Yanchun Che

An Inactivated Enterovirus 71 Vaccine in Healthy Children

BACKGROUND Enterovirus 71 (EV71) is a major cause of hand, foot, and mouth disease in children and may be fatal. A vaccine against EV71 is needed. METHODS We conducted a randomized, double-blind, placebo-controlled phase 3 trial involving healthy children 6 to 71 months of age in Guangxi Zhuang Autonomous Region, China. Two doses of an inactivated EV71 vaccine or placebo were administered intramuscularly, with a 4-week interval between doses, and children were monitored for up to 11 months. The primary end point was protection against hand, foot, and mouth disease caused by EV71. RESULTS A total of 12,000 children were randomly assigned to receive vaccine or placebo. Serum neutralizing antibodies were assessed in 549 children who received the vaccine. The seroconversion rate was 100% 4 weeks after the two vaccinations, with a geometric mean titer of 170.6. Over the course of two epidemic seasons, the vaccine efficacy was 97.4% (95% confidence interval [CI], 92.9 to 99.0) according to the intention-to-treat analysis and 97.3% (95% CI, 92.6 to 99.0) according to the per-protocol analysis. Adverse events, such as fever (which occurred in 41.6% of the participants who received vaccine vs. 35.2% of those who received placebo), were significantly more common in the week after vaccination among children who received the vaccine than among those who received placebo. CONCLUSIONS The inactivated EV71 vaccine elicited EV71-specific immune responses and protection against EV71-associated hand, foot, and mouth disease. (Funded by the National Basic Research Program and others; ClinicalTrials.gov number, NCT01569581.).

Study of the Integrated Immune Response Induced by an Inactivated EV71 Vaccine

Enterovirus 71 (EV71), a major causative agent of hand-foot-and-mouth disease (HFMD), causes outbreaks among children in the Asia-Pacific region. A vaccine is urgently needed. Based on successful pre-clinical work, phase I and II clinical trials of an inactivated EV71 vaccine, which included the participants of 288 and 660 respectively, have been conducted. In the present study, the immune response and the correlated modulation of gene expression in the peripheral blood mononuclear cells (PBMCs) of 30 infants (6 to 11 months) immunized with this vaccine or placebo and consented to join this study in the phase II clinical trial were analyzed. The results showed significantly greater neutralizing antibody and specific T cell responses in vaccine group after two inoculations on days 0 and 28. Additionally, more than 600 functional genes that were up- or down-regulated in PBMCs were identified by the microarray assay, and these genes included 68 genes associated with the immune response in vaccine group. These results emphasize the gene expression profile of the immune system in response to an inactivated EV71 vaccine in humans and confirmed that such an immune response was generated as the result of the positive mobilization of the immune system. Furthermore, the immune response was not accompanied by the development of a remarkable inflammatory response. Clinical Trial Registration: NCT01391494 and NCT01512706.

Comparative study of the immunogenicity in mice and monkeys of an inactivated CA16 vaccine made from a human diploid cell line

The coxsackie A16 virus (CA16), along with enterovirus 71 (EV71), is a primary pathogen that causes hand, foot, and mouth disease (HFMD). To control HFMD, CA16, and EV71 vaccines are needed. In this study, an experimental inactivated CA16 vaccine was prepared using human diploid cells, and the vaccine’s immunogenicity was analyzed in mice and rhesus monkeys. The results showed that the neutralizing antibody was developed in a dose-dependent manner, and was sustained for 70 days with an average GMT (geometric mean titer) level of 80 to 90 in immunized mouse and for 56 days with GMT of higher than 300 in monkeys. The neutralizing antibody had a cross-neutralizing activity against different viral strains (genotype A and B), and the specific IFN-γ-secreting cell response was activated by these virus strains in an ELISPOT assay. This study provides evidence for the potential use of inactivated CA16 as a candidate for use in vaccines.

