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Nasopharyngeal carcinoma (NPC) is an Epstein–Barr virus (EBV)-associated heterogeneous disease and is characterized by peritumoral immune infiltrate. Adoptive T-cell therapy (ACT) has emerged as a potential therapeutic strategy for NPC. However, the tumor microenvironment remains a major roadblock for the successful implementation of ACT in clinical settings. Expression of checkpoint molecules by malignant cells can inhibit the effector function of adoptively transferred EBV-specific T cells. Here we present a novel case report of a patient with metastatic NPC who was successfully treated with a combination of EBV-specific ACT and programmed cell death-1 blockade therapy. Following combination immunotherapy, the patient showed complete resolution of metastatic disease with no evidence of disease relapse for 22 months. Follow-up immunological analysis revealed dramatic restructuring of the global T-cell repertoire that was coincident with the clinical response. This case report provides an important platform for translating these findings to a larger cohort of NPC patients.

Cytomegalovirus (CMV) remains a significant burden in lung transplant recipients. Deficiencies in T-cell immunity posttransplant increase the risk of CMV-associated complications. However, it is not clear if underlying poor pretransplant immunity increases risk. To assess this, we recruited 39 prospective lung transplant patients and performed QuantiFERON-CMV on their peripheral blood. More than a third of prospective CMV-seropositive transplant recipients were CMV non-immune reactive (CMV-NIR) pretransplant. CMV-NIR status was associated with a significantly higher incidence of CMV reactivation posttransplant, demonstrating that dysfunctional CMV immunity in prospective lung transplant recipients is associated with an increased risk of viral reactivation posttransplant.

Background The recent failure of checkpoint-blockade therapies for glioblastoma multiforme (GBM) in late-phase clinical trials has directed interest towards adoptive cellular immunotherapies (ACT). In this open-label, first-in-human trial, we have assessed the safety and therapeutic potential of cytomegalovirus (CMV)-specific ACT in an adjuvant setting for patients with primary GBM, with an ultimate goal to prevent or delay recurrence and prolong overall survival. Methods Twenty-eight patients with primary GBM were recruited to this prospective study, 25 of whom were treated with in vitro-expanded autologous CMV-specific T cells. Participants were monitored for safety, progression-free survival (PFS), overall survival (OS) and immune reconstitution. Results No participants showed evidence of ACT-related toxicities. Of 25 evaluable participants, ten were alive at the completion of follow-up, while five were disease free. Reconstitution of CMV-specific T-cell immunity was evident and CMV-specific ACT may trigger bystander effect leading to additional T-cell responses to non-viral tumour-associated antigens through epitope spreading. Long-term follow-up of participants treated before recurrence showed significantly improved OS when compared to those who progressed before ACT (median 23 months, range 7-65 vs. median 14 months, range 5-19; p=0.018). Gene expression analysis of the ACT products indicated that a favourable T-cell gene signature was associated with improved long-term survival. Conclusion Data presented in this study demonstrate that CMV-specific ACT can be safely used as an adjuvant therapy for primary GBM and, if offered before recurrence, this therapy may improve overall survival of GBM patients. Trial registration anzctr.org.au: ACTRN12615000656538Funding Source:National Health & Medical Research Council (Australia)Trial registration: anzctr.org.au: ACTRN12615000656538Funding Source: Philanthropic funding &National Health & Medical Research Council (Australia).

BACKGROUND: Impaired T cell immunity in transplant recipients is associated with infection-related morbidity and mortality. We recently reported the successful use of adoptive T cell therapy (ACT) against drug-resistant/recurrent cytomegalovirus in solid-organ transplant recipients. METHODS: In the present study, we used high-throughput T cell receptor Vβ sequencing and T cell functional profiling to delineate the impact of ACT on T cell repertoire remodeling in the context of pretherapy immunity and ACT products. RESULTS: These analyses indicated that a clinical response was coincident with significant changes in the T cell receptor Vβ landscape after therapy. This restructuring was associated with the emergence of effector memory T cells in responding patients, while nonresponders displayed dramatic pretherapy T cell expansions with minimal change following ACT. Furthermore, immune reconstitution included both adoptively transferred clonotypes and endogenous clonotypes not detected in the ACT products. CONCLUSION: These observations demonstrate that immune control following ACT requires significant repertoire remodeling, which may be impaired in nonresponders because of the preexisting immune environment. Immunological interventions that can modulate this environment may improve clinical outcomes. TRIAL REGISTRATION: Australian New Zealand Clinical Trial Registry, ACTRN12613000981729. FUNDING: This study was supported by funding from the National Health and Medical Research Council, Australia (APP1132519 and APP1062074).

