Plasmid DNA Vaccine Research Articles

STING Nanoagonist Boosts Antitumor Immunity of Therapeutic DNA Vaccines.

Therapeutic DNA cancer vaccines can stimulate specific immune responses against cancer antigens but often induce suboptimal therapeutic responses. Here, we demonstrate that a manganese-doped silica nanoparticle STING agonist (MSNA) enhances the immune response of plasmid DNA vaccines, promoting the activation and migration of distinct subsets of dendritic cell (DC) and improving antitumor immunity in three animal models. MSNA coadministered with an α-fetoprotein (AFP) encoded plasmid DNA (AFP-DNA) elicited significantly higher AFP-specific CD8 T cell responses than free AFP-DNA. Animals immunized with MSNA-AFP-DNA remained tumor-free in an AFP expressing hepatocellular carcinoma challenge model. MSNA combined with a DNA plasmid encoding the human papillomavirus type 16 oncoproteins E6 and E7 induced potent E7-specific CD8 T cell responses, preventing the growth of E7-expressing solid TC-1 tumors and promoting the shrinkage of E7-expressing skin grafts. These findings together demonstrate that coadministration of MSNA can improve the efficacy of therapeutic DNA vaccines targeting cancer-specific antigens.

Revolutionizing Veterinary Health with Viral Vector-Based Vaccines.

Vaccines signify one of the economical and reasonable means to prevent and eradicate the important infectious diseases. Conventional vaccines like live attenuated and inactivated vaccines comprise of whole pathogen either in attenuated or killed form. While, new generation vaccines have been designed to elicit immune response by genetically modifying only the nucleic acid portion of that pathogen. These new generation therapeutics include mRNA vaccines, DNA plasmid vaccines, chimeric vaccines and recombinant viral vector-based vaccines. Nucleic acid based vaccines use genetic material itself thus, they are highly stable and potent in nature to induce long-lasting immune response. Amongst these novel vaccine platforms, viral vector-based vaccines is one such emerging field which has proven to be extremely effective and potent. Nowadays, veterinary medicine has also accepted this innovative vectored vaccine platform to develop an effective control strategy against certain important viral diseases of animals. Viral vector-based vaccine uses various DNA and RNA viruses of human or animal origin to carry an immunogenic transgene of target pathogen. These vaccines enhance both humoral and cell mediated immune response without use of any accessory immune-stimulants. Till today, several viruses have been modified to be characterized as vaccine vectors. Currently, large number of research programs are going on to develop vectored vaccines and novel viral vector for veterinary use. In the present review, different kinds of viral vectored vaccines having veterinary importance have been discussed.

Lipid nanoparticle-encapsulated DNA vaccine confers protection against swine and human-origin H1N1 influenza viruses.

Swine influenza A virus (IAV) is widespread and causes significant economic losses to the swine industry. Moreover, bidirectional transmission of IAV between swine and humans commonly occurs. Once introduced into the swine population, human-origin IAV often reassorts with endemic swine IAV, resulting in reassortant viruses. Thus, it is imperative to develop a vaccine that is not only effective against IAV strains endemic in swine but also capable of preventing the spillover of human-origin IAV. In this study, we developed a lipid nanoparticle-encapsulated DNA plasmid vaccine (LNP-DNA) that demonstrates efficacy against both swine- and human-origin H1N1 viruses. The LNP-DNA vaccines are non-infectious and non-viable, meeting the criteria to serve as a vaccine platform for rapidly updating vaccines. Collectively, this LNP-DNA vaccine approach holds great potential for alleviating the impact of IAV on the swine industry and preventing the emergence of reassortant IAV strains.

Lipid Nanoparticles Outperform Electroporation in Delivering Therapeutic HPV DNA Vaccines.

