New Vaccine Technologies for Rapid Response against Emerging, Reemerging Infections and Biological Threats: Lessons from COVID-19 for Future

Vaccination is the most effective method to prevent dangerous infectious diseases and save lives. The expansion of human communication, the rapid spread of emerging infections worldwide, and the creation of dangerous pandemics like COVID-19 is worrying. On the other hand, with the emergence of new technologies such as genetic engineering of microorganisms, genome editing, and synthetic biology, the possibility of abusing these tools for illegal use is the next concern. In this situation, the need for rapid vaccination technologies and programs was given special importance. Recently, new vaccine platforms such as viral vector and mRNA vaccines have shown great promise that they can be used to prepare and protect human lives against dangerous infections. One of the most important factors for vaccination is the rapid development and approval of vaccines. In this review, we have given a perspective view of new vaccine technologies to rapidly develop vaccine s to combat emerging infections and the biodefence against biological criminals. © 2023 The Author(s).

Viral vector vaccines: efforts to develop vaccines against emerging infectious diseases

Vaccines are one of the most effective means of preventing viral infections. Since Edward Jenner invented the world's first vaccine in 1796, against smallpox, various types of vaccine have been developed, including inactivated vaccines, attenuated live vaccines, recombinant protein vaccines, viral vector vaccines and nucleic acid vaccines. Viral vector vaccines and nucleic acid vaccines （mRNA vaccines and DNA vaccines） have been developed most recently. In these vaccines, genes encoding viral proteins that serve as antigens are introduced into the body. The viral vector is an excellent vaccine delivery system that efficiently delivers antigen genes to target cells, and has been utilized for vaccine development against a variety of emerging infectious diseases, including AIDS, malaria, Ebola hemorrhagic fever, dengue fever, and most recently COVID-19. Here, we provide an overview of viral vector vaccines and discuss recent efforts to develop vaccines against emerging infectious diseases.Alternate :抄録ウイルス性感染症を予防するうえで、ワクチンは最も有効な手段の一つである。1976年、エドワード・ジェンナーが世界初のワクチンである種痘を発明して以来、さまざまなウイルス性感染症に対して、不活化ワクチン、弱毒生ワクチン、組換えタンパクワクチン、ウイルスベクターワクチン、核酸ワクチンなど、多様なプラットフォームに基づくワクチン開発が進められてきた。本稿では、数あるワクチンプラットフォームの中から、ウイルスベクターワクチンに着目して、いくつかの例をあげて概説するとともに、近年、国際的な問題となっている新興感染症に対するワクチン開発などの取り組みについても述べる。

The Coming of Age of Nucleic Acid Vaccines during COVID-19.

In the 21st century, several emergent viruses have posed a global threat. Each pathogen has emphasized the value of rapid and scalable vaccine development programs. The ongoing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has made the importance of such efforts especially clear. New biotechnological advances in vaccinology allow for recent advances that provide only the nucleic acid building blocks of an antigen, eliminating many safety concerns. During the COVID-19 pandemic, these DNA and RNA vaccines have facilitated the development and deployment of vaccines at an unprecedented pace. This success was attributable at least in part to broader shifts in scientific research relative to prior epidemics: the genome of SARS-CoV-2 was available as early as January 2020, facilitating global efforts in the development of DNA and RNA vaccines within 2 weeks of the international community becoming aware of the new viral threat. Additionally, these technologies that were previously only theoretical are not only safe but also highly efficacious. Although historically a slow process, the rapid development of vaccines during the COVID-19 crisis reveals a major shift in vaccine technologies. Here, we provide historical context for the emergence of these paradigm-shifting vaccines. We describe several DNA and RNA vaccines in terms of their efficacy, safety, and approval status. We also discuss patterns in worldwide distribution. The advances made since early 2020 provide an exceptional illustration of how rapidly vaccine development technology has advanced in the last 2 decades in particular and suggest a new era in vaccines against emerging pathogens. IMPORTANCE The SARS-CoV-2 pandemic has caused untold damage globally, presenting unusual demands on but also unique opportunities for vaccine development. The development, production, and distribution of vaccines are imperative to saving lives, preventing severe illness, and reducing the economic and social burdens caused by the COVID-19 pandemic. Although vaccine technologies that provide the DNA or RNA sequence of an antigen had never previously been approved for use in humans, they have played a major role in the management of SARS-CoV-2. In this review, we discuss the history of these vaccines and how they have been applied to SARS-CoV-2. Additionally, given that the evolution of new SARS-CoV-2 variants continues to present a significant challenge in 2022, these vaccines remain an important and evolving tool in the biomedical response to the pandemic.

