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ABSTRACT |
 TOP ABSTRACT INTRODUCTIONPATIENTS AND METHODSRESULTSDISCUSSIONREFERENCES |
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/ß-arteether dissolved in peanut oil used in Indian patients. |
INTRODUCTION |
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TOPABSTRACTINTRODUCTIONPATIENTS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Phase II clinical trials of injectable Artemotil were initiated in 1993 in adult patients with Plasmodium falciparum malaria in The Netherlands and Thailand. Studies were initiated in patients with non-severe malaria to establish an optimal dosage regimen and to characterize the rapidity of the clinical response. In 1994, subjects were given Artemotil intramuscularly at a starting dose of 3.2 mg/kg, followed by 1.6 mg/kg for four days for the treatment of uncomplicated P. falciparum malaria. The regimen proved safe and had potent antimalarial efficacy, reducing parasitemia from the peripheral blood at a low dosage; similar to artemether (AM), which reduced parasitemia by 50–90%.3
In a second trial, 200 patients with severe P. falciparum malaria were studied between July 1995 and June 1997. The objective of the study was to compare the efficacy of intramuscular Artemotil to AM in patients with severe P. falciparum malaria. All patients were treated by one of the three regimens: 1) low dose of Artemotil: 3.2 mg/kg on day 0 and 1.6 mg/kg of on days 1–4, 2) low dose of AM: 3.2 mg/kg on day 0 and 1.6 mg/kg on days 1–4, or 3) high dose of Artemotil: 4.8 mg/kg on day 0, 1.6 mg/kg six hours later, and 1.6 mg/kg/day on days 1–4. The results of the study showed that the two low-dose treatments had survival percentages within the expected range: Artemotil = 83.6% and AM = 88.7%. This difference was not statistically significant. A higher initial parasite count, renal failure, and injection into the thigh resulted in a significantly lower chance of survival. Parasite clearance time (PCT) did not differ between the two treatments. Comparable means were found for fever clearance time (FCT), approximately 80 hours, which was confirmed by statistical evaluation. Coma recovery time for both low dose groups was approximately 40 hours with no significant differences found for Artemotil compared with AM in high-dose treatments.3
The phase III study was reported in 1998 from two of the three centers that studied this drug in African children with cerebral malaria. Ninety-two children were studied: 48 received Artemotil and 44 received quinine. No significant differences in survival, coma resolution time, neurologic sequelae, PCT, and fever resolution time were seen between the two regimens. Rates for negative malaria smears one month after therapy were similar in both groups. Artemotil was a well-tolerated drug in the 48 patients in this study. It appears to be at least therapeutically equivalent to quinine for the treatment of pediatric cerebral malaria and has the advantage intramuscular dosing once a day for only five days.4 A comparison study between intramuscular Artemotil and intravenous quinine has been evaluated in 102 children from Cameroon. Artemotil with sesame oil has been shown to be safe and therapeutically as effective as quinine for the treatment of cerebral malaria in children.5
Intramuscular administration of Artemotil in sesame oil yielded a low blood concentration and a long elimination half-life. It was expected that the half-life of Artemotil with sesame oil and its more lipophilic properties would be longer than with other artemisinin drugs, thus favoring accumulation in blood and brain tissue as advantages in treating treat severe and cerebral malaria.6 The comparison of the therapeutic potential of Artemotil with other artemisinin drugs in humans for 3–5-day treatments is summarized in Table 1
.3,6–27 The rates of recrudescence, fever, and parasite clearances are traditional evaluations for the efficacy of the antimalarial drugs in humans. These three parameters showed that the therapeutic potential of intramuscular Artemotil is not superior to that of oral dihydroartemisinin (DHA), oral and intravenous artesunate (AS), intramuscular AM, or even intramuscular / ß-arteether formulated with peanut oil. However, in vitro studies reported the relative antimalarial potency of artemisinin compounds as 1 (artemisinin) < 1.2–1.5 (artelinic acid) < 1.9–2.3 (artemether) < 2.2–2.6 (ß-arteether) < 3.9–4.8 (artesunate) < 3.6–5.8 (dihydroartemisinin) with culture parasites of P. falciparum.28–30 Since ß-arteether is not any less potent than artemisinin drugs in vitro, why is the antimalarial potential of Artemotil inferior to other artemisinin compounds in clinical use?
