A distinct stimulatory cDC1 subpopulation amplifies CD8+ T cell responses in tumors for protective anti-cancer immunity.

Type 1 conventional dendritic cells (cDC1) can support T cell responses within tumors but whether this determines protective versus ineffective anti-cancer immunity is poorly understood. Here, we use imaging-based deep learning to identify intratumoral cDC1-CD8+ T cell clustering as a unique feature of protective anti-cancer immunity. These clusters form selectively in stromal tumor regions and constitute niches in which cDC1 activate TCF1+ stem-like CD8+ T cells. We identify a distinct population of immunostimulatory CCR7neg cDC1 that produce CXCL9 to promote cluster formation and cross-present tumor antigens within these niches, which is required for intratumoral CD8+ T cell differentiation and expansion and promotes cancer immune control. Similarly, in human cancers, CCR7neg cDC1 interact with CD8+ T cells in clusters and are associated with patient survival. Our findings reveal an intratumoral phase of the anti-cancer T cell response orchestrated by tumor-residing cDC1 that determines protective versus ineffective immunity and could be exploited for cancer therapy.

Human TH17 cells engage gasdermin E pores to release IL-1α on NLRP3 inflammasome activation.

It has been shown that innate immune responses can adopt adaptive properties such as memory. Whether T cells utilize innate immune signaling pathways to diversify their repertoire of effector functions is unknown. Gasdermin E (GSDME) is a membrane pore-forming molecule that has been shown to execute pyroptotic cell death and thus to serve as a potential cancer checkpoint. In the present study, we show that human T cells express GSDME and, surprisingly, that this expression is associated with durable viability and repurposed for the release of the alarmin interleukin (IL)-1α. This property was restricted to a subset of human helper type 17 T cells with specificity for Candida albicans and regulated by a T cell-intrinsic NLRP3 inflammasome, and its engagement of a proteolytic cascade of successive caspase-8, caspase-3 and GSDME cleavage after T cell receptor stimulation and calcium-licensed calpain maturation of the pro-IL-1α form. Our results indicate that GSDME pore formation in T cells is a mechanism of unconventional cytokine release. This finding diversifies our understanding of the functional repertoire and mechanistic equipment of T cells and has implications for antifungal immunity.

A20 and ABIN-1 cooperate in balancing CBM complex-triggered NF-κB signaling in activated T cells.

T cell activation initiates protective adaptive immunity, but counterbalancing mechanisms are critical to prevent overshooting responses and to maintain immune homeostasis. The CARD11-BCL10-MALT1 (CBM) complex bridges T cell receptor engagement to NF-κB signaling and MALT1 protease activation. Here, we show that ABIN-1 is modulating the suppressive function of A20 in T cells. Using quantitative mass spectrometry, we identified ABIN-1 as an interactor of the CBM signalosome in activated T cells. A20 and ABIN-1 counteract inducible activation of human primary CD4 and Jurkat T cells. While A20 overexpression is able to silence CBM complex-triggered NF-κB and MALT1 protease activation independent of ABIN-1, the negative regulatory function of ABIN-1 depends on A20. The suppressive function of A20 in T cells relies on ubiquitin binding through the C-terminal zinc finger (ZnF)4/7 motifs, but does not involve the deubiquitinating activity of the OTU domain. Our mechanistic studies reveal that the A20/ABIN-1 module is recruited to the CBM complex via A20 ZnF4/7 and that proteasomal degradation of A20 and ABIN-1 releases the CBM complex from the negative impact of both regulators. Ubiquitin binding to A20 ZnF4/7 promotes destructive K48-polyubiquitination to itself and to ABIN-1. Further, after prolonged T cell stimulation, ABIN-1 antagonizes MALT1-catalyzed cleavage of re-synthesized A20 and thereby diminishes sustained CBM complex signaling. Taken together, interdependent post-translational mechanisms are tightly controlling expression and activity of the A20/ABIN-1 silencing module and the cooperative action of both negative regulators is critical to balance CBM complex signaling and T cell activation.

