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Cancer patients, particularly those receiving B cell-depleting therapy for lymphoid malignancies, are at risk of prolonged SARS-CoV-2 infection, poorer clinical outcomes, and delayed initiation or disruption of cancer-directed therapy (Lee at al., 2022, Clark et al., 2021). We first studied T-cell mediated response to the Wuhan strain of SARS-CoV-2 in a cohort of 69 patients with hematologic and solid cancers, including 18 patients who received prior B-cell depleting therapy. Patients with prolonged COVID-19 clearance, defined by a positive PCR test for longer than 30 days, had a broad but poorly converged CD8+ dominant response and a lacking CD4+ response. To conduct this analysis, we performed bulk T-cell receptor (TCR) sequencing of 121 blood samples and tracked over time TCR repertoire statistics such as clonality, convergence, breadth, and depth of COVID-19-associated TCRs during the active and convalescent periods of COVID-19 infection. These SARS-CoV-2-associated TCRs were identified leveraging immunoSEQ T-MAP database (Snyder et al., 2020), a set of TCR sequences derived from COVID-19 patients and experimentally identified as responsive to MHC Class I and II epitopes from the Wuhan SARS-CoV-2 strain using the multiplex identification of TCR antigen assay (Klinger et al., 2015). To extend our TCR repertoire analysis to other SARS-CoV-2 variants, including Omicron, we developed a deep learning (DL) method to predict TCR specificities for new SARS-CoV-2 epitopes. This DL approach also permits the identification of SARS-CoV2-responsive TCRs private to an individual. Combining this DL approach with our TCR statistics methodology, we studied the dynamics of T-cell response to COVID-19 vaccinations in a cohort of 50 patients with cancer and analyzed TCR repertoire characteristics associated with different degrees of COVID-19 severity in a cohort of 42 cancer patients who contracted the Omicron. Understanding cellular response to novel infections is critical for patient care in the context of cancer, and our novel DL-based approach can leverage existing datasets to analyze and track response to emerging viral strains.
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Introduction Patients with hematological malignancies, including multiple myeloma (MM), experience suboptimal responses to SARS-CoV-2 vaccination. Monoclonal Gammopathy of Undetermined Significance (MGUS) and Smoldering Multiple Myeloma (SMM) are precursors to MM and exhibit altered immune cell composition and function. The SARS-CoV-2 pandemic and the subsequent population-wide vaccination represent an opportunity to study the real-life immune response to a common antigen. Here, we present updated results from the IMPACT study, a study we launched in November 2020 to characterize the effect of plasma cell premalignancy on response to SARS-CoV2 vaccination (vx). Methods We performed: (i) ELISA for SARS-CoV-2-specific antibodies on 1,887 peripheral blood (PB) samples (237 healthy donors (HD), and 550 MGUS, 947 SMM, and 153 MM patients) drawn preand post-vx;(ii) single-cell RNA, T cell receptor (TCR), and B cell receptor (BCR) sequencing (10x Genomics) on 224 PB samples (26 HD, and 20 MGUS, 48 SMM, and 24 MM patients) drawn preand post-vx;(iii) plasma cytokine profiling (Olink) on 106 PB samples (32 HD, and 38 MGUS and 36 SMM patients) drawn pre- and post-vx;and (iv) bulk TCR sequencing (Adaptive Biotechnologies) on 8 PB samples from 4 patients (2 MGUS, 2 SMM) drawn pre- and post-vx. Results Patients with MGUS and SMM achieved comparable antibody titers to HD two months post-vx. However, patient titers waned significantly faster, and 4 months post-vx we observed significantly lower titers in both MGUS (Wilcoxon rank-sum, p=0.030) and SMM (p=0.010). These results indicate impaired humoral immune response in patients with MGUS and SMM.At baseline, the TCR repertoire was significantly less diverse in patients with SMM compared to HD (Wilcoxon rank-sum, p=0.039), while no significant difference was observed in the BCR repertoire (p=0.095). Interestingly, a significant increase in TCR repertoire diversity was observed post-vx in patients with SMM (paired t-test, p=0.014), indicating rare T cell clone recruitment in response to vaccination. In both HD and patients, recruited clones showed upregulation of genes associated with CD4+ naive and memory T cells, suggesting preservation of the T cell response in SMM, which was confirmed by bulk TCR-sequencing in 4 patients.Lastly, by cytokine profiling, we observed a defect in IL-1beta and IL-18 induction post-vx in patients with SMM compared to HD (Wilcoxon rank-sum, p=0.047 and p=0.015, respectively), two key monocyte-derived mediators of acute inflammation, suggesting an altered innate immune response as well. Conclusion Taken together, our findings highlight that despite the absence of clinical manifestations, plasma cell premalignancy is associated with defects in both innate and adaptive immune responses. Therefore, patients with plasma cell premalignancy may require adjusted vaccination strategies for optimal immunization.
