Atypical CD8+ cutaneous T-cell lymphoma after immunomodulatory therapy

Systemic immunomodulatory agents have recently been approved for the treatment of rheumatoid arthritis and psoriasis. Although lymphomas are known to emerge in the setting of immunosuppressive therapy, it has not been well described or established for the newer biologic immune response modifiers. Herein, we describe 2 patients who developed unusual CD8+ cutaneous lymphoproliferative disorders after treatment with efalizumab and infliximab. The mechanisms and occurrence of lymphoma after immune response modifiers are discussed.     

Cytokine therapy of cutaneous T-cell lymphoma: interferons, interleukin-12, and interleukin-2

It is well accepted that cutaneous T-cell lymphomas (CTCL), including mycosis fungoides and Sézary syndrome, represent lymphomas that are highly responsive to immune modifying agents. Furthermore, the recent emphasis on the use of cytokine-related therapeutics is based upon the exceedingly important role of the host immune response in effecting progression of disease. In this article we discuss the data that support the importance of the host immune response in the control of progression of CTCL and the role that cytokine therapy has in supporting the host immune response and the effects of this approach to induce regression of skin and systemic disease.     

Identification of HNP3 as a tumour marker in CD4+ and CD4- lymphocytes of patients with cutaneous T-cell lymphoma

Cutaneous T-cell lymphomas (CTCL) are characterized by malignant proliferation of skin homing T-cells. Although prognosis is generally good, reliable markers are needed to identify patients at risk for a more aggressive course. ProteinChip (SELDI) technology was used as a tool for the discovery of protein patterns in lymphocytes from patients with CTCL (n=25) and unaffected controls (n=25). Lymphocytes were separated in CD4+ and CD4- fractions by magnetic cell sorting (MACS). Each whole protein extract was analysed by ProteinChip technology. The resulting protein profiles were submitted for bioinformatic analysis including a clustering algorithm, a rule extraction, a rating and a rule-base construction step. For the generated combined rule base for the CD4- cell fraction, both the sensitivity and specificity for the prediction of CTCL reached 96%, while for the CD4+ fraction they were 92% and 84%, respectively, for sensitivity and specificity. The most significant peak at 3489Da could be identified as HNP3, an alpha-defensin, by immunocapturing. These results open up both the possibility for the use of this protein signature, especially HNP3, to more effectively monitor and screen CTCL, and the avenue to identify the other relevant peaks for a better understanding of the development of this tumour.     

Expression of interleukin-18 and caspase-1 in cutaneous T-cell lymphoma.

PURPOSE: Cutaneous T-cell lymphoma (CTCL) is a malignancy of skin-homing Th2 T cells. Clonal T cells and CTCL skin lesions typically express Th2 cytokines, including interleukin (IL)-4, IL-5, and IL-10, but fail to produce Th1 cytokines. However, the reason for Th2 bias is unknown. IL-18 is a pleiotropic proinflammatory cytokine produced by monocytes/macrophages lineage as well as epithelial cells, such as human keratinocytes. In the absence of IL-12, IL-18 leads to increased immunoglobulin E production from B cells and enhanced production of IL-4 and IL-13 by basophils, mast cells, and CD4(+) T cells. We have analyzed cytokines in CTCL patients, which may bias the immune response around the Th1/Th2 axis. EXPERIMENTAL DESIGN: We examined plasma of 95 CTCL patients and skin of 20 CTCL patients for IL-18, caspase-1, IL-12, and other cytokines. To identify the presence or absence of these cytokine proteins in CTCL and normal skin, we cultured explants from skin biopsies on three-dimensional matrices. RESULTS: Plasma levels of IL-18 and its converting enzyme, caspase-1, were significantly elevated in CTCL. mRNA levels for these factors were also elevated in CTCL skin lesions. Matrices populated with CTCL lesional skin produced significant amounts of IL-18 and caspase-1; however, production of IL-12 protein was barely detectable. CONCLUSIONS: We propose that the high levels of IL-18 expression in lesional CTCL skin contribute to increased plasma levels of IL-18 and that this, in the face of significantly lower levels of IL-12, may contribute to the Th2 bias seen in this disease.     

