流感减毒活疫苗临床效率的研究进展

流感病毒可以引起人类严重的呼吸系统疾病,甚至导致死亡。由于病毒具有高突变率及其抗原漂移、基因重配等特性,因此导致流感的反复暴发,甚至大流行。疫苗是预防流感最有效的手段,不同种类的流感疫苗在不同人群中会产生不同的免疫效果,本文就流感减毒活疫苗的临床效率作一综述。     

Engineering temperature sensitive live attenuated influenza vaccines from emerging viruses

The licensed live attenuated influenza A vaccine (LAIV) in the United States is created by making a reassortant containing six internal genes from a cold-adapted master donor strain (ca A/AA/6/60) and two surface glycoprotein genes from a circulating/emerging strain (e.g., A/CA/7/09 for the 2009/2010 H1N1 pandemic). Technologies to rapidly create recombinant viruses directly from patient specimens were used to engineer alternative LAIV candidates that have genomes composed entirely of vRNAs from pandemic or seasonal strains. Multiple mutations involved in the temperature-sensitive (ts) phenotype of the ca A/AA/6/60 master donor strain were introduced into a 2009 H1N1 pandemic strain rA/New York/1682/2009 (rNY1682-WT) to create rNY1682-TS1, and additional mutations identified in other ts viruses were added to rNY1682-TS1 to create rNY1682-TS2. Both rNY1682-TS1 and rNY1682-TS2 replicated efficiently at 30 °C and 33 °C. However, rNY1682-TS1 was partially restricted, and rNY1682-TS2 was completely restricted at 39 °C. Additionally, engineering the TS1 or TS2 mutations into a distantly related human seasonal H1N1 influenza A virus also resulted pronounced restriction of replication in vitro. Clinical symptoms and virus replication in the lungs of mice showed that although rNY1682-TS2 and the licensed FluMist^®-H1N1pdm LAIV that was used to combat the 2009/2010 pandemic were similarly attenuated, the rNY1682-TS2 was more protective upon challenge with a virulent mutant of pandemic H1N1 virus or a heterologous H1N1 (A/PR/8/1934) virus. This study demonstrates that engineering key temperature sensitive mutations (PB1-K391E, D581G, A661T; PB2-P112S, N265S, N556D, Y658H) into the genomes of influenza A viruses attenuates divergent human virus lineages and provides an alternative strategy for the generation of LAIVs.     

Inactivated and live, attenuated influenza vaccines protect mice against influenza: Streptococcus pyogenes super-infections

Mortality associated with influenza virus super-infections is frequently due to secondary bacterial complications. To date, super-infections with Streptococcus pyogenes have been studied less extensively than those associated with Streptococcus pneumoniae. This is significant because a vaccine for S. pyogenes is not clinically available, leaving vaccination against influenza virus as our only means for preventing these super-infections. In this study, we directly compared immunity induced by two types of influenza vaccine, either inactivated influenza virus (IIV) or live, attenuated influenza virus (LAIV), for the ability to prevent super-infections. Our data demonstrate that both IIV and LAIV vaccines induce similar levels of serum antibodies, and that LAIV alone induces IgA expression at mucosal surfaces. Upon super-infection, both vaccines have the ability to limit the induction of pro-inflammatory cytokines within the lung, including IFN-γ which has been shown to contribute to mortality in previous models of super-infection. Limiting expression of these pro-inflammatory cytokines within the lungs subsequently limits recruitment of macrophages and neutrophils to pulmonary surfaces, and ultimately protects both IIV- and LAIV-vaccinated mice from mortality. Despite their overall survival, both IIV- and LAIV-vaccinated mice demonstrated levels of bacteria within the lung tissue that are similar to those seen in unvaccinated mice. Thus, influenza virus:bacteria super-infections can be limited by vaccine-induced immunity against influenza virus, but the ability to prevent morbidity is not complete.     

