Some individuals do not return to baseline health following SARS-CoV-2 infection, leading to a condition known as long COVID. The underlying pathophysiology of long COVID remains unknown. Given that autoantibodies have been found to play a role in severity of SARS-CoV-2 infection and certain other post-COVID sequelae, their potential role in long COVID is important to investigate. Here, we apply a well-established, unbiased, proteome-wide autoantibody detection technology (T7 phage-display assay with immunoprecipitation and next-generation sequencing, PhIP-Seq) to a robustly phenotyped cohort of 121 individuals with long COVID, 64 individuals with prior COVID-19 who reported full recovery, and 57 pre-COVID controls. While a distinct autoreactive signature was detected that separated individuals with prior SARS-CoV-2 infection from those never exposed to SARS-CoV-2, we did not detect patterns of autoreactivity that separated individuals with long COVID from individuals fully recovered from COVID-19. These data suggest that there are robust alterations in autoreactive antibody profiles due to infection; however, no association of autoreactive antibodies and long COVID was apparent by this assay.
Aaron Bodansky, Chung-Yu Wang, Aditi Saxena, Anthea Mitchell, Andrew F. Kung, Saki Takahashi, Khamal Anglin, Beatrice Huang, Rebecca Hoh, Scott Lu, Sarah A. Goldberg, Justin Romero, Brandon Tran, Raushun Kirtikar, Halle Grebe, Matthew So, Bryan Greenhouse, Matthew S. Durstenfeld, Priscilla Y. Hsue, Joanna Hellmuth, J. Daniel Kelly, Jeffrey N. Martin, Mark S. Anderson, Steven G. Deeks, Timothy J. Henrich, Joseph L. DeRisi, Michael J. Peluso
Given the COVID-19 pandemic, there is interest in understanding ligand-receptor features and targeted antibody-binding attributes against emerging SARS-CoV-2 variants. Here we developed a large-scale structure-based pipeline for analysis of protein-protein interactions regulating SARS-CoV-2 immune evasion. First, we generated computed structural models of the Spike protein of three SARS-CoV-2 variants (B.1.1.529, BA.2.12.1, and BA.5) bound either to a native receptor (ACE2) or to a large panel of targeted ligands (n=282), which included neutralizing or therapeutic monoclonal antibodies. Moreover, by using the Barnes Classification, we noted an overall loss of interfacial interactions (with gain-of-new-interactions in certain cases) at the receptor-binding domain (RBD) mediated by substituted residues for neutralizing complexes in Classes 1 and 2, whereas less destabilization was observed for Classes 3 and 4. Finally, an experimental validation of predicted weakened therapeutic antibody binding was performed on a cell-based assay. Compared to the original Omicron variant (B.1.1.529), derivative variants featured progressive destabilization of antibody-RBD interfaces mediated by a larger set of substituted residues, thereby providing a molecular basis for immune evasion. This approach and findings provide a framework for rapidly and efficiently generating structural models for SARS-CoV-2 variants bound to ligands of mechanistic and therapeutic value.
Joseph H. Lubin, Christopher Markosian, D. Balamurugan, Minh T. Ma, Chih-Hsiung Chen, Dongfang Liu, Renata Pasqualini, Wadih Arap, Stephen K. Burley, Sagar D. Khare
While the development of different vaccines slowed the dissemination of SARS-CoV-2, the occurrence of breakthrough infections has continued to fuel the COVID-19 pandemic. To at least secure partial protection in majority of the population through one dose of a COVID-19 vaccine, delayed administration of boosters has been implemented in many countries. However, waning immunity and emergence of new variants of SARS-CoV-2 suggest that such measures may induce breakthrough infections due to intermittent lapses in protection. Optimizing vaccine dosing schedules to ensure prolonged continuity in protection could thus help control the pandemic. We developed a mechanistic model of immune response to vaccines as an in-silico tool for dosing schedule optimization. The model was calibrated with clinical datasets of acquired immunity to COVID-19 mRNA vaccines in healthy and immunocompromised subjects and showed robust validation by accurately predicting neutralizing antibody kinetics in response to multiple doses of COVID-19 mRNA vaccines. Importantly, by estimating population vulnerability to breakthrough infections, we predicted tailored vaccination dosing schedules to minimize breakthrough infections, especially for immunocompromised subjects. We identified that the optimal vaccination schedules vary from CDC-recommended dosing, suggesting that the model is a valuable tool to optimize vaccine efficacy outcomes during future outbreaks.
