Os resultados e discussões estão sendo apresentado em forma de artigo:
Chloroquine and desethylchloroquine levels in late parasitological
failure cases of malaria vivax from Manaus, Brazil
Authors
Marques MMa , Costa MRFa, Simões FSFa, Viera JLFd , Borges LGd ,Nascimento MTSd, Alecrim MGC.a, b, c
a
Department of Malaria– The Foundation for Tropical Medicine of Amazonas (FMTAM), Manaus, Amazonas, Brazil
b
Amazonas State University (UEA), Manaus, Amazonas, Brazil c
Nilton Lins University, Manaus, Amazonas, Brazil d
The Federal University of Pará, Belém, Pará, Brazil
Corresponding author: José Luis Fernandes Vieira The Federal University of Pará
Rua Augusto Corrêa, 01 - 66.000-000 Belém – PA, Brasil 55 (91) 8841-8678
Abstract
We report the occurrence of resistance to chloroquine administered at a dose of 25 mg / kg in seven patients with vivax malaria treated in a reference center in Manaus, Brazil, from December 2007 to June in 2008. All patients were evaluated clinically and parasitologically-blood levels of chloroquine and desetilcloroquina were obtained on days 0, 3, 7, 14 and 28 of treatment. The data point to the importance of resistance to chloroquine in Manaus, Amazonas, Brazil.
Keywords: Plasmodium vivax, chloroquine resistance, Amazonas.
1. Introduction
Monitoring the effectiveness of antimalarial treatment is crucial; therefore, resistance to antimalarial drugs in the world is one of the obstacles to controlling the disease. The spread of strains of P. vivax resistant to chloroquine, as the literature, has a broad geographical distribution in various regions of the world as Indo-Pacific, Asia and South America ( Alecrim., 2000; Baird,2004; De Santana et al., 2007; Mendis et al.,2001; OPAS., 2004; Soto et al., 2001).
In the Brazilian Amazon, chloroquine plus primaquine is the treatment of choice for malaria caused by P. vivax and they remain with a good therapeutic efficacy (Alecrim et al.,2000 ; De Santana et al., 2007). However, it is reported in this study, the occurrence of resistance to chloroquine in patients treated in the reference center in Manaus, AM, Brazil. They were analyzed for concentrations of chloroquine
and desetilcloroquina in patients with and without treatment failure, suggesting the possibility of decreased sensitivity and therapeutic failure related to strains of P. vivax, as in the cases described the blood concentrations of chloroquine and its active metabolite, were above minimum effective concentration for susceptible strains of P. vivax (MEC ≥ 100 ng / mL), as characterized previous studies that establish patterns of parasite resistance ( Baird., 2004).
2. Materials and methods
The study was conducted at the Tropical Medicine Foundation of Amazons (FMTAM), in Manaus, in the period December 2007 to June 2008. The study included 14 patients classified as having P. vivax, seven sensitive and seven resistant, from a previous study involving 154 patients that evaluated the efficacy of chloroquine in combination with primaquine.
According to the protocol of the Pan American Health Organization (OPAS., 2004), eligible patients had clinical follow-up for parasites 28 days segment of both sexes aged 12 to 60 years, with positive diagnosis of P. vivax not serious, with parasite density from 250 to 100,000 parasites / mL, axillary temperature ≥ 37.5 ° C or history of fever in the first 48 hours. The total dose of 25mg/kg was clororoquina weight, distributed in three days with daily supervised administration of drugs in D0-10 mg / kg, D1-7.5 mg / kg and D2-7.5 mg / kg and primaquine in dose 0.5 mg / kg / day for 7 days ( Dua et al.,1999). The drug used in the study were first analyzed for their physical and chemical characteristics at the Federal University of Minas Gerais (Fharmacopeia & UNITED., 1988). Patients with recurrence were treated with parasitic mefloquine under the guidelines of the Ministry of Health of Brazil (Brazilian
Ministry of Health, 2001). - 20 mg / kg - followed by administration of primaquine. The actual rate of therapeutic failure was estimated by performing diagnostic PCR in blood samples collected in D28( Snounou et al ., 1993). The determination of chloroquine (CQ) and desetilcloroquina (DCQ) in whole blood samples collected on filter paper on days D0, D3, D7, D14 and D28 were analyzed at the Federal University of Pará (UFPA) in the toxicology laboratory, using if Cromatogragfia Liquid (HPLC), according to the method described by (Patchen et al., 1983). The study was submitted and approved by the Ethics Committee of the IMT-AM Protocol 25.000.105898/2004-74 (RAVREDA, 2008). The data were analyzed using EXCEL 2007. The frequency of resistance was estimated by means of proportion to their respective confidence interval of 95% with a significance level of 5%.
