New developments in flavivirus vaccines with special attention to yellow fever

PURPOSE OF REVIEW: Here we review recent epidemiological trends in flavivirus diseases, findings related to existing vaccines, and new directions in flavivirus vaccine research. We emphasize the need for stepped-up efforts to stop further spread and intensification of these infections worldwide. RECENT FINDINGS: Although the incidence and geographic distribution of flavivirus diseases have increased in recent years, human vaccines are available only for yellow fever, Japanese encephalitis, tick-borne encephalitis and Kyasanur forest disease. Factors contributing to resurgence include insufficient supplies of available vaccines, incomplete vaccination coverage and relaxation in vector control. Research has been underway for 60 years to develop effective vaccines against dengue, and recent progress is encouraging. The development of vaccines against West Nile, virus recently introduced to North America, has been initiated. In addition, there is considerable interest in improving existing vaccines with respect to increasing safety (e.g. eliminating the newly recognized syndrome of yellow fever vaccine-associated viscerotropic adverse disease), and to reducing the cost and number of doses required for effective immunization. SUMMARY: Traditional approaches to flavivirus vaccines are still employed, while recent advancements in biotechnology produced new approaches to vaccine design, such as recombinant live virus, subunit and DNA vaccines. Live chimeric vaccines against dengue, Japanese encephalitis and West Nile based on yellow fever 17D virus (ChimeriVax) are in phase I/II trials, with encouraging results. Other chimeric dengue, tick-borne encephalitis and West Nile virus candidates were developed based on attenuated dengue backbones. To further reduce the impact of flavivirus diseases, vaccination policies and vector control programs in affected countries require revision.     

Identification of a recombinant live attenuated respiratory syncytial virus vaccine candidate that is highly attenuated in infants

BACKGROUND: Recombination technology can be used to create live attenuated respiratory syncytial virus (RSV) vaccines that contain combinations of known attenuating mutations. METHODS: Two live attenuated, recombinantly derived RSV vaccine candidates, rA2cp248/404 Delta SH and rA2cp248/404/1030 Delta SH, were evaluated in 31 adults and in 95 children >/=6 months old. rA2cp248/404/1030 Delta SH was subsequently evaluated in 44 infants 1-2 months old. These vaccine candidates share 4 attenuating genetic elements and differ only in a missense mutation (1030) in the polymerase gene. RESULTS: Both vaccines were highly attenuated in adults and RSV-seropositive children and were well tolerated and immunogenic in RSV-seronegative children. Compared with that of rA2cp248/404 Delta SH, replication of rA2cp248/404/1030 Delta SH was restricted in RSV-seronegative children (mean peak titer, 10(4.3) vs. 10(2.5) plaque-forming units [pfu]/mL), indicating that the 1030 mutation had a potent attenuating effect. Although rA2cp248/404/1030 Delta SH was well tolerated in infants, only 44% of infants who received two 10(5.3)-pfu doses of vaccine had detectable antibody responses. However, replication after administration of the second dose was highly restricted, indicating that protective immunity was induced. At least 4 of 5 attenuating genetic elements were retained in recovered vaccine viruses. CONCLUSIONS: rA2cp248/404/1030 Delta SH is the first RSV vaccine candidate to be sufficiently attenuated in young infants. Additional studies are needed to determine whether rA2cp248/404/1030 Delta SH can induce protective immunity against wild-type RSV.     

Cross-Reactive Immunity among Five Medically Important Mosquito-Borne Flaviviruses Related to Human Diseases

Flaviviruses cause a spectrum of potentially severe diseases. Most flaviviruses are transmitted by mosquitoes or ticks and are widely distributed all over the world. Among them, several mosquito-borne flaviviruses are co-epidemic, and the similarity of their antigenicity creates abundant cross-reactive immune responses which complicate their prevention and control. At present, only effective vaccines against yellow fever and Japanese encephalitis have been used clinically, while the optimal vaccines against other flavivirus diseases are still under development. The antibody-dependent enhancement generated by cross-reactive immune responses against different serotypes of dengue virus makes the development of the dengue fever vaccine a bottleneck. It has been proposed that the cross-reactive immunity elicited by prior infection of mosquito-borne flavivirus could also affect the outcome of the subsequent infection of heterologous flavivirus. In this review, we focused on five medically important flaviviruses, and rearranged and recapitulated their cross-reactive immunity in detail from the perspectives of serological experiments in vitro, animal experiments in vivo, and human cohort studies. We look forward to providing references and new insights for the research of flavivirus vaccines and specific prevention.     

