Early treatment with a combination of two potent neutralizing antibodies improves clinical outcomes and reduces virus replication and lung inflammation in SARS-CoV-2 infected macaques.

There is an urgent need for effective therapeutic interventions against SARS-CoV-2, including new variants that continue to arise. Neutralizing monoclonal antibodies have shown promise in clinical studies. We investigated the therapeutic efficacy of a combination of two potent monoclonal antibodies, C135-LS and C144-LS that carry half-life extension mutations, in the rhesus macaque model of COVID-19. Twelve young adult macaques (three groups of four animals) were inoculated intranasally and intra-tracheally with a high dose of SARS-CoV-2 and 24 hours later, treated intravenously with a high (40 mg/kg) or low (12 mg/kg) dose of the C135-LS and C144-LS antibody combination, or a control monoclonal antibody. Animals were monitored for 7 days. Compared to the control animals, animals treated with either dose of the anti-SARS-CoV-2 antibodies showed similarly improved clinical scores, lower levels of virus replication in upper and lower respiratory tract, and significantly reduced interstitial pneumonia, as measured by comprehensive lung histology. In conclusion, this study provides proof-of-concept in support of further clinical development of these monoclonal antibodies against COVID-19 during early infection.

Bispecific IgG neutralizes SARS-CoV-2 variants and prevents escape in mice.

Neutralizing antibodies that target the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein are among the most promising approaches against COVID-191,2. A bispecific IgG1-like molecule (CoV-X2) has been developed on the basis of C121 and C135, two antibodies derived from donors who had recovered from COVID-193. Here we show that CoV-X2 simultaneously binds two independent sites on the RBD and, unlike its parental antibodies, prevents detectable spike binding to the cellular receptor of the virus, angiotensin-converting enzyme 2 (ACE2). Furthermore, CoV-X2 neutralizes wild-type SARS-CoV-2 and its variants of concern, as well as escape mutants generated by the parental monoclonal antibodies. We also found that in a mouse model of SARS-CoV-2 infection with lung inflammation, CoV-X2 protects mice from disease and suppresses viral escape. Thus, the simultaneous targeting of non-overlapping RBD epitopes by IgG-like bispecific antibodies is feasible and effective, and combines the advantages of antibody cocktails with those of single-molecule approaches.

Longitudinal Serological Analysis and Neutralizing Antibody Levels in Coronavirus Disease 2019 Convalescent Patients.

BACKGROUND: Understanding the longitudinal trajectory of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies is crucial for diagnosis of prior infection and predicting future immunity. METHODS: We conducted a longitudinal analysis of coronavirus disease 2019 convalescent patients, with neutralizing antibody assays and SARS-CoV-2 serological assay platforms using SARS-CoV-2 spike (S) or nucleocapsid (N) antigens. RESULTS: Sensitivities of serological assays in diagnosing prior SARS-CoV-2 infection changed with time. One widely used commercial platform that had an initial sensitivity of >95% declined to 71% at 81-100 days after diagnosis. The trajectories of median binding antibody titers measured over approximately 3-4 months were not dependent on the use of SARS-CoV-2 N or S proteins as antigen. The median neutralization titer decreased by approximately 45% per month. Each serological assay gave quantitative antibody titers that were correlated with SARS-CoV-2 neutralization titers, but S-based serological assay measurements better predicted neutralization potency. Correlation between S-binding and neutralization titers deteriorated with time, and decreases in neutralization titers were not predicted by changes in S-binding antibody titers. CONCLUSIONS: Different SARS-CoV-2 serological assays are more or less well suited for surveillance versus prediction of serum neutralization potency. Extended follow-up should facilitate the establishment of appropriate serological correlates of protection against SARS-CoV-2 reinfection.

Evolution of antibody immunity to SARS-CoV-2.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has infected 78 million individuals and is responsible for over 1.7 million deaths to date. Infection is associated with the development of variable levels of antibodies with neutralizing activity, which can protect against infection in animal models1,2. Antibody levels decrease with time, but, to our knowledge, the nature and quality of the memory B cells that would be required to produce antibodies upon reinfection has not been examined. Here we report on the humoral memory response in a cohort of 87 individuals assessed at 1.3 and 6.2 months after infection with SARS-CoV-2. We find that titres of IgM and IgG antibodies against the receptor-binding domain (RBD) of the spike protein of SARS-CoV-2 decrease significantly over this time period, with IgA being less affected. Concurrently, neutralizing activity in plasma decreases by fivefold in pseudotype virus assays. By contrast, the number of RBD-specific memory B cells remains unchanged at 6.2 months after infection. Memory B cells display clonal turnover after 6.2 months, and the antibodies that they express have greater somatic hypermutation, resistance to RBD mutations and increased potency, indicative of continued evolution of the humoral response. Immunofluorescence and PCR analyses of intestinal biopsies obtained from asymptomatic individuals at 4 months after the onset of coronavirus disease 2019 (COVID-19) revealed the persistence of SARS-CoV-2 nucleic acids and immunoreactivity in the small bowel of 7 out of 14 individuals. We conclude that the memory B cell response to SARS-CoV-2 evolves between 1.3 and 6.2 months after infection in a manner that is consistent with antigen persistence.

