RESUMO
Tenofovir (TFV) disoproxil fumarate (TDF) and emtricitabine (FTC), two nucleos(t)ide analogs (NA), are coformulated as an anti-HIV combination tablet for treatment and preexposure prophylaxis (PrEP). TDF/FTC may have effects on the deoxynucleoside triphosphate (dNTP) pool due to their similar structures and similar metabolic pathways. We carried out a comprehensive clinical study to characterize the effects of TDF/FTC on the endogenous dNTP pool, from baseline to 30 days of TDF/FTC therapy, in both treatment-naive HIV-positive and HIV-negative individuals. dATP, dCTP, dGTP, and TTP were quantified in peripheral blood mononuclear cells (PBMC) with a validated liquid chromatography-tandem mass spectrometry (LC-MS/MS) methodology. Forty individuals (19 HIV-positive) were enrolled and underwent a baseline visit and then received TDF/FTC for at least 30 days. Longitudinal measurements were analyzed using mixed-model segmented linear regression analysis. The dNTPs were reduced by 14% to 37% relative to the baseline level within 3 days in both HIV-negative and HIV-positive individuals (P ≤ 0.003). These reductions persisted to various degrees at day 30. These findings indicate that dNTP pools are influenced by TDF/FTC therapy. This may alter cellular homeostasis and could increase the antiviral effect through a more favorable analog/dNTP ratio. Further work is needed to elucidate mechanisms, to evaluate the clinical significance of these findings, and to further probe differences between HIV-negative and HIV-positive individuals. (This study has been registered at ClinicalTrials.gov under identifier NCT01040091.).
Assuntos
Fármacos Anti-HIV/farmacologia , Nucleotídeos de Desoxiadenina/sangue , Nucleotídeos de Desoxicitosina/sangue , Nucleotídeos de Desoxiguanina/sangue , Emtricitabina/farmacologia , Infecções por HIV/tratamento farmacológico , Tenofovir/farmacologia , Nucleotídeos de Timina/sangue , Adulto , Estudos de Casos e Controles , Nucleotídeos de Desoxiadenina/antagonistas & inibidores , Nucleotídeos de Desoxicitosina/antagonistas & inibidores , Nucleotídeos de Desoxiguanina/antagonistas & inibidores , Feminino , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , HIV-1/fisiologia , Humanos , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/virologia , Modelos Lineares , Masculino , Nucleotídeos de Timina/antagonistas & inibidoresRESUMO
BACKGROUND: A series of antioxidant enzymes and non-enzymatic compounds act to protect cells from uncontrolled propagation of free radicals. It is poorly understood, though, to what extent and how their interaction is harmonized. OBJECTIVES: To explore associative interactions among a battery of urinary and blood biomarkers of oxidative stress and enzymatic and non-enzymatic markers of the antioxidant defense system in children from low income households. METHODS: For this cross-sectional descriptive study, urine, red cells, and plasma were sampled in 82 preschool children attending three daycare centers in Quetzaltenango Guatemala. The urinary oxidative stress biomarkers studied were F2-isoprostanes and 8-hydroxy-deoxy-guanosine. Red cell enzyme activities measured were: catalase, superoxide dismutase, glutathione peroxidase and glutathione reductase. Circulating non-enzymatic antioxidants selected were: retinol, tocopherols, ß-carotene and coenzymes Q9 and Q10. RESULTS: In a Spearman rank-order correlation hemi-matrix, of 55 paired combinations of the 11 biomarkers, 28 (51%) were significantly correlated among each other (p ≤ 0.05), with the strongest association being retinol and tocopherols (r = 0.697, p<0.001), and 4 associations (9%) showed a trend (p> 0.5 to ≤ 0.10). F2-isoprostanes showed the greatest number of cross-associations, having significant interactions with 8 of the 10 remaining biomarkers. Goodness-of-fit modeling improved or maintained the r value for 24 of the significant interactions and for one of the 5 borderline associations. Multiple regression backward stepwise analysis indicated that plasma retinol, ß-carotene and coenzyme Q10 were independent predictors of urinary F2-isoprostanes. CONCLUSION: Numerous significant associations resulted among biomarkers of oxidation and responders to oxidation. Interesting findings were the apparent patterns of harmonious interactions among the elements of the oxidation-antioxidation systems in this population.