Improved DNA Vaccine Delivery with Needle-Free Injection Systems.

DNA vaccines have inherent advantages compared to other vaccine types, including safety, rapid design and construction, ease and speed to manufacture, and thermostability. However, a major drawback of candidate DNA vaccines delivered by needle and syringe is the poor immunogenicity associated with inefficient cellular uptake of the DNA. This uptake is essential because the target vaccine antigen is produced within cells and then presented to the immune system. Multiple techniques have been employed to boost the immunogenicity and protective efficacy of DNA vaccines, including physical delivery methods, molecular and traditional adjuvants, and genetic sequence enhancements. Needle-free injection systems (NFIS) are an attractive alternative due to the induction of potent immunogenicity, enhanced protective efficacy, and elimination of needles. These advantages led to a milestone achievement in the field with the approval for Restricted Use in Emergency Situation of a DNA vaccine against COVID-19, delivered exclusively with NFIS. In this review, we discuss physical delivery methods for DNA vaccines with an emphasis on commercially available NFIS and their resulting safety, immunogenic effectiveness, and protective efficacy. As is discussed, prophylactic DNA vaccines delivered by NFIS tend to induce non-inferior immunogenicity to electroporation and enhanced responses compared to needle and syringe.

Immunogenicity of the DNA/Protein Combined Vaccine against COVID-19.

During the COVID-19 pandemic, the development of prophylactic vaccines, including those based on new platforms, became highly relevant. One such platform is the creation of vaccines combining DNA and protein components in one construct. For the creation of DNA vaccine, we chose the full-length spike protein (S) of the SARS-CoV-2 virus and used the recombinant receptor-binding domain (RBD) of the S protein produced in CHO-K1 cells as a protein component. The immunogenicity of the developed combined vaccine and its individual components was compared and the contribution of each component to the induction of the immune response was analyzed. The combined DNA/protein vaccine possesses the advantages of both underlying approaches and is capable of inducing both humoral (similar to subunit vaccines) and cellular (similar to DNA vaccines) immunity.

DNA and mRNA Vaccines for Chronic Viral Infections and Cancer: Rationale, Mechanisms, and Progress.

Interest in the capabilities of nucleic acid vaccines, (DNA and mRNA vaccines) for both prophylactic and therapeutic uses have greatly increased following the successful deployment of two mRNA and, on a more limited scale, one DNA vaccine for COVID-19. In addition to targeting other pathogens for prophylactic vaccines, efforts are also being made towards using them for therapies for chronic infections and cancer. An examination of past and current successes for such therapies using other technologies with an emphasis on the immunological mechanisms will be provided followed by an assessment of the relevant characteristics of DNA and mRNA vaccines to predict their utility for therapies for chronic viral infections and cancer. Efforts and progress for these targets will be described.

Optimization of In Vivo Electroporation Conditions and Delivery of DNA Vaccine Encoding SARS-CoV-2 RBD Using the Determined Protocol.

Vaccination against SARS-CoV-2 and other viral infections requires safe, effective, and inexpensive vaccines that can be rapidly developed. DNA vaccines are candidates that meet these criteria, but one of their drawbacks is their relatively weak immunogenicity. Electroporation (EP) is an effective way to enhance the immunogenicity of DNA vaccines, but because of the different configurations of the devices that are used for EP, it is necessary to carefully select the conditions of the procedure, including characteristics such as voltage, current strength, number of pulses, etc. In this study, we determined the optimal parameters for delivery DNA vaccine by electroporation using the BEX CO device. BALB/c mice were used as a model. Plasmid DNA phMGFP was intramuscular (I/M) injected into the quadriceps muscle of the left hind leg of animals using insulin syringes, followed by EP. As a result of the experiments, the following EP parameters were determined: direct and reverse polarity rectangular DC current in three pulses, 12 V voltage for 30 ms and 950 ms intervals, with a current limit of 45 mA. The selected protocol induced a low level of injury and provided a high level of GFP expression. The chosen protocol was used to evaluate the immunogenicity of the DNA vaccine encoding the receptor-binding domain (RBD) of the SARS-CoV-2 protein (pVAXrbd) injected by EP. It was shown that the delivery of pVAXrbd via EP significantly enhanced both specific humoral and cellular immune responses compared to the intramuscular injection of the DNA vaccine.

