Pathogenesis Of Cutaneous T-cell Lymphoma Research Articles

Jak3, STAT3, and STAT5 inhibit expression of miR-22, a novel tumor suppressor microRNA, in cutaneous T-Cell lymphoma.

Aberrant activation of Janus kinase-3 (Jak3) and its key down-stream effectors, Signal Transducer and Activator of Transcription-3 (STAT3) and STAT5, is a key feature of malignant transformation in cutaneous T-cell lymphoma (CTCL). However, it remains only partially understood how Jak3/STAT activation promotes lymphomagenesis. Recently, non-coding microRNAs (miRNAs) have been implicated in the pathogenesis of this malignancy. Here, we show that (i) malignant T cells display a decreased expression of a tumor suppressor miRNA, miR-22, when compared to non-malignant T cells, (ii) STAT5 binds the promoter of the miR-22 host gene, and (iii) inhibition of Jak3, STAT3, and STAT5 triggers increased expression of pri-miR-22 and miR-22. Curcumin, a nutrient with anti-Jak3 activity and histone deacetylase inhibitors (HDACi) also trigger increased expression of pri-miR-22 and miR-22. Transfection of malignant T cells with recombinant miR-22 inhibits the expression of validated miR-22 targets including NCoA1, a transcriptional co-activator in others cancers, as well as HDAC6, MAX, MYCBP, PTEN, and CDK2, which have all been implicated in CTCL pathogenesis. In conclusion, we provide the first evidence that de-regulated Jak3/STAT3/STAT5 signalling in CTCL cells represses the expression of the gene encoding miR-22, a novel tumor suppressor miRNA.

Evidence of an oncogenic role of aberrant TOX activation in cutaneous T-cell lymphoma

AbstractTOX is a nuclear factor essential for the development of CD4+ T cells in the thymus. It is normally expressed in low amounts in mature CD4+ T cells of the skin and the peripheral blood. We have recently discovered that the transcript levels of TOX were significantly increased in mycosis fungoides, the most common type of cutaneous T-cell lymphoma (CTCL), as compared to normal skin or benign inflammatory dermatoses. However, its involvement in advanced CTCL and its biological effects on CTCL pathogenesis have not been explored. In this study, we demonstrate that TOX expression is also enhanced significantly in primary CD4+CD7− cells from patients with Sézary syndrome, a leukemic variant of CTCL, and that high TOX transcript levels correlate with increased disease-specific mortality. Stable knockdown of TOX in CTCL cells promoted apoptosis and reduced cell cycle progression, leading to less cell viability and colony-forming ability in vitro and to reduced tumor growth in vivo. Furthermore, TOX knockdown significantly increased 2 cyclin-dependent kinase (CDK) inhibitors, CDKN1B and CDKN1C. Lastly, blocking CDKN1B and CDKN1C reversed growth inhibition of TOX knockdown. Collectively, these findings provide strong evidence that aberrant TOX activation is a critical oncogenic event for CTCL.

Evidence of an Oncogenic Role for Aberrant Tox Activation in Cutaneous T Cell Lymphoma

