Live Vaccine Vector Research Articles

Use of Leishmania major parasites expressing a recombinant Trypanosoma cruzi antigen as live vaccines against Chagas disease.

Trypanosoma cruzi is the protozoan parasite causing Chagas disease, a Neglected Tropical Disease that affects 8 million people and causes 12,000 deaths per year, primarily because of cardiac pathology. Effective vaccination for T. cruzi remains an elusive goal. The use of a live vaccine vector, especially one that mimics the pathogen target, may be superior to the use of recombinant protein or DNA vaccine formulations. We generated recombinant Leishmania major, a related trypanosomatid parasite, as a vaccine vehicle to express the immunogenic T. cruzi trans-sialidase (TS) antigen. The induction of T cell and antibody responses, as well as T. cruzi protective immunity generated by these vaccines were assessed in vivo. We demonstrate that mice inoculated with these recombinant TS-expressing L. major parasites mount T cell and antibody responses directed against TS and are protected against future T. cruzi infection. We also show that the partially attenuated dhfr-ts- CC1 L. major strain, previously found to induce protective immunity to virulent L. major infection without causing pathology, can also be engineered to express the TS antigen. This latter recombinant may represent a safe and effective option to explore for ultimate use in humans. Altogether, these data indicate that L. major can stably express a T. cruzi antigen and induce T. cruzi-specific protective immunity, warranting further investigation of attenuated Leishmania parasites as vaccine.

Immune gene expression analysis indicates the potential of a self-amplifying Covid-19 mRNA vaccine

Remarkable potency has been demonstrated for mRNA vaccines in reducing the global burden of the ongoing COVID-19 pandemic. An alternative form of the mRNA vaccine is the self-amplifying mRNA (sa-mRNA) vaccine, which encodes an alphavirus replicase that self-amplifies the full-length mRNA and SARS-CoV-2 spike (S) transgene. However, early-phase clinical trials of sa-mRNA COVID-19 vaccine candidates have questioned the potential of this platform to develop potent vaccines. We examined the immune gene response to a candidate sa-mRNA vaccine against COVID-19, ARCT-021, and compared our findings to the host response to other forms of vaccines. In blood samples from healthy volunteers that participated in a phase I/II clinical trial, greater induction of transcripts involved in Toll-like receptor (TLR) signalling, antigen presentation and complement activation at 1 day post-vaccination was associated with higher anti-S antibody titers. Conversely, transcripts involved in T-cell maturation at day 7 post-vaccination informed the magnitude of eventual S-specific T-cell responses. The transcriptomic signature for ARCT-021 vaccination strongly correlated with live viral vector vaccines, adjuvanted vaccines and BNT162b2 1 day post-vaccination. Moreover, the ARCT-021 signature correlated with day 7 YF17D live-attenuated vaccine transcriptomic responses. Altogether, our findings show that sa-mRNA vaccination induces innate immune responses that are associated with the development of adaptive immunity from other forms of vaccines, supporting further development of this vaccine platform for clinical application.

A recombinant baculovirus vector vaccine (BacMCP) against the infectious spleen and kidney necrosis virus (ISKNV).

The infectious spleen and kidney necrosis virus (ISKNV) is a highly lethal virus, which has brought significant losses to aquaculture. Therefore, a new vaccine against ISKNV with high efficiency, safety and convenience must be developed. While baculoviruses are more commonly used as protein expression systems for vaccine antigen production, this paper used baculovirus technology to develop a live-vector vaccine, BacMCP, which contains the coding sequence of the major capsid protein (MCP) (GenBank accession no. AF371960) of ISKNV and is driven by a CMV promoter. Real-time PCR and immunofluorescence showed that the MCP gene was successfully delivered to and expressed in fish cells and tissues inoculated with BacMCP. Immune-related gene (IgM, TGF-β, IL-1, IL-8, TNF-α) expression was induced in BacMCP-treated groups of largemouth bass compared with control groups. Specific antibodies could be detected in the serum of BacMCP injection-vaccinated largemouth bass by ELISA. After injection or immersion vaccination with BacMCP for 21 days, largemouth bass were infected with ISKNV. The immune effect of the injected immunization on fish in different sizes was evaluated. The vaccine efficacy of injection-vaccinated bass was 100% in small bass and 85.7% in large bass. The vaccine efficacy of immersion-vaccinated small bass was 77.3%. This study suggested that BacMCP can be used as a vector-based vaccine candidate to prevent the diseases caused by ISKNV infection.

