CD4+ T cells predominate in salivary gland (SG) inflammatory lesions in Sjögren’s syndrome (SS). However, their antigen specificity, degree of clonal expansion, and relationship to clinical disease features remain unknown. We used multiplex reverse-transcriptase PCR to amplify paired T cell receptor α (TCRα) and β transcripts of single CD4+CD45RA– T cells from SG and peripheral blood (PB) of 10 individuals with primary SS, 9 of whom shared the HLA DR3/DQ2 risk haplotype. TCRα and β sequences were obtained from a median of 91 SG and 107 PB cells per subject. The degree of clonal expansion and frequency of cells expressing two productively rearranged α genes were increased in SG versus PB. Expanded clones from SG exhibited complementary-determining region 3 (CDR3) sequence similarity both within and among subjects, suggesting antigenic selection and shared antigen recognition. CDR3 similarities were shared among expanded clones from individuals discordant for canonical Ro and La autoantibodies, suggesting recognition of alternative SG antigen(s). The extent of SG clonal expansion correlated with reduced saliva production and increased SG fibrosis, linking expanded SG T cells with glandular dysfunction. Knowledge of paired TCRα and β sequences enables further work toward identification of target antigens and development of novel therapies.
Michelle L. Joachims, Kerry M. Leehan, Christina Lawrence, Richard C. Pelikan, Jacen S. Moore, Zijian Pan, Astrid Rasmussen, Lida Radfar, David M. Lewis, Kiely M. Grundahl, Jennifer A. Kelly, Graham B. Wiley, Mikhail Shugay, Dmitriy M. Chudakov, Christopher J. Lessard, Donald U. Stone, R. Hal Scofield, Courtney G. Montgomery, Kathy L. Sivils, Linda F. Thompson, A. Darise Farris
Eosinophilic inflammation and Th2 cytokine production are central to the pathogenesis of asthma. Agents that target either eosinophils or single Th2 cytokines have shown benefits in subsets of biomarker-positive patients. More broadly effective treatment or disease-modifying effects may be achieved by eliminating more than one inflammatory stimulator. Here we present a strategy to concomitantly deplete Th2 T cells, eosinophils, basophils, and type-2 innate lymphoid cells (ILC2s) by generating monoclonal antibodies with enhanced effector function (19A2) that target CRTh2 present on all 4 cell types. Using human CRTh2 (hCRTh2) transgenic mice that mimic the expression pattern of hCRTh2 on innate immune cells but not Th2 cells, we demonstrate that anti-hCRTh2 antibodies specifically eliminate hCRTh2+ basophils, eosinophils, and ILC2s from lung and lymphoid organs in models of asthma and
Tao Huang, Meredith Hazen, Yonglei Shang, Meijuan Zhou, Xiumin Wu, Donghong Yan, Zhonghua Lin, Margaret Solon, Elizabeth Luis, Hai Ngu, Yongchang Shi, Arna Katewa, David F. Choy, Nandhini Ramamoorthi, Erick R. Castellanos, Mercedesz Balazs, Min Xu, Wyne P. Lee, Marissa L. Matsumoto, Jian Payandeh, Joseph R. Arron, Jo-Anne Hongo, Jianyong Wang, Isidro Hötzel, Cary D. Austin, Karin Reif
DC-based vaccines that initiate T cell responses are well tolerated and have demonstrated efficacy for tumor immunotherapy, with the potential to be combined with other therapies. Targeting vaccine antigens (Ag) directly to the DCs in vivo is more effective than cell-based therapies in mouse models and is therefore a promising strategy to translate to humans. The human CD141+ DCs are considered the most clinically relevant for initiating CD8+ T cell responses critical for killing tumors or infected cells, and they specifically express the C-type lectin-like receptor CLEC9A that facilitates presentation of Ag by these DCs. We have therefore developed a human chimeric Ab that specifically targets CLEC9A on CD141+ DCs in vitro and in vivo. These human chimeric Abs are highly effective at delivering Ag to DCs for recognition by both CD4+ and CD8+ T cells. Given the importance of these cellular responses for antitumor or antiviral immunity, and the superior specificity of anti-CLEC9A Abs for this DC subset, this approach warrants further development for vaccines.
Kirsteen M. Tullett, Ingrid M. Leal Rojas, Yoshihito Minoda, Peck S. Tan, Jian-Guo Zhang, Corey Smith, Rajiv Khanna, Ken Shortman, Irina Caminschi, Mireille H. Lahoud, Kristen J. Radford
Secreted by activated cells or passively released by damaged cells, extracellular HMGB1 is a prototypical damage-associated molecular pattern (DAMP) inflammatory mediator. During the course of developing extracorporeal approaches to treating injury and infection, we inadvertently discovered that haptoglobin, the acute phase protein that binds extracellular hemoglobin and targets cellular uptake through CD163, also binds HMGB1. Haptoglobin-HMGB1 complexes elicit the production of antiinflammatory enzymes (heme oxygenase-1) and cytokines (e.g., IL-10) in WT but not in CD163-deficient macrophages. Genetic disruption of haptoglobin or CD163 expression significantly enhances mortality rates in standardized models of intra-abdominal sepsis in mice. Administration of haptoglobin to WT and to haptoglobin gene-deficient animals confers significant protection. These findings reveal a mechanism for haptoglobin modulation of the inflammatory action of HMGB1, with significant implications for developing experimental strategies targeting HMGB1-dependent inflammatory diseases.