Chimeric live, attenuated vaccine against Japanese encephalitis (ChimeriVax-JE): phase 2 clinical trials for safety and immunogenicity, effect of vaccine dose and schedule, and memory response to challenge with inactivated Japanese encephalitis antigen

ChimeriVax-JE is a live, attenuated vaccine against Japanese encephalitis, using yellow fever (YF) 17D vaccine as a vector. In a double-blind phase 2 trial, 99 adults received vaccine, placebo, or YF 17D vaccine (YF-VAX). ChimeriVax-JE was well tolerated, with no differences in adverse events between treatment groups. Viremias resulting from administration of ChimeriVax-JE and YF-VAX were of short duration and low titer; 82 (94%) of 87 subjects administered graded doses (1.8-5.8 log(10)) of ChimeriVax-JE developed neutralizing antibodies. A second dose, administered 30 days later, had no booster effect. Previous inoculation with YF did not interfere with ChimeriVax-JE, but there was a suggestion (not statistically significant) that ChimeriVax-JE interfered with YF-VAX administered 30 days later. A separate study explored immunological memory both in subjects who had received ChimeriVax-JE 9 months before and in ChimeriVax-JE-naive subjects challenged with inactivated mouse-brain vaccine (JE-VAX). Anamnestic responses were observed in preimmune individuals. ChimeriVax-JE appears to be a safe vaccine that provides protective levels of neutralizing antibody after a single dose.     

Superiority of live attenuated compared with inactivated influenza A virus vaccines in older, chronically ill adults

Forty-eight older adults with chronic diseases were vaccinated intranasally with live attenuated influenza A/Korea/1/82 (H3N2), CR59 virus. Forty-two (88 percent) CR59 virus recipients became infected with vaccine virus without adverse effects or change in mean pulmonary function even among the 29 infected recipients with moderate to severe chronic obstructive pulmonary disease. Among control groups who received either monovalent or trivalent inactivated influenza virus vaccines intramuscularly, the rates of fourfold rises in serum antibody titer to hemagglutinin (HA) were not different from the rate following CR59 virus inoculation. However, CR59 virus was superior to inactivated virus vaccine at stimulating secretory antibody to HA. Vaccinees age 65 years and older were more likely to shed CR59 virus in nasal secretions than were younger vaccinees, but antibody responses were not different. CR59 virus vaccine was safe and immunogenic in this population and more often induced a nasal wash IgA antibody response than the inactivated virus vaccines.     

Rabies vaccine prepared in human cell cultures: progress and perspectives

Rabies vaccine prepared in human diploid cell strains is a replacement for the previously available vaccines that are prepared in animal tissues and are less immunogenic and more reactogenic. The human cel-grown vaccine made in the United States is a split-product vaccine, whereas the vaccines made in Europe are whole-virion vaccines. Both types of vaccine contain concentrated and inactivated "fixed" rabies virus. When used before exposure to rabies virus, the vaccine should be given intramuscularly in three 1-ml doses on days 0, 7, and 21. Immediately after exposure to rabies virus, a person should be given human rabies immune globulin (20 international units/kg). This treatment should be followed by five intramuscular doses of vaccine given on days 0, 3, 7, 14, and 28. For maintenance of long-term immunity in persons continously exposed to the risk of rabies, booster doses of the vaccine should be given at two-year intervals.     