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Background: Hybrid immunity is more protective than vaccination or prior infection alone. To understand the formation of hybrid immunity, we studied how SARS-CoV-2 mRNA vaccines interact with T cell memory by tracking spike (S) specific T cells in cohorts of hospitalized (n = 19) or non-hospitalized (n = 34) COVID-19 convalescents. We hypothesized that S-reactive CD4 and CD8 T cells would increase in response to serial vaccine doses and reflect prior immune exposure at the clonal level. Method(s): After vaccination, we stimulated PBMCs from 12 participants (8M/4F) with peptides spanning S. Activated cells (CD69+CD137+) were sorted and CD4/CD8 phenotype linked with paired TRB-TRA sequences at single cell resolution. S-reactive TRB sequences were mapped within 4-6 serial blood and post-booster nasal TRB repertoires to evaluate S-reactive CD4 and CD8 T cell clonotypic kinetics spanning convalescence to boost. PBMCs from 53 participants were sequenced with the ImmunoSEQ assay to evaluate S-reactive TRB breadth using a database of S-assigned TRB sequences (Adaptive Biotechnologies), comparing S-reactive TRB diagnostic breadth by hospitalization status (Wilcoxon test). Result(s): SARS-CoV-2 mRNA vaccination provoked strong T cell clonal expansion in most participants. At 8-12 months after infection, each primary mRNA dose increased the abundance and diversity of S-specific T cells. Clonal and integrated expansions were larger in CD8 than in CD4 T cells. At the convalescent time point, we observed greater diagnostic S-reactive CD4 T cell breadth in hospitalized vs. non-hospitalized patients (p< 0.01). CD4 T cell S breadth was again higher in previously hospitalized persons after the 2nd primary (p=0.02) and booster (p< 0.01) doses, suggesting that diverse CD4 T cell memory after severe infection leads to increased repertoire diversity after vaccination. S-specific T cells with identical TCRs were detectable in blood and the nasal mucosa, with specificity confirmed using a TRA/TRB transgenic T cell with the matching receptor. Conclusion(s): Although both S-specific CD8 and CD4 T cell memory are established by prior infection, S-specific CD8 T cells predominated in blood after primary vaccination, with some clonotypes showing up to 1000-fold expansion across 1-2 mRNA doses. Vaccine-reactive CD8 clonotypes were present at the barrier nasal site after booster mRNA dosing. Severe disease imprinted a highly diverse S-reactive CD4 repertoire persisting through vaccination.
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Background: Allogeneic hematopoietic cell transplantation (HCT) with ex vivo T cell receptor (TCR) alphabeta+ T cell and CD19+ B cell depletion is an effective approach for children with primary immune deficiency disorders (PIDD) as it combines advantages of high CD34+ cell dose facilitating rapid engraftment with low risk of Graft Versus Host Disease (GVHD). The ideal pre-conditioning regimen that facilitates robust donor engraftment without increasing risk of transplant related mortality has not been well defined with this approach. Method(s): We report the outcomes of 4 pediatric subjects: Chronic Granulomatous Disease (CGD) (2), Wiskott Aldrich Syndrome (WAS) (1), and RAC2 deficient Severe Combined Immunodeficiency (1) who underwent haploidentical HCT with TCRalphabeta+ T cell/CD19+ depletion at Johns Hopkins All Children's Hospital/Moffitt Cancer Center from 2020-2022 (NCT04414046). Pre-conditioning regimen consisted of distal thymoglobulin (7.5 mg/kg), fludarabine (175 mg/m2), thiotepa (10 mg/kg) and pharmacokinetic guided busulfan targeting a cumulative area under curve (cAUC) (65-75 mgxhr/L). Rituximab (200 mg/m2) was administered on day +1. Result(s): The median age at HCT was 51 months (range 10-163 months). All patients received mobilized peripheral blood stem cells from HLA- haploidentical donors (paternal=1, maternal=1 sibling=2). Median busulfan cAUC for all patients was 69 mgxhr/L (range 65-76). Median CD34 and TCR alphabeta T cell dose was 9.13x106 cells/kg (range 7.0-18.9x106) and 0.7x105 cells/kg (range 0.09-1.0x105). Median times to neutrophil and platelet engraftment were 11 days (9-12) and 11 days (range 8-15), respectively. All 4 patients are alive with median follow-up of 19.5 months (range 7-24). One patient developed late VOD without organ dysfunction that resolved with defibrotide. At last follow up, peripheral T and myeloid chimerisms exceeded 90% in all 4 patients. Average time to CD4 recovery (> 200x106/L) was 142 days. Pre-existing inflammatory bowel disease in CGD (n=1) and WAS (n=1) patients resolved immediately following transplant. There was no graft failure, and none developed Grade III-IV acute or extensive chronic GVHD. Patient with WAS developed recurrent autoimmune cytopenias requiring corticosteroids, rituximab, sirolimus and daratumumab, and ultimately resolved. Viral reactivations included EBV (n= 1), adeno (n= 1), HHV6 (n= 2), BK (n=1), norovirus (n=1), and late HSV (n=1), all responded to antivirals without disease. All patients acquired SARS-Cov-2 after transplant and recovered without sequelae. Conclusion(s): TCR alphabeta+ and CD19+ depleted haploidentical transplantation using a reduced toxicity conditioning regimen with pharmacokinetic guided busulfan, fludarabine, thiotepa and thymoglobulin is well-tolerated in young children with PIDD that results in rapid, durable engraftment with low likelihood of GVHD and graft rejection.Copyright © 2023 American Society for Transplantation and Cellular Therapy
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Background: T cells play a critical role in the adaptive immune response to SARS-CoV-2 in both infection and vaccination. Identifying T cell epitopes and understanding how T cells recognize these epitopes can help inform future vaccine design and provide insight into T cell recognition of newly emerging variants. Here, we identified SARS-CoV-2 specific T cell epitopes, analyzed epitope-specific T cell repertoires, and characterized the potency and cross-reactivity of T cell clones across different common human coronaviruses (HCoVs). Method(s): SARS-CoV-2-specific T cell epitopes were determined by IFNgamma ELISpot using PBMC from convalescent individuals with mild/moderate disease (n=25 for Spike (S), Nucleocapsid (N) and Membrane (M)), and in vaccinated individuals (n=27 for S). Epitope-specific T cells were isolated based on activation markers following a 6-hour peptide stimulation, and scRNAseq was performed for TCR repertoire analysis. T cell lines were generated by expressing recombinant TCRs in Jurkat cells and activation was measured by CD69 upregulation. Result(s): We identified multiple immunodominant T cell epitopes across S, N and M proteins in convalescent individuals. In vaccinated individuals, we detected many of the same dominant S-specific epitopes at similar frequencies as compared to convalescent individuals. T cell responses to peptide S205 (amino acids 817-831) were observed in 56% and 59% of individuals following infection and vaccination, respectively, while 20% and 19% of individuals responded to S302 (a.a. 1205-1219) following infection and vaccination, respectively. For S205, a CD4+ T cell response, we confirmed 8 unique TCRs and determined the minimal epitope to be a 9mer (IEDLLFNKV). While TCR genes TRAV8-6*01 and TRBV30*01 were commonly utilized across the TCRs, we did identify TCRs with unique immunogenetic properties with different potencies of cross-reactivity to other HCoVs. For S302, a CD8+ T cell response, we identified two unique TCRs with different immunogenetic properties that recognized the same 9mer (YIKWPWYIW) and cross-reacted with different HCoV peptides (Figure 1). Conclusion(s): These data identify immunodominant T cell epitopes following SARS-CoV-2 infection and vaccination and provide a detailed analysis of epitope-specific TCR repertoires. The prospect of developing a vaccine that broadly protects against multiple human coronaviruses is bolstered by the identification of conserved immunodominant SARS-CoV-2 T cell epitopes that cross react with multiple other HCoVs.