Huan Yang, Haichao Wang, Yaakov A. Levine, Manoj K. Gunasekaran, Yongjun Wang, Meghan Addorisio, Shu Zhu, Wei Li, Jianhua Li, Dominique P.V. de Kleijn, Peder S. Olofsson, H. Shaw Warren, Mingzhu He, Yousef Al-Abed, Jesse Roth, Daniel J. Antoine, Sangeeta S. Chavan, Ulf Andersson, Kevin J. Tracey
Central clonal deletion has been considered the critical factor responsible for the robust state of tolerance achieved by chimerism-based experimental protocols, but split-tolerance models and the clinical experience are calling this assumption into question. Although clone-size reduction through deletion has been shown to be universally required for achieving allotolerance, it remains undetermined whether it is sufficient by itself. Therapeutic Treg treatment induces chimerism and tolerance in a stringent murine BM transplantation model devoid of myelosuppressive recipient treatment. In contrast to irradiation chimeras, chronic rejection (CR) of skin and heart allografts in Treg chimeras was permanently prevented, even in the absence of complete clonal deletion of donor MHC-reactive T cells. We show that minor histocompatibility antigen mismatches account for CR in irradiation chimeras without global T cell depletion. Furthermore, we show that Treg therapy–induced tolerance prevents CR in a linked suppression–like fashion, which is maintained by active regulatory mechanisms involving recruitment of thymus-derived Tregs to the graft. These data suggest that highly efficient intrathymic and peripheral deletion of donor-reactive T cells for specificities expressed on hematopoietic cells preclude the expansion of donor-specific Tregs and, hence, do not allow for spreading of tolerance to minor specificities that are not expressed by donor BM.
Nina Pilat, Benedikt Mahr, Lukas Unger, Karin Hock, Christoph Schwarz, Andreas M. Farkas, Ulrike Baranyi, Fritz Wrba, Thomas Wekerle
To date, the major target of biologic therapeutics in systemic lupus erythematosus (SLE) has been the B cell, which produces pathogenic autoantibodies. Recently, targeting type I IFN, which is elaborated by plasmacytoid dendritic cells (pDCs) in response to endosomal TLR7 and TLR9 stimulation by SLE immune complexes, has shown promising results. pDCs express high levels of the IL-3Rα chain (CD123), suggesting an alternative potential targeting strategy. We have developed an anti-CD123 monoclonal antibody, CSL362, and show here that it affects key cell types and cytokines that contribute to SLE. CSL362 potently depletes pDCs via antibody-dependent cell-mediated cytotoxicity, markedly reducing TLR7, TLR9, and SLE serum-induced IFN-α production and IFN-α-upregulated gene expression. The antibody also inhibits TLR7- and TLR9-induced plasmablast expansion by reducing IFN-α and IL-6 production. These effects are more pronounced than with IFN-α blockade alone, possibly because pDC depletion reduces production of other IFN subtypes, such as type III, as well as non-IFN proinflammatory cytokines, such as IL-6. In addition, CSL362 depletes basophils and inhibits IL-3 signaling. These effects were confirmed in cells derived from a heterogeneous population of SLE donors, various IFN-dependent autoimmune diseases, and healthy controls. We also demonstrate in vivo activity of CSL362 following its s.c. administration to cynomolgus monkeys. This spectrum of effects provides a preclinical rationale for the therapeutic evaluation of CSL362 in SLE.
Shereen Oon, Huy Huynh, Tsin Yee Tai, Milica Ng, Katherine Monaghan, Mark Biondo, Gino Vairo, Eugene Maraskovsky, Andrew D. Nash, Ian P. Wicks, Nicholas J. Wilson
Gastrointestinal graft-versus-host-disease (GI-GVHD) is a life-threatening complication occurring after allogeneic hematopoietic cell transplantation (HCT), and a blood biomarker that permits stratification of HCT patients according to their risk of developing GI-GVHD would greatly aid treatment planning. Through in-depth, large-scale proteomic profiling of presymptomatic samples, we identified a T cell population expressing both CD146, a cell adhesion molecule, and CCR5, a chemokine receptor that is upregulated as early as 14 days after transplantation in patients who develop GI-GVHD. The CD4+CD146+CCR5+ T cell population is Th17 prone and increased by ICOS stimulation. shRNA knockdown of CD146 in T cells reduced their transmigration through endothelial cells, and maraviroc, a CCR5 inhibitor, reduced chemotaxis of the CD4+CD146+CCR5+ T cell population toward CCL14. Mice that received CD146 shRNA–transduced human T cells did not lose weight, showed better survival, and had fewer CD4+CD146+CCR5+ T cells and less pathogenic Th17 infiltration in the intestine, even compared with mice receiving maraviroc with control shRNA–transduced human T cells. Furthermore, the frequency of CD4+CD146+CCR5+ Tregs was increased in GI-GVHD patients, and these cells showed increased plasticity toward Th17 upon ICOS stimulation. Our findings can be applied to early risk stratification, as well as specific preventative therapeutic strategies following HCT.