Different Cross-Reactivities of IgM Responses in Dengue,Zika and Tick-Borne Encephalitis Virus Infections

Flaviviruses circulate worldwide and cause a number of medically relevant human diseases, such as dengue, Zika, yellow fever, and tick-borne encephalitis (TBE). Serology plays an important role in the diagnosis of flavivirus infections, but can be impeded by antigenic cross-reactivities among flaviviruses. Therefore, serological diagnosis of a recent infection can be insufficiently specific, especially in areas where flaviviruses co-circulate and/or vaccination coverage against certain flaviviruses is high. In this study, we developed a new IgM assay format, which is well suited for the specific diagnosis of TBE, Zika and dengue virus infections. In the case of TBE and Zika, the IgM response proved to be highly specific for the infecting virus. In contrast, primary dengue virus infections induced substantial amounts of cross-reactive IgM antibodies, which is most likely explained by structural peculiarities of dengue virus particles. Despite the presence of cross-reactive IgM, the standardized nature and the quantitative read-out of the assay even allowed the serotype-specific diagnosis of recent dengue virus infections in most instances.     

Immunity to influenza A virus infection in young children: a comparison of natural infection, live cold-adapted vaccine, and inactivated vaccine

Live attenuated, cold-adapted (ca) influenza A vaccines administered intranasally have been well characterized as safe and immunogenic, but comparative data on protective efficacy are required for further development. In this study, 59 young children were divided into the following four groups based on prior exposure to influenza A (H3N2) virus: natural infection, live ca vaccine given intranasally, inactivated vaccine given im, and no previous exposure. Virus challenge with homologous live ca vaccine occurred 12 months after vaccination or natural infection. Prior natural infection and live ca vaccine significantly reduced ca virus shedding after challenge compared with inactivated vaccine or no prior exposure to influenza A virus. Prechallenge nasal IgA, detected almost exclusively in subjects naturally infected or vaccinated with live ca virus, was associated with protection. Although inactivated vaccine failed to produce significant local IgA during the primary response, it seemed to prime for secondary local antibody responses after challenge with live ca virus.     

The effect of chloroquine prophylaxis on yellow fever vaccine antibody response: comparison of plaque reduction neutralization test and enzyme-linked immunosorbent assay

Weekly oral chloroquine prophylaxis for malaria has been associated with impaired antibody response to intradermal rabies vaccination. Experimental data indicate that chloroquine may inhibit yellow fever virus in vitro, yet there has been no clinical evidence to suggest that antibody response to yellow fever vaccine is impaired by concomitant oral administration of chloroquine. A prospective trial was undertaken to evaluate the antibody response to yellow fever 17D vaccine (Connaught Laboratories) of volunteers who were randomized to taking either chloroquine or no drug. Of fifty subjects, 28 were randomized to taking chloroquine, 22 were randomized to taking no drug. Yellow fever 17D vaccine was administered on day 0 and blood sampled on days 0, 14, 35 and 210. Chloroquine was administered weekly for four weeks. There was no significant difference in peak antibody titer by plaque reduction neutralization testing (PRNT) between the group that took chloroquine (mean log peak of reciprocal titer 1.43 +/- SD 0.60) with vaccine subcutaneously compared to vaccine-only group (mean log peak of reciprocal titer = 1.21 +/- 0.55). All fifty subjects seroconverted to yellow fever vaccine by day 210. ELISA testing was also performed on all subjects. The two tests showed good correlation (Spearman r = 0.675), although ELISA readings were positive by day 14 in significantly more subjects (p = .01). We conclude that routine anti-malarial doses of chloroquine do not affect antibody response to yellow fever 17D vaccine. ELISA testing, a less complex and less time-consuming test, correlates well with PRNT and is proposed for additional trials to measure yellow fever 17D vaccine response in flavivirus non-immune subjects.     

