Primary and booster vaccination in Latin American children with a DTPw-HBV/Hib combination: a randomized controlled trial

This was a randomized, partially-blind, multicenter study in three countries in Latin America (Argentina, Chile and Nicaragua) between August 2004 and September 2005. The study was approved by the appropriate local ethics committees and conducted in accordance with the Declaration of Helsinki and Good Clinical Practice guidelines. Healthy male and female infants born after a normal gestation period (between 36-42 weeks) were enrolled for first vaccination at 6-10 weeks of age. In Argentina, mothers of prospective participants were screened prenatally for the presence of hepatitis B virus surface antigen (HBsAg). Infants born to anti-HBsAg seropositive mothers were ineligible for study participation, but were offered HBV vaccination at birth. Other exclusion criteria included: previous diphtheria, tetanus, pertussis, hepatitis B, Hib or polio vaccination or disease; Bacille Calmette-Guérin vaccine given later than the first 2 weeks of life; major congenital defects or serious chronic illness; a history of allergic disease or reactions likely to be exacerbated by any component of the vaccine; a history of neurological disease, including previous seizures; any immune deficiency and prior or planned use of any blood products, immunoglobulins or immunosuppressive therapy; and acute illness at the time of enrolment. Written informed consent was obtained from the parent or guardian of each infant prior to study entry.

All vaccines were manufactured by GSK Biologicals. The diphtheria and tetanus antigens were sourced form GSK Biologicals Korlatolt Felelossegu Tarsasag (Hungary) and Pw was obtained from Commonwealth Serum Laboratory Ltd. (Australia) for the new DTPw-HBV/Hib vaccine. Both vaccines contained ≥30 international units (IU) diphtheria toxoid, ≥60 IU tetanus toxoid, ≥4 IU killed Bordetella pertussis, and 10 μg HBsAg. The new DTPw-HBV/Hib vaccine contained 2.5 μg of Hib capsular PRP polysaccharide conjugated to 5-10 μg tetanus toxoid. Tritanrix™-HBV/Hib contained 10 μg of Hib capsular PRP polysaccharide conjugated to 20-40 μg tetanus toxoid. Both vaccines were administered as a 0.5 mL intramuscular injection into the anterolateral region of the left thigh. All subjects received OPV concomitantly with each primary dose.

Parents/guardians used diary cards to record the occurrence of solicited local (pain, redness and swelling at the site of injection) and general (drowsiness, fever [defined as rectal temperature ≥38°C], irritability and loss of appetite) symptoms during a 4-day follow-up period after each vaccination. Intensity of symptoms was graded on a scale of 0-3. Grade 3 symptoms were defined as follows: for local pain as crying when limb is moved or a spontaneously painful limb; for local redness and swelling as diameter > 20 mm; for fever as rectal temperature > 40.0°C; and for all other symptoms as preventing normal daily activities. All other (unsolicited) adverse events were recorded for a period of 30 days after each vaccination. Serious adverse events (SAEs) were recorded during the entire study period. All solicited local symptoms were considered causally related to vaccination. Causality of solicited general symptoms, unsolicited symptoms and SAEs was assessed by the investigator.

The target sample size was 500 evaluable subjects for immunogenicity (125 subjects per vaccine group). Considering that approximately 15% of enrolled subjects might withdraw or be unevaluable for immunogenicity; the target enrolment size was 600 subjects (150 subjects per vaccine group).

Analysis of immunogenicity was performed on the according-to-protocol (ATP) cohort and the analysis of safety was performed on the total vaccinated cohort.

Seroprotection rates against diphtheria, tetanus, HBsAg and PRP and vaccine response rates to BPT were calculated with 95% CIs. Standardized asymptotic 95% CIs for the differences in seroprotection/vaccine response rates between the pooled DTPw-HBV/Hib groups and the Tritanrix™-HBV/Hib group one month after the third primary vaccine dose were computed. The response to the new DTPw-HBV/Hib vaccine was considered non-inferior to that of the control vaccine if the upper limit of the 95% CI for the absolute difference between groups was lower than 10% for D, T, PRP and HBs. GMCs with 95% CI were calculated by taking the log-transformation of individual concentrations and calculating the anti-log of the mean of these transformed values. Antibody concentrations below the assay cut-off were given an arbitrary value of half the cut-off value for the purpose of GMC calculation. For, D, T, PRP and HBs antigens, 95% CIs for the GMC ratio (control over pooled DTPw-HBV/Hib groups) one month after primary vaccination were computed using an ANOVA model on the logarithm in base 10 (log₁₀) transformation of the concentrations using the vaccine group as covariate, while for pertussis, the 95% CI for the GMC ratio was computed using an ANCOVA model with the vaccine group and the log₁₀ concentrations prior to vaccination as covariates.

The incidence of solicited local and general symptoms (any or Grade 3 intensity) was calculated with exact 95% CI. Differences between groups were compared using two-sided Fisher exact tests. P-values <0.05 were considered as possibly indicative of a statistically significant difference between groups.

The statistical analyses were performed using the Statistical Analysis Systems (SAS) version 8.2 on Windows XP Professional and StatXact-5 procedure on SAS.

