A Significant Contribution of the Classical Pathway of Complement in SARS-CoV-2 Neutralization of Convalescent and Vaccinee Sera.

Although high titers of neutralizing Abs in human serum are associated with protection from reinfection by SARS-CoV-2, there is considerable heterogeneity in human serum-neutralizing Abs against SARS-CoV-2 during convalescence between individuals. Standard human serum live virus neutralization assays require inactivation of serum/plasma prior to testing. In this study, we report that the SARS-CoV-2 neutralization titers of human convalescent sera were relatively consistent across all disease states except for severe COVID-19, which yielded significantly higher neutralization titers. Furthermore, we show that heat inactivation of human serum significantly lowered neutralization activity in a live virus SARS-CoV-2 neutralization assay. Heat inactivation of human convalescent serum was shown to inactivate complement proteins, and the contribution of complement in SARS-CoV-2 neutralization was often >50% of the neutralizing activity of human sera without heat inactivation and could account for neutralizing activity when standard titers were zero after heat inactivation. This effect was also observed in COVID-19 vaccinees and could be abolished in individuals who were undergoing treatment with therapeutic anti-complement Abs. Complement activity was mainly dependent on the classical pathway with little contributions from mannose-binding lectin and alternative pathways. Our study demonstrates the importance of the complement pathway in significantly increasing viral neutralization activity against SARS-CoV-2 in spike seropositive individuals.

Longitudinal Assessment of SARS-CoV-2-Specific T Cell Cytokine-Producing Responses for 1 Year Reveals Persistence of Multicytokine Proliferative Responses, with Greater Immunity Associated with Disease Severity.

Cell-mediated immunity is critical for long-term protection against most viral infections, including coronaviruses. We studied 23 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected survivors over a 1-year post-symptom onset (PSO) interval by ex vivo cytokine enzyme-linked immunosorbent spot assay (ELISpot) assay. All subjects demonstrated SARS-CoV-2-specific gamma interferon (IFN-γ), interleukin 2 (IL-2), and granzyme B (GzmB) T cell responses at presentation, with greater frequencies in severe disease. Cytokines, mainly produced by CD4+ T cells, targeted all structural proteins (nucleocapsid, membrane, and spike) except envelope, with GzmB and IL-2 greater than IFN-γ. Mathematical modeling predicted that (i) cytokine responses peaked at 6 days for IFN-γ, 36 days for IL-2, and 7 days for GzmB, (ii) severe illness was associated with reduced IFN-γ and GzmB but increased IL-2 production rates, and (iii) males displayed greater production of IFN-γ, whereas females produced more GzmB. Ex vivo responses declined over time, with persistence of IL-2 in 86% and of IFN-γ and GzmB in 70% of subjects at a median of 336 days PSO. The average half-life of SARS-CoV-2-specific cytokine-producing cells was modeled to be 139 days (~4.6 months). Potent T cell proliferative responses persisted throughout observation, were CD4 dominant, and were capable of producing all 3 cytokines. Several immunodominant CD4 and CD8 epitopes identified in this study were shared by seasonal coronaviruses or SARS-CoV-1 in the nucleocapsid and membrane regions. Both SARS-CoV-2-specific CD4+ and CD8+ T cell clones were able to kill target cells, though CD8 tended to be more potent. IMPORTANCE Our findings highlight the relative importance of SARS-CoV-2-specific GzmB-producing T cell responses in SARS-CoV-2 control and shared CD4 and CD8 immunodominant epitopes in seasonal coronaviruses or SARS-CoV-1, and they indicate robust persistence of T cell memory at least 1 year after infection. Our findings should inform future strategies to induce T cell vaccines against SARS-CoV-2 and other coronaviruses.

The First North American Experience Using Glycosorb Immunoadsorption Columns for Blood Group-Incompatible Kidney Transplantation.