Efficient Delivery of DNA Using Lipid Nanoparticles.

DNA vaccination has been extensively studied as a promising strategy for tumor treatment. Despite the efforts, the therapeutic efficacy of DNA vaccines has been limited by their intrinsic poor cellular internalization. Electroporation, which is based on the application of a controlled electric field to enhance DNA penetration into cells, has been the method of choice to produce acceptable levels of gene transfer in vivo. However, this method may cause cell damage or rupture, non-specific targeting, and even degradation of pDNA. Skin irritation, muscle contractions, pain, alterations in skin structure, and irreversible cell damage have been frequently reported. To overcome these limitations, in this work, we use a microfluidic platform to generate DNA-loaded lipid nanoparticles (LNPs) which are then characterized by a combination of dynamic light scattering (DLS), synchrotron small-angle X-ray scattering (SAXS), and transmission electron microscopy (TEM). Despite the clinical successes obtained by LNPs for mRNA and siRNA delivery, little is known about LNPs encapsulating bulkier DNA molecules, the clinical application of which remains challenging. For in vitro screening, LNPs were administered to human embryonic kidney 293 (HEK-293) and Chinese hamster ovary (CHO) cell lines and ranked for their transfection efficiency (TE) and cytotoxicity. The LNP formulation exhibiting the highest TE and the lowest cytotoxicity was then tested for the delivery of the DNA vaccine pVAX-hECTM targeting the human neoantigen HER2, an oncoprotein overexpressed in several cancer types. Using fluorescence-activated cell sorting (FACS), immunofluorescence assays and fluorescence confocal microscopy (FCS), we proved that pVAX-hECTM-loaded LNPs produce massive expression of the HER2 antigen on the cell membrane of HEK-293 cells. Our results provide new insights into the structure-activity relationship of DNA-loaded LNPs and pave the way for the access of this gene delivery technology to preclinical studies.

Immune response induced in rodents by anti-CoVid19 plasmid DNA vaccine via pyro-drive jet injector inoculation.

There is an urgent need to stop the coronavirus disease 2019 (COVID-19) pandemic through the development of efficient and safe vaccination methods. Over the short term, plasmid DNA vaccines can be developed as they are molecularly stable, thus facilitating easy transport and storage. pVAX1-SARS-CoV2-co was designed for the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) S protein. The antibodies produced led to immunoreactions against the S protein, an anti-receptor-binding-domain, and a neutralizing action of the pVAX1-SARS-CoV2-co, as previously confirmed. To promote the efficacy of the pVAX1-SARS-CoV2-co vaccine a pyro-drive jet injector (PJI) was used. An intradermally adjusted PJI demonstrated that the pVAX1-SARS-CoV2-co vaccine injection caused a high production of anti-S protein antibodies, triggered immunoreactions, and neutralized the actions against SARS-CoV-2. A high-dose pVAX1-SARS-CoV2-co intradermal injection using PJI did not cause any serious disorders in the rat model. A viral challenge confirmed that intradermally immunized mice were potently protected from COVID-19. A pVAX1-SARS-CoV2-co intradermal injection using PJI is a safe and promising vaccination method for overcoming the COVID-19 pandemic.

Adenovirus-vectored SARS-CoV-2 vaccine expressing S1-N fusion protein.

