Safety and efficacy of intermittent presumptive treatment with sulfadoxine-pyrimethamine using rapid diagnostic test screening and treatment with dihydroartemisinin-piperaquine at the first antenatal care visit (IPTp-SP+): study protocol for a randomized controlled trial

This is a phase IIIb open-label randomized controlled superiority trial of standard IPTp-SP vs. IPTp-SP plus screen-and-treat using DP (IPTp-SP+). Participants randomized to the IPTp-SP arm will receive care according to current national guidelines. Participants who are randomized to the IPTp-SP+ arm will receive standard-of-care with the addition of screen-and-treat using RDT and DP at the first ANC visit.

This study protocol has been formatted following SPIRIT reporting guidelines [17].

The trial will take place in Nchelenge District, Luapula Province, Zambia. The district is located in the northern wetlands of Zambia alongside Lake Mweru, an area of hyperendemic malaria [18]. Malaria transmission occurs throughout the year. The predominant vector is Anopheles funestus which peaks during the dry season (May-September) while both An. funestus and An. gambiae are found during the rainy season [19]. The population of the district is estimated to be over 296,000 inhabitants consisting mostly of subsistence farmers and fishermen and women. Nchelenge District has one hospital, Saint Paul’s General Hospital (SPGH), twelve rural health centers, and two health posts. The trial will recruit and follow obstetric patients and their offspring at two of the rural health centers and SPGH. SPGH is equipped with an operation theater and labor-and-delivery ward for the provision of essential obstetric services.

The following procedures will be used to ensure an unbiased assessment of treatment safety and efficacy. A computer-generated randomization schedule will be generated by the study statistician prior to the start of the study and kept in a locked cabinet accessible only to the study coordinator who will be responsible for assigning participants to a study treatment arm. The schedule will comprise blocks of varying size to allocate, in a 1:1 ratio, eligible participants to either IPTp-SP or IPTp-SP+. The trial is open-label. However, study investigators, study clinicians, and statisticians are masked to treatment assignment. Investigators will be unmasked after all participants have delivered. A Data and Safety Monitoring Board (DSMB) will be established to review safety data. Statisticians will remain masked until after the DSMB has approved the final analysis plan and the database is locked. All outcome assessors, including laboratory parasitologists and pathologists, will also be masked until the final database is locked.

The intervention will consist of IPTp-SP with or without screen-and-treat using DP at the first ANC visit. Participants randomized to the IPTp-SP+ group who test positive by rapid diagnostic test (RDT) will be given a full treatment course of DP (D-ARTEPP® 40 mg/320 mg three tablets daily for 3 days; Guilin Pharmaceutical) under direct observation (DOT) by a study nurse (day 0) or community health worker (days 1–2). All other participants will be treated with SP at the first visit, including those in the IPTp-SP+ arm with a negative RDT result, and all participants in both groups will be given SP at all subsequent visits (day 35, day 63, then monthly until delivery) under DOT. IPTp-SP will be administered according to national guidelines (G-SCOPE® 1,500 mg sulfadoxine/75 mg pyrimethamine for one dose no fewer than 4 weeks between doses; Guilin Pharmaceutical). Participants will be observed for 1 h post-dose to monitor for vomiting or other adverse reactions. If vomiting occurs within 30 min, the full dose will be readministered. If vomiting occurs after 30 min, half of the total dose will be readministered. In the case of persistent vomiting, the participant will be withdrawn from the study and referred for alternative treatment.

Treatment with either SP or DP will be administered as described above. Participants will return for follow-up on days 14, 28, 35, 42, and 63 and then monthly thereafter until delivery. They will be instructed to report to the clinic for any illness between scheduled visits.

At each scheduled or unscheduled visit, the study clinician will perform an interval history and physical examination, including an updated medication history. During the active follow-up period (up to day 63), DBS will be collected and hemoglobin will be measured. SP will be given on days 35 and 63. Additional doses of SP will be given during monthly visits until delivery with a minimum of 4 weeks between doses and laboratory samples (e.g., blood slide for microscopy, urinary tests, full blood count) will be collected if clinically indicated according to signs and symptoms. Antimalarials or antibiotics with antimalarial activity (e.g., systemic erythromycin, macrolide antibiotics, trimethoprim-sulfamethoxazole or other sulphonamides, tetracyclines, quinolones, clindamycin) will be prohibited during the active follow-up period.