Human skin-resident host T cells can persist long term after allogeneic stem cell transplantation and maintain recirculation potential.

Tissue-resident memory T cells (TRM) have recently emerged as crucial cellular players for host defense in a wide variety of tissues and barrier sites. Insights into the maintenance and regulatory checkpoints of human TRM cells remain scarce, especially due to the difficulties associated with tracking T cells through time and space in humans. We therefore sought to identify and characterize skin-resident T cells in humans defined by their long-term in situ lodgment. Allogeneic hematopoietic stem cell transplantation (allo-HSCT) preceded by myeloablative chemotherapy unmasked long-term sequestration of host T cell subsets in human skin despite complete donor T cell chimerism in the blood. Single-cell chimerism analysis paired with single-cell transcriptional profiling comprehensively characterized these bona fide long-term skin-resident T cells and revealed differential tissue maintenance for distinct T cell subsets, specific TRM cell markers such as galectin-3, but also tissue exit potential with retention of the transcriptomic TRM cell identity. Analysis of 26 allo-HSCT patients revealed profound interindividual variation in the tissue maintenance of host skin T cells. The long-term persistence of host skin T cells in a subset of these patients did not correlate with the development of chronic GvHD. Our data exemplify the power of exploiting a clinical situation as a proof of concept for the existence of bona fide human skin TRM cells and reveal long-term persistence of host T cells in a peripheral tissue but not in the circulation or bone marrow in a subset of allo-HSCT patients.

Characterization of a library of 20 HBV-specific MHC class II-restricted T cell receptors.

CD4+ T cells play an important role in the immune response against cancer and infectious diseases. However, mechanistic details of their helper function in hepatitis B virus (HBV) infection in particular, or their advantage for adoptive T cell therapy remain poorly understood as experimental and therapeutic tools are missing. Therefore, we identified, cloned, and characterized a comprehensive library of 20 MHC class II-restricted HBV-specific T cell receptors (TCRs) from donors with acute or resolved HBV infection. The TCRs were restricted by nine different MHC II molecules and specific for eight different epitopes derived from intracellularly processed HBV envelope, core, and polymerase proteins. Retroviral transduction resulted in a robust expression of all TCRs on primary T cells. A high functional avidity was measured for all TCRs specific for epitopes S17, S21, S36, and P774 (half-maximal effective concentration [EC50] <10 nM), or C61 and preS9 (EC50 <100 nM). Eight TCRs recognized peptide variants of HBV genotypes A to D. Both CD4+ and CD8+ T cells transduced with the MHC II-restricted TCRs were polyfunctional, producing interferon (IFN)-γ, tumor necrosis factor (TNF)-α, interleukin (IL)-2, and granzyme B (GrzB), and killed peptide-loaded target cells. Our set of MHC class II-restricted TCRs represents an important tool for elucidating CD4+ T cell help in viral infection with potential benefit for T cell therapy.

Peritoneal metastases of rare carcinomas treated with cytoreductive surgery and HIPEC - A single center case series.

INTRODUCTION: In selected cases, cytoreductive surgery (CRS) and hyperthermic intraperitoneal chemotherapy (HIPEC) is an established treatment for patients suffering from peritoneal metastases from colorectal, ovarian, gastric or appendiceal origin. The effectiveness of this extensive has not been elucidated within other rare diseases by now. METHODS: We conducted a retrospective analysis of patients treated with CRS for peritoneal carcinomatosis during the period between July 2010 and September 2015. Exclusion criteria were adenocarcinomas of the stomach, colon, neoplasms of the appendix, mesothelioma and ovarian cancers. Aim of this study was to examine the feasibility, complication rate and survival of patients with rare diseases. RESULTS: A total of 14 Patients were included: Four rare gynecological tumors, three adenocarcinomas of the small intestine, three retroperitoneal sarcomas, one cholangiocellular carcinoma, one neuroendocrine gastric tumor, one malignant peripheral nerve sheath tumor and one cancer of unknown primary syndrome. In 12 of 14 patients a macroscopically complete tumorresection could be achieved. No patient died during hospitalization. Seven of 14 patients experienced general complication of grade III according to NCI CTCAE V4.0, while two experienced complications of grade IV. Median follow-up and one year overall survival were 15.5 months and 46.8%, respectively. CONCLUSION: For patients with rare tumors, CRS and HIPEC is feasible with an acceptable perioperative morbidity and mortality. To improve knowledge in patient selection and outcome, rare tumors treated with CRS and HIPEC should be documented in central databases (as for example BIG RENAPE, Pierre-Benite, France).