Additional COVID-19 vaccines that are safe and immunogenic are needed for global vaccine equity. Here, we developed a recombinant type 5 adenovirus vector encoding for the SARS-CoV-2 S1 subunit antigen and nucleocapsid as a fusion protein (Ad5.SARS-CoV-2-S1N). A single subcutaneous immunization with Ad5.SARS-CoV-2-S1N induced a similar humoral response, along with a significantly higher S1-specific cellular response, as a recombinant type 5 adenovirus vector encoding for S1 alone (Ad5.SARS-CoV-2-S1). Immunogenicity was improved by homologous prime-boost vaccination, and further improved through intramuscular heterologous prime-boost vaccination using subunit recombinant S1 protein. Priming with low dose (1 × 1010 v.p.) of Ad5.SARS-CoV-2-S1N and boosting with either wild-type recombinant rS1 or B.1.351 recombinant rS1 induced a robust neutralizing response, which was sustained against Beta and Gamma SARS-CoV-2 variants. This novel Ad5-vectored SARS-CoV-2 vaccine candidate showed promising immunogenicity in mice and supports the further development of COVID-19-based vaccines incorporating the nucleoprotein as a target antigen.

Immunogenicity of genetic vaccine based on recombinant hepatitis B Core antigen and B cell multi-epitopes of Porcine epidemic diarrhea virus spike glycoprotein

In present study, in order to develop a new and effective porcine epidemic diarrhea virus (PEDV)vaccine, three B cell epitopes and the truncated S1 gene of PEDV spike protein were combined and inserted into the immunodominant region of the HBcAg. Then the constructed recombinant plasmid HBcAg-PE was transformed to E. coli BL21 (DE3) for expression. After purification and identification by Western-blot, the expressed recombinant proteins HBPE were injected into BALB/c mice as vaccine antigen with different doses through intramuscular injection and its immune effect were preliminary evaluated. The results showed that the recombinant proteins HBPE was expressed as precipitation form and it could reacted specifically with PEDV-positive serum after purification and renaturation. Besides, the RH could induce anti-PEDV specific antibodies and the related Thl and Th2 cytokines in mice. The above results indicate that the recombinant compound epitope antigen of PEDV was successfully constructed. and its immunogenicity as a new vaccine candidate was evaluated in the mice in this study. The results of this study provided a new idea for the development of PEDV genetic engineering vaccine in the future.

DNA-Vaccine-Induced Immune Response Correlates with Lower Viral SARS-CoV-2 Titers in a Ferret Model.

The COVID-19 pandemic is entering a new era with the approval of many SARS-CoV-2 vaccines. In spite of the restoration of an almost normal way of life thanks to the immune protection elicited by these innovative vaccines, we are still facing high viral circulation, with a significant number of deaths. To further explore alternative vaccination platforms, we developed COVID-eVax-a genetic vaccine based on plasmid DNA encoding the RBD domain of the SARS-CoV-2 spike protein. Here, we describe the correlation between immune responses and the evolution of viral infection in ferrets infected with the live virus. We demonstrate COVID-eVax immunogenicity as means of antibody response and, above all, a significant T-cell response, thus proving the critical role of T-cell immunity, in addition to the neutralizing antibody activity, in controlling viral spread.

A Review on DNA Vaccines in Pre-Clinical Trials Against SARS-CoV-2

COVID 19 Pandemic is caused by the viral pathogen severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Scientific fraternity worldwide swiftly developed various types of vaccines for the prevention and as mitigation measures for curbing the pandemic. Traditional inactivated vaccines, mRNA vaccines (protein subunits such as spike proteins), and viral vector vaccines (non-replicating vectors with protein subunits) have been approved by World Health Organisation (WHO) for emergency use. The emergence of many mutated variants has been a worrying factor in the fight against the pandemic. There has been continuous research in the quest for more therapeutics, especially vaccines to curb and stop the pandemic. According to WHO, there are 194 vaccines in pre-clinical trials belonging to various types out of which sixteen is DNA vaccines. In this review, we have discussed the advantages and disadvantages of the DNA vaccines for Covid-19. This article tried to explore the available information on DNA vaccines and their current status against Covid – 19 which are in pre-clinical trials. © 2022, Editorial board of Journal of Experimental Biology and Agricultural Sciences. All rights reserved.

