A Combination of Two Human Monoclonal Antibodies Limits Fetal Damage by Zika Virus in Macaques

Human infection by Zika virus (ZIKV) during pregnancy can lead to vertical transmission and fetal aberrations, including microcephaly. Prophylactic administration of antibodies can diminish or prevent ZIKV infection in animal models, but whether passive immunization can protect nonhuman primates and their fetuses during pregnancy has not been determined. Z004 and Z021 are neutralizing monoclonal antibodies to domain III of the envelope (EDIII) of ZIKV. Together the two antibodies protect nonpregnant macaques against infection even after Fc modifications to prevent antibody-dependent enhancement in vitro (ADE) and extend their half-lives. Here we report on prophylactic co-administration of the Fc-modified antibodies to pregnant rhesus macaques challenged 3 times with ZIKV during first and second trimester. The two antibodies did not entirely eliminate maternal viremia but limited vertical transmission protecting the fetus from neurologic damage. Thus, maternal passive immunization with two antibodies to EDIII can shield primate fetuses from the harmful effects of ZIKV

Mutational escape from the polyclonal antibody response to SARS-CoV-2 infection is largely shaped by a single class of antibodies

Monoclonal antibodies targeting a variety of epitopes have been isolated from individuals previously infected with SARS-CoV-2, but the relative contributions of these different antibody classes to the polyclonal response remains unclear. Here we use a yeast-display system to map all mutations to the viral spike receptor-binding domain (RBD) that escape binding by representatives of three potently neutralizing classes of anti-RBD antibodies with high-resolution structures. We compare the antibody-escape maps to similar maps for convalescent polyclonal plasma, including plasma from individuals from whom some of the antibodies were isolated. The plasma-escape maps most closely resemble those of a single class of antibodies that target an epitope on the RBD that includes site E484. Therefore, although the human immune system can produce antibodies that target diverse RBD epitopes, in practice the polyclonal response to infection is dominated by a single class of antibodies targeting an epitope that is already undergoing rapid evolution

A Combination of Two Human Monoclonal Antibodies Limits Fetal Damage by Zika Virus in Macaques

Human infection by Zika virus (ZIKV) during pregnancy can lead to vertical transmission and fetal aberrations, including microcephaly. Prophylactic administration of antibodies can diminish or prevent ZIKV infection in animal models, but whether passive immunization can protect nonhuman primates and their fetuses during pregnancy has not been determined. Z004 and Z021 are neutralizing monoclonal antibodies to domain III of the envelope (EDIII) of ZIKV. Together the two antibodies protect nonpregnant macaques against infection even after Fc modifications to prevent antibody-dependent enhancement in vitro (ADE) and extend their half-lives. Here we report on prophylactic co-administration of the Fc-modified antibodies to pregnant rhesus macaques challenged 3 times with ZIKV during first and second trimester. The two antibodies did not entirely eliminate maternal viremia but limited vertical transmission protecting the fetus from neurologic damage. Thus, maternal passive immunization with two antibodies to EDIII can shield primate fetuses from the harmful effects of ZIKV

Convergent Antibody Responses to SARS-CoV-2 Infection in Convalescent Individuals

During the COVID-19 pandemic, SARS-CoV-2 infected millions of people and claimed hundreds of thousands of lives. Virus entry into cells depends on the receptor binding domain (RBD) of the SARS-CoV-2 spike protein (S). Although there is no vaccine, it is likely that antibodies will be essential for protection. However, little is known about the human antibody response to SARS-CoV-2. Here we report on 149 COVID-19 convalescent individuals. Plasmas collected an average of 39 days after the onset of symptoms had variable half-maximal pseudovirus neutralizing titres: less than 1:50 in 33% and below 1:1,000 in 79%, while only 1% showed titres above 1:5,000. Antibody sequencing revealed expanded clones of RBD-specific memory B cells expressing closely related antibodies in different individuals. Despite low plasma titres, antibodies to three distinct epitopes on RBD neutralized at half-maximal inhibitory concentrations (IC₅₀ values) as low as single digit nanograms per millitre. Thus, most convalescent plasmas 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

A Combination of Two Human Monoclonal Antibodies Prevents Zika Virus Escape Mutations in Non-human Primates

Zika virus (ZIKV) causes severe neurologic complications and fetal aberrations. Vaccine development is hindered by potential safety concerns due to antibody cross-reactivity with dengue virus and the possibility of disease enhancement. In contrast, passive administration of anti-ZIKV antibodies engineered to prevent enhancement may be safe and effective. Here, we report on human monoclonal antibody Z021, a potent neutralizer that recognizes an epitope on the lateral ridge of the envelope domain III (EDIII) of ZIKV and is protective against ZIKV in mice. When administered to macaques undergoing a high-dose ZIKV challenge, a single anti-EDIII antibody selected for resistant variants. Co-administration of two antibodies, Z004 and Z021, which target distinct sites on EDIII, was associated with a delay and a 3- to 4-log decrease in peak viremia. Moreover, the combination of these antibodies engineered to avoid enhancement prevented viral escape due to mutation in macaques, a natural host for ZIKV

