Charles River Laboratories International : The Hunt for a Safe, Effective Vaccine

Scientists are using new strategies and movingly rapidly to find a long-lasting preventive measure against COVID-19

SARS-CoV-2, the virus that causes COVID-19, continues to spread. At the time of writing, Johns Hopkins has tracked nearly 3 million laboratory confirmed cases and more than 200,000 deaths, a number that will continue to rise. There is increasing evidence that as many as 4 out of 5 infected people only develop mild symptoms or remain asymptomatic, but are able to transmit the virus to others. Social distancing guidelines have been implemented in many states and countries to reduce the mixing of susceptible and infectious people. However, these are unlikely to be sustainable in the long term, with many states already beginning to reopen businesses. The Centers for Disease Control have indicated that we may be dealing with this virus for months or even years longer. We need a vaccine to help slow and control the spread of SARS-CoV-2.

Figure 1: Immune response following vaccination and infection. Following vaccination, there is an initial lag phase during which the immune system encounters the immunogen and generates a response. This response slowly builds to a low level, and then declines to a background memory level (blue). Once the real pathogen is encountered, the immune system can respond much more rapidly and strongly, due to the pre-existing memory response (red). This allows the body to rapidly fight the infection, hopefully completely suppressing it, before the person gets sick. (Adapted from Kuby's Immunology, 6th Edition).

Vaccines deliver an immunogen, a non-harmful full or partial version of the virus or bacteria, to train the immune system to recognize the pathogen when it is naturally encountered. This generates a slow to develop primary immune response, leading to the formation of a long-lasting memory bank of cells. When the real pathogen is encountered, the immune response is able to fight back more strongly and rapidly, protecting the individual from disease (Figure 1).

Ideally, a vaccine will stimulate both antibody-based (humoral), mediated by B cells, and cell-mediated, mainly T cells, immunity to stop a microbe before it can cause disease (Figure 2).

Most vaccines used in humans are 'whole' vaccines, composed of either inactivated (dead) or live-attenuated (weakened) versions of the pathogen so that the immunogen cannot cause disease. These strategies are being explored in three SARS-CoV-2 vaccine candidates. However, today there are many new platforms that were already in development and being tested in humans for other vaccines. These are being rapidly applied to COVID-19, with the design guided by previous attempts to produce vaccines against SARS-CoV and MERS-CoV.

According to the World Health Organizations, there are at least 70 coronavirus vaccines in development globally as of April 21, with at least three already in human trials. This number will fluctuate in the weeks to come. These candidates all utilize different vaccine technologies, but almost all rely on the SARS-CoV-2 spike (S) protein as the immunogen:

Viral Vectors

Figure 2: Immune responses stimulated by vaccination. Ideally, a vaccine will stimulate 2 arms of the adaptive immune response- humoral and cellular. The humoral response is mediated by B cells which produce antibodies. These antibodies specifically recognize the target pathogen, binding to its surface. This can aggregate the pathogen and mark it for destruction by other immune cells or may block its ability to bind to and enter host cells. The cellular immune response is largely mediated by T cells. CD4+ T cells are stimulate to produce signaling molecules, called cytokines and chemokines, to recruit and coordinate the action of other immune cells at the site of infection. CD8+ cytotoxic T cells are able to recognize and kill virus infected cells, to prevent the virus from being able to replicate and spread.

Fifteen candidates are currently being developed using recombinant viral vectors, using two different approaches. The vector itself may be genetically modified to remove proteins normally found on its surface and instead display the coronavirus S protein. This modified virus is recognized by the immune system and stimulates memory. Alternatively, the virus may be designed to deliver a piece of DNA encoding the S protein into target cells, directing the cells themselves to produce the S protein. This later strategy is the same technology being explored for gene therapy, but repurposed as a vaccine. Five candidates using adenoviruses are currently in development. This includes a Chinese vaccine which is already in Phase 2 trials to assess its ability to stimulate anti-coronavirus immunity (its immunogenicity) and its safety profile in healthy volunteers compared to a placebo control. Other candidate vaccines are being built using measles, horsepox, influenza, modified vaccinia virus Ankara, and vesicular stomatitis virus, but likely more viruses will be repurposed into coronavirus vaccine vectors. This approach has worked well in the veterinary field, with 12 viral vector vaccines currently in use. However, no viral vector vaccine has yet been approved for use in humans.

