Nivolumab plus Brentuximab vedotin +/- bendamustine combination therapy: a safe and effective treatment in pediatric recurrent and refractory classical Hodgkin lymphoma.

Introduction: Classical Hodgkin lymphoma (cHL) is the most common pediatric lymphoma. Approximately 10% of patients develop refractory or recurrent disease. These patients are treated with intensive chemotherapy followed by consolidation with radiotherapy or high-dose chemotherapy and autologous stem cell reinfusion. Although this treatment is effective, it comes at the cost of severe long-term adverse events, such as reduced fertility and an increased risk of secondary cancers. Recently, promising results of inducing remission with the immune checkpoint inhibitor nivolumab (targeting PD-1) and the anti-CD30 antibody-drug conjugate Brentuximab vedotin (BV) +/- bendamustine were published. Methods: Here we describe a cohort of 10 relapsed and refractory pediatric cHL patients treated with nivolumab + BV +/- bendamustine to induce remission prior to consolidation with standard treatment. Results and discussion: All patients achieved complete remission prior to consolidation treatment and are in ongoing complete remission with a median follow-up of 25 months (range: 12 to 42 months) after end-of-treatment. Only one adverse event of CTCAE grade 3 or higher due to nivolumab + BV was identified. Based on these results we conclude that immunotherapy with nivolumab + BV +/- bendamustine is an effective and safe treatment to induce remission in pediatric R/R cHL patients prior to standard consolidation treatment. We propose to evaluate this treatment further to study putative long-term toxicity and the possibility to reduce the intensity of consolidation treatment.

Regulatory T cells in psoriatic arthritis: an IL-17A-producing, Foxp3intCD161 + RORγt + ICOS + phenotype, that associates with the presence of ADAMTSL5 autoantibodies.

In psoriatic arthritis (PsA), predisposing class I HLA alleles, the presence of synovial clonally proliferated CD8 + T cells and autoantibodies all point towards the loss of immune tolerance. However, the key mechanisms that lead to immune dysregulation are not fully understood. In other types of inflammatory arthritis, T regulatory cell (Treg) dysfunction and plasticity at sites of inflammation were suggested to negatively affect peripheral tolerance. We here addressed if Treg variances associate with psoriatic disease. We collected clinical data, sera and peripheral blood mononuclear cells from 13 healthy controls, 21 psoriasis and 21 PsA patients. In addition, we obtained synovial fluid mononuclear cells from 6 PsA patients. We studied characteristics of CD4 + CD25 + CD127loFoxp3 + Tregs by flow cytometry and used ELISA to quantify antibodies against ADAMTSL5, a recently discovered autoantigen in psoriatic disease. In comparison with their circulating counterparts, Tregs from inflamed joints express increased levels of ICOS, CTLA-4 and TIGIT. Furthermore, synovial fluid-derived Tregs have a distinct phenotype, characterized by IL-17A production and upregulation of CD161 and RORγt. We identified a subset of Tregs with intermediate Foxp3 expression as the major cytokine producer. Furthermore, ICOS + Tregs associate with PsA disease activity as measured by PASDAS. Lastly, we observed that presence of the Foxp3int Tregs associates with an increased abundance of anti-ADAMTSL5 autoantibodies. Tregs derived from the inflammatory environment of inflamed PsA joints exhibit a distinct phenotype, which associates with loss of peripheral immune tolerance in psoriatic disease.

Preclinical Aortic Atherosclerosis in Adolescents With Chronic Disease.

