A Skin Test To Measure T-Cell Immunity to SARS-CoV-2
Understanding the level of immunity an individual has developed against SARS-CoV-2 provides useful information that can be used to support patient care, public health surveillance and vaccine trials during the pandemic. So far, measuring immunity has centred on the use of serological assays to detect antibodies. However, to overcome some of the limitations associated with this approach, interest is growing in the development and use of tests that can measure cell-mediated immunity.
Technology Networks recently had the pleasure of speaking to Seth Lederman, president and chief executive officer of Tonix Pharmaceuticals, to learn more about the T cell response to SARS-CoV-2, how it compares to the antibody response and the importance of measuring it during the pandemic. In this interview, Seth also talks about TNX-2100, a skin test designed to assess and measure T cell immune responses to SARS-CoV-2.
Anna MacDonald (AM): What do we know so far about the immune response to SARS-CoV-2?
Seth Lederman (SL): We have learned two crucial pieces of information about the immune response to SARS-CoV-2 since the onset of the pandemic. First, most people who recover from COVID-19 have protection against developing a serious case if they are reinfected. Second, we also know that most vaccinated people are protected against a serious case of COVID-19. So far, it seems that protection against serious COVID-19 from vaccination is similar to the protection from natural infection.
However, there are three important pieces of information that we still do not know. First, who are the rare “vaccine failures”, meaning people who received a vaccine but are still susceptible? Second, who are the rare people who recovered from COVID-19, but remain susceptible to getting COVID-19 again? Third, for how long does immunity last for people who were successfully vaccinated or who developed immunity after natural COVID-19?
AM: How does T cell immunity compare to antibody immunity? Why is the T cell response so important?
SL: Immunity to specific pathogens is mediated by T cells and antibodies. T cells play the lead role in fighting viruses. Antibodies play the lead role in fighting bacteria, but also have a supporting role in fighting viruses. Virus-specific T cells seek out virally infected cells in the body and kill them. Importantly T cells can kill infected cells before they become virus factories. In this process, T cells play important roles in clearing the pathogen from the body. T cell immunity lasts for years or even decades. The main challenge with T cells is that virus-specific T cells are hard to measure in laboratory tests. Although antibodies only play a supporting role in defending against viruses, they are easier to measure in blood tests. Fortunately, the presence of anti-virus antibodies is a surrogate marker for T cell immunity to viruses. Unfortunately, antibody responses are transient and typically disappear six to nine months after infection.
Monitoring T cell immunity to SARS-CoV-2 is potentially important in establishing whether someone was successfully vaccinated or developed immunity after recovering from COVID-19 for two reasons. First, the T cell response is primarily involved in fighting and clearing the virus, whereas the antibody response is mostly a surrogate marker. Second, the T cell response lasts longer than the antibody response, so only T cells can reveal whether someone was exposed or infected a year before the measurement, because if an antibody response had been present, it will likely have disappeared after a year.
AM: How could testing for T cell immunity be used in the pandemic?
SL: Measuring T cell responses to SARS-CoV-2 has the potential to inform healthcare providers about whether an individual patient has immunity and also to inform public health officials about how many people have been infected with SARS-CoV-2 and how many have been successfully or unsuccessfully vaccinated. Only relying on antibody tests to assess the exposure of the population, will underestimate the number of people who have been exposed, infected, or vaccinated. First, antibody immunity wanes over time, so a year after infection, most COVID-19 convalescent individuals would be antibody negative, even though they were likely to have been antibody positive for several months. Second, some individuals appear to respond to SARS-CoV-2 exposure or infection by mounting a T cell response without an antibody response.
From the perspective of managing the health and safety of each individual, knowledge of their T cell immunity could be an element of the COVID-19 “passport” back to work, school, public transportation, and air travel. For example, T cell immunity is expected to correlate with protection from severe COVID-19 and in reduced capacity to transmit SARS-CoV-2 to others. However, establishing the significance of T cell immunity in predicting protection and risk of forward transmission will take large studies over time.
From a public health perspective, the number of individuals with T cell immunity to SARS-CoV-2 in the population may be an important element in “herd immunity”. Herd immunity is a term that describes a level of immunity in a population when the number of immune individuals – who are protected and who do not transmit a virus efficiently – reaches a point where infection rates decline. If individuals with T cell immunity, but no antibody immunity to SARS-CoV-2 contribute to herd immunity, then the U.S. population – or at least pockets of it - may be closer to herd immunity than the antibody surveys alone suggest. Accurate profiling of the T cell immunity of a population has the potential to predict how close we are to developing herd immunity and also guiding policy about if and when resuming normal lifestyles would be advisable.
AM: Can you tell us about the TNX-2100 test?
SL: The TNX-2100 skin test is designed to measure T cell immunity using delayed-type hypersensitivity (DTH). DTH is the classic measure of antigen-specific T cell protection that has been in use for more than a century. The TNX-2100 skin test consists of three different mixtures of synthetic peptides designed to represent different protein components of the SARS-CoV-2 virus: multi-antigen peptides, spike peptides, and non-spike peptides. Since all of the COVID-19 vaccines available under Emergency Use Authorization (EUA) are based on the SARS-CoV-2 spike protein, the reason for three different peptide cocktails is to discriminate between people who are naïve, people who were vaccinated, and people who are COVID-19 convalescent.
Multi-antigen Spike peptides Non-Spike Peptides
COVID-19 naïve &unvaccinated - - -
Spike Vaccinated only + + -
Vaccinated and/or Convalescent + + +
The three mixtures of peptides are inserted into the skin at separate locations on the inner surface of a person’s arm in a clinic setting. After 48 to 72 hours, individuals who have been infected or exposed to SARS-CoV-2 would be expected to respond to all three mixtures. The measure of T cell immunity is the diameter of the raised, hardened region of the skin called “induration”. The diameter of induration is expected to be roughly proportional to the amount of T cell immunity to SARS-CoV-2, similar to how it works for in other conditions such as tuberculosis or TB using “purified protein derivative” or PPD from Mycobacteria tuberculosis.