The effect of enterovirus 71 immunization on neuropathogenesis and protein expression profiles in the thalamus of infected rhesus neonates

Enterovirus 71 (EV71) is a major pathogen that causes hand-foot-mouth disease (HFMD). Our previous studies have demonstrated that the complete process of pathogenesis, which may include tissue damage induced by host inflammatory responses and direct tissue damage caused by viral infection, can be observed in the central nervous system (CNS) of animals infected in the laboratory with EV71. Based on these observations, the neuropathogenesis and protein expression profiles in the thalamic tissues of EV71-infected animals were further analyzed in the present study. Changes in protein expression profiles following immunization with the inactivated EV71 vaccine followed by virus challenge were observed and evaluated, and their physiological roles in viral pathogenesis are discussed. Taken together, the results of these experiments provide evidence regarding the neuropathogenesis and molecular mechanisms associated with EV71 infection and identify several protein indicators of pathogenic changes during viral infection.

Immunity and clinical efficacy of an inactivated enterovirus 71 vaccine in healthy Chinese children: a report of further observations

BackgroundTo investigate the long-term effects on immunity of an inactivated enterovirus 71 (EV71) vaccine and its protective efficacy.MethodsA sub-cohort of 1,100 volunteers from Guangxi Province in China was eligible for enrolment and randomly administered either the EV71 vaccine or a placebo on days 0 and 28 in a phase III clinical trial and then observed for the following 2 years with approval by an independent ethics committee of Guangxi Zhuang Autonomous Region, China. Serum samples from the 350 participants who provided a full series of blood samples (at all the sampling points) within the 2-year period were collected. Vaccine-induced immune effects, including the neutralizing antibody titres and cross-protection against different genotypes of EV71, were examined. This study also evaluated the protective efficacy of this vaccine based upon clinical diagnosis.ResultsThis sub-cohort showed a >60 % drop-out rate over 2 years. The seroconversion rates among the 161 immunized subjects remained >95 % at the end of study. The geometric mean titres of neutralizing antibodies (anti-genotype C4) 360 days after vaccination in 350 subjects were 81.0 (subjects aged 6–11 months), 98.4 (12–23 months), 95.0 (24–35 months), and 81.8 (36–71 months). These titres subsequently increased to 423.1, 659.0, 545.0, and 321.9, respectively, at 540 days post-immunization (d.p.i.), and similar levels were maintained at 720 d.p.i. Higher IFN-γ/IL-4-specific responses to the C4 genotype of EV71 and cross-neutralization reactivity against major EV71 genotype strains were observed in the vaccine group compared to those in the placebo group. Five EV71-infected subjects were observed in the placebo-treated control group and none in the vaccine-immunized group in per-protocol analysis.ConclusionThese results are consistent with the induction of dynamic immune responses and protective efficacy of the vaccine against most circulating EV71 strains.Trial registration numberClinicaltrials.gov, NCT01569581, Trial registration date: March 2012

Analysis of the Th1/Th2 Reaction in the Immune Response Induced by EV71 Inactivated Vaccine in Neonatal Rhesus Monkeys

Although clinical trials for the enterovirus type 71 (EV71) inactivated vaccine have been progressing, the potential mechanism of EV71 infection and its associated pathogenesis are not well-characterized in terms of comprehensive analysis of the induced immune response, which is generally recognized as an important indicator of the safety of vaccines. To investigate the Th1/Th2 response following viral challenge in neonatal rhesus monkeys immunized with different doses of EV71 inactivated vaccines, the variety of different Th1 and Th2 cytokines in the organs or tissues of the monkeys were identified. The results suggest that depending on the viral challenge, the Th1/Th2 reaction induced by different doses of EV71 inactivated vaccine varies. More specifically, there is an enhanced immune response in 80EU- and 1280EU-immunized monkeys, whereas 320EU immunization induces a mild response. Although there is no direct impact on the variation in immune protection induced by the vaccine, the Th1 reaction functions in T-cell cytotoxicity, which will aid further investigation of the pathogenic characteristics of small pathological changes in the central nerves system (CNS) likely induced by the Th1 response.