Genetic stability of live attenuated vaccines against potentially pandemic influenza viruses

BackgroundEnsuring genetic stability is a prerequisite for live attenuated influenza vaccine (LAIV). This study describes the results of virus shedding and clinical isolates’ testing of Phase I clinical trials of Russian LAIVs against potentially pandemic influenza viruses in healthy adults.MethodsThree live attenuated vaccines against potentially pandemic influenza viruses, H2N2 LAIV, H5N2 LAIV and H7N3 LAIV, generated by classical reassortment in eggs, were studied. For each vaccine tested, subjects were randomly distributed into two groups to receive two doses of either LAIV or placebo at a 3:1 vaccine/placebo ratio. Nasal swabs were examined for vaccine virus shedding by culturing in eggs and by PCR. Vaccine isolates were tested for temperature sensitivity and cold-adaptation (ts/ca phenotypes) and for nucleotide sequence.ResultsThe majority of nasal wash positive specimens were detected on the first day following vaccination. PCR method demonstrated higher sensitivity than routine virus isolation in eggs. None of the placebo recipients had detectable vaccine virus replication.All viruses isolated from the immunized subjects retained the ts/ca phenotypic characteristics of the master donor virus (MDV) and were shown to preserve all attenuating mutations described for the MDV. These data suggest high level of vaccine virus genetic stability after replication in humans.During manufacture process, no additional mutations occurred in the genome of H2N2 LAIV. In contrast, one amino acid change in the HA of H7N3 LAIV and two additional mutations in the HA of H5N2 LAIV manufactured vaccine lot were detected, however, they did not affect their ts/ca phenotypes.ConclusionsOur clinical trials revealed phenotypic and genetic stability of the LAIV viruses recovered from the immunized volunteers. In addition, no vaccine virus was detected in the placebo groups indicating the lack of person-to-person transmission.LAIV TRIAL REGISTRATION at ClinicalTrials.gov: H7N3-NCT01511419; H5N2-NCT01719783; H2N2-NCT01982331.     

Comparative trials of live attenuated and detergent split influenza virus vaccines.

Comparative clinical trials of live attenuated and detergent-split subunit influenza virus vaccines were undertaken with 1048 volunteers in Western Australia. Volunteers were divided into three main groups, each of which received either live virus vaccine or a saline control administered intranasally, or subunit vaccine injected subcutaneously. No differences were recorded between the three groups in their post-vaccination symptoms. Serum samples were collected at various times up to 50 weeks after vaccination, and antibody titres were measured by haemagglutination-inhibition (HI) tests and, for 231 volunteers, by virus neutralization tests. The two vaccines were almost equivalent in inducing seroconversion in vaccinees with pre-trial HI titres of 96 or less, but the subunit vaccine stimulated a higher geometric mean HI antibody titre. The longevity of the HI antibody response was greater for the live virus vaccine. The height of the response and the longevity of neutralizing antibody were the same for both vaccines. Both vaccines provided a high degree of protection against epidemic A/England/42/72 influenza, and some protection against A/Port Chalmers/1/73 influenza.     

Shedding of live vaccine virus, comparative safety, and influenza-specific antibody responses after administration of live attenuated and inactivated trivalent influenza vaccines to HIV-infected children

HIV-infected children (N=243), >or=5 to <18 years old, receiving stable antiretroviral therapy, were stratified by immunologic status and randomly assigned to receive intranasal live attenuated influenza vaccine (LAIV) or intramuscular trivalent inactivated influenza vaccine (TIV). The safety profile after LAIV or TIV closely resembled the previously reported tolerability to these vaccines in children without HIV infection. Post-vaccination hemagglutination inhibition (HAI) antibody responses and shedding of LAIV virus were also similar, regardless of immunological stratum, to antibody responses and shedding previously reported for children without HIV infection. LAIV should be further evaluated for a role in immunizing HIV-infected children.     

