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

Abstract

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.

Keywords: COVID-19; DNA; DNA vaccines; SARS-CoV-2; nucleic acid vaccines; vaccines.

Figure 1.. Production of circular DNA constructs.

(A) Production of plasmid DNA. The designed plasmid containing the GOI is transformed to the bacterial host to generate a GMO. After fermentation, bacterial cells are harvested and lysed through chemical, physical or mechanical methods. After the removal of solids from the lysate (clarification), several steps of purification such as chromatographic purification are used for the purification of plasmid DNA. (B) Production of minicircle DNA. After the growth of the genetically modified bacteria containing the parental plasmid, the expression of recombinase and the restriction enzyme is induced by arabinose from the plasmid or bacterial genome. The recombinase initiates the site-specific recombination between its recognition sequences (attB and attP), originating the minicircle DNA and a MP consisting of the bacterial backbone. Then, MP is degraded specifically by the RE. The minicircle DNA is extracted and purified after cell harvest, cell lysis, clarification and several steps of purification. GOI: Gene of interest; GMO: Genetically modified organism; MP: Mini plasmid; RE: Recombinase.

Figure 2.. Production of linear, covalently-closed minimalistic expression DNA constructs.

(A) Production of MIDGE DNA. The purified parental plasmid containing the GOI flanking with a restriction site (e.g. EcoRI) is digested by the RE. The resulting fragments are ligated to the hairpin oligodeoxynucleotides to generate MIDGE DNA molecules. Then, unligated fragments including plasmid backbones are digested by T7 DNA polymerase and the MIDGE DNA is purified. (B) Production of Doggybone™ DNA. The purified parental plasmid containing the GOI flanking with telomeric ends (Tel-L and Tel-R) is denatured by NaOH and used as a template DNA in a RCA reaction using Phi29 DNA polymerase. The resulting DNA concatemers are cleaved and joined by an enzymatic reaction using TelN protelomerase to generate Doggybone DNA. Doggybone™ DNA molecules are then purified using chromatographic purification. The purified Doggybone DNA may be used as a template DNA for the RCA reaction. GMO: Genetically modified organism; GOI: Gene of interest; MIDGE: Minimalistic, immunologically defined gene expression; RCA: Rolling circle amplification.

Figure 3.. Overview of physical DNA vaccine delivery technologies.

(A) Delivery of DNA vaccines into skin compartments using gene gun, jet injector and microneedles. These physical methods allow for delivery of DNA vaccines into the epidermis, dermis and subcutaneous compartments by providing enhanced efficacy and great safety than conventional needle methods. (B) The use of in vivo electroporation enhances the cellular uptake of injected DNA across the cell membrane.

Figure 4.. Antigen expression by DNA vaccines.

The DNA construct encoding the transgene (vaccine antigen) translocates to the nucleus, where the transcription to mRNA takes place. Produced mRNAs are then delivered to the cytoplasm to ensure efficient translation of the vaccine antigen. MIDGE: Minimalistic, immunologically defined gene expression.

Figure 5.. Induction of adaptive immune responses by DNA vaccines.

DNA vaccines are administered by multiple routes. The injected DNA enters the somatic cells and/or APCs to produce target antigens, which is the subject of immune surveillance by both MHC class I and class II molecules of APCs. Following the presentation of antigens, activated APCs travel to the draining lymph nodes to present antigenic peptides to CD4 and CD8 T cells. This interaction allows activation and expansion of T cells, as well as antibody-producing B cells. Both humoral and cellular immune responses are induced to trigger a robust response against the target antigens. APCs: Antigen-presenting cells; BCR: B-cell receptor; MHC: Major histocompatibility complex; TCR: T-cell receptor.