Hemodynamics of Pericardial Aortic Valves: Contemporary Stented versus Stentless Valves in a Matched Comparison.

PURPOSE: Hemodynamic performance of aortic valve bioprostheses is essential for reliable function and durability. So far, the supra-annularly implanted stentless Sorin Freedom Solo (SFS) demonstrated unsurpassed hemodynamic properties. As contemporary stented and externally mounted pericardial bioprostheses, like the Labcor Dokimos Plus (LDP), also improve hemodynamic performance, these types of valves were compared in this study. METHODS: A total of 218 patients, who underwent aortic valve replacement with the LDP or the SFS, were matched retrospectively 1:1 on variables affecting hemodynamic measurements: implanted valve size, age, sex, and body surface area (BSA). With matching tolerance for valve size and gender of 0%, for age and BSA of 5%, 57 patient-pairs were yielded. Operative data, clinical, and hemodynamic outcome were analyzed. RESULTS: Except for slightly higher left ventricular function and lower procedural times in the SFS group, preoperative, operative, and postoperative characteristics of patient-pairs did not differ significantly. Mean pressure gradients, effective orifice areas (EOAs), and indexed EOAs were comparable. Corresponding to valve sizes of 21, 23, 25, and 27 mm, the indexed EOAs of the LDP and SFS prostheses were 1.08 ± 0.33, 0.92 ± 0.19, 0.93 ± 0.24, 0.99 ± 0.13 cm2/m2 and 0.81 ± 0.13, 0.92 ± 0.28, 0.95 ± 0.20, 1.04 ± 0.27 cm2/m2, respectively. CONCLUSION: Contemporary stented and stentless pericardial bioprostheses showed excellent hemodynamic properties without significant differences in EOAs and indexed EOAs.

Human T cell immune surveillance: Phenotypic, functional and migratory heterogeneity for tailored immune responses.

The human immune system constantly provides a balance between pathogen clearance as well as tolerance for autoantigens and the commensal microbiota. This is achieved by immune responses, which are highly specialized and diversified in terms of their phenotype, function, regulation and location. Despite the complexity that is inherent to human immunity, our current knowledge is primarily shaped by very reductionist insights gained from peripheral blood T cells. Since only 2% of human T cells recirculate in the blood, the vast majority remains undetected by common sampling strategies and therefore unexplored. This review highlights and discusses recent developments in human T cell immune surveillance with a particular focus on functional and migratory T cell heterogeneity and provides a critical framework for new conceptual ideas, which could serve as a starting point in the quest for novel targeted therapies for chronic tissue restricted inflammatory diseases.

Epstein-Barr viral miRNAs inhibit antiviral CD4+ T cell responses targeting IL-12 and peptide processing.

Epstein-Barr virus (EBV) is a tumor virus that establishes lifelong infection in most of humanity, despite eliciting strong and stable virus-specific immune responses. EBV encodes at least 44 miRNAs, most of them with unknown function. Here, we show that multiple EBV miRNAs modulate immune recognition of recently infected primary B cells, EBV's natural target cells. EBV miRNAs collectively and specifically suppress release of proinflammatory cytokines such as IL-12, repress differentiation of naive CD4(+) T cells to Th1 cells, interfere with peptide processing and presentation on HLA class II, and thus reduce activation of cytotoxic EBV-specific CD4(+) effector T cells and killing of infected B cells. Our findings identify a previously unknown viral strategy of immune evasion. By rapidly expressing multiple miRNAs, which are themselves nonimmunogenic, EBV counteracts recognition by CD4(+) T cells and establishes a program of reduced immunogenicity in recently infected B cells, allowing the virus to express viral proteins required for establishment of life-long infection.