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PATIENTS AND METHODS |
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TOPABSTRACTINTRODUCTIONPATIENTS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Treatment and observation. Sornchai Looareesuwan (Bangkok Hospital for Tropical Diseases, Bangkok, Thailand) manufactured the trial medication. Medication was administered intramuscularly in the gluteal region on alternating sides for five days at the low dose of 3.2 mg/kg on day 0 and 1.6 mg/kg/day on days 1–4 in 15 patients and at the high dose of 4.8 mg/kg on day 0, 1.6 mg/kg six hours after the initial injection, and 1.6 mg/kg/day on days 1–4 in 55 patients. Randomization was in blocks of five to include four high-dose Artemotil patients and one normal-dose (low dose) Artemotil patient. Clinical outcomes after a change in treatment, e.g., to intravenous AS, were also recorded in the coma recovery time.
Clinical observations were recorded at the clinic from -12 hours up to 28 days after the first drug injection or alternatively from -12 hours up to day 7 with visits on days 14, 21, and 28. Blood lactate concentrations were measured every six hours (in the same samples as blood glucose) until 24 hours after the first dose. Full physical and neurologic examinations were given before the first injection and twice a day until the patient had cleared fever, parasitemia, and coma and on days 7, 14, 21, and 28. Blood films and oral body temperature were obtained every six hours until films were negative and the temperature was < 37.5°C; thereafter, films and temperature were obtained once a day to determine recrudescence failure.
Plasma sample preparation. Blood samples (5.0 mL) for pharmacokinetic analysis were drawn repeatedly via an indwelling catheter with a heparin lock in all patients. Blood was collected into sterile plastic syringes, transferred immediately into heparinized tubes, and centrifuged at 3,000 rpm for 15 minutes at 4°C. Plasma samples were aliquoted into two glass tubes (1 mL each) and stored in dry ice or an electric freezer at –80°C. Duplicate samples were transported by thermo container containing dry ice to the laboratory at the Walter Reed Army Institute of Research in Washington, DC. On delivery, samples were inventoried and stored at –80°C until analysis could be performed. Quality control (QC) and standard curve (SC) samples were prepared in the blank plasma as close as possible to the date of sample receipt and were stored under the same conditions as the clinical samples. The QC and SC samples consisted of Artemotil and DHA in human blank plasma at different concentrations (0–1,000 ng/ mL). Six QC and 10 SC samples were assayed with each set of patient samples; they were randomly placed among clinical samples for each assay run. Six QC samples were assayed consecutively within an analysis.
A liquid-liquid extraction method was used for sample preparation as previously described.32 Artemotil and DHA concentrations were measured by reverse-phase high-performance liquid chromatography with electrochemical detection (HPLC-ED).
Analysis by HPLC-ECD. The HPLC-ECD was performed using a model BAS 200 liquid chromatography system (Bio-analytical Systems, West Lafayette, IN). This system has three mobile phase reservoirs, solenoid proportioning valves, a dual piston pump, a pulse dampener, a column and detector oven, dual thin-layer electrodes with an Ag/AgCI reference electrode, and a Rheodyne injector for manual injection, which was modified for reductive work. The system is also equipped for mobile phase heating and sparring. The HPLC-ECD was performed using minor modifications of the method of Li and others.33
Pharmacokinetic data evaluation. Extensive pharmacokinetic analysis was carried out on the first 60 patients in both regimens of dose treatment. Venous blood samples were taken through an antecubital catheter at 0, 2, 4, 8, 12, 18, 24, 48, 72, 96, 108, 120, 144, 168, and 216 hours. To determine the pharmacokinetic parameters of Artemotil and the concentration-time data of Artemotil as well as the metabolite DHA, samples collected during first day were fitted to a one-compartment model. On the last day, dosing was fitted to a two-compartment open model using a nonlinear, extended, least-square fitting procedure (WinNonlin 3.1; Scientific Consulting, Inc. Apex, NC). The area under the curve (AUC) for the drug was determined by the linear trapezoidal rule with extrapolation to infinity based on the concentration of the last time point divided by the terminal rate constant. Extrapolations to time zero were done using zero concentration for intragastric dosing and using C0 values determined from the two-compartment model equation at time zero by intravenous route. Mean clearance rate (CL) was determined by dividing the dose by the AUCinf for intravenous injection. Mean residence time (MRT) was determined by dividing the area under the first moment curve (AUMC) by the AUC. The volume of distribution at steady state (Vss) was calculated as the product of CL and MRT. The conversion ratio of DAH/Artemotil was calculated by AUCDHA/AUCArtemotil. Statistical analysis was performed with the statistical software Excel® (Microsoft, Redmond, WA) using a Student’s t-test for dependent samples to compare means of paired samples within one individual.