Prashant Dogra, Carmine Schiavone, Zhihui Wang, Javier Ruiz-Ramírez, Sergio Caserta, Daniela I. Staquicini, Christopher Markosian, Jin Wang, H. Dirk Sostman, Renata Pasqualini, Wadih Arap, Vittorio Cristini
The pathogenesis of the marked pulmonary microvasculature injury, a distinguishing feature of COVID-19 acute respiratory distress syndrome (COVID-ARDS), remains unclear. Implicated in the pathophysiology of diverse diseases characterized by endothelial damage, including ARDS and ischemic cardiovascular disease, ceramide and in particular palmitoyl ceramide (C16:0-ceramide) may be involved in the microvascular injury in COVID-19. Using deidentified plasma and lung samples from COVID-19 patients, ceramide profiling by mass spectrometry was performed. Compared with healthy individuals, a specific 3-fold C16:0-ceramide elevation in COVID-19 patient plasma was identified. Compared with age-matched controls, autopsied lungs of individuals succumbing to COVID-ARDS displayed a massive 9-fold C16:0-ceramide elevation and exhibited a previously unrecognized microvascular ceramide-staining pattern and markedly enhanced apoptosis. In COVID-19 plasma and lungs, the C16-ceramide/C24-ceramide ratios were increased and reversed, respectively, consistent with increased risk of vascular injury. Indeed, exposure of primary human lung microvascular endothelial cell monolayers to C16:0-ceramide–rich plasma lipid extracts from COVID-19, but not healthy, individuals led to a significant decrease in endothelial barrier function. This effect was phenocopied by spiking healthy plasma lipid extracts with synthetic C16:0-ceramide and was inhibited by treatment with ceramide-neutralizing monoclonal antibody or single-chain variable fragment. These results indicate that C16:0-ceramide may be implicated in the vascular injury associated with COVID-19.
Irina Petrache, Elisabet Pujadas, Aditya Ganju, Karina A. Serban, Alexander Borowiec, Beatrice Babbs, Irina A. Bronova, Nicholas Egersdorf, Patrick S. Hume, Khushboo Goel, William J. Janssen, Evgeny V. Berdyshev, Carlos Cordon-Cardo, Richard Kolesnick
Immune responses in people with multiple sclerosis (pwMS) on disease-modifying therapies (DMTs) have been of significant interest throughout the COVID-19 pandemic. Lymphocyte-targeting immunotherapies including anti-CD20 treatments and sphingosine-1-phosphate receptor (S1PR) modulators attenuate antibody responses after vaccination. Evaluation of cellular responses after vaccination is therefore of particular importance in these populations. In this study, we analysed CD4 and CD8 T cell functional responses to SARS-CoV-2 spike peptides in healthy controls and pwMS on five different DMTs by flow cytometry. Although pwMS on rituximab and fingolimod therapies had low antibody responses after both two and three vaccine doses, T cell responses in pwMS on rituximab were preserved after a third vaccination, even when an additional dose of rituximab was administered between vaccine doses two and three. PwMS taking fingolimod had low detectable T cell responses in peripheral blood. CD4 and CD8 T cell responses to SARS-CoV-2 variants of concern Delta and Omicron were lower than to the ancestral Wuhan-Hu-1 variant. Our results indicate the importance of assessing both cellular and humoral responses after vaccination and suggest that even in the absence of robust antibody responses vaccination can generate immune responses in pwMS.