Long-Term Outcome of the Sorin Freedom SOLO Stentless Aortic Valve.

BACKGROUND AND AIM OF THE STUDY: The Sorin Freedom SOLO valve is a third-generation stentless aortic valve which shows beneficial hemodynamic performance compared to stented bioprostheses. Long-term results regarding hemodynamics, the durability of the valve, and patient outcome are scarce, and their acquisition was the aim of this single-center study. METHODS: Between 2005 and 2006, a total of 68 consecutive patients (30 males, 38 females; mean age at surgery 76.1 ± 6.3 years) underwent aortic valve replacement with a Sorin Freedom SOLO prosthesis. Indications were aortic stenosis (n = 50), aortic regurgitation (n = 1) and mixed lesions (n = 17). Associated procedures were performed in 31 patients (45.6%), most of which were coronary artery bypass grafting (68.8 %). The follow up was performed by telephone interviews, and clinical and echocardiographic examinations. Morbidity, mortality and echocardiographic data were analyzed. RESULTS: The total follow up was 501.1 patient-years, with a mean follow up of 7.4 ± 3.4 years (maximum 11.2 years). The follow up was 100% complete. Hospital mortality was 4.4% (n = 3). Actuarial survival at five and 10 years was 76.5 ± 5.1% and 41.5 ± 6.5%, respectively. Reinterventions were performed in eight patients; these included three reoperations due to endocarditis, four transcatheter aortic valve implantations, and one reoperation due to structural valve deterioration (SVD). The overall freedom from valve reintervention due to SVD at five and 10 years was 97.8 ± 2.2% and 82.9 ± 7.5%, respectively. After eight years, echocardiography demonstrated peak and mean transvalvular gradients of 18 ± 11 and 10 ± 7 mmHg, respectively. The overall effective and indexed aortic valve orifice areas were 1.73 ± 0.58 cm2 and 0.92 ± 0.33 cm2/m2, respectively. CONCLUSIONS: At long-term follow up the Sorin Freedom SOLO bioprosthesis was associated with favourable hemodynamic results and survival. Freedom from SVD was not superior to that occurring with stented bioprostheses.

IL-17 and GM-CSF expression are antagonistically regulated by human T helper cells.

Although T helper 17 (TH17) cells have been acknowledged as crucial mediators of autoimmune tissue damage, the effector cytokines responsible for their pathogenicity still remain poorly defined, particularly in humans. In mouse models of autoimmunity, the pathogenicity of TH17 cells has recently been associated with their production of granulocyte-macrophage colony-stimulating factor (GM-CSF). We analyzed the regulation of GM-CSF expression by human TH cell subsets. Surprisingly, the induction of GM-CSF expression by human TH cells is constrained by the interleukin-23 (IL-23)/ROR-γt/TH17 cell axis but promoted by the IL-12/T-bet/TH1 cell axis. IL-2-mediated signal transducer and activator of transcription 5 (STAT5) signaling induced GM-CSF expression in naïve and memory TH cells, whereas STAT3 signaling blocked it. The opposite effect was observed for IL-17 expression. Ex vivo, GM-CSF(+) TH cells that coexpress interferon-γ and T-bet could be distinguished by differential chemokine receptor expression from a previously uncharacterized subset of GM-CSF-only-producing TH cells that did not express TH1, TH2, and TH17 signature cytokines or master transcription factors. Our findings demonstrate distinct and counterregulatory pathways for the generation of IL-17- and GM-CSF-producing cells and also suggest a pathogenic role for GM-CSF(+) T cells in the inflamed brain of multiple sclerosis (MS) patients. This provides not only a scientific rationale for depleting T cell-derived GM-CSF in MS patients but also multiple new molecular checkpoints for therapeutic GM-CSF suppression, which, unlike in mice, do not associate with the TH17 but instead with the TH1 axis.

