Sézary syndrome: phenotypic and functional characterization of the neoplastic cells

Phenotypic properties of the neoplastic cells in skin, blood and lymph node specimens from 5 patients with the Sézary syndrome were examined by immuno-enzymatic and -fluorescence labelling of cells and tissue sections with a monoclonal antibody panel. In 3 cases, the in vitro functional properties of the neoplastic cells (isolated from blood specimens) were also analysed using a reverse plaque-forming cell assay. 3 different immunological categories were identified as follows: T-helper/inducer neoplasms (3 patients); T-suppressor/cytotoxic neoplasms (1 patient); and neoplastic T-cells demonstrating characteristics consistent with a concept of their derivation from inducible suppressor T-cells (1 patient). These data provide conclusive evidence that Sézary syndrome is heterogeneous with respect to the immunological characteristics of the neoplastic cells.     

Ultrastructural, Immunologic, and Functional Studies on Sézary Cells: A Neoplastic Variant of Thymus-Derived (T) Lymphocytes

The vast majority of human lymphoid neoplasms examined to date have been associated with a proliferation of bone marrow-dependent (B) lymphocytes. In an effort to delineate human tumors of T-cell (thymusdependent) lineage, use was made of the peripheral blood leukocytes of sixteen subjects with various forms of mycosis fungoides. The abnormal cells in the circulation of these patients are morphologically identical to those that infiltrate their nodes and skin. On electron microscopy, such neoplastic lymphocytes (Sézary cells) had "cerebriform" nuclei and an abundance of cytoplasmic fibrils not described heretofore. Sézary cells were nonadherent and nonphagocytic and usually responded to stimulation with phytohemagglutinin, refuting earlier suggestions that the cells represent monocytes or histiocytes. In contrast to chronic lymphocytic leukemia lymphocytes, the Sézary cells lacked surface immunoglobulin and receptors for complement. Ultrastructural analysis identified Sézary cells in the center of directly formed rosettes (E-rosettes) characterizing the behavior of T lymphocytes in this test. Though some Sézary cells lacked both T and B cell-surface properties, in general, these observations support the view that the Sézary cell is a neoplastic variant of a thymusderived lymphocyte.     

The Sézary syndrome cell: surface ultrastructural characteristics.

Peripheral blood mononuclear cells from a patient with Sézary syndrome which lacked E-rosette-forming ability and surface immunoglobulins, and which displayed a markedly depressed response to a variety of mitogens, were studied by scanning electron microscopy (SEM) and transmission electron microscopy (TEM) on 3 occasions. The first peripheral blood sample (smooth) differed significantly from two later samples (moderate numbers of microvilli) when surface characteristics were examined by SEM; these differences were confirmed by TEM. The Sézary syndrome cells in this patient may be related to a T lymphocyte which has lost certain surface markers and mitogen response characteristics through a process of de-differentiation.     

Sézary syndrome associated with hepatocellular carcinoma and monoclonal gammopathy--a case report

We treated 54-year-old Japanese man with a large cell type of Sézary syndrome. He had generalized erythrodermia, superficial lymphadenopathy, atypical lymphocytes in the peripheral blood, anti-HTLV-I antibody negativity and chromosomal abnormality. The patient was a hepatitis B virus carrier, and was complicated with hepatocellular carcinoma and monoclonal gammopathy of IgG, lambda type. Sézary syndrome is a T cell malignancy, the clinical course of which is relatively mild and chronic; accordingly, this case showed no crisis under chemotherapy. However, the patient died due to rapid growth of the hepatoma. Although case reports of Sézary syndrome complicated with other malignancies are very few, the occurrence of malignancies is possible because of decreased immunological function in the patients. In this case, hepatitis B virus might participate in the hepatic oncogenesis under dysfunction of helper/inducer cells. In addition, the complication of monoclonal gammopathy was also interesting from the standpoint of the helper function of Sézary cells.     

