Tratamento , Carga ViralRESUMO
Multi-HIV, a multiepitopic protein derived from both gp120 and gp41 envelope proteins of the human immunodeficiency virus (HIV), has been proposed as a vaccine prototype capable of inducing broad immune responses, as it carries various B and T cell epitopes from several HIV strains. In this study, the immunogenic properties of a Multi-HIV expressed in tobacco chloroplasts are evaluated in test mice. BALB/c mice orally immunized with tobacco-derived Multi-HIV have elicited antibody responses, including both the V3 loop of gp120 and the ELDKWA epitope of gp41. Based on splenocyte proliferation assays, stimulation with epitopes of the C4, V3 domain of gp120, and the ELDKWA domain of gp41 elicits positive cellular responses. Furthermore, specific interferon gamma production is observed in both CD4+ and CD8+ T cells stimulated with HIV peptides. These results demonstrate that plant-derived Multi-HIV induces T helper-specific responses. Altogether, these findings illustrate the immunogenic potential of plant-derived Multi-HIV in an oral immunization scheme. The potential of this low-cost immunization approach and its implications on HIV/AIDS vaccine development are discussed.
Assuntos
Proteína gp120 do Envelope de HIV/biossíntese , Proteína gp41 do Envelope de HIV/biossíntese , Infecções por HIV/imunologia , Planticorpos/imunologia , Animais , Cloroplastos/imunologia , Epitopos de Linfócito T/imunologia , Proteína gp120 do Envelope de HIV/administração & dosagem , Proteína gp120 do Envelope de HIV/imunologia , Proteína gp41 do Envelope de HIV/administração & dosagem , Proteína gp41 do Envelope de HIV/imunologia , Infecções por HIV/prevenção & controle , Infecções por HIV/virologia , HIV-1/imunologia , HIV-1/patogenicidade , Humanos , Imunização , Camundongos , Nicotiana/citologia , Nicotiana/imunologiaRESUMO
Enfuvirtide was the first fusion inhibitor approved by the Food and Drug Administration (FDA) in 2003 for HIV-1 infection in treatment-experienced patient. It is the first approved antiviral agent to attack the HIV life cycle in its early stages. For HIV fusion to occur, the HR1 and HR2 domains in the gp41 region need to interact. Enfuvirtide is a synthetic peptide that corresponds to 36 amino acids of the HR2, which competitively binds to HR1 inhibiting the interaction with the HR2 domain thus preventing fusogenic conformation and inhibiting viral entry into host cells. Resistance to enfuvirtide is conferred by mutations occurring in the HR1 region involving residues 36-45. Mozambique, a sub-Saharan country, with an HIV prevalence of 11.5%, provides first line and second line antiretroviral therapy (ART)-based treatment. In poor resource settings such as Mozambique the lack of adequate infrastructures, the high costs of viral load tests, and the availability of salvage treatment have hindered the intended objective of monitoring HIV treatment, suggesting an important concern regarding the development of drug resistance. The general aim of this study was to evaluate naturally occurring polymorphisms and resistance-associated mutations in the gp41 region of HIV-1 isolates from Mozambique. The study included 78 patients naive to ARV treatment and 28 patients failing first line regimen recruited from Centro de Saúde Alto-Maé situated in Maputo. The gp41 gene from 103 patients was sequenced and resistance-associated mutations for enfuvirtide were screened. Subtype analysis revealed that 96% of the sequences were classified as subtype C, 2% as subtype G, 1% as subtype A1, and the other 1% as a mosaic form composed of A1/C. No enfuvirtide resistance-associated mutations in HR1 of gp41 were detected. The major polymorphisms in the HR1 were N42S, L54M, A67T, and V72I. This study suggests that this new class of antiviral drug may be effective as a salvage therapy in patients failing first line regimens in Mozambique. However, further phenotypic studies are required to determine the clinical relevance of the polymorphisms detected in this study.
