Procedia in vaccinology最新文献

Hepatitis B vaccine (Engerix B) is a recombinant vaccine containing hepatitis B surface antigen (HBsAg) produced in Saccharomyces cerevisiae. Initially, this vaccine was released using specifications that required an in vivo potency assay in mice. This paper describes the move from the in vivo potency test toward in vitro potency assays: the in vitro Auszyme test (Abbott Laboratories) and the GSK Biologicals in-house test based upon the inhibition enzyme-linked immunosorbent assay (ELISA) principle. The challenges and difficulties during the development and introduction of this in vitro assay are presented from validation studies performed by GSK Biologicals, through regulatory acceptance, to the implementation of the alternative method by the European Directorate for the Quality of Medicines (EDQM) and HealthCare Control Authorities for Official batch release. Based upon our experience introducing an in vitro model for Engerix B potency assay, we offer suggestions to facilitate future introduction of in vitro assays.

Manufacturers of veterinary vaccines frequently incorporate animal-based batch release safety tests into their quality assurance monitoring protocols to meet their internal quality standards and to conform to government regulatory requirements. These tests are conducted by vaccinating target species animals or laboratory animals with a single dose or multiple doses of the test batch, and observing the vaccinated animals for signs of local or systemic adverse reactions. Manufacturers, standard-setting bodies, animal welfare advocacy groups, and regulatory agencies are actively investigating alternative methods to reduce their reliance on animal-based methods for batch release safety testing. Approaches which have been implemented or proposed include harmonizing technical requirements, developing in vitro tests, refining the existing animal tests, improving adverse reaction monitoring (vaccinovigilance), and improving manufacturing methods and quality controls to reduce batch-to-batch variability. An approach, known as the consistency approach, is increasingly being acknowledged as a potentially viable alternative to animal-based batch release tests for vaccines. This paper will provide an overview of currently utilized batch release safety tests for veterinary vaccines, the associated regulatory requirements, and some potentially acceptable alternative approaches for reducing, refining, and replacing the use of animals in these tests.

We have developed a novel tuberculosis (TB) vaccine; a combination of the DNA vaccines expressing mycobacterial heat shock protein 65 (HSP65) and interleukin 12 (IL-12) delivered by the hemagglutinating virus of Japan (HVJ)-envelope and –liposome (HSP65 + IL-12/HVJ). This vaccine provided remarkable protective efficacy in mouse model compared to the BCG. This vaccine also provided therapeutic efficacy against multi-drug resistant TB (MDR-TB) and extremely drug resistant TB (XDR-TB) in murine models. Furthermore, we extended our studies to a cynomolgus monkey model, which is currently the best animal model of human tuberculosis. This novel vaccine provided a higher level of the protective efficacy than BCG based upon the assessment of mortality. The BCG prime and HSP65 + IL-12/HVJ vaccine (boost) by the prime-boost method showed a synergistic prophylactic effect in the monkey. Furthermore, this vaccine exerted therapeutic efficacy (100% survival) and augmentation of immune responses in the TB-infected monkeys.HVJ-Envelope/HSP65 DNA + IL-12 DNA vaccine increased the body weight of TB-infected monkeys, improved the ESR, and augmented the immuneresponses (proliferation of PBL and IL-2 production). The enhancement of IL-2 production from monkeys treated with this vaccine was correlated with the therapeutic efficacy of the vaccine. These data indicate that our novel DNA vaccine might be useful against Mycobacterium tuberculosis including XDR-TB and MDR-TB for human therapeutic clinical trials.

Two series of experiments (in vivo and in vitro) were performed to determine protective level of antibodies to tickborne encephalitis virus (TBEV). We considered, basing on the data of neutralization tests (one TBEV doze (3 lg TCID50) with different titers of specific IgG antibodies (from 1:6400 to 1:12.5)) that antibody titer 1:400 in enzymelinked immunosorbent assay (ELISA) and 1:20 in neutralization test (NT) is a minimal threshold of protective level, and 1:100 – a lower threshold of immunological memory. During the second series of experiments on neutralization of different TBEV titers (from 1 to 8 lg TCID50) with immunoglobulin (antibody titers 1:400 and 1:3200) we obtained the additional evidence that high IgG titers (1:3200) actively neutralized TBEV, and antibody titer 1:400 was the lower threshold of protection. This characteristic was used to assess an immunological effectiveness of vaccine Encepur® adult. After three years of a full vaccination course we observed high values of a specific immunity, and after five years, in half of the cases revaccination was required.

