Rectal artemisinins for malaria: a review of efficacy and safety from individual patient data in clinical studies

Background Rectal administration of artemisinin derivatives has potential for early treatment for severe malaria in remote settings where injectable antimalarial therapy may not be feasible. Preparations available include artesunate, artemisinin, artemether and dihydroartemisinin. However each may have different pharmacokinetic properties and more information is needed to determine optimal dose and comparative efficacy with each another and with conventional parenteral treatments for severe malaria. Methods Individual patient data from 1167 patients in 15 clinical trials of rectal artemisinin derivative therapy (artesunate, artemisinin and artemether) were pooled in order to compare the rapidity of clearance of Plasmodium falciparum parasitaemia and the incidence of reported adverse events with each treatment. Data from patients who received comparator treatment (parenteral artemisinin derivative or quinine) were also included. Primary endpoints included percentage reductions in parasitaemia at 12 and 24 hours. A parasite reduction of >90% at 24 hours was defined as parasitological success. Results Artemisinin and artesunate treatment cleared parasites more rapidly than parenteral quinine during the first 24 hours of treatment. A single higher dose of rectal artesunate treatment was five times more likely to achieve >90% parasite reductions at 24 hours than were multiple lower doses of rectal artesunate, or a single lower dose administration of rectal artemether. Conclusion Artemisinin and artesunate suppositories rapidly eliminate parasites and appear to be safe. There are less data on artemether and dihydroartemisinin suppositories. The more rapid parasite clearance of single high-dose regimens suggests that achieving immediate high drug concentrations may be the optimal strategy.


Background
In 1985, the Journal of Traditional Chinese Medicine described the satisfactory efficacy of qinghaosu suppositories in 100 patients with P. falciparum malaria, 4 of whom had cerebral malaria [1]. Qinghaosu (artemisinin) derivatives were soon recognised as having powerful antimalarial activity [2] and a variety of formulations have since been developed [3]. The efficacy and safety of oral and parenteral artemisinin derivatives have been widely studied for both uncomplicated and severe malaria [4][5][6][7] and these drugs form the basis of current antimalarial treatment policy in most countries in the world [8].
During the past 10 years, the WHO has directed the development of the highly active artemisinin derivative, artesunate, to assess its value in settings where it might be given rectally as a substitute for injectable treatment. The rationale for the development was that, without effective treatment, P. falciparum malaria can progress to severe malaria and death within a matter of hours. The artemisinin derivatives have been shown to have potent activity against early trophozoite forms and to rapidly reduce heavy parasite infections [3]. Therefore it was postulated that when given as a suppository in areas where patients cannot immediately access injectable therapy, they might confer therapeutic advantages in preventing parasite development to the more pathological stages that cause organ complications in severe malaria [9]. Rectal preparations have the advantage of being easy to administer in rural areas; therefore it is anticipated that rectal administration of an artemisinin derivative in remote settings might "buy time" by halting or slowing the progress of disease while a patient is being transported to a health-care facility equipped to provide definitive treatment. Their utility would consequently be greatest in areas where access to injectable therapy is poor or does not exist. The clinical evidence accumulated in the initial phase of this development focused on measures of parasite reduction -a wellestablished indicator of clinical effect in the evaluation of antimalarial drugs.
In addition to artesunate, other artemisinin derivatives formulated for rectal administration now include artemisinin, dihydroartemisinin and artemether. The available published results suggest that all achieve a rapid, consistent clinical response in regions where studies have been undertaken, despite considerable inter-individual pharmacokinetic variability [10][11][12][13][14]. However the different artemisinin derivatives have different physicochemical properties in adminstration; consequently some preparations might be more rapidly absorbed than others [11,13,[15][16][17][18]. Most clinical trials have employed small sample sizes and none have directly compared the efficacy of the different rectally administered artemisinin derivatives with one another. In addition, substantial heteroge-neity exists with respect to the dosing schedules employed. There is therefore considerable uncertainty regarding the optimal preparation and dosing schedules to use.
Given the different aims, design, location, patient demographics and disease severity of clinical trials, we review individual patient data from several trials to establish whether there are significant differences in efficacy and safety of different artemisinin-based suppositories in the first 24 hours post treatment. More specifically we evaluate: i) the efficacy of rectal artemisinins in comparison with conventional treatments for severe malaria (including parenteral quinine and parenteral artemisinins); ii) the comparative efficacy of different artemisinin derivatives for rectal use (namely, artesunate vs artemisinin vs artemether); iii) the efficacy of different dosing regimens (single vs multiple dosing) and total dose administered; iv) the overall safety profile of rectally administered artemisinins and differences between different drugs and regimens.

