Improved pruritus correlates with lower levels of IL-31 in CTCL patients under different therapeutic modalities.

Pruritus is one of the cardinal symptoms found in patients with leukemic cutaneous T cell lymphoma (CTCL). The nature of the pruritus experienced by CTCL patients is complex, involving different pathways and cell mediators, thus making it poorly responsive to conventional anti-itch therapies. Recent reports highlight the role of interleukin 31 (IL-31) as a novel cytokine involved in the pathogenesis of pruritus in atopic dermatitis and CTCL. Here we provide both in vivo and in vitro evidence suggesting that histone deacetylase (HDAC) inhibitors may mitigate itch through lowering of levels of IL-31-expressing T cells. Furthermore, we demonstrate that chemokine receptor type-4 (CCR4)-bearing T cells are a main source of IL-31 in CTCL, and that neutralizing the IL-31 pathway through targeting of the CCR4-expressing T cells may represent a promising therapeutic strategy for symptomatic relief in CTCL.

Inhibition of cell-mediated immunity by the histone deacetylase inhibitor vorinostat: implications for therapy of cutaneous T-cell lymphoma.

Several histone deacetylase inhibitors (HDACi), including vorinostat, have been approved for the therapy of cutaneous T-cell lymphoma (CTCL). Emerging data suggest that HDACi may exert immune suppressive effects which would be disadvantageous for therapy of CTCL. We describe a patient with Sezary syndrome who was monitored for drug-induced immunosuppression while undergoing treatment with vorinostat. Analysis of the patient's natural killer cell function before and after initiation of treatment confirmed inhibition of this important cell-mediated immune function. In addition, the in vitro effects of vorinostat on the immunity of healthy volunteers confirmed that this class of drug can profoundly suppress multiple arms of the cellular immune response. These findings raise concerns of increased susceptibility to infection in this high-risk population.

Immunopathogenesis and therapy of cutaneous T cell lymphoma.

Cutaneous T cell lymphomas (CTCLs) are a heterogenous group of lymphoproliferative disorders caused by clonally derived, skin-invasive T cells. Mycosis fungoides (MF) and Sezary syndrome (SS) are the most common types of CTCLs and are characterized by malignant CD4(+)/CLA(+)/CCR4(+) T cells that also lack the usual T cell surface markers CD7 and/or CD26. As MF/SS advances, the clonal dominance of the malignant cells results in the expression of predominantly Th2 cytokines, progressive immune dysregulation in patients, and further tumor cell growth. This review summarizes recent insights into the pathogenesis and immunobiology of MF/SS and how these have shaped current therapeutic approaches, in particular the growing emphasis on enhancement of host antitumor immune responses as the key to successful therapy.

Synthetic imidazoquinolines potently and broadly activate the cellular immune response of patients with cutaneous T-cell lymphoma: synergy with interferon-gamma enhances production of interleukin-12.

Advanced cutaneous T-cell lymphoma (CTCL) is typically associated with a variety of profound defects of cellular immunity, including depressed dendritic cell numbers and function. Therefore, we investigated the ability of synthetic imidazoquinolines, which are agonists for Toll-like receptors (TLRs) 7 and 8, to enhance in vitro the cell-mediated immunity of patients with leukemic CTCL and Sézary syndrome. Patients' peripheral blood mononuclear cells (PBMCs) stimulated with the TLR7 agonist 3M-001 produced high levels of interferon (IFN)-alpha, and the TLR8 agonist 3M-002 potently induced predominantly interleukin (IL)-12 and IFN-gamma. Marked upregulation of CD69 and CD25 on natural killer (NK) cells and T cells from patients and an increase in NK cytolytic activity was also observed. We further demonstrate that priming of patients' PBMCs with IFN-gamma has the ability to synergistically enhance the production of IL-12 induced by a synthetic agonist for TLR8. The underlying mechanisms of increased IL-12 production in response to priming with IFN appears to involve an increase in IL-12 p35 and IL-12 p40 gene transcription and a decrease in IL-10 levels upon stimulation with the TLR8 agonist. Our data demonstrate the ability of imidazoquinolines to potently stimulate cellular immune responses of patients with CTCL and emphasizes the benefit of using a combination of biologic modifiers to further enhance their immune responses.

