Lisa Osborne, CEO at RANA Healthcare Solutions interviews Daniel Kavanagh, PhD, Senior Scientific Advisor, Gene Therapy at WCG.
The Covid-19 pandemic has put vaccine development centre-stage. Never have citizens around the world been so focused on the length of time it takes to develop, test, approve, manufacture and distribute new vaccines. This heightened interest is because most people consider vaccine administration, and the subsequent development of herd immunity, to be the best, most viable option for returning the world to pre-pandemic norms.
Fortunately, new molecular technologies are enabling the traditional vaccine development timeline to be shortened considerably, allowing safe and effective products to be made available faster than was previously possible.
In its simplest form, a vaccine is an intervention intended to induce a protective immune response and immunological memory. A prophylactic vaccine has at least two components: an antigen, which is a molecular target or a viral protein that is recognised by the immune system, and an adjuvant, which serves as the activating signal to turn on the immune system. Sometimes one molecule can serve both purposes.
Historically, many vaccines were developed by taking a virus, growing it in culture, and then either killing it (with heat or chemical inactivation) or weakening or attenuating it using random mutations. That dead or weakened virus was then injected into a person to prime an immune response.
Those traditional vaccines were developed with an incomplete knowledge of molecular biology, so they were empirically and not rationally designed.
But the newest generation of investigational vaccines, including RNA, DNA, and viral vector vaccines, has been created by applying synthetic molecular technologies and rational design. That was possible because researchers now understand much more about how the immune system works and how viruses are built. In some cases, they can produce very targeted molecular constructs that create the desired immune response without having to grow and attenuate actual viruses. Incorporation of genetically engineered DNA or RNA in the vaccine product gives scientists tremendous flexibility and precision for induction of immune responses against a target such as a coronavirus.
To give some current examples, the Pfizer-BioNTech and Moderna Covid-19 vaccines are both RNA vaccines. Inovio has developed a DNA vaccine. Oxford-AstraZeneca and Johnson & Johnson are taking a different approach with their Covid-19 products – they are viral vector vaccines.
Developing RNA vaccines
Researchers can now take a viral sample from anywhere in the world, run it through a sequencer, and determine its genetic sequence. An expert can look at that sequence and determine what part of it is coding for the virus envelope, for example, which is a useful target for protective antibodies.
Scientists will use a variety of informatic techniques to optimise genetic code extracted from the viral sequence to make it more suitable for vaccine development. Vaccine developers can use a plug-and-play approach to insert newly acquired viral sequences into established molecular expression systems.
DNA encoding a complete expression module is produced de novo and used as a template for transcription of the genetic code from DNA into RNA.
Thus, they produce a synthetic RNA sequence reflecting a portion of the natural viral envelope that they believe is most important as an antibody target.
Most of that RNA – the open reading frame – codes for a series of amino acids that forms a protein. But there is also an untranslated section of code in front of and behind the open reading frame that provides context and tells the cell what to do, i.e., bind to a ribosome and create this protein. Often the open reading frame incorporates an export signal, which instructs the protein to leave the cell, where it is exposed to immune receptors that prime an immune response.
Several different techniques can be used to introduce that RNA into cells at the bench or in the lab. For example, electroporation, whereby the cells are exposed to a brief, high electromagnetic field, creates little openings in the membrane which allow the nucleic acids to enter.
Another approach, especially suited for clinical development, is to create a lipid nanoparticle, a lipid cage that surrounds the RNA. When the lipid nanoparticle hits the cell membrane (which is a lipid biolayer), it can squeeze through, in a similar manner to the way soap bubbles can stick to each other and coalesce.
The RNA is then introduced into the cell, and if it has the appropriate context, when it encounters a ribosome, it will be translated into the protein. As a result, the cell begins making the protein the researchers designed.
Getting that process to work in an animal or a human is trickier because there are many different filtering mechanisms and anatomical barriers involved. But the process is working quite well with RNA vaccines now.
A lot of development and quality control work must be performed before that step to ensure that not only is the correct protein produced but it also folds to resemble the target structure on a real virus.
The primary sequence needs to be designed in such a way that when the protein leaves the cell, it forms a spike that looks like the spike on the outside of a virus. It is that spike that the antibodies need to recognise.
Once the body’s macrophages and dendritic cells discover that protein, they will carry it to a lymph node and present it to white blood cells, such as B cells, which will start making antibodies, and CD4 helper T cells that activate and direct B cell responses.
RNA vaccines are classified as biologics by the US Food and Drug Administration (FDA), but because they can be produced without cell culture their manufacturing process is in many ways like that of chemical synthesis. That is one reason why Pfizer-BioNTech and Moderna have been able to ramp up production of their Covid-19 vaccines so quickly to meet the massive global demand.
Developing viral vector vaccines
The oldest vaccine in modern medicine, the smallpox vaccine, was invented by Edward Jenner in the 18th century, using a vaccinia virus from a cow. (That is where the term vaccine comes from, vacca is Latin for cow.) The vaccinia virus induces a strong immune response against smallpox and other pox viruses.