Decline in varicella-zoster virus (VZV)-specific cell-mediated immunity with increasing age and boosting with a high-dose VZV vaccine

The safety and immunogenecity of a booster dose of live attenuated varicella-zoster virus (VZV) vaccine was evaluated in 196 healthy subjects, >or=60 years old, who had already received a VZV vaccine >5 years before. This repeat booster dose was well tolerated. Cell-mediated immunity (CMI) to VZV was measured by an interferon-gamma (IFN-gamma) enzyme-linked immunosorbent spot-forming cell (ELISPOT) assay and a limiting dilution responder cell frequency (RCF) assay. Prevaccination responses decreased as a function of increasing age but were detectable in all subjects by use of the IFN-gamma ELISPOT assay. In most subjects, VZV-specific CMI was increased at 6 weeks postvaccination. The magnitude of the vaccine-induced IFN-gamma ELISPOT response was inversely related to prevaccination values. Although there was a significant correlation between the IFN-gamma ELISPOT and RCF assays, the ELISPOT assay had greater sensitivity and a wider dynamic range. A live attenuated VZV vaccine is safe and immunogenic in an elderly population, and the vaccine-induced immunity may be monitored by the IFN-gamma ELISPOT assay.     

rDEN4delta30, a live attenuated dengue virus type 4 vaccine candidate, is safe, immunogenic, and highly infectious in healthy adult volunteers

BACKGROUND: The live attenuated dengue virus type 4 (DEN-4) vaccine candidate virus rDEN4 Delta 30 was previously found to be safe and immunogenic at a dose of 10(5) plaque-forming units (pfu). METHODS: In a follow-up placebo-controlled phase 2 clinical trial, rDEN4 Delta 30 was administered as a single inoculation to 3 separate dose cohorts (10(3) pfu, 10(2) pfu, or 10(1) pfu), for further evaluation. Each dose cohort consisted of 20 vaccinees and 4 placebo recipients. Volunteers were monitored closely for adverse events, and serum was collected on study days 28 and 42 for determination of neutralizing antibody titer. RESULTS: The vaccine was well tolerated at all doses studied. The most common adverse events observed were a transient asymptomatic rash in >50% of vaccinees and a mild neutropenia in approximately 20% of vaccinees. No vaccinee developed a dengue-like illness. The vaccine was highly infectious and immunogenic, with 95%-100% of vaccinees in each dose cohort developing a >/=4-fold increase in titers of serum neutralizing antibodies against DEN-4. CONCLUSIONS: The rDEN4 Delta 30 vaccine is safe and induced an antibody response that was broadly neutralizing against genotypically diverse DEN-4 viruses. It is a promising vaccine candidate for inclusion in a tetravalent dengue vaccine formulation.     

Comparison of the immunogenicity and safety of two 17D yellow fever vaccines.

As part of the clinical validation process of a new working seed of a licensed yellow fever vaccine (new working seed PV26, Stamaril; Pasteur Mérieux Connaught, Lyon, France), the immunogenicity and safety of two batches of this vaccine (PM-YF) were compared with those of another commercially available vaccine (Arilvax; Evans Medical-Wellcome, Liverpool, United Kingdom) in 211 healthy adults. While the geometric mean titer values at days 10-14 and day 28 after vaccination were higher in the PM-YF group, the vaccines provided equivalent seroprotection (titers > or = 1/10) one month after a single vaccine dose (100% PM-YF versus 99% W-YF; P = 0.001, by one-sided equivalence test). Both vaccines were safe. There were no serious local or systemic reactions reported, nor any clinically significant hepatic function abnormalities associated with the use of either vaccine. These two 17D yellow fever vaccines from different European vaccine manufacturers were highly immunogenic and safe, and provided equivalent seroprotection.     

Safety and immunogenicity of an inactivated thimerosal‐free influenza vaccine in infants and children