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T-cell recognition of antigenic peptides presented by HLA molecules at the cell surface is critical for mounting an efficient adaptive immune response during acute viral infection such as COVID-19 caused by SARS-CoV-2. Recent data suggest that the depth of peptide coverage and the breadth of T cells that are able to respond are both important parameters associated with disease outcome. Strong T-cell responses against SARS-CoV-2 have also been reported in unexposed individuals, pointing to a possible role of heterologous immunity. In this study, we performed immunosequencing of the TCR CDR3beta region in a large cohort of 116 alloHSCT recipients and their corresponding healthy donors collected prior to the emergence of SARS-CoV-2. We used bioinformatics analyses and a large database of about 150,000 SARS-CoV-2 specific T-cell sequences in order to investigate the composition of the TCR repertoire regarding the presence of SARS-CoV-2 specific clonotypes in unexposed subjects among the more than 3.5 million CDR3beta sequences that we retrieved by immunosequencing. We also performed peptide binding predictions based on the reference proteome of the virus and by using the HLA class I high resolution typing data of the 116 patients. We could show that every individual is equipped with a large and diverse repertoire of clonotypes sharing their CDR3beta sequence with a SARS-CoV-2 specific T cell. Furthermore, the composition of the anti-SARSCoV- 2 repertoire was very similar among individuals, in healthy donors but also in the context of immune reconstitution in recipients, despite significant differences previously reported when accounting for the whole repertoire or for CMV-specific clonotypes only. In addition, each individual had the potential to cover a diverse repertoire of SARS-CoV-2 derived peptides (i.e., a few thousands strong and weak binders), but, interestingly, some interindividual differences were observed when only accounting for a strong affinity level of binding.
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T cells, and especially cytotoxic T cells are at the forefront of the fight against viral infection. The killer cells are able not only to distinguish between self and foreign peptides, but also to engage in the fight to clear the viral infection by eliminating the infected cells. Our lab is focused on understanding how T cells engage with viral peptide antigens, that are presented by highly polymorphic HLA molecules. T cells have receptors on their surface called T cell receptors (TCRs) that allow them to recognize the composite surface of the peptide- HLA complex. Using x-ray crystallography we can understand at the atomic level both peptide antigens presentation and TCR recognition, both important to determine the quality of the subsequent immune response. We can then link that structural information with our cellular assay that determines the strength and magnitude of the anti-viral response, providing the basis for peptide modification to reach stronger response or an understanding of viral mutation that led to viral escape. Our current work compared the T cell response, at the antigen level against 32 single epitope derived from spike, between COVID-19 recovered and vaccinated donors. We have shown that the booster shot (3rd dose) increases the antigen-specific T cell response, increases the level of T cell cross-reactivity against variant of SARS-CoV-2, but also alters the phenotype of the T cell. Those results are important to future guide vaccination advise and better understand the immune response to SARS-CoV-2 infection. POSTER PRESENTATIONS Autoimmunity, Infection, Reproduction and Cancer.
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This study aimed to review zinc's effectiveness as an antivirus in treating herpes simplex virus infection. The authors use international journals published from 2000-2022, and use search engines such as Google Scholar, PubMed, and Science Direct with the keywords "zinc and herpes simplex virus". The herpes simplex virus that often causes symptoms in humans are HSV type 1 and type 2. The lesions appear as vesicles which then rupture into ulcers. Zinc is one of the most abundant nutrients or metals in the human body besides iron. Studies about the effects of zinc on HSV have shown that it has the function of inhibiting the viral life cycle. HSV attaches to the host cells to replicate and synthesize new viral proteins. Zinc can inhibit this process by depositing on the surface of the virion and inactivating the enzymatic function which is required for the attachment to the host cell, disrupting the surface glycoprotein of the viral membrane so it could not adhere and carry out the next life cycle, it can also inhibit the function of DNA polymerase that works for viral replication in the host cell. This article showed that zinc has effectiveness as an antivirus against the herpes simplex virus, therefore, patients infected with HSV can be treated with zinc as an alternative to an antivirus drug.Copyright © 2022 The Authors. Published by Innovare Academic Sciences Pvt Ltd.
