Executive summary... 1 Background... 3 EPI Serology Study Design... 5 Results Pooled Analysis... 12

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1 WHO Advisory Committee on serological responses to Expanded Programme on Immunization vaccines in infants receiving Intermittent Preventive Treatment for malaria (IPTi) FINAL REPORT October 8, 2009

2 Contents Executive summary...1 Background...3 EPI Serology Study Design...5 Study sites and study population...5 Study design...6 WHO Advisory Committee...11 Results Pooled Analysis...12 Pooled SP vs Placebo comparison for measles:...12 Pooled drugs (SP, LapDap, SP-ART, AQ-ART, MQ) vs placebo comparison for measles...13 Pooled Analysis for other antigens...15 Results Yellow Fever (Navrongo)...16 Conclusions of the WHO Advisory Committee...18 Annex 1: Report of the Technical Consultation on Intermittent Preventive Treatment in Infants (IPTi), Technical Expert Group (TEG) on Preventive Chemotherapy, April 23-24, 2009 Geneva Annex 2: Membership of WHO Advisory Committee on serological responses to Expanded Programme on Immunization vaccines in infants receiving Intermittent Preventive Treatment for malaria...20 Annex 3: Final Pooled Analysis: Assessment of Serological Responses to Expanded Programme on Immunization Vaccines in Infants Receiving Intermittent Preventive Treatment (v.3 submitted July 3, 2009)...21 Annex 4: Summaries of Individual Study Results...22 Navrongo, Ghana...22 Manhiça, Mozambique...24 Bungoma, Kenya...26 Kisumu, Kenya...29 Kilimanjaro, Tanzania...32

3 Executive summary Intermittent Preventive Treatment in infants (IPTi) is the administration of a full course of an antimalarial treatment to infants at specified time points, regardless of the presence of parasites. The objective of IPTi is to reduce the infant malaria burden in the first year of life. Administering IPTi at the time of routine immunization is proposed to be the main delivery strategy. It is therefore of critical importance to confirm that IPTi does not have any adverse interaction with the serological responses to EPI vaccines. As a partner of the IPTi Consortium, and with funding from the Bill & Melinda Gates Foundation, WHO undertook serology assessments in five IPTi efficacy trials conducted in Ghana (Navrongo), Kenya (Bungoma and Kisumu), Mozambique (Manhica), and Tanzania (Kilimanjaro). Infants eligible for the serology studies were selected from the larger study population at each site. To oversee and guide this work, in 2003 WHO established a 5-member, independent Ad Hoc Advisory Committee (see Annex 2 for Terms of Reference and Membership). The Advisory Committee assisted with the design of the project, selection of the subcontractors, and review of all the data arising from five EPI serology studies. The Advisory Committee guided WHO accordingly, and issued an Interim Report in July The Interim Report, based on data at the time from the Navrongo (Ghana), Manhica (Mozambique) and Bungoma (Kenya) studies, concluded that IPTi-SP did not have an adverse impact on serological responses to vaccination against measles, diphtheria, tetanus, pertussis, polio serotypes 1 and 3, Haemophilus influenzae type b and hepatitis B. The studies using other drug combinations and analysis of all the samples have now been completed and reviewed. This Final Report of the Advisory Committee describes the methodology of the IPTi serology studies, summarizes the results of the pooled statistical analysis, and presents the Advisory Committee's final conclusions based on a compilation of all the data available. The report is primarily based on the final pooled analysis of the results, and supported by selected analyses of the individual trials where no pooled analyses were planned (i.e. yellow fever and Hib vaccines). While it cannot be held responsible for the conduct of the field trials or laboratory work, the Advisory Committee concludes the following: 1. Serological data from studies in Navrongo, Manhica, and Kilmanjaro provide strong evidence that IPTi with SP, does not have an adverse impact on serological responses to measles vaccine; 2. Though very limited, the available serological data from Navrongo provides evidence that SP does not have a negative impact on antibodies following vaccination against yellow fever; 3. Serological data from studies in Kisumu and Kilimanjaro provide strong evidence that IPTi with SP-ART, AQ-ART, MQ, or LapDap do not have an adverse impact on serological responses to measles vaccination; 1

4 4. Serological data from Manhica and Kisumu provide strong evidence that IPTi with SP, SP-ART, AQ-ART, or LapDap do not have an adverse impact on serological responses to DTP, polio, Hib, and HepB vaccines. 5. The pooled analyses provide further evidence that IPTi treatments do not impair serological responses to EPI antigens. Overall conclusion: Studies have demonstrated that there is no adverse impact on the serological responses to DTP, polio, Hib, HepB, and measles vaccines when the IPTi drugs SP, SP-ART, AQ- ART, and LapDap are administered to infants at the time of routine vaccination. Concomitant administration of IPTi-SP at the time of yellow fever vaccination, and IPTi- MQ at the time of measles vaccination, have also been shown to have no negative effect on the serological responses. 2

