Human neonatal rotavirus vaccine (RV3-BB) targets rotavirus from birth

Background A birth dose strategy using a neonatal rotavirus vaccine to target early prevention of rotavirus disease may address remaining barriers to global vaccine implementation. Methods We conducted a randomized, placebo-controlled trial in Indonesia to evaluate the efficacy of an oral human neonatal rotavirus vaccine (RV3-BB) to prevent rotavirus gastroenteritis. Healthy newborns received three doses of RV3-BB administered in a neonatal schedule at 0-5 days, 8 and 14 weeks or infant schedule at 8, 14 and 18 weeks, or placebo. Laboratory-confirmed rotavirus gastroenteritis was graded using a modified Vesikari score. The primary analysis was efficacy against severe rotavirus gastroenteritis from two weeks after all doses to 18 months in the combined vaccine group (neonatal and infant schedule) compared with placebo. Results Vaccine efficacy against severe rotavirus gastroenteritis to 18 months was 63% in the combined vaccine group (95% CI 34, 80; p<0.001), 75% in the neonatal vaccine group (95% confidence interval [CI] 44, 91; p<0.001) and 51% in the infant vaccine group (95% CI 7, 76; p=0.03) in the per protocol analysis, with similar results in the intention-to-treat analysis. Vaccine efficacy to 12 months was 94% in the neonatal vaccine group (95%CI 56, 99; p=0.006). Vaccine take occurred in 78/83 (94%) in the neonatal vaccine group and 83/84 (99%) in the infant vaccine group. The vaccine was well tolerated, with similar incidence of adverse events in vaccine and placebo recipients. Conclusion RV3-BB was efficacious, immunogenic and well-tolerated when administered in a neonatal or infant schedule in Indonesia.

Despite evidence of success of rotavirus vaccines, over 90 million infants still lack 93 access to a rotavirus vaccine (1,2). Barriers to global implementation include cost, 94 sub-optimal efficacy in low-income countries and lingering safety concerns (3,4). An 95 oral rotavirus vaccine administered at birth has potential to address these challenges. 96 Rotavirus disease occurs early in life in infants in low-income countries (5). A birth 97 dose rotavirus vaccine would provide early protection and maximize the opportunity to 98 complete a full vaccine schedule (6). Birth presents a unique opportunity that may 99 assist the uptake of an oral vaccine as gastric acid is limited and environmental 100 enteropathy not yet established (7,8). As intussusception is rare in newborns, a birth 101 dose administration may offer a safety advantage (9). 102 RV3-BB vaccine was developed from the human neonatal rotavirus strain, RV3 103 (G3P [6]), identified in the stool of asymptomatic infants (10). Wild-type infection with 104 RV3 provided protection from severe gastroenteritis in the first 3 years of life, with 105 strong heterotypic serological responses to community rotavirus strains (11,12). RV3 106 appears to be naturally attenuated and adapted to the newborn gut, replicating well 107 despite the presence of maternal antibodies and breastfeeding (13). RV3-BB vaccine 108 aims to take advantage of the intrinsic characteristics of this novel strain to target a 109 birth dose vaccination strategy. RV3-BB was well tolerated and immunogenic when 110 delivered in a neonatal or infant schedule in a phase IIa trial in New Zealand (14). 111 The primary objective of this study was to assess the efficacy of three doses of RV3-112 BB against severe rotavirus gastroenteritis to 18 months of age. Secondary objectives 113 included assessment of efficacy, immunogenicity and safety of RV3-BB when 114 delivered in a neonatal schedule (first dose 0-5 days of age), or an infant schedule (first dose 8-10 weeks of age), compared with placebo, efficacy to 12 months, against 116 rotavirus gastroenteritis of any severity and all-cause severe gastroenteritis. 117

Trial design and oversight 119
This phase IIb, randomized, double-blind, placebo-controlled trial involving 1649 120 participants was conducted from January 2013 to July 2016 in primary health centers 121 and hospitals in Central Java and Yogyakarta, Indonesia. Indonesia is a low-middle 122 income country with an under-5 mortality rate in Yogyakarta and Central Java of 30-123 38 per 1000 live births and per capita gross regional product of USD $2,164-$2,326 124 score was applied where intravenous, naso-gastric rehydration or 6-hours of 175 supervised oral rehydration was scored as hospitalization, whether administered 176 within a primary health center or hospital. Gastroenteritis of any severity, defined as 177 three or more stools looser than normal for that child within a 24-hour period. 178