Participants who present during the follow-up period with any signs or symptoms of malaria will undergo testing by thick smear, and DBS will be collected from those with positive microscopy. The signs and symptoms that we will consider indicative of possible clinical malaria include fever, chills, weakness, fatigue, myalgia, arthralgia, headache, anorexia, nausea, vomiting, abdominal pain, or diarrhea. All confirmed uncomplicated malaria cases will be treated with artemether-lumefantrine according to national guidelines (Coartem™ 20 mg/120 mg four tablets taken at 0, 8, 24, 36, 48, and 60 h). Participants who develop severe malaria defined according to WHO criteria will be referred to SPGH for inpatient management [20].

On the day of delivery, the participant and infant will be evaluated as soon as possible after delivery. The birthweight will be measured, and an assessment for congenital abnormalities will be done. Obstetric complications including but not limited to hemorrhage, premature rupture of membranes, cesarean section, and others will be captured as adverse events. Congenital abnormalities, if present, will also be recorded. Thick and thin peripheral blood films, DBS, and hemoglobin measurement will be done on both the participant and infant. In addition, cord blood, placental blood films for thick and thin smear microscopy, placental DBS, and placental biopsy for histopathological analysis will be collected.

Infants will be reassessed at 1, 6, 9, and 12 months postpartum for focused history and physical examination, including interval medication and hospitalization history, growth, and assessment of developmental milestones. At the first monthly visit, hemoglobin measurement and malaria parasitology by thick smear and PCR testing using DBS prepared from heel stick blood will be performed.

Drug safety will be monitored at every visit during the course of the study in compliance with International Conference on Harmonization Good Clinical Practice guidelines for adverse events (AEs) and serious adverse events (SAEs) [26]. All expected and unexpected AEs will be collected and reported in trial publications. SAEs will be reported to the sponsor and the ethics committee within 24 working hours of the study staff first becoming aware. Investigators will determine the relationship between safety signals and the study drugs and define outcomes, according to standard classifications [26].

The sample size was determined based on available data and with the objective of achieving a detectable difference of a 50% reduction in the incidence of MIP. In 2019, Nchelenge District recorded 7791 live births (unpublished data). Prior studies in the same population found PCR prevalence of P. falciparum parasitemia to be 22–26% in pregnant women [14, 15]. A sample size of 324 pregnant women (n=162 per arm) will afford 80% power to detect a 50% difference of effect between study arms with a two-tailed alpha of 0.05. To allow for loss to follow-up of up to 20%, a total of 392 (n=196 per arm) will be recruited.

Source data will be recorded using paper case report forms (CRFs) which will then be double-entered into a password-secure electronic database. The CRFs and database will be routinely checked for accuracy by the investigators during the data collection period. The final database will be locked after resolution of all queries. All paper CRFs will be filed and kept in lockable cabinets in offices accessible only to study personnel. At the conclusion of the study, the files will be transferred to the Tropical Diseases Research Centre where they will be stored for a period of 5 years after study completion.

To facilitate retention in the study, participants will be issued study visit cards that include the dates of their follow-up visits. We will collect participants’ telephone numbers and physical addresses to allow the study team to contact participants in case of missed visits, advising them to report to the clinic for their scheduled visit within the window period of 3 days. Participants are allowed to leave the study at any time for any reason without any consequences. Participants will be withdrawn from the study only if they withdraw informed consent.