5 Background Intermittent Preventive Treatment in infants (IPTi), is the administration of a full course of an antimalarial treatment to infants at specified time points, regardless of the presence of parasites. The objective of IPTi is to reduce the infant malaria burden in the first year of life. Administering IPTi at the time of routine immunization is one of the delivery strategies. Over the past 10 years, IPTi has been the focus of a number of research efforts. With funding from the Bill & Melinda Gates Foundation, in 2002 the IPTi Consortium (a collaboration of 17 research institutions plus WHO and UNICEF) was formed to undertake a coordinated and comprehensive research agenda on IPTi in order to inform policy development 1. This has included clinical trials in southern-african countries (Gabon, Ghana, Kenya, Tanzania, Mozambique) and large-scale pilot implementation studies involving over 300,000 infants a year in districts of Benin, Ghana, Madagascar, Mali, Malawi, Tanzania, and Senegal. The available evidence on the safety, efficacy and other relevant aspects of IPTi with sulfadoxine-pyrimethamine (SP-IPTi) delivered through the Expanded Programme on Immunization (EPI) has been reviewed independently in 2008 by the Institute of Medicine (IOM) 2 and three times by WHO's Technical Expert Group (TEG) on Preventive Chemotherapy 3 most recently in April 2009 (Annex 1). On the basis of a pooled analysis of 6 published studies 4, IPTi-SP was found to be safe and decreased: incidence of clinical malaria episodes by 30% (95% CI: 19.8%; 39.4%) (similar to the levels of efficacy observed with the use of insecticide-treated bednets); anaemia (<8g/dl) overall by 21.3% (95% CI: 8.3%; 32.5%); all cause hospital admissions in the first year of life by 23% (95% CI: 10.0%; 34.0%) (noting that admissions were not, however, all due to severe malaria). 1 See web site: and IPTi Fact Sheet (Feb. 2009) for more information. 2 Institute of Medicine. Assessment of the Role of Intermittent Preventive Treatment for Malaria in Infants: Letter Report. Committee on the Perspectives on the Role of Intermittent Preventive Treatment for Malaria in Infants, July 11, ( 3 WHO. Report of the Technical Consultation on Intermittent Preventive Treatment in Infants (IPTi), Technical Expert Group (TEG) on Preventive Chemotherapy, April 23-24, 2009 Geneva. (Annex 1). 4 Aponte, John. J, Schellenberg, David, et al. Intermittent Preventive Treatment for Malaria Control in African Infants: Pooled analysis of safety and efficacy in six randomized controlled trials. Accepted for publication by Lancet 2009, forthcoming. 3

6 Both the IOM and WHO's TEG have recommended that IPTi-SP be considered for implementation through EPI as an additional malaria control intervention in countries in Africa south of the Sahara with moderate to high malaria transmission, and where drug resistance to SP is low. As part of the IPTi Consortium research to support this policy recommendation, since 2002 WHO has been coordinating a project designed to investigate the impact of IPTi with SP and a range of other antimalarial drugs 5 on infant serological responses to EPI vaccines. Confirmation that IPTi does not have any adverse interaction with the serological responses to vaccination was critical to ensuring that EPI could safely be used as the delivery mechanism for IPTi. Early results from the first IPTi-SP trial in Ifakara, Tanzania 6 raised concerns that seropositivity to measles and pertussis vaccines was lower in infants receiving IPTi compared to placebo. To oversee and guide this area of work, WHO established a 5-member, independent Ad Hoc Advisory Committee (see Annex 2 for Terms of Reference and Membership). The Advisory Committee assisted with the design of the project, selection of the subcontractors, and review of all the data arising from five EPI serology studies. The Advisory Committee guided WHO accordingly, and issued an Interim Report in July The Interim Report, based on data at the time from the Navrongo (Ghana), Manhica (Mozambique) and Bungoma (Kenya) studies, concluded that IPTi-SP did not have an adverse impact on serological responses to vaccination against measles, diphtheria, tetanus, pertussis, polio serotypes 1 and 3, Haemophilus influenzae type b and hepatitis B. This Final Report of the Advisory Committee describes the methodology of the IPTi serology studies, summarizes the results of the pooled statistical analysis, and presents the Advisory Committee's final conclusions based on all the data available. 5 See Table 1 for list of drugs studied and their abbreviations in the footnote of the table. 6 Schellenberg, D. et al. (2001). Intermittent treatment for malaria and anaemia control at time of routine vaccinations in Tanzanian infants: a randomized, placebo-controlled trial. Lancet, 357: The Interim Report summarized the results as of 2006 from Navrongo, Manhica and Bungoma, a pooled analysis of Navrongo and Manhica data, and the conclusions of the Advisory Committee. This Final Report includes all data from all trials and as such supercedes the analysis of the Interim Report. 4

7 EPI Serology Study Design Study sites and study population The EPI serology work was designed as "nested sub-studies" within five randomized controlled studies of the IPTi Consortium and one other on-going trial to assess the protective efficacy of IPTi against episodes of clinical malaria and anemia. The EPI serology assessments were included in the IPTi trials conducted in Ghana (Navrongo)8, Kenya (Bungoma9 and Kisumu), Mozambique (Manhica), and Tanzania (Kilimanjaro). Infants eligible for the serology studies were selected 10 from the larger study population at each site. Infants were randomized into: i) placebo or SP in Navrongo, Manhica and Bungoma, ii) placebo, SP-ART, AQ-ART and LapDap in Kisumu and iii) placebo, SP, MQ and LapDap in Kilimanjaro. All sites performed serological testing for measles vaccine. Only Navrongo undertook serological assessment for yellow fever vaccine. Bungoma, Manhica, and Kisumu performed serological testing on all other antigens. Table 1 summarizes the specific details of each of the serological studies including the drugs used, the IPTi dosing schedule, sample size, timing of blood samples, and EPI antigens assessed. In each of the serological studies, IPTi (using a number of antimalarial drug options) or placebo was administered on three occasions 11 during the first year of life at the time of routine EPI vaccination (doses 2 and 3 of diphtheria, tetanus and pertussis (DTP); and measles vaccination). Detailed information was obtained on the vaccination status (including BCG and birth doses of polio) of each infant recruited to the serology studies. 8 Note: The Navrongo study was nearly completed when the protocol for the EPI serology studies was designed. Since Navrongo provided a unique opportunity to obtain information on serological responses to yellow fever vaccination, a decision was made to retrospectively measure the serological responses on a selection of blood samples already taken. 9 This trial was supported by WHO/TDR and not part of the IPTi Consortium. 10 Notes on sample selection: Navrongo: As this trial was already completed, the EPI serology study used stored samples. Those children with pre and post vaccination paired samples of sufficient volume were selected. Bungoma: Samples from all children in the study were used unless there was insufficient volume or sample was not taken. Kisumu: This study was designed with a sample size of 379 per arm at the beginning. When the serology study was proposed the trial was unable to increase the sample size to 500 per arm for budgetary and operational reasons. The serology study used samples collected from all children unless the sample was of insufficient volume or missed. Manhica: The trial was underway before the EPI serology study was developed. Serological responses to EPI vaccines were assessed in a subsample of study infants consecutively selected as they came to the clinic until the estimated sample size of 300 children per arm (for the main trial) was completed. 11 The Navrongo trial included a fourth dose of IPTi at 12 months of age which was not linked with EPI. 5