Asthma exacerbations among asthmatic children receiving live attenuated versus inactivated influenza vaccines

ObjectiveTo investigate whether there is a difference in the risk of asthma exacerbations between children with pre-existing asthma who receive live attenuated influenza vaccine (LAIV) compared with inactivated influenza vaccine (IIV).Material and methodsWe identified IIV and LAIV immunizations occurring between July 1, 2007 and March 31, 2014 among Kaiser Permanente Northern California members aged 2 to <18 years with a history of asthma, and subsequent asthma exacerbations seen in the inpatient or Emergency Department (ED) setting. We calculated the ratio of the odds (OR) of an exacerbation being in the risk interval (1–14 days) versus the comparison interval (29–42 days) following immunization, separately for LAIV and IIV, and then examined whether the OR differed between children receiving LAIV and those receiving IIV (“difference-in-differences”).ResultsAmong 387,633 immunizations, 85% were IIV and 15% were LAIV. Children getting LAIV vs. IIV were less likely to have “current or recent, persistent” asthma (25% vs. 47%), and more likely to have “remote history” of asthma (47% vs. 25%). Among IIV-vaccinated asthmatic children, the OR of an inpatient/ED asthma exacerbation was 0.97 (95% CI: 0.82–1.15). Among LAIV-vaccinated asthmatic children the OR was 0.38 (95% CI: 0.17–0.90). In the difference-in-differences analysis, the odds of asthma exacerbation following LAIV were less than IIV (Ratio of ORs: 0.40, CI: 0.17–0.95, p value: 0.04).ConclusionAmong children ≥2 years old with asthma, we found no increased risk of asthma exacerbation following LAIV or IIV, and a decreased risk following LAIV compared to IIV.     

Immunogenicity and efficacy of the monovalent,trivalent and quadrivalent intranasal live attenuated influenza vaccines containing different pdmH1N1 strains

A ferret challenge study was conducted to address the efficacy of the egg-based and Madin-Darby canine kidney (MDCK)-based live attenuated influenza vaccine (LAIV) strains. Vaccines derived as 6:2 reassortants from the A/Leningrad/134/17/57 master donor strain and the HA and NA components from the A/California/07/2009 (A/Cal)- and A/Michigan/45/2015 (A/Mich)-like strains of type A H1N1 influenza virus were used in the study. Monovalent, trivalent and quadrivalent formulations of the LAIV containing either of the two H1N1 strains were analysed. A total of ten groups of six animals each were immunised intranasally (i.n.) with a single dose of 0.5-ml vaccine formulation or placebo and challenged on day 28 with the homologous wild-type A/Cal or A/Mich strain. Immune response post immunisation and virus replication post challenge were studied.Both the strains derived from embryonated eggs or MDCK cells, irrespective of the vaccine valency, were capable of rendering complete protection from virus replication in the lung. The A/Mich vaccine strain showed higher immune titres and efficacy than the A/Cal vaccine strain in all the vaccine formulations. The haemagglutination inhibition and virus neutralisation antibody titres were induced, and the reduction in the virus load in the respiratory tract was observed to be higher in animals treated with the monovalent formulation compared to the trivalent and quadrivalent formulations. Overall, it appears that the monovalent formulations render better protection from infection and would therefore be the best candidate during a pandemic.     

Efficacy of live attenuated vaccines against 2009 pandemic H1N1 influenza in ferrets

The advent of the H1N1 influenza pandemic (pH1N1) in 2009 triggered the rapid production of pandemic influenza vaccines, since seasonal influenza vaccines were expected and demonstrated not to provide significant cross-protection against the newly emerged pandemic virus. To increase vaccine production capacity and further evaluate the effectiveness of different candidate pandemic influenza vaccines, the World Health Organization stimulated the evaluation of different vaccination concepts including the use of live attenuated influenza vaccines (LAIVs). Therefore, we have immunized ferrets intranasally with a single dose of pH1N1-LAIV from different manufacturers. They all induced adequate serum HI antibody titers in the ferrets and protected them against intratracheal wild-type pH1N1 virus challenge: pH1N1 virus replication in the upper respiratory tract and lungs was reduced and no disease signs or severe broncho-interstitial pneumonia were observed in any of the vaccinated ferrets. These data together with the relatively efficient production process emphasize the potential of the LAIV concept for pandemic preparedness.     

Cold-attenuated live influenza vaccines, a risk-benefit assessment.

