Antibodies to SARS-CoV-2: Are Some People Already Immune?
Some people have antibody profiles that could indicate repeat exposures or existing immunity to COVID-19.
The immune response to SARS-CoV-2 has turned out to be a key factor in the severity of the resulting illness COVID-19. The first studies looking at the antibody-mediated response to this coronavirus are now available. Although some of these are preprint articles that have not yet been peer-reviewed, collectively they provide key information about the diversity in the immune response to the virus.
Understanding the antibody-mediated response is key to determining if exposure will generate immunity, how long that immunity will last, and how to test for such immunity. Additionally, variations in the antibody-mediated response could contribute to disease severity through a process called antibody-dependent enhancement (ADE).
Unexpectedly Fast Appearance of IgG Antibodies
Suthar and colleagues examined antibody responses to the part of the virus protein than interacts with the receptor on human cells. This virus protein is called the Spike (S) protein and the region that interacts with the human cell receptor is called the receptor-binding domain (RBD). S is the protein that gives coronaviruses their name, because in images of the virus S looks like a crown surrounding the virus core.
All coronaviruses have an S protein, but the amino acids that make up this protein differ in each specific virus. The coronavirus most similar to SARS-CoV-2, the virus that causes COVID-19, is called SARS-CoV. The S proteins of SARS-CoV and SARS-CoV-2 are only 73% identical in amino acids.
These proteins are also modified by sugars that are attached by enzymes in the infected cells. These sugar modifications are referred to as glycosylation and the pattern of glycosylation can also be different between different S proteins from different coronaviruses. This is part of the reason why infection with one coronavirus does not necessarily generate an immune response against a different coronavirus. The amino acids of the proteins that are exposed on the virus surface and their glycosylation patterns are not identical between even closely related coronaviruses.
However, parts of these proteins could be similar enough to generate cross-reacting antibodies that recognize multiple coronaviruses. Thus, a previous exposure to a coronovirus could result in some antibodies that recognize SARS-CoV-2. Indeed, there are 4 common coronaviruses that infect people: HCoV-HKU1, HCoV-OC43, HCoV-NL63, and HCoV-229E.
The reason for investigating antibodies against S protein is that this protein is exposed on the surface of the viral particle. Additionally, antibodies that target the RBD should interfere with the interaction with S and the human cell receptor, which is a protein called ACE2. Thus, antibodies that recognize this part of the S protein should prevent entry of the virus into cells and let the immune system to remove the virus. Such antibodies would be considered “neutralizing antibodies.”
Another important aspect of the study by Suthar and colleagues is that they examined a particular type of antibody called IgG. There are 5 types of antibodies. The are classified by the type of heavy chain. Part of the heavy chain and the light chain work together to recognize foreign proteins.
IgM is the type that are made upon the first exposure to a pathogen; IgG is the type that are produced and recognize pathogens when re-exposure occurs. By monitoring IgG antibodies that recognize the RBD of S protein, Suthar and colleagues were tracking the antibodies most likely to be neutralizing and provide immunity.
Initially, Suthar and colleagues looked at the RBD antibodies in 44 patients with COVID-19. For the initial study with the 44 patients, the control was blood plasma samples collected from 12 healthy people early in 2019 before SARS-CoV-2 emerged. To test for antibodies specific to RBD, the researchers generated the RBD from S protein. This was only a part of the entire S protein: amino acids 319 to 541. They purified the RBD peptide and then tested the serum from patients for the ability to bind to this purified RBD peptide. The assay that they used to detect the RBD-specific antibodies also indicated what type of antibody was present.
The researchers detected IgG, IgM, and IgA antibodies in people who hospitalized for COVID-19 but not in the healthy control samples. Of the 44 infected patients, 36 had detectable amounts of RBD-specific IgG antibodies. Intriguingly, the amount of the antibodies was highly variable.
Antibodies are measured as “titer,” which is related to how much a solution with the antibody can be diluted before it is undetectable. The IgG RBD antibody titers in the 36 patients ranged from less than 100 to 142,765. (A lower titer means a lower concentration of antibodies.) The amounts of the IgM and IgA antibodies recognizing RBD were also highly variable, and the average amounts of IgM and IgA were less than the average amounts of IgG.
Some of the patients, including the ones with either very low RBD-specific IgG titers or undetectable RBD-specific IgG antibodies, had higher amounts of IgM antibodies recognizing RBD than IgG antibodies recognizing RBD. Other patients had higher titers of IgG than IgM. This is an intriguing finding, because the proportion of the different types of antibodies changes over time after exposure to a virus.