8 All studies were approved by the relevant local and international (including WHO) Ethical Review Boards. Written informed consent was obtained from the parents or guardians of all participating infants. 12 Study design The serological assessments were designed as non-inferiority studies, with the objective to demonstrate that IPTi does not reduce serological responses to EPI vaccines by more than a proportion deemed to be clinically important. Primary endpoint: The primary endpoint was serological responses to measles vaccine. If the proportion of infants attaining protective levels (> 120IU/L) of measles antibody postvaccination was reduced by 5% in the IPTi group compared to the placebo group, it would not be considered that IPTi had an adverse impact on serological responses to measles. The sample size of per arm was calculated with the aim of rejecting, with 80% power and a one-sided significance level of 5%, the null hypothesis that the difference between the groups was > 5%. Secondary endpoints: The secondary endpoints were serological responses to all other EPI vaccines. If the proportion of infants attaining protective levels (see Table 3) of antibody post-vaccination was reduced by 10% in the IPTi group compared to the placebo group, it would not be considered that IPTi had an adverse impact on serological responses to EPI vaccines. The sample size of 250 per arm was calculated with the aim of rejecting, with 80% power and a one-sided significance level of 5%, the null hypothesis that the difference between the groups was > 10%. Plotting of reverse cumulative distribution functions (RCDF) of the proportion of infants attaining different antibody titres was an important graphic tool for visualizing the full distribution of data and comparison of results. Close agreement of RCDFs from infants given IPTi with those given placebo supports the conclusion that antibody responses are similar in the two populations over the full range of antibody concentrations. Postvaccination geometric mean concentrations in the treatment and placebo groups were compared. 12 See Annex 4 page 27 for details about Bungoma trial having to re-do written consent retrospectively. 13 At the outset the Kisumu sample size was planned to be less (379) because they knew it was not possible to obtain a target of 500 before the conclusion of the trial. 6

9 Table 1. IPTi studies that include assessment of serological responses to EPI vaccines Study site Study design Drug(s) for IPTi Age at IPTi drug dosing Sample size EPI immunization schedule Timing of blood samples Serological information available from each study Navrongo, Ghana RCT SP Placebo 10 weeks (DTP2) 14 weeks (DTP3) 9 months (measles) 12 months 500 SP 500 placebo BCG: birth DTP, hepb, Hib: 6, 10, 14 weeks Polio: birth, 6, 10, 14 weeks Measles, yellow fever: 9 months 9 months 12 months Measles, yellow fever Bungoma, Kenya RCT SP Placebo 10 weeks (DTP2) 14 weeks (DTP3) 9 months (measles) 500 SP 500 placebo BCG: birth DTP, hepb, Hib: 6, 10, 14 weeks Polio: birth, 6, 10, 14 weeks Measles: 9 months 6 weeks 18 weeks 9 months 10 months DTP, polio, hepatitis B, Hib, measles 7

10 Manhica, Mozambique RCT SP Placebo 12 weeks (DTP2) 16 weeks (DTP3) 9 months (measles) 500 SP 500 placebo BCG: birth DTP, hepb: 8, 12, 16 weeks Polio: birth, 8, 12, 16 weeks Measles: 9 months 20 weeks 9 months 12 months DTP, polio, hepatitis B measles Kisumu, Kenya RCT SP-ART AQ-ART 10 weeks (DTP2) 14 weeks (DTP3) 379 x 3 intervention 379 placebo As for Bungoma 6 weeks 18 weeks DTP, polio, hepatitis B, Hib, measles LapDap 9 months (measles) 9 months Placebo 12 months SP BCG: birth Kilimanjaro, Tanzania RCT MQ LapDap 8 weeks (DTP2) 12 weeks (DTP3) 500 x 3 intervention 500 placebo DTP, hepb: 4, 8, 12 weeks Polio: birth, 4, 8, 12 weeks 9 months 10 months Measles Placebo 9 months (measles) Measles: 9 months Legend: RCT: randomised controlled trial; SP: sulfadoxine-pyrimethamine; SP-ART: sulfadoxine-pyrimethamine plus artesunate; MQ: mefloquine; LapDap: chlorproguanildapsone; AQ-ART: amodiaquine plus artesunate; DTP: diphtheria, tetanus, pertussis; hepb: hepatitis B; Hib: Haemophilus influenzae type b 8