Assuntos
Antioxidantes/metabolismo , Nucleotídeos de Desoxiguanina , F2-Isoprostanos , Estresse Oxidativo , Oxirredutases/sangue , 8-Hidroxi-2'-Desoxiguanosina , Biomarcadores/sangue , Biomarcadores/urina , Criança , Pré-Escolar , Estudos Transversais , Nucleotídeos de Desoxiguanina/sangue , Nucleotídeos de Desoxiguanina/urina , F2-Isoprostanos/sangue , F2-Isoprostanos/urina , Feminino , Guatemala , Humanos , Masculino , Fatores SocioeconômicosRESUMO
Abacavir (ABC) has been associated with ischaemic cardiovascular events in HIV-infected patients, but the pathogenic mechanisms are unknown. Aim of our study was to assess whether ABC induces in vivo platelet activation and ex vivo platelet hyper-reactivity. In a retrospective, case-control study, in vivo platelet activation markers were measured in 69 HIV-infected patients, before starting therapy and after 6-12 months of either ABC (n=35) or tenofovir (TDF) (n=34), and compared with those from 20 untreated HIV-infected patients. A subgroup of patients was restudied after 28-34 months for ex vivo platelet reactivity. In vivo platelet activation markers were assessed by ELISA or flow cytometry, ex vivo platelet reactivity by light transmission aggregometry (LTA) and PFA-100®. Thein vitro effects of the ABC metabolite, carbovir triphosphate, on aggregation and intra-platelet cGMP were also studied. sPLA2, sPsel and sGPV increased significantly 6-12 months after the beginning of ABC, but not of TDF or of no treatment. Ex vivo platelet function studies showed enhanced LTA, shorter PFA-100® C/ADP closure time and enhanced platelet expression of P-sel and CD40L in the ABC group. The intake of ABC blunted the increase of intraplatelet cGMP induced by nitric oxide (NO) and acutely enhanced collagen-induced aggregation. Preincubation of control platelets with carbovir triphosphate in vitro enhanced platelet aggregation and blunted NO-induced cGMP elevation. In conclusion, treatment with ABC enhances in vivo platelet activation and induces platelet hyperreactivity by blunting the inhibitory effects of NO on platelets. These effects may lead to an increase of ischaemic cardiovascular events.
Assuntos
Fármacos Anti-HIV/efeitos adversos , Didesoxinucleosídeos/efeitos adversos , Infecções por HIV/sangue , Infecções por HIV/tratamento farmacológico , Ativação Plaquetária/efeitos dos fármacos , Adenina/efeitos adversos , Adenina/análogos & derivados , Adulto , Fármacos Anti-HIV/sangue , Plaquetas/efeitos dos fármacos , Plaquetas/metabolismo , Ligante de CD40 , Estudos de Casos e Controles , GMP Cíclico/sangue , Nucleotídeos de Desoxiguanina/efeitos adversos , Nucleotídeos de Desoxiguanina/sangue , Didesoxinucleosídeos/sangue , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Óxido Nítrico/sangue , Organofosfonatos/efeitos adversos , Selectina-P/sangue , Fosfolipases A2 Secretórias/sangue , Agregação Plaquetária/efeitos dos fármacos , Estudos Retrospectivos , TenofovirRESUMO