Egress of HSV-1 capsid requires the interaction of VP26 and a cellular tetraspanin membrane protein

HSV-1 viral capsid maturation and egress from the nucleus constitutes a self-controlled process of interactions between host cytoplasmic membrane proteins and viral capsid proteins. In this study, a member of the tetraspanin superfamily, CTMP-7, was shown to physically interact with HSV-1 protein VP26, and the VP26-CTMP-7 complex was detected both in vivo and in vitro. The interaction of VP26 with CTMP-7 plays an essential role in normal HSV-1 replication. Additionally, analysis of a recombinant virus HSV-1-UG showed that mutating VP26 resulted in a decreased viral replication rate and in aggregation of viral mutant capsids in the nucleus. Together, our data support the notion that biological events mediated by a VP26 - CTMP-7 interaction aid in viral capsid enveloping and egress from the cell during the HSV-1 infectious process.

The Gene Expression Profile of Peripheral Blood Mononuclear Cells from EV71-Infected Rhesus Infants and the Significance in Viral Pathogenesis

Enterovirus 71 (EV71) is the major pathogen responsible for fatal hand, foot and mouth disease (HFMD). Our previous work reported on an EV71-infected rhesus monkey infant model that presented with histo-pathologic changes of the central nervous system (CNS) and lungs. This study is focused on the correlated modulation of gene expression in the peripheral blood mononuclear cells (PBMCs) from EV71-infected rhesus monkey infants. The expression of more than 500 functional genes associated with multiple pathways was modulated. The expression of genes associated with immune inflammatory responses was up-regulated during the period from days 4 to 10 post-infection. The expression of two genes (TAC1 and IL17A), which play major roles in inflammatory reactions, was remarkably up-regulated during the infection period. Furthermore, a higher expression level of the TAC1 gene was identified in the CNS compared to the lungs, but a high expression level of the IL-17A gene was observed in the lungs and not in the CNS. The results of this study suggest at least two facts about EV71 infection, which are that: the TAC1 gene that encodes substance P and neurokinin-A is present in both PBMCs and the hypothalamus; and the up-regulation of IL-17A is sustained in the peripheral blood.

Coxsackievirus A 16 infection does not interfere with the specific immune response induced by an enterovirus 71 inactivated vaccine in rhesus monkeys

Hand, foot and mouth disease is usually caused by enterovirus 71 (EV71) and coxsackievirus A 16 (CA16), which are members of the Picornaviridae family. In the present study, the characteristics of the immune response induced by an EV71 inactivated vaccine (made from human diploid cells) were explored in the presence of CA16 infection, based on the previously established neonatal rhesus monkey model. The typical clinical manifestations, including body temperature, viral viremia and virus shedding in the mouth, pharynx and feces, were characterized. A specific neutralizing antibody assay showed that the specific immune response induced by the EV71 inactivated vaccine was active against EV71 but not against CA16. No remarkable fluctuation in proinflammatory cytokine release was identified in the serum of immunized monkeys with EV71 vaccine and CA16 infections subsequently. The results showed that the specific immune response induced by the EV71 inactivated vaccine is effective against EV71 infection but is not affected by CA16 infection.

Functional analysis of transcriptional regulation of herpes simplex virus type 1 tegument protein VP22

The herpes simplex virus type 1 (HSV-1) tegument proteins have important functions in the viral replication process. In order to investigate the role of the HSV-1 tegument protein VP22 in viral replication, its transcriptional regulation of viral promoters was investigated using the chloramphenicol acetyltransferase (CAT) assay. The results indicate that VP22 exerts a dose-dependent transcriptional inhibitory effect on the HSV-1 α4, TK, and gC gene promoters. VP22 had the capacity to repress transcriptional activation of promoters via different viral transcription regulatory factors such as VP16 and ICP0, as evidenced by the specific repression of the TK and gC gene promoters by ICP0. In addition, VP22 was capable of inhibiting the promotion of ICP0 transcriptional activation in the presence of HAT PCAF, which is even more remarkable than the VP22 repression of ICP0 transcriptional activation. Finally, the transcriptional inhibitory effect of VP22 on other viral promoters was demonstrated by the analysis of β-galactosidase activities in internal controls.