Therapeutic HPV vaccines that induce potent HPV-specific cellular immunity and eliminate pre-existing infections remain elusive. Among various candidates under development, those based on DNA constructs are considered promising because of their safety profile, stability, and efficacy. However, the use of electroporation (EP) as a main delivery method for such vaccines is notorious for adverse effects like pain and potentially irreversible muscle damage. Moreover, the requirement for specialized equipment adds to the complexity and cost of clinical applications. As an alternative to EP, lipid nanoparticles (LNPs) that are already commercially available for delivering mRNA and siRNA vaccines are likely to be feasible. Here, we have compared three intramuscular delivery systems in a preclinical setting. In terms of HPV-specific cellular immune responses, mice receiving therapeutic HPV DNA vaccines encapsulated with LNP demonstrated superior outcomes when compared to EP administration, while the naked plasmid vaccine showed negligible responses, as expected. In addition, SM-102 LNP M exhibited the most promising results in delivering candidate DNA vaccines. Thus, LNP proves to be a feasible delivery method in vivo, offering improved immunogenicity over traditional approaches.

Module-combinatorial design and screening of multifunctional polymers based on polyaspartic acid for DNA delivery

It is crucial to develop non-viral gene vectors that can efficiently and safely transfect plasmid DNA into cells. Low transfection efficiency and high cytotoxicity of cationic polymers hinder their application as gene carriers. Modification of cationic polymers has emerged as an attractive strategy for efficient and safe nucleic acids delivery. In this study, a simple and rapid method is developed to synthesize a series of multifunctional polymers by utilizing biodegradable polyaspartic acid as the backbone and modifying it with three modules. This one-component polymer possesses capabilities for nucleic acid condensation, cellular uptake, and endosomal escape. Polymers containing imidazole, triazole, or pyridine group exhibited promising transfection activity. Substituted with dodecylamine or 2-hexyldecan-1-amine enhance cellular uptake and subsequent transfection. Furthermore, the influence of ionizable amine side chains on gene delivery is investigated. Two optimal polymers, combined with the avian encephalomyelitis virus (AEV) plasmid vaccine, induced robust specific antibody responses and cellular immune responses in mice and chickens. Through module-combination design and screening of polyaspartamide polymers, this study presents a paradigm for the development of gene delivery vectors.

Preclinical Development of a Novel Epitope-based DNA Vaccine Candidate against SARS-CoV-2 and Evaluation of Immunogenicity in BALB/c Mice.

The protective efficacies of current licensed vaccines againstCOVID-19 have significantly reduced as a result of SARS-CoV-2 variants of concern(VOCs) which carried multiple mutations in the Spike (S) protein. Considering thatthese vaccines were developed based on the S protein of the original SARS-CoV-2Wuhan strain, we designed a recombinant plasmid DNA vaccine based on highlyconserved and immunogenic B and T cell epitopes against SARS-CoV-2 Wuhan strainand the Omicron VOC. Literature mining and bioinformatics were used toidentify 6 immunogenic peptides from conserved regions of the SARS-CoV-2 S andmembrane (M) proteins. Nucleotide sequences encoding these peptides representinghighly conserved B and T cell epitopes were cloned into a pVAX1 vector to form thepVAX1/S2-6EHGFP recombinant DNA plasmid vaccine. The DNA vaccine wasintranasally or intramuscularly administered to BALB/c mice and evaluations ofhumoral and cellular immune responses were performed. The intramuscularadministration of pVAX1/S2-6EHGFP was associated with a significantly higherpercentage of CD8+ T cells expressing IFN-γ when compared with the empty vectorand PBS controls. Intramuscular or intranasal administrations of pVAX1/S2-6EHGFPresulted in robust IgG antibody responses. Sera from mice intramuscularly immunizedwith pVAX1/S2-6EHGFP were found to elicit neutralizing antibodies capable ofSARS-CoV-2 Omicron variant with the ACE2 cell surface receptor. This studydemonstrated that the DNA vaccine construct encoding highly conserved immunogenicB and T cell epitopes was capable of eliciting potent humoral and cellular immuneresponses in mice.