Search strategy and selection criteria
Electronic searches of the National Library of Medicine's MEDLINE database, Current Contents database and manual searches of selected specialty journals were performed to identify all the pertinent literature. MEDLINE database engines (Ovid, PubMed and GratefulMed) were used with the keywords "rectal", "artemisinin", and "treatment". The search was further refined with the words "artesunate", "artemether", "artemisinin", "dihydroartemisinin", "suppositories", and "rectocaps" from 1980 to March 2006. Reference lists from qualitative topic reviews and published clinical trials in English were also searched. We attempted to obtain the original individual patient data from all studies, regardless of publication status.
Selection criteria for inclusion of studies were clinical trials that assessed the efficacy of a rectal artemisinin-based preparation where individual patient data (inclusion/ exclusion criteria, age, parasitological status at baseline and parasitology post treatment) were available. Comparative and non-comparative studies were included, regardless of study design, geographical area, patient age group, disease severity or the artemisinin derivative used. Safety data and information on dosage was specifically requested from the individual study investigators if this was not provided or evident from a publication. Data extraction was conducted by two investigators (MP, IR) for efficacy data and two investigators (MW, MG) for safety.

Efficacy endpoints
The main focus of the review was on the assessment of parasite reduction and clinical response of patients during the first 24 hours following treatment. This was considered most appropriate, given the intended indication as emergency pre-referral treatment, where ameliorating disease progression within 24 hours, i.e. while the patient is being transported to a clinic or hospital, is likely to be most important.
All efficacy definitions used are consistent with the scientific literature. Parasite reduction ratio (PRR) at 12 and 24 hours was assessed as the percent reduction of parasitaemia at 12 and 24 hours from baseline parasitaemia. Parasitological success was defined as the absence of clinical deterioration from baseline and a PRR at 24 hours of ≥ 90%. In the analysis (see further below) both the continuous variable PRR and the binary variable of ≥ 90% parasite reduction were used, the former to define efficacy at 12 hours, and the latter to define efficacy at 24 hours.
Because definitions of parasite and fever clearance times differed from study to study, these time-to-event variables were re-derived from the serial parasite density estimations and temperatures for individual patients. Parasite clearance and fever clearance time were defined as the time at which the first negative blood smear or normal temperature (<37.5°C axillary or <38.0°C oral) was recorded. The effect of consolidation treatment on recrudescence of the infection during follow-up was evaluated from baseline to the reappearance of parasitaemia.

Safety endpoints
Except in two trials, laboratory markers of safety (including haematological and biochemical indices) were not available. Safety analyses were consequently restricted to clinical descriptions of reported adverse events. In the absence of prospective standardised methods for defining, assessing, reporting and classifying adverse events across all trials, and due to inherent difficulties in clinically distinguishing drug side-effects from manifestations of malarial infection, principal investigators who contributed data were asked to re-review individual patient data retrospectively and reassess all reported adverse events. Ideally this was performed directly from case record forms where archived data were accessible. Each reported event was thus re-classified by the clinical investigator as being either "unlikely", "possibly", "probably" or "definitely" due to the treatment. Those events considered possibly, probably or definitely drug-related were thereafter re-classified as "potentially drug-related" for the purposes of the pooled analysis.