CD95 ligand mediates T-cell receptor-induced apoptosis of a CD4+ CD8+ double positive thymic lymphoma

Tumors in the thymus can be of different cellular origin. Among the most common tumors are thymoma and lymphoma, which are derived from transformed thymic epithelial cells and transformed lymphocytes, respectively. Thymic lymphoma and their response to apoptotic stimuli are poorly characterized. Here, we analyse apoptosis events in the thymic lymphoma cell line Thy278, which expresses cell surface antigens characteristic of immature double positive thymocytes. Upon T-cell receptor (TCR)/CD3 stimulation, Thy278 cells die by apoptosis, similar as primary thymocytes during negative selection. Caspases are crucial for deletion of both Thy278 cells and normal thymocytes. Moreover, we show that deletion of primary thymocytes and Thy278 cells upon CD3 stimulation is considerably impaired by neutralizing CD95L antibody. Thus, our results not only demonstrate that TCR-induced apoptosis is still functional in transformed thymocytes, but also suggest that Thy278 cells are a helpful model for the molecular analysis of negative selection.     

The effects of trastuzumab on the CD4+CD25+FoxP3+ and CD4+IL17A+ T-cell axis in patients with breast cancer

In addition to the direct targeting effects on HER2-positive cells, trastuzumab may have a therapeutic role modulating the activity of the cellular immune system in patients with breast cancer. To investigate this further, the balance of T-regulatory (T_reg), Th17, natural killer (NK) and NK T (NKT) cells before, during and after trastuzumab therapy was investigated. Sequential frequencies of circulating T_reg cells, Th17 cells, NK and NKT cells were measured in peripheral blood of breast cancer patients and normal controls throughout therapy. Individuals with breast cancer had significantly higher T_reg frequencies of peripheral blood compared with healthy controls (9.2 or 8.6 vs 6%; P<0.05), and no significant differences in T_reg frequencies were observed between HER2-positive and HER2-negative individuals. The number of Th17 cells was lowest in HER2-positive patients compared with both healthy controls and HER2-negative patients (0.31 vs 0.75% or 0.84%; P=0.01). There appeared to be an inverse relationship between T_reg and Th17 frequencies in metastatic breast cancer (MBC) with T_reg levels significantly reduced during treatment with trastuzumab (P=0.04), whereas Th17 frequencies were concomitantly increased (P=0.04). This study supports earlier data that T_reg cells are present at higher frequencies in breast cancer patients compared with healthy individuals. For the first time, we show that HER2-positive individuals with breast carcinomas have reduced numbers of circulating Th17 cells, which appear, in turn to have an inverse relationship with T_reg frequency in MBC. The change in balance of the T_reg : Th17 ratio appears to characterise the cancer state, and furthermore, is disrupted by trastuzumab therapy.     

皮肤T淋巴细胞瘤的研究进展

随着对皮肤T细胞淋巴瘤(CTCL)病理发生机制研究的深入以及肿瘤治疗技术的进步,近年来对CTCL诊断及治疗方法也有了一定的进展.早期CTCL首选局部治疗及免疫治疗.全身皮肤电子束治疗、靶向治疗及化疗、联合治疗、维持治疗是进展期CTCL的主要治疗手段.近年来新的药物及治疗方法不断涌现,本文对皮肤T细胞淋巴瘤的临床特点及其...     