3. Results
Of the 14 patients selected for the study are predominantly male with a mean age of patients resistant to 34 ± 9.4 years and the tender of 36 ± 7 years. The parasite density of resistant participants, 7050.26±expressed as geometric mean and standard deviation, the D0 was 2526.65 parasitas/mm3 (16 to 16,465 parasitos/mm3), decreasing to 437.19 ± 1282.57 D1, 236, 30 ± 400 in D2. The patients showed reactivation of parasitemia at D28 with a mean parasite density of 2926.75 ± 2216.54 interval from 543 to 6980 parasitos/mm3. Since the parasite density of sensitive participants was 2730.10 ± 2581.49 parasitos/mm3 (125 - 8400) at baseline (D0), showing a decrease from D1 to become negative on D7. The average blood concentrations of CQ at D3, D7, D14 and D28 in sensitive patients were 2519.04 ± 2013.8 ng / mL, 1251.22 ± 1250.5 ng / mL, 1275.67 ± 642.05 ng /
mL and 203.49 ± 361.5 ng / mL, respectively. As for DCQ were 793 ± 637 ng / mL, 657.2 ± 498 ng / mL, 709.34 ± 586 ng / mL and 580.3 ± 432.4 ng / mL. The blood concentrations of CQ at D3, D7, D14 and D28 in patients with recurrent parasite were 670 ± 629.7 ng / mL, 411.8 ± 381.2 ng / mL, 384.3 ± 324.3 ng / mL and 209.9 ± 187.94 ng / mL, respectively. For DCQ values were 495.5 ± 306 ng / mL, 873.8 ± 767 ng / mL, 660 ± 456.4 ng / mL and 506 ± 289 ng / mL, respectively. There was no significant difference in average levels of CQ and DCQ between the groups in several days of study (p> 0.05). However, there was significant difference in blood levels of CQ + DCQ only D3. The average amount of blood concentrations of CQ + DCQ in the days of follow-up are illustrated in Table 1.
TABLE 1: Comparison between the average concentrations of CQ + DCQ in sensitive and resistant patients.
Dia CQ+DCQ Valores de p Pacientes Sensíveis (x ± DP) Pacientes Resistentes (x ± DP) D3 3311,1 ± 2361,8 1069,8 ± 810,64 0,04892 D7 1914,2 ± 1700 1321,5 ± 922,6 0,4631 D14 1351,4 ± 1763 1044,3 ± 667,6 0,6794 D28 783,8 ± 530,3 647,4 ± 227,2 0,5433 4. Discussion
All subjects in this study were adequately treated medically supervised and controlled the factors related to tolerance to the drug and its intake, a situation characterized by the concentration of CQ and DCQ in D3 in both groups. The average blood concentrations of CQ and DCQ in D3 sensitive patients was 2519 ±
2013.8 ng / mL and 793 ± 637 ng / mL, respectively. Similar results were found in 61 Indian patients treated orally with 25 mg chloroquine base / kg body weight for 3 days and reaching the D2 average blood concentration of CQ, 1980 ng / mL, and 1510 ng / mL present in red cells and 470 ng / mL in plasma from the same patients
(Dua et al.,1999) . In the same study the blood concentration of DCQ was 760 ng /
mL, erythrocyte levels of 540 ng / mL plasma and 220 ng / mL. These figures demonstrate that the CQ was well absorbed from the gastrointestinal tract, indicating that there were other factors that may influence the kinetics of the drug, including rapid excretion or poor absorption (Dua et al.,1999). Assessing levels of QC 785.4 ± 800.1 ng / ml in patients of Manaus. Eleven patients were resistant to P. vivax, with concentrations of 356.6 ± 296.1 ng / mL, lower than those found in patients sensitive
( De Santana et al ., 2007). Among the resistant patients, two had blood