Statistical analysis
Original data from different studies were merged into a master data set. In the analysis mainly hierarchical models were used, with treatment arm as the second level of clustering. In some cases within-study comparisons of different treatment arms was possible and methods for metaanalysis were then accordingly applied in the analysis of PRR. A hierarchical mixed model applying the DerSimonian & Laird method was used with the estimate of heterogeneity being taken from the inverse variance fixed-effect model [19].
Most of the analyses, however, were based on comparisons of treatment arms from different studies, here called pooled analyses, and conducted as follows: For PRR, linear mixed effect models with random intercepts were applied. In the analysis at 12 hours, an identity link (normal distribution) was used. At 24 hours a large proportion of the PRR values for the artemisinins were close to 100% at 24 hours, and these were then categorised into binary observations (PRR >90% versus <90%). A logistic link was then used in the analysis. It should be noted that crude mean values of PRR at 24 hours using the identity link are also given for description. We systematically examined the effect (stepwise backward elimination of covariates not reaching a significance level of p = 0.05) of the following covariates within all analyses: baseline parasitaemia, age, region, total treatment dose provided within the initial 12 and 24 hours, and severity of disease as defined by the evaluating clinician.
Time-to-event analyses, including time to parasite and fever clearance and the efficacy of the consolidated treatment in suppressing parasitaemia post treatment (reappearance of parasites), were represented using crude Kaplan-Meier plots (these are provided as graphs only where the difference in estimates was significant). Hazardratios were estimated using a non-hierarchical Cox regression model applying the same stepwise backward elimination of covariates as above. Intra-rectal treatments were compared to parenteral treatment when followed by the same consolidation treatment in one analysis and intrarectal treatments followed by different consolidation treatments were compared in a second analysis. Time to parasite clearance, age, region, disease severity, parasitaemia at 72 hours (often the start of consolidation treatment) were assessed as covariates through stepwise backward elimination of covariates.
All statistical computations were performed with Stata for Windows (version 8 and 9.2, Stata Corporation, College Station, Texas-USA).
Altogether the studies included in this analysis enrolled a total of 1167 patients in 37 separate treatment arms. Five patients who simultaneously received rectal artesunate and quinine were excluded, leaving a total of 1162 patients included in the analysis. Of these 786 had been treated with rectal administration of an artemisinin derivative and 376 with a comparator drug which was either a parenteral artemisinin (236) oral artesunate (17) or parenteral quinine (123). The majority of included patients treated with a rectal artemisinin were from mainland South East Asia (487) or Papua New Guinea (89), with relatively fewer patients from Africa (210). Thirtyone percent of patients were children under 5, 11.8% children aged 6-10, 21.3% adolescents aged 11-20 years and 36% adults over 20 years of age. Treatment exposure information is provided in Table 2.

Efficacy 1 comparisons with quinine
Two studies (Molyneux1997-8 and Barnes1998) contributed to a standard meta analysis as both studies compared clinical and parasitological response of artesunate 10 mg/ kg versus quinine 10 mg/kg. The log-transformed PRR at 24 h with a single dose of rectal artesunate was significantly better than quinine, weighted mean difference 0.60 (95% CI 0.32-0.89, p ≤ 0.0001). The pooled analyses showed that the artemisinin derivatives, regardless of route of administration and number of doses, were superior to quinine in reducing parasitaemia at 12 and 24 hours ( Figure 1 and Table 3). In the model, parasitological efficacy was partly dependent on age and severity of disease but independent of baseline parasitaemia and region of use.
Time to clearance of parasitaemia for the different drugs are given in Figure 1. Multiple Cox regression analysis showed a significant difference in time to clearance between parenteral artemisinins and parenteral quinine (Hazard Ratio HR = 4.1; p ≤ 0.0001), between single dose rectal artesunate and parenteral quinine (HR = 2.7; p ≤ 0.0001) and between single dose artemisinin suppositories and quinine (HR = 2.4;p = 0.03) with parasitaemia at baseline being a significant covariate in the parenteral comparison (HR = 0.99; p ≤ 0.0001) and in the comparison with single dose artesunate (HR = 0.99; p = 0.042), but not in the comparison with single dose artemisinin suppository treatment.