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Objetivos: As neoplasias Linfoproliferativas (NLP) T sao um grupo heterogeneo de doencas oncohematologicas cujo diagnostico e desafiador devido ao amplo espectro de achados clinicos, morfologicos e histologicos que frequentemente se sobrepoem aos encontrados em condicoes reacionais. A determinacao de clonalidade e necessaria e pode ser realizada por deteccao dos rearranjos de genes do TCR por PCR ou NGS. No entanto, tais tecnicas apresentam alto custo e tempo prolongado para resultado. A imunofenotipagem por citometria de fluxo (CMF) e utilizada rotineiramente para diagnostico das NLP. O TCR e composto por cadeias alfa e beta com dois dominios constante da cadeia beta, TRBC 1 e TRBC2 que sao mutuamente exclusivos. O anticorpo JOVI-1 apresenta alta especificidade contra o TRBC1. Celulas normais ou reacionais demonstram um misto de positividade e negatividade para TRBC1, ou seja, um perfil policlonal, enquanto celulas T monoclonais demonstram expressao restrita de algum TRBC. A inclusao do anticorpo JOVI-1 nos paineis de CMF e, portanto, atrativa para determinacao da clonalidade das NLP-T. O objetivo desse trabalho e avaliar e reportar a inclusao do JOVI-1 nos paineis de CMF em casos suspeitos de NLP-T. Materiais e Metodos: Avaliacao de perfil de expressao de TCR Beta 1 atraves da inclusao com anticorpo JOVI-1 aos paineis utilizados em pesquisa de NLP-T. O JOVI-1 foi tambem utilizado na avaliacao de Leucemias Agudas T (LLA) de forma a ser um grupo controle para o ensaio, uma vez que linfoblastos imaturos nao apresentam expressao de TCR. Resultados: O JOVI-1 foi realizada em 18 casos suspeitos de NLP. 6 amostras de medula ossea e 12 de sangue periferico. A idade mediana dos suspeitos foi de 50,5 anos (1-71 anos) e a proporcao de individuos do genero masculino para o feminino foi de 2,4:1,0. 10 casos (55%) demonstraram perfil policlonal sendo que em 3 desses casos confirmaram-se infeccoes virais (1 por Covid-19 e 2 por Citomegalovirus) e 7 tiveram outros diagnosticos que nao NLP. Ja em 45% dos casos observou-se perfil monoclonal: 4 diagnosticos de Leucemia de Grandes Linfocitos Granulares T, 2 diagnosticos de Linfoma/Leucemia de Celulas T do Adulto, 1 diagnostico de Sindrome de Sezary e 1 caso de Linfoma T gama-delta. Foram avaliados 6 casos de LLA T, no qual apenas um foi positivo para o marcador e tratava-se de LLA T madura. Discussao e Conclusao: Nesse trabalho demonstramos que a adicao do JOVI-1 ao painel habitual de diagnostico torna a CMF uma boa ferramenta diagnostica para NLP-T cronicas obtendo estudo de clonalidade de forma relativamente simples e rapida, similar ao que acontece nas NLP de linhagem B ha decadas. Copyright © 2022
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SESSION TITLE: Unique Inflammatory and Autoimmune Complications of COVID-19 Infections SESSION TYPE: Rapid Fire Case Reports PRESENTED ON: 10/19/2022 12:45 pm - 1:45 pm INTRODUCTION: Viral infections can induce an immune cascade which may incite varied autoimmune disease to take action. One such disease is dermatomyositis, a rare inflammatory disease with multisystemic involvement. As we enter the post pandemic era, several unique complications related to COVID-19 are now surfacing. Here we present a case of a 57-year-old female who developed dermatomyositis after recent Covid-19 infection. CASE PRESENTATION: Patient is a 57-year-old female who presented to our pulmonary clinic with complaints of cough and shortness of breath (SOB). Patient reported feeling ill since contracting COVID-19 in November 2020. Two months after being diagnosed with COVID-19 she started to experience generalized muscle and joint pain;she underwent extensive rheumatological workup which was consistent with Sjogren disease for which she was started on hydroxychloroquine. A month after initiation of medication she started to experience worsening cough and SOB and underwent pulmonary function testing (PFT) in our clinic which showed evidence of restrictive lung disease. Chest CT was consistent with interstitial changes, therefore a diagnosis of ILD (interstitial lung disease) due to connective tissue disorder was made. Further assessment revealed positive CPK and anti-Jo1 antibodies indicative of dermatomyositis as a cause of her ILD. She was started on oral steroids which helped improve her symptoms. DISCUSSION: Several viruses including EBV, Hepatitis C, Rubella, HTLV-1 and Parvovirus have been associated with development of autoimmune diseases. Viral infections, like COVID-19 have shown to trigger an intense immune response which in turn may lead to autoimmune activity against host antigen. SARS-CoV2 has been found to enter muscle cells through ACE-2 receptors, allowing for transfer of genetic material and skeletal muscle damage. Another proposed mechanism of COVID induced myopathy has been T-cell clonal expansion by the virus up regulating TLR4 receptors increasing expression of ACE2, therefore facilitating entry of viruses leading to further inflammation. Identification of very specific T cell receptor epitopes for SARS-COv2 in patients with dermatomyositis has suggested COVID-19 as a trigger for CD8-T cells which leads to dermatomyositis in these patients. Autoimmune reactions occur from varying mechanisms like epitope spreading and bystander activation to molecular mimicry triggered by viral infections. CONCLUSIONS: SARS-COv2 presented with several challenges in the field of medicine. As we enter the post COVID period in medicine, we will continue to face several challenges proposed by the inflammatory surge caused by this disease. It is therefore important clinicians recognize and report these rare cases to increase awareness regarding several post covid diseases. Reference #1: HUSSEIN, H. M.;RAHAL, E. A. The role of viral infections in the development of autoimmune diseases. Critical Reviews in Microbiology, [s. l.], v. 45, n. 4, p. 394–412, 2019. DOI 10.1080/1040841X.2019.1614904. Disponível em: https://search-ebscohost-com.proxy.lib.wayne.edu/login.aspx?direct=true&db=a9h&AN=138199390&site=ehost-live&scope=site. Acesso em: 4 abr. 2022. DISCLOSURES: No relevant relationships by Kevser Akyuz No relevant relationships by Ranim Chamseddin No relevant relationships by Padmini Giri No relevant relationships by verisha khanam No relevant relationships by Emad Shehada No relevant relationships by Abdullah Yesilyaprak