Antibody potency, effector function, and combinations in protection and therapy for SARS-CoV-2 infection in vivo.

SARS-CoV-2, the causative agent of COVID-19, has been responsible for over 42 million infections and 1 million deaths since its emergence in December 2019. There are few therapeutic options and no approved vaccines. Here, we examine the properties of highly potent human monoclonal antibodies (hu-mAbs) in a Syrian hamster model of SARS-CoV-2 and in a mouse-adapted model of SARS-CoV-2 infection (SARS-CoV-2 MA). Antibody combinations were effective for prevention and in therapy when administered early. However, in vitro antibody neutralization potency did not uniformly correlate with in vivo protection, and some hu-mAbs were more protective in combination in vivo. Analysis of antibody Fc regions revealed that binding to activating Fc receptors contributes to optimal protection against SARS-CoV-2 MA. The data indicate that intact effector function can affect hu-mAb protective activity and that in vivo testing is required to establish optimal hu-mAb combinations for COVID-19 prevention.

Enhanced SARS-CoV-2 neutralization by dimeric IgA.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus that causes coronavirus disease 2019 (COVID-19), primarily infects cells at mucosal surfaces. Serum neutralizing antibody responses are variable and generally low in individuals that suffer mild forms of COVID-19. Although potent immunoglobulin G (IgG) antibodies can neutralize the virus, less is known about secretory antibodies such as IgA that might affect the initial viral spread and transmissibility from the mucosa. Here, we characterize the IgA response to SARS-CoV-2 in a cohort of 149 convalescent individuals after diagnosis with COVID-19. IgA responses in plasma generally correlated with IgG responses. Furthermore, clones of IgM-, IgG-, and IgA-producing B cells were derived from common progenitor cells. Plasma IgA monomers specific to SARS-CoV-2 proteins were demonstrated to be twofold less potent than IgG equivalents. However, IgA dimers, the primary form of antibody in the nasopharynx, were, on average, 15 times more potent than IgA monomers against the same target. Thus, dimeric IgA responses may be particularly valuable for protection against SARS-CoV-2 and for vaccine efficacy.

COVID-19 antibody development fueled by HIV-1 broadly neutralizing antibody research.

PURPOSE OF REVIEW: The coronavirus disease 2019 (COVID-19) pandemic has caught the world unprepared, with no prevention or treatment strategies in place. In addition to the efforts to develop an effective vaccine, alternative approaches are essential to control this pandemic, which will most likely require multiple readily available solutions. Among them, monoclonal anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies have been isolated by multiple laboratories in record time facilitated by techniques that were first pioneered for HIV-1 antibody discovery. Here, we summarize how lessons learned from anti-HIV-1 antibody discovery have provided fundamental knowledge for the rapid development of anti-SARS-CoV-2 antibodies. RECENT FINDINGS: Research laboratories that successfully identified potent broadly neutralizing antibodies against HIV-1 have harnessed their antibody discovery techniques to isolate novel potent anti-SARS-CoV-2 antibodies, which have efficacy in animal models. These antibodies represent promising clinical candidates for treatment or prevention of COVID-19. SUMMARY: Passive transfer of antibodies is a promising approach when the elicitation of protective immune responses is difficult, as in the case of HIV-1 infection. Antibodies can also play a significant role in post-exposure prophylaxis, in high-risk populations that may not mount robust immune responses after vaccination, and in therapy. We provide a review of the recent approaches used for anti-SARS-CoV-2 antibody discovery and upcoming challenges in the field.

Antibody potency, effector function and combinations in protection from SARS-CoV-2 infection in vivo.

SARS-CoV-2, the causative agent of COVID-19, is responsible for over 24 million infections and 800,000 deaths since its emergence in December 2019. There are few therapeutic options and no approved vaccines. Here we examine the properties of highly potent human monoclonal antibodies (hu-mAbs) in a mouse adapted model of SARS-CoV-2 infection (SARS-CoV-2 MA). In vitro antibody neutralization potency did not uniformly correlate with in vivo activity, and some hu-mAbs were more potent in combination in vivo . Analysis of antibody Fc regions revealed that binding to activating Fc receptors is essential for optimal protection against SARS-CoV-2 MA. The data indicate that hu-mAb protective activity is dependent on intact effector function and that in vivo testing is required to establish optimal hu-mAb combinations for COVID-19 prevention.

Longitudinal analysis of clinical serology assay performance and neutralising antibody levels in COVID19 convalescents.