Comparisons between artemisinin derivatives
Mixed model estimates comparing efficacy of the different artemisinin derivatives are provided in Table 4.

Parenteral versus rectal administration Parenteral artesunate or artemether versus single-dose artemisinin or artesunate rectal administration
Mean PRR using either a single dose artesunate or single dose artemisinin was higher than parenteral artemisinins at 12 hours (65.9% for rectal treatment compared with 60.0% for parenteral treatment). This was observed also in the adjusted model presented in Table 4, in which severe disease was an important covariate in response prior to 12 hours. At 24 hours the difference was in the same direction, with 90.0% reduction in parasitaemia with rectal artemisinins compared with 83.8% parasite reduction with parenteral artemisinins. The proportion PRR>90 at 24 hours gave an OR = 1.75 (p = 0.072) in favour of rectal administration. However, the Kaplan Meier survival plot provided in Figure 2 demonstrates the overall superiority of parenteral administration beyond the 24-hour period (unadjusted log rank test of survival function p ≤ 0.0001 for a single dose of rectal artesunate/ artemisinin versus parenteral treatment). Baseline parasite

Effect of consolidation treatment on recrudescence
Consolidation treatment varied (Table 5). A multiple Cox regression analysis of time-to-recrudescence gave a nonsignificant difference between an intra-rectal and parenteral artemisinin derivative when both were followed by mefloquine (HR = 0.78, p = 0.639) but parasitaemia at 72 hours was a significant covariate in the analysis (HR = 0.99, p = 0.003). However, in a comparison of intra-rectal treatment followed by sulphadoxinepyrimethamine (SP) versus intra-rectal treatment followed by mefloquine, the Kaplan Meier analysis (log-rank test p ≤ 0.0001) and Cox regression analysis (HR = 0.36, p = 0.006) of time to recrudescence between the two treatments showed a significant difference in favour of mefloquine, with parasitaemia at 72 hours (HR = 0.99, p = 0.005) and region of study (HR = 0.16, p = 0.001) being significant covariates in the analysis (Figure 3).

Safety
A total of 196 adverse events were reported in 140 (17.8%) of the 786 patients treated with rectal artemisinins ( In summary, the total incidence of adverse events considered by clinicians to be possibly drug-related was estimated at being between 2.7% and 9.0% of all rectal artemisinin-treated patients, compared with 22% of qui-  nine-treated patients. The majority of possibly drugrelated adverse events in rectal artemisinin-treated patients involved either the gastrointestinal system or were generalized and non-specific in nature and were not severe.