Significance of circulating T-cell clones in Sezary syndrome

Identification of malignant Sézary cells by T-cell receptor (TCR) clonality studies is routinely used for the diagnosis of Sézary syndrome, but T-cell clones expressed in a single patient have never been accurately characterized. We previously reported that CD158k expression delineates Sézary syndrome malignant cells, and, more recently, we identified vimentin at the surface membranes of Sézary cells and normal activated lymphocytes. In the present study, T-cell clones from 13 patients with Sézary syndrome were identified by immunoscopy and further characterized in the blood according to their TCR Vbeta, CD158k, and vimentin cell-surface expression. We found in most patients a unique malignant T-cell clone that coexpressed CD158k and vimentin and that, when patients were tested, was also present in the skin. However, in some patients we detected the presence of a nonmalignant circulating clone expressing high amounts of vimentin and lacking CD158k. These results indicate that clonal expansion may originate from circulating malignant and nonmalignant CD4(+) T cell populations in patients with Sézary syndrome. Identification of the malignant cells in Sézary syndrome cannot be achieved by T-cell clonality studies or by TCR Vbeta monoclonal antibody (mAb) analysis alone; it also relies on CD158k phenotyping.     

The Sézary syndrome--an erythrodermic T-cell-lymphoma]

Report on a 75-year-old patient with Sézary syndrome which is characterized by the following leading symptoms: Erythrodermia, leukocytosis, circulating atypical lymphocytes with cerebriformous nuclei (Sézary cells) and swollen lymph nodes. Light microscopic and electron microscopic findings refer to a transitory situation of the Sézary cells from the reactive character to the (pre)neoplastic one. The disease is classified by T-(helper)cells as a low grade non-Hodgkin-lymphoma in parallel to mycosis fungoides and in general it finishes lethally after a course of 5 to 8 years. Apart from local steroid ointments and UV-irradiations the internal application of steroids, in the late stage if necessary in combination with cytostatics is recommended for therapy.     

Detection of human T-lymphotropic virus-like particles in cultures of peripheral blood lymphocytes from patients with mycosis fungoides.

Because of seronegativity and absence of a leukemic phase in most patients with mycosis fungoides, a role for the human T-lymphotropic virus type I (HTLV-I) in this disease has remained tenuous. Virus particles are not seen in fresh isolates of skin or blood lymphocytes and the malignant cells (Sézary cells) have been difficult to culture. The availability of growth factors and biomolecular techniques have prompted a renewed attempt to find evidence of virus infection in these patients. We report here the successful culture of blood lymphocytes of 17 patients with mycosis fungoides and 1 patient with the Sézary syndrome. The cells of 2 additional patients failed to grow after 4-6 weeks in vitro. Ultrastructural analysis of the cultures showed an abundance of HTLV-like particles in the specimens of 18 of the 20 patients. Preliminary immunohistochemical studies carried out with various antisera directed against HTLV-I and the polymerase chain reaction utilizing a probe for a conserved region of the pol gene of HTLV-I were positive on only a portion of the specimens. Although definitive characterization of this organism awaits further analysis, it seems likely that circulating lymphocytes of all patients with mycosis fungoides harbor a virus that morphologically resembles HTLV-I.     

A prospective study on the evolution of the T-cell repertoire in patients with Sézary syndrome treated by extracorporeal photopheresis

Sézary syndrome is a leukemic form of epidermotropic cutaneous T-cell lymphoma related to the malignant proliferation of clonal CD4(+) T cells. Extracorporeal photochemotherapy may induce a transient improvement of the clinical signs, but its efficiency is discussed. To investigate the frequency of the T-cell clone in the peripheral blood of patients with Sézary syndrome and to monitor its evolution in patients treated using extracorporeal photopheresis or chemotherapy, we used the immunoscope technique. In one patient, we observed a decrease of the relative frequency of the clone from 15.6% to 0%, paralleling a complete remission of the clinical disease and a disappearance of the circulating Sézary cells. In the other cases, the evolution of the relative frequency paralleled the initial improvement of the clinical status and the absence of long-term efficiency in patients treated with extracorporeal photopheresis or chemotherapy. We observed a quick-acting direct cytotoxicity of the association 8MOP + UVA on the T-cell clone. The immunoscope technique appears to be an efficient tool to appreciate the amount of tumoral cells and to monitor the evolution of the clonal component in the Sézary syndrome.     