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Background: The role of T cell immunity in protection against COVID-19 in immunosuppressed patients who failed to mount serological responses remains ill defined. Hypothesis: Vaccine-based intradermal skin test (IDT) serves as a surrogate marker of T cell responses in seronegative immunosuppressed patients. Methods: We compared anti-SARS-CoV-2 antibodies and cellular responses in vaccinated immunosuppressed (IS) patients (n = 58), healthy unvaccinated naive controls (NC, n = 8) and healthy vaccinated controls (VC, n = 32) by Luminex, IFN-γ ELIPSOT and IDT 3 to 6 months after vaccination. In 3 VC we performed a skin biopsy 24h after IDT and performed single-cell RNAseq of the skin-infiltrating CD45+ cells Results: Seronegative NC had no detectable T cell responses and negative IDR, whereas VC had anti-SARS-CoV-2 antibodies (100%), positive ELIPSOT (90%) and IDR (90%). Overall IS patients had significantly less antibodies up to 39 weeks after vaccination compared to VC but similar ELIPSOT responses. ELISPOT was positive in 33.3 % and 66.6 % and IDR in 62.5% and 90.5% of seronegative vs seropositive IS patients respectively. Conversely, patients with negative IDR had significantly lower T cell responses and IgG titers than those with positive IDR. Importantly, the TCR repertoire of infiltrating skin lymphocytes revealed 18/1064 clonotypes with known specificities against SARS-CoV-2. Conclusion: Our results indicate that local reaction to IDR is partially composed of SARS-CoV-2-specific T cells. IDR represents a promising tool to cost-effectively monitor SARS-CoV-2 specific T cell immunity in IS patients.
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Patients with haematological malignancies are prioritized for COVID-19 vaccine due to their high risk for SARS-CoV-2 infection related disease severity and mortality. Immune defects associated with malignancy and their treatment could influence vaccine driven immune response in these cancer patients. Thus, to understand T cell immunity, its long-term persistence, and correlation with antibody response, we evaluated the BNT162b2 SARS-CoV-2 mRNA vaccine-specific immune response in chronic lymphocytic leukaemia (CLL) and myeloid dysplastic syndrome (MDS) patients. Longitudinal analysis of CD8+ T cells using DNA-barcoded peptide-MHC multimers covering the full SARS-CoV-2 spike-protein showed vaccine-specific T cell activation and persistence of memory T cells up to 6 months post-vaccination. Surprisingly, a higher frequency of vaccine-induced antigen-specific CD8+ T cell was observed in the patient group compared to the healthy donor group. Furthermore, and importantly, immunization with the second booster dose significantly increased the frequency of antigen-specific CD8+ T cells as well as the total number of T cell specificities. Altogether, 23 immunogenic epitopes were identified and a strong immunodominance was observed for the two of the spike-specific T cell epitopes restricted to HLA-A24 (NYNYLYRLF) and HLA-A2 (YLQPRTFLL) with a prevalence of 100% and 80% respectively. In summary, we mapped the vaccine-induced antigen-specific CD8+ T cells and showed a booster-specific activation and enrichment of memory T cells that could be crucial for long-term disease protection for this patient group.
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Background: Vaccination against SARS-CoV-2 is a highly effective strategy to protect against infection, which is predominantly mediated by vaccine-induced antibodies. Postvaccination antibodies are robustly produced by those with inflammatory bowel disease (IBD) even on immune-modifying therapies but are blunted by anti-TNF therapy. In contrast, T-cell response which primarily determines long-term efficacy against disease progression, , is less well understood. We aimed to assess the post-vaccination T-cell response and its relationship to antibody responses in patients with inflammatory bowel disease (IBD) on immunemodifying therapies. Methods: We evaluated IBD patients who completed SARS-CoV-2 vaccination using samples collected at four time points (dose 1, dose 2, 2 weeks after dose 2, 8 weeks after dose 2). T-cell clonal analysis was performed by T-cell Receptor (TCR) immunosequencing. The breadth (number of unique sequences to a given protein) and depth (relative abundance of all the unique sequences to a given protein) of the T-cell clonal response were quantified using reference datasets and were compared to antibody responses. Results: Overall, 303 subjects were included (55% female;5% with prior COVID) (Table). 53% received BNT262b (Pfizer), 42% mRNA-1273 (Moderna) and 5% Ad26CoV2 (J&J). The Spike-specific clonal response peaked 2 weeks after completion of the vaccine regimen (3- and 5-fold for breadth and depth, respectively);no changes were seen for non-Spike clones, suggesting vaccine specificity. Reduced T-cell clonal depth was associated with chronologic age, male sex, and immunomodulator treatment, and was preserved by nonanti- TNF biologic therapies;augmented clonal depth was associated with anti-TNF treatment (Figure). TCR depth and breadth were associated with vaccine type;after adjusting for age and gender, Ad26CoV2 (J&J) exhibited weaker metrics than mRNA-1273 (Moderna) (p= 0.01 for each) or BNT262b (Pfizer) (p=0.056 for depth). Antibody and T-cell responses were only modestly correlated;while those with robust humoral responses also had robust TCR clonal expansion, a substantial fraction of patients with high antibody levels had only a minimal T-cell clonal response (Figure). Conclusion: Age, sex and select immunotherapies are associated with the T-cell clonal response to SARS-CoV-2 vaccines, and T-cell responses are low in many patients despite high antibody levels. These factors, as well as differences seen by vaccine type may help guide reimmunization vaccine strategy in immune-impaired populations. Further study of the effects of anti-TNF therapy on vaccine responses are warranted. (Table Presented)
