DNA Vaccines: Mechanism, Benefits, and Advances

DNA Vaccines Definition

A DNA vaccine is actually a ring-shaped plasmid DNA constructed from a plasmid expression vector and a target gene for the intended antigen, which is injected into human cells and expresses the antigen encoded by the target gene, thus activating the immune response of the body. From the central law of molecular biology, it is known that DNA stores genetic information in the cell nucleus, transcribes to form mRNA, which directly guides protein translation, while proteins perform their functions in life activities, and immunogenic substances that can activate the body's immune response are proteins. Compared with proteins, plasmid DNA has the advantages of higher stability, easier production and long-term preservation. More than 30 years ago, scientists introduced DNA plasmids directly into the body, and were surprised to find that the target genes injected into the body expressed target proteins and stimulated the body to produce corresponding antibodies and other immune responses, and DNA vaccine technology was born.

What is Recombinant DNA Vaccine?

Using gene recombination technology, the protective antigen genes encoding pathogenic microorganisms are recombined into bacteria, yeast, and cells, and after culture, proliferation, extraction and purification of the expressed protective antigen made of a vaccine. For example, recombinant hepatitis B vaccine, bivalent, quadrivalent and nine-valent human papillomavirus vaccine, etc. Unlike traditional vaccines that use weakened or inactivated forms of pathogens, recombinant DNA vaccines involve the insertion of DNA sequences coding for pathogen-specific antigens into a vector, typically plasmid DNA.

What is Plasmid DNA?

Plasmid DNA (pDNA) is an extrachromosomal stable genetic factor, ranging in size from 1-200kb, which is a double-stranded, closed-loop DNA molecule and exists in the host cell in a superhelical state. Plasmids are mainly found in bacterial, actinomycetes and fungal cells, where they have the ability to replicate and transcribe autonomously, maintain a constant copy number in the progeny cells, and express the genetic information they carry. In the context of DNA vaccines, plasmids are modified to contain the genetic code for specific antigens. These plasmids are designed to be stable and efficiently enter the host cell where they direct the cell to produce the target antigen. This antigen then triggers an immune response without causing disease.

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How DNA Vaccines Work?

DNA vaccines have the advantage of activating both cellular and humoral immune responses, with a portion of the DNA vaccine entering epidermal or myocyte cells, where it expresses antigenic peptides or proteins to stimulate a CD4+ T cell response, and a portion entering directly into antigen-presenting cells to stimulate CD4+ T and CD8+ T cell responses. On the other hand, once the B-cell receptor recognizes the antigenic protein, the humoral immune response is simultaneously stimulated and antibodies are produced with the help of activated antigen-specific CD4+ T cells. These induced immune responses do not directly neutralize and remove pathogens that invade the body, thus achieving a preventive effect. DNA vaccines operate through a series of well-defined steps:

Mechanisms of DNA vaccines. (Hobernik, D.; et al, 2018)

• Gene Encoding: A plasmid DNA vector is engineered to include the genetic code for a pathogen-specific antigen.
• Delivery: The plasmid DNA is delivered into the host cells using various methods such as electroporation, gene gun, or viral vectors.
• Antigen Production: Once inside the host cells, the plasmid DNA is transcribed and translated into the antigen protein.
• Immune Response: The antigen protein is presented on the cell surface or secreted, prompting the host's immune system to recognize and mount a response against it.
• Memory Formation: The immune system develops memory cells that recognize the antigen, providing long-term protection against future infections by the same pathogen.

DNA Vaccine vs mRNA Vaccine

Similar to DNA vaccines, mRNA vaccines deliver genetic material to human cells to synthesize one or more viral or bacterial proteins. While there are some similarities between DNA vaccines and mRNA vaccines, there are clear differences between these genetic vaccines. For a DNA vaccine to be effective, plasmid DNA must cross the cell membrane, enter the cytoplasm, and then travel through the nuclear membrane to the nucleus. In contrast, RNA vaccines only need to cross the cell membrane to enter the cytoplasm. The cytoplasm contains enzymes that use genetic information in mRNA molecules to synthesize bacterial or viral proteins. Because DNA vaccines need to go through additional steps to get into the nucleus, they produce a much lower immune response than mRNA vaccines. However, a plasmid DNA can produce a large number of copies of mRNA. Once the plasmid enters the nucleus, it can produce more bacterial or viral proteins than a single molecule of an mRNA vaccine. DNA vaccines do not inherently have the same immune-stimulating effects as mRNA vaccines, but it is unclear whether this is a drawback, as the inflammation of mRNA vaccines may limit their use. While people may tolerate muscle inflammation and other side effects caused by RNA vaccines in the context of the COVID-19 pandemic, these side effects may limit their use for non-pandemic diseases. In addition, mRNA vaccines are fragile and need to be stored and transported at low or ultra-low temperatures. In contrast, DNA vaccines are more stable and easier to store and transport than mRNA vaccines. The logistics of storing and transporting mRNA vaccines hamper the distribution of vaccines to low-income countries. Temperature-stabilized DNA vaccines offer a viable alternative. For example, the DNA vaccine ZyCoV-D remains stable for at least 3 months at room temperature and even longer at 2-8°C, making it invaluable for resource-limited environments. However, the medical community has long been concerned about the safety of DNA vaccines. Regulators worry about foreign DNA recombining or integrating with DNA from home. Current mRNA vaccine technology has a more direct path to success, as it can be "translated" directly into the protein without having to "translate" it to the nucleus.

