Objectives:Despite the effective roll-out of COVID-19 vaccines, immunocompromised patients have a higher risk of morbidity and mortality following SARS-CoV-2 infection. Allogeneic adoptive T-cell immunotherapy is now established as an effective approach to treat viral diseases in immunocompromised patients. The objective of this study was to assess the safety of allogeneic virus-specific T-cell therapy in patients with COVID-19. Methods:Using a repository of SARS-CoV-2-specific T cells generated from healthy exposed volunteers, we conducted an open-label phase I trial to assess the feasibility and safety of allogeneic SARS-CoV-2-specific T cells in immune-compromised cancer patients with COVID-19. Results:Six participants at risk of severe COVID-19 were enrolled and received SARS-CoV-2-specific T-cell therapy within 4 days of recruitment. The first five participants who received two infusions of allogeneic SARS-CoV-2-specific T-cell therapy experienced no adverse events following treatment. Four of the six participants showed improvement in viral load following treatment and were alive at 12-week follow-up. One participant died 6 days after their second infusion, because of established pulmonary parenchymal damage following prolonged COVID infection. Another, who had underlying lupus nephritis, developed cytokine release syndrome and diffuse alveolar haemorrhage following a single infusion and was withdrawn from the study. They subsequently recovered from this serious adverse event. Conclusion:Allogeneic SARS-CoV-2-specific T-cell therapy provides a platform to rapidly administer T cells to high-risk COVID-19 patients. It was associated with a reduced viral load and increased SARS-CoV-2-specific T-cell responses in the majority of treated patients.

Adoptive T-cell immunotherapy holds great promise for the treatment of viral complications in immunocompromised patients resistant to standard anti-viral strategies. We present a retrospective analysis of 78 patients from 19 hospitals across Australia and New Zealand, treated over the last 15 years with "off-the-shelf" allogeneic T cells directed to a combination of Epstein-Barr virus (EBV), cytomegalovirus (CMV), BK polyomavirus (BKV), John Cunningham virus (JCV) and/or adenovirus (AdV) under the Australian Therapeutic Goods Administration's Special Access Scheme. Most patients had severe post-transplant viral complications, including drug-resistant end-organ CMV disease, BKV-associated haemorrhagic cystitis and EBV-driven post-transplant lymphoproliferative disorder. Adoptive immunotherapy is well tolerated with few adverse effects. Importantly, 46/71 (65%) patients show definitive clinical improvement including reduction in viral load, clinical symptoms and complete resolution of end-organ disease. In addition, seven high-risk patients remain disease free. Based on this long-term encouraging clinical experience, we propose that a dedicated nationally funded centre for anti-viral cellular therapies should be considered to provide T cell therapies for critically ill patients for compassionate use. Adoptive T-cell immunotherapy offers promise to patients who are resistant to standard anti-viral strategies. Here the authors describe clinical observations in patients with viral complications treated with adoptive immunotherapy over the last 15 years.

AbstractObjectivesDysregulation of Epstein–Barr virus (EBV)‐specific cellular immunity has been hypothesised as one of the contributing factors in the pathogenesis of systemic lupus erythematosus (SLE). Lupus nephritis is a major risk factor for overall morbidity in SLE. Immune‐based strategies directed to EBV have been proposed as potential therapeutic strategy for SLE and lupus nephritis.MethodsAutologous EBV latent antigen‐specific CD4+ and CD8+ T cells were expanded in vitro and adoptively transferred to a lupus nephritis patient.ResultsThis adoptive immunotherapy had no immediate adverse effects, and the patient was subsequently treated with the anti‐CD20 antibody, obinutuzumab. The patient showed a reduction in anti‐dsDNA antibodies and improved glomerular filtration rate but remained nephrotic. These observations were coincident with a reduction in anti‐viral and global T‐cell activation.ConclusionTo our knowledge, this is the first report of the use of EBV‐specific adoptive immunotherapy to treat a patient with lupus nephritis.

Abstract T cell receptor (TCR) diversity analysis is critical for understanding the complex role of T cells in disease pathogenesis. In this study we profile the CD4+ and CD8+ T cell compartments in SARS-CoV-2-recovered participants and use TCR beta variable (TRBV) receptor deep sequencing to explore the association between the T cell repertoire and severity of COVID-19. We compare mild and severe COVID-19 cohorts and identify two unique populations of shared TRBV sequences that are significantly enriched in patients following severe disease or in patients with post-acute sequelae of COVID-19 (PASC). These enriched TRBV sequences display a clear association with human leukocyte antigen (HLA) class I and class II alleles, yet are distinct from the SARS-CoV-2-specific T cell repertoire. Exploration of additional disease datasets confirms that these TRBV populations are present in unexposed individuals and potentially represent a TRBV signature associated with either restricting efficient viral control or exacerbating disease symptoms. Subject terms: COVID-19, T cell immunity, T cell receptor

Gating strategy for CD8+ T-cell subsets. CM: Central memory; EM: Effector memory; TEMRA: Effector memory expressing CD45RA

Raw scanned data from ELISpot plate scanning reader (AID ELISpot Reader Classic); data was then processed using the AID ELISpot software to standardise and analyse the spot counts using intensity and size measurements, to provide the final data presented in Supplementary Figure 2.