AM: How does the test compare to other methods of detecting T cell immunity? What advantages does it offer?
SL: The major potential advantages of TNX-2100 is that it is easier to perform than other available tests, does not require complex laboratory equipment and is a measure of in vivo functional immunity. The typical methods of detecting T cell immunity to SARS-CoV-2 require expensive, multi-step sample preparation and in vitro T cell stimulation using methods that have not yet been standardized and require highly specialized laboratories. These techniques involve drawing relatively large amounts of blood, isolating T cells from the collected blood, stimulating the isolated T cells with SARS-CoV-2 peptides in tissue culture, and finally analyzing the cultured T cells using sophisticated assays. In the most common technique, called intracytoplasmic cytokine staining (ICS) the analysis instrument is called a multi-parameter flow cytometer.
The TNX-2100 skin test is designed to employ the classic DTH reaction which has been used for TB for approximately 120 years. The DTH skin test does not require specialized techniques. The skin test is designed to be administered in a clinic setting like doctor’s office, one of the growing number of retail pharmacy-based clinics, or a work-based employee health center. Given the simplicity of the procedure and the expected stability of the peptides, it could be administered at a remote outpost without running water or in inclement or extreme weather. The skin test involves one visit for the administration of the test and a second visit for a health care professional to measure the induration. In the future, with sufficient validation and testing, it is possible that the second visit might be replaced by a mobile-phone image that can be transmitted to a healthcare professional for interpretation and documentation.
AM: Are SARS-CoV-2 variants likely to affect the performance of the skin test?
SL: The effect of SARS-CoV-2 variants on the performance of the skin test will need to be studied, but we believe that the T cell response to the currently identified variants is more likely to be constant between the variants than the antibody responses. So far, the variants appear to be mutating to change peptide epitopes that are positioned on the outside of the spike protein, which is where antibodies bind. In contrast, T cells recognize peptide epitopes that are often from the insides of proteins, because proteins need to be chewed up into fragments before T cells can see them. The number of changes in potential T cell epitopes between the variants appears small. Their patterns of altered spike amino acids suggest that the variants are mutating to evade antibody responses. The success of the variants in evading antibody responses has been shown in studies where Wuhan-type COVID-19 convalescent serum has reduced capacity to neutralize the variants. Together, these facts and observations suggest that the skin test will be relatively unaffected by the variants.
AM: What further steps are needed before the test could become widely available?
SL: In February, we received the written response from the U.S. Food and Drug Administration (FDA) to Type B pre-investigational new drug (IND) meeting questions for TNX-2100. This document provides guidance for product development and clinical testing. TNX-2100 has not been studied yet, so the next step is to submit an IND application to get FDA clearance to test this technology in the clinic. Our team at Tonix is currently finalizing the design of a proof-of-concept study. We expect this will be followed rapidly by a larger study. Provided the test performs as we expect, we believe development will progress quickly. When fully developed, the TNX-2100 skin test is expected to provide clinicians, patients, employers, and public health officials with information to guide patient care and public health decisions.
AM: Aside from the development of TNX-2100, what other ways is Tonix Pharmaceuticals involved in the COVID-19 response?
SL: We are developing TNX-1800, a live viral vector vaccine against SARS-CoV-2. TNX-1800 is a modified horsepox virus that is engineered to express the SARS-CoV-2 spike protein. Recent DNA analysis reported a 99.7% colinear identity between a circa 1860 U.S. smallpox vaccine and the horsepox virus, that we are developing as a vector platform. The finding that a vaccine so similar to our vector, was already used as a smallpox vaccine, supports the case for human safety. It also supports our belief that horsepox is closer to the vaccine virus that Edward Jenner used in 1798 to vaccinate against smallpox than the modern vaccines called “vaccinia” today. We have formed a strategic collaboration with Southern Research to develop TNX-1800 involving animal testing. We recently announced that TNX-1800 induced a strong immune response against SARS-CoV-2 in non-human primates and protected the upper and lower airways from SARS-CoV-2 challenge.
From a manufacturing perspective, we are engaged in a manufacturing partnership with FUJIFILM Diosynth Biotechnologies (FDBT) for production of TNX-1800 at FDBT’s College Station, Texas site. We also recently announced plans to build-out our own process development facility in New Bedford, MA and to build a vaccine production facility in Hamilton, MT, which is home to the National Institutes of Health’s Rocky Mountain National Labs.
The TNX-2100 skin test was a byproduct of our efforts to develop and test the TNX-1800 vaccine, because the TNX-1800 vaccine is designed to elicit a strong T cell response to SARS-CoV-2. The potential advantages of a strong T cell response include durability of protection and the inhibition of forward transmission should a vaccinated person be subsequently infected with SARS-CoV-2. We are optimistic about the prospects of TNX-1800 because Edward Jenner’s smallpox vaccine was not just the first vaccine, but also remains the most successful vaccine in history. Jenner’s vaccine eradicated smallpox, which is the only infectious disease ever eradicated. It is unlikely that SARS-CoV-2 will be eradicated by any strategy since it is already known to infect a variety of other animals. It is more likely that SARS-CoV-2 will become endemic. For that reason, our short-term goal is to develop TNX-1800 as a vaccine for adults, but our long-term goal is to test and develop TNX-1800 as a potential vaccine for children to be included with the other childhood immunizations.
Seth Lederman was speaking to Anna MacDonald, Science Writer for Technology Networks.