Assuntos
Humanos , Masculino , Feminino , Pré-Escolar , Adulto , Pessoa de Meia-Idade , Polimorfismo Genético , Proteína gp41 do Envelope de HIV/genética , Infecções por HIV/virologia , HIV-1/genética , Fragmentos de Peptídeos , Filogenia , RNA Viral , Dados de Sequência Molecular , Análise por Conglomerados , Alinhamento de Sequência , Sequência de Aminoácidos , HIV-1/classificação , Falha de Tratamento , Análise de Sequência de DNA , Fármacos Anti-HIV , Mutação de Sentido Incorreto , Farmacorresistência Viral , Enfuvirtida , Genótipo , MoçambiqueRESUMO
The effectiveness of switching from enfuvirtide to raltegravir in HIV-1-infected patients on a suppressive antiretroviral regimen has been poorly studied in the clinical practice of developing countries. We conducted a multicenter retrospective cohort study in HIV-1-infected, multidrug-experienced adults (≥18 years old) with plasma HIV-1-RNA <400 copies/ml for at least 4 months on an enfuvirtide-containing therapy between 2005 and 2010, in whom the attending physician switched from enfuvirtide to raltegravir. Effectiveness endpoints were measured at week 48 after switch. Analyses were conducted on an intent-to-treat basis and two strategies for handling missing outcome data were used (hereafter, strategies 1 and 2). Overall, 87 patients were eligible for analysis. At baseline, the median CD4(+) T cell count was 400 cells/µl and 91.9% of patients had <50 HIV-1-RNA copies/ml. At week 48, the proportions of patients with plasma HIV-1-RNA <50 and <400 copies/ml were, respectively, 86.2% (95% CI=77.1; 92.7%) and 88.5% (95% CI=79.9; 94.3%) (strategies 1 and 2) and 89.7% (95% CI=81.3; 95.2%) and 90.8% (95% CI=82.7; 95.9%) (strategies 1 and 2). This was a -10.3% (95% CI=-2.8; -17.9%) and -9.2% (95% CI=-2; -16.4%) difference from baseline in the proportion of patients with plasma HIV-1-RNA <400 copies/ml. The median increase in CD4(+) T cell counts was 41 and 64 cells/µl (p<0.001) (strategies 1 and 2). No patient withdrew raltegravir or developed opportunistic infections, but one was diagnosed with HIV-related dementia. In conclusion, switching from enfuvirtide to raltegravir in patients on a virologically suppressive regimen is an effective strategy even in a Brazilian clinical setting.
Assuntos
Fármacos Anti-HIV/uso terapêutico , Farmacorresistência Viral , Proteína gp41 do Envelope de HIV/uso terapêutico , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , Fragmentos de Peptídeos/uso terapêutico , Pirrolidinonas/uso terapêutico , Adolescente , Adulto , Brasil , Contagem de Linfócito CD4 , Estudos de Coortes , Enfuvirtida , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , RNA Viral/sangue , Raltegravir Potássico , Resultado do Tratamento , Carga Viral , Adulto JovemRESUMO
The selective pressure of antiretroviral drugs (ARVs) targeting HIV-1 pol can promote drug resistance mutations in other genomic regions, such as env. Drug resistance among women should be monitored to avoid horizontal and mother-to-child transmission. To describe natural resistance to T-20 (enfuvirtide), gp41 env polymorphisms, mutations in pol and HIV-1 subtypes, 124 pregnant women were recruited. For 98 patients, the gp41 env, protease (PR) and reverse transcriptase (RT) fragments were sequenced. The patients were ARV naïve (n = 30), taking mother-to-child transmission prophylaxis (n = 50), or being treated with highly active ARV therapy/HAART (n = 18). The Stanford and IAS/USA databases and other sources were used to analyze PR/RT, gp41 env resistance mutations. The HIV-1 genetic diversity was analyzed by REGA/phylogenetic analyses. The patients' median age was 25 years (range, 16-42), 18.4% had AIDS. The frequency of natural resistance to T-20 (N42D, L44M, and R46M-low-impact mutations) was 6.1% (6/98); 20.4% (20/98) had compensatory mutations in HR2. The prevalence of transmitted drug resistance in the pol was 13.3% (4/30), and the prevalence of secondary drug resistance was 33.3% (6/18). Two patients were infected with multidrug resistant/MDR viruses. The analysis of HIV-1 subtypes (PR/RT/gp41) revealed that 61.2% (60/98) were subtype B, 12.2% (12/98) were subtype C, 4.1% (4/98) were subtype F1, and 22.4% (22/98) were possible recombinants (BF1 = 20.4%; BC = 2%). Natural resistance to T-20 was not associated with pol resistance or previous ARV use. The high rate of secondary resistance, including MDR, indicates that the number of women that may need T-20 salvage therapy may be higher than anticipated.