Populations of IFNγ high and IFNγ low in response to tumour/peptide stimulation in MLTC at day 34 in patients (A) D05 and (B) D14

Whole -exome alignment of PABPC3 compared to the PABPC1 sequence (A): PABPC3 sequencing reads alignment with PABPC1 and PABPC3 reference sequence. (B): PABPC3 dinucleotide change resulting in R -->Q amino acid change in the protein sequence.

Background & Aim: Despite the successful implementation of vaccines worldwide, COVID-19 remains a risk in patients with a compromised immune system.Emerging viral variants have also reduced the effectiveness of monoclonal antibody therapies in these patients.New treatment options are therefore required to improve clinical outcomes. Methods, Results & Conclusion:T cell immunotherapy has proven effective for the treatment of a number of refractory viral diseases in patients with a compromised immune system.We have now completed the manufacture of a bank of SARS-CoV-2 specific T cells and commenced an open-label phase I clinical trial at the Royal Brisbane and Women's Hospital, Australia.Patients enrolled in the study receive two doses of partially HLA-matched allogeneic T cells at a fortnightly interval.We have thus far recruited and treated three immune compromised patients with SARS-CoV-2 T cells.In two of the three patients treated thus far, the administration of T cell therapy was coincident with the clearance of viral load after 28 days.Viral clearance in these patients was also associated with an increase in circulating SARS-CoV-2 specific T cells.Our preliminary observations suggest that SARS-CoV-2 specific T cell therapy is well tolerated and has the potential to impact viral control in immune compromised patients.

Total raw dataset of assessment of potential neoepitope sequences through epitope prediction programs for D05, D14, D41, A02 and A06.

Raw data from the initial peptide screen across MLTC assessed by IFNγ ELISpot. The yellow line delineates 30 spots and the red line marks 40 spots. Each bar represents wells assessed in triplicate, averaged and the average background measurement subtracted. The top measurement was capped at 200 spots.

Cell line information and Sanger sequence confirmed mutations in each cell line.

Pulmonary hypertension is a hemodynamic state defined by a mean pulmonary arterial pressure >20 mmHg and a pulmonary vascular resistance ≥3 WU, subdivided into 5 groups. Chronic Thromboembolic Pulmonary Hypertension (CTEPH) corresponds to group 4. The antiphospholipid syndrome is one of the most associated thrombophilia, with a prevalence of CTEPH of 2%-50%. A case-control study was conducted where data from the Right Cardiac Catheterization Registry of the PH Clinic were collected, with a diagnosis of CTEPH in patients aged 18-60 years and any sex. Antiphospholipid Syndrome (APLS) patients were separated from those with only CTEPH. It was developed in a statistical analysis based on frequencies, means, and standard deviation. The variables were evaluated using the Kolmogorov-Smirnov, Student's T, Mann-Whitney U, and Chi-Square tests with a 95% confidence interval. A total of 12 patients with APLS diagnosis and 30 without it were identified. The comparison between both groups shows that the patients with APLS were younger (38 ± 14.35 vs 51.63 ± 15.02 years, P 0.010) and had a significant association with autoimmune diseases (25% vs 0%, P 0.003). The patients diagnosed with APLS were primarily men (7 vs 5), and no statistically significant difference was found between laboratory and hemodynamic parameters. Patients diagnosed with CTEPH and APLS are mainly male, younger mean age, and have a greater significant association with autoimmune diseases than patients with CTEPH.

Vaccines have curtailed the devastation wrought by COVID-19. Nevertheless, emerging variants result in a high incidence of breakthrough infections. Here we assess the impact of vaccination and breakthrough infection on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) T cell immunity. We demonstrate that COVID-19 vaccination induces robust spike-specific T cell responses that, within the CD4(+) compartment, display comparable IFN-gamma responses to SARS-CoV-2 infection without vaccination. Vaccine-induced CD8(+) IFN-gamma responses however, were significantly greater than those primed by SARSCoV-2 infection alone. This increased responsiveness is associated with induction of novel HLA-restricted CD8(+) T cell epitopes not primed by infection alone (without vaccination). Despite these augmented responses, breakthrough infection still induced de novo T cell responses against additional SARS-CoV-2 CD8(+) epitopes that display HLA-associated immunodominance hierarchies consistent with those in unvaccinated COVID-19 convalescent individuals. This study demonstrates the unique modulation of anti-viral T cell responses against multiple viral antigens following consecutive yet distinct priming events in COVID-19 vaccination and breakthrough infection.

