Clinical oxidative stress during leprosy multidrug therapy:impact of dapsone oxidation

This study aims to assess the oxidative stress in leprosy patients under multidrug therapy (MDT; dapsone, clofazimine and rifampicin), evaluating the nitric oxide (NO) concentration, catalase (CAT) and superoxide dismutase (SOD) activities, glutathione (GSH) levels, total antioxidant capacity, lipid peroxidation, and methemoglobin formation. For this, we analyzed 23 leprosy patients and 20 healthy individuals from the Amazon region, Brazil, aged between 20 and 45 years. Blood sampling enabled the evaluation of leprosy patients prior to starting multidrug therapy (called MDT 0) and until the third month of multidrug therapy (MDT 3). With regard to dapsone (DDS) plasma levels, we showed that there was no statistical difference in drug plasma levels between multibacillary (0.518±0.029 μg/mL) and paucibacillary (0.662±0.123 μg/mL) patients. The methemoglobin levels and numbers of Heinz bodies were significantly enhanced after the third MDTsupervised dose, but this treatment did not significantly change the lipid peroxidation and NO levels in these leprosy patients. In addition, CAT activity was significantly reduced in MDT-treated leprosy patients, while GSH content was increased in these patients. However, SOD and Trolox equivalent antioxidant capacity levels were similar in patients with and without treatment. These data suggest that MDT can reduce the activity of some antioxidant enzyme and influence ROS accumulation, which may induce hematological changes, such as methemoglobinemia in patients with leprosy. We also explored some redox mechanisms associated with DDS and its main oxidative metabolite DDS-NHOH and we explored the possible binding of DDS to the active site of CYP2C19 with the aid of molecular modeling software

Dapsone effect in generation of oxidative stress in patients with leprosy in use of multidrug therapy

Inflammation caused by Mycobacterium leprae infection and drugs with oxidative properties such as dapsone, are risk factors to induce the oxidative stress in leprosy patients. This study aims to determine plasma concentrations of dapsone in leprosy patients in use of multidrug therapy (MDT), correlating the development of oxidative stress. For the study, healthy individuals and leprosy patients were selected, followed before (D0) and after the third MDT-supervised dose (D3). The plasma concentrations of dapsone in patients under treatment (D3) were evaluated by high performance liquid chromatography. The oxidative stress was performed by methemoglobin (MetHb) and Heinz bodies determination, concentrations of reduced glutathione (GSH), nitric oxide (NO), malondialdehyde (MDA), total antioxidant capacity (TEAC) and enzymatic activity of catalase (CAT), superoxide dismutase (SOD) in blood. In the study were obtained 23 samples from leprosy patients in D0 and 13 in D3; and 20 healthy and without leprosy subjects. In patients before treatment (D0) were observed increase of NO (D0 = 18.91 ± 2.39, control = 6.86 ± 1.79mM) and significant reduction of the activity of SOD enzyme (D0 = 69.88 ± 12.26, control = 138.42 ± 14.99 nmol/mL). In MDT-treated patients (D3), the dapsone concentration in plasma was of 0,552 ± 0,037 μg / mL, and they showed Heinz bodies presence and significant increase in the MetHb percentage (D3 = 3.29 ± 0.74, control = 0051 ± 0.66%). In this patients also were observed increase of the GSH levels (D3=7.01 ± 1.9; control =3.33 ± 1.9 μg/mL) and decrease of the CAT activity (D3 = 10.29 ± 02.02, control = 19:52 2:48 ± U / g protein). However, MDA levels in D0 and D3 patients didn’t show changed, while TEAC levels, in this patients, significantly increased (D0 = 2.90 ± 0.42; D3 = 3.04 ± 0:52, control = 1.42 ± 0.18 μmol / mL). These data suggest that the MDT is mainly cause for oxidative stress, because it induced the generation of oxidative damage identified by Heinz bodies’ presence and increase in the MetHb percent.FAPESPA - Fundação Amazônia de Amparo a Estudos e PesquisasCNPq - Conselho Nacional de Desenvolvimento Científico e TecnológicoO processo inflamatório decorrente da infecção por Mycobacterium leprae e a administração de fármacos com propriedades oxidativas, como a dapsona, são fatores de riscos ao estresse oxidativo ocasionado em pacientes com hanseníase. Este trabalho visa determinar as concentrações plasmáticas de dapsona em pacientes com hanseníase em uso de poliquimioterapia (PQT), correlacionando ao desenvolvimento do estresse oxidativo. Para o estudo, foram selecionados indivíduos saudáveis e pacientes com hanseníase, acompanhados antes (D0) e após a terceira dose supervisionada de PQT (D3). As concentrações plasmáticas de dapsona dos pacientes sob tratamento foram mesuradas por cromatografia liquida de alta eficiência. A avaliação do estresse oxidativo foi realizada através da determinação de metemoglobina (MetHb) e das concentrações de glutationa reduzida (GSH), óxido nítrico (NO), malondialdeído (MDA), capacidade antioxidante total (TEAC) e avaliação das atividades das enzimas catalase (CAT), superóxido dismutase (SOD) e da presença de corpúsculo de Heinz em esfregaços sanguíneos. No período de estudo foram obtidas 23 amostras de pacientes com hanseníase D0, 13 pacientes em D3 e 20 de indivíduos saudáveis e sem a doença. As alterações no pacientes antes do tratamento estavam associadas ao aumento de NO (D0=18.91± 2.39; controle= 6.86±1.79mM) e a redução significativa na enzima SOD (D0=69.88±12.26;controle= 138.42±14.99nmol/mL). Com relação aos pacientes sob tratamento, a concentração de dapsona no plasma foi 0.552± 0.037 μg/mL e as principais alterações observadas foram o aumento significativo no percentual de MetHb (D3= 3.29±0.74;controle=0.66±0.051%) e presença de corpúsculo de Heinz. Nos pacientes em tratamento também se observou aumento nos níveis de GSH (7.01±1.09μg/mL) quando comparados ao controle (3.33±1.09g/mL) e diminuição da atividade de CAT (D3= 10.29± 2.02; controle= 19.52±2.48 U/g de proteínas). Os níveis de MDA antes e durante o PQT não mostraram alteração, enquanto os níveis de TEAC aumentaram significativamente neste pacientes (D0=2.90 ± 0.42; D3=3.04±0.52; controle= 1.42±0.18μmol/mL). Estes dados sugerem que a PQT é a principal responsável pelo desenvolvimento de estresse oxidativo, através da geração de danos oxidativos identificados pela presença de corpúsculo de Heinz e aumento no percentual de MetHb

Determination of nitric oxide (NO) in serum (a), percentage of blood methemoglobin (b) levels of malondialdehyde (MDA) in plasma (c) of patients with untreated leprosy (MDT 0) and after the third dose supervised treatment (MDT 3).

<p>Figures in the chart are expressed as mean ± SD. *p<0.05 compared to MDT0 (ANOVA).</p

Representation of the CYP 2C19.

<p>Area covered (depicted as <i>light blue</i> transparent <i>solid surface</i>) by the two internal cavities (catalytic site cavity and adjacent cavity) of human microsomal cytochrome P450 (CYP) 2C19 (<i>brown</i>). The heme prosthetic group is represented in <i>green</i> and the molecule of dapsone in the color <i>red</i> (<b>a</b>). Delimitation of the area occupied by the two internal cavities (catalytic site cavity and adjacent cavity) of the CYP2C19 (<b>b</b>).</p