BACKGROUND: Here, we aimed to investigate the pharmacokinetics of abacavir and carbovir triphosphate (CBV-TP) with darunavir/ritonavir 900/100 mg once daily or raltegravir 400 mg twice daily. METHODS: HIV-infected subjects on abacavir (600 mg once daily) underwent steady-state pharmacokinetic assessments without and with darunavir/ritonavir or raltegravir. Within-subject changes in plasma and intracellular pharmacokinetic parameters were evaluated by geometric mean ratios (GMRs) and 90% CIs. RESULTS: A total of 19 patients completed the study. With darunavir/ritonavir (versus abacavir alone), abacavir GMRs (90% CI) were 0.73 (0.66, 0.80), 0.62 (0.50, 0.77) and 0.78 (0.69, 0.87) for area under the curve (AUC), trough concentration (C(trough)) and maximum concentration (C(max)), respectively. With raltegravir, they were 1.03 (0.97, 1.10), 0.83 (0.62, 1.11) and 1.06 (0.95, 1.18), respectively. Intracellular CBV-TP GMRs (90% CI) were 0.88 (0.72, 1.07), 0.68 (0.48, 0.95) and 0.98 (0.79, 1.23) for AUC, C(trough) and C(max), respectively, with darunavir/ritonavir, and 0.96 (0.76, 1.20), 0.57 (0.33, 1.00) and 1.07 (0.85, 1.35), respectively, with raltegravir. CONCLUSIONS: There was a 27% decrease in abacavir plasma exposure with darunavir/ritonavir and no changes with raltegravir. CBV-TP C(trough) was significantly decreased with darunavir/ritonavir (32%) and showed a high inter-individual variability with raltegravir.
Assuntos
Nucleotídeos de Desoxiguanina/farmacocinética , Didesoxinucleosídeos/farmacocinética , Infecções por HIV/tratamento farmacológico , Pirrolidinonas/farmacocinética , Ritonavir/farmacocinética , Sulfonamidas/farmacocinética , Adulto , Fármacos Anti-HIV/sangue , Fármacos Anti-HIV/farmacocinética , Área Sob a Curva , Estudos Cross-Over , Darunavir , Nucleotídeos de Desoxiguanina/sangue , Didesoxinucleosídeos/sangue , Esquema de Medicação , Quimioterapia Combinada , Feminino , Infecções por HIV/sangue , Infecções por HIV/virologia , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Pirrolidinonas/sangue , RNA Viral/análise , Raltegravir Potássico , Ritonavir/sangue , Sulfonamidas/sangue , Carga Viral/efeitos dos fármacosRESUMO
Studies were conducted to determine if there is a mechanistic basis for reports of suboptimal virologic responses and concerns regarding the safety of regimens containing the combination of tenofovir (TFV) disoproxil fumarate (TDF) and didanosine (ddI) by assessing the pharmacokinetic consequences of coadministration of these drugs on intracellular nucleotides. This was a prospective and longitudinal study in HIV-1-infected patients of adding either TDF or ddI to a stable antiretroviral regimen containing the other drug. Intracellular concentrations of the nucleotide analogs TFV diphosphate (TFV-DP) and ddATP and the endogenous purine nucleotides dATP and 2'-dGTP in peripheral blood mononuclear cells were measured. A total of 16 patients were enrolled into the two study arms and a study extension. Intracellular TFV-DP concentrations (median, 120 fmol/10(6) cells) and ddATP concentrations (range, 1.50 to 7.54 fmol/10(6) cells in two patients) were unaffected following addition of ddI or TDF to a stable regimen containing the other drug. While coadministration of ddI and TDF for 4 weeks did not appear to impact dATP or dGTP concentrations, cross-sectional analysis suggested that extended therapy with ddI-containing regimens, irrespective of TDF coadministration, may decrease dATP and ddATP concentrations. Addition of TDF or ddI to a stable regimen including the other drug, in the context of ddI dose reduction, did not adversely affect the concentration of dATP, dGTP, TFV-DP, or ddATP. The association between longer-term ddI therapy and reduced intracellular nucleotide concentrations and this observation's implication for the efficacy and toxicity of ddI-containing regimens deserve further study.