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BACKGROUND: In response to COVID-19 vaccination, cytotoxic and cytokine effector T cell immune responses are elicited in the T-cell compartment, based on recognition of epitopes presented by Class I or Class II MHC molecules, respectively. The levels of these distinct T-cell responses may have significant implications for immunization strategies and risk assessment. Knowledge of these two responses after vaccination is still largely unknown, especially in the context of immunomodulatory treatment regimens. METHODS: We performed T-cell receptor (TCR) immunosequencing (Adaptive Biotechnologies, Seattle WA) of IBD patients (N=303) and health care worker controls (HCW, N=224) at up to four time points (prior to dose 1, prior to dose 2, 2 weeks after dose 2, 8 weeks after dose 2). Two metrics of TCR response, breadth (# of unique antigen-specific sequences) and depth (expansion of antigen-specific sequences), were calculated for all sequences and Class I- and Class II-specific sequences, and compared to demographics, IBD treatment, and vaccine type. Subjects with exceptional Class I or Class II responses were calculated as significant residuals relative to the Class I vs. Class II regression line. Similar associations were observed for both breadth and depth: breadth is presented here for brevity. RESULTS: Both Class I- and Class II-specific T-cell responses peaked 2 weeks after dose 2, and significantly correlated with lower age, female gender, and mRNA vaccine type (mRNA-1273/Moderna and BNT262b/Pfizer, versus vector vaccine AD26CoV2/J&J) (FIGURE). Class II responses comprised ~85% of detected TCR response in both IBD and HCW subjects. Among IBD patients, there was a significant elevation of the class I response with anti-TNF treatment (p=0.04). This effect was most pronounced at later timepoints, suggesting that anti-TNF permitted a more persistent Class I-specific response. Among patients with exceptionally high or low Class I TCR response, there were significant differences in TCR metrics across vaccine types (p=0.0035). 21% of AD26CoV2 patients were highly Class I-biased (Zscore>1, 9.4% and 7.3% for BNT162 and mRNA-1273, respectively), and this was correlated with lower anti-spike serology 2 and 8 weeks after vaccination (p<1E-10). Conversely, mRNA- 1273 patients were Class I-deficient, representing 25.3% of patients but 44.1% of highly Class I-deficient patients (Zscore<1, 0% for AD26CoV2). CONCLUSION: The T-cell clonal response to SARS-CoV-2 vaccine is Class II-predominant, but the Class I-response is augmented by anti-TNF therapy and vector vaccine type. These factors may help guide reimmunization vaccine strategy in immune-impaired populations, and warrant further study of the effects of anti-TNF therapies on vaccine efficacy.(Figure Presented)Figure: TCR response time course (left);effect of anti-TNF (middle);effect of vaccine type (right). Breadth was predominantly Class II for most patients, with maximum response at 2 weeks after full vaccination (left). The balance of Class I vs. Class II response was significantly biased towards Class I at 8 weeks after full vaccination for patients receiving anti-TNF treatment for IBD (asterisk, p=0.036). Patients receiving AD26CoV2 vaccines were significantly increased in Class I responses, while patients receiving mRNA-1273 vaccines were significantly reduced for Class I responses (t-tests: p=0.0036 at 8 weeks [asterisk], p=0.051 at 2 weeks).
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Background: Inflammatory bowel disease (IBD) is a chronic inflammatory disorder of the gastrointestinal tract characterized by immune dysregulation and decreased T cell receptor (TCR) repertoire diversity. Patients with immune-mediated disorders such as IBD have attenuated convalescent antibody responses after COVID-19 infection. We sought to understand the immune configuration associated with high versus low convalescent SARS-CoV- 2 antibodies in patients with IBD using single-cell immunophenotyping. Methods: We performed a study of 9 patients with IBD who were SARS-CoV-2 convalescent (recovered from COVID-19 and converted RNA positive to negative) and 9 matched SARS-CoV-2 naïve controls (no prior COVID-19, confirmed RNA negative). We measured plasma SARS-CoV- 2 antibody (N protein IgG, S1RBD IgG, S1RBD IgA) levels from patients with IBD two months after recovering from COVID-19 (RNA negative). We selected three patients with the highest SARS-CoV-2 antibodies and three matched (for age, sex, IBD subtype and disease activity, medications, COVID-19 severity) patients with the lowest antibodies and performed their peripheral blood mononuclear cell (PBMC) single-cell transcriptomics with paired TCR and BCR sequencing using 10X Genomics. Normalization, dimensionality reduction, and clustering were performed using Seurat. TCR and BCR immune repertoire analyses were performed using Immunarch. Results: SARS-CoV-2 convalescent patients with IBD had detectable but variable SARS-CoV-2 antibody levels (range 0-469 U/mL), whereas SARSCoV- 2 naïve IBD patients had no detectable antibodies. The mean SARS-CoV-2 antibody concentration among the three IBD patients with the highest and three patients with the lowest groups differed by more than 10-fold (206.0 vs 17.5 U/mL, P<0.001). PBMC singlecell immunophenotyping revealed decreased naïve CD4+ T cell and increased CD14+ monocyte and memory CD4+ T cell proportions in IBD patients in the low versus high SARSCoV- 2 antibody group. There were higher numbers of HLA-DQA1+ B cells and CD8 T cells and lower GPR183+ B cells and CD8 T cells in the high SARS-CoV-2 antibody group. There was a trend towards decreased TCR and BCR repertoire diversity in the low SARS-COV-2 antibody group. Finally, we identified immunoglobulin gene signatures (IGHV1-69D/IGLV3- 25, IGHV3-48, IGHV3-7/IGKV41/IGLV1-47, IGHV3-7/IGKV4-1, IGHV3-7/IGKV4-44) that were enriched only in the high SARS-CoV-2 antibody group. Conclusions: Single-cell immunophenotyping of PBMC from convalescent patients with IBD reveal differences in CD4+ T cell, CD14+ monocyte, and HLA-DQA1+ and GPR183+ B and CD8 T cell immunophenotypes, immune repertoire diversity, and immunoglobulin gene signatures in patients with high versus low SARS-CoV-2 antibody levels.(Figure Presented)Figure 1. SARS-COV-2 Antibodies in Convalescent Patients with IBD and Single-Cell Immunophenotypes. A) SARS-COV-2 antibody levels in COVID-19 convalescent versus SARS-CoV-2 naïve patients with IBD B) T-SNE plot of PBMC immunophenotypes in all convalescent patients with IBD C) Differences in proportion of single-cell PBMC immunophenotypes in high versus low SARS-COV-2 antibody patients D) Differences in HLA-DQA1 and GPR183 immunophenotypes in high versus low SARS-COV-2 antibody patients.