Cutaneous T cell lymphoma (CTCL) consists of two main subtypes: mycosis fungoides (MF), which primarily affects the skin; and Sézary syndrome (SS), which is characterized by the presence of circulating malignant Sézary cells. Although defective apoptosis and cell cycle progression have been implicated in CTCL disease development, the precise molecular mechanism underlying these abnormalities is only partially understood. Recently our group showed that TOX, a transcription factor transiently expressed during thymocyte development, was ectopically expressed in the malignant CD4+ T cells in MF skin lesions. Normally in mature CD4+ T cells, TOX is expressed at very low levels. Not only is TOX highly expressed in the infiltrating CD4+ T cells in MF skin lesions, but its expression levels also inversely correlate with the clinical outcome of CTCL patients. However, its involvement in other types of CTCL remains unknown and its biological effects on CTCL pathogenesis have not been explored. In this study, we sought to examine if aberrant TOX expression occurs in SS (a leukemic variant of CTCL), and whether it contributes to CTCL pathogenesis. We found that TOX expression is also enhanced significantly in primary CD4+CD7- cells from SS patients (n=12), and its enhanced transcript levels correlate with higher disease-specific mortality (P=0.039). To investigate the role of TOX in CTCL, we corrected the ectopic TOX expression by lentiviral-shRNA-mediated TOX gene knock-down in CTCL cell lines (Hut78, HH, and SZ4). In all three cell lines, this resulted in markedly increased apoptosis (measured using Annexin V apoptosis assay, 2-3 fold, P<0.05) and reduced cellular proliferation (assessed by BrdU incorporation assay and colony formation in 3D cultures, 1.5-12 fold, P<0.05). In line with these phenotypes, TOX knock-down led to an increase in cleaved caspases (3, 9) and enhanced expression of multiple cell cycle inhibitors (CDKN1B, CDKN1C). Moreover, to test the effect of TOX suppression on the tumor-inducing ability of CTCL cells in vivo, we injected TOX-suppressed cells or control CTCL cells (1x106 per injection site) subcutaneously into NOD-SCID IL2rγ null (NSG) mice and monitored local tumor formation. Control CTCL cells (Hut78, HH) induced local tumor growth in six out of six mice within 2 weeks of injection, with enlargement progressively to 1.5cm in diameter within 4 weeks of injection. In contrast, TOX silencing effectively reduced the tumor volume induced by HH cells (n=6) and abolished the tumor-forming ability of Hut78 cells (n=6). Since TOX is a DNA-binding nuclear protein, we next sought to explore the potential downstream molecular partners of TOX by performing comparative transcriptome analysis between TOX-suppressed cells and control CTCL cells (Hut78, HH). The expression of several tumor suppressor genes, including SMAD3 and FOXO3, was increased upon TOX knock-down in both Hut78 and HH cell lines. Pathway analyses of the differentially-expressed genes uncovered activation of apoptotic pathways and cell cycle arrest processes upon TOX suppression. These findings were validated using quantitative RT-PCR. In conclusion, our results indicate that ectopic expression of TOX in CTCL cells causes the proliferation/apoptosis balance to shift toward proliferation through suppression of the transcription of multiple tumor suppressors. Therefore abolishing TOX activity may be a promising treatment strategy for CTCL. DisclosuresNo relevant conflicts of interest to declare.

Ectopic Expression of Cancer–Testis Antigens in Cutaneous T-cell Lymphoma Patients

The pathogenesis of cutaneous T-cell lymphoma (CTCL) remains only partially understood. A number of recent studies attempted to identify novel diagnostic markers and future therapeutic targets. One group of antigens, cancer-testis (CT) antigens, normally present solely in testicular germ cells, can be ectopically expressed in a variety of cancers. Currently, only a few studies attempted to investigate the expression of CT antigens in CTCL. In the present work, we test the expression of CT genes in a cohort of patients with CTCL, normal skin samples, skin from benign inflammatory dermatoses, and in patient-derived CTCL cells. We correlate such expression with the p53 status and explore molecular mechanisms behind their ectopic expression in these cells. Our findings demonstrate that SYCP1, SYCP3, REC8, SPO11, and GTSF1 genes are heterogeneously expressed in patients with CTCL and patient-derived cell lines, whereas cTAGE1 (cutaneous T-cell lymphoma-associated antigen 1) was found to be robustly expressed in both. Mutated p53 status did not appear to be a requirement for the ectopic expression of CT antigens. While T-cell stimulation resulted in a significant upregulation of STAT3 and JUNB expression, it did not significantly alter the expression of CT antigens. Treatment of CTCL cells in vitro with vorinostat or romidepsin histone deacetylase inhibitors resulted in a significant dose-dependent upregulation of mRNA but not protein. Further expression analysis demonstrated that SYCP1, cTAGE1, and GTSF1 were expressed in CTCL, but not in normal skin or benign inflammatory dermatoses. A number of CT genes are ectopically expressed in patients with CTCL and can be used as biomarkers or novel targets for immunotherapy.

