Safety and efficacy of primaquine in patients with Plasmodium vivax malaria from South Asia: a systematic review and individual patient data meta-analysis

Background The optimal dosing of primaquine to prevent relapsing Plasmodium vivax malaria in South Asia remains unclear. We investigated the efficacy and safety of different primaquine regimens to prevent P. vivax relapse. Methods A systematic review identified P. vivax efficacy studies from South Asia published between 1 January 2000 and 23 August 2021. In a one-stage meta-analysis of available individual patient data, the cumulative risks of P. vivax recurrence at day 42 and 180 were assessed by primaquine total mg/kg dose and duration. The risk of recurrence by day 180 was also determined in a two-stage meta-analysis. Patients with a >25% drop in haemoglobin to <70 g/L, or an absolute drop of >50 g/L between days 1 and 14 were categorised by daily mg/kg primaquine dose. Results In 791 patients from 7 studies in the one-stage meta-analysis, the day 180 cumulative risk of recurrence was 61.1% (95% CI 42.2% to 80.4%; 201 patients; 25 recurrences) after treatment without primaquine, 28.8% (95% CI 8.2% to 74.1%; 398 patients; 4 recurrences) following low total (2 to <5 mg/kg) and 0% (96 patients; 0 recurrences) following high total dose primaquine (≥5 mg/kg). In the subsequent two-stage meta-analysis of nine studies (3529 patients), the pooled proportions of P. vivax recurrences by day 180 were 12.1% (95% CI 7.7% to 17.2%), 2.3% (95% CI 0.3% to 5.4%) and 0.7% (95% CI 0% to 6.1%), respectively. No patients had a >25% drop in haemoglobin to <70 g/L. Conclusions Primaquine treatment led to a marked decrease in P. vivax recurrences following low (~3.5 mg/kg) and high (~7 mg/kg) total doses, with no reported severe haemolytic events. PROSPERO registration number CRD42022313730.


ADDITIONAL FILE 1
Verma R et al, Safety and efficacy of primaquine in patients with P. vivax malaria from South Asia: A systematic review and individual patient data meta-analysis Page Checklist S1.PRISMA-IPD 2

Box S1. Search Strategy 7
Table S1.Studies included in the one stage individual patient data meta-analysis 8 Table S2.Study testing and inclusion criteria 9 Table S3.Study sites included in the one stage individual patient data metaanalysis Table S4.Reasons for studies not being included in the one stage efficacy metaanalysis Table S5.Studies targeted for the one stage efficacy analysis but not included Table S6: Comparison of baseline characteristics between studies included and studies targeted for the one stage meta-analysis Table S7: Risk of bias assessment in randomised controlled studies in one or two stage meta-analyses Table S8: Risk of bias assessment in single arm observational studies in one or two stage meta-analyses Table S9: Sensitivity analysis for cumulative risk of first P. vivax recurrence between day 7 to 42 and between day 7 to 180 Provide an explicit statement of the questions being addressed with reference, as applicable, to participants, interventions, comparisons, outcomes and study design (PICOS).Include any hypotheses that relate to particular types of participant-level subgroups.

8
BMJ Publishing Group Limited (BMJ) disclaims all liability and responsibility arising from any reliance Supplemental material placed on this supplemental material which has been supplied by the author(s) BMJ Glob Health doi: 10.  • Use of a one-stage or two-stage approach.
• How effect estimates were generated separately within each study and combined across studies (where applicable).
• Specification of one-stage models (where applicable) including how clustering of patients within studies was accounted for.
• Use of fixed or random effects models and any other model assumptions, such as proportional hazards.
• Methods for quantifying statistical heterogeneity (such as I 2 and t 2 ).
• How studies providing IPD and not providing IPD were analysed together (where applicable).
• How missing data within the IPD were dealt with (where applicable).

11-12
Exploration of variation in effects

A2
If applicable, describe any methods used to explore variation in effects by study or participant level characteristics (such as estimation of interactions between effect and covariates).State all participant-level characteristics that were analysed as potential effect modifiers, and whether these were pre-specified.Give numbers of studies screened, assessed for eligibility, and included in the systematic review with reasons for exclusions at each stage.Indicate the number of studies and participants for which IPD were sought and for which IPD were obtained.For those studies where IPD were not available, give the numbers of studies and participants for which aggregate data were available.Report reasons for non-availability of IPD.Include a flow diagram.

Study characteristics 18
For each study, present information on key study and participant characteristics (such as description of interventions, numbers of participants, demographic data, unavailability of outcomes, funding source, and if applicable duration of follow-up).Provide (main) citations for each study.Where applicable, also report similar study characteristics for any studies not providing IPD.

IPD integrity A3
Report any important issues identified in checking IPD or state that there were none.

Risk of bias within studies 19
Present data on risk of bias assessments.If applicable, describe whether data checking led to the up-weighting or downweighting of these assessments.Consider how any potential bias impacts on the robustness of meta-analysis conclusions.
Tables S7  and S8 Results of individual studies 20 For each comparison and for each main outcome (benefit or harm), for each individual study report the number of eligible participants for which data were obtained and show simple summary data for each intervention group (including, where applicable, the number of events), effect estimates and confidence intervals.These may be tabulated or included on a forest plot.
18 and Figure 3 Results of syntheses 21 Present summary effects for each meta-analysis undertaken, including confidence intervals and measures of statistical heterogeneity.State whether the analysis was pre-specified, and report the numbers of studies and participants and, where applicable, the number of events on which it is based.