Human Antibody Response to Flaviviruses and SARS-Coronavirus-2

The last decade has seen a number of viral epidemics and global pandemic. Increased global connectivity makes the likelihood of future mass outbreaks ever greater. Understanding the adaptive immune response to these pathogens will expand basic understanding of viral immunity mechanisms and can help inform vaccine and therapeutic development. This thesis describes the characterization of the human antibody response to several important viral pathogens, all of which are either emerging pathogens posing significant public health burdens or are already responsible for epidemic outbreaks. These include tick-borne encephalitis virus (TBEV), Powassan virus (POWV), Zika virus (ZIKV) and severe acute respiratory syndrome coronavirus (SARS-CoV-2). The first part of this thesis focuses on encephalitis-causing tick-borne flaviviruses, specifically TBEV and POWV. Both TBEV and POWV are emerging human pathogens that cause potentially fatal disease; there are no disease-specific treatments. Mouse monoclonal antibodies are protective against TBEV and POWV, but little is known about the human antibody response to active infection. Although 10,000 to 20,000 cases of TBEV infection are reported per year, study of the immune response to POWV is additionally complicated by the paucity of confirmed cases. Here I report on the human neutralizing antibody response to TBEV in a cohort of infected and vaccinated individuals, and to POWV in a cohort of infected individuals. Memory B cells expressing anti- TBEV envelope domain-III (EDIII) antibodies were isolated using a combination of single-cell sorting and nested polymerase chain reaction (PCR). Expanded clones of memory B cells expressing closely related anti-EDIII antibodies were found in both groups of volunteers. Antibodies representative of clonal and unique sequences found in TBEV-infected and -vaccinated donors were expressed as monoclonals and characterized functionally. In reporter virus particle (RVP) neutralization assays, the most potently neutralizing antibodies with IC50s below 1ng/mL were found only in convalescent donors. Neutralization assays using authentic TBEV in vitro confirmed potent neutralization activity in at least the four antibodies tested. Select antibodies also neutralized RVPs representative of other emerging tick-borne flaviviruses including Langat, louping ill, Omsk hemorrhagic fever, Kyasanur forest disease, and Powassan viruses. Prophylactic or early therapeutic antibody administration was effective at low doses in mice that were lethally infected with TBEV. Shifting focus to the ongoing SARS-CoV-2 pandemic, this virus continues to be a global problem in part because of the emergence of variants of concern that evade neutralization by antibodies elicited by Wuhan-Hu-1 infection or vaccination. I report on the human neutralizing antibody and memory responses to the Gamma variant in a cohort of hospitalized individuals from Manaus, Brazil. Plasma from infected individuals potently neutralized viruses pseudotyped with the Gamma SARS-CoV-2 spike protein, but neutralizing activity against Wuhan-Hu-1, Beta, Delta, or Omicron was significantly lower. Memory B cells expressing anti-Gamma receptor binding domain (RBD) antibodies were again isolated using a combination of single-cell sorting and nested polymerase chain reaction (PCR). Monoclonal antibodies representative of clonal and unique antibody sequences were expressed and characterized functionally. In an in vitro neutralization assay using pseudovirus, antibodies from Gamma-infected individuals were shown to neutralize Gamma and Beta pseudoviruses more effectively than Wuhan-Hu-1. Sixty-nine % and 34% of Gamma-neutralizing antibodies failed to neutralize Delta or Wuhan-Hu-1. In contrast to class 1 and 2 antibodies that dominate the response to Wuhan-Hu-1 or Beta, 43% of antibodies elicited by Gamma infection recognized Class 3 epitopes. These results have implications for variant-specific vaccines, suggesting that these strategies would have limited efficacy. In the course of characterization, specific antibodies with unique properties were identified. In the final part of this thesis, I return to these antibodies to investigate them in more detail. They include antibody T025, which was shown to be efficacious at preventing disease in an in vivo model of infection. Structural analysis revealed a conserved epitope near the lateral ridge of EDIII adjoining the EDI-EDIII hinge region. Antibody T036, also from a TBEV-infected donor, was shown separately to enhance RVP and authentic virus infection in vitro in a Fc receptor-independent manner. Enhancement could be ablated by addition of 4G2, a mouse anti-fusion loop monoclonal antibody. T036 was shown to bind a cryptic epitope on the TBEV EDIII, suggesting that T036 binds the virion during or after conformational change of the E protein. Separately, Z039 and Z015 had been shown in previous studies of anti-ZIKV EDIII antibodies in ZIKV-exposed cohorts to exhibit unexpectedly broad cross-reactivity. Z039 and Z015 were confirmed to bind ZIKV EDIII, as well as Dengue virus 1-4 EDIIIs, and yellow fever virus and West Nile virus EDIIIs, respectively. in vitro neutralization assays using authentic virus confirmed corresponding patterns of antibody-mediated neutralization. Z039 was shown to recognize an epitope on the flavivirus EDIII similar to that of Z021, an anti-Zika antibody with comparatively limited cross-reactivity, although Z039 appears to make contacts with the EDIII backbone which may partially explain its breadth of activity. Z015, on the other hand, does not compete with Z021 or Z004 and recognizes a distinct but cryptic epitope on the yellow fever virus EDIII