Protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and SARS-related emergent zoonotic coronaviruses is urgently needed. We made homotypic nanoparticles displaying the ...receptor binding domain (RBD) of SARS-CoV-2 or co-displaying SARS-CoV-2 RBD along with RBDs from animal betacoronaviruses that represent threats to humans (mosaic nanoparticles with four to eight distinct RBDs). Mice immunized with RBD nanoparticles, but not soluble antigen, elicited cross-reactive binding and neutralization responses. Mosaic RBD nanoparticles elicited antibodies with superior cross-reactive recognition of heterologous RBDs relative to sera from immunizations with homotypic SARS-CoV-2-RBD nanoparticles or COVID-19 convalescent human plasmas. Moreover, after priming, sera from mosaic RBD-immunized mice neutralized heterologous pseudotyped coronaviruses as well as or better than sera from homotypic SARS-CoV-2-RBD nanoparticle immunizations, demonstrating no loss of immunogenicity against particular RBDs resulting from co-display. A single immunization with mosaic RBD nanoparticles provides a potential strategy to simultaneously protect against SARS-CoV-2 and emerging zoonotic coronaviruses.
Current influenza vaccines do not elicit broadly protective immune responses against multiple strains. New strategies to focus the humoral immune response to conserved regions on influenza antigens ...are therefore required for recognition by broadly neutralizing antibodies. It has been suggested that B-cells with receptors that recognize conserved epitopes would be preferentially stimulated through avidity effects by mosaic particles presenting multiple forms of a variable antigen. We adapted SpyCatcher-based platforms, AP205 virus-like particles (VLPs) and mi3 nanoparticles (NPs), to covalently co-display SpyTagged hemagglutinin (HA) trimers from group 1 and group 2 influenza A strains. Here we show successful homotypic and heterotypic conjugation of up to 8 different HA trimers to both VLPs and NPs. We characterized the HA-VLPs and HA-NPs by cryo-electron tomography to derive the average number of conjugated HAs and their separation distances on particles, and compared immunizations of mosaic and homotypic particles in wild-type mice. Both types of HA particles elicited strong antibody responses, but the mosaic particles did not consistently elicit broader immune responses than mixtures of homotypic particles. We conclude that covalent attachment of HAs from currently-circulating influenza strains represents a viable alternative to current annual influenza vaccine strategies, but in the absence of further modifications, is unlikely to represent a method for making a universal influenza vaccine.
The HIV-1 envelope (Env) glycoprotein, a trimer of gp120–gp41 heterodimers, relies on conformational flexibility to function in fusing the viral and host membranes. Fusion is achieved after gp120 ...binds to CD4, the HIV-1 receptor, and a coreceptor, capturing an open conformational state in which the fusion machinery on gp41 gains access to the target cell membrane. In the well-characterized closed Env conformation, the gp120 V1V2 loops interact at the apex of the Env trimer. Less is known about the structure of the open CD4-bound state, in which the V1V2 loops must rearrange and separate to allow access to the coreceptor binding site. We identified two anti–HIV-1 antibodies, the coreceptor mimicking antibody 17b and the gp120–gp41 interface-spanning antibody 8ANC195, that can be added as Fabs to a soluble native-like Env trimer to stabilize it in a CD4-bound conformation. Here, we present an 8.9-Å cryo-electron microscopy structure of a BG505 Env–sCD4–17b–8ANC195 complex, which reveals large structural rearrangements in gp120, but small changes in gp41, compared with closed Env structures. The gp120 protomers are rotated and separated in the CD4-bound structure, and the three V1V2 loops are displaced by ∼40 Å from their positions at the trimer apex in closed Env to the sides of the trimer in positions adjacent to, and interacting with, the three bound CD4s. These results are relevant to understanding CD4-induced conformational changes leading to coreceptor binding and fusion, and HIV-1 Env conformational dynamics, and describe a target structure relevant to drug design and vaccine efforts.