Background Adolescents with chronic disease are often exposed to inflammatory, metabolic, and hemodynamic risk factors for early atherosclerosis. Since postmortem studies have shown that atherogenesis starts in the aorta, the CDACD (Cardiovascular Disease in Adolescents with Chronic Disease) study investigated preclinical aortic atherosclerosis in these adolescents. Methods and Results The cross-sectional CDACD study enrolled 114 adolescents 12 to 18 years old with chronic disorders including juvenile idiopathic arthritis, cystic fibrosis, obesity, corrected coarctation of the aorta, and healthy controls with a corrected atrial septal defect. Cardiovascular magnetic resonance was used to assess aortic pulse wave velocity and aortic wall thickness, as established aortic measures of preclinical atherosclerosis. Cardiovascular magnetic resonance showed a higher aortic pulse wave velocity, which reflects aortic stiffness, and higher aortic wall thickness in all adolescent chronic disease groups, compared with controls (P<0.05). Age (ß=0.253), heart rate (ß=0.236), systolic blood pressure (ß=-0.264), and diastolic blood pressure (ß=0.365) were identified as significant predictors for aortic pulse wave velocity, using multivariable linear regression analysis. Aortic wall thickness was predicted by body mass index (ß=0.248) and fasting glucose (ß=0.242), next to aortic lumen area (ß=0.340). Carotid intima-media thickness was assessed using ultrasonography, and was only higher in adolescents with coarctation of the aorta, compared with controls (P<0.001). Conclusions Adolescents with chronic disease showed enhanced aortic stiffness and wall thickness compared with controls. The enhanced aortic pulse wave velocity and aortic wall thickness in adolescents with chronic disease could indicate accelerated atherogenesis. Our findings underscore the importance of the aorta for assessment of early atherosclerosis, and the need for tailored cardiovascular follow-up of children with chronic disease.

Immunometabolic factors in adolescent chronic disease are associated with Th1 skewing of invariant Natural Killer T cells.

Invariant Natural Killer T (iNKT) cells respond to the ligation of lipid antigen-CD1d complexes via their T-cell receptor and are implicated in various immunometabolic diseases. We considered that immunometabolic factors might affect iNKT cell function. To this end, we investigated iNKT cell phenotype and function in a cohort of adolescents with chronic disease and immunometabolic abnormalities. We analyzed peripheral blood iNKT cells of adolescents with cystic fibrosis (CF, n = 24), corrected coarctation of the aorta (CoA, n = 25), juvenile idiopathic arthritis (JIA, n = 20), obesity (OB, n = 20), and corrected atrial septal defect (ASD, n = 25) as controls. To study transcriptional differences, we performed RNA sequencing on a subset of obese patients and controls. Finally, we performed standardized co-culture experiments using patient plasma, to investigate the effect of plasma factors on iNKT cell function. We found comparable iNKT cell numbers across patient groups, except for reduced iNKT cell numbers in JIA patients. Upon ex-vivo activation, we observed enhanced IFN-γ/IL-4 cytokine ratios in iNKT cells of obese adolescents versus controls. The Th1-skewed iNKT cell cytokine profile of obese adolescents was not explained by a distinct transcriptional profile of the iNKT cells. Co-culture experiments with patient plasma revealed that across all patient groups, obesity-associated plasma factors including LDL-cholesterol, leptin, and fatty-acid binding protein 4 (FABP4) coincided with higher IFN-γ production, whereas high HDL-cholesterol and insulin sensitivity (QUICKI) coincided with higher IL-4 production. LDL and HDL supplementation in co-culture studies confirmed the effects of lipoproteins on iNKT cell cytokine production. These results suggest that circulating immunometabolic factors such as lipoproteins may be involved in Th1 skewing of the iNKT cell cytokine response in immunometabolic disease.

Tissue-Resident Memory CD8+ T Cells From Skin Differentiate Psoriatic Arthritis From Psoriasis.

OBJECTIVE: To compare immune cell phenotype and function in psoriatic arthritis (PsA) versus psoriasis in order to better understand the pathogenesis of PsA. METHODS: In-depth immunophenotyping of different T cell and dendritic cell subsets was performed in patients with PsA, psoriasis, or axial spondyloarthritis and healthy controls. Subsequently, we analyzed cells from peripheral blood, synovial fluid (SF), and skin biopsy specimens using flow cytometry, along with high-throughput transcriptome analyses and functional assays on the specific cell populations that appeared to differentiate PsA from psoriasis. RESULTS: Compared to healthy controls, the peripheral blood of patients with PsA was characterized by an increase in regulatory CD4+ T cells and interleukin-17A (IL-17A) and IL-22 coproducing CD8+ T cells. One population specifically differentiated PsA from psoriasis: i.e., CD8+CCR10+ T cells were enriched in PsA. CD8+CCR10+ T cells expressed high levels of DNAX accessory molecule 1 and were effector memory cells that coexpressed skin-homing receptors CCR4 and cutaneous lymphocyte antigen. CD8+CCR10+ T cells were detected under inflammatory and homeostatic conditions in skin, but were not enriched in SF. Gene profiling further revealed that CD8+CCR10+ T cells expressed GATA3, FOXP3, and core transcriptional signature of tissue-resident memory T cells, including CD103. Specific genes, including RORC, IFNAR1, and ERAP1, were up-regulated in PsA compared to psoriasis. CD8+CCR10+ T cells were endowed with a Tc2/22-like cytokine profile, lacked cytotoxic potential, and displayed overall regulatory function. CONCLUSION: Tissue-resident memory CD8+ T cells derived from the skin are enhanced in the circulation of patients with PsA compared to patients with psoriasis alone. This may indicate that aberrances in cutaneous tissue homeostasis contribute to arthritis development.