Prognostic significance of tumor burden in the blood of patients with erythrodermic primary cutaneous T-cell lymphoma

Erythrodermic cutaneous T-cell lymphoma (CTCL) includes patients with erythrodermic mycosis fungoides who may or may not exhibit blood involvement and Sézary syndrome and in whom hematological involvement is, by definition, present at diagnosis. These patients were stratified into 5 hematologic stages (H0-H4) by measuring blood tumor burden, and these data were correlated with survival. The study identified 57 patients: 3 had no evidence of hematologic involvement (H0), 8 had a peripheral blood T-cell clone detected by polymerase chain reaction (PCR) analysis of the T-cell receptor gene and less than 5% Sézary cells on peripheral blood smear (H1), and 14 had either a T-cell clone detected by Southern blot analysis or PCR positivity with more than 5% circulating Sézary cells (H2). Twenty-four patients had absolute Sézary counts of more than 1 x 10(9) cells per liter (H3), and 8 patients had counts in excess of 10 x 10(9) cells per liter (H4). The disease-specific death rate was higher with increasing hematologic stage, after correcting for age at diagnosis. A univariate analysis of 30 patients with defined lymph node stage found hematologic stage (P =.045) and lymph node stage (P =.013) but not age (P =.136) to be poor prognostic indicators of survival. Multivariate analysis identified only lymph node stage to be prognostically important, although likelihood ratio tests indicated that hematologic stage provides additional information (P =.035). Increasing tumor burden in blood and lymph nodes of patients with erythrodermic CTCL was associated with a worse prognosis. The data imply that a hematologic staging system could complement existing tumor-node-metastasis staging criteria in erythrodermic CTCL.     

Extracorporeal photopheresis in Sézary syndrome: hematologic parameters as predictors of response

Data were analyzed from 23 patients with Sézary syndrome (defined by erythroderma, more than 10% circulating atypical mononuclear cells, and peripheral blood T-cell clone) undergoing monthly extracorporeal photopheresis as the sole therapy for up to 1 year. The cohort showed a significant reduction of skin scores during treatment (P =.001). Thirteen patients (57%) achieved a reduction in skin score greater than 25% from baseline at 3, 6, 9, or 12 months (responders). Reduction in skin score correlated with reduction in the Sézary cell count as a percentage of total white cell count (P =.03). Responders and nonresponders were compared. None of the measured parameters was significantly different between the 2 groups. It was assessed whether any of the baseline parameters predicted outcome. A higher baseline lymphocyte count was significantly associated with a decrease in skin score at 6 months (P <.05). A higher baseline Sézary cell count as a percentage of total white cell count predicted a subject was more likely to be a responder after 6 months of treatment (P =.021). No other parameters predicted responder status. These data show that the modest falls in CD4, CD8, and Sézary cell counts were seen in all patients and might have resulted from lymphocyte apoptosis. This mechanism could explain the more favorable response seen in patients with higher percentages of Sézary cells in the peripheral blood. Alternatively, minimum tumor burden might be required for the induction of a cytotoxic response. Analysis of tumor-specific cytotoxic T cells is needed to investigate these possibilities further.     

Differential expression of CD24-related epitopes in mycosis fungoides/Sézary syndrome: a potential marker for circulating Sézary cells.