Assuntos
Fármacos Anti-HIV/farmacologia , Farmacorresistência Viral , Variação Genética , Proteína gp41 do Envelope de HIV/genética , HIV-1/genética , Complicações Infecciosas na Gravidez/virologia , Produtos do Gene pol do Vírus da Imunodeficiência Humana/genética , Adolescente , Adulto , Brasil , Enfuvirtida , Feminino , Genótipo , Proteína gp41 do Envelope de HIV/farmacologia , Infecções por HIV/virologia , HIV-1/classificação , HIV-1/isolamento & purificação , Humanos , Dados de Sequência Molecular , Mutação de Sentido Incorreto , Fragmentos de Peptídeos/farmacologia , Gravidez , Prevalência , Análise de Sequência de DNA , Adulto JovemRESUMO
The method used by YAGYU et al. for the subtype-specific polymerase chain reaction (PCR) amplification of the gp41 transmembrane region of the human immunodeficiency virus type-1 (HIV-1) env gene, was tested. HIV-1 proviral DNA from 100 infected individuals in Itajaí, South Brazil was used to analyze this method. Seventy individuals were determined according to this method as having PCR products at the expected size for subtypes B, C, D and F. Of these individuals, 26 (37.1%) were observed as having the expected amplification for subtype C, and 42 (60%) were observed as having the expected products for subtypes B and D. Of the subtype B and D amplicons, 16 (22.9%) were classified as subtype D, and 26 (37.1%) were classified as subtype B. Two individuals (2.9%) had amplicons that were observed after subtype F-specific amplification was performed. Sequencing and comparing the patient sequences to reference sequences confirmed the classification of sequences of subtypes C and B. However, sequences that were falsely determined as being D and F in the PCR assay were determined as being subtypes C and B, respectively, by sequence analysis. For those individuals from whom no amplified products were obtained, a low viral load that was indicated in their patient history may explain the difficulty in subtyping by PCR methods. This issue was demonstrated by the results of ANOVA when testing the effect of viral load on the success of PCR amplification. The alignment of the obtained sequences with HIV-1 reference sequences demonstrated that there is high intra-subtype diversity. This indicates that the subtype-specific primer binding sites were not conserved or representative of the subtypes that are observed in the Brazilian populations, and that they did not allow the correct classification of HIV-1 subtypes. Therefore, the proposed method by YAGYU et al. is not applicable for the classification of Brazilian HIV-1 subtypes.
Assuntos
DNA Viral/genética , Proteína gp41 do Envelope de HIV/genética , Infecções por HIV/virologia , HIV-1/genética , Adulto , Brasil , Feminino , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Reprodutibilidade dos Testes , Análise de Sequência de DNA , Carga Viral , Adulto JovemRESUMO
Entry of HIV-1 into a host cell is a multi-step process, with the viral envelope gp120 and gp41 acting sequentially to mediate the viral attachment, CD4 binding, coreceptor binding, and fusion of the viral and host membranes. The emerging class of antiretroviral agents, collectively known as entry inhibitors, interfere in some of these steps. However, viral diversity has implications for possible differential responses to entry inhibitors, since envelope is the most variable of all HIV genes. Different HIV genetic forms carry in their genomes genetic signatures and polymorphisms that could alter the structure of viral proteins which are targeted by drugs, thus impairing antiretroviral binding and efficacy. This review will examine current research that describes subtype differences in envelope at the genetic level and the effects of mutations on the efficacy of current entry inhibitors.