Assuntos
Adenina/análogos & derivados , Fármacos Anti-HIV/farmacocinética , Fármacos Anti-HIV/uso terapêutico , Didanosina/farmacocinética , Didanosina/uso terapêutico , Infecções por HIV/tratamento farmacológico , Nucleotídeos/sangue , Organofosfonatos/farmacocinética , Organofosfonatos/uso terapêutico , Adenina/farmacocinética , Adenina/uso terapêutico , Adulto , Idoso , Cromatografia Líquida , Nucleotídeos de Desoxiadenina/sangue , Nucleotídeos de Desoxiguanina/sangue , Didesoxinucleotídeos/sangue , Feminino , Infecções por HIV/sangue , Humanos , Masculino , Pessoa de Meia-Idade , Espectrometria de Massas em Tandem , Tenofovir , Adulto JovemRESUMO
OBJECTIVES: To evaluate a potential pharmacodynamic/pharmacokinetic interaction between abacavir (ABC) and tenofovir disoproxil fumarate (TDF). DESIGN AND METHODS: This randomized trial compared 7 days of ABC or TDF monotherapy, separated by a 35-day washout, with 7 days of ABC + TDF dual-therapy in treatment-naive, HIV-1-infected patients. During each 7-day course, the slope of the phase I viral decay was estimated and steady-state intracellular concentrations of carbovir triphosphate (CBV-TP), deoxyguanosine triphosphate (dGTP), tenofovir diphosphate (TFV-DP) and deoxyadenosine triphosphate (dATP) were determined. RESULTS: Twenty-one participants were randomized to initial monotherapy with ABC (n = 11) or TDF (n = 10). The addition of TDF did not increase the slope of viral decay compared to ABC alone (-0.15 log10 per day vs. -0.16 log10 per day, respectively). No decrease in CBV-TP or TFV-DP between monotherapy and dual-therapy was observed. However, intracellular dATP concentrations increased between monotherapy and dual-therapy [median dATP (fmol/10 cells) 3293 vs. 4638; P = 0.08], although this difference was significant only among patients randomized to TDF [median dATP (fmol/10 cells) 3238 vs. 4534; P = 0.047]. A lower TFV-DP-to-dATP ratio was associated with reduced viral decay during dual-therapy (rho = -0.529; P = 0.045). CONCLUSION: In this study, the viral decay during ABC and TDF dual-therapy was similar to that during ABC therapy alone, suggesting a nonadditive antiviral effect. This negative pharmacodynamic interaction was not explained by changes in CBV-TP or TFV-DP concentrations. Rather, modest increases in endogenous dATP pools were associated with reduced antiviral potency of TDF during co-administration with ABC.
Assuntos
Adenina/análogos & derivados , Fármacos Anti-HIV/farmacocinética , Didesoxinucleosídeos/farmacocinética , Infecções por HIV/tratamento farmacológico , HIV-1/efeitos dos fármacos , Organofosfonatos/farmacocinética , RNA Viral/efeitos dos fármacos , Adenina/administração & dosagem , Adenina/farmacocinética , Adulto , Fármacos Anti-HIV/administração & dosagem , Nucleotídeos de Desoxiguanina/sangue , Didesoxinucleosídeos/administração & dosagem , Interações Medicamentosas , Quimioterapia Combinada , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Organofosfonatos/administração & dosagem , Tenofovir , Resultado do TratamentoAssuntos
Nucleotídeos de Desoxiguanina/sangue , Infecções por HIV/tratamento farmacológico , Líquido Intracelular/química , Adulto , Coleta de Amostras Sanguíneas/normas , Técnicas de Laboratório Clínico/normas , Didesoxinucleosídeos/sangue , Infecções por HIV/sangue , Humanos , Leucócitos Mononucleares/química , Masculino , Pessoa de Meia-Idade , Reprodutibilidade dos TestesRESUMO
Our previous negative ESI-LC-MS/MS method developed for nucleoside reverse transcriptase inhibitor (NRTI) triphosphate (-TP) measurements in human peripheral blood mononuclear cells (PBMC) encountered some specificity problems for several NRTI-TP and simultaneous endogenous nucleotide triphosphates analysis. As LC-MS/MS offers several possibilities to circumvent such problems, we have investigated the contribution of the positive electrospray ionization mode in enhancing the specificity of the intracellular analyses of triphosphate metabolites of lamivudine, abacavir, and tenofovir. For intracellular NRTI-TP analysis, after disruption of PBMCs, concentrated supernatants were directly injected into the LC-MS/MS system, dimethylhexylamine being used as ion-pairing agent to resolve NRTI-TP. MS/MS detection was performed after positive electrospray ionization. Total run time was 12 min instead of 26 min for NRTI-TP analysis. The validation parameters of the method met the international requirements, and endogenous chromatographic interferences were eliminated. The use of positive ESI, offering a better specificity and a slightly better sensitivity than the negative ESI mode for these compounds, resulted in specificity enhancement and more robust assay methods.