Upcoming immunotherapeutic combinations for B-cell lymphoma.

After initial introduction for B-cell lymphomas as adjuvant therapies to established cancer treatments, immune checkpoint inhibitors and other immunotherapies are now integrated in mainstream regimens, both in adult and pediatric patients. We here provide an overview of the current status of combination therapies for B-cell lymphoma, by in-depth analysis of combination therapy trials registered between 2015-2020. Our analysis provides new insight into the rapid evolution in lymphoma treatment, as propelled by new additions to the treatment arsenal. We conclude with prospects on upcoming clinical trials which will likely use systematic testing approaches of more combinations of established chemotherapy regimens with new agents, as well as new combinations of immunotherapy and targeted therapy. Future trials will be set up as basket or umbrella-type trials to facilitate the evaluation of new drugs targeting specific genetic changes in the tumor or associated immune microenvironment. As such, lymphoma patients will benefit by receiving more tailored treatment that is based on synergistic effects of chemotherapy combined with new agents targeting specific aspects of tumor biology and the immune system.

DNAM1 and TIGIT balance the T cell response, with low T cell TIGIT expression corresponding to inflammation in psoriatic disease.

Objectives: Signals at the contact site of antigen-presenting cells (APCs) and T cells help orchestrate the adaptive immune response. CD155 on APCs can interact with the stimulatory receptor DNAM1 or inhibitory receptor TIGIT on T cells. The CD155/DNAM1/TIGIT axis is under extensive investigation as immunotherapy target in inflammatory diseases including cancer, chronic infection and autoimmune diseases. We investigated a possible role for CD155/DNAM1/TIGIT signaling in psoriatic disease. Methods: By flow cytometry, we analyzed peripheral blood mononuclear cells of patients with psoriasis (n = 20) or psoriatic arthritis (n = 21), and healthy individuals (n = 7). We measured CD155, TIGIT, and DNAM1 expression on leukocyte subsets and compared activation-induced cytokine production between CD155-positive and CD155-negative APCs. We assessed the effects of TIGIT and DNAM1 blockade on T cell activation, and related the expression of CD155/DNAM1/TIGIT axis molecules to measures of disease activity. Results: High CD155 expression associates with tumor necrosis factor (TNF) production in myeloid and plasmacytoid dendritic cells (DC). In CD1c+ myeloid DC, activation-induced CD155 expression associates with increased HLA-DR expression. CD8 T cells - but not CD4 T cells - express high levels of TIGIT. DNAM1 blockade decreases T cell pro-inflammatory cytokine production, while TIGIT blockade increased T cell proliferation. Finally, T cell TIGIT expression shows an inverse correlation with inflammation biomarkers in psoriatic disease. Conclusion: CD155 is increased on pro-inflammatory APCs, while the receptors DNAM1 and TIGIT expressed on T cells balance the inflammatory response by T cells. In psoriatic disease, low TIGIT expression on T cells is associated with systemic inflammation.

Broad proteomic screen reveals shared serum proteomic signature in patients with psoriatic arthritis and psoriasis without arthritis.

