Rupali Das

Blockade of interleukin-6 signaling augments regulatory T-cell reconstitution and attenuates the severity of graft-versus-host disease.

Graft-versus-host disease (GVHD) is the major complication after allogeneic bone marrow transplantation and is characterized by the overproduction of proinflammatory cytokines. In this study, we have identified interleukin-6 (IL-6) as a critical inflammatory cytokine that alters the balance between the effector and regulatory arms of the immune system and drives a proinflammatory phenotype that is a defining characteristic of GVHD. Our results demonstrate that inhibition of the IL-6 signaling pathway by way of antibody-mediated blockade of the IL-6 receptor (IL-6R) markedly reduces pathologic damage attributable to GVHD. This is accompanied by a significant increase in the absolute number of regulatory T cells (Tregs) that is due to augmentation of thymic-dependent and thymic-independent Treg production. Correspondingly, there is a significant reduction in the number of T helper 1 and T helper 17 cells in GVHD target organs, demonstrating that blockade of IL-6 signaling decreases the ratio of proinflammatory T cells to Tregs. These studies demonstrate that antibody blockade of the IL-6R serves to recalibrate the effector and regulatory arms of the immune system and represents a novel, potentially clinically translatable, strategy for the attenuation of GVHD.

Treatment of Epstein Barr virus-induced haemophagocytic lymphohistiocytosis with rituximab-containing chemo-immunotherapeutic regimens

Haemophagocytic lymphohistiocytosis (HLH) is a life threatening complication of Epstein–Barr virus (EBV) infection. The anti‐CD20 antibody rituximab depletes B cells, leading to improved outcomes for patients with EBV‐associated B‐lymphoproliferative disorders. To gather data on the use of rituximab in EBV‐HLH, we performed a retrospective investigation involving 42 EBV‐HLH patients who had received treatment with rituximab‐containing regimens. On average, patients received 3 rituximab infusions (range 1–10) at a median dose of 375 mg/m2. In all patients, rituximab was administered with other HLH‐directed medications, including steroids, etoposide and/or ciclosporin. Rituximab‐containing regimens appeared well tolerated and improved clinical status in 43% of patients. Examination of laboratory data obtained prior to and within 2–4 weeks after the first rituximab dose revealed significant reductions in EBV load (median load pre‐rituximab: 114 200 copies/ml, median post‐rituximab: 225 copies/ml, P = 0·0001) and serum ferritin levels (median ferritin pre‐rituximab: 4260 μg/l, median post‐rituximab: 1149 μg/l, P = 0·001). Thus, when combined with conventional HLH‐directed therapies, rituximab improves symptoms, reduces viral load and diminishes inflammation. These data support the incorporation of rituximab into future prospective clinical trials for patients with EBV‐HLH.

X-linked lymphoproliferative syndrome: a genetic condition typified by the triad of infection, immunodeficiency and lymphoma.

X‐linked lymphoproliferative disease (XLP) is an inherited immunodeficiency characterized by the clinical triad of increased susceptibility to primary Epstein–Barr virus (EBV) infection, dysgammaglobulinaemia and lymphoma. Most cases are caused by germline mutations in the SH2D1A gene, which encodes the adaptor molecule Signalling Lymphocytic Activation Molecule (SLAM)‐associated protein (SAP). Recently, a subset of patients with an XLP‐like phenotype was found to carry mutations in XIAP, the gene encoding the X‐linked inhibitor of apoptosis protein (XIAP). Studies of XLP patients and Sap−/− mice reveal that loss of SAP expression impairs immune cell activities, such as natural killer and CD8+ T cell cytotoxicity, T cell cytokine production, activation‐induced cell death, germinal centre formation and natural killer T cell development. Efforts to dissect the diverse roles of SAP and XIAP are enhancing our understanding of immune cell biology and defining how genetic defects in these molecules predispose to EBV‐specific as well as more general cellular and humoral immune dysfunction. These studies are also highlighting critical signalling pathways that might be amenable to pharmacological targeting to improve the treatment of XLP and other disorders associated with impaired antiviral and antitumour immunity.