Asia-Sophia Wolf, Anthony Ravussin, Marton König, Mathias H. Øverås, Guri Solum, Ingrid Fadum Kjønstad, Adity Chopra, Trygve Holmøy, Hanne F. Harbo, Silje Watterdal Syversen, Kristin Kaasen Jørgensen, Einar A. Høgestøl, John T. Vaage, Elisabeth G. Celius, Fridtjof Lund-Johansen, Ludvig A. Munthe, Gro Owren Nygaard, Siri Mjaaland
BACKGROUND. The Omicron BA.5 subvariant of SARS-CoV-2 markedly escapes neutralizing antibodies induced by vaccination due to mutations in the Spike (S) protein. Solid organ transplant recipients (SOTRs) suffer high COVID-19 morbidity and demonstrate poor Omicron strain recognition after COVID-19 vaccination. T cell responses may provide a crucial second line of defense. Therefore, it is critical to understand which vaccine regimens induce robust, conserved T cell responses. METHODS. We evaluated anti-S IgG titers, subvariant pseudo-neutralization, and S-specific CD4+ and CD8+ T cell responses from SOTRs in a national, prospective observational trial (n=75). Participants were selected if they received 3 doses of mRNA (homologous boosting) or two doses of mRNA followed by Ad26.COV2.S (heterologous boosting). RESULTS. Homologous boosting with three mRNA doses induced the highest anti-S IgG titers. However, antibodies induced by both vaccine regimens demonstrated significantly lower pseudo-neutralization against BA.5 compared to the ancestral strain. In contrast, vaccine-induced S-specific T cells maintained cross-reactivity against BA.5 compared to ancestral recognition. Homologous boosting induced higher frequencies of activated polyfunctional CD4+ T cell responses, with polyfunctional IL-21+ peripheral T follicular helper cells increased in mRNA-1273 compared to BNT¬¬162b2. IL-21+ cells robustly correlated with antibody titers. Heterologous boosting with Ad26.COV2.S did not increase CD8+ responses compared to homologous boosting. CONCLUSIONS. These data demonstrate that boosting with the ancestral strain can induce cross-reactive T cell responses against emerging variants of concern in SOTRs, but alterative vaccine strategies are required to induce robust CD8+ T cell responses. TRIAL REGISTRATION. IRB00248540 FUNDING. U01AI138897, U54CA260492, Emory COVID-19 research repository
Elizabeth A. Thompson, Wabathi Ngecu, Laila Stoddart, T. Scott Johnston, Amy Chang, Katherine Cascino, Jennifer L. Alejo, Aura T. Abedon, Hady Samaha, Nadine Rouphael, Aaron A.R. Tobian, Dorry L. Segev, William A. Werbel, Andrew H. Karaba, Joel N. Blankson, Andrea L. Cox
Currently authorized COVID-19 vaccines induce humoral and cellular responses to epitopes in the SARS-CoV-2 spike protein, though the relative roles of antibodies and T cells in protection are not well understood. To understand the role of vaccine-elicited T cell responses in protection, we established a T cell–only vaccine using a DC-targeted lentiviral vector expressing single CD8+ T cell epitopes of the viral nucleocapsid, spike, and ORF1. Immunization of angiotensin-converting enzyme 2–transgenic mice with ex vivo lentiviral vector–transduced DCs or by direct injection of the vector induced the proliferation of functional antigen-specific CD8+ T cells, resulting in a 3-log decrease in virus load upon live virus challenge that was effective against the ancestral virus and Omicron variants. The Pfizer/BNT162b2 vaccine was also protective in mice, but the antibodies elicited did not cross-react on the Omicron variants, suggesting that the protection was mediated by T cells. The studies suggest that the T cell response plays an important role in vaccine protection. The findings suggest that the incorporation of additional T cell epitopes into current vaccines would increase their effectiveness and broaden protection.
Takuya Tada, Ju-Yi Peng, Belinda M. Dcosta, Nathaniel R. Landau
Multiple randomized, controlled clinical trials have yielded discordant results regarding the efficacy of convalescent plasma in outpatients, with some showing an approximate two-fold reduction in risk and others showing no effect. We quantified binding and neutralizing antibody levels in 492 of the 511 participants from the C3PO trial of a single unit of COVID-19 convalescent plasma (CCP) vs. saline infusion. In a subset of 70 participants, peripheral blood mononuclear cells were obtained to define the evolution of B and T cell responses through day 30. Binding and neutralizing antibody responses were measurably higher one hour post-infusion in recipients of CCP compared to saline plus multivitamin, but levels achieved by the native immune system by day 15 were much higher than seen immediately after CCP administration. Infusion of CCP did not block generation of the host antibody response or skew B or T cell phenotype or maturation. Activated CD4+ and CD8+ T cells were associated with more severe disease outcome. These data show that CCP leads to a measurable boost in anti-SARS-CoV-2 antibodies, but that the boost is modest and may not be sufficient to alter disease course.