concentrations of DCQ on D0 characterizing drug intake before the start of treatment, without reporting the inclusion of the study. One of those reporting the highest parasite density in D28. It is noteworthy that one of the patients had previously acquired malaria by at least 7 times, justifying the levels found. However, the resistant patients had lower concentrations of CQ and DCQ in D3 (p> 0.05) with values of 670 ± 629.7 ng / mL and 495.5 ± 306 ng / mL, respectively, but without statistical significance, the which may be linked to the number of study participants. Assessing resistant patients had a mean CQ in the blood of 880 ng / mL, with 420 ng / mL found in red blood cells and 320 ng / mL in plasma. The blood concentration of DCQ was 440 ng / mL with 260 ng / mL present in red blood cells and 180 ng / mL in plasma ( Dua et al ., 2000). Was found a large interindividual difference of chloroquine in whole blood of patients in both groups, in agreement with previous studies ( Baird ., 2004; Dua et la ., 1999; Dua et al ., 2000).The average blood
concentrations of CQ + DCQ in sensitive and resistant patients in D3 was 3311.1 ± 2361.8 and 1069.8 ± 810.64 ng / mL, respectively. Statistically significant difference between the two groups with p = 0.04892. This difference was not observed on days 7, 14 and 28. In the study evaluating the mean concentration of CQ + DCQ in erythrocytes and plasma of patients was significantly more sensitive than in resistant patients with p <0.0001 and p <0.02, respectively ( Dua et al., 2000).However, on day 7 no significant difference between the sensitive and resistant cases. Similarly, the mean concentration of CQ in plasma of patients was more sensitive than in resistant patients (p> 0.05), ( Karim et al., 1992), blood concentrations of CQ in D7 were lower in patients resistant than the sensitive ( Hellgren et al .,1989). Confirming the results found in this study.
The relatively low concentration of CQ in resistant patients may be caused by changes in absorption or the inter-individual variability in the pharmacokinetics of CQ ( Dua et al ., 1999).In D28 the average concentrations of CQ+DCQ in sensitive and resistant patients was 783.8 ± 530.3 ng / mL and 647.4 ± 227.2 ng/mL, respectively.
The resistance of P. vivax chloroquine characterized as parasitic recurrence within 28 days of clinical follow-parasites in subjects treated adequately with chloroquine (total dose - 25 mg / kg), confirming that the total levels of chloroquine (CQ + DCQ) in plasma or blood are above the minimum effective concentration of already standardized reference: ≥ 100 ng / mL in blood and plasma ≥ 10ng/mL ( Baird et al., 1996). In this study all patients had concentrations of CQ + DCQ above 100 ng / ml in D28 showing the existence of P. vivax resistant to CQ in Manaus, Amazonas. Although the study data confirm the emergence of P. vivax resistant to CQ in Manaus, the level of resistance is still not so high as to require a change in treatment
policy. The monitoring of therapeutic levels of this drug is useful for alerting the authorities to control the levels of resistance and determine the extent of the problem in the region, given that P. vivax is responsible for the largest number of malaria cases in Brazil.
Acknowledgments:
This study was part of a scientific protocol for programming multi Amazon Network for Monitoring Antimalarial Drug Resistance (RAVREDA) Secretariat of Health Surveillance / Ministry of Health, with financial support also from the Laboratory of Toxicology, Federal University of Pará (UFPA ) and FMTAM, serving also as a topic for dissertation in the Masters in Tropical and Infectious Diseases at the University of the State of Amazonas (UEA)
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