11 It was planned that each study site would attain the requisite sample size for the primary and secondary endpoints, and that data from all sites be pooled to provide overall summary measures of even greater precision. Blood sampling: A minimum of 0.5mls of whole blood was taken by finger prick or venous sampling at intervals indicated in Table 2. Samples were centrifuged, and sera frozen at minus 20 degrees Centigrade. Serological assays: Following a tendering process, the WHO Advisory Committee assessed quotations from four internationally recognized public health reference laboratories for carrying out the serological assays for these studies. The Health Protection Agency (HPA), UK, was chosen on the basis of its competitive pricing, expertise in functional (plaque reduction neutralization) assays, and capacity to deal with large numbers of samples within the requisite timeframe. The laboratory was blind to the allocation status (IPTi or placebo) of all samples. Geometric Mean Titres (GMTs) were measured by plaque reduction neutralization (measles and yellow fever), microneutralization (polio serotypes 1 and 3) and enzyme linked immunosorbent assay (ELISA - all other EPI antigens). All assays were run in duplicate, using standardized reagents or validated test kits. For measles and yellow fever, pre-vaccination blood samples were assayed to check whether subjects had been exposed to these diseases prior to vaccination. For all other EPI antigens, measurement was confined to the post-vaccination blood sample 14. In instances where there was insufficient blood to carry out all of the assessments, assays were carried out in the following order: week sample i. Haemophilus influenzae type b ii. Diphtheria iii. Polio serotype 3 iv. Hepatitis B v. Pertussis toxin vi. Tetanus vii. Polio serotype 1 viii. Pertussis filamentous haemagglutinin month sample i. Measles ii. Yellow fever 14 Pre-vaccination samples were collected, but as these were likely to contain maternal antibodies they were stored in the event that that the post-vaccination results were equivocal and analysis of the pre-vaccination samples was needed. 9

12 Table 2. Serological assays Vaccine Pre or postvaccination sample Timing of blood samples Type of test Protective level Measles Pre 9 months Plaque reduction neutralisation (PRN) 1-2 dilutions To check for presence of measles antibodies pre-vaccination Post 10 months or 12 months PRN 6 dilutions (GMT) 120 IU/l Yellow fever Pre 9 months PRN 1-2 dilutions To check for presence of YF antibodies prevaccination Post 10 months or 12 months PRN 6 dilutions (GMT) 1:5 (PRN titre) Diphtheria Pre At time of DTP1 Store in freezer* Post One month post DTP3 Quantitative ELISA (GMT) Tetanus Pre At time of DTP1 Store in freezer* Post One month post DTP3 Quantitative ELISA (GMT) 0.1 IU/ml 0.1 IU/ml Pertussis (PT and FHA only) Polio (serotypes 1 and 3 only) Pre At time of DTP1 Store in freezer* Post One month post DTP3 Quantitative ELISA (GMT) Pre At time of DTP1 Store in freezer* Post One month post DTP3 PRN 6 dilutions (GMT) Protective levels not defined 1:8 (PRN titre) Haemophilus influenzae type b Pre At time of DTP1 Store in freezer* Post One month post DTP3 Quantitative ELISA (GMT) Hepatitis B Pre At time of DTP1 Store in freezer* Post One month post DTP3 Quantitative ELISA (GMT) 0.15 µg/ml (1.0 µg/ml will be used as a secondary descriptive) 10 IU/l * Pre-vaccination GMTs will only be obtained if the post-vaccination results are equivocal; PRN: Plaque reduction neutralisation; GMT: Geometric mean titre; PT: Pertussis toxin; FHA: Filamentous haemagglutinin 10

13 This ranking was based on the relative immunogenicity of each antigen, with those of lower immunogenicity being assigned a higher rank, so as to increase the likelihood of detection of any potential interference. Haemophilus influenzae type b (Hib) was assigned top rank since regulatory authorities are particularly concerned about serological responses to Hib vaccine. Interference with responses to this vaccine have been demonstrated in different settings, particularly in relation to fever 15. Any infants failing to attain protective levels of antibody post-vaccination were offered revaccination. Statistical analysis: A competitive tender was won by the London School of Tropical Medicine and Hygiene (LSHTM) Tropical Epidemiology Group. LSHTM was sent the results of all the laboratory serology, relevant clinical data from each study site plus the randomization codes. Post-vaccination geometric mean antibody concentrations (GMCs) and the proportion of infants attaining the protective level for each EPI antigen were compared in the IPTi and placebo groups. Reverse cumulative curves were plotted for each EPI antigen. The results for each of the trials are provided in individual statistical reports for each study. These final reports are on file with WHO and have been shared with the study teams for inclusion in the publication of their trial results. A final pooled analysis of all the study results was prepared in early Data from the trial in Bungoma were excluded due to previously established concerns about data quality. The final pooled analysis (March 27, 2009) that was accepted by the Advisory Committee is provided in Annex 3. WHO Advisory Committee The Committee met several times over the course of this project, particularly in 2003 to advise on the design of the protocol and selection of the laboratory. In later years, the Committee has used teleconferences to conduct its work (See Annex 2 for complete listing of dates of meetings and teleconferences). The Advisory Committee reviewed and discussed each of the statistical reports of the five studies. Often the Committee requested revisions or additional statistical analyses in order to facilitate their interpretation and conclusions. A complete record of the minutes of these teleconference is on file with WHO. Brief summaries of the results for each of the five studies can be found in Annex 4. Prior to issuing their final conclusions, the Advisory Committee requested an independent audit of the laboratory work carried out by HPA. A post study audit was performed on the laboratory records in Jan and March Several discrepancies were noted and HPA was requested to take corrective action on these points. Subsequently, a second follow-up audit was conducted in June 2009 with the objective to further investigate the discrepancies noted in the first audit and to evaluate the corrective actions taken. Overall, the HPA labs were 15 Usen S, Milligan P, Ethevenaux C, Greenwood B, Mulholland K. Effect of fever on the serum antibody response of Gambian children to Haemophilus influenzae type b conjugate vaccine. Pediatr Infect Dis J 2000;19(5):