Longitudinal data will be displayed using the nonparametric Kaplan-Meier estimations of the survival function. The primary analysis will be comparison of the 42-day incidence of MIP using multivariate models to estimate the relative hazard between the IPTp-SP and IPTp-SP+ groups according to intention-to-treat (ITT). Testing for proportionality of hazards will be done using formal statistical testing with Schoenfeld residuals as well as visual inspection of the nonparametric survival estimates. For non-proportional hazards, an extended Cox model will be applied. To handle interval censoring due to participants who miss one or more follow-up visits, we will use parametric models for interval-censored survival-time data. The relative hazard will be estimated in parametric models accounting for recurrent events by using a robust variance estimator. For the primary safety analysis, we will compare the proportions of serious and non-serious AEs using logistic regression models and relative hazards of AEs using Cox regression methods as above. Secondary analyses of continuous outcomes (hemoglobin concentration, birthweight) will be examined in linear regression models and in logistic regression models for binary outcomes (prevalence of low birth weight, neonatal mortality, placental malaria, congenital malaria, maternal anemia, congenital anemia, drug resistance allele frequency). Prespecified subgroup analyses will be done according to the following strata: primigravidae, multigravidae, gestational age at enrollment, delivery before or after the day 63 visit, treatment dose on a mg/kg basis, positive P. falciparum PCR on enrollment, and presence or absence of quintuple and sextuple mutations. In a secondary analysis, we will compare the proportions of treatment failures and prevention failures according to the antimalarial drug given at the first encounter (SP or DP) in an on-treatment analysis using logistic regression. Treatment failures will be defined as recurrent infections that are determined by genotyping to be recrudescent parasites from the initial infection in participants who tested positive by PCR at the initial visit. Prevention failures will be defined as PCR-confirmed parasitemia in participants who initially tested negative by PCR at the first visit, or who tested positive by PCR at the first visit but who experienced reinfection with a new parasite as determined by genotyping. No interim analysis is planned. Every effort will be made to collect complete data for all participants and minimize missing data. Missing data will be handled using multiple imputation.

This protocol has been approved by the European and Developing Countries Clinical Trial Partnership Ethics Review Committee (ERC) and locally by the Tropical Diseases Research Centre ERC, the Zambia Medicine Regulatory Authority, and the National Health Research Ethics Board of the Ministry of Health of the Government of the Republic of Zambia. Any amendment to the study protocol will be submitted to the ERC for approval before its implementation. Data will be de-identified for purposes of publication, data sharing, and secondary analyses. Records containing names or other personal identifiers will be securely stored separately from de-identified data. Expedited or full review, as appropriate, by an ethical review board will be sought for secondary studies. Secondary studies may use stored biological samples from participants who provided consent for future use of blood samples using de-identified data [27].

Four site visits will be conducted by an independent external monitor who will carry out a minimum of 20% source data verification. A data and safety monitoring board (DSMB) composed of four members will be established. The DSMB will meet quarterly to review progress and provide independent assessments of the quality of the data produced and the safety of study treatments.

Prevention of malaria during pregnancy is important to maternal and child health in endemic areas, but the spread of drug resistant P. falciparum has compromised the effectiveness of the current standard approach of IPTp-SP [28–32]. Clinical trials of alternative strategies are therefore needed. This trial protocol describes a hybrid IPTp-SP approach that combines standard IPTp-SP with screening and treatment using DP at the first ANC visit.

There are four justifications to this approach. First, higher density infections that occur early in gestation around the time of the first ANC visit are believed to pose a greater risk to pregnancy than infections that occur later in pregnancy or are lower in density [33–35]. These earlier, higher density infections are often detectable by RDT and are more effectively cleared by curative artemisinin-based combination therapies (ACTs) than by SP, particularly in areas with high quintuple and sextuple mutants. Second, compared to other ACTs, DP provides a period of post-treatment prophylaxis similar to SP owing to piperaquine’s long half-life [36–38]. ACTs with shorter elimination half-lives than DP may be insufficient to bridge the period between ANC visits. Third, screening and treatment during the first visit only—rather than all ANC visits—has the ability of detecting over 50% of all P. falciparum infections diagnosed during pregnancy and it is believed to minimize programmatic complexity and resource consumption without sacrificing utility [34]. Mathematical models that compared screening and treatment at the first visit only to screening and treatment at every visit predicted only a marginal difference between the two approaches [33]. Fourth, drug resistance in P. falciparum to antifolates is a graduated, rather than all-or-none, phenomenon. Even in areas of prevalent SP resistance genetic markers, IPTp-SP retains a clinically significant level of effectiveness [11]. Some of its effectiveness may be independent of its antimalarial properties. Thus, abandoning SP altogether may not be advisable because it would relinquish a still useful drug while exposing other antimalarials to a greater potential of emerging resistance.