Wei Li, Liangyi Liu, Aurelie Gomez, Jilu Zhang, Abdulraouf Ramadan, Qing Zhang, Sung W. Choi, Peng Zhang, Joel K. Greenson, Chen Liu, Di Jiang, Elizabeth Virts, Stephanie L. Kelich, Hong Wei Chu, Ryan Flynn, Bruce R. Blazar, Helmut Hanenberg, Samir Hanash, Sophie Paczesny
BACKGROUND. Some adult patients presenting with unexplained pyrexia, serositis, skin rashes, arthralgia, myalgia, and other symptoms commonly found in autoinflammatory disorders may not fit a specific diagnosis, either because their clinical phenotype is nondiagnostic or genetic tests are negative. We used the term undifferentiated systemic autoinflammatory disorder (uSAID) to describe such cases. Given that well-defined autoinflammatory diseases show responses to IL-1 blockade, we evaluated whether anakinra was useful for both diagnosing and treating uSAID patients.
METHODS. We performed a retrospective analysis of consecutive patients presenting with uSAID between 2012–2015 who were treated with the recombinant IL-1 receptor antagonist anakinra. uSAID was diagnosed after excluding malignancy, infection, and pathogenic mutations in known hereditary fever syndromes (HFS) genes and where clinical criteria for adult onset Still’s disease (AOSD) were not met.
RESULTS. A total of 11 patients presented with uSAID (5 males and 6 females), with a mean time to diagnosis of 3.5 years (1–8 years). Patients were unresponsive or only partially controlled on disease-modifying antirheumatic drug (DMARD)/steroid treatment. Anakinra controlled symptoms within 4–6 weeks of starting treatment in 9 of 11 cases. Two patients discontinued therapy — one due to incomplete response and another due to severe injection-site reactions.
CONCLUSION. This retrospective case series demonstrates that the spectrum of poorly defined autoinflammatory disorders that show responsiveness to anakinra is considerable. Anakinra seems a viable treatment option for these patients, who are unresponsive to standard steroid/DMARD treatments. Moreover, given the mechanisms of action, response to anakinra implicates underlying IL-1 dysregulation in the disease pathogenesis of responding uSAIDs patients.
Stephanie R. Harrison, Dennis McGonagle, Sharmin Nizam, Stephen Jarrett, Jeroen van der Hilst, Michael F. McDermott, Sinisa Savic
Rheumatoid arthritis (RA) is a debilitating disease characterized by persistent accumulation of leukocytes within the articular cavity and synovial tissue. Metabololipidomic profiling of arthritic joints from omega-3 supplemented mice identified elevated levels of specialized proresolving lipid mediators (SPM) including resolvin D1 (RvD1). Profiling of human RA synovial fluid revealed physiological levels of RvD1, which — once applied to human neutrophils — attenuated chemotaxis. These results prompted analyses of the antiarthritic properties of RvD1 in a model of murine inflammatory arthritis. The stable epimer 17
Lucy V. Norling, Sarah E. Headland, Jesmond Dalli, Hildur H. Arnardottir, Oliver Haworth, Hefin R. Jones, Daniel Irimia, Charles N. Serhan, Mauro Perretti
Immunotherapies targeting the immune checkpoint receptor programmed cell death protein 1 (PD-1) have shown remarkable efficacy in treating cancer. CD4+CD25hiFoxP3+ Tregs are critical regulators of immune responses in autoimmunity and malignancies, but the functional status of human Tregs expressing PD-1 remains unclear. We examined functional and molecular features of PD-1hi Tregs in healthy subjects and patients with glioblastoma multiforme (GBM), combining functional assays, RNA sequencing, and cytometry by time of flight (CyTOF). In both patients with GBM and healthy subjects, circulating PD-1hi Tregs displayed reduced suppression of CD4+ effector T cells, production of IFN-γ, and molecular signatures of exhaustion. Transcriptional profiling of tumor-resident Tregs revealed that several genes coexpressed with PD-1 and associated with IFN-γ production and exhaustion as well as enrichment in exhaustion signatures compared with circulating PD-1hi Tregs. CyTOF analysis of circulating and tumor-infiltrating Tregs from patients with GBM treated with PD-1-blocking antibodies revealed that treatment shifts the profile of circulating Tregs toward a more exhausted phenotype reminiscent of that of tumor-infiltrating Tregs, further increasing IFN-γ production. Thus, high PD-1 expression on human Tregs identifies dysfunctional, exhausted Tregs secreting IFN-γ that exist in healthy individuals and are enriched in tumor infiltrates, possibly losing function as they attempt to modulate the antitumoral immune responses.
Daniel E. Lowther, Brittany A. Goods, Liliana E. Lucca, Benjamin A. Lerner, Khadir Raddassi, David van Dijk, Amanda L. Hernandez, Xiangguo Duan, Murat Gunel, Vlad Coric, Smita Krishnaswamy, J. Christopher Love, David A. Hafler
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