Now using genetic engineering, parts of the native viral code in the vaccinia virus can be replaced with DNA encoding coronavirus antigens. Then, if a person is injected with that vector, in theory, it will induce an immune response against the coronavirus (as well as some poxviral antigens). That is the basic concept of a viral vector.
Some researchers are using vaccinia for that purpose, but there are a dozen other viruses from different virus families being used for experimental vaccines.
Johnson & Johnson and Oxford-AstraZeneca are both using adenoviruses as the vector for their Covid-19 vaccines. Adenoviruses are well known and have been studies extensively. Wildtype adenoviruses can cause a range of illnesses in humans, including the common cold and flu-like symptoms, according to the US Centers for Disease Control and Prevention (CDC).1
When developing a viral vector vaccine, the first step is to take the virus and attenuate it, i.e., knock out the disease-causing pieces of the genome. While it is no longer a disease-causing virus, it can still cause a brief infection. Then, the signals, sequences, and coding for the antigen that they want to induce an immune response against need to be introduced.
That approach frequently produces a robust immune response against the target vaccinal antigen and sometimes against vector antigens, which is both good and bad. It is good because these viruses induce a strong immune response. The problem is that there is also off-target immune reactivity against the vector. Different approaches have been developed to try and avoid that – they include stripping down the vector to minimise the presence of vector proteins and designing a prime-boost approach where the same viral vector is not used in repeated doses. This type of prime-boost system is not being used in the current leading Covid-19 vaccine candidates, but might prove useful in the future, for example in helping to protect vaccinated individuals from emerging novel coronavirus infections.
Moderna is working on a “boost” but as it has a vector-less vaccine, that is a different concept. Moderna plans to adjust the RNA sequence in its vaccine to match the new Covid-19 variants more closely and then introduce it as a third dose in the regimen.
Preparing for a modern vaccine clinical trial
When any investigational product that contains genetically modified DNA or RNA (which includes all vaccines mentioned above) is being introduced into a person, the US National Institutes of Health (NIH) defines it as a human gene transfer trial.
When NIH funding is involved, that trial is subject to the NIH Guidelines for Research Involving Recombinant or Synthetic Nucleic Acid Molecules, and it must have Institutional Biosafety Committee (IBC) approval at each clinical trial site before the research can begin. The IBC is responsible for reviewing the site’s facilities, training, procedures, and the research protocol. It also assesses the site’s compliance with both NIH and CDC guidelines for handling recombinant synthetic and infectious materials. The IBC’s purview is to protect clinic staff, visitors, the public and the environment from any unintended consequences of the proposed research.
IBCs were originally formed at US universities in the 1970s at the NIH’s behest. Each NIH-funded institution involved in this type of research was required to form an IBC composed of its own scientists, plus two people from the community that were unaffiliated with the university, to review all the projects, and then vote on whether to approve them. That is still the process followed at many major universities.
But when hospitals and clinics around the globe want to conduct a vaccine study, they rarely have a team of scientists and a compliance office with the requisite experience on site to form an IBC and oversee it.
WCG has developed a centrally administered IBC model to address that need. The company establishes an IBC for each site, staffed with its own experienced scientists and local community members (ideally persons involved in public health, clinical care, or biosafety), who are not affiliated with the research site. It then registers each IBC with the NIH. An IBC typically includes experts in areas such as microbiology, molecular biology, gene therapy, and biosafety; depending on research under review, the committee may also include experts in plant or animal science, agriculture, or medical specialties.
It typically takes six to eight weeks for each new IBC to be formally recognised by the NIH, but efforts are being made to accelerate that process for Covid-19 research.
The IBC reviews the protocol to identify potential biohazards before any work begins. IBC scientists must answer crucially important questions, such as: Does the research plan to employ a novel infectious agent? Does the researchers’ proposed RNA sequence inadvertently code for a toxin? Can the new product replicate itself? What is the risk of viral shedding and does it constitute a public health risk?
Partnering with IRBs
While they have different areas of responsibility and expertise, it can benefit all parties if the IBC collaborates with the institutional/independent review board (IRB) reviewing the clinical research protocol on issues relating to participant safety.
The IRB is tasked with ensuring that the research protocol does not present any ethical issues and that the highest standards of human participant protection are maintained. The IRB can give a study-level review of a protocol.
While the IBC can conduct a central review of the protocol, it still needs to perform on-site assessments to determine, for example, how each research site disposes of its used needles and other sharps. The IBC also makes sure that all staff wear the correct personal protective equipment (PPE) for the agent they are working with.
Then, the IBC at each site convenes to deliberate and vote on whether to approve the research proposal.
Conducting vaccine trials
Once the necessary approvals have been obtained, the clinical study can begin. Vaccine clinical trials usually follow the standard phase 1 to 3 drug development approach. A Phase 1 trial is typically a single arm trial designed to test the vaccine’s safety and immunogenicity. A Phase 2 trial is usually a placebo-controlled study to determine the optimal vaccine dosing and further investigate its safety and efficacy. Phase 3 trials are generally pivotal randomised controlled trials powered to detect whether the vaccine has sufficient safety and efficacy to justify marketing approval.