Oral Immunization with Attenuated Salmonella Choleraesuis Expressing the P42 and P97 Antigens Protects Mice against Mycoplasma hyopneumoniae Challenge.

Mycoplasma hyopneumoniae (M. hyopneumoniae, Mhp) is the etiological agent of swine enzootic pneumonia (EP), which has been associated with considerable economic losses due to reduced daily weight gain and feed efficiency. Adhesion to the cilia is important for Mhp to colonize the respiratory epithelium. Therefore, a successful vaccine must induce broad Mhp-specific immune responses at the mucosal surface. Recombinant attenuated Salmonella strains are believed to act as powerful live vaccine vectors that are able to elicit mucosal immune responses against various pathogens. To develop efficacious and inexpensive vaccines against Mhp, the immune responses and protection induced by recombinant attenuated Salmonella vaccines based on the P42 and P97 antigens of Mhp were evaluated. In general, the oral inoculation of recombinant rSC0016(pS-P42) or rSC0016(pS-P97) resulted in strong mucosal immunity, cell-mediated immunity, and humoral immunity, which was a mixed Th1/Th2-type response. In addition, the levels of specific IL-4 and IFN-γ in the immunized mice were increased, and the proliferation of lymphocytes was also enhanced, confirming the production of a good cellular immune response. Finally, both vaccine candidate strains were able to improve the weight loss of mice after a challenge and reduce clinical symptoms, lung pathological damage, and the inflammatory cell infiltration. These results suggest that the delivery of protective antigens with recombinant attenuated Salmonella vectors may be an effective means by which to combat Mhp infection. IMPORTANCE Mhp is the main pathogen of porcine enzootic pneumonia, a highly infectious and economically significant respiratory disease that affects pigs of all ages. As the target tissue of Mhp infections are the mucosal sites of the respiratory tract, the induction of protective immunity at the mucosal tissues is the most efficient strategy by which to block disease transmission. Because the stimulation of mucosal immune responses is efficient, Salmonella-vector oral vaccines are expected to be especially useful against mucosal-invading pathogens. In this study, we expressed the immunogenic proteins of P42 and P97 with the attenuated Salmonella Choleraesuis vector rSC0016, thereby generating a low-cost and more effective vaccine candidate against Mhp by inducing significant mucosal, humoral and cellular immunity. Furthermore, rSC0016(pS-P42) effectively prevents Mhp-induced weight loss and the pulmonary inflammation of mice. Because of the effectiveness of rSC0016(pS-P42) against Mhp infection in mice, this novel vaccine candidate strain shows great potential for its use in the pig breeding industry.

Construction of constitutive expression of Eimeria tenella eukaryotic initiation factor U6L5H2 on the surface of Lactobacillus plantarum and evaluation of its immunoprotective efficiency against chicken coccidiosis

Lactobacillus strains exhibit preferable properties that make them attractive candidates for vaccine delivery systems because of their ability to regulate intestinal mucosal immunity in the body. To date, live Lactobacillus delivery vaccines reported for the defense against Eimeria tenella have been inducer-dependent systems whose applications are significantly limited due to their unattainable induction conditions in vivo. Here, a constitutive expression of Lactobacillus plantarum NC8 surface display system was constructed. Then, this system was used to prepare a live oral vaccine to constitutively express the E. tenella U6L5H2 (EtU6) protein on the NC8 surface and to evaluate its protective efficacy against E. tenella challenge in chickens. The results showed that the heterologous protein (EGFP or EtU6) was successfully expressed on the surface of L. plantarum NC8 without any inducer. The immunoprotection of EtU6 with constitutive expression in L. plantarum NC8 system (NC8/Pc-EtU6) was significantly stronger than that of EtU6 with induced expression of L. plantarum NC8 system (NC8/Pi-EtU6) (ACI: 168.28 vs. 152.74) as evidenced by increased body weight, decreased oocyst output and lesion scores. Furthermore, the constitutive system NC8/Pc-EtU6 produced higher levels of specific cecal SIgA, serum IgG, transcription of cytokines IFN-γ and IL-2, and lymphocyte proliferation than the induced system NC8/Pi-EtU6. These results indicate that, compared to the inducible system, the constitutive surface display system of L. plantarum has the advantages of continuously expressing antigens in vivo and stimulating the host immune system. It could be an ideal platform for vaccine expression. The live vector vaccine for coccidiosis constructed by this constitutive system greatly improves the application potential in chicken production and provides a novel platform for the prevention of coccidiosis in chickens.