Assuntos
HIV-1/fisiologia , Produtos do Gene env do Vírus da Imunodeficiência Humana/química , Produtos do Gene env do Vírus da Imunodeficiência Humana/metabolismo , Farmacorresistência Viral/genética , Proteína gp120 do Envelope de HIV/química , Proteína gp120 do Envelope de HIV/metabolismo , Proteína gp41 do Envelope de HIV/química , Proteína gp41 do Envelope de HIV/metabolismo , Inibidores da Fusão de HIV/farmacologia , HIV-1/efeitos dos fármacos , Humanos , Internalização do Vírus/efeitos dos fármacos , Produtos do Gene env do Vírus da Imunodeficiência Humana/genéticaRESUMO
OBJECTIVE: One approach to identifying HIV-1 vaccine candidates is to dissect the natural antiviral immune response in treatment-naïve individuals infected for over ten years, considered slow progressor patients (SPs). It is suspected that SP plasma has strongly neutralizing antibodies (NAb) targeting specific HIV viral epitopes. METHODS: NAbs levels of 11 HIV-1-infected SPs were detected by PBMC-based neutralization assays. To investigate SP NAb epitope, this study used a biopanning approach to obtain mimotopes of HIV-1 that were recognized by SP plasma NAbs. IgG was purified from high-titer NAb SP plasma, and used as the ligand for three rounds of biopanning to select HIV-specific mimotopes from a phage-displayed random peptide library. Double-antibody sandwich ELISA, competitive inhibition assays, and peptide sequence analysis were used to evaluate the characteristics of phage-borne mimotopes. RESULTS: SPs had significantly more plasma neutralizing activity than typical progressors (TPs) (p=0.04). P2 and P9 plasma, which have highest-titer HIV-NAb, were selected as ligands for biopanning. After three rounds of biopanning, 48 phage clones were obtained, of which 22 clones were consistent with requirement, binding with HIV-1 positive plasma and unbinding with HIV-1 negative plasma. Compared with linear HIV-1 protein sequence and HIV-1 protein structure files, only 12 clones were possible linear mimotopes of NAbs. In addition, the C40 clone located in gp41 CHR was found to be a neutralizing epitope, which could inhibit pooled HIV-1 positive plasma reaction. CONCLUSION: Biopanning of serum IgG can yield mimotopes of HIV-1-related antigen epitopes. This methodology provides a basis for exploration into HIV-1-related antigen-antibody interactions and furthers NAb immunotherapy and vaccine design.
Assuntos
Anticorpos Neutralizantes/imunologia , Epitopos/imunologia , Anticorpos Anti-HIV/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , China , Reações Cruzadas , Ensaio de Imunoadsorção Enzimática , Proteína gp120 do Envelope de HIV/imunologia , Proteína gp41 do Envelope de HIV/imunologia , Humanos , Biblioteca de PeptídeosRESUMO
The present study describes a follow-up of a prospective and observational cohort of patients infected with HIV-1 and treated with raltegravir for salvage therapy in Brazil. Two groups of patients were analyzed: switching from T20 to RAL (Group 1, n = 9) and salvage therapy containing RAL (Group 2, n = 10). Blood samples were drawn for CD4(+) T-cell counts and HIV-1 viral load determinations. Protease, reverse transcriptase, and integrase genotyping were performed at baseline and at the time of virologic failure. CD4(+) T-cells increased at 6 and 12 months in both groups; HIV-1 viral load was continuously suppressed for Group 1, and for Group 2 it significantly decreased after starting a RAL-containing regimen. Three out of 10 patients from Group 2 could not suppress HIV-1 viral load. The mutations Q148H + G140S were observed for two patients and for the third patient only mutations to PR/RT inhibitors were detected. The genotypic sensitivity score (GSS) was analyzed for all patients of Group 2 and both patients who developed resistance to raltegravir presented a GSS < 2.0 for the RAL-containing scheme, which could be associated to the lack of effectiveness of the proposed scheme. The present study describes, for the first time in Brazil, the close follow-up of a series of patients using a raltegravir-containing HAART, showing the safety of the enfuvirtide switch to RAL and the effectiveness of a therapeutic regimen with RAL in promoting immune reconstitution and suppressing HIV replication, as well as documenting the occurrence of resistance to integrase inhibitors in the country.