Benefits of being prepared
Global vaccine development against Covid-19 has progressed quickly in part because researchers have been preparing for a potential pandemic for almost two decades and they also had prior experience with severe acute respiratory syndrome (SARS).
In May 2018, the World Health Organisation (WHO) published a book on managing epidemics. It referenced several major epidemics from the 21st century, including old diseases, such as cholera, plague, and yellow fever, that had returned, and new ones that had emerged, such as SARS, pandemic influenza, Middle East respiratory syndrome (MERS), Ebola and Zika.2 WHO first identified SARS in February 2003.3
As a result, when the Covid-19 pandemic hit, researchers had tested strategies that they could draw upon. They already knew how to sequence a coronavirus and create an RNA molecule that resembled the virus’ characteristic protein spike to trigger the required immune response. They had also designed the nanoparticle carriers and validated storage methods that would preserve the vaccine long enough for the doses to be divided and distributed efficiently. A lot of the potential problems had been solved by researchers who were preparing to make novel vaccines in anticipation of a pandemic.
Vaccine regulatory approvals
In addition, global regulatory agencies are becoming increasingly accustomed to evaluating biologics license applications for gene transfer vaccines.
The first gene transfer vaccine for an infectious disease was approved by the FDA in June 2016. It was a PaxVax genetically modified bacteria to provide protection against cholera.4 Then, the FDA approved a Merck & Co. live, attenuated vaccine that had been genetically engineered to contain a protein from the Zaire ebolavirus for the prevention of Ebola virus disease in December 2019.5 But Pfizer-BioNTech6 and Moderna6 were the first companies to receive FDA authorisation, in the form of an emergency use authorisation (EUA), for an RNA vaccine. They were granted EUAs for their Covid-19 RNA vaccines on December 11 and December 18, 2020, respectively. The FDA’s Vaccines and Related Biological Products Advisory Committee voted unanimously on February 26, 2021 to recommend that the FDA authorise Johnson & Johnson’s single dose viral vector Covid-19 vaccine for emergency use in adults. The FDA followed their advice and granted that EUA the next day.8
Global collaboration has been a central tenet of the Covid-19 response and has helped maximise the impact of all the technological advances that have been made. The flexible, modular technology platforms that were used to design and manufacture the Covid-19 vaccines will also allow vaccines to be developed for other indications. RNA vaccines are currently being studied for a wide range of infectious diseases including seasonal flu, Zika, rabies, cytomegalovirus, and Nipah virus. Not only are these advances proving to be an asset now, but they will ensure that the world is better positioned to respond to future epidemics or pandemics.
About the Authors
Dr. Kavanagh serves as scientific lead in WCG’s IBC Services division. Previously, Dr. Kavanagh was assistant professor of medicine at Harvard Medical School (HMS) engaged in preclinical and clinical vaccine development, assistant immunologist at the Massachusetts General Hospital (MGH), IBC Vice Chair, HMS/ MGH, and a principal investigator studying infectious diseases at the Ragon Institute of MGH, MIT, and Harvard. He has a PhD in molecular microbiology and immunology from the Oregon Health and Science University, and RAC credential through the Regulatory Affairs Professionals Society.
Ms. Osborne is CEO of Rana Healthcare Solutions, a boutique public relations (PR) agency. She has 25 years’ experience as a medical reporter, editor, and PR consultant. She received a BSc in biological sciences from Birmingham University, UK.
- CDC Adenovirus Symptoms https://www.cdc.gov/adenovirus/about/symptoms.html
- WHO Severe Acute Respiratory Syndrome (SARS) overview. who.int/health-topics/severe-acute-respiratory-syndrome#tab=tab_1
- Managing epidemics. Key facts about major deadly diseases. WHO. May 2018. who.int/emergencies/diseases/managing-epidemics/en/
- FDA approves vaccine to prevent cholera for travelers. Press release. June 10, 2016. fda.gov/news-events/press-announcements/fda-approves-vaccine-prevent-cholera-travelers
- First FDA-approved vaccine for the prevention of Ebola virus disease. Press release. December 19, 2019. fda.gov/news-events/press-announcements/first-fda-approved-vaccine-prevention-ebola-virus-disease-marking-critical-milestone-public-health
- Pfizer-BioNTech Covid-19 Vaccine EUA. fda.gov/emergency-preparedness-and-response/coronavirus-disease-2019-covid-19/pfizer-biontech-covid-19-vaccine
- Moderna Covid-19 Vaccine EUA. https://www.fda.gov/emergency-preparedness-and-response/coronavirus-disease-2019-covid-19/moderna-covid-19-vaccine
- Johnson & Johnson’s Covid-19 Vaccine EUA. https://www.fda.gov/news-events/press-announcements/fda-issues-emergency-use-authorization-third-covid-19-vaccine