Atopic dermatitis-like non-erythrodermic leukemic variant of CD3(-/+dim) CD4(+) cutaneous T-cell lymphoma preceded by cutaneous papular xanthomatosis

We report a patient with cutaneous papular xanthomatosis who 4 years later developed a CD3(-/+dim)/CD4(+) T-cell lymphoma. Pruritic xerotic non-erythrodermic skin, eosinophilia and hyper-IgE were present and erroneously classified as atopic dermatitis. Flow cytometry and DNA ploidy analysis of both blood and skin lymphocytes, skin histology and blood T-cell receptor gene rearrangement studies confirmed diagnosis of T-cell lymphoma. Monoclonal CD3(-/+dim)/CD4(+) T-cells were especially prone to the synthesis of IL-13, a cytokine that is involved in IgE-secretion, and comprised both a medium (diploid) and large (hyperploid) sized T-cell populations with a similar immunophenotype. The majority of the normal residual T-cells were large granular lymphocytes, expressed activation-related and natural-killer-associated markers and secreted high levels of interferon gamma, suggesting that they might correspond to active cytotoxic cells directed against the neoplastic T-lymphocytes.     

Development of IgG lambda multiple myeloma in a patient with cutaneous CD30+ anaplastic T-cell lymphoma.

We report a patient with an epidermotropic cutaneous T-cell lymphoma which transformed into an anaplastic cutaneous CD30+ T-cell lymphoma. Repeated relapses required prolonged systemic PUVA therapy. Two years after diagnosis, the patient had several episodes of infections of the respiratory tract. Serum electrophoresis now revealed significantly reduced polyclonal immunglobulin production and an additional band in the gamma fraction corresponding to IgG lambda monoclonal gammopathy. Thereafter, the patient suffered a pathologic fracture of the dorsolateral 5th rib on the right side and an accumulation of monoclonal plasma cells in the bone marrow confirmed the diagnosis of multiple myeloma (IgG lambda). Accordingly, 6 cycles of cytoreductive chemotherapy (alkeran, decortin) were given. After one year of steady state disease the patient lost weight and bone pain increased while only a few papular eruptions were detectable. Radiography showed multiple small osteolytic areas. A few months later he died with signs of bone marrow insufficiency.     

淋巴瘤细胞EL-4培养上清液对CD4＋CD25＋调节性T细胞比例的调节作用

目的探讨肿瘤细胞是否能上调CD4＋CD25＋调节性T细胞（Treg细胞）的比例。方法将小鼠淋巴瘤细胞株EL4培养上清液与正常小鼠脾脏淋巴细胞混合培养72h,流式细胞仪检测其中CD4＋CD25＋Treg细胞含量,RT—PCR检测Foxp3mRNA的表达,实验重复3次。结果和EL-4培养上清液混合培养的正常小鼠脾脏淋巴细胞中CD4＋CD25＋Treg细胞的比例高于对照组（P〈0．05）,在CD3单抗刺激的同时加入EL-4培养上清液后,小鼠脾脏中CD4＋CD25＋Treg细胞仍呈同步增加（P〈0．05）;且Foxp3mRNA的表达增加。结论淋巴瘤细胞分泌的免疫调节因子能诱导CD4＋CD25＋Treg细胞的增生,提示肿瘤可以上调CD4＋CD25＋Treg细胞比例。     

Peripheral T-cell tolerance associated with prostate cancer is independent from CD4+CD25+ regulatory T cells

CD4(+)CD25(+)Foxp3(+) regulatory T cells (Treg) are thought to suppress the natural and vaccine-induced immune response against tumor-associated antigens (TAA). Here, we show that Treg accumulate in tumors and tumor-draining lymph nodes of aging transgenic adenocarcinoma of the mouse prostate (TRAMP) male mice, which spontaneously develop prostate cancer. TAA overexpression and disease progression associate also with induction of TAA-specific tolerance. TAA-specific T cells were found in the lymphoid organs of tumor-bearing mice. However, they had lost the ability to release IFN-gamma and kill relevant targets. Neither in vivo depletion of Treg by PC61 monoclonal antibody followed by repeated vaccinations with antigen-pulsed dendritic cells nor the combined treatment with 1-methyl-L-tryptophan inhibitor of the enzyme indoleamine 2,3-dyoxigenase, PC61 antibody, and dendritic cell vaccination restored the TAA-specific immune response. Treg did not seem to control the early phases of tolerance induction, as well. Indeed, depletion of Treg, starting at week 6, the age at which TRAMP mice are not yet tolerant, and prolonged up to week 12, did not avoid tolerance induction. A similar accumulation of Treg was found in the lymph nodes draining the site of dendritic cell vaccination both in TRAMP and wild-type animals. Hence, we conclude that Treg accrual is a phenomenon common to the sites of an ongoing immune response, and in TRAMP mice in particular, Treg are dispensable for induction of tumor-specific tolerance.     