Assuntos
Fármacos Anti-HIV/uso terapêutico , Farmacorresistência Viral/genética , Infecções por HIV/tratamento farmacológico , HIV-1/genética , Pirrolidinonas/uso terapêutico , Terapia de Salvação/métodos , Terapia Antirretroviral de Alta Atividade , Brasil/epidemiologia , Contagem de Linfócito CD4 , Cicloexanos/uso terapêutico , Enfuvirtida , Seguimentos , Técnicas de Genotipagem , Proteína gp41 do Envelope de HIV/uso terapêutico , Infecções por HIV/epidemiologia , Infecções por HIV/virologia , Integrase de HIV/genética , Protease de HIV/genética , Transcriptase Reversa do HIV/genética , HIV-1/enzimologia , HIV-1/fisiologia , Humanos , Maraviroc , Mutação , Fragmentos de Peptídeos/uso terapêutico , Estudos Prospectivos , RNA Viral/sangue , Raltegravir Potássico , Resultado do Tratamento , Triazóis/uso terapêutico , Carga Viral , Replicação ViralRESUMO
Antiretroviral naive patients (n=49) were recruited in central western Brazil (Campo Grande City/Mato Grosso do Sul State, located across the Bolivia and Paraguay borders). HIV-1 protease (PR), reverse transcriptase (RT), and env gp41 HR1 fragments were sequenced. Genetic diversity was analyzed by REGA/phylogenetic analyses. Intersubtype recombinants were identified by SimPlot/phylogenetic trees. PR/RT resistance was analyzed by Calibrated Population Resistance/Stanford databases. T-20 resistance in gp41 was assessed by Stanford, Los Alamos, and other sources. Of HIV-1 subtypes 65.3% were B(PR)B(RT), 10.2% were C(PR)C(RT), and 8.2% were F1(PR)F1(RT). Intersubtype recombinants were 16.3%: four B/F1 and four B/C (two were "CRF31_BC-like"). The Pol-RT V75M mutation was detected in two homosexual partners; one patient had the T215S revertant mutation. T-20/gp41 resistance mutations were L44M (n=2) and V38A (n=1). The high percentage of non-B isolates (â¼35%) highlights the importance of molecular surveillance studies in settings distant from the origin of the epidemic. Our data help elaborate the molecular epidemiological map of HIV-1 in Brazil.