Role of IL-13 and its receptors in the pathogenesis of cutaneous T-cell lymphomas (TUM7P.961)

Abstract Cutaneous T-cell lymphomas (CTCL) affect primarily skin and are characterized by proliferation of mature CD4+ T-helper cells. Diagnose is difficult in the absence of specific markers for malignant lymphocytes. The pattern of cytokine production in the skin and blood is considered to be of major importance for the pathogenesis of CTCL. Abnormal cytokine expression in CTCL may be responsible for enhanced proliferation of the malignant cells and/or the depression of the anti-tumor immune response. Cytokine IL-13 promotes growth or survival of several tumors as well as suppresses immunosurveillance. Here we show that IL-13 and its receptors IL-13Rα1 and IL-13Rα2 are highly expressed in the clinically involved skin of CTCL patients. We also show that the tumor cells, identified by the expression of CD4 and TOX (thymus high-mobility group box), in the skin and blood of CTCL patients produce IL-13 and express both receptors. This finding is intriguing since CTCL is a T-cell origin tumor yet T cells do not express IL-13 receptors. Thus IL-13 is implicated as a possible autocrine factor for CTCL. Furthermore expression of the decoy IL-13Rα2 by CTCL cells may represent a novel marker of CTCL malignancy. Soluble human IL-13Rα2 and an anti-IL-13 monoclonal antibody are being tested for inhibition of tumor cell proliferation in the peripheral blood of patients. Altogether these results identify potential tumor markers for the early diagnosis of CTCL as well as novel therapeutic targets.

Increased Expression Of IL-15 Promotes Cutaneous T-Cell Lymphomagenesis Via The Upregulation Of Histone Deacetylases: Evidence For Successful Preclinical Targeting