Objective Few prospective studies of inactivated split virion influenza vaccine have been conducted in infants and children. Our objective was to evaluate the safety, reactogenicity and immunogenicity of a thimerosal‐free inactivated influenza vaccine (Fluvax^®; CSL Limited, Parkville, Australia) in children aged 6 months to <9 years. Methods A prospective, open‐label, phase III clinical trial was conducted in 298 healthy children previously unvaccinated with influenza, commencing in the Southern Hemisphere 2005 autumn. Participants were divided into two groups (Group A: ≥6 months to <3 years; Group B: ≥3 years to <9 years), and received two doses of the 2005 vaccine, and one dose of the 2006 vaccine one year later (Group A: 0·25 ml per dose; Group B: 0·5 ml per dose). Vaccine safety and reactogenicity was evaluated for 30 days after each dose. Immunogenicity was assessed using hemagglutination inhibition and single radial hemolysis assays. Results There were no withdrawals due to adverse events (AEs). The majority of solicited local and systemic AEs were of mild severity. A maximum intensity of severe was reported for injection site pain and fever by only 3·0% and 3·4% of participants, respectively. The vaccine was immunogenic for all antigens, with ≥95% of both younger and older children achieving seroprotection after dose 2. Conclusions This thimerosal‐free inactivated influenza vaccine had a favorable safety profile and was immunogenic in children aged ≥6 months and <9 years. Primary and booster vaccination produced consistently immunogenic responses including in children under 3 years of age receiving 0·25 ml doses of vaccine.     

Future vaccines against enteric pathogens

A small number of bacterial agents, including enterotoxigenic Escherichia coli (ETEC), enteropathogenic E. coli (EPEC), Shigella and Vibrio cholerae 01 and one virus, rotavirus, combine to cause a major proportion of the diarrheal illness of public health importance worldwide. Salmonella typhi is by far the major cause of enteric fever. Attempts to develop safe, practical, and effective vaccines against these agents are under way. Examples of vaccine candidates include live oral vaccines against S. typhi, V. cholerae, Shigella, and rotavirus, and inactivated, submit vaccines given parentally or orally against S. typhi, V. cholerae or ETEC. Although the oral attenuated S. typhi vaccine is ready for commercial license, it will be several years before the other vaccines are proven to be practically safe and effective in children and adults.     

Nanoparticles in influenza subunit vaccine development: Immunogenicity enhancement

The threat of novel influenza infections has sparked research efforts to develop subunit vaccines that can induce a more broadly protective immunity by targeting selected regions of the virus. In general, subunit vaccines are safer but may be less immunogenic than whole cell inactivated or live attenuated vaccines. Hence, novel adjuvants that boost immunogenicity are increasingly needed as we move toward the era of modern vaccines. In addition, targeting, delivery, and display of the selected antigens on the surface of professional antigen‐presenting cells are also important in vaccine design and development. The use of nanosized particles can be one of the strategies to enhance immunogenicity as they can be efficiently recognized by antigen‐presenting cells. They can act as both immunopotentiators and delivery system for the selected antigens. This review will discuss on the applications, advantages, limitations, and types of nanoparticles (NPs) used in the preparation of influenza subunit vaccine candidates to enhance humoral and cellular immune responses.     

Dengue virus type 2 vaccine: reactogenicity and immunogenicity in soldiers

A live dengue virus type 2 (dengue-2) vaccine (PR-159/S-1) was tested for reactogenicity and immunogenicity in a placebo-controlled, double-blind clinical trial involving 98 soldiers. Seroconversion rates based on the development of neutralizing antibody to dengue-2 were 90% in 70 recipients with immunity to yellow fever and 61% in 28 vaccinees without such immunity (P less than .01). Peak titers of neutralizing antibody were three times higher in recipients with antibody to yellow fever virus and persisted in most for at least 18 months. Individuals seroconverting to the vaccine virus more frequently experienced systemic symptoms than those who received placebo (P less than .02). Future users of this dengue-2 vaccine may wish to employ immunization schedules that include preliminary immunization against yellow fever and must be prepared to accept mild vaccine-related symptoms in some recipients.     