In the hematopoietic system, the B-cell associated antigen CD24 is expressed at high density on B cells, B-cell precursors, and B-cell malignancies as well as at low density on peripheral blood polymorphonuclear leukocytes. The 42-Kd sialoglycoprotein has not been previously demonstrated to be expressed on T cells, thymocytes, or T-cell malignancies. We identified three patients with mycosis fungoides/Sézary syndrome that showed low density expression of the CD24-related epitope recognized by antibody BA-1 on circulating T cells. All three patients had Sézary cells by morphologic assessment and clonal T-cell populations in the peripheral blood by gene rearrangement studies. In two of these patients, indirect immunofluorescence with a panel of six anti-CD24 monoclonal antibodies demonstrated reactivity for two of six antibodies in one case and only one of six antibodies in the other. The biologic significance of CD24-related epitope expression on circulating T cells in mycosis fungoides/Sézary syndrome is unclear. However, these findings suggest that differential, low density expression of CD24-related epitopes (BA-1+, OKB2-) may be a useful phenotypic marker for identifying circulating Sézary cells.     

Renal insufficiency caused by leukemic cell infiltration in Sézary syndrome

A 77-year-old woman was admitted to our hospital because of pneumonia and heart failure in July 2002. She had been diagnosed as having with Sézary syndrome in 1993, and had been treated with a combination of prednisolone, methotrexate, and cyclosporin A with subsequent stable disease but persistent generalized erythroderma. On admission, the white blood count was 23.9 X 10(9)/L with 28% Sézary cells, and serum creatinine levels were within normal limits. One month after admission, the pneumonia and heart failure improved remarkably with antibiotics and diuretics. However, at the same time, her renal function deteriorated with increasingly high serum creatinine levels. She died of anuria in September, 2002. An autopsy showed marked perivascular and peritubular infiltration of abnormal lymphocytes with degenerative nephrotubuli in the kidneys. This patient may be the first reported case of Sézary syndrome with renal failure caused by leukemic infiltration.     

Unclassified mature T cell leukemia with cerebriform nuclei

A 53 year-old male visited our hospital for evaluation of his leukocytosis, which was first diagnosed more than 6 years previously. He was asymptomatic and there were no remarkable findings on physical and laboratory examinations except for the lymphocytosis. Abnormal lymphocytes with deep folded nuclei were seen on light microscopy, whose phenotype was CD3+, CD4-, CD8-, CD7-, CD16 , CD56-, CD45RO+ and CD45RA- . Electron microscopy revealed 'cerebriform nuclei' which were characteristic of Sézary cells. Adult T cell leukemia (ATL) and Sézary syndrome (SS) were ruled out because of the negative HTLV-1 test and the absence of skin lesions, respectively. T-prolymphocytic leukemia (T-PLL), which is characterized by a marked increase in leukocytes having a CD7-phenotype and a progressive fatal course, was also excluded. Recently, the TCL1 onco-protein has been shown to be overexpressed in progressive T-PLL but not in other mature T cell leukemias including Sézary syndrome. Peripheral mononuclear cells in the present patient did not overexpress TCL1. In its morphology and phenotypes, our case resembled 'Sézary cell leukemia (SCL)' but the clinical course was much more indolent. This case did not match any of the mature T cell leukemias defined in the WHO classification.     