What are the Disadvantages of DNA Vaccines?

According to the guidelines on DNA vaccine development updated by WHO in August 2020, the most prominent advantages of DNA vaccines are proposed as follows:

1. Preparation of DNA vaccines requires only recombination operations of nucleic acid substances, which is fast and simple;

2, The use of coding gene fragments do not replicate and do not integrate, adverse reactions are minimal;

3, Access to the body can simultaneously activate cellular and humoral immune response;

4, Stable at room temperature and easy to store, and easy to mass production;

5, Flexible and variable design to cope with viral mutations more quickly.

However, there is one concern that always lingers in the mind, which is the potential risk of integrating genomic DNA. In fact, this concern can be eliminated. The guidelines for the development of DNA vaccines recognize that one of the advantages of DNA is that "the use of encoded gene fragments does not replicate and does not integrate, and adverse reactions are rare. The reason is that DNA vaccine design follows the following four principles:

1. The plasmid DNA does not contain homologous sequences such as those found in the human body;

2. It does not contain elements for viral integration;

3, It does not contain gene-integrating enzymes;

4, It does not contain self-replicating elements.

The safety of DNA vaccines is guaranteed according to the above principles. In addition, many scientific studies have shown that even in the most severe cases of "integration", the resulting mutations are at least three orders of magnitude lower than the number of spontaneous mutations produced by the host, which means that they are completely negligible.

Examples of DNA Vaccine Programs and Research

INO-4800

INO-4800 is a notable DNA vaccine developed by INOVIO Pharmaceuticals for COVID-19. The vaccine is based on a plasmid DNA platform that encodes the spike protein of the SARS-CoV-2 virus. INO-4800 has demonstrated promising results in clinical trials, showing the ability to induce robust immune responses in participants. The vaccine is designed to be administered via intradermal injection using the CELLECTRA electroporation device, which enhances the delivery of the plasmid DNA into cells. This technology has positioned INOVIO as a leader in DNA vaccine development.

VGX-3100

VGX-3100 is another significant DNA vaccine developed by INOVIO Pharmaceuticals, targeting HPV (human papillomavirus) infection. Specifically designed for the treatment of HPV-induced precancerous lesions, VGX-3100 has shown the potential to induce strong cellular immune responses. Clinical trials have indicated that the vaccine can reduce or eliminate precancerous lesions in the cervix, offering a promising therapeutic option for HPV-related cancers.

VX-104

VX-104 is a DNA vaccine developed by PaxVax, targeting various infectious diseases. The vaccine is currently in advanced stages of development, showcasing the flexibility of DNA vaccine technology. VX-104 is designed to provide protection against diseases where traditional vaccines have limitations. The ongoing research aims to assess the vaccine's efficacy and safety in preventing infections and to explore its potential applications in different disease contexts.

ZyCoV-D

ZyCoV-D, developed by Zydus Cadila, is a pioneering DNA vaccine for COVID-19. It is notable for being the world's first DNA vaccine to receive emergency use authorization. ZyCoV-D uses a plasmid DNA vector to encode the spike protein of SARS-CoV-2. The vaccine is administered through a needle-free device, enhancing patient comfort and accessibility. Clinical trials have demonstrated that ZyCoV-D is effective in inducing a strong immune response and has a favorable safety profile.

Optimizing Strategy for DNA Vaccines

• Enhanced Delivery Methods: Developing more effective delivery systems, such as improved electroporation techniques or novel viral vectors, can enhance the uptake of plasmid DNA by host cells.
• Plasmid Design: Engineering plasmids to improve stability, expression levels, and antigen presentation can increase vaccine efficacy. This includes optimizing promoters and codon usage.
• Adjuvants and Boosters: Incorporating adjuvants or additional booster doses can strengthen the immune response and increase the duration of protection.
• Safety and Efficacy Testing: Rigorous preclinical and clinical testing is essential to ensure the safety and effectiveness of DNA vaccines. Ongoing research focuses on minimizing any potential adverse effects and maximizing immune responses.
• Regulatory Pathways: Navigating regulatory requirements effectively can streamline the approval process and facilitate faster vaccine availability to the public.

Reference

1. Hobernik, D.; et al. A DNA Vaccines-How Far From Clinical Use?. Int. J. Mol. Sci. 2018, 19(11): 3605.

DNA Vaccines: Mechanism, Benefits, and Advances

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