Cutaneous lymphoma is a heterogeneous group of neoplasms of skin-homing malignant lymphocytes. Cutaneous T-cell Lymphoma (CTCL) represents 70-80% of all cutaneous lymphoma and its pathogenesis is largely unknown. Previous studies have shown that interleukin (IL)-15 is a potent stimulant and growth factor for CTCL cells in vitro. In order to investigate the intrinsic levels of IL-15 in CTCL patients, malignant CD4+ T-cells were analyzed for expression of IL-15. Relative quantitation of IL-15 transcript in patient vs normal donor (ND) CD4+ cells showed overexpression of IL-15 in patients (fold increase mean ± SD = 5.36 ± 4.39, N=13, P<0.001). Increase in IL-15 transcript was directly proportional to disease severity in patients i.e. fold increase mean ± SD in IL-15 in Stage I =3.28 ± 1.42, N=3 each, P=0.0047 vs. Stage III patients = 7.42 ± 1.30, N=3 each, P=0.0073. Further, cutaneous lesions in patients stained positive for IL-15 protein in atypical lymphoid cells and Pautrier's microabscess. We next investigated the role of IL-15 in CTCL development using IL-15 transgenic (tg) mice. Within 4-6 weeks of birth, IL-15 tg mice developed extensive patch/plaque skin lesions, progressive alopecia, and severe pruritus. Adult IL-15 tg mice developed extensive involvement with cutaneous lymphoma that was fatal in 100% of the mice (P=0.0003). Antibodies staining revealed that CD4+ skin resident T-cells in IL-15 tg mice were CD3+CD62L-CD44hiCCR4+CLA+. Flow cytometric analysis of single cell suspension of skin showed ∼25-fold increase in CD3+ T-cells in IL-15 tg compared to WT controls (Mean ± SD of absolute number of cells= 3.80 ± 6.97, N=14 vs. 0.15 ± 0.26, N=8 respectively, P<0.001). Lymphoma cells from IL-15 tg mouse skin engrafted and mimicked the primary disease in immune deficient SCID mice upon adoptive transfer. CD4+ T-cells from CTCL patients showed increased histone deacetylases (HDAC) 1, HDAC2 and HDAC6 transcripts over ND CD4+ T-cells and immunoblot analysis of ND CD4+ T-cells exposed to 100ng/ml IL-15 showed upregulation of HDAC1, HDAC2 and HDAC6 ex vivo. IL-15 stimulation of ND CD4+ T-cells resulted in loss of expression of the downstream HDAC1/2 target tumor suppressor, p21 in vitro, and knock down of HDAC6 in IL-15 stimulated ND CD4+ T-cells inhibited their migration in vitro; suggesting that IL-15 mediated upregulation of HDAC6 is critical for T-cell migration. Considering these observations, we used specific novel HDAC inhibitors (HDACi) to target HDAC1/2 (JQ12) and/or HDAC6 (WT161) in IL-15 tg mice to determine if we could prevent CTCL in vivo. IL-15 tg mice were treated with 50mg/kg of either or both the inhibitors, 5 days/week for 4 weeks (n=4 each). While placebo treated IL-15 tg mice progressively developed lesions during the course of treatment, IL-15 tg mice treated with JQ12 and/or WT161 showed no clinical signs of disease. This was further corroborated by histopathology analysis of skin sections from treated mice (Figure 1). Thus, our data suggest that inhibiting HDAC1, HDAC2 and/or HDAC6 pathways inhibits the development of CTCL in IL-15tg mice. In addition to the prevention study, we assessed the ability of a novel pan-HDACi, AR42, to treat active and progressive disease in our model. IL-15 tg mice with established CTCL were randomized to receive either AR42 or placebo feed (n=6 each) for 12 days. The IL-15 tg mice treated with AR42 showed remarkable improvement compared to the placebo mice whose disease progressed. Histopathology analysis of the AR42-treated IL-15 tg mice showed an impressive clearance of the CD3+ and CD4+ atypical lymphocytic infiltrate compared to placebo-treated mice (Figure 2). In summary we provide evidence that IL-15 has a causal role in the pathogenesis of CTCL; that IL-15 tg mice provide a novel model for studying disease pathogenesis and for evaluating potential therapies; that HDACi targeting specific HDACs may be effective in preventing CTCL and a novel pan-HDACi can reverse severe dermatologic disease in this CTCL model. [Display omitted] [Display omitted] Disclosures:No relevant conflicts of interest to declare.

Expression of Human Endogenous Retrovirus-W Including Syncytin-1 in Cutaneous T-Cell Lymphoma

The pathomechanism of mycosis fungoides (MF), the most common type of primary cutaneous T-cell lymphomas (CTCLs) and a malignancy of non-recirculating, skin-resident T-cells, is unknown albeit underlying viral infections have been sought for. Human endogenous retroviruses (HERVs) are ancient retroviral sequences in the human genome and their transcription is often deregulated in cancers. We explored the transcriptional activity of HERV sequences in a total of 34 samples comprising MF and psoriasis skin lesions, as well as corresponding non-malignant skin using a retrovirus-specific microarray and quantitative RT-PCR. To identify active HERV-W loci, we cloned the HERV-W specific RT-PCR products, sequenced the cDNA clones and assigned the sequences to HERV-W loci. Finally, we used immunohistochemistry on MF patient and non-malignant inflammatory skin samples to confirm specific HERV-encoded protein expression. Firstly, a distinct, skin-specific transcription profile consisting of five constitutively active HERV groups was established. Although individual variability was common, HERV-W showed significantly increased transcription in MF lesions compared to clinically intact skin from the same patient. Predominantly transcribed HERV-W loci were found to be located in chromosomes 6q21 and 7q21.2, chromosomal regions typically altered in CTCL. Surprisingly, we also found the expression of 7q21.2/ERVWE1-encoded Syncytin-1 (Env) protein in MF biopsies and expression of Syncytin-1 was seen in malignant lymphocytes, especially in the epidermotropic ones, in 15 of 30 cases studied. Most importantly, no Syncytin-1 expression was detected in inflammatory dermatosis (Lichen ruber planus) with skin-homing, non-malignant T lymphocytes. The expression of ERVWE1 mRNA was further confirmed in 3/7 MF lesions analyzed. Our observations strengthen the association between activated HERVs and cancer. The study offers a new perspective into the pathogenesis of CTCL since we demonstrate that differences in HERV-W transcription levels between lesional MF and non-malignant skin are significant, and that ERVWE1-encoded Syncytin-1 is expressed in MF lymphoma cells.

