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Publication on the Development of Recombinant Neutralizing Antibodies that Block SARS-CoV-2 Infection

Recombinant Antibodies Paper Summary
 

July 21, 2020

The coronavirus disease 2019 (COVID-19) pandemic is caused by a novel coronavirus called the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has spread worldwide and currently has no treatment and no vaccine.

Treatment development, as well as diagnostic tools and vaccines, necessitate high-quality antibodies that can be easily generated in a reproducible and scalable manner. Antibodies that neutralize infection could be used for passive immunity. This means that patients would be treated with antibodies that recognize the virus and prevent or limit the severity of the disease, instead of needing to produce their own antibodies after they are infected. High-quality antibodies can also be used to detect the presence of SARS-CoV-2 in diagnostic assays as well as being controls for diagnostic tests for whether a person is producing their own anti-SARS-CoV-2 antibodies in response to infection.

A recent article in the journal Cell Reports describes the results of a study in which a team of scientists that included researchers from Fudan University and Active Motif China purified memory B cells from 11 COVID-19 patients and obtained 729 human immunoglobulins (Igs) by single-cell cloning that recognized the SARS-CoV-2 spike protein S1 and its receptor-binding domain (RBD). 17 of these Igs were strong binders to S1 or the RBD and 8 were able to effectively neutralize the infectivity of SARS-CoV-2. These antibodies potently neutralized SARS-CoV-2, with the best one having a neutralizing IC50 of 1.75 nM, suggesting that they could be developed for use as therapeutics against SARS-CoV-2.

Are Antibodies the Answer to the COVID-19 Pandemic?

The COVID-19 pandemic has already contributed to catastrophic consequences across the world, including a high death rate, forcing countries into lockdown, and ongoing economic crises. Even though the pandemic was halted in a lot of countries, it is still a significant public health issue in the United States, and there are concerns that the end of the lockdowns worldwide could cause a second wave of infections.

So, how do we get back to “normal life” while preventing another repeat pandemic? Different solutions are already in place in the countries that have removed their lockdowns.

Improved COVID-19 Testing Can Slow Its Spread

Large-scale testing programs are being used in the countries that are successfully controlling COVID-19 to detect infected patients as soon as possible. This allows the epidemiologists and public health officials to identify a potential cluster of infections and institute quarantines before SARS-CoV-2 can spread very far.

SARS-CoV-2 infection can be detected by three types of tests: identifying the viral genome using qPCR, identifying antibodies to the virus produced by the immune response using ELISA-like techniques, or identifying viral proteins using antibodies. Each type of test has its own pros and cons, so it is best to consult your healthcare provider to get more information if you are thinking about getting tested.

Investing in a Cure for COVID-19

A lot of resources are being dedicated to finding a cure or therapies for COVID-19. Different clinical trials (DisCoVeRy, RECOVERY) have been launched to test existing therapeutic approaches, but they have not been successful so far.

Treatment of patients with plasma from people that had COVID-19 but recovered, which is referred to as convalescent plasma, showed positive results on patients’ health as well as a reduced viral load. Nevertheless, convalescent plasma is only available in limited quantity. The development of recombinant neutralizing antibodies could be a good therapeutic alternative to plasma therapy because it is amenable to high-scale production.

A COVID-19 Vaccine is Our Best Hope – But Takes Time

The development of a vaccine is a priority. A good vaccine could both block new infections (referred to as a prophylactic vaccine) as well as be used to treat patients already infected with the virus (which is referred to as a therapeutic vaccine).

Although the development of a successful vaccine is important, it will take a long time and there is no guarantee that a vaccine will be developed in time to help end this pandemic.

Therefore, the development of recombinant neutralizing antibodies to treat SARS-CoV-2 infection seems to currently be the most cost-effective, fast, and promising solution in the fight against COVID-19.

Characterization of Recombinant Human Antibodies that Block SARS-CoV-2 Infection

In a paper recently published in the journal Cell Reports, a team of researchers from Fudan University, Active Motif China, and their collaborators reported on the development and characterization of recombinant antibodies that neutralized SARS-CoV-2 infectivity, making them potential therapeutic candidates. The scientists had access to blood from 11 patients that recovered from COVID-19 and they cloned the memory B cells from these patients and characterized the antibodies they produced. Their goal was to identify and produce antibodies that neutralized SARS-CoV-2.

The antibodies were isolated and produced using the proprietary AbEpic™ technology developed by Active Motif. After peripheral blood mononuclear cell (PBMC) isolation, single antigen-specific memory B cells were sorted using His-tagged S1 bait and specific B cell markers to obtain only RBD- and S1-binding memory B cells. Then, heavy and light chains were amplified, cloned, sequenced, and characterized. Once cloned, heavy and light chains of human monoclonal antibodies were expressed in HEK293E cells and purified with protein A. 729 paired antibody genes were obtained from the 11 patients.

178 of these antibodies showed RBD and S1 binding specificity by ELISA, with some demonstrating a sub-nanomolar EC50. However, the recognition and binding of the antigen by an antibody are not necessarily enough to neutralize the virus. So, the researchers tested neutralization properties of the antibodies in pseudoviral and live viral infection assays. HEK293T cells expressing the ACE2 receptor (the receptor required for entry by SARS-CoV-2) were infected with pseudovirus. 12 antibodies neutralized infectivity of the pseudovirus and were competed by 50 nM of ACE2. The best antibody (clone 414-1) displayed an IC50 of 3 nM in the pseudovirus neutralization assays, with a strong affinity to the RBD domain.

The antibodies that neutralized the pseudovirus were then tested in live viral neutralization assays in Vero-E6 cells overexpressing ACE2. The clone 414-1 antibody was able to block live viral entry with IC50 of 1.75 nM. The combination of the clone 414-1 antibody and another antibody, clone 553-15, decreased the IC50 to 0.45 nM. Additional antibodies showed neutralization potential, inhibiting live SARS-CoV-2 with IC50s of <10 nM.

These results indicate that recombinant human antibody clone 414-1 potently neutralizes SARS-CoV-2 infectivity, making it a strong therapeutic candidate. Furthermore, the authors demonstrated that combinations of clones 414-1 and 553-15 resulted in significantly enhanced neutralization properties.

Summary: Advancing COVID-19 Research, Diagnostics, and Therapeutics with Recombinant Antibodies

COVID-19 became a worldwide pandemic in just a few months after the novel coronavirus that causes the disease was discovered. The research projects studying SARS-CoV-2 and COVID-19 have progressed rapidly to identify and characterize the unknown virus, determine how it enters the cells, and which mechanisms could be targeted for a potential cure or vaccine.

A team of researchers from Fudan University and Active Motif China and their collaborators was able to isolate and characterize human antibodies recognizing and neutralizing SARS-CoV-2. Most of the antibodies they developed demonstrated no cross-reactivity with other coronaviruses, making them ideal therapeutic candidates due to this high level of virus specificity.

These antibodies display a high affinity and specificity towards the RBD domain of the SARS-CoV-2 Spike S1 protein. Therefore, these antibodies could be developed for diagnostic purposes because of their sensitive detection of the virus in infected tissue or other biological samples by ELISA-like techniques.

One antibody alone was able to block live virus entry, and these neutralization properties were enhanced further by combining antibodies.

A fully human recombinant neutralization antibody like this could be used in approaches to develop therapeutics to treat SARS-CoV-2 infection with a direct effect on the viral load. Further testing is needed to validate the use of these types of antibodies in a therapeutic setting in the clinic, including in vivo and clinical trials, but taken together, these data give high hopes regarding the possibility of COVID-19 treatment and the hopefully the end of this pandemic.


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