Assuntos
Adenina/análogos & derivados , Citidina Trifosfato/análogos & derivados , Nucleotídeos de Desoxiguanina/sangue , Didesoxinucleotídeos/sangue , Lamivudina/análogos & derivados , Leucócitos Mononucleares/química , Organofosfonatos/sangue , Espectrometria de Massas por Ionização por Electrospray/métodos , Espectrometria de Massas em Tandem/métodos , Adenina/sangue , Adenina/química , Fármacos Anti-HIV/farmacocinética , Citidina Trifosfato/sangue , Didesoxinucleosídeos/farmacocinética , Infecções por HIV/metabolismo , Humanos , Lamivudina/sangue , Lamivudina/farmacocinética , Leucócitos Mononucleares/metabolismo , Organofosfonatos/química , Reprodutibilidade dos Testes , TenofovirRESUMO
Arterial dysfunction is a hallmark of early atherosclerosis; however, its behavior in patients with metabolic syndrome (MS) is still unclear. We investigated the role of oxidative stress on ischemia-induced flow-mediated dilatation (FMD) in patients with MS. FMD and oxidative stress, as assessed by serum levels of 8-hydroxy-2-deoxy-2-deoxyguanosine (8-OHdG), were studied in 18 MS and 30 control subjects. Thereafter, in the 18 MS patients, FMD was assessed after iv infusion of 1 g vitamin C or placebo in a randomized, double-blind, crossover design; serial blood samples were taken in peripheral circulation before and after FMD to analyze 8-OHdG. Compared to controls, MS patients had higher 8-OHdG (p<0.001) and lower FMD (p<0.001); 8-OHdG and FMD were inversely correlated (R=-0.74; p<0.01). In MS patients, placebo administration did not change FMD, whereas vitamin C significantly enhanced it (p<0.001). After placebo, ischemia-induced FMD was associated with a significant increase in 8-OHdG (p<0.001), an effect that was counteracted by vitamin C. Vitamin C infusion was associated with an inverse correlation between the changes in FMD and oxidative stress (R=-0.67; p<0.01). The present study shows that arterial dilatation is impaired and that enhanced oxidative stress may play a key role in patients with MS.
Assuntos
Ácido Ascórbico/uso terapêutico , Síndrome Metabólica/tratamento farmacológico , Síndrome Metabólica/patologia , Estresse Oxidativo , 8-Hidroxi-2'-Desoxiguanosina , Adulto , Idoso , Artérias/efeitos dos fármacos , Artérias/patologia , Estudos Transversais , Nucleotídeos de Desoxiguanina/sangue , Humanos , Masculino , Síndrome Metabólica/sangue , Síndrome Metabólica/epidemiologia , Pessoa de Meia-IdadeRESUMO
In this paper, we describe the development and use of enzymatic assays to determine intracellular lamivudine triphosphate (3TCTP) and carbovir triphosphate (CBVTP) concentrations in peripheral blood mononuclear cells (PBMCs) from human immunodeficiency virus (HIV)-infected patients. The assays involve inhibition of HIV reverse transcriptase (RT), which normally incorporates radiolabeled deoxynucleoside triphosphates into a synthetic template primer. For the 3TCTP assay, a preincubation procedure was added whereby 3TCTP becomes incorporated before [(3)H]dCTP. At a 1:400 template primer dilution, control product formation was reduced by 88.0% with 0.8 pmol of 3TCTP. Standard 3TCTP inhibition curves were performed using this procedure. For the CBVTP assay, 0.1 pmol of CBVTP inhibited control product formation with and without the use of a preincubation step, so inhibition curves were constructed using both procedures. However, reduced template primer stability with assays using preincubation steps led to a single-incubation procedure being adopted for future studies. The presence of PBMC extracts interfered with the 3TCTP assay. However, this was overcome by the addition of CuSO(4). PBMC extracts did not interfere with the CBVTP assay. Intracellular 3TCTP and CBVTP concentrations were determined in PBMCs from HIV-infected patients over 24 h or greater. Peak concentrations were obtained 6 to 8 h after dosing, and the half-lives of the anabolites suggested the possibility of once-daily dosing. These assays are currently being used for determination of 3TCTP and CBVTP concentrations in clinical studies.