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RESULTS |
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8. Eight of the 70 patients had acute renal failure and required hemodialysis. Another 27 of these 70 patients had anemia and required blood transfusions.
All patients showed a slow initial response (mean PCT = 75.8 hours, mean FCT = 77.2 hours in low-dose patients, and mean PCT = 64.4 hours, mean FCT = 70.1 hours in high-dose patients). One patient (151), who likely received intravenous drug administration, showed a rapid initial response (PCT = 36.0 hours, FCT = 4.0 hours). The low and high doses of Artemotil did not clear the hyperparasitemia completely and resulted in a high recrudescence of 77% in the low-dose patients and 46% in high-dose patients. All recrudescence patients showed recrudescence during follow-up on days 17–21 (Table 2
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Evaluation of toxicity. The evaluation of adverse experiences, injection site inspection, electrocardiographic recordings, neurologic examinations, and clinical laboratory tests did not show abnormalities due to any of the two study medications. Clinical observations were recorded at the clinic from –12 hours up to 28 days after the first drug injection or alternatively from –12 hours up to day 7 with visits on days 14, 21, and 28.
Stability of plasma samples. Quality control showed frozen samples of Artemotil in plasma to be stable. The concentration of Artemotil did not change in the first nine months while stored at –80°C. Artemotil levels were slightly lower (1.4–3.9%) when compared with freshly prepared samples after one year at –80°C. The Artemotil concentration of frozen samples was lower (6.6–13.2%) when stored at –80°C for two years. Dihydroartemisinin is less stable than Artemotil in the frozen plasma; its half-life of stability was 14.5 months at –80°C. In the present study, the QC and SC samples were prepared shortly before the date of sample receipt and stored frozen under the same conditions as the clinical samples. The assay quantitation of clinical samples of Artemotil and DHA were not affected by long-term storage.
Pharamcokinetic analysis of Artemotil. The data sets obtained with the pharmacokinetic modeling program were used for the daily dosing interval fitting in patients receiving the low-dose regimen, for the twice dosing interval fitting on day 0 (0 and 6 hours), and for the daily dosing interval fitting on days 1–4 in the high-dose subjects. Good graphic fits of the measured plasma concentration curves obtained for both regimens, indicating that the procedure performed well in estimating the pharmacokinetic parameters. Low inter-parameter correlation and small confidence intervals were obtained with good coefficients of determination (r2 = 0.934–0.971).
The individual levels of Artemotil and the mean plasma concentrations of DHA are shown in Figure 1. The main pharmacokinetic parameters following low- and high-dose administrations are shown in Table 3. The maintaining intramuscular dose of Artemotil on day 4 was half and one-fourth of that on day 0 (loading dose) in the low-dose and high-dose groups, respectively. However, the plasma concentration of Artemotil on day 4 was three times higher than those on day 0 in both dose groups. Drug accumulations of Artemotil and DHA in the blood were evident. Due to daily increases of the drug levels, the steady state plasma concentrations of Artemotil and DHA were not reached until after the final dose (96 hours).
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). The elimination half-lives calculated for artemotil on day 4 are not true elimination half-lives. Prolonged absorption of artemotil from sesame oil will extend the plasma concentrations and make the elimination half-life appear longer than the true elimination half-life.
Pharmacokinetic analysis of DHA.