CD164 identifies CD4+ T cells highly expressing genes associated with malignancy in Sézary syndrome: the Sézary signature genes, FCRL3, Tox, and miR-214.

Sézary syndrome (SS), a leukemic variant of cutaneous T-cell lymphoma (CTCL), is associated with a significantly shorter life expectancy compared to skin-restricted mycosis fungoides. Early diagnosis of SS is, therefore, key to achieving enhanced therapeutic responses. However, the lack of a biomarker(s) highly specific for malignant CD4+ T cells in SS patients has been a serious obstacle in making an early diagnosis. We recently demonstrated the high expression of CD164 on CD4+ T cells from Sézary syndrome patients with a wide range of circulating tumor burdens. To further characterize CD164 as a potential biomarker for malignant CD4+ T cells, CD164+ and CD164-CD4+ T cells isolated from patients with high-circulating tumor burden, B2 stage, and medium/low tumor burden, B1-B0 stage, were assessed for the expression of genes reported to differentiate SS from normal controls, and associated with malignancy and poor prognosis. The expression of Sézary signature genes: T plastin, GATA-3, along with FCRL3, Tox, and miR-214, was significantly higher, whereas STAT-4 was lower, in CD164+ compared with CD164-CD4+ T cells. While Tox was highly expressed in both B2 and B1-B0 patients, the expression of Sézary signature genes, FCRL3, and miR-214 was associated predominantly with advanced B2 disease. High expression of CD164 mRNA and protein was also detected in skin from CTCL patients. CD164 was co-expressed with KIR3DL2 on circulating CD4+ T cells from high tumor burden SS patients, further providing strong support for CD164 as a disease relevant surface biomarker.

CD164 and FCRL3 are highly expressed on CD4+CD26- T cells in Sézary syndrome patients.

Sézary syndrome (SS) cells express cell surface molecules also found on normal activated CD4 T cells. In an effort to find a more specific surface marker for malignant SS cells, a microarray analysis of gene expression was performed. Results showed significantly increased levels of mRNA for CD164, a sialomucin found on human CD34+ hematopoietic stem cells, and FCRL3, a molecule present on a subset of human natural T regulatory cells. Both markers were increased in CD4 T cells from SS patients compared with healthy donors (HD). Flow cytometry studies confirmed the increased expression of CD164 and FCRL3 primarily on CD4+CD26- T cells of SS patients. Importantly, a statistically significant correlation was found between an elevated percentage of CD4+CD164+ T cells and an elevated percentage of CD4+CD26- T cells in all tested SS patients but not in patients with mycosis fungoides and atopic dermatitis or HD. FCRL3 expression was significantly increased only in patients with high tumor burden. CD4+CD164+ cells displayed cerebriform morphology and their loss correlated with clinical improvement in treated patients. Our results suggest that CD164 can serve as a marker for diagnosis and for monitoring progression of cutaneous T-cell lymphoma (CTCL)/SS and that FCRL3 expression correlates with a high circulating tumor burden.

IL-21 enhances antitumor responses without stimulating proliferation of malignant T cells of patients with Sézary syndrome.

IL-21, a common gamma-chain cytokine secreted by activated CD4+ T cells, influences both humoral and cell-mediated immune responses through the regulation of T, B, dendritic, and natural killer (NK) cells. Sézary syndrome is an advanced form of cutaneous T-cell lymphoma, a clonally derived malignancy of CD4+ T cells that is characterized by profound defects in host cellular immune function. As a modulator of both innate and adaptive immune responses, IL-21 could play an important role in augmenting cell-mediated immunity in these patients. Normal donor and Sézary syndrome patient peripheral blood mononuclear cells were cultured with IL-21 and tested for CD8+ T- and NK-cell activation, NK-cell cytotoxicity, and tumor cell proliferation and apoptosis. IL-21 resulted in a modest increase in CD8+ T- and NK-cell activation, associated with a marked increase in cytolytic activity against both K562 and malignant CD4+ T-cell targets. Although IL-21 failed to demonstrate pro-apoptotic effects on the malignant CD4+ T cells, it is noteworthy that it had no demonstrable proliferative effects on these cells. Thus, IL-21 may play an important role in enhancing the host immune response of Sézary syndrome patients through the increased cytolytic activity of T and NK cells.