Administration of live varicella vaccine to HIV-infected children with current or past significant depression of CD4(+) T cells

BACKGROUND: Varicella can be a severe illness in human immunodeficiency virus (HIV)-infected children. The licensed, live attenuated varicella vaccine is safe and immunogenic in HIV-infected children with minimal symptoms and good preservation of CD4(+) T cells (Centers for Disease Control and Prevention immunologic category 1). METHODS: To study the safety and immunogenicity of this vaccine in varicella-zoster virus (VZV)-naive, HIV-infected children with moderate symptoms and/or more pronounced past or current decreases in CD4(+) T cell counts, such children (age, 1-8 years) received 2 doses of vaccine 3 months apart. The children were observed in a structured fashion for adverse events. Blood was tested for VZV antibody and VZV-specific cell-mediated immunity (CMI) at baseline, 8 weeks after each dose, and annually for 3 years. Subjects who had no evidence of immunity 1 year after vaccination received a third dose and were retested. RESULTS: The vaccine was well tolerated; there were no vaccine-related, serious adverse events. Regardless of immunologic category, at least 79% of HIV-infected vaccine recipients developed VZV-specific antibody and/or CMI 2 months after 2 doses of vaccine, and 83% were responders 1 year after vaccination. CONCLUSIONS: HIV-infected children with a CD4(+) T cell percentage of > or =15% and a CD4(+) T cell count of > or =200 cells/ microL are likely to benefit from receiving varicella vaccine.     

Combined effects of immunomodulators and specific inactivated vaccines against alpha- and flavivirus infections

The combined effects of immunomodulators, such as ridostin, polyribonate, glucose muramyl dipeptide, and peptidoglycan-160, and specific vaccines on survival of mice with alpha- (eastern equine encephalitis) and flavivirus (tick-borne encephalitis (TBE) infections were found to be significant. In alphavirus infection, the combined effects of the specific vaccine and ridostin were accompanied by increases in specific humoral immunity (specific antibodies) and cellular immunity (adoptive transfer of immune lymphocytes). The concurrent use of the specific vaccine and the immunomodulator ridostin is recommended in clinical trials of TBE in the foci of infection.     

Efficacy and Duration of Immunity after Yellow Fever Vaccination: Systematic Review on the Need for a Booster Every 10 Years

Current regulations stipulate a yellow fever (YF) booster every 10 years. We conducted a systematic review of the protective efficacy and duration of immunity of YF vaccine in residents of disease-endemic areas and in travelers to assess the need for a booster in these two settings and in selected populations (human immunodeficiency virus–infected persons, infants, children, pregnant women, and severely malnourished persons). Thirty-six studies and 22 reports were included. We identified 12 studies of immunogenicity, 8 of duration of immunity, 8 of vaccine response in infants and children, 7 of human-immunodeficiency virus–infected persons, 2 of pregnant women, and 1 of severely malnourished children. Based on currently available data, a single dose of YF vaccine is highly immunogenic and confers sustained life-long protective immunity against YF. Therefore, a booster dose of YF vaccine is not needed. Special considerations for selected populations are detailed.     

减毒猪流感病毒与灭活猪流感病毒的体液免疫及细胞免疫效果的观察

摘要：目的 研究减毒A/California/07/2009ca流感病毒及A/California/07/2009灭活流感病毒之间的体液免疫及细胞免疫效果的差异。方法 蔗糖密度梯度离心纯化减毒A/California/07/2009ca流感病毒经滴鼻途径免疫BALB/c小鼠,以灭活病毒肌肉注射组为对照。应用ELISA的方法检测小鼠血清中IgG1、IgG2a及脾淋巴细胞上清液中的IL-6、IL-4和IFN-γ的效价。结果 通过对小鼠血清中IgG1、IgG2a及脾淋巴细胞上清液中的IL-6、IL-4及IFN-γ细胞因子的检测,结果显示,灭活流感病毒的IgG1/IgG2a高于减毒流感病毒,同时灭活病毒产生的IL-6和IL-4高于减毒病毒,但IFN-γ低于减毒病毒。结论 A/California/07/2009灭活病毒能够刺激小鼠机体产生较好的体液免疫,但细胞免疫较差,而减毒A/California/07/2009ca流感病毒能够较好激发小鼠体内的体液免疫和细胞免疫反应。通过这些数据我们可以初步断定,A/California/07/2009ca减毒病毒具有较全面的免疫效果,这为流感喷鼻疫苗的研制提供了理论数据参考。