Breakdown of thymic tolerance - an etiologic link between acute and chronic graft-versus-host disease

Allogeneic haematopoietic stem cell transplantation (HSCT) is the standard therapy for many disorders of the haematological system. Its use is limited by the major complication acute graft-versus-host disease (GVHD). Development of acute GVHD predisposes to chronic GVHD whose autoimmune manifestations are integral components of disease. It remains uncertain, however, whether and how autoimmunity is linked to antecedent alloimmunity. A hallmark of murine acute GVHD is the de novo generation of autoreactive T cells that suggests breakdown of thymic tolerance induction. Central tolerance is dependent on the intrathymic expression of a full scope of tissue-restricted self-antigens (TRA), which is a distinct property of mature medullary thymic epithelial cells (mTEChigh). The ectopic expression of TRA in mTEChigh is partly controlled by the autoimmune regulator (Aire). Since the thymus epithelium is a target of donor T-cell alloimmunity, I hypothesised that thymic acute GVHD interfered with the mTEChigh capacity to sustain TRA diversity. I found that reductions in mTEChigh compartment sizes are universal manifestations of thymic acute GVHD in murine models of haploidentical, fully MHC-disparate and MHC-identical allogeneic HSCT. Moreover, acute GVHD weakens the platform for central tolerance induction because individual TRA are purged from the total repertoire secondary to a decline in the Aire+mTEChigh subset. The most substantially reduced TRA are enriched for genes specific for known target tissues of chronic GVHD. I provide direct evidence in a transgenic mouse system using ovalbumin (OVA) as a model neo-TRA that the de novo production of TRA-specific CD4+ T cells during acute GVHD is a consequence of impaired ectopic TRA expression. OVA-specific CD4+ T cells are present in the periphery in mice with acute GVHD. Peritransplant administration of an epithelial cytoprotective agent, fibroblast growth factor-7, maintains a stable pool of Aire+mTEChigh, which is due to enhanced proliferation of cells within the total mTEC compartment. In parallel, Fgf7 improves the TRA transcriptome despite acute GVHD. Taken together, these data indicate the presence of an etiologic link between acute GVHD and autoimmunity during subsequent chronic GVHD. The present results also suggest that approaches for epithelial cytoprotection may prove to prevent the emergence of thymus-dependent autoreactive T cells

Graft-versus-host disease reduces lymph node display of tissue-restricted self-antigens and promotes autoimmunity

Acute graft-versus-host disease (GVHD) is initially triggered by alloreactive T cells, which damage peripheral tissues and lymphoid organs. Subsequent transition to chronic GVHD involves the emergence of autoimmunity although the underlying mechanisms driving this process are unclear. Here, we tested the hypothesis that acute GVHD blocks peripheral tolerance of autoreactive T cells by impairing lymph node (LN) display of peripheral tissue-restricted antigens (PTA). At the initiation of GVHD, LN fibroblastic reticular cells (FRC) rapidly reduced expression of genes regulated by DEAF1, an Autoimmune Regulator-like transcription factor required for intra-nodal expression of PTA. Subsequently, GVHD led to the selective elimination of the FRC population, and blocked the repair pathways required for its regeneration. We used a transgenic mouse model to show that the loss of presentation of an intestinal PTA by FRC during GVHD resulted in the activation of auto-aggressive T cells and gut injury. Finally, we show that FRC normally expressed a unique PTA gene signature that was highly enriched for genes expressed in the target organs affected by chronic GVHD. In conclusion, acute GVHD damages and prevents repair of the FRC network, thus disabling an essential platform for purging auto-reactive T cells from the repertoire

Mocravimod, a Selective Sphingosine-1-Phosphate Receptor Modulator, in Allogeneic Hematopoietic Stem Cell Transplantation for Malignancy