Elucidating the role of interleukin-17F in cutaneous T-cell lymphoma

AbstractInappropriately 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.

STAT5-mediated expression of oncogenic miR-155 in cutaneous T-cell lymphoma

The pathogenesis of cutaneous T-cell lymphoma (CTCL) remains elusive. Recent discoveries indicate that the oncogenic microRNA miR-155 is overexpressed in affected skin from CTCL patients. Here, we address what drives the expression of miR-155 and investigate its role in the pathogenesis of CTCL. We show that malignant T cells constitutively express high levels of miR-155 and its host gene BIC (B cell integration cluster). Using ChIP-seq, we identify BIC as a target of transcription factor STAT5, which is aberrantly activated in malignant T cells and induced by IL-2/IL-15 in non-malignant T cells. Incubation with JAK inhibitor or siRNA-mediated knockdown of STAT5 decreases BIC/miR-155 expression, whereas IL-2 and IL-15 increase their expression in cell lines and primary cells. In contrast, knockdown of STAT3 has no effect, and BIC is not a transcriptional target of STAT3, indicating that regulation of BIC/miR-155 expression by STAT5 is highly specific. Malignant proliferation is significantly inhibited by an antisense-miR-155 as well as by knockdown of STAT5 and BIC.In conclusion, we provide the first evidence that STAT5 drives expression of oncogenic BIC/miR-155 in cancer. Moreover, our data indicate that the STAT5/BIC/miR-155 pathway promotes proliferation of malignant T cells, and therefore is a putative target for therapy in CTCL.

Elucidating the molecular pathogenesis of cutaneous T-cell lymphoma

Elucidating the molecular pathogenesis of cutaneous T-cell lymphoma

The Role of Molecular Analysis in Cutaneous Lymphomas

The purpose of this review is to summarize the most important molecular techniques for the diagnosis of cutaneous lymphomas. When making a diagnosis, we are looking for the solid clinicopathological correlation. Molecular analysis includes immunophenotyping and clonality analysis, and is important for 2 principal reasons: (1) to confirm the diagnosis in cases where the clinical and/or pathological presentations are nondiagnostic, and (2) to further characterize the nature of the lymphoma. More specifically, we are trying to discern whether the lymphoma is primarily cutaneous or systemic with secondary skin involvement, and we are also attempting to subclassify the tumor. Recently, many techniques have provided a more accurate diagnosis of cutaneous lymphomas and some prognostic implications, including polymerase chain reaction, fluorescence in situ hybridization, and flow cytometry. Fluorescence in situ hybridization is not routinely used in the diagnosis of cutaneous lymphoma, but many studies have shown potential future applications in various areas. Other techniques, such as comparative genomic hybridization, are still confined to the research arena, but have added some insight into the molecular pathogenesis of cutaneous T-cell lymphoma.