14 found to be operating in a manner that meets relevant GLP criteria for this GCP study. The audit satisfied the Advisory Committee that the analyses were carried out appropriately. This Final Report presents the final conclusions of the Advisory Committee and is primarily based on the final pooled analysis of the results, and supported by selected analyses of the individual trials where no pooled analyses was planned (i.e. yellow fever and Hib vaccines). Results Pooled Analysis The pooled analysis (see Annex 3 for full report) was conducted on data from four of the study sites Navrongo, Manhica, Kisumu, and Kilimanjaro. Data from the fifth trial, conducted in Bungoma, was excluded due to concerns about data quality. Data from different trials and treatment groups were pooled only after establishing, by means of appropriate interaction tests, that treatment effects were not heterogeneous. If there was evidence of heterogeneity, pooling was not done and trial-specific treatment effects were reported. Both Intention-to-Treat (ITT) and According-to-Protocol (ATP) analyses were undertaken. All children with matched pre and post measles vaccination samples were included in the ITT analysis. Children with incomplete drug dosing were excluded from the ATP analysis. Hence, only Navrongo children with all four drug doses taken, and Manhica, Kisumu and Kilmanjaro children with all three drug doses taken were considered for the ATP. Separate analyses for measles were undertaken excluding children with: i) detectable and ii) protective pre-vaccination levels. For all other antigens all children with a post vaccination sample were included in the ITT analysis, whereas children with incomplete drug dosing were excluded from the ATP analysis. For all antigens analyses were conducted on post-vaccination antibody concentrations, using: i) the continuous concentration variable, summarized by its geometric mean (GMC), and ii) the binary protected/unprotected variable, based on whether antibody concentrations were above or below the pre-defined threshold of protection for each antigen, where appropriate. Pooled SP vs Placebo comparison for measles: Pooled data from Navrongo, Manhica and Kilmanjaro provided matched pre and post vaccination measurements for 2,015 children (997 in placebo and 1,018 in SP groups). Comparison of the geometric mean concentrations (GMC) for measles antibodies in the two treatment groups before and after vaccination, as well as the median post-vaccination concentration, found no evidence of a difference in the GMC between SP and placebo groups in any of the sub-population investigations 16. (Annex 3) The formal test of non-inferiority of the null hypothesis (that the difference in percentage unprotected between groups was > 5%) gave strong evidence that SP is not inferior to placebo in ITT and ATP analyses, excluding children with detectable/with protective prevaccination concentration levels. For example, for the ATP analysis (excluding those 16 With and without detectable concentration pre-vaccination; with and without protective measles antibody level pre-vaccination; all children. 12

15 children with detectable concentration at pre-vaccination) the actual difference (SP minus placebo) between the groups was -0.15% with a 95% CI (-2.33, 2.04) (p<0.0001). Finally, the reverse cumulative distribution function for measles antibody concentrations for the ITT cohort (excluding children with detectable antibody levels at pre-vaccination), shows that the curves for placebo and SP are nearly identical (Figure 1). FIGURE 1: Pooled Analysis (Navrongo, Manhica, Kilimanjaro) Reverse cumulative distribution function for measles antibody concentrations for the ITT cohort excluding children with detectable antibody levels at pre-vaccination Reverse empirical cumulative distribution function (vertical line indicates assumed protective threshold) log(10) measles antibody concentration post-vaccination Placebo SP Pooled drugs (SP, LapDap, SP-ART, AQ-ART, MQ) vs placebo comparison for measles For the Kilimanjaro (SP+MQ+LapDap) measles comparison data were pooled for both analyses, resulting in 397 children in placebo and 1,141 in the single combined treatment group. No evidence was found to support a difference in the GMC between the combined treatment group and placebo. The formal test of non-inferiority, gave strong evidence to reject the null hypothesis that the proportion unprotected was at least 5% higher than in the placebo group. The actual difference (combined treatment minus placebo) found in the ATP analysis, excluding those with detectable concentration at pre-vaccination, was 0.21% with a 95% CI (-2.67, 3.08) (p=0.0001). For the Kisumu (SP-ART+AQ-ART+LapDap) measles comparison, the treatment groups were pooled for the analysis of GMCs, resulting in 284 children in placebo and 838 in the combined treatment group. No evidence was found to support a difference in the measles GMC between the combined treatment group and placebo. For the analysis of proportions 13

16 unprotected, however, the three treatment groups were heterogeneous (owing to the much lower proportion unprotected in the LapDap group), so the three treatment groups were not pooled (the separate analyses for the three treatment groups all rejected the null hypothesis that the proportion unprotected is at least 5% higher than for placebo; see Annex 4). A secondary pooled analysis of the Kisumu measles data excluding the LapDap group was undertaken, to compare SP-ART + AQ-ART versus placebo. This strongly rejected the null hypothesis that treatment is inferior to placebo. A final pooled analysis for measles combining all treatment groups (except the Kisumu LapDap group, for the reasons given above) versus placebo across the four trials (Navrongo, Manhica, Kisumu and Kilimanjaro) resulted in 1,281 children in placebo and 2,363 in the combined group. No evidence was found of a difference in the measles GMC between the all combined treatment group and placebo. The formal test of non-inferiority (difference in unprotected proportion of children is >5%), provided strong evidence that the combined group was not inferior to placebo. For the ATP analysis, excluding those children with detectable concentration at pre-vaccination, the actual difference (combined treatments minus placebo) was 0.53% with a 95% CI (-1.18, 2.22) (p=0.0001). The reverse cumulative distribution function of measles antibody concentrations for the ITT cohort, excluding children with detectable pre-vaccination antibody levels,provides additional support that there is little difference between the intervention group (all treatments except LapDap Kisumu) and placebo (Figure 2). FIGURE 2: Pooled Analysis All Sites (Navrongo, Manhica, Kisumu and Kilimanjaro), All Treatments Combined (except Kisumu LapDap) Reverse cumulative distribution function for measles for the ITT cohort excluding children with detectable antibody levels at pre-vaccination Reverse empirical cumulative distribution function (vertical line indicates assumed protective threshold) log(10) measles antibody concentration post-vaccination Placebo Combined 14

