Structure and recognition of a novel HIV-1 gp120-gp41 interface antibody that caused MPER exposure through viral escape.

CAPRISA, 2017.Abstract available in pdf

Most neutralizing human monoclonal antibodies target novel epitopes requiring both Lassa virus glycoprotein subunits

Lassa fever is a severe multisystem disease that often has haemorrhagic manifestations. The epitopes of the Lassa virus (LASV) surface glycoproteins recognized by naturally infected human hosts have not been identified or characterized. Here we have cloned 113 human monoclonal antibodies (mAbs) specific for LASV glycoproteins from memory B cells of Lassa fever survivors from West Africa. One-half bind the GP2 fusion subunit, one-fourth recognize the GP1 receptor-binding subunit and the remaining fourth are specific for the assembled glycoprotein complex, requiring both GP1 and GP2 subunits for recognition. Notably, of the 16 mAbs that neutralize LASV, 13 require the assembled glycoprotein complex for binding, while the remaining 3 require GP1 only. Compared with non-neutralizing mAbs, neutralizing mAbs have higher binding affinities and greater divergence from germline progenitors. Some mAbs potently neutralize all four LASV lineages. These insights from LASV human mAb characterization will guide strategies for immunotherapeutic development and vaccine design

Passive antibody transfer from pregnant women to their fetus are maximized after SARS-CoV-2 vaccination irrespective of prior infection

Background: Pregnancy is associated with a higher risk of adverse symptoms and outcomes for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection for both mother and neonate. Antibodies can provide protection against SARS-CoV-2 infection and are induced in pregnant women after vaccination or infection. Passive transfer of these antibodies from mother to fetus in utero may provide protection to the neonate against infection. However, it is unclear whether the magnitude or quality and kinetics of maternally derived fetal antibodies differs in the context of maternal infection or vaccination. Objective: We aimed to determine whether antibodies transferred from maternal to fetus differed in quality or quantity between infection- or vaccination-induced humoral immune responses. Methods: We evaluated 93 paired maternal and neonatal umbilical cord blood plasma samples collected between October 2020 and February 2022 from a birth cohort of pregnant women from New Orleans, Louisiana, with histories of SARS-CoV-2 infection and/or vaccination. Plasma was profiled for the levels of spike-specific antibodies and induction of antiviral humoral immune functions, including neutralization and Fc-mediated innate immune effector functions. Responses were compared between 4 groups according to maternal infection and vaccination. Results: We found that SARS-CoV-2 vaccination or infection during pregnancy increased the levels of antiviral antibodies compared to naive subjects. Vaccinated mothers and cord samples had the highest anti-spike antibody levels and antiviral function independent of the time of vaccination during pregnancy. Conclusions: These results show that the most effective passive transfer of functional antibodies against SARS-CoV-2 in utero is achieved through vaccination, highlighting the importance of vaccination in pregnant women

Altered COVID-19 immunity in children with asthma by atopic status

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection causes a spectrum of clinical outcomes that may be complicated by severe asthma. Antiviral immunity is often compromised in patients with asthma; however, whether this is true for SARS-CoV-2 immunity and children is unknown. Objective: We aimed to evaluate SARS-CoV-2 immunity in children with asthma on the basis of infection or vaccination history and compared to respiratory syncytial viral or allergen (eg, cockroach, dust mite)-specific immunity. Methods: Fifty-three children from an urban asthma study were evaluated for medical history, lung function, and virus- or allergen-specific immunity using antibody or T-cell assays. Results: Polyclonal antibody responses to spike were observed in most children from infection and/or vaccination history. Children with atopic asthma or high allergen-specific IgE, particularly to dust mites, exhibited reduced seroconversion, antibody magnitude, and SARS-CoV-2 virus neutralization after SARS-CoV-2 infection or vaccination. TH1 responses to SARS-CoV-2 and respiratory syncytial virus correlated with antigen-respective IgG. Cockroach-specific T-cell activation as well as IL-17A and IL-21 cytokines negatively correlated with SARS-CoV-2 antibodies and effector functions, distinct from total and dust mite IgE. Allergen-specific IgE and lack of vaccination were associated with recent health care utilization. Reduced lung function (forced expiratory volume in 1 second ≤ 80%) was independently associated with (SARS-CoV-2) peptide-induced cytokines, including IL-31, whereas poor asthma control was associated with cockroach-specific cytokine responses. Conclusion: Mechanisms underpinning atopic and nonatopic asthma may complicate the development of memory to SARS-CoV-2 infection or vaccination and lead to a higher risk of repeated infection in these children

Fine mapping of the CAP248-2B epitope.