OBJECTIVE: To identify novel serum proteins involved in the pathogenesis of PsA as compared with healthy controls, psoriasis (Pso) and AS, and to explore which proteins best correlated to major clinical features of the disease. METHODS: A high-throughput serum biomarker platform (Olink) was used to assess the level of 951 unique proteins in serum of patients with PsA (n = 20), Pso (n = 18) and AS (n = 19), as well as healthy controls (HC, n = 20). Pso and PsA were matched for Psoriasis Area and Severity Index (PASI) and other clinical parameters. RESULTS: We found 68 differentially expressed proteins (DEPs) in PsA as compared with HC. Of those DEPs, 48 proteins (71%) were also dysregulated in Pso and/or AS. Strikingly, there were no DEPs when comparing PsA with Pso directly. On the contrary, hierarchical cluster analysis and multidimensional scaling revealed that HC clustered distinctly from all patients, and that PsA and Pso grouped together. The number of swollen joints had the strongest positive correlation to ICAM-1 (r = 0.81, P < 0.001) and CCL18 (0.76, P < 0.001). PASI score was best correlated to PI3 (r = 0.54, P < 0.001) and IL-17 receptor A (r = -0.51, P < 0.01). There were more proteins correlated to PASI score when analysing Pso and PsA patients separately, as compared with analysing Pso and PsA patients pooled together. CONCLUSION: PsA and Pso patients share a serum proteomic signature, which supports the concept of a single psoriatic spectrum of disease. Future studies should target skin and synovial tissues to uncover differences in local factors driving arthritis development in Pso.

Increased intra-articular granzyme M may trigger local IFN-λ1/IL-29 response in rheumatoid arthritis.

OBJECTIVES: Granzymes are serine proteases involved in eliminating tumour cells and virally infected cells. In addition, extracellular granzyme levels are elevated in inflammatory conditions, including several types of infection and autoimmune diseases, such as rheumatoid arthritis (RA). While GrA and GrB have been associated with RA, a role for the other three granzymes (GrH, GrK, and GrM) in this disease remains unclear. Here, we aimed to investigate the presence and role of GrM and GrK in serum and synovial fluid of patients with RA, psoriatic arthritis, and osteoarthritis. METHODS: Granzyme levels were determined in serum, synovial fluid, peripheral blood mononuclear cells (PBMCs) and synovial fluid mononuclear cells (SFMCs) of RA patients and relevant control groups. In addition, the link between GrM and inflammatory cytokines in synovial fluid was investigated. RESULTS: Serum GrM and GrK levels were not affected in RA. GrM, but not GrK, levels were elevated in synovial fluid of RA patients. GrM was mainly expressed by cytotoxic lymphocytes in SFMCs with a similar expression pattern as compared with PBMCs. Intra-articular GrM expression correlated with IL-25, IL-29, XCL1, and TNFα levels. Intriguingly, purified GrM triggered the release of IL-29 (IFN-λ1) from human fibroblasts in vitro. CONCLUSIONS: These data indicate that GrM levels are increased in RA synovial fluid and that GrM can stimulate proinflammatory IL-29 release from fibroblasts, suggesting a role of GrM in the pathogenesis of RA.

Emerging molecular biomarkers for predicting therapy response in psoriatic arthritis: A review of literature.

Psoriatic arthritis (PsA) is a heterogeneous, chronic inflammatory musculoskeletal disorder that affects ~0.1% of the population. PsA may severely impact quality-of-life and constitutes a significant economic burden on our health care system. While early effective treatment is deemed essential to prevent irreversible joint damage and functional impairment, not all patients respond to the same disease modifying anti-rheumatic drugs (DMARDs). DMARD options for PsA are rapidly evolving, yet only 50-60% of patients show a satisfactory response to their first-line DMARD therapy. Hence, there is an urgent medical need to predict which patients benefit from a particular treatment. To this end, molecular biomarkers capable of predicting therapeutic response are currently being scrutinized in clinical studies, that together should build a framework for clinical guidelines that improve personalized targeted treatment. In this review new developments within the field of biomarker discovery for predicting therapeutic response to DMARDs in PsA are examined.

CXCL4 is a driver of cytokine mRNA stability in monocyte-derived dendritic cells.