Mutations in PLCG1 Is a Frequent Event in Cutaneous T-Cell Lymphomas

AbstractAbstract 300 Background:Cutaneous T-cell lymphoma (CTCL) is a heterogeneous group of diseases characterized by clonal expansion of malignant T-cells in the skin. The two predominant clinical forms of CTCL are mycosis fungoides (MF) and Sezary syndrome (SS). Tumor-stage MF has an unfavorable prognosis with a 10-year survival of approximately 40%. The molecular pathogenesis of CTCL is still basically unknown, although some data suggest that signalling from T-cell receptor (TCR) is a driving force. However, the molecular mechanisms responsible for this activation have not been fully clarified. Methods:Based on the hypothesis that TCR activation may depend, at least in part, on somatic mutations, we have investigated this in a selection of genes belonging to TCR, or related pathways, such as NFkB, JAK/STAT, by means of deep sequencing. A Target Enrichment method using SureSelect system (Agilent) has been used to enrich in exons and regulatory regions of 524 genes belonging to these pathways.DNA from 2 tumoral-MF, 5 erythrodermic-MF and 4 SS patients, both normal and tumoral, were processed and sequenced with Genome Analyzer GA2 (Illumina) (PE-42bp). Sequencing data were first checked by FastQC and aligned to the human reference genome (GRCh37) using BWA and BFAST alignments. Somatic variants were identified using GATK. Thus, SNPs available at dbSNP 135 (hg19) and 1000 Genomes Project were filtered out from VCF output files. The GATK-QUAL field was employed for ranking selected somatic variants. Biological impact predictions for detected variants were obtained from Ensembl Variant Effect Predictor. Putative variants were manually reviewed and validated by capillary sequencing.Immunohistochemical analysis for NFAT, p50, p52 and STAT·p was also performed.qPCR-genotyping for specific variants was performed in a new cohort of 60 CTCL patients including SS and tumoral MFs. Results:Several mutations were found in essential genes belonging to pathways implicated in the Treg and Th17 regulatory pathways, NFkB and JAK/STAT, among others. PLCG1 was found mutated in three samples, two of them sharing the same mutation affecting one of the PLCG1 protein catalytic domains. This mutation was further analyzed by qPCR-genotyping in the new series of patients, being detected in 20% of samples.PLCG mutated cases showed a strong paraffin immunostaining for nuclear NFAT, p50 and p52.Additionally, immunological studies performed by flow cytometry in CTCL cell lines show aberrant coexpression of TH17 and Treg phenotypes. Conclusions:Activation of the TCR in CTCL might be partially dependent on the acquisition of somatic mutations in the coding region of genes known to play an essential role in T-cell differentiation processes and acquisition of TH17 and Treg phenotypes. Especially relevant is the finding that the catalytic domain of PLCG1 is frequently mutated in tumoral MF samples. Disclosures:No relevant conflicts of interest to declare.

Pathophysiology of chemokines and chemokine receptors in dermatological science: A focus on psoriasis and cutaneous T-cell lymphoma

Skin is an immunological organ with a delicate immunological network that governs the homeostasis and homing of the pro- and anti-inflammatory immune cells. Dysregulated immune response may result in psoriasis, a common inflammatory skin disease. Inappropriate immune surveillance, on the other hand, may cause the development of cutaneous lymphomas. Such homing of the immune cells likely depends on the interactions of chemokine and its receptors. Chemokine receptors and their corresponding chemokine ligands play key roles in the migration and localization of normal T cells in psoriasis and neoplastic T cells in cutaneous T-cell lymphoma (CTCL). While important in immune cell homing in psoriatic skin, chemokines and chemokine receptors may also be used for arrest, homing, and survival of neoplastic T cells in CTCL. In this review, we discuss roles of chemokine receptors, including those of CCR4 and CCR10 in the pathogenesis of CTCL, and of CCR6 in the pathogenesis of psoriasis. Targeting chemokines and chemokine receptors may lead to more effective treatments in these two skin diseases.