The emergence of the SARS-CoV-2 Omicron variant has raised concerns of escape from vaccine-induced immunity. A number of studies have demonstrated a reduction in antibody-mediated neutralization of the Omicron variant in vaccinated individuals. Preliminary observations have suggested that T cells are less likely to be affected by changes in Omicron. However, the complexity of human leukocyte antigen genetics and its impact upon immunodominant T cell epitope selection suggests that the maintenance of T cell immunity may not be universal. In this study, we describe the impact that changes in Omicron BA.1, BA.2 and BA.3 have on recognition by spike-specific T cells. These T cells constitute the immunodominant CD8+ T cell response in HLA-A*29:02+ COVID-19 convalescent and vaccinated individuals; however, they fail to recognize the Omicron-encoded sequence. These observations demonstrate that in addition to evasion of antibody-mediated immunity, changes in Omicron variants can also lead to evasion of recognition by immunodominant T cell responses.

Understanding the immune response to severe acute respiratory syndrome coronavirus (SARS-CoV-2) is critical to overcome the current coronavirus disease (COVID-19) pandemic. Efforts are being made to understand the potential cross-protective immunity of memory T cells, induced by prior encounters with seasonal coronaviruses, in providing protection against severe COVID-19. In this study we assessed T-cell responses directed against highly conserved regions of SARS-CoV-2. Epitope mapping revealed 16 CD8(+) T-cell epitopes across the nucleocapsid (N), spike (S), and open reading frame (ORF)3a proteins of SARS-CoV-2 and five CD8(+) T-cell epitopes encoded within the highly conserved regions of the ORF1ab polyprotein of SARS-CoV-2. Comparative sequence analysis showed high conservation of SARS-CoV-2 ORF1ab T-cell epitopes in seasonal coronaviruses. Paradoxically, the immune responses directed against the conserved ORF1ab epitopes were infrequent and subdominant in both convalescent and unexposed participants. This subdominant immune response was consistent with a low abundance of ORF1ab encoded proteins in SARS-CoV-2 infected cells. Overall, these observations suggest that while cross-reactive CD8(+) T cells likely exist in unexposed individuals, they are not common and therefore are unlikely to play a significant role in providing broad preexisting immunity in the community. IMPORTANCE T cells play a critical role in protection against SARS-CoV-2. Despite being highly topical, the protective role of preexisting memory CD8(+) T cells, induced by prior exposure to circulating common co ronavirus strains, remains less clear. In this study, we established a robust approach to specifically assess T cell responses to highly conserved regions within SARS-CoV-2. Consistent with recent observations we demonstrate that recognition of these highly conserved regions is associated with an increased likelihood of milder disease. However, extending these observations we observed that recognition of these conserved regions is rare in both exposed and unexposed volunteers, which we believe is associated with the low abundance of these proteins in SARS-CoV-2 infected cells. These observations have important implications for the likely role preexisting immunity plays in controlling severe disease, further emphasizing the importance of vaccination to generate the immunodominant T cells required for immune protection.

Adoptive T-cell immunotherapy has provided promising results in the treatment of viral complications in humans, particularly in the context of immunocompromised patients who have exhausted all other clinical options. The capacity to expand T cells from healthy immune individuals is providing a new approach to anti-viral immunotherapy, offering rapid off-the-shelf treatment with tailor-made human leukocyte antigen (HLA)-matched T cells. While most of this research has focused on the treatment of latent viral infections, emerging evidence that SARS-CoV-2-specific T cells play an important role in protection against COVID-19 suggests that the transfer of HLA-matched allogeneic off-the-shelf virus-specific T cells could provide a treatment option for patients with active COVID-19 or at risk of developing COVID-19. We initially screened 60 convalescent individuals and based on HLA typing and T-cell response profile, 12 individuals were selected for the development of a SARS-CoV-2-specific T-cell bank. We demonstrate that these T cells are specific for up to four SARS-CoV-2 antigens presented by a broad range of both HLA class I and class II alleles. These T cells show consistent functional and phenotypic properties, display cytotoxic potential against HLA-matched targets and can recognize HLA-matched cells infected with different SARS-CoV-2 variants. These observations demonstrate a robust approach for the production of SARS-CoV-2-specific T cells and provide the impetus for the development of a T-cell repository for clinical assessment.

Abstract The emergence of the SARS-CoV-2 Omicron variant has raised concerns of escape from vaccine-induced immunity. A number of studies have demonstrated a reduction in antibody-mediated neutralization of the Omicron variant in vaccinated individuals. Preliminary observations have suggested that T cells are less likely to be affected by changes in Omicron. However, the complexity of human leukocyte antigen genetics and its impact upon immunodominant T-cell epitope selection suggests that the maintenance of T-cell immunity won’t be universal. In this study, we describe the impact that changes in Omicron have on recognition by spike-specific T cells. These T cells constitute the immunodominant CD8+ T-cell response in HLA-A*29:02+ COVID-19 convalescent and vaccinated individuals; however, they fail to recognize the Omicron-encoded sequence. We also failed to detect other immunodominant responses in vaccinated HLA-A*29:02+ individuals, suggesting that the Omicron variant may have an increased capacity to escape immune recognition in this population.