Faculty Bibliography

Patients with cutaneous T-cell lymphoma (CTCL) are frequently colonized with Staphylococcus aureus (SA). Eradication of SA is, importantly, associated with significant clinical improvement suggesting that SA promotes the disease activity but the underlying mechanisms remain poorly characterized. Here we show that SA isolates from involved skin express staphylococcal enterotoxins (SEs) which induce cross-talk between malignant and benign T cells leading to Stat3-mediated IL-10 production by the malignant T cells. The SEs did not stimulate the malignant T cells directly. Instead, SEs triggered a cascade of events involving cell-cell and asymmetric cytokine interactions between malignant and benign T cells, which stimulated the malignant T cells to express high levels of IL-10. Much evidence supports that malignant activation of the Stat3/IL-10 axis plays a key role in driving the immune dysregulation and severe immunodeficiency that characteristically develops in CTCL patients. The present findings thereby establish a novel link between SEs and immune dysregulation in CTCL strengthening the rationale for antibiotic treatment of colonized patients with severe or progressive disease.

Antibody repertoires are known to be shaped by selection for antigen binding. Unexpectedly, we now show that selection also acts on a non-antigen-binding antibody region: the heavy-chain variable (VH)-encoded "elbow" between variable and constant domains. By sequencing 2.8 million recombined heavy-chain genes from immature and mature B-cell subsets in mice, we demonstrate a striking gradient in VH gene use as pre-B cells mature into follicular and then into marginal zone B cells. Cells whose antibodies use VH genes that encode a more flexible elbow are more likely to mature. This effect is distinct from, and exceeds in magnitude, previously described maturation-associated changes in heavy-chain complementarity determining region 3, a key antigen-binding region, which arise from junctional diversity rather than differential VH gene use. Thus, deep sequencing reveals a previously unidentified mode of B-cell selection.

The IL-23 pathway is genetically linked to autoimmune disease in humans and is required for pathogenic Th17 cell function in mice. However, because IL-23R-expressing mature Th17 cells are rare and poorly defined in mice at steady-state, little is known about IL-23 signaling. In this study, we show that the endogenous CCR6(+) memory T cell compartment present in peripheral lymphoid organs of unmanipulated mice expresses Il23r ex vivo, displays marked proinflammatory responses to IL-23 stimulation in vitro, and is capable of transferring experimental autoimmune encephalomyelitis. The prolyl-tRNA synthetase inhibitor halofuginone blocks IL-23-induced Stat3 phosphorylation and IL-23-dependent proinflammatory cytokine expression in endogenous CCR6(+) Th17 cells via activation of the amino acid starvation response (AAR) pathway. In vivo, halofuginone shows therapeutic efficacy in experimental autoimmune encephalomyelitis, reducing both established disease progression and local Th17 cell effector function within the CNS. Mechanistically, AAR activation impairs Stat3 responses downstream of multiple cytokine receptors via selective, posttranscriptional suppression of Stat3 protein levels. Thus, our study reveals latent pathogenic functions of endogenous Th17 cells that are regulated by both IL-23 and AAR pathways and identifies a novel regulatory pathway targeting Stat3 that may underlie selective immune regulation by the AAR.

Phosphorylation of signal transducer and activator of transcription 3 (STAT3) is essential for cell survival, proliferation and differentiation. STAT3 phosphorylation results from signaling by cytokines and growth factors, and constitutive STAT3 activity is characteristic of a number of human malignancies, including Cutaneous T Cell Lymphoma (CTCL). Furthermore, we now know that STAT3 is also required for the initiation and maintenance of the Th17 differentiation program. Th17 cells are a subset of CD4 T helper cells that have been implicated in chronic inflammatory conditions like rheumatoid arthritis and psoriasis. Mycosis fungoides (MF) and the leukemic variant of this disease, Sezary syndrome (SS), are the most frequently encountered forms of CTCL and in both of these diseases, the cell of origin - as far as the type of Teffector cell involved, has not been defined. Recent results from our laboratory and that of our colleagues have lead us to believe that Th17 cells may either be the cells of origin in CTCL or may act as critical mediators of chronic inflammation that creates a favorable environment for tumor growth in the context of this malignancy. In an effort to elucidate the role of STAT3 as a transforming factor in T cell malignancies, we generated a mouse model wherein T cell specific expression of a hyper-active STAT3 mutant protein (STAT3C) leads to the development of a lymphoproliferative disease that is highly reminiscent of CTCL. We are now taking advantage of this unique mouse model, patient biospecimens and carefully characterized CTCL cell lines to dissect the role of STAT3 signaling cascade in the malignant transformation and maintenance of CTCL. Most recently, our attention has been focused on understanding the mechanism of action of epigenetic therapy in the form of histone deacetylase inhibitors (HDACi), which is highly effective in the treatment of CTCL. We hypothesize that HDAC inhibitors affect the STAT3 mediated Th17 differentiation and thus have clinical efficacy in !