Interleukin-23 secretion by donor antigen-presenting cells is critical for organ-specific pathology in graft-versus-host disease

Damage to the gastrointestinal tract during graft-versus-host disease (GVHD) from the conditioning regimen in conjunction with alloreactive donor T cells plays a pivotal role in the pathogenesis of this disease. In this study, we have identified secretion of interleukin-23 (IL-23) by donor antigen-presenting cells (APCs) as a critical event in the induction of GVHD of the colon linking conditioning regimen-induced mucosal injury and lipopolysaccharide (LPS) translocation to subsequent proinflammatory cytokine production and GVHD-associated pathologic damage. In the absence of donor APC-derived IL-23 secretion, there is a selective and profound reduction in pathologic damage as well as a marked reduction in LPS and proinflammatory cytokine production in the colon microenvironment. The downstream proinflammatory effects of IL-23 are dependent upon donor-derived secretion of interferon-gamma (IFN-gamma), but are independent of donor IL-17 production. These findings define a novel organ-specific role for IL-23 in the pathophysiology of GVHD and demonstrate that IL-23 can direct tissue-specific pathology within the context of a systemic inflammatory disorder. Furthermore, these studies also identify IL-23 as a potential therapeutic target for the prevention of this life-threatening disorder.

Transcriptional control of invariant NKT cell development

Summary:  Invariant natural killer T (iNKT) cells comprise a rare lymphocyte sublineage with phenotypic and functional properties similar to T and NK cells. Akin to conventional αβ T cells, their development occurs primarily in the thymus, where they originate from CD4+ CD8+ double positive (DP) progenitors. However, the selection of iNKT cells is unique in that it is mediated by homotypic interactions of DP cells and recognition of glycolipid antigen‐CD1d complexes. Additionally, iNKT cells acquire an activated innate‐like phenotype during development that allows them to release cytokines rapidly following antigen exposure. Given their hybrid features, it is not surprising that the developmental program of iNKT cells partially overlaps with that of T and NK cells. Several recent reports have provided new and exciting insights into the developmental mechanisms that direct natural killer T (NKT) cell lineage commitment and maturation. In this review, we provide a discussion of the NKT cell developmental program with an emphasis on the signaling mechanisms and transcription factors that influence the ontogeny of this lineage. Continued investigations into the complex interplay of these transcription factors and their relationship with other extracellular and intracellular signaling molecules will undoubtedly provide important clues into the biology of this unusual T‐cell lineage.

Janus kinase inhibition lessens inflammation and ameliorates disease in murine models of hemophagocytic lymphohistiocytosis.

Hemophagocytic lymphohistiocytosis (HLH) comprises an emerging spectrum of inherited and noninherited disorders of the immune system characterized by the excessive production of cytokines, including interferon-γ and interleukins 2, 6, and 10 (IL-2, IL-6, and IL-10). The Janus kinases (JAKs) transduce signals initiated following engagement of specific receptors that bind a broad array of cytokines, including those overproduced in HLH. Based on the central role for cytokines in the pathogenesis of HLH, we sought to examine whether the inhibition of JAK function might lessen inflammation in murine models of the disease. Toward this end, we examined the effects of JAK inhibition using a model of primary (inherited) HLH in which perforin-deficient (Prf1(-∕-)) mice are infected with lymphocytic choriomeningitis virus (LCMV) and secondary (noninherited) HLH in which C57BL/6 mice receive repeated injections of CpG DNA. In both models, treatment with the JAK1/2 inhibitor ruxolitinib significantly lessened the clinical and laboratory manifestations of HLH, including weight loss, organomegaly, anemia, thrombocytopenia, hypercytokinemia, and tissue inflammation. Importantly, ruxolitinib treatment also significantly improved the survival of LCMV-infectedPrf1(-∕-)mice. Mechanistic studies revealed that in vivo exposure to ruxolitinib inhibited signal transducer and activation of transcription 1-dependent gene expression, limited CD8(+)T-cell expansion, and greatly reduced proinflammatory cytokine production, without effecting degranulation and cytotoxic function. Collectively, these findings highlight the JAKs as novel, druggable targets for mitigating the cytokine-driven hyperinflammation that occurs in HLH. These observations also support the incorporation of JAK inhibitors such as ruxolitinib into future clinical trials for patients with these life-threatening disorders.

Blockade of interleukin-23 signaling results in targeted protection of the colon and allows for separation of graft-versus-host and graft-versus-leukemia responses.