Simultaneous generation of CD8+ and CD4+ melanoma-reactive T cells by retroviral-mediated transfer of a single T-cell receptor

Adoptive immunotherapy of cancer requires the generation of large numbers of tumor antigen-reactive T cells for transfer into cancer patients. Genes encoding tumor antigen-specific T-cell receptors can be introduced into primary human T cells by retroviral mediated gene transfer as a potential method of providing any patient with a source of autologous tumor-reactive T cells. A T-cell receptor-specific for a class I MHC (HLA-A2)-restricted epitope of the melanoma antigen tyrosinase was isolated from a CD4(+) tumor-infiltrating lymphocyte (TIL 1383I) and introduced into normal human peripheral blood lymphocytes by retroviral transduction. T-cell receptor-transduced T cells secreted various cytokines when cocultured with tyrosinase peptide-loaded antigen-presenting cells as well as melanoma cells in an HLA-A2-restricted manner, and could also lyse target cells. Furthermore, T-cell clones isolated from these cultures showed both CD8(+) and CD4(+) transduced T cells could recognize HLA-A2(+) melanoma cells, giving us the possibility of engineering class I MHC-restricted effector and T helper cells against melanoma. The ability to confer class I MHC-restricted tumor cell recognition to CD4(+) T cells makes the TIL 1383I TCR an attractive candidate for T-cell receptor gene transfer-based immunotherapy.     

Depletion of CD4+CD25+ regulatory T cells enhances interleukin-2-induced antitumor immunity in a mouse model of colon adenocarcinoma

Interleukin 2 (IL)-2 induces antitumor immunity and clinical responses in melanoma and renal cell carcinoma. However, IL-2 also increases the number of CD4(+)CD25(+) regulatory T (Treg) cells that suppress antitumor immune responses. The aim of the present study was to elucidate the effect of depletion of Treg cells on IL-2-induced antitumor immunity. IL-2-transfected mouse colon adenocarcinoma (MC38/IL-2) cells were implanted subcutaneously or intrahepatically into male C57BL/6 mice, and tumor growth and the proportion of tumor-infiltrating lymphocytes with Treg-cell depletion in response to treatment with anti-CD25 monoclonal antibody (PC61) were determined. In mice treated with phosphate-buffered saline, 40-60% of MC38/IL-2 tumors were rejected. In contrast, all MC38/IL-2 tumors were rejected in mice treated with PC61. The number of tumor-infiltrating CD8(+) T cells in mice treated with PC61 was approximately twice that in mice treated with PBS. The numbers of tumor-infiltrating CD4(+) and natural killer cells were also increased significantly. To test the antimetastatic effects of IL-2 treatment in combination with Treg-cell depletion, human recombinant IL-2 (rIL-2) and PC61 were administered to mice implanted with MC38/mock cells in the spleen, and hepatic metastasis was investigated. The average liver weight in mice treated with rIL-2 plus PC61 was 1.04 +/- 0.03 g, less than that in mice treated with rIL-2 (2.04 +/- 0.51 g) or PC61 alone (1.81 +/- 0.38 g). We conclude that IL-2-induced antitumor immunity is enhanced by Treg-cell depletion and is due to expansion of the tumor-infiltrating cytotoxic CD8(+) T-cell population.     

Recognition of adult T-cell leukemia/lymphoma cells by CD4+ helper T lymphocytes specific for human T-cell leukemia virus type I envelope protein.