Tissue-Specific Homing of Immune Cells in Malignant Skin Tumors

Tissue-specific migration of immune cells involved both in physiological and pathological immune responses is a current research subject for medical science. Several homing molecules have been identified orchestrating extravasation of immune cells to certain peripheral non-lymphoid tissues such as gut, lung and skin. Regarding lymphocyte homing to skin, the first-line defense of human body cutaneous lymphocyte associated antigen (CLA) and a group of chemokine-chemokine receptor pairs are considered to be of crucial importance. The aim of the present review is to summarize existing knowledge about skin- and tumor-specific migration of immune cells playing a major pathogenetic role in host immune responses induced by non-lymphoid malignant skin tumors as well as in the development of primary cutaneous T-cell lymphomas (CTCL). Melanoma malignum, squamous and basal cell carcinoma evoke host immune responses and consequently a subset of reactive immune cells is recruited to site of the tumor. Regarding migratory process and exact functional role of these cells a growing number of data is available in literature. On the other hand tissue-specific immune cell homing is regarded as a key process in the pathogenesis of CTCL where malignant T-lymphocytes can be found in circulation and symptomatic skin. Hereby homing mechanism of malignant T-cells in mycosis fungoides and Sézary-syndrome as separate clinical entities of CTCL is discussed. A precise insight into the molecular background of skin- and tumor-specific immune cell migration can contribute to developing efficient vaccine therapies in non-lymphoid malignant skin tumors and beneficial treatment modalities in CTCL.

Up-Regulation of the Chemokine CCL18 by Macrophages Is a Potential Immunomodulatory Pathway in Cutaneous T-Cell Lymphoma

Mycosis fungoides (MF) is the most frequent form of cutaneous T-cell lymphoma (CTCL), which can deteriorate from patch stage to dermal-based tumors and systemic involvement in years. The interaction of chemokines in the skin with CTCL cells might have implications for the pathogenesis of the disease. In this study, we show by PCR analysis and immunofluorescence staining that the chemokine CCL18 is present in skin biopsy specimens of patients with MF and its precursor form parapsoriasis en plaque but not in healthy tissue. In addition, the serum levels of CCL18 were increased threefold in MF patients compared with those in healthy controls. In skin, CCL18 was specifically expressed by CD163(+) CD209(+) macrophages at the invasive margin of the tumor and not expressed by mature CD208(+) dendritic cells in the center of the tumor. The chemokine CCL17 was, by contrast, ubiquitously expressed. Furthermore, CCL18 promoted the chemotaxis but not the proliferation of CTCL cells. CCL18 inhibited proliferation of tumor cells and abolished the CXCL12-induced growth of a CTCL cell line. These data link the increased expression of CCL18 with CTCL and suggest an immunomodulatory effect of the chemokine in the pathogenesis of CTCL.

Structural alterations of the FAS gene in cutaneous T-cell lymphoma (CTCL)

FAS (TNF receptor superfamily member 6, also known as CD95) plays a major role in T-cell apoptosis and is often dysregulated in CTCL. We searched for structural alterations of the FAS gene with the potential to affect its function. Although several heterozygous FAS promoter single nucleotide polymorphisms (SNPs) were detected, the only homozygous one was the −671 GG SNP present in 24/80 CTCL cases (30%). This SNP maps to an interferon response element activated by STAT-1. EMSA and supershift EMSA showed decreased CTCL nuclear protein/STAT-1 binding to oligonucleotides bearing this SNP. Luciferase reporters showed significantly less interferon-alfa responsive expression by FAS promoter constructs containing this SNP in multiple CTCL lines. Finally, FAS was upregulated by interferon-alfa in wildtype CTCL cells but not those bearing the −671 GG SNP. These findings indicate that many CTCL patients harbor the homozygous FAS promoter −671 GG SNP capable of blunting its response to interferon. This may have implications for CTCL pathogenesis, racial incidence and the response of patients to interferon-alfa therapy. In contrast, functionally significant mutations in FAS coding sequences were detected uncommonly. Among CTCL lines with the potential to serve as models of FAS regulation, FAS-high MyLa had both FAS alleles, FAS-low HH was FAS-hemizygous and FAS-negative SeAx was FAS-null.