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Background: Vaccine-induced protection against SARS-CoV-2 infection is predominantly mediated by humoral immunity;protection against disease progression is primarily determined by cellular immunity. Patients with inflammatory bowel disease (IBD) have high rates of post-vaccination anti-Spike IgG [IgG(S)] seroconversion, but postvaccination immune responses relative to non-IBD controls have not been well described. We aimed to assess post-vaccination humoral (antibody) and cellular (T-cell) responses in IBD relative to healthcare worker (HCW) controls. Methods: We evaluated IBD patients enrolled in a US registry referred from 26 centers at 2, 8, and 16 weeks after completing 2 doses of SARSCoV- 2 mRNA vaccination and compared results to non-IBD non-immunosuppressed HCW participating in a parallel study. We analyzed plasma antibodies to the receptor binding domain of the viral spike protein using the SARS-CoV-2 IgG-II assay (Abbott Labs, Abbott Park, IL);IgG(S) > 50 AU/mL was defined as positive. Those with prior COVID were excluded. We also performed T-cell clonal analysis by T-cell receptor (TCR) immunosequencing at 8 weeks (Adaptive Biotechnologies, Seattle, WA). The breadth (number of unique sequences to a given protein) and depth (relative abundance of all the unique sequences to a given protein) of the T-cell clonal response were quantified using reference datasets. Analyses were adjusted for age, sex and vaccine type. Results: Overall, 1805 subjects were included (IBD n=1074 (65% Crohn's disease, 35% ulcerative colitis);HCW n=731). Age and sex were similar between both cohorts;Hispanic ethnicity and Asian race were less common among IBD than HCW (Table). Vaccine type included BNT162b2 (Pfizer) (75% of IBD, 98% of HCW) and the remainder mRNA-1274 (Moderna). IBD treatments included anti- TNF (46%), other biologics (33%), other immune suppressing therapy (9%), and no immune suppression (12%). Postvaccination antibody levels were lower among IBD than HCW both before and after adjusting for vaccine type (p<0.0001 each timepoint;Figure). After further restricting the IBD cohort to those on no immune-suppressive therapies, antibodies remained lower in IBD vs HCW at 2w (p=0.008) and 8w (p<0.0001), but not 16w (p=0.07). Among 321 subjects with available whole cell samples at 8 weeks (IBD n=163, HCW =158), Spikespecific TCR responses were similar between IBD and HCW for both clonal breadth and depth in both unadjusted and adjusted analyses;sub-analyses of those on biologics yielded similar results. Conclusion: Patients with IBD have dampened humoral responses, but similar cellular responses, after SARS-CoV-2 mRNA vaccination relative to HCW. These findings suggest a potentially greater risk of infection, but not of disease progression, among those with IBD, and should be considered to help guide booster dosing strategies for the IBD population. (Figure Presented) (Figure Presented) Figure: Post-vaccination immune responses: (A) Antibody responses are lower in IBD relative to non-IBD healthcare workers at 2, 8, and 16 weeks (p<0.0001 at each timepoint). In contrast, post-vaccination Spike-specific T-cell receptor clonal breadth (B1) and clonal depth (B2) at 8 weeks are similar in IBD compared to healthcare workers.
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RATIONALE: The proteomic responses of hospitalized patients with SARS Co-V-2 infection may provide insight into risk, time course, and mechanisms associated with this infection. We used a high throughput proteomic platform to examine proteins that were differentially expressed relative to the length of hospital stay (LOS). METHOD:26 patients, hospitalized with SARS CoV-2 infection (mean age 48 yrs, 44% women) had blood samples obtained within 72 hours of admission. Initial plasma samples were analyzed from patients who were hospitalized for < 3 days (n=6), < 7days (n=12) and > 7 days (n=8) of LOS and compared to healthy controls (HC, n=8). Samples were analyzed with the modified aptamer-based array (SomaScan) that measures more than 7,000 human proteins representing different molecular pathways and gene families. Differentially regulated proteins with > 1.5 fold change and a false discovery rate of 5% were analyzed using the Ingenuity Pathway Analysis (IPA). Unique protein categories associated with LOS were assessed. RESULT: Compared to HC, differentially expressed proteins were detected among the 3 groups: 461 at < 3 days, 1,635 proteins at < 7 days and 1,738 proteins in >7 days. 407 proteins were common among all hospitalized COVID 19 individuals independent of LOS and 12, 250 and 361 proteins were uniquely present at < 3 days, < 7 days and > 7 days respectively compared to HC. The table below demonstrates the top highly enriched canonical pathway, molecular function and upstream regulator of differentially expressed proteins. The temporal sequence of these protein networks varied with LOS. Representative examples include early responses;platelet membrane glycoprotein GP6 signaling pathway that involves the FcR gamma-chain and the Src kinases linked to platelet aggregation, signaling involved in T cell receptor-mediated IL-2 production (TEC kinase). Less than 7 days include diacylglycerol associated with T cell activation, carnitine palmitoyltransferase associated with mitochondrial beta-oxidation of long chain fatty acids. CXCR4 a receptor for stromal -cell derived factor 1 and associated with COVID-19 prognosis. Late responses after 7 days include pathways involved in remodeling of epithelial adherens junctions. CONCLUSIONS : A high throughput proteomic approach provides insight into the dynamic regulation of protein pathways associated with the progression of SARS-Co-V2 infection. This may provide additional insight into risk and mechanisms associated with outcomes in COVID. (Table Presented).