Combined biolistic and cell penetrating peptide delivery for the development of scalable intradermal DNA vaccines

Physical-based gene delivery via biolistic methods (such as the Helios gene gun) involve precipitation of nucleic acids onto microparticles and direct transfection through cell membranes of exposed tissue (e.g. skin) by high velocity acceleration. The glycosaminoglycan (GAG)-binding enhanced transduction (GET) system exploits novel fusion peptides consisting of cell-binding, nucleic acid condensing, and cell-penetrating domains, which enable enhanced transfection across multiple cell types. In this study, we combined chemical (GET) and physical (gene gun) DNA delivery systems, and hypothesized the combination would generate enhanced distribution and effective uptake in cells not initially transfected by biolistic penetration. Physicochemical characterization, optimization of bullet contents and transfection experiments in vitro in cell monolayers and engineered tissue demonstrated these formulations transfected efficiently, including DC2.4 dendritic cells. We incorporated these formulations into a biolistic format for gene gun by forming fireable dry bullets obtained via lyophilization (freeze drying). This system is simple and with enhanced scalability compared to conventional methods to generate bullets. Flushed GET bullet contents retained their ability to mediate transfection (17-fold greater and 13-fold greater reporter gene expression than standard spermidine bullets in the absence and presence of serum, respectively). Fired GET bullets in vitro (in cells and collagen gels) and in vivo (mice) showed increased reporter gene transfection compared to untreated controls, whilst maintaining cell viability in vitro and having no obvious toxicity in vivo. Lastly, a SARS-CoV-2 plasmid DNA vaccine with spike (S) protein-receptor binding domain (S-RBD) was delivered by gene gun using GET bullets. Specific T cell and antibody responses comparable to the conventional system were generated. The non-physical and physical combination of GET‑gold-DNA carriers using gene gun shows potential as an alternative DNA delivery method that is scalable for mass deployable vaccination and intradermal gene delivery.

Transformed Salmonella typhimurium SL7207/pcDNA-CCOL2A1 as an orally administered DNA vaccine

The use of attenuated bacteria for oral delivery of DNA vaccines is a recent innovation. We designed and constructed the naked plasmid DNA vaccine pcDNA-CCOL2A1, which effectively prevented and treated a rheumatoid arthritis model by inducing immunotolerance. We aimed to ensure a reliable, controllable dosage of this oral DNA vaccine preparation and establish its stability. We transformed pcDNA-CCOL2A1 via electroporation into attenuated Salmonella typhimurium SL7207. A resistant plate assay confirmed the successful construction of the transformed strain of the SL7207/pcDNA-CCOL2A1 oral DNA vaccine. We verified its identification and stability in vitro and in vivo. Significant differences were observed in the characteristics of the transformed and blank SL7207 strains. No electrophoretic restriction patterns or direct sequencing signals were observed in the original extract of the transformed strain. However, target gene bands and sequence signals were successfully detected after PCR amplification. CCOL2A1 expression was detected in the ilea of BALB/c mice that were orally administered SL7207/pcDNA-CCOL2A1. The pcDNA-CCOL2A1 plasmid of the transformed strain was retained under the resistant condition, and the transformed strain remained stable at 4 °C for 100 days. The concentration of the strain harboring the pcDNA-CCOL2A1 plasmid was stable at 109 CFU/mL after 6–8 h of incubation. The results demonstrated that the transformed strain SL7207/pcDNA-CCOL2A1 can be expressed in vivo, has good stability, and may be used to prepare the oral DNA vaccine pcDNA-CCOL2A1 with a stable, controllable dosage and the capacity to provide oral immunization. This vehicle can effectively combine both oral immunotolerance and DNA vaccination.

Isolation and Characterization of Neutralizing Monoclonal Antibodies from a Large Panel of Murine Antibodies against RBD of the SARS-CoV-2 Spike Protein.

The COVID-19 pandemic, once a global crisis, is now largely under control, a testament to the extraordinary global efforts involving vaccination and public health measures. However, the relentless evolution of SARS-CoV-2, leading to the emergence of new variants, continues to underscore the importance of remaining vigilant and adaptable. Monoclonal antibodies (mAbs) have stood out as a powerful and immediate therapeutic response to COVID-19. Despite the success of mAbs, the evolution of SARS-CoV-2 continues to pose challenges and the available antibodies are no longer effective. New variants require the ongoing development of effective antibodies. In the present study, we describe the generation and characterization of neutralizing mAbs against the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein by combining plasmid DNA and recombinant protein vaccination. By integrating genetic immunization for rapid antibody production and the potent immune stimulation enabled by protein vaccination, we produced a rich pool of antibodies, each with unique binding and neutralizing specificities, tested with the ELISA, BLI and FACS assays and the pseudovirus assay, respectively. Here, we present a panel of mAbs effective against the SARS-CoV-2 variants up to Omicron BA.1 and BA.5, with the flexibility to target emerging variants. This approach ensures the preparedness principle is in place to address SARS-CoV-2 actual and future infections.