Analysis of p53 and mdm-2 expression in 18 patients with Sézary syndrome

Sézary syndrome is a leukaemic form of cutaneous T-cell lymphoma which presents with multiple cytogenetic abnormalities and responds poorly to chemotherapy. Because of the importance of the p53 tumour suppressor in maintaining genomic stability and in sensitizing transformed cells to DNA damaging agents, we looked for alterations which may affect p53 functions in 18 patients with Sézary syndrome. Cytogenetic analysis suggested frequent p53 gene inactivation since 6/18 patients had loss of one copy of 17p. However, single-strand conformational polymorphism (SSCP) revealed that p53 gene mutations are relatively rare, occurring in only two of 18 Sézary patients. Neither of these two patients was missing a copy of 17p. Possible abnormalities of p53 pathway function through mdm-2 over-expression were also investigated. Although all 18 patients had normal levels of mdm-2 RNA, 4/18 over-expressed mdm-2 protein. One patient with advanced disease and the highest percentage of malignant cells overexpressed mdm-2 protein and possessed a nonsense p53 gene mutation. The five patients with abnormalities of p53 or mdm-2 were found to have significantly higher absolute lymphocyte counts and higher absolute numbers of Sézary cells (P = 0.021 and 0.027 respectively). In summary, molecular alterations of 17p and potential p53 pathway abnormalities are a common event in Sézary syndrome and appear to be associated with more advanced disease.     

Complication of topoisomerase II inhibitor-related acute promyelocytic leukemia with t(1;10) (q21;q26) in a patient with Sézary syndrome

A 74-year-old man was diagnosed as having Sézary syndrome in 1999. Treatment with combination chemotherapy could not completely control both the erythroderma and Sézary cells. However, treatment with oral administration of etoposide was able to maintain the patient in a good condition for about 4 years. In June 2004, he developed topoisomerase II inhibitor-related acute promyelocytic leukemia. Chromosomal analysis demonstrated abnormalities of t(1;10) (q21;q26) and t(15;17) (q22;q12) in 17 of 20 cells. Despite treatment with ATRA and combination chemotherapy, the patient died of brain hemorrhage.     

Successful treatment of chemotherapy-refractory Sézary syndrome with alemtuzumab (Campath-1H)

INTRODUCTION: Sézary syndrome (SS) is a cutaneous T-cell lymphoma characterized by erythroderma, lymphadenopathy and circulating atypical T cells. Median survival after diagnosis is 10 yr, with chemotherapy resistance being a major problem in advanced disease. Alemtuzumab (Campath-1H) is a monoclonal antibody directed against the lymphocytic antigen CD52, expressed on B- and T-cells. Alemtuzumab is approved for relapsing chronic B-cell leukemia and seems to be active also in T-cell lymphomas such as T-cell prolymphocytic lymphoma, SS and mycosis fungiodes. CASE HISTORY: A 32-yr-old male patient presented with advanced stage, extensively pretreated SS with heavily itching erythroderma, peripheral lymphadenopathy, circulating Sézary cells and bone marrow infiltration. The disease had not responded to PUVA/interferon-alpha and progressed on chemotherapy with CHOP, 2-CDA, vinorelbine, etoposide and liposomal doxorubicin. Following treatment with alemtuzumab (30 mg i.v. three times per week for 10 wk), itching resolved rapidly and an almost complete remission was achieved within 3 months after starting this treatment. At 12-month follow up, no disease progression was present. CONCLUSION: In accordance with previous data, this single case underlines the potent activity of alemtuzumab in advanced, chemotherapy-refractory SS.     

Sézary syndrome showing a stable clinical course for more than four years after oral administration of etoposide and methotrexate

A 62-year-old woman was admitted because of leukocytosis, systemic lymph node swelling and erythroderma. Laboratory data disclosed a WBC count of 15,600/microliter (CD4-positive cells: 91%). CD25 and HTLV-1 were negative. A skin biopsy revealed the involvement of T cells. These data and findings were consistent with a diagnosis of Sézary syndrome. Although the patient was treated with 2 courses of CHOP, the leukocyte count did not decrease. We then treated the patient orally with etoposide (25 mg/day) and methotrexate (10 mg/week), and this resulted in reduction of the leukocyte count and lymph node swelling, and amelioration of the erythroderma. As Sézary syndrome is an extremely rare disease, it is worthwhile reporting cases like the present one, where therapy was successful.     