Broadly neutralizing monoclonal antibodies protect against infection with HIV-1 in animal models, suggesting that a vaccine that elicits these antibodies would be protective in humans. However, it ...has not yet been possible to induce adequate serological responses by vaccination. Here, to activate B cells that express precursors of broadly neutralizing antibodies within polyclonal repertoires, we developed an immunogen, RC1, that facilitates the recognition of the variable loop 3 (V3)-glycan patch on the envelope protein of HIV-1. RC1 conceals non-conserved immunodominant regions by the addition of glycans and/or multimerization on virus-like particles. Immunization of mice, rabbits and rhesus macaques with RC1 elicited serological responses that targeted the V3-glycan patch. Antibody cloning and cryo-electron microscopy structures of antibody-envelope complexes confirmed that immunization with RC1 expands clones of B cells that carry the anti-V3-glycan patch antibodies, which resemble precursors of human broadly neutralizing antibodies. Thus, RC1 may be a suitable priming immunogen for sequential vaccination strategies in the context of polyclonal repertoires.
Antibody-mediated immunity plays a crucial role in protection against SARS-CoV-2 infection. We isolated a panel of neutralizing anti-receptor-binding domain (RBD) antibodies elicited upon natural ...infection and vaccination and showed that they recognize an immunogenic patch on the internal surface of the core RBD, which faces inwards and is hidden in the "down" state. These antibodies broadly neutralize wild type (Wuhan-Hu-1) SARS-CoV-2, Beta and Delta variants and some are effective against other sarbecoviruses. We observed a continuum of partially overlapping antibody epitopes from lower to upper part of the inner face of the RBD and some antibodies extend towards the receptor-binding motif. The majority of antibodies are substantially compromised by three mutational hotspots (S371L/F, S373P and S375F) in the lower part of the Omicron BA.1, BA.2 and BA.4/5 RBD. By contrast, antibody IY-2A induces a partial unfolding of this variable region and interacts with a conserved conformational epitope to tolerate all antigenic variations and neutralize diverse sarbecoviruses as well. This finding establishes that antibody recognition is not limited to the normal surface structures on the RBD. In conclusion, the delineation of functionally and structurally conserved RBD epitopes highlights potential vaccine and therapeutic candidates for COVID-19.
Many anti-severe acute respiratory syndrome coronavirus 2 (anti-SARS-CoV-2) neutralizing antibodies target the angiotensin-converting enzyme 2 (ACE2) binding site on viral spike receptor-binding ...domains (RBDs). Potent antibodies recognize exposed variable epitopes, often rendering them ineffective against other sarbecoviruses and SARS-CoV-2 variants. Class 4 anti-RBD antibodies against a less-exposed, but more-conserved, cryptic epitope could recognize newly emergent zoonotic sarbecoviruses and variants, but they usually show only weak neutralization potencies. Here, we characterize two class 4 anti-RBD antibodies derived from coronavirus disease 2019 (COVID-19) donors that exhibit breadth and potent neutralization of zoonotic coronaviruses and SARS-CoV-2 variants. C118-RBD and C022-RBD structures reveal orientations that extend from the cryptic epitope to occlude ACE2 binding and CDRH3-RBD main-chain H-bond interactions that extend an RBD β sheet, thus reducing sensitivity to RBD side-chain changes. A C118-spike trimer structure reveals rotated RBDs that allow access to the cryptic epitope and the potential for intra-spike crosslinking to increase avidity. These studies facilitate vaccine design and illustrate potential advantages of class 4 RBD-binding antibody therapeutics.
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•Donor-derived antibodies C118 and C022 recognize a highly conserved RBD epitope•Both antibodies cross-react and neutralize sarbecoviruses and SARS-CoV-2 VOCs•C118-RBD and C022-RBD crystal structures show long CDRH3s that extend RBD β sheet•C118-S cryo-EM structure suggests intra- and inter-spike crosslinking by C118 IgG
Jette et al. characterize antibodies derived from convalescent COVID-19 donors that broadly recognize sarbecoviruses and neutralize ACE2-tropic strains, including all SARS-CoV-2 variants of concern. Structures reveal binding to a highly conserved RBD epitope using long CDRH3 loops, with an orientation that inhibits ACE2 binding to the RBD and allows IgG avidity.