Phase 2 trial of a DNA vaccine (pTVG-HP) and nivolumab in patients with castration-sensitive non-metastatic (M0) prostate cancer

PurposeWe have previously reported that a plasmid DNA vaccine encoding prostatic acid phosphatase (pTVG-HP) had greater clinical activity when given in combination with pembrolizumab to patients with metastatic, castration-resistant prostate...

A novel SARS-CoV-2 Beta RBD DNA vaccine directly targeted to antigen-presenting cells induces strong humoral and T cell responses

Throughout the COVID-19 pandemic, several variants of concern (VoC) of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have evolved, affecting the efficacy of the approved COVID-19 vaccines. To address the need for vaccines that induce strong and persistent cross-reactive neutralizing antibodies and T cell responses, we developed a prophylactic SARS-CoV-2 vaccine candidate based on our easily and rapidly adaptable plasmid DNA vaccine platform. The vaccine candidate, referred to here as VB2129, encodes a protein homodimer consisting of the receptor binding domain (RBD) from lineage B.1.351 (Beta) of SARS-CoV-2, a VoC with a severe immune profile, linked to a targeting unit (human LD78β/CCL3L1) that binds chemokine receptors on antigen-presenting cells (APCs) and a dimerization unit (derived from the hinge and CH3 exons of human IgG3). Immunogenicity studies in mice demonstrated that the APC-targeted vaccine induced strong antibody responses to both homologous Beta RBD and heterologous RBDs derived from Wuhan, Alpha, Gamma, Delta, and Omicron BA.1 variants, as well as cross-neutralizing antibodies against these VoC. Overall, preclinical data justify the exploration of VB2129 as a potential booster vaccine that induces broader antibody- and T cell-based protection against current and future SARS-CoV-2 VoC.

Targeting neoantigens to APC-surface molecules improves the immunogenicity and anti-tumor efficacy of a DNA cancer vaccine.

Tumor-specific mutations generate neoepitopes unique to the cancer that can be recognized by the immune system, making them appealing targets for therapeutic cancer vaccines. Since the vast majority of tumor mutations are patient-specific, it is crucial for cancer vaccine designs to be compatible with individualized treatment strategies. Plasmid DNA vaccines have substantiated the immunogenicity and tumor eradication capacity of cancer neoepitopes in preclinical models. Moreover, early clinical trials evaluating personalized neoepitope vaccines have indicated favorable safety profiles and demonstrated their ability to elicit specific immune responses toward the vaccine neoepitopes. By fusing in silico predicted neoepitopes to molecules with affinity for receptors on the surface of APCs, such as chemokine (C-C motif) ligand 19 (CCL19), we designed an APC-targeting cancer vaccine and evaluated their ability to induce T-cell responses and anti-tumor efficacy in the BALB/c syngeneic preclinical tumor model. In this study, we demonstrate how the addition of an antigen-presenting cell (APC) binding molecule to DNA-encoded cancer neoepitopes improves neoepitope-specific T-cell responses and the anti-tumor efficacy of plasmid DNA vaccines. Dose-response evaluation and longitudinal analysis of neoepitope-specific T-cell responses indicate that combining APC-binding molecules with the delivery of personalized tumor antigens holds the potential to improve the clinical efficacy of therapeutic DNA cancer vaccines. Our findings indicate the potential of the APC-targeting strategy to enhance personalized DNA cancer vaccines while acknowledging the need for further research to investigate its molecular mechanism of action and to translate the preclinical results into effective treatments for cancer patients.