John F. McDyer, Mahzad Azimpouran, Valerie L. Durkalski-Mauldin, Robert G. Clevenger, Sharon D. Yeatts, Xutao Deng, William Barsan, Robert Silbergleit, Nahed El Kassar, Iulia Popescu, Dimiter Dimitrov, Wei Li, Emily J. Lyons, Sophia C. Lieber, Mars Stone, Frederick K. Korley, Clifton W. Callaway, Larry J. Dumont, Philip J. Norris
People with HIV (PWH) appear at higher risk for suboptimal pathogen responses and worse COVID-19 outcomes, but the effects of host factors and COVID-19 on the humoral repertoire remain unclear. We assessed the antibody isotype/subclass and Fc-receptor binding Luminex arrays of non-SARS-CoV-2 and SARS-CoV-2 humoral responses among ART-treated PWH. Among the entire cohort, COVID-19 infection was associated with higher CMV responses (vs COVID-negative), potentially signifying increased susceptibility or a consequence of persistent inflammation. Among the COVID-positive, 1) higher BMI was associated with a striking amplification of SARS-CoV-2 responses, suggesting exaggerated inflammatory responses, and 2) lower nadir CD4 was associated with higher SARS-CoV-2 IgM and FcgRIIB binding capacity, indicating poorly functional extrafollicular and inhibitory responses. Among the COVID-negative, female sex, older age, and lower nadir CD4 were associated with unique repertoire shifts. In this first comprehensive assessment of the humoral repertoire in a global cohort of PWH, we identify distinct SARS-CoV-2-specific humoral immune profiles among PWH with obesity or lower nadir CD4+ T-cell count, underlining plausible mechanisms associated with worse COVID-19-related outcomes in this setting. Host factors associated with the humoral repertoire in the COVID-negative cohort enhance our understanding of these important shifts among PWH.
Samuel R. Schnittman, Wonyeong Jung, Kathleen V. Fitch, Markella V. Zanni, Sara McCallum, Jessica Shih-Lu Lee, Sally Shin, Brandon J. Davis, Evelynne S. Fulda, Marissa R. Diggs, Francoise Giguel, Romina Chinchay, Anandi N. Sheth, Carl J. Fichtenbaum, Carlos D. Malvestutto, Judith A. Aberg, Judith Currier, Douglas A. Lauffenburger, Pamela S. Douglas, Heather J. Ribaudo, Galit Alter, Steven K. Grinspoon
We assessed vaccine-induced antibody responses to the SARS-CoV2 ancestral virus and Omicron variant before and after booster immunization in 57 patients with B-cell malignancies. Over one third of vaccinated patients at the pre-booster timepoint were seronegative, and these patients were predominantly on active cancer therapies such as anti-CD20 monoclonal antibody. While booster immunization was able to induce detectable antibodies in a small fraction of seronegative patients, the overall booster benefit was disproportionately evident in patients already seropositive and not receiving active therapy. While ancestral and Omicron-reactive antibody levels among individual patients were largely concordant, neutralizing antibodies against Omicron tended to be reduced. Interestingly, in all patients, including those unable to generate detectable antibodies against SARS-CoV2 spike, we observed comparable levels of EBV and influenza reactive antibodies demonstrating that B cell-targeting therapies primarily impair de novo but not pre-existing antibody levels. These findings support rationale for vaccination prior to cancer treatment.
Joseph H. Azar, John P. Evans, Madison H. Sikorski, Karthik B. Chakravarthy, Selah McKenney, Ian Carmody, Cong Zeng, Rachael Teodorescu, No-Joon Song, Jamie L. Hamon, Donna Bucci, Maria Velegraki, Chelsea Bolyard, Kevin P. Weller, Sarah A. Reisinger, Seema A. Bhat, Kami J. Maddocks, Nathan Denlinger, Narendranath Epperla, Richard Gumina, Anastasia N. Vlasova, Eugene Oltz, Linda Saif, Dongjun Chung, Jennifer A. Woyach, Peter G. Shields, Shan-Lu Liu, Zihai Li, Mark P. Rubinstein
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