<p>(A) Cartoon of the CAP248-2B paratope (shown as a mirror image of the docked model in B, and colored as in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006074#ppat.1006074.g003" target="_blank">Fig 3</a>), showing amino acids that have affinity matured relative to the predicted germline genes with main chain Cα spheres. (B) A surface view schematic of gp41 (dark grey) and proximal regions in gp120 (light grey) showing the predicted location of CAP248-2B CDRs and FW3, coloured as in A. The approximate location of the viral membrane is indicated. (C) The envelope trimer is shown in cartoon view with the viral membrane estimated as in B. Regions of Env predicted to form part of the CAP248-2B epitope are coloured and labelled. Point mutants shown to significantly affect CAP248-2B neutralization are shown with black spheres. (D) Table showing neutralization IC₅₀ titers for CAP248-2B, PGT151, VRC34, 3BC315, 35O22, 10E8, and 8ANC195 against CAP45 and various mutants. The location of each mutant in either gp41 or the gp120 C terminus is shown on the left and coloured as in C. Fold effects on IC₅₀ are colored, with warmer colours to indicated increasingly negative effect.</p

Escape mutations from CAP248-2B accumulate in the gp120 C terminus.

<p>(A) Sequence alignment of the gp120 C-terminus (positions 500–511) from CAP248 autologous viruses at nine weeks (study enrolment), 1, 2, 3 and 3.5 years post-infection. The primary (position 511) and secondary (position 504) gp160 cleavage sites are indicated with arrows. The total number of viruses with identical amino acid sequence within this region are indicated in brackets to the right. Residues undergoing significant selection pressure are indicated with the asterisks. (B) Sequence logograms showing variation within the gp120 C-terminus for all clades, and clade C only, from the LANL HIV sequence database, as well as from CAP248-2B at 3.5 years post-infection, colored and labelled as in A. The global frequencies for each of the autologous mutations identified in CAP248 sequences were: 500 (E8.88%, K46.56%, G3.56%), 502 (K79.35%, R18.73%, Q1.37%), 505 (V98.98%, A0.55%, M0.08%, L<0.01%), 507 (E47.5%, G3.87%, A0.7%), 508 (R98.24%, K1.49%), 509 (E95.11%, G1.53%, A0.27%). (C) Neutralization by CAP248-2B of the heterologous strain CAP45, when compared to gp120 C-terminal mutant viruses with changes identified from autologous CAP248 Env sequences. Data was plotted as percent inhibition (y-axis) against antibody concentration (x-axis). The wild-type virus is shown in black. Dotted lines indicate y-axis intersections for IC₈₀, IC₅₀, and IC₂₀. (D) Neutralization by CAP248-2B of CAP45 wild-type and mutant viruses with the additional gp41 changes identified from CAP248 autologous sequences, plotted as in C. (E) Binding to cleaved (solid bars) or uncleaved (speckled bars) cell-surface expressed Env measured by flow cytometry. Median fluorescence intensity (MFI) is shown on the y-axis, and Palivizumab was used as an HIV-1 negative control. (F) An SDS-PAGE gel of a single SOSIP trimer sample that was divided into two and subsequently captured from suspension by either CAP248-2B or CAP256-VRC26.09. Samples were run in the presence or absence of dithiothreitol (DTT) to assess the level of furin cleavage.</p

Crystal structure of CAP248-2B reveals an unusually long, protruding CDR-L3, with a hydrophobic tip.

<p>(A) Sequence alignment of the CAP248-2B heavy and light chain with their predicted V- and J-gene precursors. The CDRs are shaded, labelled, and colored. The heavy chain FR3 is similarly indicated in blue. (B) Crystal structure of the CAP248-2B Fab. The light and heavy chains are colored olive and forest green respectively, while CDR loops and FR-H3 are colored according to A. Two views are shown around a ~45° axis to highlight the long CDR-L3 (yellow). Insets show the conformational differences between the CDR-H3 and CDR-L3 loops between Fab structure 1 (shown here) and Fab structure 2 (shown in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006074#ppat.1006074.s001" target="_blank">S1 Fig</a>). Two Fabs were present in both asymmetric units, so four loops are shown per inset, two for Fab1 CDR-H3 (red) or L3 (yellow), and two for Fab2 CDR-H3 and L3 (both grey). Asp^100B and Asp^100C in the heavy chain and Phe^95C and Phe^95D in the light chain are shown with stick representations to highlight the conformational divergence between the two structures. Due to crystal packing all downstream analyses were based on the 3.1 Å Fab1 structure.</p

Escape mutations from CAP248-2B enhance the neutralization of broadly neutralizing antibodies that bind to gp41.