Allogeneic stem cell transplantation is the most potent form of effective adoptive immunotherapy. The graft-versus-leukemia (GVL) effect mediated by the allogeneic graft, however, is typically coexpressed with graft-versus-host disease (GVHD), which is the major complication of allogeneic stem cell transplantation. In this study, we used genetic and antibody-based strategies to examine the effect that blockade of interleukin 23 (IL-23) signaling had on GVH and GVL reactivity in murine transplantation recipients. These studies demonstrate that the selective protection of the colon that occurs as a consequence of inhibition of IL-23 signaling reduces GVHD without loss of the GVL effect. The separation of GVH and GVL reactivity was noted in both acute and chronic hematologic malignancy models, indicating that this approach was not restricted by the kinetic profile of the underlying leukemia. Furthermore, a potent GVL response could be mounted in the colon under conditions where tumor cells migrated to this site, indicating that this organ did not serve as a sanctuary site for subsequent systemic relapse in GVHD-protected animals. These studies demonstrate that blockade of IL-23 signaling is an effective strategy for separating GVH and GVL responses and identify IL-23 as a therapeutic target for the regulation of alloresponses in humans.

iNKT cell cytotoxic responses control T-lymphoma growth in vitro and in vivo .

Bassiri and colleagues demonstrate that the cytotoxic responses of invariant natural killer T (iNKT) cells are sufficient to limit the growth of T lymphomas, highlighting the potential utility of iNKT cells in the immunotherapy of CD1d+ cancers. Invariant natural killer T (iNKT) cells comprise a lineage of CD1d-restricted glycolipid-reactive T lymphocytes with important roles in host immunity to cancer. iNKT cells indirectly participate in antitumor responses by inducing dendritic cell maturation and producing cytokines that promote tumor clearance by CD8+ T and NK cells. Although iNKT cells thereby act as potent cellular adjuvants, it is less clear whether they directly control the growth of tumors. To gain insights into the direct contribution of iNKT cells to tumor immune surveillance, we developed in vitro and in vivo systems to selectively examine the antitumor activity of iNKT cells in the absence of other cytolytic effectors. Using the EL4 T-lymphoma cell line as a model, we found that iNKT cells exert robust and specific lysis of tumor cells in vitro in a manner that is differentially induced by iNKT cell agonists of varying T-cell receptor (TCR) affinities, such as OCH, α-galactosyl ceramide, and PBS44. In vitro blockade of CD1d-mediated lipid antigen presentation, disruption of TCR signaling, or loss of perforin expression significantly reduce iNKT cell killing. Consistent with these findings, iNKT cell reconstitution of T, B, and NK cell–deficient mice slows EL4 growth in vivo via TCR-CD1d and perforin-dependent mechanisms. Together, these observations establish that iNKT cells are sufficient to control the growth of T lymphoma in vitro and in vivo. They also suggest that the induction of iNKT cell cytotoxic responses in situ might serve as a more effective strategy to prevent and/or treat CD1d+ cancers, such as T lymphoma. Cancer Immunol Res; 2(1); 59–69. ©2013 AACR.

Invariant NKT cells: Killers and conspirators against cancer

Although invariant natural killer T (iNKT) cells influence antitumor responses indirectly by secreting cytokines and promoting the cytolytic functions of T and NK cells, we find that iNKT cells mediate direct tumoricidal activity in vitro and significantly inhibit tumor growth in vivo, even in the absence of other cytotoxic lymphocytes.

Interleukin 17 Is Not Required for Autoimmune-Mediated Pathologic Damage during Chronic Graft-versus-Host Disease

The transition from acute to chronic graft-versus-host disease (aGVHD, cGVHD) is characterized by the progressive loss of self-tolerance and the development of autoimmune manifestations. Interleukin 17 (IL-17) is a proinflammatory cytokine that has been shown to play a prominent role in autoimmune disorders in the nontransplant setting, but the extent to which IL-17 is necessary for the autoimmunity that occurs as a consequence of GVHD is not known. In this study, we demonstrate using a combination of antibody-based and genetic approaches that IL-17 is not required for the loss of self-tolerance and resulting CD4(+)T cell-dependent pathologic damage that occurs during the evolution from aGVHD to cGVHD. Rather, T(H)1 cells and other proinflammatory cytokines are fully competent to promote autoimmune-mediated tissue damage. Thus, the selective targeting of IL-17 may not be a viable clinical strategy for preventing the autoimmune manifestations that develop during cGVHD.