NICEATM and ICCVAM convened an international workshop to review the state of the science of human and veterinary vaccine potency and safety testing methods, and to identify opportunities to advance new and improved methods that can further reduce, refine, and replace animal use (the 3Rs). Six topics were addressed in detail by speakers and workshop participants and are reported in a series of reports. This workshop report, the fifth in the series, addresses methods and strategies for human vaccine post-licensing safety testing that can reduce, refine, and replace animal use. It also provides recommendations for priority research and other activities necessary to advance the development and/or implementation of 3Rs methods for human vaccine post-licensing safety testing. Workshop participants agreed that future 3Rs activities should give highest priority to vaccine safety tests that (1) use the most animals per test and for which many vaccine lots are tested annually, (2) produce high variability and/or require frequent repeat tests, (3) are associated with severe animal pain and distress, and/or (4) involve nonhuman primates. Based on these criteria, safety tests for diphtheria, pertussis, oral polio, and tetanus vaccines were identified as the highest priorities. Recommended priority research, development, and validation activities included (1) expanding use of the transgenic mouse model for oral polio vaccine, and (2) developing alternatives to the monkey neurovirulence test for preclinical safety and lot release neurovirulence testing of mumps vaccines. Implementation of the workshop recommendations is expected to advance alternative in vitro methods for human vaccine post-licensing safety testing that will benefit animal welfare while ensuring continued production of safe human vaccines and protection of human and animal health.

Biologics are usually produced from live organisms, and the manufacturing process often involves a degree of natural variability. Characterization of biologics such as vaccines is inherently difficult due to the complex molecular structure of the antigens they contain and the presence of excipients such as preservatives and adjuvants that can interfere with testing. Therefore, each batch, lot, or serial produced must be tested before market release to ensure that the product complies with regulatory standards. This batch release testing emphasizes quality control of the final product and may be characterized by an extensive use of laboratory animals. The consistency approach is based upon the principle that the quality of a biologic is the result of the strict application of a quality system and consistent production. Subsequent batches are determined to be similar to clinically evaluated batches and therefore acceptable for release through the in-process testing that comprises this quality system. The European Centre for Validation of Alternative Methods (ECVAM) organized international workshops in 2006 and 2010 to discuss the consistency approach and its potential to reduce the number of animals used in testing of biological products. This paper provides an overview of these workshops.

Mucosal immunization with the conserved Shigella protein IpaD formulated with the two-component adjuvant IC31® provided protection in the murine lung model. Neither the recombinant IpaD alone nor IC31® with an irrelevant protein antigen was effective. IpaD-specific serum IgG and IgA levels obtained by IC31®-adjuvantation were comparable to those induced by known mucosal adjuvants.