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) remains the sole curative option for patients with acute myelogenous leukemia. Outcomes are limited by leukemia relapse, graft-versus-host disease (GVHD), and abnormal immune reconstitution. Mocravimod (KRP203) is an oral sphingosine-1-phosphate receptor (S1PR) modulator that blocks the signal required by T cells to egress from lymph nodes and other lymphoid organs. Mocravimod retains T cell effector function, a main differentiator to immunosuppressants. In preclinical models, mocravimod improves survival by maintaining graft-versus-leukemia (GVL) activity while reducing GVHD. In patients undergoing allo-HSCT for hematological malignancies, mocravimod is postulated to prevent GVHD by redistributing allogeneic donor T cells to lymphoid tissues while allowing a sufficient GVL effect in the lymphoid, where malignant cells usually reside. The primary objective of this study was to assess the safety and tolerability of mocravimod in patients undergoing allo-HSCT for hematologic malignancies. Secondary objectives were to determine the pharmacokinetic profiles of mocravimod and its active metabolite mocravimod-phosphate in this patient group, as well as to assess GVHD-free, relapse free survival at 6 months after the last treatment. In this 2-part, single- and 2-arm randomized, open-label trial, we evaluated the safety, tolerability, and pharmacokinetics of mocravimod in allo-HSCT recipients (ClinicalTrials.gov identifier NCT01830010). Patients received either 1 mg or 3 mg mocravimod per day on top of standard of care GVHD prophylaxis with either cyclosporine A/methotrexate or tacrolimus/methotrexate. We found that mocravimod can be safely added to standard treatment regimens in patients with hematologic malignancies requiring allo-HSCT. Mocravimod resulted in a significant reduction of circulating lymphocyte numbers and had no negative impact on engraftment and transplantation outcomes. Our results indicate that mocravimod is safe and support a larger study to investigate its efficacy in a homogeneous acute myelogenous leukemia patient population undergoing allo-HSCT

Fibroblastic Reticular Cells Control Conduit Matrix Deposition during Lymph Node Expansion.

Lymph nodes (LNs) act as filters, constantly sampling peripheral cues. This is facilitated by the conduit network, a tubular structure of aligned extracellular matrix (ECM) fibrils ensheathed by fibroblastic reticular cells (FRCs). LNs undergo rapid 3- to 5-fold expansion during adaptive immune responses, but these ECM-rich structures are not permanently damaged. Whether conduit flow or filtering function is affected during LN expansion is unknown. Here, we show that conduits are partially disrupted during acute LN expansion, but FRC-FRC contacts remain connected. We reveal that polarized FRCs deposit ECM basolaterally using LL5-β and that ECM production is regulated at transcriptional and secretory levels by the C-type lectin CLEC-2, expressed by dendritic cells. Inflamed LNs maintain conduit size exclusion, and flow is disrupted but persists, indicating the robustness of this structure despite rapid tissue expansion. We show how dynamic communication between peripheral tissues and LNs provides a mechanism to prevent inflammation-induced fibrosis in lymphoid tissue

Tbet promotes NK cell egress from the bone marrow and CXCR6 expression in immature NK cells

Tbet-deficient mice have reduced NK cells in blood and spleen, but increased NK cells in bone marrow and lymph nodes, a phenotype that is thought to be due to defective migration. Here, we revisit the role of Tbet in NK cell bone marrow egress. We definitively show that the accumulation of NK cells in the bone marrow of Tbet-deficient (Tbx21-/-) animals occurs because of a cell-intrinsic migration defect. We identify a profile of gene expression, co-ordinated by Tbet, which affects the localisation of NK cells in the bone marrow. Tbet promotes Cxcr6 expression and immature NK cells accumulate in the bone marrow of CXCR6-deficient mice. This suggests that CXCR6 is among the mediators of migration, controlled by Tbet, that co-ordinate NK cell bone marrow egress.</jats:p

Tbet promotes CXCR6 expression in immature natural killer cells and natural killer cell egress from the bone marrow