The Cooperation of IL-29 and PLGA Nanoparticles Improves the Protective Immunity of the gD-1 DNA Vaccine Against Herpes Simplex Virus Type 1 in Mice

ABSTRACT In clinical practice, the low immunogenicity and low stability of the DNA plasmid vaccine candidates are two significant shortcomings in their application against infectious diseases. To overcome these two disadvantages, the plasmid expressing IL-29 (pIL-29) as a genetic adjuvant and polylactic-co-glycolic acid (PLGA) as a non-viral delivery system were used, respectively. In this study, the pIL-29 encapsulated in PLGA nanoparticles (nanoIL-29) and the pgD1 encapsulated in PLGA nanoparticles (nanoVac) were simultaneously applied to boost immunologic responses against HSV-1. We generated spherical nanoparticles with encapsulation efficiency of 75 ± 5% and sustained the release of plasmids from them. Then, Balb/c mice were subcutaneously immunized twice with nanoVac+nanoIL-29, Vac+IL-29, nanoVac, Vac, nanoIL-29, and/or IL-29 in addition to negative and positive control groups. Cellular immunity was evaluated via lymphocyte proliferation assay, cytotoxicity test, and IFN-γ, IL-4, and IL-2 measurements. Mice were also challenged with 50X LD50 of HSV-1. The nanoVac+nanoIL-29 candidate vaccine efficiently enhances CTL and Th1-immune responses and increases the survival rates by 100% in mice vaccinated by co-administration of nanoVac and nanoIL-29 against the HSV-1 challenge. The newly proposed vaccine is worth studying in further clinical trials, because it could effectively improve cellular immune responses and protected mice against HSV-1.

A DNA vaccine (EG95-PT1/2/3-IL2) encoding multi-epitope antigen and IL-2 provokes efficient and long-term immunity to echinococcosis

Echinococcosis is a highly prevalent global zoonosis, and vaccines are required. The commercial vaccine based on a protein-based subunit (EG95), however, is limited by its insufficient cellular immunity, a short protection period, and limited prevention against novel mutant strains. Herein, we applied bioinformatics to develop a DNA vaccine (pEG95-IL2) expressing both multi-epitope-based antigens (EG95-PT1/2/3) and an IL-2 adjuvant to regulate T cell differentiation and memory cell response. EG95-PT1/2/3 was screened with hierarchical structure prediction from the epitope conformation of B cells with high confidence across various species to guarantee immunogenicity. Importantly, cationic arginine-rich lipid nanoparticles (RNP) were utilized as a delivery vehicle to form lipoplexes that had a transfection efficiency of nearly two orders of magnitude greater than that of commercial reagents (Lipofectamine 2000 and polyethyleneimine) with both immune and nonimmune cells (DC2.4 and L929 cells, respectively). RNP/pEG95-IL2 lipoplexes displayed a robust and long-term antigen expression, as well as adjuvant effects during the immunization. Consequently, intramuscular injection of RNP/pEG95-IL2 elicited similar humoral immune responses and significantly greater cellular responses in mice when compared with those of the commercial vaccine. In addition, the inoculation protocol of RNP/pEG95-IL2 with sequential booster further strengthens cellular immunity in comparison with the homologous booster. Those findings provide a promising strategy for improving plasmid vaccine efficacy.

A Tailored Approach to Leishmaniases Vaccination: Comparative Evaluation of the Efficacy and Cross-Protection Capacity of DNA vs. Peptide-Based Vaccines in a Murine Model.

Zoonotic leishmaniases are a worldwide public health problem for which the development of effective vaccines remains a challenge. A vaccine against leishmaniases must be safe and affordable and should induce cross-protection against the different disease-causing species. In this context, the DNA vaccine pHisAK70 has been demonstrated to induce, in a murine model, a resistant phenotype against L. major, L. infantum, and L. amazonensis. Moreover, a chimeric multiepitope peptide, HisDTC, has been obtained by in silico analysis from the histone proteins encoded in the DNA vaccine and has showed its ability to activate a potent CD4+ and CD8+ T-cell protective immune response in mice against L. infantum infection. In the present study, we evaluated the plasmid DNA vaccine pHisAK70 in comparison with the peptide HisDTC (with and without saponin) against L. major and L. infantum infection. Our preliminary results showed that both formulations were able to induce a potent cellular response leading to a decrease in parasite load against L. infantum. In addition, the DNA candidate was able to induce better lesion control in mice against L. major. These preliminary results indicate that both strategies are potentially effective candidates for leishmaniases control. Furthermore, it is important to carry out such comparative studies to elucidate which vaccine candidates are the most appropriate for further development.

Exploring the Potential of a Genome-Reduced Escherichia coli Strain for Plasmid DNA Production.