PURPOSE: Human T-cell leukemia virus type I (HTLV-I) can cause an adult T-cell leukemia/lymphoma (ATLL). Because ATLL is a life-threatening lymphoproliferative disorder and is resistant to chemotherapy, the establishment and enhancement of T-cell immunity to HTLV-I through the development of therapeutic vaccines could be of value. Thus, the identification of HTLV-I epitopes for both CD8(+) and CD4(+) T cells should facilitate the development of effective vaccines. Although numerous HTLV-I epitopes for CTLs have been identified, few epitopes recognized by CD4(+) helper T cells against this virus have been described. EXPERIMENTAL DESIGN: Synthetic peptides prepared from several regions of the HTLV-I envelope (Env) sequence that were predicted to serve as helper T-cell epitopes were prepared with use of computer-based algorithms and tested for their capacity to trigger in vitro helper T-cell responses using lymphocytes from normal volunteers. RESULTS: The results show that the HTLV-I-Env(317-331), and HTLV-I-Env(384-398)-reactive helper T lymphocytes restricted by HLA-DQw6 and HLA-DR15, respectively, could recognize intact HTLV-I+ T-cell lymphoma cells and, as a consequence, secrete lymphokines. In addition, HTLV-I Env(196-210)-reactive helper T lymphocytes restricted by HLA-DR9 were able to directly kill HTLV-I+ lymphoma cells and recognize naturally processed antigen derived from killed HTLV-I+ lymphoma cells, which was presented to the helper T cells by autologous antigen-presenting cells. CONCLUSIONS: The present findings hold relevance for the design and optimization of T-cell epitope-based immunotherapy against HTLV-I-induced diseases such as ATLL.     

Attenuation of CD8(+) T-cell function by CD4(+)CD25(+) regulatory T cells in B-cell non-Hodgkin's lymphoma

The underlying mechanisms by which tumor cells are resistant to CTL-mediated apoptosis are not clear. Using a human model of B-cell non-Hodgkin's lymphoma (B-cell NHL), we show that intratumoral T(reg) cells inhibit the proliferation and granule production of activated autologous infiltrating CD8(+) T cells. Our results also show that degranulation and subsequent cytotoxic activity of infiltrating CD8(+) T cells exposed to lymphoma B cells is completely attenuated by the presence of intratumoral T(reg) cells. Furthermore, we show that increased numbers of intratumoral T(reg) cells correlates with the number of CD8(+) T cells in biopsy specimens from patients with B-cell NHL, supporting the in vitro findings that intratumoral T(reg) cells inhibit proliferation of infiltrating CD8(+) T cells. Taken together, these data indicate that human lymphoma B cells are sensitive to autologous CTL-mediated cell death but are protected by the inhibitory function of intratumoral T(reg) cells.     

Distinct and overlapping roles of interleukin-10 and CD25+ regulatory T cells in the inhibition of antitumor CD8 T-cell responses

Lack of antitumor immunity is often related to impaired CD8 T-cell responses that could result from a poor priming capacity by tumor-infiltrating dendritic cells (TIDC) and/or further inhibition by regulatory T cells (T(reg)). Interleukin-10 (IL-10) has been implicated in the inhibition of TIDC as well as in the generation and functions of T(reg). Here, we address some of the respective and possibly overlapping roles of IL-10 and CD25+ T(reg) in CD8 antitumor immunity. Whereas tumor antigen-specific CD8 T cells proliferated in vivo in the presence of IL-10 or T(reg), optimal effector functions were observed in mice lacking both IL-10 and T(reg). Indeed, tumors grown in normal but not in IL-10-deficient or CD25-depleted mice induced tumor antigen-specific CD8 suppressor T cells. Suppression involved transforming growth factor-beta. Similarly, both IL-10 and T(reg) were responsible for impaired CD8 T cell priming by TIDCs, but IL-12 production by TIDCs was prevented only by T(reg)-independent IL-10. Subsequently, IL-10 defect and T(reg) depletion were required to achieve optimal induction of CD8 T-cell effectors by TIDC following CpG activation. Our results point out major redundant and nonredundant roles for IL-10 and T(reg) in the inhibition of TIDC-mediated generation of antitumor CD8 T-cell response.