Translating a Thin-Film Rehydration Method to Microfluidics for the Preparation of a SARS-CoV-2 DNA Vaccine: When Manufacturing Method Matters.

Previous investigations conducted on a liposomal formulation for a SARS-CoV-2 DNA vaccine manufactured using the thin-film layer rehydration method showed promising immunogenicity results in mice. The adaptation of the liposomal formulation to a scalable and reproducible method of manufacture is necessary to continue the investigation of this vaccine candidate. Microfluidics manufacture shows high potential in method translation. The physicochemical characterization of the blank liposomes produced by thin-film layer rehydration or microfluidics were shown to be comparable. However, a difference in lipid nanostructure in the bilayer resulted in a significant difference in the hydration of the thin-film liposomes, ultimately altering their complexation behavior. A study on the complexation of liposomes with the DNA vaccine at various N/P ratios showed different sizes and Zeta-potential values between the two formulations. This difference in the complexation behavior resulted in distinct immunogenicity profiles in mice. The thin-film layer rehydration-manufactured liposomes induced a significantly higher response compared to the microfluidics-manufactured samples. The nanostructural analysis of the two samples revealed the critical importance of understanding the differences between the two formulations that resulted in the different immunogenicity in mice.

Identification of T cell epitopes in BALB/c mice immunized with COVID-19 vaccine

Objective: To identify the T cell epitopes of the COVID-19 vaccine carrying SARS-CoV-2 S, N and M genes in BALB/c mice.

COVID-19 vaccine and hematopoietic stem cell transplantation

In the pandemic era of coronavirus disease 2019 (COVID-19), vaccines have been developed and approved to control the pandemic that might reduce the COVID-19 mortality. Transplant recipients are among the high-risk groups and are more susceptible to COVID-19 infection. According to the available data about COVID-19 vaccines, some platform technologies include vector-based, inactivated, protein subunit, virus-like particles, mRNA, and DNA vaccines (1). There are several guidelines about vaccination in immunocompromised individuals for both non-live- and live vaccines. However, there are still limited evidence-based data about COVID-19 vaccines in the hematopoietic stem cell transplantation (HSCT), and establishing a proper recommendation for vaccination in these patients would be challenging (2, 3). Transplant recipients may have shown lesser responses to the vaccines compared with the general population, and it is unknown to what extent the vaccine is effective in this group of patients. Also, in many countries, the vaccination schedule is not adjustable by the patients or physicians, and selecting a particular time window for the best efficacy of immunization is impossible. In this regard, the main concern in the patients treated with immunosuppressive drugs is not worsening symptoms and disease following vaccination. The most critical issue is determining the best time for vaccination to increase its efficacy. Here are some considerations about vector-based, inactivated, and mRNA- nanoparticle vaccines, but most evidence is not based on the results of cohort or clinical trial studies. Before HSCT, patients could receive the COVID-19 vaccine if they are not already immunosuppressed. According to evidence about other inactivated vaccines, such as the influenza vaccine, the interval to start the conditioning regimen could be considered 2 - 4 weeks following the vaccination (4). In autologous HSCT patients, COVID-19 vaccination can be considered 1 - 3 months after transplantation if there has been a community outbreak. If acquiring or transmitting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was well controlled, vaccination could be withheld after six months of transplantation. In the current pandemic, COVID-19 vaccination in allogeneic HSCT patients could be considered at least three months after transplantation. If transmission of SARS-CoV-2 was controlled, vaccination could be withheld after six months of transplantation (4-6). Vaccination of patients with chronic graft versus host disease (cGVHD) receiving less than 20 mg/day prednisolone (or equivalent) for less than 2 weeks, can be considered similar to the HSCT recipients with no GVHD (5). Vaccines in HSCT recipients with active SARS-CoV-2 infection are not effective thus, receiving the vaccine is not recommended. If an HSCT recipient has received the COVID-19 vaccine before HSCT, re-vaccination after transplantation is suggested (6). The administration of the vaccine is considered when the immune system acquired functional competence. Transplant donation should not be delayed due to the vaccination of the donor to protect the patients in case the transplant is urgent (6). It was reported that recipients who have received anti-B cell antibodies might get the vaccine at 3 - 6 months after the administration and four weeks before the next course of B cell-depleting therapy. If this time window was not possible, vaccination can be regarded under B-cell depleting therapy, considering a suboptimal response to the vaccine (7). It should be noted that the effects of rituximab may last for six months or even a year. Also, the decision to order vaccines following the use of rituximab should be based on the level of immunoglobulins and CD19. There is no strong evidence for the short duration of vaccination following the use of rituximab (such as 3 to 6 months). However, despite the low efficacy of the vaccine in such conditions, it is recommended to get the vaccine whenever available. It is reasonable that recipients who have received therapy with antithy