17 Pooled Analysis for other antigens Manhica (SP) and Kisumu (SP-ART, AQ-ART, LapDap) were the only sites assessing antigens other than measles and yellow fever (polio types 1 & 3, diphtheria, tetanus, hepatitis B, pertussis FHA & Toxin) 17. Serology data for these antigens was available for a total of 634 for the ITT and 567 for the ATP analysis. Complete data on all antigens was available for 150 children. The comparison of placebo groups in Manhica and Kisumu found that Kisumu had a significantly higher percentage of unprotected children for polio types 1 & 3 and diphtheria when compared to Manhica. For tetanus and HepB there was no evidence of a difference. These results were the same in ITT, ATP, and other sub-population analyses. For diphtheria, pertussis toxin and FHA, tetanus and HepB Manhica children had on average a higher geometric mean concentration after vaccination when compared to Kisumu children. In this context, the Advisory Committee observed that there appeared to be marked variation across the study sites, and that some study sites seemed to be "high responders" and others (particularly Kisumu) were "low responders". The Committee speculated that there could be differences in the health status and weight of children between the sites, or that the vaccines were not in their best condition in the low responder sites (perhaps owing to cold chain issues) 18. As data on antigens other than measles and yellow fever is available from only two sites, Manhica and Kisumu, the individual analyses for these studies is summarized. In Manhica, the proportion of infants achieving protective titres post-vaccination was similar in the SP and placebo groups for diphtheria, tetanus, polio type 1 & 3, and HepB (Hib vaccine was not at the time included in the Mozambique vaccination schedule). The test of non-inferiority (with a 10% threshold) was significant for each antigen, providing evidence that SP treatment had no adverse impact on serological responses to EPI vaccines. There are no know serological correlates of protection for pertussis. For all antigens, post-vaccination GMCs and the reverse cumulative curves were similar in both the SP and placebo groups. In Kisumu (Placebo, SP-ART, AQ-ART, LapDap): the post-vaccination GMC's for diphtheria, tetanus, Hib and pertussis toxin were similar in the four treatment groups in both ITT & ATP analyses. For HepB, both in the ITT & ATP analyses, there was evidence to reject the null hypothesis of equal GMC in all four groups. The hypothesis that the proportions unprotected in IPTi-treatment groups were at least 10% higher than in the placebo group was rejected, in both ITT & ATP analyses, for diphtheria, tetanus, Hib, HepB and polio type Hib was assessed only in the Kisumu trial. Other antigens were also assessed in Bungoma, but these data were excluded from pooled analyses as stated previously. 18 Differences were also observed in the proportions of children protected against measles within the placebo groups in Kilimanjaro, Kisumu, Manhica and Navrongo. The proportion unprotected was highest in Manhica and lowest in Navrongo. 15

18 For polio type 3 the null hypothesis of inferiority (with respect to proportions unprotected) of the LapDap and SP-ART groups compared to placebo for both ITT & ATP analyses could not be rejected. The reverse cumulative distribution functions were similar for all antigens, except HepB. Given the difficulties of interpreting the HepB results an additional post-hoc analysis which used 100 IU/L (instead of 10 IU/L) 19 for inferiority was conducted. The HepB analysis was compromised by the small sample size (n=222 which meant only around 50 in each arm). There was no evidence of any problem with SP-ART, some evidence that AQ-ART was better than placebo, and some very weak evidence that LapDap might be worse than placebo, the proportions under 100 IU/L being quite high. However, no definite conclusions could be drawn owing to the lack of power. In Kisumu, a comparison of GMCs in the combined treatment group (SP-ART+AQ- ART+LapDap) versus placebo group was conducted for diphtheria, pertussis FHA & toxin, and tetanus, but not for hepatitis B. The GMCs in the combined treatment group were similar to those for the placebo group. The reverse cumulative distribution functions were also similar. For proportions unprotected,, it was possible to pool the treatment groups for polio type 1 & 3 and diphtheria. The null hypothesis that the proportion unprotected is more than 10% greater in the treatment group than in the placebo group was rejected for all antigens, in ITP and ATP analyses. Results Yellow Fever (Navrongo) Only one study site, Navrongo (Ghana) was able to provide data on serological responses to yellow fever vaccination in infants randomized to receive SP or placebo. Unfortunately, many of the samples selected had insufficient volumes for testing, and additionally there was a miscommunication with the lab about the dilution, which further reduced the available samples to only 136, when the protocol specified 250. Owing to many complicating factors (small sample size; lack of clarity about the correlation between protective titres and IU/ml concentration values; suspected interference with crossreactive antibodies for other flaviviruses) the analysis of the yellow fever data was limited to differences between the groups post-vaccination.. Focusing only on the post-immunization samples, a comparison of the two groups was undertaken on the entire cohort (not excluding any of the children with inconclusive replicate samples, or detectable or protective antibody levels prior to immunization). The proportions protected were similar in both groups. The formal test of non-inferiority rejected the null hypothesis that the proportion unprotected in the treatment groups was more than 10% greater than in the placebo group; the actual difference (SP minus placebo) was 4.21%. Three methods adjusting and not for clustering produced similar results. The IU/L is the level at which long term immunity is conferred; 10 IU/L is the level for seroconversion. 16