<p>(A) Neutralization of CAP45 and a mutant variant that includes the six gp120 C-terminal mutations identified in CAP248 autologous sequences (CS-Mut), by broadly neutralizing antibodies with epitopes in V2, V3, the CD4bs, the MPER, and the gp120-gp41 interface. Fold changes less than three are within the variation of the assay (no effect). Fold enhancement in neutralization sensitivity to CAP248-2B is indicated in orange (3–10 fold increased sensitivity) and red (> 10 fold increased sensitivity). Conferred resistance to the mutant virus at IC₅₀ is shown by grey shading. (B) Neutralization of five paired WT (blue) and CS-Mut (red) viruses by 35O22, 10E8, and 4E10.</p

The CAP248-2B CDR-L3 interacts with the viral membrane.

<p>Two docking orientations for the CAP248-2B Fab are modelled with (A) the CDR-L3 in close proximity to the viral membrane, and (B) the CDR-L3 in close proximity to the fusion peptide. The trimer is coloured as in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006074#ppat.1006074.g004" target="_blank">Fig 4</a>, and the Fab heavy and light chains shown in forest and olive green respectively, and the approximate location of the viral membrane is indicated with dotted lines. In the zoomed panel insets, the CDR-H1 (pink) and CDR-L3 (yellow) are shown in their predicted binding locations for each model. The fusion peptide is colored purple and shown with surface representation. (C) Neutralization of three heterologous viruses by CAP248-2B and related CDR-H1 and CDR-L3 mutants. Percentage inhibition was plotted on the y-axis versus antibody concentration on the x-axis. Dotted lines indicate y-axis intersections for IC₈₀, IC₅₀, and IC₂₀. (D) ELISA showing binding of CAP248-2B and related mutant antibodies to the BG505(gp120)-CAP45(gp41) chimeric SOSIP trimer. Absorbance readings are plotted on the y-axis and antibody concentration on the x-axis. CAP256-VRC26.09 and F105 are used are positive and negative control antibodies. (E) Anti-cardiolipin antibody ELISA, labelled as in D. (F) HEp-2 cell reactivity assays comparing a no antibody control to 50 μg/mL concentrations of either 4E10 (positive control), 35O22 (negative control), or CAP248-2B.</p

Isolated antibody CAP248-2B exhibits low neutralization plateaus, but recapitulates the plasma neutralization breadth.

<p>(A) Bar graph showing percentage neutralization breadth of CAP248 plasma at ID₅₀ titers of >1:100 (y-axis) on a 45 virus panel at one, two, and three years post-infection (x-axis). The total breadth at each time point is indicated above the bars. (B) Comparison between the neutralization breadth of CAP248 plasma at three years, and CAP248-2B at IC₂₀. The maximum percentage neutralization (neutralization plateau) reached by CAP248-2B is indicated. Palivizumab, a monoclonal antibody specific for Respiratory Syncytial Virus (RSV), was used as an IC₂₀ negative control. Titers are colored yellow, orange and red by potency. A strong inverse spearman correlation with a rho value of -0.802 (p-value <0.00001) indicates good concordance between CAP248 plasma ID₅₀ and CAP248-2B monoclonal antibody IC₂₀. (C) Percentage neutralization breadth of monoclonal antibody CAP248-2B (y-axis) on the same 45 virus panel as in A, when measured at IC₈₀, IC₅₀, and IC₂₀ (x-axis). (D) Neutralization curves of CAP248-2B against four viral strains (CAP45, CNE52, CAP228, and ZM249) plotted as percentage inhibition (y-axis) versus antibody concentration (x-axis). Dotted lines indicate y-axis intersections for IC₈₀, IC₅₀, and IC₂₀. The maximum inhibitory percentage achieved against each virus is listed to the right of each curve.</p