The peak plasma concentration of DHA, an active metabolite of Artemotil, was much lower than its parent drug (Figures 1 and 2). The pharmacokinetic parameters showed that the AUC value of Artemotil during the entire treatment period was 5.3-fold higher than that of DHA in low-dose patients and 6.2-fold higher in high-dose patients (Table 3). The ratio of AUCDHA to AUCArtemotil was 0.19 ± 0.14 (mean ± SD) in the low-dose group and 0.16 ± 0.07 in the high-dose group during the entire treatment period (Table 3).
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). The Cmax increased 1.6-fold from day 0 (12.7 ng/mL) to day 4 (20.3 ng/mL) in the low-dose group and 3.3-fold in the high-dose group (Table 3). On day 4, AUC values were 1,273 ng·hr/mL in the low-dose group (a 14.7-fold increase) and 1,411 ng·hr/ml in high-dose group (a 10.7-fold increase). The half-lives of DHA also increased from 14.7–16.0 hours on day 0 to 23.8–24.4 hours on day 4 in the both dose groups (Table 4). The MRT of DHA (33.2–34.0 hours) was shorter than that of Artemotil (44.9–61.4 hours) on day 4, suggesting that DHA in the human body does not last longer that the parent drug Artemotil (Tables 3 and 4).
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, indicating that his pharmacokinetic profiles were different from other patients. Patient 151 at first dosing seemed to be incidentally injected through a vessel and showed an intravenous injection profile with very high Cmax concentration right after injection and a rapid distribution phase. Artemotil and DHA plasma concentrations decreased significantly after first dosing with an elimination t1/2 of 1.1 hours in the first six hours. After the second intramuscular injection at the sixth hour, the elimination t1/2 of Artemotil was immediately increased to 9.35 hours as a daily half-life (Table 3). The Cmax (2,407 ng/mL) was approximately 22-fold higher in patient 151 than the mean value (110 ng/mL) reported in other high-dose patients. The AUC value (12,259 ng·hr/mL) was 2.6 times higher than the mean value (7,403 ng·hr/mL) reported in other subjects in the high-dose group. The results were also reported for increasing DHA levels in patient 151. |
DISCUSSION |
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The data in this accumulation study confirm and extend that of earlier studies in humans and animal species following multiple intramuscular administrations of Artemotil or AM with sesame oil.2,34–37 A rat study demonstrated that the absorption of Artemotil from muscle (injection site) was incomplete on the first day and the bioavailability of Artemotil was 23.4% following single intramuscular administration. At 24 hours after dosing, up to 38% of the total single dose of Artemotil remained at the injection site; at 48 hours, 22% of the total single dose remained at the injection site.9 Based on the calculated half-life (26.3 hours) of the slow absorption phase in muscle, a single Artemotil dose (25 mg/kg) would be completely absorbed in 6–7 days. Acute toxicity data has shown that animals receiving a single high dose of Artemotil in sesame oil died between days 5 and 11.35 The data suggests that the low bioavailability of Artemotil after intramuscularly administration, as well as the delayed onset of toxicity and death in rats, is due to slow and prolonged absorption.