Nonmalignant T cells stimulate growth of T-cell lymphoma cells in the presence of bacterial toxins

AbstractBacterial toxins including staphylococcal enterotoxins (SEs) have been implicated in the pathogenesis of cutaneous T-cell lymphomas (CTCLs). Here, we investigate SE-mediated interactions between nonmalignant T cells and malignant T-cell lines established from skin and blood of CTCL patients. The malignant CTCL cells express MHC class II molecules that are high-affinity receptors for SE. Although treatment with SE has no direct effect on the growth of the malignant CTCL cells, the SE-treated CTCL cells induce vigorous proliferation of the SE-responsive nonmalignant T cells. In turn, the nonmalignant T cells enhance proliferation of the malignant cells in an SE- and MHC class II–dependent manner. Furthermore, SE and, in addition, alloantigen presentation by malignant CTCL cells to irradiated nonmalignant CD4+ T-cell lines also enhance proliferation of the malignant cells. The growth-promoting effect depends on direct cell-cell contact and soluble factors such as interleukin-2. In conclusion, we demonstrate that SE triggers a bidirectional cross talk between nonmalignant T cells and malignant CTCL cells that promotes growth of the malignant cells. This represents a novel mechanism by which infections with SE-producing bacteria may contribute to pathogenesis of CTCL.

P53 protein expression in cutaneous T-cell lymphomas.

p53 is an oncosuppressor gene located on chromosome 17p. Point mutations of the p53 gene are seen frequently in human malignancies, and are closely associated with malignant transformation under in vitro conditions. Mutated p53 protein shows a slow cell turnover rate, and accumulates in cells at the nuclear and/or cytoplasmic level. As a result, mutated p53 protein can be detected more readily by immunohistology than the wild-type protein. In this study, we used a monoclonal anti-p53 antibody (clone DO7) to examine the expression of p53 protein in 25 cutaneous T-cell lymphomas (CTCL) of low- and high-grade malignancy, i.e. mycosis fungoides (n = 6), Sézary's syndrome (n = 2), and large cell lymphomas of pleomorphic (n = 14) or anaplastic (n = 3) subtype. The results showed that easily detectable p53 protein was present in many of the neoplastic cells in half of the high-grade lymphomas. In contrast, in the low-grade lymphomas no, or only very few, p53-positive neoplastic cells could be detected. These findings suggest that molecular and/or genetic alterations of p53 may be implicated in the pathogenesis of high-grade CTCL, but are unlikely to be of critical importance in low-grade CTCL.

Biological Insights into the Pathogenesis of Cutaneous T-Cell Lymphomas (CTCL)

Mycosis fungoides and Sezary syndrome, collectively known as cutaneous T-cell lymphomas (CTCLs), are low-grade, indolent, clonal, non-Hodgkin's lymphomas consisting of CD4+ CD45RO+ T cells with a CLA+ CCR4+ skin-homing phenotype. There are several variants of primary CTCLs with differences in clinical behavior and prognosis. Currently, the precise etiologies of mycosis fungoides and Sezary syndrome are unknown. This article reviews our current understanding of the pathogenetic abnormalities involving genomic mutations, abnormal cDNA expression, and dysregulation of signaling pathways in CTCL.

Pilot study of etanercept in patients with relapsed cutaneous T-cell lymphomas

BackgroundTumor necrosis factor (TNF)-α has been implicated in the pathogenesis of cutaneous T-cell lymphoma (CTCL). ObjectiveTo assess the toxicity, safety, and efficacy of etanercept (soluble TNF receptor) in patients with relapsed CTCL. MethodsEtanercept was administered twice weekly at a dose of 25 mg subcutaneously. Patients with improvement after two months could be continued on treatment. ResultsTwelve out of the 13 patients enrolled on study were evaluable (Stage I-IIA, 3 patients; Stage IIB-IV disease, 9 patients). The median number of previous therapies was 7 (range, 3-12). Etanercept induced partial remission in one patient (8%) and minor response in one patient (8%), both of whom had Stage IB disease. Most patients experienced no side effects. ConclusionThis pilot study suggests that etanercept is safe and generally well tolerated in patients with CTCL. The effect of etanercept in a larger cohort of patients with early disease merits investigation.