A brief review on DNA vaccines in the era of COVID-19

This article provides a brief overview of DNA vaccines. First, the basic DNA vaccine design strategies are described, then specific issues related to the industrial production of DNA vaccines are discussed, including the production and purification of DNA products such as plasmid DNA, minicircle DNA, minimalistic, immunologically defined gene expression (MIDGE) and Doggybone™. The use of adjuvants to enhance the immunogenicity of DNA vaccines is then discussed. In addition, different delivery routes and several physical and chemical methods to increase the efficacy of DNA delivery into cells are explained. Recent preclinical and clinical trials of DNA vaccines for COVID-19 are then summarized. Lastly, the advantages and obstacles of DNA vaccines are discussed.

Increased Potential of Bone Formation with the Intravenous Injection of a Parathyroid Hormone-Related Protein Minicircle DNA Vector

Osteoporosis is commonly treated via the long-term usage of anti-osteoporotic agents; however, poor drug compliance and undesirable side effects limit their treatment efficacy. The parathyroid hormone-related protein (PTHrP) is essential for normal bone formation and remodeling; thus, may be used as an anti-osteoporotic agent. Here, we developed a platform for the delivery of a single peptide composed of two regions of the PTHrP protein (1–34 and 107–139); mcPTHrP 1–34+107–139 using a minicircle vector. We also transfected mcPTHrP 1–34+107–139 into human mesenchymal stem cells (MSCs) and generated Thru 1–34+107–139-producing engineered MSCs (eMSCs) as an alternative delivery system. Osteoporosis was induced in 12-week-old C57BL/6 female mice via ovariectomy. The ovariectomized (OVX) mice were then treated with the two systems; (1) mcPTHrP 1–34+107–139 was intravenously administered three times (once per week); (2) eMSCs were intraperitoneally administered twice (on weeks four and six). Compared with the control OVX mice, the mcPTHrP 1–34+107–139-treated group showed better trabecular bone structure quality, increased bone formation, and decreased bone resorption. Similar results were observed in the eMSCs-treated OVX mice. Altogether, these results provide experimental evidence to support the potential of delivering PTHrP 1–34+107–139 using the minicircle technology for the treatment of osteoporosis.

Synthesis and Characterization of Mannosylated Formulations to Deliver a Minicircle DNA Vaccine

DNA vaccines still represent an emergent area of research, giving rise to continuous progress towards several biomedicine demands. The formulation of delivery systems to specifically target mannose receptors, which are overexpressed on antigen presenting cells (APCs), is considered a suitable strategy to improve the DNA vaccine immunogenicity. The present study developed binary and ternary carriers, based on polyethylenimine (PEI), octa-arginine peptide (R8), and mannose ligands, to specifically deliver a minicircle DNA (mcDNA) vaccine to APCs. Systems were prepared at various nitrogen to phosphate group (N/P) ratios and characterized in terms of their morphology, size, surface charge, and complexation capacity. In vitro studies were conducted to assess the biocompatibility, cell internalization ability, and gene expression of formulated carriers. The high charge density and condensing capacity of both PEI and R8 enhance the interaction with the mcDNA, leading to the formation of smaller particles. The addition of PEI polymer to the R8-mannose/mcDNA binary system reduces the size and increases the zeta potential and system stability. Confocal microscopy studies confirmed intracellular localization of targeting systems, resulting in sustained mcDNA uptake. Furthermore, the efficiency of in vitro transfection can be influenced by the presence of R8-mannose, with great implications for gene expression. R8-mannose/PEI/mcDNA ternary systems can be considered valuable tools to instigate further research, aiming for advances in the DNA vaccine field.

Increased Potential of Bone Formation With Intravenous Injection of Parathyroid Hormone-minicircle DNA Vector

Background: Osteoporosis is usually treated with long-term usage of anti-osteoporotic agents. However, poor drug compliance and emerging side effects sometimes are limitations for the treatment of osteoporosis. Parathyroid hormone-related protein (PTHrP) is needed for normal bone formation and remodeling. We attempted a new method using minicircle vectors (mc) encoding PTHrP analogs. Methods: We generated mc encoding the infusion of PTHrP 1-34 with 107-139 (mc 1-34+107-139). Ovariectomized (OVX) model was induced in 12-week-old C57BL/6 female mice. mc 1-34+107-139 was administered three times weekly via intravenous injections. Results: mc 1-34+107-139 significantly increased bone formation compared with the OVX group and decreased the bone resorption. PTHrP mc DNA vector was effective in increasing the quality of trabecular bone structure. Conclusions: These results provide experimental evidence for the therapeutic potential of minicircle DNA vectors in OVX model. This study is a first attempted proof-of-concept gene therapy using minicircle vectors for the treatment of osteoporosis.

Evaluation of BMP-2 Minicircle DNA for Enhanced Bone Engineering and Regeneration

Background: To date, the significant osteoinductive potential of bone morphogenetic protein 2 (BMP-2) non-viral gene therapy cannot be fully exploited therapeutically. This is mainly due to weak gene delivery and brief expression peaks restricting the therapeutic effect. Objective: Our objective was to test the application of minicircle DNA, allowing prolonged expression potential. It offers notable advantages over conventional plasmid DNA. The lack of bacterial sequences and the resulting reduction in size, enables safe usage and improved performance for tissue regeneration. Methods: We inserted an optimized BMP-2 gene cassette with minicircle plasmid technology. BMP-2 minicircle plasmids were produced in E. coli yielding plasmids lacking bacterial backbone elements. Comparative studies of these BMP-2 minicircles and conventional BMP-2 plasmids were performed in vitro in cell systems, including bone marrow derived stem cells. Tests performed included gene expression profiles and cell differentiation assays. Results: A C2C12 cell line transfected with the BMP-2-Advanced minicircle showed significantly elevated expression of osteocalcin, alkaline phosphatase (ALP) activity, and BMP-2 protein amount when compared to cells transfected with conventional BMP-2-Advanced plasmid. Furthermore, the plasmids show suitability for stem cell approaches by showing significantly higher levels of ALP activity and mineralization when introduced into human bone marrow stem cells (BMSCs). Conclusion: We have designed a highly bioactive BMP-2 minicircle plasmid with the potential to fulfil clinical requirements for non-viral gene therapy in the field of bone regeneration.