Vaccine Take and Immunogenicity 179
Vaccine take was assessed in the first cohort recruited (n=282). Blood was collected 180 from the cord (baseline for neonatal schedule comparison), immediately prior to IP 181 dose 2 (baseline for infant schedule comparison), 28 days after IP dose 3 and 28 days 182 after IP dose 4. Serum rotavirus immunoglobulin A (IgA) antibody titers and serum 183 neutralizing antibody titers were measured using previously described methods (14, 184 18). RV3-BB shedding in stool was detected using a rotavirus VP6 specific reverse 185 transcription-polymerase chain reaction assay and confirmed by sequence analysis 186 (14). Positive vaccine take was defined as a serum immune response (≥3 fold increase 187 in titer from baseline in anti-rotavirus IgA or serum neutralising antibodies) 28 days 188 following IP administration, or RV3-BB shedding on days 3-7 following IP administration. Cumulative vaccine take was defined as a positive vaccine take 190 following IP dose of 1, 2 or 3 for the neonatal vaccine group, and following IP doses 2, 191 3 or 4 for the infant vaccine group. 192

Safety 193
Vital signs were assessed prior to, and in the 30 minutes after IP administration. population (all randomized participants), comparing events from randomisation to 18 209 months. Vaccine efficacy is presented as 1-risk ratio x 100 with its exact 95% 210 confidence interval based on the Clopper-Pearson method (19). Based on local data we assumed 3% of placebo participants would experience an 221 episode of severe rotavirus gastroenteritis during the study (20,21) and calculated a 222 sample size of 549 participants in each group would provide 80% power to reject the 223 null hypothesis of no difference between the combined vaccine and placebo groups if 224 the true efficacy was 60% (one-sided test with alpha of 0.1), allowing for 10% non-225 adherence. We calculated 282 participants were required to reject the null hypothesis 226 of no difference in the proportion with a positive vaccine take (two-sided test with alpha 227 of 0.05) assuming 25% of placebo participants would be exposed to rotavirus (14) and 228 50% in each vaccine group would have a positive vaccine take, allowing for 10% non-229

232
Of the 1649 newborns randomized, 1640 received at least one dose of IP (safety 233 population) and 1588 (96%) were followed to 18 months. The primary efficacy analysis 234 was performed on 1513 (92%) in the PP population (Figure 1b). The demographic 235 characteristics of the study population and age of receipt of first dose of IP were similar 236 across all groups (Appendix : Table S1). 237

Vaccine Efficacy 238
Severe rotavirus gastroenteritis occurred in 28  Ghana, and Kenya) (23,24) and Rotavac (Bharat Biotech) was 55.1% (India) (25). 289 Three doses of Rotasil (Serum Institute of India) had an efficacy of 66.7% at a mean 290 follow up of 9.8 months of age in Niger (26). If the 75% protective efficacy for the 291 neonatal schedule of RV3-BB translates into effectiveness throughout Indonesia, it 292 has the potential to avert an estimated 5,450 deaths, 117,110 hospitalizations and 293 >300,000 outpatient clinic visits each year due to rotavirus gastroenteritis in children 294 under 5 years (27). 295 The concept of a birth dose strategy for vaccination is not new. Birth is an established 296 immunization time-point in many countries. A neonatal dose was investigated in the 297 early phase of rotavirus vaccine development but not pursued due to concerns 298 regarding inadequate immune responses and safety (28)(29)(30). The VP4 protein of 299 human neonatal P [6] strains have specific residues at the basal surface of VP8* that 300 may allow them to adhere to cell surface receptors in the newborn gut (31). This may 301 provide an advantage for a birth dose schedule. The P [6] VP4 of RV3-BB may also 302 offer an advantage in Africa and Asia where the Lewis-negative phenotype is common 303 (32). Lewis (FUT3) and secretor (FUT2) genes appear to mediate susceptibility to 304 rotavirus infection (32). P [8] rotaviruses only infect individuals who are Lewis-positive 305 and secretor-positive whereas P [6] rotaviruses infect individuals irrespective of their 306 Lewis and secretor status (33). This may explain the high proportion of disease caused 307 by P [6] rotaviruses in Africa and the lower efficacy of vaccines with a P [8] genotype in 308 these region (34). RV3-BB is currently the only vaccine with a P[6] VP4. 309 Unlike IgG, IgA is not transferred via the placenta, and the newborn may not mount a 310 significant serum IgA response following the birth dose of an oral vaccine, such as 311 RV3-BB, despite evidence that the neonatal schedule is efficacious (35). Similar 312 dissonance has been demonstrated with other vaccines administered in the newborn 313 period (36). An equine-like G3P [8] strain was responsible for most episodes of severe 314 gastroenteritis in this study and reflects the global emergence of this strain (37). Based on the strong heterotypic serological responses to community strains (G1,G2 316 dominant) provided by the parent strain RV3 (11), it is anticipated that RV3-BB will 317 also offer protection against a range of circulating rotavirus strain but this could not be 318 assessed in this study.