The principle of live attenuated influenza vaccines has been known for many decades. However, the pharmaceutical and clinical development according to current regulations, of modern live influenza vaccines based on cold adapted influenza viruses (CAIV) started only recently and these vaccines will most probably become an alternative within the next couple of years to licensed inactivated influenza vaccines that have been used routinely since the early 1940s. In contrast to contemporary trivalent inactivated influenza vaccines, which are administered intramuscularly, trivalent CAIV-based vaccines will be administered intranasally as a spray. Quality, safety and efficacy aspects related to CAIV vaccines as well as possible risks linked to the widespread use of these vaccines will be discussed in the following overview and compared to established influenza vaccines. Moreover, issues of practicality of CAIV vaccines focusing on the necessity of an annual update of influenza vaccines are addressed.     

WHO-supported comparative studies of attenuated live measles virus vaccines

Under the auspices of WHO a comparative study of attenuated live measles vaccines was made by teams in five countries: Canada, Czechoslovakia, Nigeria, Switzerland and Yugoslavia. The reactions and antibody responses to Enders'' Edmonston B vaccine with or without gamma-globulin and to the Schwarz, Beckenham and Milovanović vaccine were measured. In each trial Enders'' Edmonston B vaccine with gamma-globulin was used as a ”standard” vaccine, and in each trial there was also an unvaccinated control group. There were altogether 1685 children in the study—1317 vaccinated and 368 controls.     

A randomized, double-blind noninferiority study of quadrivalent live attenuated influenza vaccine in adults

Background

Trivalent seasonal influenza vaccines contain 2 A strains and 1 B strain. B strains of 2 antigenically distinct lineages, Yamagata and Victoria, have been co-circulating annually, and the B strain included in vaccines often has not been a lineage match to the major circulating strain. Thus, a vaccine containing B strains from both lineages could broaden protection against influenza. Quadrivalent live attenuated influenza vaccine (Q/LAIV) is an investigational 4-strain formulation of LAIV that contains 2 A strains, A/H1N1 and A/H3N2, and 2 B strains, 1 from each lineage.

Methods

A randomized, double-blind, active-controlled study of Q/LAIV was conducted in 1800 adults aged 18-49 years to compare the immunogenicity and safety of Q/LAIV to trivalent LAIV (T/LAIV). Subjects were randomized 4:1:1 to receive an intranasal dose of Q/LAIV (n = 1200) or 1 of 2 matching T/LAIV vaccines, each containing 1 of the B strains included in Q/LAIV (n = 600 total). The primary endpoint was the comparison of the post-vaccination strain-specific geometric mean titers (GMT) of hemagglutination inhibition antibody in Q/LAIV recipients to those in T/LAIV recipients, with immunologic noninferiority of Q/LAIV to be demonstrated if the upper bound of the 2-sided 95% confidence interval (CI) for the ratio of the GMTs [T/LAIV divided by Q/LAIV] was ≤1.5 for all strains.

Results and Conclusion

Q/LAIV met the criteria for noninferiority: the ratios of the GMTs for the A/H1N1, A/H3N2, B/Yamagata, and B/Victoria strains were 1.09 (95% CI, 1.01-1.18), 1.05 (95% CI, 0.96-1.14), 1.10 (95% CI, 0.97-1.25), and 0.92 (95% CI, 0.82-1.03), respectively. Solicited symptoms and adverse events were similar in the Q/LAIV and T/LAIV arms. Q/LAIV may confer increased protection against influenza by targeting B strains from both lineages.     

Cross-clade protective immune responses of NS1-truncated live attenuated H5N1 avian influenza vaccines

BackgroundH5N1 highly pathogenic avian influenza (HPAI) has raised global concern for causing huge economic losses in poultry industry, and an effective vaccine against HPAI is highly desirable. Live attenuated influenza vaccine with trunctated NS1 protein as a potential strategy will be extremely useful for improving immune efficacy.MethodsA series of H5N1 avian influenza virus reassortants harboring amino-terminal 48, 70, 73, and 99 aa in NS1 proteins, along with a modified low pathogenic HA protein was generated, and named as S-HALo/NS48, S-HALo/NS70, S-HALo/NS73, and S-HALo/NS99, respectively. In addition, their biological and immunological characteristics were further analyzed.ResultsThe viruses S-HALo/NS70, S-HALo/NS73, and S-HALo/NS99, but not S-HALo/NS48, had a comparable growth property with the full-length NS1 virus, S-HALo/NSFu. Mice and chickens studies demonstrated that the viruses with truncated NS1 protein were further attenuated when compared to the virus S-HALo/NSFu. Vaccination with the virus S-HALo/NS73 in chickens induced significant cross-protection against homologous clade 2.3.4 H5 virus and heterologous clade 7.2, 2.3.2.1, and 2.3.4.4 H5 viruses.ConclusionA 70-aa amino-terminal fragment of NS1 protein may be long enough for viral replication. The recombinant virus S-HALo/NS73 is a broad-spectrum live attenuated H5N1 avian influenza vaccine candidate in chickens.     