The chemokine CXCL4 has been implicated in several immune diseases. Exposure of monocyte-derived dendritic cells (moDCs) to CXCL4 potentiates the production of inflammatory cytokines in the presence of TLR3 or TLR7/8 agonists. Here we investigated the transcriptional and post-transcriptional events underlying the augmented inflammatory responses in CXCL4-moDCs. Our results indicate that CXCL4-moDCs display an increased expression and secretion of IL-12, IL-23, IL-6 and TNF upon TLR3 activation. Analysis of the cytokine transcripts for the presence of AU-rich elements (ARE), motifs necessary for ARE-mediated mRNA decay, revealed that all these cytokine transcripts are, at least in silico, possibly regulated at the level of mRNA stability. In vitro assays confirmed that mRNA stability of IL6 and TNF, but not IL12B and IL23A, is increased in CXCL4-moDCs. We next screened the expression of ARE-binding proteins (ARE-BPs) and found that TLR stimulation of CXCL4-moDCs induced tristetraprolin (TTP or ZFP36). Increased TTP mRNA expression was found to be a consequence of TTP phospho-mediated inactivation, which over time causes the protein to degrade its own mRNA. Concomitantly with TTP inactivation, we observed increased MAPK p38 signalling, upstream of TTP, in stimulated CXCL4-moDCs. P38 inhibition restored TTP activation and subsequently reduced the production of inflammatory cytokines. Finally, TTP knockdown in moDCs resulted in an increased production of IL6 and TNF after TLR stimulation. Overall, our study shows that the pro-inflammatory phenotype of CXCL4-moDCs relies in part on enhanced cytokine mRNA stability dictated by TTP inactivation.

An update on the 'danger theory' in inhibitor development in hemophilia A.

INTRODUCTION: Nowadays, one of the most serious treatment complications in hemophilia A is the formation of neutralizing antibodies against coagulation factor VIII (FVIII). These so-called inhibitors develop in about 30% of all patients with severe hemophilia A. Once formed, inhibitors reduce FVIII efficacy in blood coagulation, which has a negative impact on patients' health and quality of life and significantly increases hemophilia A treatment costs. The pathophysiology of inhibitor development is a complex and multi-causal process, in which both genetic factors as well as environmental factors participate. So-called 'danger signals' are considered contributors to inhibitor formation, and can be triggered by surgery, joint bleeds or infections. A pro-inflammatory tissue micro-environment is thereby established, which is characterized by the upregulation of costimulatory molecules on antigen-presenting cells (APCs), that can facilitate the alloimmunization to FVIII and thereby inhibitor formation. Here, the authors will discuss evidence from (pre)clinical studies about this theory in hemophilia A. Areas covered: In this review, the current knowledge regarding the 'danger theory' with regard to inhibitor development in hemophilia A is summarized. Expert opinion: Danger signals might contribute to inhibitor development; however, the evidence is scarce and not conclusive. Future studies, like multinational registries, are warranted but challenging.

Bystander T-Cells Support Clonal T-Cell Activation by Controlling the Release of Dendritic Cell-Derived Immune-Stimulatory Extracellular Vesicles.

Extracellular vesicles (EV) that are released by immune cells are studied intensively for their functions in immune regulation and are scrutinized for their potential in human immunotherapy, for example against cancer. In our search for signals that stimulate the release of functional EV by dendritic cells we observed that LPS-activated human monocyte-derived dendritic cells (moDC) changed their morphological characteristics upon contact with non-cognate activated bystander T-cells, while non-activated bystander T-cells had no effect. Exposure to activated bystander T-cells also stimulated the release of EV-associated proteins by moDC, particularly CD63, and ICAM-1, although the extent of stimulation varied between individual donors. Stimulation of moDC with activated bystander T-cells also increased the release of EV-associated miR155, which is a known central modulator of T-cell responses. Functionally, we observed that EV from moDC that were licensed by activated bystander T-cells exhibited a capacity for antigen-specific T-cell activation. Taken together, these results suggest that non-cognatei interactions between DC and bystander T-cells modulates third party antigen-specific T-cell responses via EV.

NFκB and MHC-1 Interplay in Neuroblastoma and Immunotherapy.