Assuntos
Farmacorresistência Viral Múltipla/genética , Proteína gp41 do Envelope de HIV/genética , Soropositividade para HIV/virologia , HIV-1/genética , Mutação de Sentido Incorreto , Produtos do Gene pol do Vírus da Imunodeficiência Humana/genética , Adulto , Brasil/epidemiologia , Feminino , Variação Genética , Genótipo , Soropositividade para HIV/epidemiologia , HIV-1/isolamento & purificação , Humanos , Masculino , Pessoa de Meia-Idade , Epidemiologia Molecular , Dados de Sequência Molecular , Filogenia , RNA Viral , Análise de Sequência de DNARESUMO
OBJECTIVE: To evaluate the polymorphisms and resistance mutations in gp41 HR1 region of HIV-1. METHODS: The study included 28 HIV-positive patients undergoing enfuvirtide (ENF) treatment or not from Porto Alegre, Rio Grande do Sul state, and Rio de Janeiro, Rio de Janeiro state, between 2006 and 2009. Resistance mutations and polymorphisms of the gp41 HR1 region were detected using the genomic DNA of 12 ENF-untreated patients and 16 patients in ENF treatment, encompassing subtypes B, C, and F1. Sample subtypes were determined by neighbor-joining phylogenetic analysis with a Kimura's two-parameter correction. RESULTS: A high prevalence of polymorphisms unrelated to resistance was observed. Among ENF-untreated patients, 16% showed mutations related with resistance. Among patients in ENF treatment, 50% had resistance-related mutations. Overall, 17% of all isolates showed the N42S polymorphism related to ENF hypersusceptibility. The presence of ENF resistance mutations in the group of treated patients reduced viral load. The V38A substitution was the most frequent among treatment-experienced patients followed by the G36D/E, N42D, and V38M substitutions. CONCLUSIONS: The V38A substitution in the gp41 HR region was the most common resistance mutation among ENF-treated patients and was associated with increased viral load.
Assuntos
Fármacos Anti-HIV/farmacologia , Farmacorresistência Viral , Proteína gp41 do Envelope de HIV/genética , Proteína gp41 do Envelope de HIV/farmacologia , Infecções por HIV/virologia , HIV-1/genética , Fragmentos de Peptídeos/farmacologia , Polimorfismo Genético , Substituição de Aminoácidos/genética , Fármacos Anti-HIV/uso terapêutico , Análise por Conglomerados , Enfuvirtida , Genótipo , Proteína gp41 do Envelope de HIV/uso terapêutico , Infecções por HIV/tratamento farmacológico , HIV-1/isolamento & purificação , Humanos , Dados de Sequência Molecular , Mutação de Sentido Incorreto , Fragmentos de Peptídeos/uso terapêutico , Filogenia , Análise de Sequência de DNARESUMO
Entry inhibitor is a new class of drugs that target the viral envelope protein. This region is variable; hence resistance to these drugs may be present before treatment. The aim of this study was to analyze the frequency of patients failing treatment with transcriptase reverse and protease inhibitors that would respond to the entry inhibitors Enfuvirtide, Maraviroc, and BMS-806. The study included 100 HIV-1 positive patients from one outpatient clinic in the city of Sao Paulo, for whom a genotype test was requested due to treatment failure. Proviral DNA was amplified and sequenced for regions of gp120 and gp41. A total of 80 could be sequenced and from those, 73 (91.3%), 5 (6.3%) and 2 (2.5%) were classified as subtype B, F, and recombinants (B/ F and B/C), respectively. CXCR4 co-receptor use was predicted in 30% of the strains. Primary resistance to Enfuvirtide was found in 1.3%, following the AIDS Society consensus list, and 10% would be considered resistant if a broader criterion was used. Resistance to BMS-806 was higher; 6 (7.5%), and was associated to non-B strains. Strikingly, 27.5% of samples harbored one or more mutation among A316T, I323V, and S405A, which have been related to decreased susceptibility of Maraviroc; 15% of them among viruses predictive to be R5. A more common mutation was A316T, which was associated to the Brazilian B strain harboring the GWGR motif at the tip of V3 loop and their derivative sequences. These results may be impact guidelines for genotype testing and treatment in Brazil.