Bexarotene blunts malignant T-cell chemotaxis in Sezary syndrome: reduction of chemokine receptor 4-positive lymphocytes and decreased chemotaxis to thymus and activation-regulated chemokine.

The malignant cells in Sezary syndrome express the skin trafficking molecules' cutaneous lymphocyte associated antigen (CLA) and chemokine receptor 4 (CCR4). High levels of the CCR4 ligand, thymus, and activation-regulated chemokine (TARC), have been reported in the blood and skin of patients. The rexinoid X-receptor specific retinoid, bexarotene, has contributed to the resolution of cutaneous disease among patients. To evaluate the effects of bexarotene on skin trafficking molecule expression and chemotaxis, peripheral blood mononuclear cells from Sezary syndrome patients and healthy controls were treated with bexarotene in vitro. CCR4 and CLA expression levels and chemotaxis in response to TARC (6.25 ng/ml) were evaluated among lymphocytes before and after treatment with bexarotene (10 microM). Flow cytometric analysis was performed to evaluate CD4, CD26, CLA, and CCR4 cell surface expression. Transwell migration assays were performed to evaluate chemotaxis to TARC. Prior to treatment, malignant cells exhibited higher CCR4 expression (45-90%) and greater than four times more chemotaxis to TARC compared with healthy controls. After treatment with bexarotene for 36-96 hr, a 28% reduction in CCR4 expression was noted (P < 0.05) among the malignant population with an associated 9% decrease in chemotaxis to TARC (P < 0.05). Our results show that bexarotene may inhibit malignant cell trafficking to the skin through an ability to suppress CCR4 expression among Sezary syndrome lymphocytes.

Enhancement of the host immune responses in cutaneous T-cell lymphoma by CpG oligodeoxynucleotides and IL-15.

Patients with advanced cutaneous T-cell lymphoma (CTCL) exhibit profound defects in cell-mediated immunity. Host immune functions appear to play an integral role in mediating disease-controlling responses in CTCL, therefore we investigated the effects of synthetic oligode-oxynucleotides with CpG motifs (CpG ODN), which have been recognized as immune stimulatory by virtue of activation of dendritic cells (DCs) following binding to Toll-like receptor (TLR) 9. Peripheral blood mononuclear cells (PBMCs) from patients with advanced CTCL (erythroderma with circulating malignant T cells) and healthy volunteers were cultured with either CpG-A or CpG-B ODN. Patients' PBMCs exhibited marked induction of interferon-alpha (IFN-alpha) release following culture with CpG-A. Similarly significant activation of NK cells and CD8 T cells occurred as assessed by up-modulation of CD69 expression and by natural killer lytic activity. Nevertheless, the PBMCs of patients exhibited blunted responses to CpG-A compared to healthy volunteers. In such cases, IL-15 was capable of producing levels of NK activation that were superior to CpG-A, while the combined effects of CpG-A plus IL-15 induced maximal activation of NK cells and further enhanced activation of CD8 T cells. These findings have important implications for the potential enhancement of antitumor immunity among patients with advanced CTCL.

The use of cytokines, fusion proteins and antibodies to treat cutaneous T-cell lymphoma.

It has long been known that certain immune augmenting therapeutics, particularly interferon alpha, can exert profound salutary effects on the clinical progress of patients with cutaneous T-cell lymphoma. Emerging evidence that the host immune response may play an important role in the control of this disorder has led to the clinical application of other cytokines including interleukin-12 and interferon gamma. In this review, the authors will summarize current knowledge regarding the use of cytokines, fusion proteins and antibodies for the treatment of cutaneous T-cell lymphoma.