Molecular biology and genetic studies in combating the new coronavirus (COVID-19) outbreak

The new type of coronavirus (SARS-CoV-2), which is transmitted from person to person and causes Severe Acute Respiratory Distress Syndrome (SARS), emerged in Wuhan, China in December 2019. The definitive diagnosis of the coronavirus, which is transmitted from person to person through droplets, is given through PCR-based tests. The continuation of the COVID-19 pandemic has made it necessary to develop an effective vaccine against SARS-CoV-2. Vaccines developed against COVID-19 can be classified as inactivated/live virus vaccines, recombinant protein vaccines/vectored vaccines or RNA/DNA vaccines. This review aims to give information about the molecular structure and genetic features of SARSCoV- 2 virus, laboratory diagnostic methods, potential therapeutic drugs and vaccine studies.

A Nucleic Acid-Based Orthopoxvirus Vaccine Targeting the Vaccinia Virus L1, A27, B5, and A33 Proteins Protects Rabbits against Lethal Rabbitpox Virus Aerosol Challenge

In the age of COVID, nucleic acid vaccines have garnered much attention, at least in part, because of the simplicity of construction, production, and flexibility to adjust and adapt to an evolving outbreak. Orthopoxviruses remain a threat on multiple fronts, especially as emerging zoonoses. In response, we developed a DNA vaccine, termed 4pox, that protected nonhuman primates against monkeypox virus (MPXV)-induced severe disease. Here, we examined the protective efficacy of the 4pox DNA vaccine delivered by intramuscular (i.m.) electroporation (EP) in rabbits challenged with aerosolized rabbitpox virus (RPXV), a model that recapitulates the respiratory route of exposure and low dose associated with natural smallpox exposure in humans. We found that 4pox-vaccinated rabbits developed immunogen-specific antibodies, including neutralizing antibodies, and did not develop any clinical disease, indicating protection against aerosolized RPXV. In contrast, unvaccinated animals developed significant signs of disease, including lesions, and were euthanized. These findings demonstrate that an unformulated, nonadjuvanted DNA vaccine delivered i.m. can protect against an aerosol exposure. IMPORTANCE The eradication of smallpox and subsequent cessation of vaccination have left a majority of the population susceptible to variola virus or other emerging poxviruses. This is exemplified by human monkeypox, as evidenced by the increase in reported endemic and imported cases over the past decades. Therefore, a malleable vaccine technology that can be mass produced and does not require complex conditions for distribution and storage is sought. Herein, we show that a DNA vaccine, in the absence of a specialized formulation or adjuvant, can protect against a lethal aerosol insult of rabbitpox virus.

Path to Success and Future Impact of Nucleic Acid Vaccines: DNA and mRNA

The rapid development of mRNA vaccines for COVID-19 has both astonished the world and raised concerns about their safety, perhaps because many people do not realize the decades’ long efforts for nucleic acid vaccines, both mRNA and DNA vaccines, including the licensure of several veterinary DNA vaccines. This manuscript traces the milestones for nucleic acid vaccine research and development (R&D), with a focus on the immune and safety issues they both raised and answered. The characteristics of the two entities are compared, demonstrating the similarities and differences between them, the advantages and disadvantages, which might lead toward using one or the other technology for different indications. In addition, as the SARS-CoV-2 pandemic has once again highlighted the importance of One Health, that is, the interactions between animal and human pathogens, focus will also be given to how DNA vaccine utilization and studies both in large domestic animals and in wildlife pave the way for more integrated approaches for vaccines to respond quickly to, and prevent, the global impacts of emerging diseases.