ABSTRACT
Prior to the COVID-19 pandemic, the music industry was already experiencing uncertainty as musicians experimented with new modes of dissemination and monetization following developments in telecommunications. The arrival of the internet instigated varying, sometimes contradictory, cultural concerns including dispersion, dissipation, preservation, development, homogenization and heterogeneity. Guitar players have traditionally formed local networks and communities in geo-located domains. However, in the twenty-first century, community domains also include virtual spaces. An immersive netnographic study investigated activities in online guitar communities from the perspective of UNESCO’s Convention for the Safeguarding of Intangible Cultural Heritage. Data were analysed using the protocols of Inductive Thematic Analysis generating seven codes, regarding the impact of COVID-19 on virtual guitar communities. Without doubt, the pandemic has had serious negative effects on individual musicians’ and live venues’ income streams. However, it does seem to have acted as a catalyst for fresh vigour within online communities seeking new ways to connect. With more artists sharing and interacting, the result could be a richer environment in the future. However, without a strong recognition of cultural responsibility this richness may result in a homogenous melting pot. Alternatively, it may also bring to light cultural expressions previously suppressed © Equinox Publishing Ltd 2022, Office 415, The Workstation, 15 Paternoster Row, Sheffield S1 2BX
ABSTRACT
Background: The coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a pandemic that has taken millions of lives around the globe. Treatment of patients with moderate and severe COVID-19 disease has included dexamethasone, tocilizumab, Remdesivir, convalescent plasma, and targeted antibodies, however, currently, there are no FDA approved targeted cellular therapies in the treatment of mild or moderate SARS-CoV-2 disease. Virus-specific cytotoxic T cell lymphocytes (vCTLs) have shown therapeutic efficacy in immunocompromised patients with viral infections. We developed a multicenter and multidisciplinary Viral Cytotoxic T-Cell Consortium (VIRCTLC) to investigate the use of vCTLs manufactured by direct enrichment using the Cytokine Capture System (CCS) on the CliniMACS® Prodigy device. SARS-CoV-2 specific PepTivator Peptides consist of overlapping peptides that span the entire sequence of the protein (Protein N and M), or the length of its immunodominant domain (Protein S). The peptides can bind to either MHC class I or MHC class II molecules and are therefore able to target both CD4 and CD8 T cells. Objective: To screen, manufacture, and characterize SARS-CoV-2 vCTLs generated from convalescent COVID-19 donors using the CliniMACS® Cytokine Capture System on the CliniMACS® Prodigy device. Methods: Donor screening was done utilizing PBMNCs from 15 convalescent COVID-19 donors after informed consent. PBMNCs were stimulated with a mix of PepTivator peptides (Miltenyi Biotech®) contained in the S, M and N proteins. IFN-γ levels were examined in CD3, CD4, and CD8 T cells by flow cytometry analysis. After informed consent, PBMNCs from three convalescent COVID-19 donors who screened positively to the PepTivator® peptide pools of SARS-CoV-2 Proteins M, N and S were collected by apheresis using the SPECTRA Optia® apheresis instrument. PBMNCs were incubated with the PepTivator® peptide pools for 4 hours. After incubation, the SARS-CoV-2 vCTLs were enriched using the CliniMACS Cytokine Capture System as we have previously described (Flower/Cairo, et al, ASTCT, 2020). Samples were taken from the enriched vCTLs and tested in gram stains, sterility cultures, cell counts, viability and IFN-γ cytokine staining (CD3/CD4/CD8/IFN-γ marker panel) by flow cytometry. Amplification and sequencing of TCRβ CDR3 regions of pre-stimulated PBMNC, stimulated PBMNCs samples taken from the QC bag (QC samples) and the enriched SARS-CoV-2 vCTLs were performed on the ImmunoSEQ platform using ImmunoSEQ® TCRB Assay kit (Adaptive Biotechnologies, Seattle, WA, USA). Characterization of immune subsets was done by mass cytometry analysis with 41 Immunophenotypic markers. Transcriptome of the immune landscape of QC samples, and enriched vCTLs was compared with the pre samples using the human nCounter PanCancer Immune Profiling Panel on the nCounter system. Results: We demonstrate that 93.3% of convalescent donor blood samples passed the screening criteria for clinical manufacture. Three validation runs resulted in enriched T cells that consisted of 79% + 21% (mean + SEM) IFNγ + T cells (Fig.1). TCRβ sequencing showed that convalescent COVID-19 donors have a highly diverse TCR repertoire and we identified TCRβ CDR3 clones that are known to be associated with SARS-CoV-2 T cell responses. Immunophenotyping analysis demonstrated more CD4 T cells than CD8 T cells in the SARS CoV-2 vCTLs, an increase in memory CD8 and CD4 cells, especially CD8 T EM, CD4 T cm and CD4 T EMRA cells (Fig.2) and an increase DC cells in the SARS CoV-2 vCTL products as compared to pre-stimulated PBMNCs. Expression of the exhaustion markers was not enhanced in the SARS CoV-2 vCTLs as compared to pre-stimulated PBMNCs. Transcriptome analysis showed increased gene expression in T-cell function, interleukin, pathogen defense, and TNF superfamily pathway genes in the SARS CoV-2 vCTLs as compared to pre-stimulated PBMNCs. Conclusion: Our study demonstrates that highly functional SARS-CoV-2 vCTLs can be rapidly generat d by direct cytokine enrichment from convalescent donor peripheral blood mononuclear cells. These data serve as pre-clinical validation for an ongoing clinical trial utilizing related HLA-matched and haplo-identical SARS CoV-2 vCTLs for the treatment of patients with mild and moderate SARS-CoV-2 disease (IND #27260, NCT# 04896606). [Formula presented] Disclosures: Lee: Kiadis Pharma: Divested equity in a private or publicly-traded company in the past 24 months, Honoraria, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties, Research Funding;Courier Therapeutics: Current holder of individual stocks in a privately-held company. Johnson: Miltenyi Biotec: Research Funding. Cairo: Jazz Pharmaceutical: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau;Amgen: Speakers Bureau;Sanofi: Speakers Bureau;Servier: Speakers Bureau;Sobi: Speakers Bureau;Omeros: Membership on an entity's Board of Directors or advisory committees;Nektar: Membership on an entity's Board of Directors or advisory committees.