The global demand for nucleic acid-based vaccines, including plasmid DNA (pDNA) and mRNA vaccines, needs efficient production platforms. However, conventional hosts for plasmid production have encountered challenges related to sequence integrity due to the presence of insertion sequences (ISs). In this study, we explored the potential of a genome-reduced Escherichia coli as a host for pDNA production. This strain had been constructed by removing approximately 23% of the genome which were unessential genes, including the genomic unstable elements. Moreover, the strain exhibits an elevated level of NADPH, a coenzyme known to increase plasmid production according to a mathematical model. We hypothesized that the combination of genome reduction and the abundance of NADPH would significantly enhance pDNA production capabilities. Remarkably, our results confirmed a three-fold increase in pDNA production compared to the widely employed DH5α strain. Furthermore, the genome-reduced strain exhibited heightened sensitivity to various antibiotics, bolstering its potential for large scale industrial pDNA production. These findings suggest the genome-reduced E. coli as an exciting candidate for revolutionizing the pDNA industry, offering unprecedented efficiency and productivity.

Advancement in Polymer-based Carrier for DNA Vaccine.

A novel strategy that has the potential to solve the drawbacks of the present conventional vaccines is the development of DNA vaccines. DNA vaccines offer a versatile and adaptable platform for treating a wide variety of diseases, as immunization targets may be easily adjusted by altering the gene sequences encoded in the plasmid DNA delivered. Due to their ability to elicit both humoral and cellular immune responses, their stability, and the ease with which they may be produced, plasmid DNA vaccines are quickly becoming the vaccine of choice, they are frequently safer than conventional vaccinations. Despite the highly encouraging outcomes of ongoing clinical trials, these vaccines' immunogenicity is compromised by a few factors. The use of various vaccine delivery techniques, the use of various polymer-based carriers, and the use of adjuvants are some of the several approaches that might be examined to better the immunogenicity of DNA vaccines made from plasmids. These advancements taken together might allow plasmid DNA vaccines to be successfully used in clinical settings.

A phase II study of concurrent WOKVAC vaccination with neoadjuvant chemotherapy and HER2-targeted monoclonal antibody therapy.

TPS636 Background: HER2+ breast cancer (BCa) is an aggressive breast cancer subtype. The treatment of Stage I-III HER2+ BCa with neoadjuvant chemotherapy and HER2 monoclonal antibodies (trastuzumab and pertuzumab) induces a pathologic complete response in 43-67% of patients. Residual HER2+ disease (i.e. residual cancer burden) after neoadjuvant therapy is associated with a significantly higher risk of recurrence. Evidence supports that HER2 immunity is progressively lost in HER2 oncogenesis and that restoration of HER2 specific CD4+ Th1 immunity is associated with a lower risk of recurrence. Trastuzumab can partially restore HER2 specific immunity, but only in a minority of patients. We have demonstrated that HER2 specific vaccination can augment HER2 specific immunity. While taxane chemotherapy can deplete immunosuppressive cells (MDSCs) 13 – 17 days after treatment creating an optimal time for vaccination. The WOKVAC vaccine is a plasmid vaccine encoding Th1 selective epitopes from HER2 and two other high-risk cancer antigens IGFBP-2 and IGF-1R. The Phase I trial of this vaccine reported that it is safe and all doses induced antigen specific Type I immune responses. We hypothesize that addition of WOKVAC to neoadjuvant chemo/HER2 targeted therapy will induce a significant increase in the number of patients with Th1 CD4+ and CD8+ T cells after neoadjuvant vaccine/chemo/HER2 therapy. Methods: Trial Design: Phase II single arm trial of WOKVAC vaccination with concurrent neoadjuvant taxane based chemotherapy, trastuzumab, and pertuzumab in patients with Stage I-III HER2+ (HR+/-) breast cancer. Enrolled patients will receive WOKVAC vaccine 150mcg on day 13 of cycles 1-3. WOKVAC vaccination may be given with either TCHP or THP. A maximum of 16 patients will be enrolled in this study. Objectives: The primary objective is to evaluate whether concurrent WOKVAC vaccine with chemotherapy and HER2 antibody therapy can significantly increase T-bet+ CD4+ and CD8+ TIL. Secondary objectives are to determine: (1) whether concurrent WOKVAC with neoadjuvant chemo/HER2 therapy is safe, (2) if the magnitude of vaccine induced antigen specific Th1 immunity after neoadjuvant therapy is associated with pathologic response. Statistical Methods: (1) In order to detect a 30% increase in the number of patients with Tbet+ CD4+ and CD8+ TIL in the enrolled patients with 80% power and a one-side alpha of 0.05, our enrollment goal for this study will be 16 patients. (2) Safety will be assessed per CTCAE v5.0, Benchmarks for safety needed to advance this trial concept to a Phase III study will be a grade 3 toxicity rate of ≤13% and a grade 4 toxicity rate of ≤ 7%. (3) The induction of Th1 immunity will be compared against the presence or absence of a complete pathologic response to determine if there is a significant correlation using Pearson r correlation analysis. Targeted Accrual: The study will accrue up to 16 patients. Clinical trial information: NCT04329065 .