Discussion
This review addresses the lack of any data directly comparing the therapeutic efficacy and safety of the different rectal preparations of artemisinin derivatives. The pooled analysis of individual patient data suggest that artemisinin and artesunate suppositories rapidly eliminate parasites and are safe. There is far less evidence for artemether [32] and no studies of dihydroartemisinin suppositories were available to be included in this analysis. The results indicate that both artemisinin and artesunate, whether as single or multiple dose regimens, induce a superior parasitological response than parenteral quinine over the 24 hours following initiation of treatment. Regimens employing a higher single dose of rectal artesunate were five times as likely to result in >90% parasite reductions at 24 hours than were multiple lower doses of rectal artesunate or than a single administration of artemether. These results imply that dosage regimens that result in immediate high blood concentrations of drug [10,30,34,38] are those best able to reduce parasitaemia in patients with evolving severe malaria and that sustained drug exposure achieved by sequential treatment with moderate doses [20,21,35,39] offers no therapeutic advantage.
The analysis used the rate of parasite clearance in the first 24 hours following treatment as the primary endpoint to compare therapeutic efficacy of alternative drugs and regimens. This endpoint has been commonly used in studies of antimalarial efficacy, particularly for treatments intended for severe malaria [40]. Parasite clearance is a surrogate marker of clinical response but it cannot be assumed that superior parasite clearance equates with improved clinical outcome and lower mortality. Although parenteral artemisinin derivatives have long been recognised as having superior parasite clearance to quinine, it remained uncertain until recently whether this characteristic converted into a survival benefit. A recent trial comparing iv artesunate with parenteral quinine demonstrated a 30% lower mortality with artesunate, confirming that more rapid initial parasite clearance may translate to reduced mortality in severe adult malaria [41]. Therefore, although the current pooled analysis was not powered to assess mortality as an endpoint, the differences in parasite clearance rates between rectal artemisinins and parenteral quinine, and between different rectal artemisinin dosing regimens, should be regarded as important indicators of possible real differences in therapeutic efficacy and clinical benefit. It should also be noted that any future study designed to use mortality as an end-Cumulative probability of having parasites: parenteral artem-isinins versus rectal artemisinins Figure 2 Cumulative probability of having parasites: parenteral artemisinins versus rectal artemisinins. Cumulative probability of recrudescing with consolidation treatment Figure 3 Cumulative probability of recrudescing with consolidation treatment. Rectal artemisinins followed by Sulphadoxine-pyrimethamine point to compare different rectal artemisinins (or to demonstrate non-inferiority of a rectal artemisinin with a parenteral preparation such as iv artesunate) would require such a large sample size that it is unlikely ever to be implemented. Therefore the surrogate marker of parasite clearance used in this analysis is likely to remain the best available evidence on which to base comparisons of treatment efficacy for the rectal artemisinins.  There has been a recent systematic review of published data on rectal artemisinin derivatives with a focus on pharmacokinetics of various preparations and a summary of efficacy [42]; however there were less data and limited capacity to standardize definitions and account for statistical heterogeneity. In contrast, the current meta-analysis synthesizes individual patient data from studies meeting well-defined inclusion criteria and for whom standardized end-points were calculated. The analysis has enabled a robust and statistically powerful comparison of efficacy outcomes between rectal artemisinins, parenterally administered artemisinins and parenteral quinine that has had the capacity to examine and allow for the influence of covariates such as age, geographic origin and disease severity. Given that only a small number of direct comparative trials have been performed, this meta-analysis of 1162 individual patients represents a significant contribution to the available comparative efficacy data on rectal artemisinins. In particular, its results showing that early parasite clearance of rectal artemisinins is clearly superior to that of quinine, and appears equivalent to that of parenteral artemisinins is an important observation, given the results of the recent SEQUAMAT trial [41].
There are methodological limitations inherent in making comparisons of safety across several trials and in attributing causality to adverse events in patients with malaria.
Overall the data suggest that the artemisinin based suppositories studied have a benign safety profile, consistent with that of the artemisinins in general [43,44]. There were no special concerns related specifically to the rectal route of administration and there were no reports suggestive of serious neurotoxicity. Neurotoxicity of the artemisinin derivatives has been described in animals but this now appears to be associated with sustained exposure to the central nervous system rather than peak levels [45]. Therefore a single dose as pre-referral treatment (rather than multiple dosing) may also have additional theoretical benefits in terms of safety as well as efficacy. Pharmacokinetic data have largely been derived from studies of artesunate [10,20,21,30,31,38,47] and artemisinin [14][15][16][17][18]48,49] with little data on rectal formulations of artemether [32] or dihydroartemisinin [22]. In the absence of sufficient pharmacokinetic information it cannot be assumed that all rectal preparations have the same efficacy or safety profile. Well-designed clinical trials that directly compare the efficacy, safety and pharmacokinetic profile of the different suppository formulations are needed.
The evidence from this analysis supports the WHO recommendation for the use of artesunate and artemisinin as initial pre-referral treatment [8]. The analysis was not designed to assess long-term cure rates and there are insufficient data on which to substantiate the use of rectal treatment for full management of severe malaria.

Conclusion
Early effective treatment with artemisinin based suppositories has potential as a lifesaving intervention, particularly at the periphery of the health-care system, where suppositories might be administered early in lieu of parenteral treatment in remote communities by relatively untrained personnel. Combined with accurate diagnosis and artemisinin combination therapy, rectal artemisinins have been effectively used to reduce malaria incidence and mortality in Asia [3,50,51], an approach which holds great promise for malaria control elsewhere.