DNA Plasmids

This strategy places the gene for the spike protein inside of a DNA plasmid, a circular piece of DNA that is maintained outside of the normal genome of the cell and that is used to transfer in new genes. However, it is hard for naked DNA to enter into cells, so many vaccine candidates are paired with devices or substances to increase uptake. For example, an electroporation device may be used to gently and quickly 'shock' the cells to create temporary pores in the membrane, allowing the vaccine DNA to enter easily. The pores are quickly closed, the cell produces the S protein, and this then stimulates the immune response. This strategy is also in early clinical trials for COVID-19 and at least 5 other DNA plasmid candidates are in the planning stages. As with viral vectors, this strategy is already used in veterinary medicine, including for vaccines against West Nile Virus in horses and for canine melanoma, but no vaccines using this technology have yet been approved for humans.

RNA Platforms

Eleven COVID-19 vaccine candidates are composed of a messenger RNA molecule encoding the S protein. Similar to the DNA plasmid vaccines, the mRNA drives the cell to produce the S protein to stimulate immune responses. However, the delivery method is often different. Many candidates are being packaged inside of lipid nanoparticles, essentially a shell of fat molecules, to protect the RNA and help deliver it efficiently into the target cell. mRNA vaccines are expected to be faster, easier, and cheaper to produce than many other vaccine types, which has allowed them to also already reach Phase 1 trials. The biggest challenge, however, is that no mRNA vaccine has yet been approved for either humans or animals, and so long term data on their efficacy and safety is not available as it is for the other platforms.

Subunit Vaccines

Twenty-seven vaccine candidates are based the SARS-CoV-2 proteins themselves. Full length viral proteins (usually the S protein) or shortened pieces of the proteins called peptides are delivered. Other candidates rely on viral-like particles, entirely synthetic versions of the virus containing only the outer structural proteins without any of the replication machinery. These proteins are directly recognized as foreign by immune cells, which mount a response, without requiring the cell to first produce the protein. Subunit vaccines are generally safe and easy to produce, but often require the incorporation of adjuvants to boost the immune response as they are not typically immunogenic enough on their own. Many subunit vaccines are already used in humans, including for Hepatitis B, HPV, Shingles, pertussis, Haemophilus influenzae, and others, perhaps explaining why so many candidates are taking this approach.

When will one of these vaccines be available?

Vaccines often take 10-20 years to develop. For ebolavirus, researchers were able to bring a vaccine to the clinic in about 5 years. Optimistically, for COVID-19, scientists think they can accelerate this even further and produce a vaccine in only 12-18 months. Making a vaccine today containing some putative immunogen is trivially easy due to advances in biotechnology and the wide variety of platforms available. For example, one vaccine candidate reached its first human volunteer only 63 days after the SARS-CoV-2 genome was published. Animal and clinical trials are being performed concurrently, instead of sequentially, to further speed up time lines. They cannot be skipped, however, as this is the only way we can understand a vaccine's immunogenicity and safety profile before it is given to millions of people.

Poor safety profiles have plagued some of the previous coronavirus vaccine attempts. Both SARS-CoV and MERS-CoV vaccine candidates have led to the development of a phenomenon called antibody dependent enhancement. The vaccines did stimulate virus-specific antibodies, but instead of blocking the virus, they actually enhanced the infection, resulting in significant damage to the liver and the lungs in animal models. The mechanism of this is not completely understood, but it is thought that these 'non-neutralizing' antibodies when bound to the virus are recognized by receptors on immune cells. The immune cells then internalize the virus, accidentally infecting themselves. An ongoing question is if antibody dependent enhancement occurs in the context of COVID-19 and, if so, how should a vaccine be designed to avoid this?

A variety of other open-ended questions remain with regards to developing a vaccine, including:

• Will the vaccines stimulate strong anti-coronavirus immune responses in humans?
• What kinds of immune responses will be most effective at preventing future infections?
• How long will a vaccine confer protection? Can people who previously had COVID-19 be re-infected?
• Will the virus mutate so that a vaccinated person's immune system can no longer recognize the new form of the virus?

The good news is that as we continue to study patients who have recovered from COVID-19 and as the data from these vaccine trials start to come in, we will start to better understand how the immune system responds to this virus. Hopefully, this will lead us to a safe and effective vaccine. The world is waiting.