In human studies, no plasma accumulation has been found for artemisinin drugs by other administration routes. Four artemisinin drugs (artemisinin, AS, AM, and DHA) have shown a decreasing concentration in plasma during multiple oral treatments in malaria patients and healthy subjects. When comparing the final dose day to day 0 in these studies, the Cmax and AUC values were markedly reduced from one-third to one-seventh. The decrease in drug exposure during treatment was not disease-related since the pharmacokinetics of artemisinin on day 0 was similar to that reported in healthy subjects.38–43 It is suggested that artemisinin drugs may induce CYP2C19 in an increase of metabolic capacity and undergo auto-induction of the first-pass effect; this may result in a decrease in bioavailability after repeated doses.42,44
In vitro studies have addressed parasite viability after exposure to artemisinin; killing or suppressive effects of the compound in relation to its concentrations and exposure time were assessed. Concentrations of 10–6-10–5 M (280–2,800 ng/ mL) artemisinin showed an effect after only three hours of drug exposure, whereas 10–7 M (28 ng/mL) artemisinin required 24 hours of exposure for a killing effect. At concentrations of 10–8 M (2.8 ng/mL), artemisinin had no appreciable effect on the parasites.45 The low-dose range would not be selected in human treatments due to the very short half-life of artemisinin drugs; as described previously, the longer exposure time at low blood concentrations was more likely to cause fatal neurotoxicity than higher levels of exposure.46 Longer exposure times and low blood levels did not increase efficacy. Longer half-lived Artemotil (12–42 hours) did not increase efficacy. In treating patients with severe and complicated malaria, patient survival is the most important factor. A curative effect is best achieved by switching off the vital functions of the parasite as soon as possible.47 There is no evidence that rapid parasite killing is harmful to the patient, and the therapeutic blood drug concentration should be as high as possible without risking toxicity.48
During the Artemotil clinical trial, patient 151 inadvertently obtained a full or fractional intravenous injection with the intramuscularly formulation in the first treatment. The pharmacokinetic data for this patient showed an intravenous concentration versus time profile and extremely high drug level in plasma (Figure 2). Compared with the mean values of other patients who were in same regimen group, the Cmax was 22-fold higher in this patient than in the other 44 subjects. The AUC of patient 151 was 2.6 times higher than that of other patients on day 0. Due to high drug levels, this patient obtained very successful clinical therapy with a PCT of 36 hours and an FCT of 4 hours (which are close to the therapeutic potency of an artesunate injection) compared with other subjects (PCT = 64 hours and FCT = 70 hours). If an improved formulation for intramuscular Artemotil showed more rapid and reliable absorption to the extent seen in patient 151, the therapeutic potential of Artemotil would be much improved and at least as good as other artemisinin drugs.
Another reason for the poor efficacy of Artemotil could be due to the extremely high-count parasitemia in the patients in this study. Mean parasitemias of 436,633 and 378,045/µL were detected in the low-dose and high-dose patients, respectively. These were approximately two times higher than the numbers for hyperparasitemia (> 200,000 parasites/µL or > 5% infected red blood cells) in patients who were also unable to swallow tablets, with coma, anemia, and/or renal failure as the indication of severe malaria.49 The low drug concentration and longer exposure time of Artemotil with sesame oil did not seem to be suitable for the severe malaria patients. Previous pharmacokinetic data demonstrated that the plasma concentration of AM was more than three-fold higher than that of ß-arteether in rats at a same dose level; both drugs were formulated in sesame oil.33 Similar data that showed that Artemotil has a slower rate of absorption than AM was reported in humans.45 It may be that AM has less lipophilic properties than Artemotil (ß-arteether), favoring absorption from muscle. The better efficacy of /ß-arteether was found in the peanut oil formulation from India, suggesting that peanut oil may result in easier absorption of this drug in the humans. Further efficacy studies need to be done on the new intramuscular formulation of Artemotil.
Sesame oil and cremophore were used to evaluate the efficacy and pharmacokinetics of Artemotil in two different intramuscular vehicles in the rat.46 Data showed that the Cmax was 13-fold higher after using 25 mg/kg of arteether (AE) with cremophore (AECM) than after using Artemotil at same dose level on day 0. The bioavailability of AECM following dosing was 74.5 ± 12.1% (mean ± SD) (3.7-fold higher than Artemotil). The high bioavailability or the high exposure level contributed to a marked increase of the efficacy in mice. When AECM and Artemotil were administered intramuscularly once a day for three consecutive days in P. berghei-infected mice, the cure dose in 50% of the animals (CD50) was 34.1 (95% confidence interval [CI] = 13.3–66.9) mg/kg for Artemotil and 14.2 (95% CI = 6.8–21.5) mg/kg for AECM. The data confirm that changing the formulation has the ability to change the efficacy of AE.46
Intramuscular Artemotil has the advantage of simple application and fewer undesirable side effects in cases where the patient is not able to retain food. In particular, treatment with intramuscular Artemotil can save time and lives in small children living in areas poorly served by health services. Also, Artemotil has the lowest possible cost and will be made available to developing malaria-endemic countries.37 However, the recrudescence rate is too high (47–71%) compared with other antimalarial regimens for the treatment of severe malaria in these regions. Mefloquine reduces the recrudescence rate from 24% to 5% with intravenous artesunate, from 45% to 20% with intramuscular artemether, and quinine and tetracycline had a recrudescence rate of 4%.50 The fact that the recrudescence rates are too high may be related to the lower exposure level during treatment with Artemotil. Therapy with Artemotil for severe malaria might be used in combination with a long-acting antimalarial such as mefloquine to avoid this problem with recrudescence.