International audienceTbet-deficient mice have reduced natural killer (NK) cells in blood and spleen, but increased NK cells in bone marrow and lymph nodes, a phenotype that is thought to be the result of defective migration. Here, we revisit the role of Tbet in NK cell bone marrow egress. We definitively show that the accumulation of NK cells in the bone marrow of Tbet-deficient Tbx21-/- animals occurs because of a migration defect and identify a module of genes, co-ordinated by Tbet, which affects the localization of NK cells in the bone marrow. Cxcr6 is approximately 125-fold underexpressed in Tbx21-/- , compared with wild-type, immature NK cells. Immature NK cells accumulate in the bone marrow of CXCR6-deficient mice, and CXCR6-deficient progenitors are less able to reconstitute the peripheral NK cell compartment than their wild-type counterparts. However, the CXCR6 phenotype is largely confined to immature NK cells, whereas the Tbet phenotype is present in both immature and mature NK cells, suggesting that genes identified as being more differentially expressed in mature NK cells, such as S1pr5, Cx3cr1, Sell and Cd69, may be the major drivers of the phenotype

Tbet promotes NK cell bone marrow egress via CXCR6 expression

Tbet-deficient mice have reduced NK cells in blood and spleen, but increased NK cells in bone marrow and lymph nodes, a phenotype that is thought to be due to a defect in S1PR5-mediated migration. Here, we revisit the role of Tbet in NK cell bone marrow egress. We definitively show that the accumulation of NK cells in the bone marrow of Tbet-deficient (Tbx21 −/− ) animals occurs because of a cell-intrinsic migration defect. We identify a profile of gene expression, co-ordinated by Tbet, which affects the localisation of NK cells in the bone marrow. The most underexpressed gene in the absence of Tbet was Cxcr6, and we confirmed that CXCR6 protein was also not expressed in Tbet-deficient NK cells. Cxcr6-expressing ILC progenitors and immature NK cells accumulate in the bone marrow of CXCR6-deficient mice. This suggests that CXCR6 is among the mediators of migration, controlled by Tbet, that work together to coordinate NK cell bone marrow egress. Understanding NK cell bone marrow egress will become increasingly important as we move into an era in which NK cell immunotherapies are being developed

Impaired thymic expression of tissue-restricted antigens licenses the de novo generation of autoreactive CD4+ T cells in acute GVHD

During acute graft-versus-host disease (aGVHD) in mice, autoreactive T cells can be generated de novo in the host thymus implying an impairment in self-tolerance induction. As a possible mechanism, we have previously reported that mature medullary thymic epithelial cells (mTEC(high)) expressing the autoimmune regulator are targets of donor T-cell alloimmunity during aGVHD. A decline in mTEC(high) cell pool size, which purges individual tissue-restricted peripheral self-antigens (TRA) from the total thymic ectopic TRA repertoire, weakens the platform for central tolerance induction. Here we provide evidence in a transgenic mouse system using ovalbumin (OVA) as a model surrogate TRA that the de novo production of OVA-specific CD4(+) T cells during acute GVHD is a direct consequence of impaired thymic ectopic OVA expression in mTEC(high) cells. Our data, therefore, indicate that a functional compromise of the medullary mTEC(high) compartment may link alloimmunity to the development of autoimmunity during chronic GVHD

The obese liver environment mediates conversion of NK cells to a less cytotoxic ILC1-like phenotype

The liver contains both NK cells and their less cytotoxic relatives, ILC1. Here, we investigate the role of NK cells and ILC1 in the obesity-associated condition, non-alcoholic fatty liver disease (NAFLD). In the livers of mice suffering from NAFLD, NK cells are less able to degranulate, express lower levels of perforin and are less able to kill cancerous target cells than those from healthy animals. This is associated with a decreased ability to kill cancer cells in vivo. On the other hand, we find that perforin-deficient mice suffer from less severe NAFLD, suggesting that this reduction in NK cell cytotoxicity may be protective in the obese liver, albeit at the cost of increased susceptibility to cancer. The decrease in cytotoxicity is associated with a shift towards a transcriptional profile characteristic of ILC1, increased expression of the ILC1-associated proteins CD200R1 and CD49a, and an altered metabolic profile mimicking that of ILC1. We show that the conversion of NK cells to this less cytotoxic phenotype is at least partially mediated by TGFβ, which is expressed at high levels in the obese liver. Finally, we show that reduced cytotoxicity is also a feature of NK cells in the livers of human NAFLD patients