Pediatric neuroblastoma tumors are notorious nonimmunogenic cancers. In contrast to adult tumor types, neuroblastoma cells express low MHC-1, a derivative of its embryonic cell origin expressing little MHC-1. We here address the role of the nuclear factor kappa B (NFκB) pathway in controlling MHC-1 expression in embryonic neural crest cells, during differentiation of healthy cells, and in neuroblastoma tumors. Implications for immunotherapy are discussed.

Nedd4-Binding Protein 1 and TNFAIP3-Interacting Protein 1 Control MHC-1 Display in Neuroblastoma.

: Neuroblastoma is the second most common tumor in children. The cause of neuroblastoma is thought to lie in aberrant development of embryonic neural crest cells and is accompanied by low MHC-1 expression and suppression of the NF-κB transcription factor, thereby gearing cells toward escape from immunosurveillance. Here, we assess regulation of the MHC-1 gene in neuroblastoma to enhance its immunogenic potential for therapeutic T-cell targeting. A genome-wide CRISPR screen identified N4BP1 and TNIP1 as inhibitory factors of NF-κB-mediated MHC-1 expression in neuroblastoma. Patients with advanced stage neuroblastoma who expressed high levels of TNIP1 and N4BP1 exhibited worse overall survival. Depletion of N4BP1 or TNIP1 increased NF-κB and MHC-1 expression and stimulated recognition by antigen-specific CD8+ T cells. We confirmed that TNIP1 inhibited canonical NF-κB member RelA by preventing activation of the RelA/p50 NF-κB dimer. Furthermore, N4BP1 inhibited both canonical and noncanonical NF-κB through binding of deubiquitinating enzyme CEZANNE, resulting in stabilization of TRAF3 and degradation of NF-κB-inducing kinase NIK. These data suggest that N4BP1/CEZANNE or TNIP1 may be candidate targets for immunotherapy in neuroblastoma tumors and should lift NF-κB suppression, thereby triggering increased peptide/MHC1-mediated tumor reactivity to enhance therapeutic T-cell targeting. SIGNIFICANCE: Aberrant regulation of NF-κB and MHC-1 in neuroblastoma tumors provides new targets for immunotherapeutic approaches against neuroblastoma.

Increased risk of hematologic malignancies in primary immunodeficiency disorders: opportunities for immunotherapy.

Primary immunodeficiency disorders (PIDs) convey increased susceptibility to infections and sometimes to malignancies, particularly lymphomas. Such cancer development can be contributed by immune impairments resulting in weakened immunological surveillance against (pre)malignant cells and oncogenic viruses. Molecular defects in PID-patients are therefore being clarified, identifying new targets for innovative immunotherapy. Particularly pediatric cancers are being scrutinized, where over one-third of cancer-related deaths are accounted for by leukemia and lymphomas. Here we review how immunopathogenic mechanisms of several PIDs might associate with lymphoma development. We furthermore delineate existing immunotherapy strategies in adults for potential therapeutic application in childhood leukemia and lymphomas.

Endocytosed soluble cowpox virus protein CPXV012 inhibits antigen cross-presentation in human monocyte-derived dendritic cells.

Viruses may interfere with the MHC class I antigen presentation pathway in order to avoid CD8+ T cell-mediated immunity. A key target within this pathway is the peptide transporter TAP. This transporter plays a central role in MHC class I-mediated peptide presentation of endogenous antigens. In addition, TAP plays a role in antigen cross-presentation of exogenously derived antigens by dendritic cells (DCs). In this study, a soluble form of the cowpox virus TAP inhibitor CPXV012 is synthesized for exogenous delivery into the antigen cross-presentation route of human monocyte-derived (mo)DCs. We show that soluble CPXV012 localizes to TAP+ compartments that carry internalized antigen and is a potent inhibitor of antigen cross-presentation. CPXV012 stimulates the prolonged deposition of antigen fragments in storage compartments of moDCs, as a result of reduced endosomal acidification and reduced antigen proteolysis when soluble CPXV012 is present. Thus, a dual function can be proposed for CPXV012: inhibition of TAP-mediated peptide transport and inhibition of endosomal antigen degradation. We propose this second function for soluble CPXV012 can serve to interfere with antigen cross-presentation in a peptide transport-independent manner.