Immunogenicity and protective efficacy of cold-adapted X-31 live attenuated pre-pandemic H5N1 influenza vaccines

Despite global efforts to control influenza viruses, they have taken a heavy toll on human public health worldwide. Among particular threats is highly pathogenic avian H5N1 influenza virus (HPAI) due to not only its high mortality in humans but also possible human-to-human transmission either through reassortment with other human influenza viruses such as 2009 pandemic H1N1 influenza virus, or by genetic mutations. With the aim of developing effective vaccines against the H5N1 viruses, we generated two live attenuated H5N1 vaccine candidates against A/Indonesia/05/2005 (clade 2.1) and A/chicken/Korea/ES/2003 (clade 2.5) strains, in the genetic background of the cold-adapted donor strain of X-31. In mice, a single dose of immunization with each of the two vaccines was highly immunogenic inducing high titers of serum viral-neutralizing and hemagglutinin-inhibiting antibodies against the homologous H5N1 strain. Furthermore, significant levels of cross-clade antibody responses were induced by the vaccines, suggesting a broad-spectrum cross-reactivity against the heterologous H5N1 strains. The immunizations provided solid protections against heterologous lethal challenges with H5N2 virus, significantly reducing the morbidity and challenge virus replications in the respiratory tracts. The robustness of the antibody responses against both the homologous and heterologous strains, together with efficient protection against the lethal H5N2 challenge, strongly support the protection against wild type H5N1 infections. These results could serve as an experimental basis for the development of safe and effective H5N1 pre-pandemic vaccines while further addressing the biosecurity concerns associated with H5N1 HPAI.     

Safety of live, attenuated oral vaccines in HIV-infected Zambian adults: Oral vaccines in HIV

Background

Current recommendations are that HIV-infected persons should not be given live vaccines. We set out to assess potential toxicity of three live, attenuated oral vaccines (against rotavirus, typhoid and ETEC) in a phase 1 study.

Methods

Two commercially available oral vaccines against rotavirus (Rotarix) and typhoid (Vivotif) and one candidate vaccine against Enterotoxigenic Escherichia coli (ACAM2017) were given to HIV seropositive (n = 42) and HIV seronegative (n = 59) adults. Gastrointestinal symptoms were sought actively by weekly interview up to 1 month of vaccination. In rotavirus vaccine recipients, intestinal biopsies were collected by endoscopy and evaluated for expression of IL-8 and pro-inflammatory cytokines.

Results

No difference was observed between symptoms in HIV infected and HIV uninfected vaccinees, except for diarrhoea reported more than 7 days after the last dose of vaccine. If only diarrhoea episodes within 7 days of vaccination are included, diarrhoea was not more frequent in HIV seropositive than in HIV seronegative vaccinees (OR 6.7, 95% CI 1.2–67; P = 0.09). However, if later episodes of diarrhoea are included, a significant increase in diarrhoea was demonstrated (OR 5.3, 95% CI 0.98–53; P = 0.04). All episodes were mild and transient. IL-8 was slowly up-regulated over the week following vaccination (P = 0.02), but IL-β, IFNγ or TNFα were not.

Conclusions

No evidence was found of adverse events following administration of these three vaccines, except for late episodes of diarrhoea which may not be attributable to vaccination. Our data do not support the need for a prohibition on oral administration of live, attenuated vaccines to all HIV infected adults, though further work on severely immunocompromised adults and children are required.