Seroprotection/vaccine response rates and antibody GMCs for all vaccine antigens are shown in Table 1. The new DTPw-HBV/Hib vaccine was found to be non-inferior to Tritanrix™-HBV/Hib in terms of seroprotection against diphtheria, tetanus, HBs and PRP antigens following primary vaccination. One month after completion of the 3-dose primary course, 99.0% of subjects in the DTPw-HBV/Hib group and 100% of those in the Tritanrix™-HBV/Hib group had anti-PRP antibody concentrations ≥0.15 μg/mL, with anti-PRP antibody concentrations ≥1.0 μg/mL in 97.3% and 100% of subjects in the two groups, respectively. At least 97.7% of subjects in the DTPw-HBV/Hib group and 93.9% of those in the Tritanrix™-HBV/Hib group had seroprotective levels of antibodies against diphtheria, tetanus and hepatitis B and a vaccine response to the pertussis component at this time. Post-primary vaccination GMCs for anti-diphtheria and anti-HBs antibodies were significantly higher (non-overlapping CI) in the DTPw-HBV/Hib group than in the Tritanrix™-HBV/Hib control group, while anti-PRP GMC was significantly higher in the Tritanrix™-HBV/Hib group than in the DTPw-HBV/Hib group (21.4 and 12.5 μg/mL, respectively; non-overlapping CI).

Prior to booster administration at 18-24 months, anti-PRP antibody concentrations remained ≥0.15 μg/mL in 95.5% of subjects in the DTPw-HBV/Hib and 100% of those in the Tritanrix™-HBV/Hib group. More than 81.7% and 75.8% of subjects in the two groups, respectively, still had seroprotective levels of antibodies to diphtheria (by ELISA), tetanus and hepatitis B at this time, with 32.7% and 36.4% of subjects remaining seropositive for anti-BPT antibodies (data not shown). Testing of seronegative samples by the Vero cell neutralisation assay showed a global seroprotection rate against diphtheria of 90.5% and 82.4% in the two groups, respectively. No differences in pre-booster antibody GMCs were seen between the two groups, with the exception of a significantly higher anti-PRP GMC in the Tritanrix™-HBV/Hib group (2.1 compared with 1.0 μg/mL in the DTPw-HBV/Hib group; non-overlapping CI).

A marked booster response to all vaccine antigens was observed in both groups. One month after booster administration, all subjects had anti-PRP antibody concentrations ≥0.15 μg/mL, with anti-PRP antibody concentrations ≥1.0 μg/mL in all but one subject in the DTPw-HBV/Hib vaccine group. At least 99.1% of subjects in the DTPw-HBV/Hib group and 97.1% of those in the Tritanrix™-HBV/Hib group had seroprotective levels of antibodies against diphtheria, tetanus and hepatitis B and a booster response to the pertussis component following booster administration. Post-booster GMCs did not appear to differ between the two groups, with the exception of anti-PRP GMC which remained significantly higher in the Tritanrix™-HBV/Hib group than in the DTPw-HBV/Hib group (66.6 and 33.4 μg/mL, respectively; non-overlapping CI).

The overall incidence of solicited local and general symptoms following primary and booster doses is shown in Table 2. Pain was the most frequent local solicited symptom in both groups. Grade 3 pain appeared to be more common in the DTPw-HBV/Hib group than in the Tritanrix™-HBV/Hib group after both primary and booster doses (21.8% and 15.4% in the two groups, respectively, following primary vaccination; non-overlapping CI, and 43.4% and 23.5%, respectively, after the booster dose). Irritability and fever were the most common general solicited symptoms. One subject in each group withdrew due to Grade 3 irritability following administration of the first primary dose. Irritability occurred more frequently after the booster dose in the DTPw-HBV/Hib group than in the Tritanrix™-HBV/Hib group (71.7% versus 47.1%, respectively). The incidence of fever (rectal temperature ≥38.0°C) was higher in the DTPw-HBV/Hib group than in the Tritanrix™-HBV/Hib group following both primary and booster vaccination (56.9% versus 46.7%; non-overlapping CI and 70.8% versus 58.8%, respectively). However, the incidence of rectal temperature ≥40.0°C was low and similar in both groups after all doses. Very few subjects required medical advice for solicited symptoms following either primary or booster doses (Table 2).

The incidence of unsolicited symptoms considered related to vaccination did not differ between groups (17.7% of the doses administered in the DTPw-HBV/Hib group and 17.1% in the Tritanrix™-HBV/Hib group after primary vaccination and none were reported in either of the groups after the booster dose). The most common unsolicited symptom considered related to vaccination after primary vaccination was injection site reactions in both groups. Injection site induration was reported following 15.9% and 15.6% of the vaccine doses in the DTPw-HBV/Hib and in the Tritanrix™-HBV/Hib groups respectively. Two SAEs considered related to vaccination were reported during the primary course. Both of these events occurred in the DTPw-HBV/Hib group and required hospitalization, but resolved without sequelae. One subject who presented persistent crying on the day of administration of the first primary dose remained to complete the 3-dose primary vaccination course. The other subject experienced erythema and induration at the vaccine injection site six days after vaccination. The subject was diagnosed as having left thigh abscess and was treated. Following discharge the subject recovered, but was subsequently withdrawn from the study. No SAEs considered related to vaccination were reported after booster administration.