Pharmacokinetics and pharmacodynamics of (+)-primaquine and (-)-primaquine enantiomers in rhesus macaques (Macaca mulatta).

Primaquine (PQ) remains the sole available drug to prevent relapse of Plasmodium vivax malaria more than 60 years after licensure. While this drug was administered as a racemic mixture, prior studies suggested a pharmacodynamic advantage based on differential antirelapse activity and/or toxicities of its enantiomers. Oral primaquine enantiomers prepared using a novel, easily scalable method were given for 7 days to healthy rhesus macaques in a dose-rising fashion to evaluate their effects on the blood, liver, and kidneys. The enantiomers were then administered to Plasmodium cynomolgi-infected rhesus macaques at doses of 1.3 and 0.6 mg/kg of body weight/day in combination with chloroquine. The (-)-PQ enantiomer had higher clearance and apparent volume of distribution than did (+)-PQ and was more extensively converted to the carboxy metabolite. There is evidence for differential oxidative stress with a concentration-dependent rise in methemoglobin (MetHgb) with increasing doses of (+)-PQ greater than that seen for (-)-PQ. There was a marked, reversible hepatotoxicity in 2 of 3 animals dosed with (-)-PQ at 4.5 mg/kg. (-)-PQ in combination with chloroquine was successful in preventing P. cynomolgi disease relapse at doses of 0.6 and 1.3 mg/kg/day, while 1 of 2 animals receiving (+)-PQ at 0.6 mg/kg/day relapsed. While (-)-PQ was also associated with hepatotoxicity at higher doses as seen previously, this has not been identified as a clinical concern in humans during >60 years of use. Limited evidence for increased MetHgb generation with the (+) form in the rhesus macaque model suggests that it may be possible to improve the therapeutic window for hematologic toxicity in the clinic by separating primaquine into its enantiomers.

Randomized, double-blind, placebo-controlled clinical trial of a two-day regimen of dihydroartemisinin-piperaquine for malaria prevention halted for concern over prolonged corrected QT interval.

Dihydroartemisinin-piperaquine, the current first-line drug for uncomplicated malaria caused by Plasmodium falciparum and Plasmodium vivax in Cambodia, was previously shown to be of benefit as malaria chemoprophylaxis when administered as a monthly 3-day regimen. We sought to evaluate the protective efficacy of a compressed monthly 2-day treatment course in the Royal Cambodian Armed Forces. The safety and efficacy of a monthly 2-day dosing regimen of dihydroartemisinin-piperaquine were evaluated in a two-arm, randomized, double-blind, placebo-controlled cohort study with 2:1 treatment allocation. Healthy military volunteers in areas along the Thai-Cambodian border where there is a high risk of malaria were administered two consecutive daily doses of 180 mg dihydroartemisinin and 1,440 mg piperaquine within 30 min to 3 h of a meal once per month for a planned 4-month period with periodic electrocardiographic and pharmacokinetic assessment. The study was halted after only 6 weeks (69 of 231 projected volunteers enrolled) when four volunteers met a prespecified cardiac safety endpoint of QTcF (Fridericia's formula for correct QT interval) prolongation of >500 ms. The pharmacodynamic effect on the surface electrocardiogram (ECG) peaked approximately 4 h after piperaquine dosing and lasted 4 to 8 h. Unblinded review by the data safety monitoring board revealed mean QTcF prolongation of 46 ms over placebo at the maximum concentration of drug in serum (Cmax) on day 2. Given that dihydroartemisinin-piperaquine is one of the few remaining effective antimalarial agents in Cambodia, compressed 2-day treatment courses of dihydroartemisinin-piperaquine are best avoided until the clinical significance of these findings are more thoroughly evaluated. Because ECG monitoring is often unavailable in areas where malaria is endemic, repolarization risk could be mitigated by using conventional 3-day regimens, fasting, and avoidance of repeated dosing or coadministration with other QT-prolonging medications. (This study has been registered at ClinicalTrials.gov under registration no. NCT01624337.).

Pharmacokinetics and pharmacodynamics of oral artesunate monotherapy in patients with uncomplicated Plasmodium falciparum malaria in western Cambodia.

Artemisinin-resistant malaria along the Thailand-Cambodian border is an important public health concern, yet mechanisms of drug action and their contributions to the development of resistance are poorly understood. The pharmacokinetics and pharmacodynamics of oral artesunate monotherapy were explored in a dose-ranging trial in an area of emerging artesunate resistance in western Cambodia. We enrolled 143 evaluable subjects with uncomplicated Plasmodium falciparum malaria in an open label study of directly observed artesunate monotherapy at 3 dose levels (2, 4, and 6 mg/kg of body weight/day) for 7 days at Tasanh Health Center, Tasanh, Cambodia. Clinical outcomes were similar among the 3 groups. Wide variability in artesunate and dihydroartemisinin concentrations in plasma was observed. No significant dose-effect or concentration-effect relationships between pharmacokinetic (PK) and parasite clearance parameters were observed, though baseline parasitemia was modestly correlated with increased parasite clearance times. The overall parasite clearance times were prolonged compared with the clearance times in a previous study at this site in 2006 to 2007, but this did not persist when the evaluation was limited to subjects with a comparable artesunate dose (4 mg/kg/day) and baseline parasitemia from the two studies. Reduced plasma drug levels with higher presentation parasitemias, previously hypothesized to result from partitioning into infected red blood cells, was not observed in this population with uncomplicated malaria. Neither in vitro parasite susceptibility nor plasma drug concentrations appeared to have a direct relationship with the pharmacodynamic (PD) effects of oral artesunate on malaria parasites. While direct concentration-effect relationships were not found, it remains possible that a population PK modeling approach that allows modeling of greater dose separation might discern more-subtle relationships.

A simplified liquid chromatography-mass spectrometry assay for artesunate and dihydroartemisinin, its metabolite, in human plasma.

Artesunate (AS) is a potent antimalarial that is used worldwide for the treatment of malaria. A simple method with a total run time of 12 min was developed and validated for the quantification of AS and dihydroartemisinin (DHA), its active metabolite, in human (heparinized) plasma based on one-step protein precipitation in acetonitrile using artemisinin (ARN) as an internal standard, followed by liquid chromatography with a single quadrupole mass spectrometry system connected to a C18 column. Peak area ratio responses were fitted to the 2nd-order curve type, polynomial equation with weighting (1/concentration) over a quantification range between 3.20/5.33-3,000/5,000 nM (1.23/1.52-1153/1422 ng/mL) of AS/DHA showing linearity with very good correlation (r2>0.999). Single ion recordings of 5 µL injections of plasma extracts allowed for limits of detection of 1.02 nM (0.39 ng/mL) for AS and 0.44 nM (0.13 ng/mL) for DHA. The inter-assay and intra-assay accuracy and precision of the method was very good with an inaccuracy of ±12.4% and coefficients of variation of ≤10.7% at all tested concentrations. The recovery of the analytes from plasma was ≥95%. Other commonly used antimalarials including mefloquine, quinine, and chloroquine, did not interfere with the analysis. Post-preparative tests over 24 h in an autosampler (10 °C) showed that the DHA response was only 2.1% of AS from auto-hydrolysis, and ß-DHA was the major, stable epimer that was used for quantification of DHA. In contrast, α-DHA increased steadily up to 600%. Artesunate and DHA in plasma were stable through three freeze/thaw cycles for up to 6 h at room temperature and up to one year at -80 °C.