Nonhuman Primates Are Protected against Marburg Virus Disease by Vaccination with a Vesicular Stomatitis Virus Vector-Based Vaccine Prepared under Conditions to Allow Advancement to Human Clinical Trials.

Vaccines are needed to disrupt or prevent continued outbreaks of filoviruses in humans across Western and Central Africa, including outbreaks of Marburg virus (MARV). As part of a filovirus vaccine product development plan, it is important to investigate dose response early in preclinical development to identify the dose range that may be optimal for safety, immunogenicity, and efficacy, and perhaps demonstrate that using lower doses is feasible, which will improve product access. To determine the efficacious dose range for a manufacturing-ready live recombinant vesicular stomatitis virus vaccine vector (rVSV∆G-MARV-GP) encoding the MARV glycoprotein (GP), a dose-range study was conducted in cynomolgus macaques. Results showed that a single intramuscular injection with as little as 200 plaque-forming units (PFUs) was 100% efficacious against lethality and prevented development of viremia and clinical pathologies associated with MARV Angola infection. Across the vaccine doses tested, there was nearly a 2000-fold range of anti-MARV glycoprotein (GP) serum IgG titers with seroconversion detectable even at the lowest doses. Virus-neutralizing serum antibodies also were detected in animals vaccinated with the higher vaccine doses indicating that vaccination induced functional antibodies, but that the assay was a less sensitive indicator of seroconversion. Collectively, the data indicates that a relatively wide range of anti-GP serum IgG titers are observed in animals that are protected from disease implying that seroconversion is positively associated with efficacy, but that more extensive immunologic analyses on samples collected from our study as well as future preclinical studies will be valuable in identifying additional immune responses correlated with protection that can serve as markers to monitor in human trials needed to generate data that can support vaccine licensure in the future.

Construction and Immunogenicity of a Recombinant Pseudorabies Virus Expressing SARS-CoV-2-S and SARS-CoV-2-N.

Coronavirus (CoV) is an important pathogen of humans and animals, which can infect humans or animals through the respiratory mucosal route. Syndrome coronavirus 2 (SARS-CoV-2) is quite similar to syndrome coronavirus (SARS-CoV) with the same receptor, angiotensin-converting enzyme 2 (ACE2). The S and N proteins are the most important protective antigens of the SARS-CoV-2. The S protein on the viral membrane mediates the virus attachment with the host cells, and the N protein is the most abundant expression during infection. In this study, the recombinant viruses expressing the S and N proteins of SARS-CoV-2 were successfully constructed by Red/ET recombinant technology using Pseudorabies virus (PRV) strain Bartha-K61 as a vector. Genetic stability and growth kinetics analysis showed that the recombinant viruses rPRV-SARS-CoV-2-S and rPRV-SARS-CoV-2-N had similar genetic stability and proliferation characteristics to the parental PRV. The immunoassay results showed that mice immunized with recombinant viruses could produce total IgG antibodies. Therefore, PRV is feasible and promising as a viral vector to express SARS-CoV-2-S and SARS-CoV-2-N genes. This study can provide a reference for future research on live vector vaccines for domestic animals, pets, and wild animals.

Construction and immunological evaluation of live vector vaccine based on attenuated Listeria monocytogenes vector against Vibrio parahaemolyticus infection

Vibrio parahaemolyticus is considered to be one of the most threatening aquatic pathogens in aquaculture. Therefore, the development of an effective vaccine is crucial to prevent and control it. In this report, we identified the target antigen VP2309 with strong immunogenicity from four outer membrane proteins of V. parahaemolyticus. Furthermore, the live vector vaccine strain EGD-e△actA/inlB-VP2309 based on attenuated Listeria monocytogenes (Lm) was successfully constructed, consisting of secretory signal peptide of P60 protein, strong promoter Plmo2196 of Lm, and VP2309. It was able to express and secrete VP2309 efficiently. However, its expression retarded the growth of the host strain. The results of immune protection evaluation showed that both EGD-e△actA/inlB and EGD-e△actA/inlB-VP2309 strains reduced the mortality of zebrafish infected with V. parahaemolyticus. However, the vaccine strain carrying the VP2309 did not show better immune protection than the empty vector. Thus, we found that the attenuated Lm vector could offer a certain immune protection to zebrafish infected with V. parahaemolyticus, and is a potential vaccine vector that can be used in fish vaccine.