B cells require licensing by dendritic cells to serve as primary antigen-presenting cells for plasmid DNA

ABSTRACT DNA vaccines have been an attractive approach for treating cancer patients, however have demonstrated modest immunogenicity in human clinical trials. Dendritic cells (DCs) are known to cross-present DNA-encoded antigens expressed in bystander cells. However, we have previously reported that B cells, and not DCs, serve as primary antigen-presenting cells (APCs) following passive uptake of plasmid DNA. Here we sought to understand the requirements for B cells to present DNA-encoded antigens, to ultimately increase the immunogenicity of plasmid DNA vaccines. Using ovalbumin-specific OT-1 CD8+ T cells and isolated APC populations, we demonstrated that following passive uptake of plasmid DNA, B cells but not DC, can translate the encoded antigen. However, CD8 T cells were only activated by B cells when they were co-cultured with DCs. We found that a cell-cell contact is required between B cells and DCs. Using MHCI KO and re-purification studies, we demonstrated that B cells were the primary APCs and DCs serve to license this function. We further identified that the gene expression profiles of B cells that have been licensed by DCs, compared to the B cells that have not, are vastly different and have signatures similar to B cells activated with a TLR7/8 agonist. Our data demonstrate that B cells transcribe and translate antigens encoded by plasmid DNA following passive uptake, however require licensing by live DC to present antigen to CD8 T cells. Further study of the role of B cells as APCs will be important to improve the immunological efficacy of DNA vaccines.

Antigen presenting cell targeted T cell DNA vaccine inducing strong and specific cellular responses across multiple T cell epitopes of SARS-COV-2

Abstract The severe respiratory syndrome coronavirus 2 (SARS-CoV-2) causing COVID-19 has continuously evolved with successive new virus variants of concern (VOC). A key challenge is to develop vaccines that can prevent infection and/or protect against severe disease caused VOCs that evade vaccine induced Spike neutralizing antibodies. T cell responses likely contribute to the efficacy of approved vaccines against VOCs and provide the rationale for the development of T-cell based vaccines. VB10.2210 is a T-cell based vaccine that was developed using Nykode’s easily adaptable DNA plasmid (pDNA) vaccine platform. VB10.2210 pDNA encodes homodimers consisting of i) a targeting unit that binds chemokine receptors on antigen-presenting cells, ii) a dimerization unit, and iii) an antigenic unit consisting of a selection of validated immunogenic SARS-CoV-2-specific T cell epitopes. The T cell epitopes were identified by Adaptive using T cell receptor sequencing of more than 6500 samples from COVID-19 individuals representing diverse geographies. The vaccine candidate was designed with broad HLA coverage and contains a diversity of both MHCI and MHCII T-cell epitopes across multiple viral proteins and known VOCs. In vitro characterization of the VB10.2210 pDNA showed that intact protein was expressed and secreted in human cell culture. The immunogenicity of the vaccine candidate was evaluated in transgenic HLA-A2.1, C57BL/6 and BALB/c mice. Preclinical data in these three mouse models demonstrate that VB10.2210 consistently induced strong, broad, dose-dependent, and persistent T cell responses across multiple T cell epitopes. VB10.2210 is currently being evaluated for safety and immunogenicity in a Phase I clinical trial (NCT05069623).