To combat future severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants and spillovers of SARS-like betacoronaviruses (sarbecoviruses) threatening global health, we designed mosaic ...nanoparticles that present randomly arranged sarbecovirus spike receptor-binding domains (RBDs) to elicit antibodies against epitopes that are conserved and relatively occluded rather than variable, immunodominant, and exposed. We compared immune responses elicited by mosaic-8 (SARS-CoV-2 and seven animal sarbecoviruses) and homotypic (only SARS-CoV-2) RBD nanoparticles in mice and macaques and observed stronger responses elicited by mosaic-8 to mismatched (not on nanoparticles) strains, including SARS-CoV and animal sarbecoviruses. Mosaic-8 immunization showed equivalent neutralization of SARS-CoV-2 variants, including Omicrons, and protected from SARS-CoV-2 and SARS-CoV challenges, whereas homotypic SARS-CoV-2 immunization protected only from SARS-CoV-2 challenge. Epitope mapping demonstrated increased targeting of conserved epitopes after mosaic-8 immunization. Together, these results suggest that mosaic-8 RBD nanoparticles could protect against SARS-CoV-2 variants and future sarbecovirus spillovers.
Members of the KDM5 histone H3 lysine 4 demethylase family are associated with therapeutic resistance, including endocrine resistance in breast cancer, but the underlying mechanism is poorly defined. ...Here we show that genetic deletion of KDM5A/B or inhibition of KDM5 activity increases sensitivity to anti-estrogens by modulating estrogen receptor (ER) signaling and by decreasing cellular transcriptomic heterogeneity. Higher KDM5B expression levels are associated with higher transcriptomic heterogeneity and poor prognosis in ER+ breast tumors. Single-cell RNA sequencing, cellular barcoding, and mathematical modeling demonstrate that endocrine resistance is due to selection for pre-existing genetically distinct cells, while KDM5 inhibitor resistance is acquired. Our findings highlight the importance of cellular phenotypic heterogeneity in therapeutic resistance and identify KDM5A/B as key regulators of this process.
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•KDM5 activity modulates response and resistance to endocrine therapies•Endocrine resistance is due to selection for pre-existing distinct cell populations•Acquired KDM5 inhibitor resistance is epigenetic, including gain of ER signaling•Transcriptomic but not genetic heterogeneity is associated with higher KDM5B
Hinohara et al. demonstrate that histone demethylases KDM5A and KDM5B are key regulators of phenotypic heterogeneity in estrogen receptor (ER)-positive breast cancer. Inhibition of KDM5 activity increases sensitivity to endocrine therapy by modulating ER signaling.
The goal of the gene normalization task is to link genes or gene products mentioned in the literature to biological databases. This is a key step in an accurate search of the biological literature. ...It is a challenging task, even for the human expert; genes are often described rather than referred to by gene symbol and, confusingly, one gene name may refer to different genes (often from different organisms). For BioCreative II, the task was to list the Entrez Gene identifiers for human genes or gene products mentioned in PubMed/MEDLINE abstracts. We selected abstracts associated with articles previously curated for human genes. We provided 281 expert-annotated abstracts containing 684 gene identifiers for training, and a blind test set of 262 documents containing 785 identifiers, with a gold standard created by expert annotators. Inter-annotator agreement was measured at over 90%.
Twenty groups submitted one to three runs each, for a total of 54 runs. Three systems achieved F-measures (balanced precision and recall) between 0.80 and 0.81. Combining the system outputs using simple voting schemes and classifiers obtained improved results; the best composite system achieved an F-measure of 0.92 with 10-fold cross-validation. A 'maximum recall' system based on the pooled responses of all participants gave a recall of 0.97 (with precision 0.23), identifying 763 out of 785 identifiers.
Major advances for the BioCreative II gene normalization task include broader participation (20 versus 8 teams) and a pooled system performance comparable to human experts, at over 90% agreement. These results show promise as tools to link the literature with biological databases.
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