14-20
When exploring variation in effects due to patient or study characteristics, present summary interaction estimates for each characteristic examined, including confidence intervals and measures of statistical heterogeneity.State whether the analysis was pre-specified.State whether any interaction is consistent across trials.
Provide a description of the direction and size of effect in terms meaningful to those who would put findings into practice.

Risk of bias across studies 22
Present results of any assessment of risk of bias relating to the accumulated body of evidence, including any pertaining to the availability and representativeness of available studies, outcomes or other variables. Ref

Search strategy
All prospective antimalarial efficacy studies of uncomplicated P. vivax with a minimum of 28 days follow up, published between Jan 1, 2000 and August 23, 2021 were identified by the application of the key terms (listed below) through Medline (Pubmed), Web of Science, Embase and the Cochrane Central.Abstracts of all references containing any mention of antimalarial drugs were manually checked to confirm prospective clinical trials, with review of full text when needed.Studies on prevention, prophylaxis, reviews, animal studies, patients with severe malaria, where schizontocidal treatment was unsupervised or where data were extracted retrospectively from medical records outside of a planned trial were excluded.Inclusion criteria included studies undertaken in South Asia including Bangladesh, Bhutan, India, Nepal, Pakistan, Sri Lanka.Studies were included if they had at least one treatment arm with primaquine commencing within 7 days of schizontocidal treatment, had data on age, sex and parasitemia on day 0, data on schizontocidal and primaquine dosing and reported parasite presence or absence during follow up.The year of the study was taken as the year in which the paper was published, although the start and end date of patient enrolment were also recorded.The review process was undertaken by two independent investigators who also performed data extraction (RV and RJC), and the original review process is documented in more detail in, Commons RJ, Thriemer K, Humphreys G, et al.

Figure S1 .Figure S4 .
Figure S1.Location of study sites in efficacy analysis Figure S2.Mg/kg total dose of primaquine administered Figure S3.Mg/kg drug dosing of primaquine by body weight Figure S4.Flowchart for two stage meta-analysis Specify inclusion and exclusion criteria including those relating to participants, interventions, comparisons, outcomes, study design and characteristics (e.g. years when conducted, required minimum follow-up).Note whether these were applied at the study or individual level i.e. whether eligible participants were included (and ineligible participants excluded) from a study that included a wider population than specified by the review inclusion criteria.The rationale for criteria should be stated.Describe all methods of identifying published and unpublished studies including, as applicable: which bibliographic databases were searched with dates of coverage; details of any hand searching including of conference proceedings; use of study registers and agency or company databases; contact with the original research team and experts in the field; open adverts and surveys.Give the date of last search or elicitation.
8Present the full electronic search strategy for at least one database, including any limits used, such that it could be repeated.10Describe how IPD were requested, collected and managed, including any processes for querying and confirming data with investigators.If IPD were not sought from any eligible study, the reason for this should be stated (for each such study).

Table S1
Give results of any additional analyses (e.g.sensitivity analyses).If applicable, this should also include any analyses that incorporate aggregate data for studies that do not have IPD.If applicable, summarise the main meta-analysis results following the inclusion or exclusion of studies for which IPD were not available.

Table S1 . Studies included in the one stage individual patient data meta-analysis
Publishing Group Limited (BMJ) disclaims all liability and responsibility arising from any reliance Supplemental material placed on this supplemental material which has been supplied by the author(s) BMJACTartemisinin-based combination treatment; Cqchloroquine; GIgastrointestinal; PQ/Pqprimaquine; Primaquine treatment was classified as supervised if all doses were directly observed, partially supervised if >1 dose but not all doses were observed, and not-supervised if 1 dose was observed; *Treatment code describes (schizontocidal drug)_(hypnozoitocidal drug)_(total primaquine dose)_(duration of primaquine treatment eg 14d = 14 days)_(primaquine start day); # Reference 36 is a follow up study up to 450 days including some of the patient cohort in references 35 and 37.

Table S2 . Study testing and inclusion criteria
Hbhaemoglobin; Hcthaematocrit.* Reference 36 is a follow up study up to 450 days including some of the patient cohort in references 35 and 37.

Table S6 : Comparison of baseline characteristics between studies included and studies targeted for the one stage meta-analysis Characteristic Included studies N = 7 Targeted studies that were not included N = 11 Year of Enrolment
Age and female percentage of targeted studies frequency weighted according to number of patients treated with chloroquine alone (No primaquine) and different dosages of primaquine.Year of the enrolment defined as the year study enrolment completed.Age, and female percentage of targeted studies calculated using frequency weighted mean or median according to number of patients.*Mean or median age not available for 5 studies.#Percentage not available for 3 studies.BMJ Publishing Group Limited (BMJ) disclaims all liability and responsibility arising from any reliance Supplemental material placed on this supplemental material which has been supplied by the author(s)

Table S7 . Risk of bias assessment in randomised controlled studies in one or two stage meta-analyses Author-year Bias from randomisation Bias due to deviation from intervention Bias from missing outcome Bias in measurement of the outcome Bias in selection of the reported results Overall bias Follow up to 180 days Table S8. Risk of bias assessment in single arm observational studies in one or two stage meta-analyses Figure S3: Mg/kg drug dosing of primaquine by body weight (n=512)
BMJ Publishing Group Limited (BMJ) disclaims all liability and responsibility arising from any reliance Supplemental material placed on this supplemental material which has been supplied by the author(s)