19 geometric mean concentrations were found to be very similar in both groups. Finally, the reverse cumulative distribution functions suggest that there is no reduction in antibody concentrations in the SP group. 17

20 Conclusions of the WHO Advisory Committee The Advisory Committee was established 6 years ago to review data generated by existing studies. While it cannot be held responsible for the conduct of the field trials or laboratory work, the Committee concludes the following: 1. Serological data from studies in Navrongo, Manhica, and Kilmanjaro provide strong evidence that IPTi with SP, does not have an adverse impact on serological responses to measles vaccine; 2. Though very limited, the available serological data from Navrongo provides evidence that SP does not have a negative impact on antibodies following vaccination against yellow fever; 3. Serological data from studies in Kisumu and Kilimanjaro provide strong evidence that IPTi with SP-ART, AQ-ART, MQ, or LapDap do not have an adverse impact on serological responses to measles vaccination; 4. Serological data from Manhica and Kisumu provide strong evidence that IPTi with SP 20, SP-ART, AQ-ART, or LapDap do not have an adverse impact on serological responses to DTP, polio, Hib, and HepB vaccines. 5. The pooled analyses provide further evidence that IPTi treatments do not impair serological responses to EPI antigens. Overall conclusion: Studies have demonstrated that there is no adverse impact on the serological responses to DTP, polio, Hib, HepB, and measles vaccines when the IPTi drugs SP, SP-ART, AQ- ART, and LapDap are administered to infants at the time of routine vaccination. Concomitant administration of IPTi-SP at the time of yellow fever vaccination, and IPTi- MQ at the time of measles vaccination, have also been shown to have no negative effect on the serological responses. 20 Data from Bungoma, although weaker, also suggest that there is no negative interaction between SP and EPI antigens. 18

21 Annex 1: Report of the Technical Consultation on Intermittent Preventive Treatment in Infants (IPTi), Technical Expert Group (TEG) on Preventive Chemotherapy, April 23-24, 2009 Geneva.. 19

22 Technical Expert Group meeting on Preventive chemotherapy WHO HEADQUARTERS, GENEVA, APRIL 2009 Report of the Technical Consultation on Intermittent Preventive Treatment in Infants (IPTi)

23 Technical Expert Group meeting on Preventive chemotherapy WHO HEADQUARTERS, GENEVA, april 2009 Report of the technical consultation on Intermittent Preventive Treatment in Infants (IPTi)

24 Contents 1. Background Conclusions Recommendations Other recommendations References List of participants...10 WHO Library Cataloguing-in-Publication Data Technical expert group meeting on preventive chemotherapy : report of the technical consultation on intermittent preventive treatment in infants (IPTi) Geneva, April Malaria, Falciparum - prevention and control. 2.Malaria, Falciparum - drug therapy. 3.Infants. 4.Drug administration schedule. 5.Pyrimethamine - therapeutic use. 6.Sulfadoxine - therapeutic use. 7.Treatment outcomes. 8.Meta-analysis 9.Guidelines. I.World Health Organization. ISBN (NLM classification: WC 765) World Health Organization All rights reserved. The designations employed and the presentation of the material in this publication do not imply the expression of any opinion whatsoever on the part of the World Health Organization concerning the legal status of any country, territory, city or area or of its authorities, or concerning the delimitation of its frontiers or boundaries. Dotted lines on maps represent approximate border lines for which there may not yet be full agreement. The mention of specific companies or of certain manufacturers products does not imply that they are endorsed or recommended by the World Health Organization in preference to others of a similar nature that are not mentioned. Errors and omissions excepted, the names of proprietary products are distinguished by initial capital letters. All reasonable precautions have been taken by the World Health Organization to verify the information contained in this publication. However, the published material is being distributed without warranty of any kind, either expressed or implied. The responsibility for the interpretation and use of the material lies with the reader. In no event shall the World Health Organization be liable for damages arising from its use.

25 WHO Technical Expert Group on Preventive Chemotherapy 1 1. Background Intermittent preventive treatment in infancy (IPTi) is defined as: the administration of a full course of an effective antimalarial treatment at specified time points to infants at risk of malaria, regardless of whether or not they are parasitaemic, with the objective of reducing the infant malaria burden. In October 2006 and October 2007, WHO convened meetings of the Technical Expert Group (TEG) on Intermittent Preventive Treatment in Infants to review the available evidence on the safety, efficacy and other relevant aspects of IPTi with sulfadoxine-pyrimethamine (SP-IPTi) delivered through the Expanded Programme for Immunization (EPI). At the time, six randomised, placebo-controlled clinical trials with SP-IPTi were being, or had been, conducted in areas of Africa, south of the Sahara with relatively high malaria endemicity (see Table). TEG 2006 concluded that SP-IPTi held promise as a potential malaria control intervention, noting that three of the studies were yet ongoing or unpublished. 1 At the TEG 2007, at which time the six studies had been completed, the committee concluded that, though IPTi remains a potential intervention for malaria control, the use of SP-IPTi cannot be recommended as a strategy for general deployment based on the assessment of the risks and benefits, and advised a future review of further evidence when available. 2 In the current expert review of the evidence on SP-IPTi, TEG 2009 reviewed the evidence available on SP-IPTi including additional data that were generated since the TEG-2007 meeting, with a view to making a definitive policy recommendation on this intervention for malaria control. The new information reviewed was the following: 1. An in-depth analysis conducted by the IPTi Consortium 3, of the severe skin reactions associated with SP-IPTi reported previously. 4,5 2. Two additional randomized placebo controlled trials on the safety and efficacy of IPTi, which have been submitted for publication. 6,7 3. The experience of implementation studies conducted by UNICEF in selected districts of six countries in Africa, south of the Sahara 8 and another by the IPTi Consortium 9 11 with respect to the feasibility of implementation, and its safety, monitored through active and passive observations on adverse reactions.