TCR-gene-transfer is an efficient strategy to produce therapeutic T cells of defined antigen specificity. However, there are substantial variations in the cell surface expression levels of human TCRs, which can impair the function of engineered T cells. Here we demonstrate that substitutions of 3 amino acid residues in the framework of the TCR variable domains consistently increase the expression of human TCRs on the surface of engineered T cells.The modified TCRs mediate enhanced T cell proliferation, cytokine production and cytotoxicity, while reducing the peptide concentration required for triggering effector function up to 3000-fold. Adoptive transfer experiments in mice show that modified TCRs control tumor growth more efficiently than wild-type TCRs. Our data indicate that simple variable domain modifications at a distance from the antigen-binding loops lead to increased TCR expression and improved effector function. This finding provides a generic platform to optimize the efficacy of TCR gene therapy in humans., Increasing TCR cell surface expression can potentiate T cell responses to low-concentrations of antigen. Here the authors identify aminoacids in human TCR variable domains that impact its surface expression, and demonstrate how editing these residues can improve T cell activation and effector function without altering antigen specificity.

Objectives There is emerging evidence that SARS‐CoV‐2‐specific memory T‐cell responses are likely to provide critical long‐term protection against COVID‐19. Strategies to rapidly assess T‐cell responses are therefore likely to be important for assessing immunity in the global population. Methods Here, we have developed a rapid immune‐monitoring strategy to assess virus‐specific memory T‐cell responses in the peripheral blood of COVID‐19 convalescent individuals. We validated SARS‐CoV‐2‐specific memory T‐cell responses detected in whole blood using in vitro expansion with SARS‐CoV‐2 proteins. Results T‐cell immunity characterised by the production of IFN‐γ and IL‐2 could be consistently detected in the whole blood of recovered participants. T cells predominantly recognised structural SARS‐CoV‐2 proteins. In vitro expansion demonstrated that while CD8+ T cells recognised nucleocapsid protein, spike protein and ORF3a, CD4+ T cells more broadly targeted multiple SARS‐CoV‐2 proteins. Conclusion These observations provide a timely monitoring approach for identifying SARS‐CoV‐2 cellular immunity and may serve as a diagnostic for the stratification of risk in immunocompromised and other at‐risk individuals., T‐cell immunity is likely critical for the control of COVID‐19. We have developed a rapid assay to detect T‐cell immunity in whole blood, and demonstrate the effectiveness of this assay in individuals who have recovered from COVID‐19.

Objectives Clinical response to antibody‐based immunotherapies targeting checkpoint inhibitors is critically dependent on the tumor immune microenvironment (TIME). However, the precise impact of the TIME on adoptive cellular immunotherapy remains unexplored. Here we have conducted a long‐term follow‐up analysis of patients with recurrent glioblastoma multiforme (GBM) who were treated with autologous CMV‐specific T‐cell therapy to delineate the potential impact of the TIME on their clinical response. Methods Multiplexed immunohistochemical analysis of CD3, PD‐L1 and Sox‐2 in GBM tissue biopsies obtained before autologous T‐cell therapy was carried out and correlated with long‐term survival of GBM patients adoptively treated with T‐cell therapy. Results Tumor microenvironment analyses revealed that the pre‐treatment cellular composition of the tumor tissue may influence the subsequent response to adoptive T‐cell therapy. GBM patients who showed prolonged overall survival following T‐cell therapy had a significantly lower number of tumor‐infiltrating CD3+ T cells in recurrent tumors than that in patients with short‐term survival. Furthermore, long‐term surviving patients showed low or undetectable PD‐L1 expression in tumor cells in recurrent GBM biopsies. Conclusion We hypothesise that lack of PD‐L1‐mediated immunosuppression in the TIME may allow efficient immune control following adoptive T‐cell therapy. Future studies combining anti‐PD‐L1 or genetically modified T cells with PD‐1 receptor knockdown could be considered to improve clinical responses in patients who have high PD‐L1 expression in their tumors., The precise role of tumor immune microenvironment (TIME) on clinical response to adoptive cellular immunotherapies remains largely unexplored. In this paper, we have analysed the impact of TIME on the clinical response of glioblastoma patients who showed either long‐term or short‐term overall survival following adoptive T‐cell therapy.

BACKGROUND. Increasing evidence indicates a role for EBV in the pathogenesis of multiple sclerosis (MS). EBV-infected autoreactive B cells might accumulate in the CNS because of defective cytotoxic CD8+ T cell immunity. We sought to determine the feasibility and safety of treating progressive MS patients with autologous EBV-specific T cell therapy. METHODS. An open-label phase I trial was designed to treat 5 patients with secondary progressive MS and 5 patients with primary progressive MS with 4 escalating doses of in vitro–expanded autologous EBV-specific T cells targeting EBV nuclear antigen 1, latent membrane protein 1 (LMP1), and LMP2A. Following adoptive immunotherapy, we monitored the patients for safety and clinical responses. RESULTS. Of the 13 recruited participants, 10 received the full course of T cell therapy. There were no serious adverse events. Seven patients showed improvement, with 6 experiencing both symptomatic and objective neurological improvement, together with a reduction in fatigue, improved quality of life, and, in 3 patients, reduced intrathecal IgG production. All 6 patients receiving T cells with strong EBV reactivity showed clinical improvement, whereas only 1 of the 4 patients receiving T cells with weak EBV reactivity showed improvement (P = 0.033, Fisher’s exact test). CONCLUSION. EBV-specific adoptive T cell therapy was well tolerated. Clinical improvement following treatment was associated with the potency of EBV-specific reactivity of the administered T cells. Further clinical trials are warranted to determine the efficacy of EBV-specific T cell therapy in MS. TRIAL REGISTRATION. Australian New Zealand Clinical Trials Registry, ACTRN12615000422527. FUNDING. MS Queensland, MS Research Australia, Perpetual Trustee Company Ltd., and donations from private individuals who wish to remain anonymous.