Upon the first exposure to a virus, B cells make IgM antibodies, then some B cells switch and start making IgA antibodies or IgG antibodies. IgM antibodies and the IgG antibodies are present in the circulation; whereas IgA antibodies are mostly in mucus. So, it was not surprising that IgA antibodies against RBD would be lower than IgG or IgM antibodies against RBD in the blood samples. It was also not surprising that some patients had higher amounts of IgM than IgG.
The IgM antibodies would be the first ones to appear. So, patients with their first infection of SARS-CoV-2 and early in the stage of infection (within about 2 weeks) would have a lot of IgM and not many (or any) detectable IgG antibodies against the virus. Thus, the balance of IgM and IgG antibodies can indicate how long ago a person was infected with the virus, whether the person was mounting a response to the first infection with the virus or a subsequent infection, or had antibodies that recognized the virus from infection with a different related virus.
How could some COVID-19 patients have very high titers of IgG and very low titers of IgM? Three possibilities include:
- The patients were late their in first exposure, so their B cells had mostly switch from IgM to IgG.
- The patients had been exposed to SARS-CoV-2 previously and were now mounting the re-exposure response. This is consistent with the reports of asymptomatic people who test positively for the virus. These patients could have had an asymptomatic infection that was cleared and were now infected for a second (or more) time.
- The patients had IgG from memory B cells from a previous coronavirus infection, not the SARS-CoV-2 virus, and these cells were producing antibodies that cross-reacted with a protein on SARS-CoV-2.
Because the patients in the study had symptoms for different lengths of time, it is possible that some patients were responding to SARS-CoV-2 for the first time and some for a subsequent exposure. Some patients were tested after 3 days of symptoms and some after 30 days. Some patients had the RBD IgG antibodies after only 5 days, and some had high titers of these antibodies only 2 days after testing positive for COVID-19. It would be unusual to have IgG antibodies specific for this virus so early in the course of the infection, assuming that symptom onset and time of testing positive for the virus are indicative of the period of exposure.
Antibodies that Neutralize SARS-CoV-2
Another important aspect of the study by Suthar and colleagues is that they tested for the ability of the patients’ antibodies to neutralize the virus. The assay involved mixing diluted samples of the patient’s plasma with SARS-CoV-2 and then using the mixture to infect cells in culture. Of the 44 COVID-19 patients, 40 had neutralizing antibodies in their plasma.
The amount of the neutralizing antibodies was highly variable. Some could be diluted only 55 times and some as much as 5,763 times.
Only some of the antibodies against a virus will be neutralizing. Some will recognize other viral proteins that are not exposed on the surface of the virus but get exposed as infected cells are attacked by the immune system. Some antibodies will recognize other parts of surface viral proteins but do not interfere with entry of the virus into the cells and replication of the virus in the cells. So, just detecting antibodies that recognize a viral protein or even a key part of a viral protein is not enough to know if any of the antibodies will be neutralizing antibodies.
Predicting Neutralizing Antibodies from RBD-Specific IgG Antibody Titer
To determine if the presence of RBD-specific IgG antibodies was sufficient to predict neutralizing activity, the researchers performed a test with a larger set of confirmed COVID-19 patient samples. These 231 samples were collected from patients within 22 days of testing positive for SARS-CoV-2. The samples were grouped according to the time after testing positive: 0–3 days was one group, 4–6 days was another, and 7–22 days was the last group.
The researchers tested the relationship between the presence of the RBD-specific antibodies and neutralizing antibody activity for each of these three groups. Their analysis indicated that using this antibody test had a negligible fast positive rate and produced no false negatives. Thus, this would be a good system for assaying for the presence of antibodies that could potentially provide some level of immunity to SARS-CoV-2.
This study has important implications for timing the testing of people for the presence of neutralizing antibodies. Additionally, it raises the question of whether some of the hospitalized COVID-19 patients are actually not experiencing the virus for the first time. Instead, they are mounting a secondary response dominated by IgG either from a previous asymptomatic SARS-CoV-2 infection. Alternatively, the antibody response could be from a previous infection with another coronavirus.
It is important to note that this study did not assess whether these antibodies confer protection to a subsequent exposure or reduce disease severity. Another point worth mentioning is that not all of the COVID-19 patients had detectable antibodies. Out of the 44 initially tested, 8 did not have detectable antibodies. Either these patients were tested before antibodies developed, or the antibodies that were produced did not recognize the S proteins and so were not detected in the assay. The highlighted study is a preprint that has not yet been peer reviewed.
M. S. Suthar, et al., Rapid Generation of Neutralizing Antibody Responses in COVID-19 Patients. MedRxiv 2020.05.03.20084442 (8 May 2020). https://doi.org/10.1101/2020.05.03.20084442
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