NICEATM and ICCVAM convened an international workshop to review the state of the science of human and veterinary vaccine potency and safety testing methods and to identify opportunities to advance new and improved methods that can further reduce, refine, and replace animal use. Six topics were addressed in detail by speakers and workshop participants and are reported in a series of six reports. This workshop report, the second in the series, provides recommendations for current and future use of non-animal methods and strategies for veterinary vaccine potency testing. Workshop participants recommended that future efforts to replace animal use give priority to vaccines (1) that use large numbers of animals per test and for which many serials are produced annually, (2) that involve significant animal pain and distress during procedures, (3) for which the functional protective antigen has been identified, (4) that involve foreign animal/zoonotic organisms that are dangerous to humans, and (5) that involve pathogens that can be easily spread to wildlife populations. Vaccines identified as the highest priorities were those for rabies, Leptospira spp., Clostridium spp., Erysipelas, foreign animal diseases (FAD), poultry diseases, and fish diseases. Further research on the identification, purification, and characterization of vaccine protective antigens in veterinary vaccines was also identified as a priority. Workshop participants recommended priority research, development, and validation activities to address critical knowledge and data gaps, including opportunities to apply new science and technology. Recommendations included (1) investigations into the relative impact of various adjuvants on antigen quantification assays, (2) investigations into extraction methods that could be used for vaccines containing adjuvants that can interfere with antigen assays, and (3) review of the current status of rabies and tetanus human vaccine in vitro potency methods for their potential application to the corresponding veterinary vaccines. Workshop participants recommended enhanced international harmonization and cooperation and closer collaborations between human and veterinary researchers to expedite progress. Implementation of the workshop recommendations is expected to advance alternative in vitro methods for veterinary vaccine potency testing that will benefit animal welfare and replace animal use while ensuring continued protection of human and animal health.

The norovirus P particle, a subviral particle (∼20 nanometers in diameter) formed by 24 protrusion (P) domains of the norovirus capsid protein, is easily made, stable, and highly immunogenic and thus an excellent vaccine candidate against noroviruses. Each P domain has three surface loops that have been shown useful for antigen presentation. We have successfully inserted a number of small (5 aa) to large (238 aa) antigens into these loops without affecting P particle formation and production. Increased immune responses were demonstrated by improved antibody titers induced by the P particle presented antigens compared to free antigens. Significantly increased neutralization of virus and/or protection against influenza virus and rotavirus challenges have also been demonstrated in mice after immunization with chimeric P particle vaccines containing flu M2e and rotavirus VP8 antigens, compared to free M2e and VP8 antigens, respectively. The chimeric P particle-induced antibodies also blocked binding of noroviruslike particles (VLPs) to histo-blood group antigen (HBGA) receptors, indicating a potential dual vaccine against norovirus in addition to rotavirus and influenza virus. Taken together, the P particle appears to be an excellent platform for antigen presentation for vaccine development. The multiple surface loops and the large capacity of foreign antigen insertion suggest that this platform may have a wide application in vaccine development against different infectious diseases.

NICEATM and ICCVAM convened an international workshop to review the state of the science of human and veterinary vaccine potency and safety testing methods and to identify opportunities to advance new and improved methods that can further reduce, refine, and replace animal use. Topics were addressed in detail by speakers and workshop participants and are reported in a series of six reports. This workshop report, the third in the series, addresses methods and strategies for human vaccine potency testing that can refine animal use to lessen pain and distress, improve animal welfare, and reduce animal use. Workshop participants agreed that the following potency tests for human vaccines should have the highest priority for development of reduction and/or refinement methods: (1) potency tests for vaccines that are most commonly used, (2) potency tests that require the largest number of animals, (3) potency tests that cause severe animal pain and distress, (4) tests where the knowledge base of each antigen is advanced, and (5) potency tests for which alternative methods already exist or are in development. Based on these criteria, the highest-priority human vaccines were identified as diphtheria and tetanus vaccines, pertussis vaccines (whole cell and acellular), rabies vaccine, anthrax vaccine, and complex combination vaccines (containing diphtheria, tetanus, and pertussis together with other antigens such as IPV, Hib, and HepB). For successful implementation of reduction and refinement alternatives, further research is required into the development and broader use of humane endpoints, serological potency methods, and approaches that would reduce the number of animals used in currently approved potency assays. Because the workshop focused on both human and veterinary vaccines, workshop participants also recommended that human vaccine potency testing methods for rabies and tetanus be reviewed for their potential application to the corresponding veterinary vaccines. Participants agreed that achieving broader acceptance and use of alternative methods, requires that the general principles and procedures for the validation of alternative methods for vaccines be standardized/harmonized internationally. The research, development, validation, and harmonization activities recommended at this workshop are expected to lead to new reduction and/or refinement of animal use in human vaccine potency testing methods and more widespread adoption of existing methods that can reduce animal use and improve animal welfare while ensuring the continued safety and efficacy of human vaccines.