This study did not find evidence of differences in posttreatment blood sulfadoxine-pyrimethamine concentrations between cases with ACPR and LTF treatment outcomes. The clinically relevant therapeutic range for sulfadoxine-pyrimethamine is not well established, but a previous study found that nonimmune patients (travelers) with RII (patients still parasitemic at day 7 after treatment) SP-resistant P. falciparum
infections from Tanzania in 1986, who attained maximum sulfadoxine and pyrimethamine concentrations below the range of 62 to 115 μg/ml and below 49.7 ng/ml, respectively, developed secondary parasitemia (10
). Clearly, in the present study, most subjects (on day 3 only), irrespective of treatment outcome, sustained SDX and PYR concentrations (mean of 73.2 μg/ml [range, 51 to 89 μg/ml] and mean of 290 ng/ml [95% CI, 182 to 398], respectively) that seemed to be the same or below those that were associated with failure in the 1986 Tanzanian study. In contrast, Chulay et al. found that sulfadoxine and pyrimethamine concentrations of ≥60 μg/ml and ≥15 ng/ml, respectively, were effective against K39, a resistant strain of P. falciparum
in vitro (2
). Blood sulfadoxine-pyrimethamine levels attained in the present study were above the concentrations found by Chulay et al. Thus, values for parasite-growth-inhibitory concentrations for resistant or sensitive P. falciparum
isolates obtained in vitro may not always be an accurate reference for assessing the drug's efficacy in vivo.
In the univariate analysis, we found that among patients infected with the quintuple mutants, day 3 blood pyrimethamine concentrations were higher in those who cleared the infection than in those who did not. This difference could not be detected, however, in multivariate analysis (with Bonferroni adjustment), probably because there were numerous explanatory variables between the two groups (those that cleared quintuple mutants and those that failed to clear quintuple mutants) with only less than half (n
= 74) of the enrolled subjects analyzed. It is therefore possible that a greater sample size for the analysis would have been required to find that low blood pyrimethamine concentration is indeed a risk factor for failure to clear the quintuple mutants. The pharmacologic activity of drugs is a function of the unbound drug concentration. Since sulfadoxine and pyrimethamine are highly protein bound (88% and 93%, respectively [8
]) and since our drug assays looked only at the total drug, we might not have been able to notice the difference in plasma concentrations between the groups compared.
In a separate multivariate analysis that included all patients that had LTF and ACPR outcomes (n = 115), whether they had quintuple-mutant infections or not, it was found that low day 3 blood concentration of PYR (OR, 1.011 [1.003 to 1.024]; P = 0.018) was a risk factor for late treatment failure. The magnitude of the odds ratio from this analysis and the borderline significance of the results from the univariate analysis of the subgroup infected with quintuple mutants suggested that our observation that higher blood pyrimethamine concentrations enhance the ability of patients to clear resistant P. falciparum must be interpreted with caution and needs further validation.
There was substantial interindividual variation in sulfadoxine-pyrimethamine disposition. The cause of these differences is unknown, but the interindividual variations may reflect the effect of disease severity and/or differences in the time of day at which posttreatment day 3 blood samples were collected. Blood samples were collected at any time during working hours on day 3 between 8 a.m. and 5 p.m. The observed ranges of day 3 sulfadoxine and pyrimethamine concentrations (59 to 75 μg/ml and 208 to 264 ng/ml, respectively) in this study were, nevertheless, similar to the ranges (sulfadoxine range: 51 to 89 μg/ml [mean, 73.2 μg/ml]; pyrimethamine range: 252 to 484 ng/ml [mean, 368 ng/ml]) found by Hellgren et al. in Tanzania (3
) and Bustos et al. (1
) in the Philippines. In contrast to our results and those of Hellgren et al., Bustos et al. found a mean sulfadoxine day 3 concentration of 184 ± 40 μg/ml (1
). In our study population, back-extrapolated sulfadoxine C0
was found to fall within the previously observed range of the highest blood concentration levels of a drug after administration, 51 to 169 μg/ml, following oral administration of the standard SP dose (1
). The apparent elimination half-life of sulfadoxine agreed with previous reports of a range of 4 to 11 days (1
Sulfadoxine-pyrimethamine acts through a two-step synergistic blockade of plasmodial division. A decline in the concentration of one of the component drugs of this combination to a concentration below that required for effective synergy would result in the loss of the antiplasmodial synergistic action. We found a trend towards lower sulfadoxine levels in patients with treatment failure than in those with ACPR (P
= 0.061). We observed that after day 3 posttreatment (Fig. ), sulfadoxine concentrations decreased rapidly (in both ACPR and LTF) to concentration levels that are below those required to kill resistant P. falciparum
isolates in vivo (i.e., below the range of 62 to 115 μg/ml). This is of concern, since therapeutic concentrations need to be maintained for three life cycles to eradicate P. falciparum
). Additional studies are needed to determine whether after day 3 the pyrimethamine concentration remained above that required for effective synergistic action against resistant parasites. However, we postulate that the rapid decline of sulfadoxine levels below the concentrations required for synergy immediately after day three should result in an overall loss of the required effective synergy between pyrimethamine and sulfadoxine as a combination. Such a situation would result in differential pyrimethamine susceptibility among parasites that would otherwise be susceptible to the combination, resulting in the selection of the low-grade resistant parasites observed in this study.
For this study population, it has been shown that predicted SP therapeutic levels were sustained, at least until day 3 after treatment, in subjects who cleared quintuple-mutant infections as well as in those who could not clear them and in ACPR and LTF groups in general. However, therapeutic levels are very poorly defined by the small numbers of patients with uncomplicated malaria for whom published data on the association of pharmacokinetics and treatment response (pharmacodynamics) are available (1
). This is further complicated by the rightward shift in the dose response curve as resistance spreads as well as by the effects of other factors, including humoral immunity (7
) and blood folate levels (13
), on therapeutic response. We therefore suggest that larger pharmacokinetic-pharmacodynamic studies are needed to elucidate factors that may account for interindividual variation in the observed SP therapeutic responses.