The therapeutic efficacy was higher in the patients treated the high-dose regimens; a significant difference was found in the malaria cure rates (23% in low-dose patients and 48% in high-dose patients). The rate of recrudescence was higher in low-dose patients (77%) than in high-dose patients (46%). The pharmacokinetic data showed that the AUC of Artemotil in the high-dose group was 1.2-fold higher than that of the low-dose group. Similar results were also exhibited for the AUC value of DHA. This further indicated the high drug concentration (drug exposure level) produced a better therapeutic efficacy for Artemotil. Although the larger loading dose (high-dose regimen) for Artemotil gave higher AUC and Cmax values, it was also highly variable. This may be a particular problem in severe malaria, where peripheral blood flow may be reduced, and absorption from the depot sites may be further impeded. A better solution for reliable bioavailability would be an improved formulation with more rapid absorption.
Considerable interindividual variation in the pharmacokinetics of both parent drug and metabolite were reported. Dihydroartemisinin is the major plasma metabolite of Artemotil, which has been shown in vitro to be a five-fold more potent antimalarial than the parent drug.51,52 Thus, evaluating the conversion rate of Artemotil to DHA is very important for assessing the efficacy of Artemotil. In the present study, the two dose regimens showed a very low and similar conversion range (no significant difference) of Artemotil to DHA with 0.16–0.19 DHA to Artemotil ratios in rats. The low concentration and extended half-life of DHA did not contribute to the efficacy observed in this study, further suggesting that drug exposure level is more important than drug exposure time.
In conclusion, due to slow and prolonged absorption, Artemotil produced low peak concentrations, high accumulations, and longer exposure times in blood. Although Artemotil is therapeutically at least as effective as quinine for the treatment of cerebral malaria in children,4 the two dose regimens in this study failed to show a better therapeutic effect than that of other artemisinin drugs. In vitro and in vivo research has demonstrated that a high drug concentration in blood at first exposure is important for the extermination of the parasites, especially in conditions of severe malaria. The PCT seen in patient 151 shows the potential therapeutic effect of an intramuscular Artemotil, which is related to direct injection into a blood vessel with a rapid absorption and very high peak concentration. Therefore, a better solution for potential efficacy of this drug would be an improved formulation with more rapid and complete absorption.
Received February 5, 2004. Accepted for publication June 30, 2004.
Financial support: This work was supported by the UNDP/World Bank/World Health Organization Special Program for Research and Training in Tropical Diseases (TDR) and a Mahidol University Grant.
Authors’ addresses: Qigui Li, and Wilbur K. Milhous, Division of Experimental Therapeutics, Walter Reed Army Institute of Research, 503 Robert Grant Avenue, Silver Spring, MD 20910-7500, Telephone: 301-319-9351, Fax: 301-319-7360, E-mail: qigui.li{at}na.amedd.army.mil. Charles B. Lugt, ARTECEF BV, PO Box 5, NL-3600, AA Maarssen, The Netherlands. Sornchai Looareesuwan, Srivicha Krudsood, Polrat Wilairatana, Suparp Vannaphan, and Kobsiri Chalearmrult, Faculty of Tropical Medicine, Mahidol University, Hospital for Tropical Diseases, 420/6 Ratchavithi Road, 10400 Bangkok, Thailand, Telephone: 66-2-247-1688, Fax: 66-2-245-7288.
Reprint requests: Qigui Li, Division of Experimental Therapeutics, Walter Reed Army Institute of Research, 503 Robert Grant Avenue, Silver Spring, MD 20910-7500.
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REFERENCES |
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TOPABSTRACTINTRODUCTIONPATIENTS AND METHODSRESULTSDISCUSSIONREFERENCES |
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/ß-arteether in acute falciparum malaria. Trans R Soc Trop Med Hyg 89: 299–301.[Medline]
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| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