BACKGROUND: Blood group incompatibility (ABOi) is the most common barrier to living donor kidney transplantation. Options for such recipients include kidney paired donation (KPD) or desensitization methodology to reduce blood antibody response. OBJECTIVE: The objective of this study is to report on the first North America experience in ABOi living donor kidney transplantation using Glycosorb ABO immunoadsorption columns. DESIGN: Retrospective observational cohort study. SETTING: Renal transplant program at St. Michael's Hospital, Unity Health Toronto, University of Toronto. PATIENTS: Twenty-six ABOi living donor transplants from August 2011 through February 2020 were undertaken at our center. MEASUREMENTS: Renal allograft and patient survival postdesensitization for ABOi living donor transplants and isohemagglutinin titer reduction. METHODS: Preoperative immunosuppressive regimen consisted of a single dose of Rituximab 375 mg/m2 IV on day -28; tacrolimus, mycophenolic acid, and prednisone to start on day -7. Immunoadsorption treatments with Glycosorb A or B columns were performed on day -7 through day -1 based on anti-A or anti-B titers on Spectra Optia Apheresis System. Immunosuppression included basiliximab, solumedrol followed by oral prednisone, once-daily tacrolimus, and mycophenolic acid. The mean follow-up was 53 months (3-96 months). RESULTS: A total of 26 individuals underwent an attempt at desensitization of whom 24 patients underwent immediate transplant. One patient had a rebound in titers and subsequently was transplanted from a blood group compatible living donor. A second patient had an unrelated medical issue and desensitization was discontinued. Five-year patient survival was 96% and death censored allograft survival was 92%. Posttransplant anti-A or anti-B titers were monitored daily for the first 7 days posttransplant and every 2 days from days 7 to 14. There were no acute rejections seen in this cohort of transplant recipients. LIMITATIONS: As our protocol was first initiated as proof of concept, a few recipients had low initial isohemagglutinin titers. This may have contributed to improved clinical outcomes. CONCLUSIONS: ABO column immunoadsorption with specific columns is a safe and effective method for ABOi living donor kidney transplantation, and an option when KPD is less than ideal.Trial not registered as this was a retrospective cohort review.

CONTEXTE: L'incompatibilité du système ABO (ABOi) est l'obstacle le plus fréquent à la transplantation d'un rein provenant d'un donneur vivant. Un don croisé ou une désensibilisation visant à atténuer la réponse immunitaire constituent les seules options pour les receveurs de ce type de greffe. OBJECTIF: Faire état de la première expérience nord-américaine d'utilisation des colonnes d'immunoadsorption Glycosorb ABO pour la transplantation d'un rein ABOi provenant d'un donneur vivant. TYPE D'ÉTUDE: Étude de cohorte observationnelle rétrospective. CADRE: Le programme de transplantation rénale du centre hospitalier universitaire St Michael's de l'Unity Health Toronto. SUJETS: L'étude porte sur les 26 transplantations de reins ABOi provenant de donneurs vivants pratiquées à notre centre entre août 2011 et février 2020. MESURES: La survie du patient et de l'allogreffe après une désensibilisation en vue de la transplantation d'un rein ABOi provenant d'un donneur vivant, ainsi que la réduction du titre d'isohémagglutinine. MÉTHODOLOGIE: Le traitement préopératoire immunosuppressif consistait en une dose unique de 375 mg/m2 de Rituximab par voie intraveineuse (IV) au jour -28; et l'administration de tacrolimus, d'acide mycophénolique et de prednisone à partir du jour -7. Les traitements d'immunoadsorption avec les colonnes Glycosorb A ou B ont été effectués du jour -7 au jour -1 en fonction des titres anti-A ou anti-B obtenus avec le système d'apharèse Spectra Optia. Le traitement immunosuppressif était constitué de basiliximab, de solumédrol suivi de prednisone par voie orale, et d'une dose quotidienne de tacrolimus et d'acide mycophénolique. Le suivi s'est étalé sur une moyenne de 53 mois (3 à 96 mois). RÉSULTATS: En tout, 26 patients avaient tenté une désensibilisation, desquels 24 ont immédiatement subi une transplantation. Un rebond des titres a été observé chez un patient, lequel a par la suite été transplanté avec un organe provenant d'un donneur de groupe sanguin compatible. La désensibilisation a dû être interrompue chez un autre patient en raison d'un problème médical non relié. Cinq ans après la greffe, 96% des patients et 92% des allogreffes avaient survécu. Les titres d'anti-A et d'anti-B post-transplantation avaient été mesurés quotidiennement pour les sept premiers jours suivant l'intervention, puis tous les deux jours entre le jour 7 et le jour 14. Aucun rejet aigu n'est survenu dans la cohorte étudiée. LIMITES: Notre protocole ayant d'abord été utilisé comme preuve de concept, certains patients présentaient de faibles titres initiaux d'isohémagglutinine, ce qui pourrait avoir contribué à l'amélioration des résultats cliniques. CONCLUSIONS: L'immunoadsorption sur colonne ABO avec colonnes spécifiques s'avère une méthode sûre et efficace pour la transplantation d'un rein ABOi provenant d'un donneur vivant, et constitue une option valable lorsque le don croisé n'est pas idéal.Essai non enregistré puisqu'il s'agit d'une étude de cohorte rétrospective.