When plasmid DNA was discovered, its function was not even known. Today its importance goes beyond microbial evolution, as it plays a fundamental role in the production of vaccines, gene therapies and cell therapies.
What is plasmid DNA?
A plasmid is a very special small genetic element. It consists of a circular double-stranded DNA present in many bacterial species capable of self-replicating independently of chromosomal DNA. This capacity already gives it the category of “special“, but there is more. Often, the genes found in plasmids encode for proteins necessary for bacterial survival or possess genes that provide genetic advantages. An example of this are antibiotic resistance genes, which enable bacteria to adapt quickly to conditions that limit their growth. But plasmids not only enable adaptation by providing new functions, but also by varying their copy number. This can lead to high gene expression and a high mutation rate, which can be of use to the bacterium. But why are plasmids so widely used today?
From discovery to biotechnology application
Although plasmid DNA was isolated for the first time in 1967, its existence was already known in the 1940s. Its function was unknown to scientists at that time, but that did not prevent them from naming it. The first definition of plasmid DNA had only four words: “any extrachromosomal hereditary element” and was coined by Joshua Lederberg in 1952. It is not surprising that this first definition is not retained, because, although it was a good attempt, we now know that some plasmids are temporarily integrated into chromosomal DNA.
A few years later, Rosalind Franklin’s discovery of the structure of DNA gave a boost to plasmid DNA and its future biotechnological application. Once the structure was known, it took only a few years to figure out the function. When it was realized that DNA was the carrier of genetic information, it became clear that plasmids could be used for gene transmission. This discovery led to the conclusion that plasmids had been fundamental to bacterial evolution, and that we could also put them to use. Once they were isolated, they began to be used as genetic engineering tools. It could not be otherwise, since they were stable and easy to modify genetically, and genes from other species could be inserted into them. At this point, there would be a large window of possibilities.
One of the first biotechnological applications of plasmids was achieved in the 1970s. It was possible to produce human insulin in Escherichia coli thanks to the use of a plasmid as an expression vector. Thus began the era of recombinant DNA. As the years went by, advances in expression and transfection vector techniques increased and consequently so did their applications.
Applications of plasmid DNA
In 1991, the use of plasmid DNA for gene therapy began to be explored. This application is based on the administration of plasmids coding for a therapeutic protein. To express the protein, first an in vivo administration is performed and then transfection into the target cells takes place. The plasmids function as a transport system for the gene, resulting in the in situ production of the therapeutic protein. Compared to the use of viruses, plasmids are a better alternative because they have a low risk of oncogenesis and immunogenicity. They are also more stable and easier to produce in large quantities. Perhaps the major drawback of plasmid DNA is its low efficacy in in vivo studies. By bringing together these two alternatives, viruses and plasmids, the technique used today in most gene therapies is achieved: administration of viral vectors.
In the 1990s, when gene therapy research was just beginning, none of the researchers imagined that plasmids would also have an application in the virtual world, in the metaverse that is so fashionable today. It turns out that in the video game “Bioshock” players can acquire new abilities by injecting plasmids into their bodies. The video game also refers to genetic recombination as a way to modify one’s own genetic structure. Although an anecdotal application, it is an effective way for children to understand how plasmids work and deserves to be mentioned.
The stability and ease of gene editing that plasmids present is what has made them so successful in the biotechnology industry.
On the other hand, the ability of plasmid DNA to stimulate our immune system is perfect for vaccine development. It turns out that, upon sensing the entry of foreign genetic material, our body activates cellular immunity (B and T cells) and in some cases our humoral response (antibodies) can also be activated. This stimulation of the immune response is one of the main advantages that DNA vaccines offer over traditional vaccines. They also eliminate the need to inject infectious agents, they are more stable at different temperatures, so they are easier to store and transport, and they can be manufactured on a large scale and relatively inexpensively. All these advantages have made it possible for DNA vaccines to be developed for various diseases, including several types of cancer and neurological diseases.
Another growing application is the use of plasmids for the production of almost anything. The synthesis of expression vectors is easier than ever and they can be designed for the production of antibodies, viral particles or even CRISPR/Cas elements. In addition, codon optimization can be performed to achieve better production. However, there are also drawbacks. While the immune response it triggers is a positive effect for some of its applications, it is not for all of them. Nor should we forget that it is a tool under development and that there are still logistical challenges in the production of plasmid DNA for clinical applications.
Demand is increasing, will it be possible to meet it?
It is clear that the stability and easy gene editing of plasmids is what has made them so successful in the biotechnology industry. They are fundamental tools for the development of biologic drugs and gene therapies. In fact, many plasmid DNA therapies are advancing rapidly in clinical development and are expected to be on the market in a very short time. Moreover, research into DNA vaccines, as well as personalized cell therapies, will further increase the demand for plasmid DNA. As a result of these advances, plasmid manufacturing capacity will have to increase exponentially. Such a high demand cannot be met by a single company. A whole business ecosystem will be needed. This network will have to be formed not only by manufacturing companies, but also by companies specialized in the manufacture of specific machinery for the production of plasmid DNA. This growing demand is more present than ever and the biotechnology industry will have to be prepared.
53Biologics is a CDMO with extensive experience in biologics production. The company offers end-to-end services for the biopharma industry, including process development, clinical manufacturing and analytical testing to support every step up to commercialization of biotherapeutics.
For more information on the company and its services, please visit 53Biologics.com.
Patricia De La Madrid
Marketing & Business Development Manager at 53Biologics