Various alternatives to standard IPTp-SP are under consideration including the hybrid approach we will test in this protocol [16]. Alternative strategies evaluated in sub-Saharan Africa have so far included IPTp using alternative antimalarials (e.g., DP, artemether-lumefantrine, mefloquine, azithromycin-chloroquine, amodiaquine), intermittent screening and treatment in pregnancy (ISTp) with SP or other agents at one or more ANC visits, or a combination of these approaches [39–47]. Mefloquine (MQ) and DP were shown to be superior to SP for IPTp in reducing incident parasitemia and clinical malaria in areas of widespread SP resistance [42–44]. MQ, however, was less well tolerated and DP was associated with lower birth weight compared to SP [43, 44]. Prior studies also identified the rapid emergence of purported genetic markers of piperaquine resistance within IPTp-DP cohorts [48, 49]. ISTp with artemisinin-based combination therapy at the first ANC visit, in which those who screened negative received no drug at all, was associated with a higher proportion of MIP compared to standard IPTp-SP, attributed to P. falciparum parasitemia that went undiagnosed and untreated due to the incomplete sensitivity of RDTs used in screening [44–46].

A combined approach of ISTp at the first ANC visit and IPTp-SP at subsequent visits was proposed by WHO as one potential cost-effective MIP prevention strategy to be tested in areas with very high SP resistance [50, 51]. One such approach was implemented as national policy in Tanzania for surveillance purposes in 2014, referred to as “single screen and treat in pregnancy” [52]. Women are screened using an RDT at their first ANC visit, and those who screen positive are treated with either quinine (for women in their first trimester) or an artemisinin-based combination therapy (for women in their second or third trimester) while those who screen negative receive standard IPTp-SP starting from the second trimester. This hybrid strategy was recently evaluated in mathematical models by Walker et al. [33]. The authors modeled five different MIP prevention strategies: IPTp-SP, IPTp-DP, ISTp-DP, and two hybrid strategies of IPTp-SP combined with ISTp-DP either at the first visit only or at every visit. In modeled scenarios that assumed a high frequency of quintuple resistance, the two hybrid strategies performed near equally to each other, while IPTp-DP performed the best and IPTp-SP and IST-DP performed the poorest [33]. The modeled predictions of ISTp and IPTp have been borne out in clinical trials [44, 45], while hybrid strategies have been less closely studied to date.

There are limitations to this study design. A number of previous trials of IPTp included pregnant women who had symptoms of malaria at the time of enrollment. We will exclude women who are symptomatic at the time of their first ANC visit because they must undergo routine clinical evaluation, which includes testing for malaria, and therefore cannot be withheld from malaria screening as would be required if they were randomized to the control group. We will also exclude women who received an antimalarial drug during the current pregnancy to control for contamination bias. These exclusions will reduce the generalizability of the study. Due to the nature of the experimental intervention—ISTp-DP added to standard IPTp-SP—there will be participants who are randomized to the intervention arm but who will receive identical care to that of the comparator arm apart from a single instance of RDT screening. That is, participants randomized to the IPTp-SP+ arm but who screen negative will receive IPTp-SP. This is anticipated to attenuate the observed protective effect of the intervention on the subsequent incidence of MIP due to the mixing of participants who receive DP with those who receive SP at the first ANC visit. However, this has been accounted for in the sample size calculation, and has the advantage of mimicking the real-world application of the strategy to test its effectiveness. We will also account for this by conducting a subgroup analysis comparing IPTp-SP to IPTp-SP+ among participants with a positive P. falciparum PCR at the first visit.

Alternative chemopreventive strategies for MIP are urgently needed given the widespread prevalence of SP resistant P. falciparum in sub-Saharan Africa. This trial will test a strategy of adding screening and treatment with DP at the first antenatal visit to the standard IPTp-SP to reduce the incidence of MIP and thereby protect against malaria-attributable complications of pregnancy and child development.

Additional file 1. : Supplementary Table S1. SPIRIT figure