Auxotrophic Mycobacterium bovis BCG: Updates and Perspectives.

Mycobacterium bovis BCG has been used for a century as the only licensed vaccine against tuberculosis. Owing to its strong adjuvant properties, BCG has also been employed as an oncological immunotherapeutic as well as a live vaccine vector against other pathogens. However, BCG vaccination has limited efficacy in protecting against adult forms of tuberculosis (TB), raises concerns about its safety in immunocompromised populations, compromises the diagnosis of TB through the tuberculin test and lacks predictability for successful antigen expression and immune responses to heterologous antigens. Together, these factors propelled the construction and evaluation of auxotrophic BCG strains. Auxotrophs of BCG have been developed from mutations in the genes required for their growth using different approaches and have shown the potential to provide a model to study M. tuberculosis, a more stable, safe, and effective alternative to BCG and a vector for the development of recombinant live vaccines, especially against HIV infection. In this review, we provide an overview of the strategies for developing and using the auxotrophic BCG strains in different scenarios.

Cytolysin A-mediated protein exportation efficiency and its role in enhancing the fitness of live recombinant Salmonella Typhi vaccine strain.

The genetic fusion of cytolysin A (clyA) to heterologous antigen expressed in live Salmonella vector demonstrated efficient translocation into periplasmic space and extracellular medium. Accumulating evidence has shown that clyA-mediated antigen delivery improved growth fitness and enhanced immunogenicity of live vector vaccine, but the factors influencing this protein exportation has not been investigated. In this study, Toxoplasma gondii antigen fused at C-terminal of clyA protein was expressed in live S. Typhi vector via both plasmid and chromosomal-based expressions. The bivalent strains showed comparable growth rates as monovalent strains, but in varies antigen exportation efficiency. ClyA-fusion antigen with positive charges was translocated to the extracellular spaces, whereas those with negative charges were retained in the cytoplasm. Furthermore, excessive cellular resources expenditure on antigen expression, especially antigen with larger size, could limit the clyA-fusion antigen exportation, resulting in undesirable metabolic burden that eventually affects the growth fitness. Altogether, the present work indicates potential linkage of factors mainly on antigen properties and expression platforms that may affect clyA-mediated antigen delivery to enhance the growth fitness of live vector strain.

Measles-based Zika vaccine induces long-term immunity and requires NS1 antibodies to protect the female reproductive tract

Zika virus (ZIKV) can cause devastating effects in the unborn fetus of pregnant women. To develop a candidate vaccine that can protect human fetuses, we generated a panel of live measles vaccine (MV) vectors expressing ZIKV-E and -NS1. Our MV-based ZIKV-E vaccine, MV-E2, protected mice from the non-lethal Zika Asian strain (PRVABC59) and the lethal African strain (MR766) challenge. Despite 100% survival of the MV-E2 mice, however, complete viral clearance was not achieved in the brain and reproductive tract of the lethally challenged mice. We then tested MV-based vaccines that expressed E and NS1 together or separately in two different vaccines. We observed complete clearance of ZIKV from the female reproductive tract and complete fetal protection in the lethal African challenge model in animals that received the dual antigen vaccines. Additionally, MV-E2 and MV-NS1, when administered together, induced durable plasma cell responses. Our findings suggest that NS1 antibodies are required to enhance the protection of ZIKV-E antibodies in the female reproductive tract.

An Engineered Outer Membrane-Defective Escherichia coli Secreting Protective Antigens against Streptococcus suis via the Twin-Arginine Translocation Pathway as a Vaccine.

Live bacterial vector vaccines are one of the most promising vaccine types and have the advantages of low cost, flexibility, and good safety. Meanwhile, protein secretion systems have been reported as useful tools to facilitate the release of heterologous antigen proteins from bacterial vectors. The twin-arginine translocation (Tat) system is an important protein export system that transports fully folded proteins in a signal peptide-dependent manner. In this study, we constructed a live vector vaccine using an engineered commensal Escherichia coli strain in which amiA and amiC genes were deleted, resulting in a leaky outer membrane that allows the release of periplasmic proteins to the extracellular environment. The protective antigen proteins SLY, enolase, and Sbp against Streptococcus suis were targeted to the Tat pathway by fusing a Tat signal peptide. Our results showed that by exploiting the Tat pathway and the outer membrane-defective E. coli strain, the antigen proteins were successfully secreted. The strains secreting the antigen proteins were used to vaccinate mice. After S. suis challenge, the vaccinated group showed significantly higher survival and milder clinical symptoms compared with the vector group. Further analysis showed that the mice in the vaccinated group had lower burdens of bacteria load and slighter pathological changes. Our study reports a novel live bacterial vector vaccine that uses the Tat system and provides a new alternative for developing S. suis vaccine.