Antigen receptor editing-a process of secondary rearrangements of antigen receptor genes in autoreactive lymphocytes-is a well-established tolerance mechanism in B cells, whereas its role in T cells remains controversial. Here, we investigated this issue using a novel Tcra knock-in locus, which ensured appropriate timing of TCRalpha expression and allowed secondary rearrangements. Under these conditions the only response to self-antigen that could be unambiguously identified was negative selection of CD4/CD8 double positive thymocytes. No evidence could be obtained for antigen-induced TCR editing, whereas replacement of the transgenic TCRalpha chain by ongoing gene rearrangement occurred in some cells irrespective of the presence or absence of self-antigen.

Th17 cells are a proinflammatory subset of effector T cells that have been implicated in the pathogenesis of asthma. Their production of the cytokine IL-17 is known to induce local recruitment of neutrophils, but the direct impact of IL-17 on the lung epithelium is poorly understood. In this study, we describe a novel mouse model of spontaneous IL-17-driven lung inflammation that exhibits many similarities to asthma in humans. We have found that STAT3 hyperactivity in T lymphocytes causes an expansion of Th17 cells, which home preferentially to the lungs. IL-17 secretion then leads to neutrophil infiltration and lung epithelial changes, in turn leading to a chronic inflammatory state with increased mucus production and decreased lung function. We used this model to investigate the effects of IL-17 activity on airway epithelium and identified CXCL5 and MIP-2 as important factors in neutrophil recruitment. The neutralization of IL-17 greatly reduces pulmonary neutrophilia, underscoring a key role for IL-17 in promoting chronic airway inflammation. These findings emphasize the role of IL-17 in mediating neutrophil-driven pulmonary inflammation and highlight a new mouse model that may be used for the development of novel therapies targeting Th17 cells in asthma and other chronic pulmonary diseases.

In patients with cutaneous T-cell lymphoma (CTCL) bacterial infections constitute a major clinical problem caused by compromised skin barrier and a progressive immunodeficiency. Indeed, the majority of patients with advanced disease die from infections with bacteria, e.g., Staphylococcus aureus. Bacterial toxins such as staphylococcal enterotoxins (SE) have long been suspected to be involved in the pathogenesis in CTCL. Here, we review links between bacterial infections and CTCL with focus on earlier studies addressing a direct role of SE on malignant T cells and recent data indicating novel indirect mechanisms involving SE- and cytokine-driven cross-talk between malignant- and non-malignant T cells.

Inappropriately regulated expression of interleukin (IL)-17A is associated with the development of inflammatory diseases and cancer. However, little is known about the role of other IL-17 family members in carcinogenesis. Here, we show that a set of malignant T-cell lines established from patients with cutaneous T-cell lymphoma (CTCL) spontaneously secrete IL-17F and that inhibitors of Janus kinases and Signal transducer and activator of transcription 3 are able to block that secretion. Other malignant T-cell lines produce IL-17A but not IL-17F. Upon activation, however, some of the malignant T-cell lines are able to coexpress IL-17A and IL-17F, leading to formation of IL-17A/F heterodimers. Clinically, we demonstrate that IL-17F messenger RNA expression is significantly increased in CTCL skin lesions compared with healthy donors and patients with chronic dermatitis. IL-17A expression is also increased and a significant number of patients express high levels of both IL-17A and IL-17F. Concomitantly, we observed that the expression of the IL-17 receptor is significantly increased in CTCL skin lesions compared with control subjects. Importantly, analysis of a historic cohort of 60 CTCL patients indicates that IL-17F expression is associated with progressive disease. These findings implicate IL-17F in the pathogenesis of CTCL and suggest that IL-17 cytokines and their receptors may serve as therapeutic targets.

Mice lacking the Jak tyrosine kinase member Tyk2 become progressively obese due to aberrant development of Myf5+ brown adipose tissue (BAT). Tyk2 RNA levels in BAT and skeletal muscle, which shares a common progenitor with BAT, are dramatically decreased in mice placed on a high-fat diet and in obese humans. Expression of Tyk2 or the constitutively active form of the transcription factor Stat3 (CAStat3) restores differentiation in Tyk2(-/-) brown preadipocytes. Furthermore, Tyk2(-/-) mice expressing CAStat3 transgene in BAT also show improved BAT development, normal levels of insulin, and significantly lower body weights. Stat3 binds to PRDM16, a master regulator of BAT differentiation, and enhances the stability of PRDM16 protein. These results define Tyk2 and Stat3 as critical determinants of brown fat lineage and suggest that altered levels of Tyk2 are associated with obesity in both rodents and humans.