Pathogen-induced human TH17 cells produce IFN-γ or IL-10 and are regulated by IL-1ß.

IL-17-producing CD4+ T helper cells (TH17) have been extensively investigated in mouse models of autoimmunity. However, the requirements for differentiation and the properties of pathogen-induced human TH17 cells remain poorly defined. Using an approach that combines the in vitro priming of naive T cells with the ex vivo analysis of memory T cells, we describe here two types of human TH17 cells with distinct effector function and differentiation requirements. Candida albicans-specific TH17 cells produced IL-17 and IFN-γ, but no IL-10, whereas Staphylococcus aureus-specific TH17 cells produced IL-17 and could produce IL-10 upon restimulation. IL-6, IL-23 and IL-1ß contributed to TH17 differentiation induced by both pathogens, but IL-1ß was essential in C. albicans-induced TH17 differentiation to counteract the inhibitory activity of IL-12 and to prime IL-17/IFN-γ double-producing cells. In addition, IL-1ß inhibited IL-10 production in differentiating and in memory TH17 cells, whereas blockade of IL-1ß in vivo led to increased IL-10 production by memory TH17 cells. We also show that, after restimulation, TH17 cells transiently downregulated IL-17 production through a mechanism that involved IL-2-induced activation of STAT5 and decreased expression of ROR-γt. Taken together these findings demonstrate that by eliciting different cytokines C. albicans and S. aureus prime TH17 cells that produce either IFN-γ or IL-10, and identify IL-1ß and IL-2 as pro- and anti-inflammatory regulators of TH17 cells both at priming and in the effector phase.

Significant response after treatment with the mTOR inhibitor sirolimus in combination with carboplatin and paclitaxel in metastatic melanoma patients.

Melanoma is highly resistant to chemotherapy. In melanoma, the PI3K-AKT-mTOR signaling pathway is constitutively activated through multiple mechanisms. Several experimental studies suggest that targeting the PI3K-AKT-mTOR signaling pathway is a promising strategy to overcome chemoresistance. This is the first report describing a chemosensitizing effect of mTOR inhibition in patients with melanoma. We report two cases of patients with metastatic melanoma who showed significant remission after combination of carboplatin and paclitaxel with the mTOR inhibitor sirolimus. Our case report, together with the literature discussed, suggests that mTOR inhibition possibly enhances the sensitivity of melanoma cells to chemotherapy and should prompt in-depth and clinical investigation.

Naive CD4+ T cells from lupus-prone Fas-intact MRL mice display TCR-mediated hyperproliferation due to intrinsic threshold defects in activation.

Autoreactive T cell activation is a consistent feature of murine lupus; however, the mechanism of such activation remains unclear. We hypothesized that naive CD4+ T cells in lupus have a lower threshold of activation through their TCR-CD3 complex that renders them more susceptible to stimulation with self-Ags. To test this hypothesis, we compared proliferation, IL-2 production, and single cell calcium signaling of naive CD4+ T cells isolated from Fas-intact MRL/+(Fas-lpr) mice with H-2k-matched B10.BR and CBA/CaJ controls, following anti-CD3 stimulation in the presence or absence of anti-CD28. We also assessed the responsiveness of naive CD4+ T cells isolated from Fas-intact MRL and control mice bearing a rearranged TCR specific for amino acids 88-104 of pigeon cytochrome c to cognate and low affinity peptide Ags presented by bone marrow-matured dendritic cells. TCR transgenic and wild-type CD4+ T cells from MRL mice displayed a lower threshold of activation than control cells, a response that was class II MHC dependent. The rise in intracellular calcium in MRL vs controls was enhanced and prolonged following anti-CD3 triggering, suggestive of proximal defects in TCR-engendered signaling as the mechanism for the observed hyperactivity. These findings were observed as early as 1-2 mo postweaning and, based on analysis of F1 T cells, appeared to be dominantly expressed. This genetically altered threshold for activation of MRL T cells, a consequence of a proximal defect in CD3-mediated signal transduction, may contribute to the abrogation of T cell tolerance to self-Ags in lupus.