Plasmodium falciparum Antigen Expression in Leishmania Parasite: A Way Forward for Live Attenuated Vaccine Development.

Live attenuated vaccines (LAVs) are among the most critical interventions in modern medicine and have already proven their potential to save millions of lives. LAVs are always explored as potential vaccine candidates since they induce an immune response, which is as good as the wild type pathogen. For parasitic diseases, the efficacy of LAVs is still under investigation and needs extensive research to mark their presence in the field. In malaria, live attenuated sporozoites have been evaluated for a vaccine against the liver stage. This vaccine approach is limited due to the highly cumbersome technique of sporozoite isolation and related relapse issues. We have developed a novel vaccine against malaria by expressing Plasmodium falciparum antigens in Leishmania donovani promastigotes. These hybrid, recombinant L. donovani parasites mimicking P. falciparum parasite antigens were analyzed for their anti-malarial efficacy in preclinical studies. We demonstrate the potential of Leishmania spp. parasites in developing an important live vector vaccine against malaria for the induction of protective immune responses. Herein, we describe a method to express malaria parasite antigens in L. donovani promastigotes and analyze its potential for a vaccine against malaria. This methodology can be extended to live, attenuated Leishmania promastigotes parasites to develop LAV against malaria.

A single intranasal dose of a live-attenuated parainfluenza virus-vectored SARS-CoV-2 vaccine is protective in hamsters

Single-dose vaccines with the ability to restrict SARS-CoV-2 replication in the respiratory tract are needed for all age groups, aiding efforts toward control of COVID-19. We developed a live intranasal vector vaccine for infants and children against COVID-19 based on replication-competent chimeric bovine/human parainfluenza virus type 3 (B/HPIV3) that express the native (S) or prefusion-stabilized (S-2P) SARS-CoV-2 S spike protein, the major protective and neutralization antigen of SARS-CoV-2. B/HPIV3/S and B/HPIV3/S-2P replicated as efficiently as B/HPIV3 invitro and stably expressed SARS-CoV-2 S. Prefusion stabilization increased S expression by B/HPIV3 invitro. In hamsters, a single intranasal dose of B/HPIV3/S-2P induced significantly higher titers compared to B/HPIV3/S of serum SARS-CoV-2-neutralizing antibodies (12-fold higher), serum IgA and IgG to SARS-CoV-2 S protein (5-fold and 13-fold), and IgG to the receptor binding domain (10-fold). Antibodies exhibited broad neutralizing activity against SARS-CoV-2 of lineages A, B.1.1.7, and B.1.351. Four weeks after immunization, hamsters were challenged intranasally with 104.5 50% tissue-culture infectious-dose (TCID50) of SARS-CoV-2. In B/HPIV3 empty vector-immunized hamsters, SARS-CoV-2 replicated to mean titers of 106.6 TCID50/g in lungs and 107 TCID50/g in nasal tissues and induced moderate weight loss. In B/HPIV3/S-immunized hamsters, SARS-CoV-2 challenge virus was reduced 20-fold in nasal tissues and undetectable in lungs. In B/HPIV3/S-2P-immunized hamsters, infectious challenge virus was undetectable in nasal tissues and lungs; B/HPIV3/S and B/HPIV3/S-2P completely protected against weight loss after SARS-CoV-2 challenge. B/HPIV3/S-2P is a promising vaccine candidate to protect infants and young children against HPIV3 and SARS-CoV-2.