Assuntos
Cicloexanos/farmacologia , Farmacorresistência Viral , Inibidores da Fusão de HIV/farmacologia , Infecções por HIV/tratamento farmacológico , HIV-1/efeitos dos fármacos , Mutação de Sentido Incorreto , Triazóis/farmacologia , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética , Adulto , Instituições de Assistência Ambulatorial , Fármacos Anti-HIV/uso terapêutico , Brasil , DNA Viral/química , DNA Viral/genética , Enfuvirtida , Feminino , Proteína gp41 do Envelope de HIV/farmacologia , Inibidores da Fusão de HIV/administração & dosagem , Infecções por HIV/virologia , Inibidores da Protease de HIV , Transcriptase Reversa do HIV , HIV-1/isolamento & purificação , Humanos , Masculino , Maraviroc , Pessoa de Meia-Idade , Dados de Sequência Molecular , Fragmentos de Peptídeos/farmacologia , Piperazinas/farmacologia , Prevalência , Provírus/genética , Análise de Sequência de DNA , Falha de TratamentoRESUMO
The study was carried out in a Hospital of Reference in Infectious Diseases of Ceará, from February 2006 to February 2007, and and aimed at identifying the socio-demographic profile of Enfuvirtide users and their main difficulties/facilities in previous and current treatment scheme. The initial sample analyzed the medical record of 23 patients;18 agreed to participate, comprising the final sample. The descriptive analysis of quantitative data was carried out through the distribution of frequencies and quantitative data, submitted for content analysis. It was observed that 83% were male, 78% were single and the majority was between 30 and 52 years old and, in average, eight years and a half of antiretroviral treatment. From qualitative data, two categories emerged: (1) Previous treatment: difficulties and adversities and (2) Current treatment: from cognition to ability. The difficulties to conduct previous treatments were related to the size and high amount of tablets and side effects. As for facility, the easy drug administration was indicated. Regarding the current treatment, the difficulties were self administration and nodules on the sites where Enfuvirtide was applied and the facilities were absence of gastrointestinal effects and improvement of viral load. It's important to implement an interdisciplinary work that helps patients overcome the difficulties of the treatment, in addition to works in groups in order to better address the difficulties and help increase adhesion to treatment.
Assuntos
Síndrome da Imunodeficiência Adquirida/tratamento farmacológico , Proteína gp41 do Envelope de HIV/uso terapêutico , Inibidores da Fusão de HIV/uso terapêutico , Fragmentos de Peptídeos/uso terapêutico , Adulto , Enfuvirtida , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
The aim of this study was to determine the rate and risk factors of HIV-1 mother-to-child transmission (MTCT), the timing of transmission and the transmitted subtype in a population where subtypes B and C co-circulate. One hundred and forty-four babies born to HIV-1-infected mothers were studied. Subtype and timing of transmission were determined by a nested polymerase chain reaction of the gp41 gene. Seven children were infected (4.9%): four were infected intrautero and one intrapartum. The higher frequency of intrautero transmission was statistically significant (P = 0.001). Use of antiretrovirals (ARVs) in the three stages of gestation was a protective risk factor for MTCT (PR = 0.42; CI: 0.21-0.83; P = 0.013). A higher HIV viral load at delivery was the only independent risk factor for MTCT. Early and universal access to ARVs during pregnancy are the most important measures to decrease vertical HIV-1 transmission even in areas where HIV clade distribution differs.