26 2 WHO Geneva, April 2009 Table Summary of site-specific information on the six published SP-IPTi trials considered for the pooled analysis* Study site Study period Transmission pattern Iron supplementation # Infants studied SP/ placebo** Ifakara, UR Tanzania i Navrongo, Ghana ii Perennial Yes 350/ Seasonal Yes 1183/1203 Manhica, Perennial/ Mozambique iii seasonal peaks None 748/755 Kumasi, Ghana iv Perennial None 535/535 Tamale, Perennial/ Ghana v seasonal peaks Lambarene, Perennial/ Gabon vi seasonal peaks None 600/600 None 504/507 i ) Schellenberg D et al. (2001). Intermittent treatment for malaria and anaemia control at time of routine vaccinations in Tanzanian infants: a randomised, placebocontrolled trial. Lancet, 357: ii) Chandramohan D et al. (2005). Cluster randomized trial of intermittent preventive treatment for malaria in infants in area of high, seasonal transmission in Ghana. Br Med J, 331: iii) Macete E et al. (2006). Intermittent preventive treatment for malaria control administered at the time of routine vaccinations in Mozambican infants: a randomized, placebo controlled trial. J Inf Dis, 194: iv) Kobbe R. et al. (2007). A randomized controlled trial of extended intermittent preventive antimalarial treatment in infants. Clin Infect Dis, 45: v) Mockenhaupt FP et al. (2007). Intermittent Preventive Treatment in Infants as a Means of Malaria Control: a Randomized, Double-Blind, Placebo-Controlled Trial in Northern Ghana. Antimicrob Agents Chemother, 51: vi) Grobusch M. et al. (2007). Intermittent preventive treatment in infants against malaria in Gabon a randomised, double-blind, placebo-controlled trial. J Inf Dis, 196: * The pooled analysis excludes the most recent study6 which was accepted for publication after the meeting, although its data were made available to the meeting. ** Who received at least the first dose of SP-IPTi

27 WHO Technical Expert Group on Preventive Chemotherapy 3 2. Conclusions The TEG 2009 concluded that: 1. The previous safety concerns about SP-IPTi, specifically with respect to the reported severe skin reactions were mitigated by the evidence from the larger observational studies and retrospective in-depth examination by the Consortium of the severe skin reactions reported in previous studies. 2. The benefits of SP-IPTi in areas where SP remains effective against Plasmodium falciparum malaria parasites, were upheld as providing a 30% (95% CI: 19.8% 39.4%) overall protection against clinical malaria episodes and a variable reduction (overall 21.3%) (95% CI: 8.3% 32.5%) in anaemia (< 8 g/dl) in a pooled analysis of data from 6 published studies (see Table). 12 The reduction in all cause hospital admissions by 23% (95% CI: 10.0% 34.0%), was noted as a potential benefit. The admissions, however, were not all due to severe malaria, and this therefore cannot be equated to a similar reduction in the incidence of severe malaria. The pooled analysis excludes the most recent study 6 which was accepted for publication after the meeting, although its data were made available to the meeting. The protective efficacy of SP-IPTi against clinical malaria episodes in this study was 6.7% (95% CI: ). 3. Where effective, SP-IPTi offers a personal protection against clinical malaria for a period of approximately 35 days following the administration of each dose. There is no evidence for an individual cumulative protective effect beyond this period until the next dose. The mechanism of action appears to be predominantly chemoprophylaxis related to the half-life of the medicine and the susceptibility of the prevalent malaria parasites. 4. The protective efficacy of SP-IPTi is dependent upon the efficacy of SP, to which there is increasing parasite resistance in Africa and worldwide, but the threshold of parasite resistance to SP at which IPTi ceases to be effective is still not known. SP-IPTi was reported to provide benefit when the in vivo therapeutic failure rate of SP at day 14 was 31% (measured

28 4 WHO Geneva, April 2009 in children with symptomatic malaria) 13 and the population prevalence of Pfdhps + Pfdhfr quintuple mutants (molecular markers of parasite resistance to SP) was 50% 14 but there was no benefit when the in vivo SP therapeutic failure rate was 82% at day 28, and the prevalence of the quintuple mutants was 90% Uncertainties remain on the potential impact, or lack thereof, of SP-IPTi on the incidence of severe malaria or malaria mortality. 6. Uncertainties also remain on the impact of SP-IPTi at low levels of malaria transmission (either natural or resulting from effective control interventions). 7. A rebound effect by way of greater susceptibility to malaria following the termination of SP-IPTi was not evident in the pooled analysis. However, this warrants further observation in view of the fact that three of the studies reported an increase in either malaria infections associated high density parasitaemia 15 ; anaemia (< 7.5 g/dl) 4 ; or severe malaria and severe malarial anaemia (Hb < 5g/dl) 5 during the post-intervention period in children who had received SP compared to the placebo group. 8. SP-IPTi was deemed a safe addition to EPI because there was no evidence of an adverse effect of SP-IPTi on infants serological response to EPI vaccines against DTP, Polio, Hepatitis B, Hib, yellow fever and measles. 16 Limited implementation studies suggest that SP-IPTi incurs only marginally additional costs to EPI, and that it has a favorable effect on EPI coverage. The panel comprised of 15 independent Experts. The Consultation was attended by observers from UNICEF, the Bill and Melinda Gates foundation, and the IPTi Consortium (Appendix 1, List of participants).