The ability to decode antigen specificities encapsulated\ud in the sequences of rearranged T-cell receptor\ud (TCR) genes is critical for our understanding of\ud the adaptive immune system and promises signifi-\ud cant advances in the field of translational medicine.\ud Recent developments in high-throughput sequencing\ud methods (immune repertoire sequencing technology,\ud or RepSeq) and single-cell RNA sequencing\ud technology have allowed us to obtain huge numbers\ud of TCR sequences from donor samples and link them\ud to T-cell phenotypes. However, our ability to annotate\ud these TCR sequences still lags behind, owing\ud to the enormous diversity of the TCR repertoire and\ud the scarcity of available data on T-cell specificities. In\ud this paper, we present VDJdb, a database that stores\ud and aggregates the results of published T-cell speci-\ud ficity assays and provides a universal platform that\ud couples antigen specificities with TCR sequences.\ud We demonstrate that VDJdb is a versatile instrument\ud for the annotation of TCR repertoire data, enabling a\ud concatenated view of antigen-specific TCR sequence\ud motifs. VDJdb can be accessed at https://vdjdb.cdr3.\ud net and https://github.com/antigenomics/vdjdb-db.

Adoptive T cell therapy has emerged as a powerful strategy to treat human cancers especially haematological malignancies. Extension of these therapies to solid cancers remains a significant challenge especially in the context of defining immunological correlates of clinical responses. Here we describe results from a clinical study investigating autologous Epstein-Barr virus (EBV)-specific T cells generated using a novel AdE1-LMPpoly vector to treat patients with nasopharyngeal carcinoma (NPC) either pre-emptively in at-risk patients with no or minimal residual disease (N/MRD) or therapeutically in patients with active recurrent/metastatic disease (ARMD). Tolerability, safety and efficacy, including progression-free survival (PFS) and overall survival (OS), were evaluated following adoptive T-cell immunotherapy. Twenty-nine patients, including 20 with ARMD and nine with N/MRD, successfully completed T-cell therapy. After a median follow-up of 18.5 months, the median PFS was 5.5 months (95% CI 2.1 to 9.0 months) and the median OS was 38.1 months (95% CI 17.2 months to not reached). Post-immunotherapy analyses revealed that disease stabilization in ARMD patients was significantly associated with the functional and phenotypic composition of in vitro-expanded T cell immunotherapy. These included a higher proportion of effector CD8+ T-cells and an increased number of EBV-specific T-cells with broader antigen specificity. These observations indicate that adoptive immunotherapy with AdE1-LMPpoly-expanded T cells stabilizes relapsed, refractory NPC without significant toxicity. Promising clinical outcomes in N/MRD patients further suggest a potential role for this approach as a consolidation treatment following first-line chemotherapy.

Background: Current peptide vaccines may select suboptimal antigen-specific T-cells from polyclonal populations. Results: A combinatorial peptide library screen was used to generate an optimal ligand that could preferentially activate a known effective T-cell clonotype. Conclusion: Rationally designed altered peptide ligands may enable the preferential selection of high quality, antigen-sensitive T-cell clonotypes. Significance: This proof-of-principle study could facilitate the development of more effective peptide vaccination strategies., Altered peptide antigens that enhance T-cell immunogenicity have been used to improve peptide-based vaccination for a range of diseases. Although this strategy can prime T-cell responses of greater magnitude, the efficacy of constituent T-cell clonotypes within the primed population can be poor. To overcome this limitation, we isolated a CD8+ T-cell clone (MEL5) with an enhanced ability to recognize the HLA A*0201-Melan A27–35 (HLA A*0201-AAGIGILTV) antigen expressed on the surface of malignant melanoma cells. We used combinatorial peptide library screening to design an optimal peptide sequence that enhanced functional activation of the MEL5 clone, but not other CD8+ T-cell clones that recognized HLA A*0201-AAGIGILTV poorly. Structural analysis revealed the potential for new contacts between the MEL5 T-cell receptor and the optimized peptide. Furthermore, the optimized peptide was able to prime CD8+ T-cell populations in peripheral blood mononuclear cell isolates from multiple HLA A*0201+ individuals that were capable of efficient HLA A*0201+ melanoma cell destruction. This proof-of-concept study demonstrates that it is possible to design altered peptide antigens for the selection of superior T-cell clonotypes with enhanced antigen recognition properties.