Fecal samples were negative for the presence of rotavirus antigen in all the animals. No gross or microscopic histopathological changes were detected in either sex. All the animals were positive for rotavirus AC220 in vitro antibodies before administration of the vaccine and remained positive 43 days after vaccination. The IgA was determined by using enzyme-linked immunosorbent assay (ELISA) as Modulators described previously [19]. Thus, SII hexavalent BRV vaccine did not cause any toxicity when administered as single and repeated dose by the oral route in Wistar rats and New Zealand

white rabbits. The studies also proved that along with the antigens, the formulation which contains stabilizers and antacid is safe. These results opened prospects for human clinical studies on the vaccine. Considering rotavirus serotype distribution in India, a pentavalent formulation which comprised of G1, G2, G3, G4 and G9 serotypes was used for clinical development (Fig. 1). Three clinical studies (Phase I, Phase IIa and Phase IIb) have been conducted on SII BRV-PV in India (Registration numbers CTRI/2009/091/000821 and CTRI/2010/091/003064). The study populations included adults, toddlers and infants. All studies were approved by the Drug Controller General of India (DCGI) and institutional ethics committees. They complied with all the national regulatory and ethical standards

as well as the ICH good clinical practices (GCP). An independent Data Safety Monitoring Board (DSMB) monitored the safety and rights of the study subjects. The sera samples Z-VAD-FMK in vitro for rotavirus specific IgA antibodies were tested using IgA ELISA at the Christian Medical College, Vellore (India) [19] and stool samples for shedding were tested using rotavirus antigen detection kit (Generic Assays, Germany) at Metropolis Laboratory, Pune. Seroconversion was defined as a change in IgA concentration from <20 U/ml to ≥20 U/ml, or ≥3 fold rise in IgA titers in case of baseline titers ≥20 U/ml. The Phase I study was a randomized, double-blind, placebo controlled study to assess the safety of a single oral dose of SII BRV-PV sequentially in healthy adults, very toddlers and infants. The study also assessed

the immunogenicity and shedding of the vaccine. A single oral dose of the vaccine containing 106 FFU/serotype was investigated in 54 subjects (18 adults, 18 toddlers and 18 infants) who received vaccine or placebo in 2:1 ratio. BRV-PV was found safe and well tolerated in all three age groups. There was no serious adverse event (SAE). The few adverse events reported were mild and transient. Vaccine related events included nausea, loss of appetite, diarrhea and vomiting (Table 1). Except for a few minor changes, the hematology, biochemistry and urine analysis results remained normal in all the groups. No shedding was seen in stool samples. As expected, the single dose of the vaccine did not show immune response in adults and toddlers.