T598 and T601 phosphorylation sites of canine parvovirus NS1 are crucial for viral replication and pathogenicity

Canine parvovirus-2 (CPV-2) is an important pathogen causing severe diseases in dogs and other wild carnivores. Phosphorylation of NS1 may be related to CPV-2 pathogenicity, but the exact mechanism is unclear. Here, we conducted parvovirus disease surveillance in Shaanxi Province of China and 51 fecal swabs were detected to be infected with CPV-2. The 7 CPV-2 strains were identified, all of which belonged to CPV-2c. The complete genome sequence of one of the strains (CPV-2c XY) was cloned into pKQLL plasmid to construct a full-length infectious clone plasmid pX-CPV-2c, which carried a genetic marker. The plasmid pX-CPV-2c was transfected into F81 cells for virus rescue. And the rescued virus, which was designed as X-CPV-2c, showed the similar biological property to parental CPV-2c XY in vitro and in vivo. We further constructed four NS1 phosphorylation site mutant strains (X-CPV-2cT584A, X-CPV-2cS592A, X-CPV-2cT598A/T601A and X-CPV-2cT617A) on the basis of X-CPV-2c. After the analysis and comparison of biological characteristics, the low pathogenic strain X-CPV-2cT598A/T601A was further screened out, which emphasized the importance of phosphorylation sites 598 T/601 T for the pathogenicity of CPV-2. Overall, our data indicated that T598 and T601, the C-terminal phosphorylation site of CPV-2 NS1, play important roles in viral pathogenicity and laid the foundation for the development of new attenuated live vaccine vectors.

Approaches to quality control, preclinical and clinical studies of live recombinant viral vector vaccines

At present, there are not much data on the clinical use of live recombinant viral vector vaccines. Characteristics of new vaccines should be factored into requirements/recommendations for quality control, preclinical and clinical studies of vaccines in order to enable further risk/benefit assessment. The aim of this study was to analyse current approaches to quality control, preclinical and clinical studies of live recombinant viral vector vaccines. The paper provides an overview of the licensed live viral vector vaccines and those at various stages of clinical trials. The authors analysed Russian, European, American, and Japanese guidelines related to quality issues, preclinical and clinical studies of live viral vector vaccines. The analysis demonstrated that the regulatory requirements for live recombinant viral vector vaccines include assessment of a detailed rationale for vaccine development, including information on the choice of the vector, the origin of the heterologous antigen gene(s), elements related to the transgene(s) expression, as well as assessment of the genetic and phenotypic stability of the recombinant virus, the risk of reversion to virulence or recombination with wild type strains, the potential for virus genome integration into the host cell chromosome, the pre-existing immunity to the vector, the intensity of the immune response elicited by the vector, and the reusability of the vector. The choice and number of applicable toxicological and pharmacological models will depend on these aspects. The results of the analysis of approaches to quality control, preclinical and clinical studies of live recombinant viral vector vaccines may be used in the development of Russian regulatory guidelines harmonised with the international norms and regulations.

Evaluation of an attenuated Listeria monocytogenes as a vaccine vector to control Helicobacter pylori infection

The increasing resistance of Helicobacter pylori (H. pylori) to antibiotics has limited the efficacy of antibiotic therapy in the treatment of H. pylori-associated gastric diseases. The vaccine as an alternative method is becoming a safe and effective way to address this problem. In previous studies, live vector vaccines have proved to be effective in controlling H. pylori infection. Attenuated Listeria monocytogenes (L. monocytogenes) is a potential candidate vector applied in clinical trials, which can deliver foreign antigens and induce a broad immune response. To further explore the effectiveness of L. monocytogenes as a vaccine vector against H. pylori, attenuated L. monocytogenes-based vaccine EGDeΔactA/inlB(EGDeAB)-MECU was constructed to secrete a multi-epitope chimeric antigen (MECU) containing multiple B cell epitopes from H. pylori antigens. EGDeAB-MECU could secrete MECU stably. After immunized by gavage and intravenous injection, both EGDeAB and EGDeAB-MECU could significantly decrease gastric H. pylori colonization and induce a high level of specific antibodies against H. pylori. In conclusion, attenuated L. monocytogenes had an immunotherapeutic effect on H. pylori-infected mice, indicating its further development as a promising candidate vaccine vector for the H. pylori vaccine.

Physiopathology and effectiveness of therapeutic vaccines against human papillomavirus