OBJECTIVES: To investigate longitudinal trajectory of SARS-CoV-2 neutralising antibodies and the performance of serological assays in diagnosing prior infection and predicting serum neutralisation titres with time Design Retrospective longitudinal analysis of a COVID19 case cohort . Setting NHS outpatient clinics Participants Individuals with RT-PCR diagnosed SARS-CoV-2 infection that did not require hospitalization Main outcome measures The sensitivity with which prior infection was detected and quantitative antibody titres were assessed using four SARS-CoV-2 serologic assay platforms. Two platforms employed SARS-CoV-2 spike (S) based antigens and two employed nucleocapsid (N) based antigens. Serum neutralising antibody titres were measured using a validated pseudotyped virus SARS-CoV-2 neutralisation assay. The ability of the serological assays to predict neutralisation titres at various times after PCR diagnosis was assessed. Results The three of the four serological assays had sensitivities of 95 to100% at 21-40 days post PCR-diagnosis, while a fourth assay had a lower sensitivity of 85%. The relative sensitivities of the assays changed with time and the sensitivity of one assay that had an initial sensitivity of >95% declined to 85% at 61-80 post PCR diagnosis, and to 71% at 81-100 days post diagnosis. Median antibody titres decreased in one serologic assay but were maintained over the observation period in other assays. The trajectories of median antibody titres measured in serologic assays over this time period were not dependent on whether the SARS-CoV-2 N or S proteins were used as antigen source. A broad range of SARS-CoV-2 neutralising titres were evident in individual sera, that decreased over time in the majority of participants; the median neutralisation titre in the cohort decreased by 45% over 4 weeks. Each of the serological assays gave quantitative measurements of antibody titres that correlated with SARS-CoV-2 neutralisation titres, but, the S-based serological assay measurements better predicted serum neutralisation potency. The strength of correlation between serologic assay results and neutralisation titres deteriorated with time and decreases in neutralisation titres in individual participants were not well predicted by changes in antibody titres measured using serologic assays. CONCLUSIONS: SARS-CoV-2 serologic assays differed in their comparative diagnostic performance over time. Different assays are more or less well suited for surveillance of populations for prior infection versus prediction of serum neutralisation potency. Continued monitoring of declining neutralisation titres during extended follow up should facilitate the establishment of appropriate serologic correlates of protection against SARS-CoV-2 reinfection.

Measuring SARS-CoV-2 neutralizing antibody activity using pseudotyped and chimeric viruses.

The emergence of SARS-CoV-2 and the ensuing explosive epidemic of COVID-19 disease has generated a need for assays to rapidly and conveniently measure the antiviral activity of SARS-CoV-2-specific antibodies. Here, we describe a collection of approaches based on SARS-CoV-2 spike-pseudotyped, single-cycle, replication-defective human immunodeficiency virus type-1 (HIV-1), and vesicular stomatitis virus (VSV), as well as a replication-competent VSV/SARS-CoV-2 chimeric virus. While each surrogate virus exhibited subtle differences in the sensitivity with which neutralizing activity was detected, the neutralizing activity of both convalescent plasma and human monoclonal antibodies measured using each virus correlated quantitatively with neutralizing activity measured using an authentic SARS-CoV-2 neutralization assay. The assays described herein are adaptable to high throughput and are useful tools in the evaluation of serologic immunity conferred by vaccination or prior SARS-CoV-2 infection, as well as the potency of convalescent plasma or human monoclonal antibodies.

Convergent antibody responses to SARS-CoV-2 in convalescent individuals.

During the coronavirus disease-2019 (COVID-19) pandemic, severe acute respiratory syndrome-related coronavirus-2 (SARS-CoV-2) has led to the infection of millions of people and has claimed hundreds of thousands of lives. The entry of the virus into cells depends on the receptor-binding domain (RBD) of the spike (S) protein of SARS-CoV-2. Although there is currently no vaccine, it is likely that antibodies will be essential for protection. However, little is known about the human antibody response to SARS-CoV-21-5. Here we report on 149 COVID-19-convalescent individuals. Plasma samples collected an average of 39 days after the onset of symptoms had variable half-maximal pseudovirus neutralizing titres; titres were less than 50 in 33% of samples, below 1,000 in 79% of samples and only 1% of samples had titres above 5,000. Antibody sequencing revealed the expansion of clones of RBD-specific memory B cells that expressed closely related antibodies in different individuals. Despite low plasma titres, antibodies to three distinct epitopes on the RBD neutralized the virus with half-maximal inhibitory concentrations (IC50 values) as low as 2 ng ml-1. In conclusion, most convalescent plasma samples obtained from individuals who recover from COVID-19 do not contain high levels of neutralizing activity. Nevertheless, rare but recurring RBD-specific antibodies with potent antiviral activity were found in all individuals tested, suggesting that a vaccine designed to elicit such antibodies could be broadly effective.