Histopathologic staging at initial diagnosis of mycosis fungoides and the Sézary syndrome. Definition of three distinctive prognostic groups

STUDY OBJECTIVE: To determine the optimal staging evaluation at the time of initial diagnosis of mycosis fungoides or the Sézary syndrome. DESIGN: Retrospective review of a uniformly staged inception cohort. SETTING: Single-institution tertiary care center. PATIENTS: 152 consecutive patients who had mycosis fungoides with or without the Sézary syndrome within 6 months of the initial definitive diagnosis. INTERVENTION: A detailed staging evaluation including physical examination, routine laboratory studies, chest roentgenogram, lymphangiogram, peripheral blood smear, lymph node biopsy, bone marrow aspirate or biopsy, and liver biopsy in selected patients. MEASUREMENTS AND MAIN RESULTS: Univariate adverse prognostic features at initial diagnosis in patients with mycosis fungoides included (P less than 0.01) one or more cutaneous tumors or generalized erythroderma, adenopathy, blood smear involvement with Sézary cells, lymph node effacement, eosinophilia, and visceral involvement. Important, independent prognostic factors in a multivariate analysis are the presence of visceral disease and type of skin involvement. CONCLUSIONS: A staging system based on histopathologic evaluation of skin, lymph nodes, blood, and visceral sites provides more comprehensive prognostic information than clinical evaluation of skin disease and adenopathy. Patients may be divided at initial presentation into three prognostic groups: good-risk patients, who have plaque-only skin disease without lymph node, blood, or visceral involvement (median survival, greater than 12 years); intermediate-risk patients, who have cutaneous tumors, erythroderma, or plaque disease with node or blood involvement but no visceral disease or node effacement (median survival, 5 years); and poor-risk patients, who have visceral involvement or node effacement (median survival, 2.5 years).     

Gastrointestinal involvement in the Sézary syndrome.

A case of documented Sézary syndrome, a cutaneous T-cell lymphoma, with gastrointestinal lymphocytic infiltration is presented. The symptom of diarrhea waxed and waned with the course of the disease process. Radiographic studies were normal, and no evidence of malabsorption was elicited. The diagnosis was established by endoscopic and biopsy techniques, with light and electron microscopy of colonic and small bowel biopsies demonstrating extensive infiltration with typical Sézary cells. To our knowledge this is the first reported case of gastrointestinal involvement in the Sézary syndrome.     

The use of anti-T-cell receptor-Vbeta antibodies for the estimation of treatment success and phenotypic characterization of clonal T-cell populations in cutaneous T-cell lymphomas

Sézary syndrome and Mycosis fungoides are the most common forms of cutaneous T-cell lymphomas. To assess the response to different therapies especially in Sézary syndrome, it is helpful to monitor the percentage of circulating tumour cells in the blood. The use of T-cell receptor (TCR)-Vbeta specific monoclonal antibodies provides a suitable tool for detecting Sézary cells. In this study, we analysed the levels of clonal CD4+Vbeta+ cells of seven patients with various treatment modalities using flow cytometry and investigated the immunophenotype of the clonal cells by double staining with a panel of antibodies recognizing lymphatic surface markers. Additionally, a polymerase chain reaction-denaturing gradient gel electrophoresis assay was performed on clonal CD4+Vbeta2+ cells, showing that these cells carry a Vgamma10/11, JgammaP1/2 TCR rearrangement. Follow-up studies revealed close association of the Vbeta+ clone developmentwith the clinical response to different therapiesinsixpatients. Intwo cases, the CD4+Vbeta+ cells decreased accompanied by partial regression or even complete remission. In four cases, a stable or increasing clonal CD4+Vbeta+ population reflected well a stable or progressing course of the disease. Double staining of Vbeta+ cells revealed the following pattern, CD3+, CD5+, CD7+, CD28+, CD80-, CD86+ and human leucocyte antigen (HLA) class I+. In contrast, HLA-DR was heterogeneously expressed. We conclude that identification and monitoring of CD4+Vbeta+ clonal T cells by fluorescence-activated cell sorting with double staining is a suitable method to assess clinical responses to different therapies.