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Background: Recent studies have shown that vaccinated individuals harbor cross-reactive T cell responses that can cross-recognize SARS-CoV-2 and endemic human common cold coronaviruses (HCoVs). However, it is still unknown whether CD4+ T cells from vaccinated individuals recognize peptides from bat coronaviruses that may have the potential of causing future pandemics. In this study, we identified a SARS-CoV-2 spike protein epitope (S815-827) that is conserved in coronaviruses from different genera and subgenera including SARS-CoV, MERS-CoV, multiple bat coronaviruses and a feline coronavirus. We hypothesized that S815-827 is recognized by vaccinated individuals, and that S815-827-reactive T cells can cross-recognize homologues bat coronaviruses. Methods: To evaluate CD4+ T cell responses, we isolated CD8 depleted PBMCs from COVID-19 vaccinated individuals and performed IFN-γ ELISPOT assays. To assess T cell cross-reactivity, S815-827-reactive T cell lines were re-stimulated with homologous coronavirus peptides and cytokine production was assessed with flow cytometry. Additionally, the Vira-FEST assay (which utilizes TCR Vβ CDR3 sequencing) was performed to identify cross-reactive CD4+ T cell clones. Statistical comparisons were done using Mann-Whitney test, Wilcoxon matched-pairs signed rank test or Friedman test with Dunn's multiple comparison as appropriate. Results: Our results show that 16 out of 38 (42%) of vaccinated participants in our study who received the Pfizer-BioNTech (BNT162b2) or Moderna (mRNA-1273) COVID-19 vaccines had robust CD4+ T cell responses to S815-827. All responders also recognized homologous peptides from at least 2 other coronaviruses, and 8 out of 11 responders recognized peptides from at least 6 out of the 9 other coronaviruses tested. To determine T cell cross-reactivity, we re-stimulated S815-827 specific T cell lines with homologous coronavirus peptides. We found that S815-827 specific T cells had a robust increase in IFN-γ+ TNF-α+ expression upon re-stimulation with other peptides. We next used the Vira-FEST assay to confirm cross-reactivity by assessing if the same CD4+ T receptor clonotypes recognize both S815-827 and homologous bat coronavirus peptides. In all 3 participants tested, we identified multiple cross-reactive T cell receptors that recognize both S815-827 and homologous bat coronavirus peptides. Conclusion: Our results suggest that current mRNA vaccines elicit T cell responses that can cross-recognize bat coronaviruses, and thus might induce protection.
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Background: About 10% of individuals with mild infection with SARS-CoV-2 suffer from Long COVID-19, defined as signs and symptoms developed during or following COVID-19 that continue for more than twelve weeks and cannot be explained by an alternative diagnosis. In this study, we analyzed the ADCC response and the reactivation of CMV and EBV in Long COVID-19 syndrome, in comparison with patients who completely recovered from mild COVID-19 Methods: 30 patients with Long COVID-19 (Long COVID-19) and 20 individuals who suffered mild COVID-19 and were completely recovered (Recovered) were recruited for this study. Specific anti-SARS-CoV-2 IgG titers were analyzed by direct ELISA and their neutralizing capability was measured by using pseudovirus neutralization assay. Phenotype of CD4+ and CD8+ T cells, NK, NKT, and B cells in peripheral blood was analyzed by flow cytometry. ADCC activity was analyzed using rituximab-coated Raji cells as target. EBV and CMV reactivation in plasma was analyzed by qPCR. Results: 1) 86.6% and 55.50% participants were female in Long COVID-19 and Recovered cohorts, respectively. Median age at COVID-19 diagnosis was 42y(IQR 37-46) and 45y (IQR 28-57), respectively. 2) Similar levels of CD4+ T cells were observed in both groups. However, Tregs were increased 2.8-fold in Long COVID-19 participants (p=0.0007). 3) CD8+ T cells, CD8+TCRγδ and CD8+TCRγδ were increased 1.3-(p=0.0005), 2.0-(p=0.049), and 2.5-fold (p=0.005) in Long COVID-19 individuals. 4) Expression of CD56 in NK cells and CD3-CD56+CD16+ cells were increased 1.7-(p=0.0005) and 1.7-fold (p=0.032) in Long COVID-19, respectively. 5) Specific anti-SARS-CoV-2 IgG titers were increased 2.3-fold in Long COVID-19 individuals (p=0.02) and their neutralizing capacity was increased 4.2-fold (p=0.034) in this cohort. However, ADCC activity was decreased 1.4-fold (p=0.0044). 6) Resting memory B cells were increased 2.3-fold during Long COVID-19, whereas plasmablasts were reduced 3.1-fold. 7) EBV was reactivated in 33.3% of Long COVID-19 individuals (p<0.0001), whereas CMV was not reactivated in any individual. Conclusion: Despite high levels of neutralizing antibodies and cytotoxic immune populations, an impaired antibody-dependent cytotoxic activity was observed in PBMCs from individuals with Long COVID-19. This defective cytotoxic immune response may impede viral clearance, which may also contribute to EBV reactivation observed in these individuals, thereby influencing on the persistent COVID-19 symptoms.