Human papillomavirus (HPV) is a well-known sexually transmitted disorder globally. Human papillomavirus (HPV) is the 3rd most common cancer that causes cervical carcinoma, and globally it accounts for 275,000 deaths every year. The load of HPV-associated abrasions can be lessened through vaccination. At present, three forms of prophylactic vaccines, Cervarix, Gadrasil, and Gardasil 9, are commercially accessible but all these prophylactic vaccines have not the ability to manage and control developed abrasions or infections. Therefore, a considerable amount of the population is not secured from HPV infectivity. Consequently, the development of therapeutic HPV vaccines is a crucial requirement of this era, for the treatment of persisting infections, and to stop the progression of HPV-associated cancers. Therapeutic vaccines are a developing trial approach. Because of the constitutive expression of E6 and E7 early genes in cancerous and pre-cancerous tissues, and their involvement in disturbance of the cell cycle, these are best targets for this therapeutic vaccine treatment. For the synthesis and development of therapeutic vaccines, various approaches have been examined comprising cell-based vaccines, peptide/protein-based vaccines, nucleic acid–based vaccines, and live-vector vaccines all proceeding towards clinical trials. This review emphasizes the development, progress, current status, and future perspective of several vaccines for the cure of HPV-related abrasions and cancers. This review also provides an insight to assess the effectiveness, safety, efficacy, and immunogenicity of therapeutic vaccines in the cure of patients infected with HPV-associated cervical cancer.

Establishment of Recombinant Eimeria acervulina Expressing Multi-Copies M2e Derived from Avian Influenza Virus H9N2.

The potential of Eimeria parasites as live vaccine vectors has been reported with successful genetic manipulation on several species like E. tenella, E. mitis and E. necatrix. Among seven Eimeria species infecting chickens, E. acervulina is a highly prevalent, moderately pathogenic species. Thus, it is valuable for the study of transfection and for use as a potential as vaccine vector. In this study, a plasmid containing expression cassette with enhanced yellow fluorescent protein (EYFP), red fluorescent protein (RFP) and 12 copies of extracellular domain of H9N2 avian influenza virus M2 (M2e) protein was used for the transfection. Nucleofected sporozoites were inoculated into birds through wing vein. Recombinant E. acervulina oocysts with 0.1% EYFP+ and RFP+ populations were collected from the feces of the inoculated birds. The fluorescent rate of transgenic parasites reached over 95% after nine successive propagations with a pyrimethamine selection in vivo and fluorescent-activated cell sorting (FACS) of progeny oocysts. The expression of M2e in the transgenic parasites (EaM2e) was confirmed by Western blot and its cytoplasm localization in sporozoites was displayed by an indirect immunofluorescent assay (IFA). Meanwhile, we found that the fecundity of EaM2e was equivalent to that of wild type E. acervulina (EaWT). Taken together, the stable transfection of E. acervulina was successfully established. Future studies will focus on whether transgenic E. acervulina can serve as a live vaccine vector.

Identification of four insertion sites for foreign genes in a pseudorabies virus vector

BackgroundPseudorabies virus (PRV) is a preferred vector for recombinant vaccine construction. Previously, we generated a TK&gE-deleted PRV (PRVΔTK&gE−AH02) based on a virulent PRV AH02LA strain. It was shown to be safe for 1-day-old piglets with maternal PRV antibodies and 4 ~ 5 week-old PRV antibody negative piglets and provide rapid and 100 % protection in weaned pigs against lethal challenge with the PRV variant strain. It suggests that PRVTK&gE−AH02 may be a promising live vaccine vector for construction of recombinant vaccine in pigs. However, insertion site, as a main factor, may affect foreign gene expression.ResultsIn this study, we constructed four recombinant PRV-S bacterial artificial chromosomes (BACs) carrying the same spike (S) expression cassette of a variant porcine epidemic diarrhea virus strain in different noncoding regions (UL11-10, UL35-36, UL46-27 or US2-1) from AH02LA BAC with TK, gE and gI deletion. The successful expression of S gene (UL11-10, UL35-36 and UL46-27) in recombinant viruses was confirmed by virus rescue, PCR, real-time PCR and indirect immunofluorescence. We observed higher S gene mRNA expression level in swine testicular cells infected with PRV-S(UL11-10)ΔTK/gE and PRV-S(UL35-36)ΔTK/gE compared to that of PRV-S(UL46-27)ΔTK/gE at 6 h post infection (P < 0.05). Moreover, at 12 h post infection, cells infected with PRV-S(UL11-10)ΔTK/gE exhibited higher S gene mRNA expression than those infected with PRV-S(UL35-36)ΔTK/gE (P = 0.097) and PRV-S(UL46-27)ΔTK/gE (P < 0.05). Recovered vectored mutant PRV-S (UL11-10, UL35-36 and UL46-27) exhibited similar growth kinetics to the parental virus (PRVΔTK&gE−AH02).ConclusionsThis study focuses on identification of suitable sites for insertion of foreign genes in PRV genome, which laids a foundation for future development of recombinant PRV vaccines.