Assuntos
Técnicas de Química Analítica/métodos , Nucleotídeos de Desoxiguanina/sangue , Infecções por HIV/sangue , Lamivudina/análise , Infecções por HIV/metabolismo , Humanos , Controle de Qualidade , Padrões de ReferênciaRESUMO
Using HPLC methods, we measured the concentrations of nucleosides and nucleotides for a patient with no purine nucleoside phosphorylase (PNP; EC 2.4.2.1) enzymatic activity. Concentrations of inosine and guanosine were abnormally high in urine and plasma, whereas guanosine diphosphate (GDP) and guanosine triphosphate (GTP) concentrations in erythrocytes were depleted. The unusual presence of deoxyribonucleosides (deoxyinosine and deoxyguanosine) and deoxyribonucleotides (dGDP and dGTP) was also notable. Thus, HPLC represents an accurate and useful tool for the study of purine metabolic disorders.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Purina-Núcleosídeo Fosforilase/deficiência , Erros Inatos do Metabolismo da Purina-Pirimidina/metabolismo , Nucleotídeos de Desoxiadenina/sangue , Nucleotídeos de Desoxiguanina/sangue , Eritrócitos/metabolismo , Guanosina/sangue , Guanosina/urina , Guanosina Difosfato/sangue , Guanosina Trifosfato/sangue , Humanos , Lactente , Inosina/sangue , Inosina/urina , MasculinoRESUMO
Cell-mediated immunity in human subjects is affected adversely as a result of zinc deficiency. The mechanism by which a deficiency of zinc may affect lymphocyte proliferation and functions, is not well understood at present. Nucleoside phosphorylase (NPase), a purine catabolic pathway enzyme, is zinc dependent, and a congenital deficiency of this enzyme is known to affect adversely cell-mediated immunity. This effect has been related to an accumulation of toxic nucleotides in lymphocytes as a result of NPase deficiency. Inasmuch as the effect of zinc deficiency on the activity of NPase and the levels of nucleotides in human lymphocytes has not been previously reported, we assayed these parameters in human subjects with zinc deficiency before and after zinc supplementation. A mild deficiency of zinc was diagnosed in those having decreased zinc in two out of three cell lineages (less than 42 micrograms in granulocytes, less than 48 micrograms in lymphocytes, and less than 1.70 microgram in platelets, per 10(10) cells). In comparison with five subjects with sufficient zinc, six subjects with zinc deficiency showed a decrease in the activity of NPase (p = 0.01), an increase in adenosine diphosphate (ADP) level (p = 0.008), a decreased adenosine triphosphate (ATP)-to-ADP ratio (p = 0.0001), and an increase in both guanosine triphosphate (GTP) (p = 0.02) and deoxyadenosine triphosphate (dGTP) (p = 0.04 in the lymphocytes.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Linfócitos/metabolismo , Nucleotídeos/sangue , Pentosiltransferases/sangue , Purina-Núcleosídeo Fosforilase/sangue , Zinco/deficiência , Difosfato de Adenosina/sangue , Trifosfato de Adenosina/sangue , Plaquetas/metabolismo , Nucleotídeos de Desoxiguanina/sangue , Granulócitos/metabolismo , Guanosina Trifosfato/sangue , Humanos , Purina-Núcleosídeo Fosforilase/deficiência , Zinco/uso terapêuticoRESUMO