Assuntos
Infecções por HIV/transmissão , HIV-1 , Transmissão Vertical de Doenças Infecciosas , Adulto , Antirretrovirais/uso terapêutico , Brasil , Feminino , Proteína gp41 do Envelope de HIV/genética , Humanos , Recém-Nascido , Transmissão Vertical de Doenças Infecciosas/prevenção & controle , Masculino , Reação em Cadeia da Polimerase , Gravidez , Carga ViralRESUMO
We analyzed the gp41 sequences of 80 HIV-infected enfuvirtide-naive individuals who were eligible to receive this antiretroviral according to Brazilian guidelines. We analyzed the genetic diversity of pol and the heptad repeat 1 and 2 (HR1 and HR2) regions of gp41, and compared the genetic profile of HR1 and HR2 found in PBMCs with the profile found in plasma. The similarity between sequences obtained from DNA and RNA in the HR1 and HR2 regions was, on average, 98.6% and 98.9%, respectively. We detected mutations related to enfuvirtide resistance (L44M or N43K) in HR1 DNA samples from three individuals (3.8%) and RNA samples from three individuals (4.6%). Other polymorphisms frequently detected were E137K (10% and 13.8%), L130I (8.8% and 9.2%), S129N (6.3% and 10.8%), L44M (2.5% and 4.6%), S138A (2.5% and 1.5%), and N43K (1.3% and 0%) in DNA and RNA, respectively. Subtype B was identified in 68.8% of the samples [protease (PR) B, reverse transcriptase (RT) B, gp41 B], subtype F in 5.0%, subtype C in 1.3%, and the remaining sequences presented with a mosaic profile. These results suggest that genotyping the gp41 region prior to introducing an expensive and complex approach, such as enfuvirtide, may be cost effective. Moreover, assessment of proviral DNA may be less expensive than RNA, as well as being sufficient for this purpose.
Assuntos
Farmacorresistência Viral/genética , Proteína gp41 do Envelope de HIV/genética , Inibidores da Fusão de HIV/uso terapêutico , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , HIV-1/genética , Fragmentos de Peptídeos/uso terapêutico , Polimorfismo de Nucleotídeo Único , Adulto , Brasil/epidemiologia , DNA Viral/análise , DNA Viral/sangue , DNA Viral/genética , Enfuvirtida , Variação Genética , Proteína gp41 do Envelope de HIV/uso terapêutico , Infecções por HIV/sangue , Infecções por HIV/epidemiologia , HIV-1/efeitos dos fármacos , HIV-1/metabolismo , Humanos , Leucócitos Mononucleares/virologia , Pessoa de Meia-Idade , RNA Viral/análise , RNA Viral/sangue , RNA Viral/genética , Resultado do Tratamento , Produtos do Gene pol do Vírus da Imunodeficiência Humana/genéticaRESUMO
HIV-1 resistance mutations to antiretroviral (ARV) drugs (nucleoside reverse transcriptase inhibitors, NRTI; nonnucleoside reverse transcriptase inhibitors, NNRTI; and protease inhibitors, PI) represent a challenge for sustainable virologic and immunologic responses. HIV-1 phylogenetic diversity and ARV resistance mutations associated with protease (PR) and reverse transcriptase (RT) were assessed among 48 ARV-experienced patients from Goiania/Goias, central west Brazil. The gp41 resistance mutations to the fusion inhibitor (T-20) were analyzed among multidrug-resistant (MDR) patients. PR, partial RT, and gp41 genes were amplified and sequenced from plasma RNA. HIV-1 subtype was assigned by phylogenetic analysis. ARV mutations in PR/RT were analyzed by the Stanford Database. T-20 resistance mutations in gp41 were identified by Stanford, the Los Alamos Database, and other sources. Subtype B represented 79.2% (38/48), subtype F1 4.2% (2/48), subtype C 2.1% (1/48), F1(PR)/B(RT) 8.3% (4/48), and B(PR)/F1(RT) 6.3% (3/48). Secondary drug resistance was observed in 79% (38/48): NRTI resistance (n = 1), NNRTI resistance (n = 1), PI + NRTI or NRTI + NNRTI resistance (n = 20), and PI + NRTI +NNRTI resistance, considered MDR (n = 16). Almost half of the MDR patients had viral loads below 10,000 copies/ml. The gp41 sequences from 14 MDR revealed one F1(PR)/B(RT)/F1(ENV) recombinant and 13 subtype B(PR)/B(RT)/B(ENV) isolates. G36E, N42T, and N43S T-20 resistance mutations were observed in three MDR patients, two of them previously treated with T-20 and the other who had never used T-20. Our data among ARV-experienced patients showed a high proportion of drug-resistance mutations and MDR. T-20 resistance mutations were detected among MDR patients corroborating the importance of T-20 genotyping for clinical management and salvage therapy.