Low ATP/ADP ratios have been reported consistently for nucleotide levels of mononuclear cells separated from peripheral blood by conventional techniques. We have established that these low values (mean 2.3:1) were not due to cell damage or poor viability, but resulted from heavy platelet contamination, which is unavoidable when heparinized blood is used. The results reflect the low ATP/ADP ratios (mean 1.6:1) characteristic of platelets. Platelet-free extracts from defibrinated blood had very high ATP/ADP ratios (mean 17.4:1). The initial finding of detectable amounts of deoxy-ATP and deoxy-GTP in mononuclear cells from children with two distinct inherited immunodeficiency disorders [adenosine deaminase (ADA) and purine nucleoside phosphorylase (PNP) deficiency respectively] many have been due to contamination by nucleated erythrocytes as well as platelets in non-defibrinated preparations. Defibrination before nucleotide extraction of mononuclear cells from a patient with T-cell leukaemic/lymphoma treated with the ADA inhibitor deoxycoformycin enabled the demonstration of grossly raised deoxy-ATP levels relative to deoxy-ADP levels (ratio 16.1:1), associated with severe ATP depletion. This reciprocal relationship between ATP and dATP was found by us previously in the erythrocytes in inherited ADA deficiency. These findings underline the importance of extracts uncontaminated by platelets, or nucleated erythrocytes, in the evaluation of lymphocyte nucleotide levels in inherited or acquired immunodeficiency syndromes.
Assuntos
Difosfato de Adenosina/sangue , Trifosfato de Adenosina/sangue , Síndromes de Imunodeficiência/sangue , Linfócitos/metabolismo , Adenosina Desaminase/deficiência , Inibidores de Adenosina Desaminase , Plaquetas/metabolismo , Separação Celular , Criança , Coformicina/análogos & derivados , Coformicina/farmacologia , Nucleotídeos de Desoxiadenina/sangue , Nucleotídeos de Desoxiguanina/sangue , Humanos , Pentostatina , Purina-Núcleosídeo Fosforilase/deficiênciaAssuntos
Adenosina Desaminase/deficiência , Síndromes de Imunodeficiência/enzimologia , Nucleosídeo Desaminases/deficiência , Pentosiltransferases/deficiência , Purina-Núcleosídeo Fosforilase/deficiência , Regulação Alostérica , Nucleotídeos de Desoxiadenina/sangue , Nucleotídeos de Desoxiguanina/sangue , Humanos , Ribonucleotídeo Redutases/antagonistas & inibidores , S-Adenosilmetionina/metabolismoRESUMO
Purine nucleoside phosphorylase (PNP) deficiency is associated with a severe defect in thymus-derived (T)-lymphocyte function combined with normal bone marrow-derived (B)-lymphocyte function. To investigate the role of this enzyme deficiency in the resulting immune dysfunction, we measured the levels of ribonucleoside and deoxyribonucleoside triphosphates in erythrocytes from two unrelated PNP-deficient, T-lymphocyte-deficient patients. Both PNP-deficient patients have abnormally high levels of deoxyguanosine triphosphate (deoxy-GTP) in their erythrocytes (5 and 8 nmol/ml packed erythrocytes). In contrast, normal controls and adenosine deaminase-deficient, immunodeficient patients do not have detectable amounts of deoxyGTP (<0.5 nmol/ml packed erythrocytes). We propose that deoxyguanosine, a substrate of PNP, is the potentially lymphotoxic metabolite in PNP deficiency. The mechanism of toxicity involves phosphorylation of deoxyguanosine to deoxyGTP, which acts as a potent inhibitor of mammalian ribonucleotide reductase.