Mycosis Fungoides Patients Research Articles

Expression of Fas and Fas-ligand in primary cutaneous T-cell lymphoma (CTCL): association between lack of Fas expression and aggressive types of CTCL.

Fas (CD95; APO-1) is a transmembrane protein that mediates apoptosis upon cross-linking with Fas-ligand (Fas-L). Interaction of Fas-L expressed by cytotoxic T cells with Fas-expressing tumour cells plays an important part in antitumour immune responses. We aimed to investigate Fas and Fas-L expression in frozen and paraffin-embedded material from a large group of patients with cutaneous T-cell lymphoma (CTCL). Immunostaining with monoclonal antibodies against Fas and Fas-L was performed in material from 23 patients with mycosis fungoides (MF), 10 with lymphomatoid papulosis (LyP), 10 with CD30-positive primary cutaneous large T-cell lymphoma (LTCL) and nine with CD30-negative LTCL. The results were correlated with the type and stage of CTCL and clinical features. Expression of Fas by the large majority of the neoplastic T cells was observed in 15 of 15 cases of plaque-stage MF, 10 of 10 cases of LyP and 10 of 10 cases of CD30-positive LTCL, but only in four of 12 cases of tumour-stage MF and two of nine cases of CD30-negative LTCL. In three of four MF patients in whom both plaques and tumours could be studied, a significant decrease in Fas expression was observed with progression from plaque-stage to tumour-stage disease. Fas-L was expressed by > 50% of the neoplastic T cells in 46 of 56 biopsies, and no clear relationship with type of CTCL and clinical behaviour was observed. This study demonstrates loss of Fas expression in aggressive types of CTCL, but not in indolent types of CTCL. These data suggest that loss of Fas receptor expression may be one of the mechanisms that allow tumour cells to escape an effective immune response, and may contribute to the unfavourable prognosis of some types of CTCL.

Transmission of human T-cell lymphotropic virus type 1 tax to rabbits by tax-only-positive human cells.

The human T-cell lymphrotropic virus type 1 (HTLV-1) is causally related to adult T-cell leukemia and lymphoma and the neurodegenerative diseases tropical spastic paraparesis and HTLV-1-associated myelopathy. In the United States the prevalence of infection has been estimated to range from 0.016 to 0.1% on the basis of serologic tests for antibodies to the viral structural proteins. Blood from donors positive for antibodies to HTLV-1 or HTLV-2 is not used for transfusion. However, patients with the cutaneous T-cell lymphoma mycosis fungoides (MF) are HTLV-1 and -2 seronegative yet harbor proviral sequences identical to those that encode the HTLV-1 transactivating and transforming gene product p40tax in their peripheral blood mononuclear cells (PBMCs), and they usually have antibodies to p40(tax). Moreover, a study of 250 randomly selected blood donors revealed that approximately 8% of these seronegative individuals also had HTLV-1 tax sequences and antibodies to p40(tax), while they lacked sequences and antibodies related to gag, pol, or env. Thus, it seemed important to determine whether the "tax-only" state can be transmitted by transfusion. To this end, PBMCs from HTLV-1 and -2 seronegative tax-only-positive MF patients or from healthy tax-only-positive blood donors were injected into adult rabbits, an established animal model for HTLV-1 infection. The PBMCs of all injected rabbits became tax sequence positive. These observations suggest that HTLV-1 tax can be transmitted by tax-only-positive mononuclear cells.

Photopheresis in Cutaneous T-Cell Lymphoma: Five-Year Experience

Cutaneous T-cell lymphoma (CTCL) includes several lymphoproliferative disorders involving mature T-lymphocyte proliferation initially confined to the cutis. These affections, after variable periods, may progress to the blood, limph nodes and visceral organs. Mycosis fungoides (MF) is the most frequent form of CTCL and has an indolent clinical course. The therapy of CTCL depends on the stage of the disease and the patient's general conditions. For advanced cases it includes chemotherapy, retinoids, and interferon-alpha. Since 1987 extracorporeal photochemotherapy (ECP), a novel immunomodulatory approach based on apheresis and photoirradiation of leukocytes, has been successfully introduced for the treatment of advanced CTCL. It can prolong survival of patients with erythrodermic CTCL without significant side effects. To review our five-year experience with ECP in CTCL. Since June 1994, 33 CTCL patients have been recruited for ECP, using two different regimens: two procedures on two consecutive days at four-week intervals for six months, or at two-week intervals for three months with progressive tapering in the second three-month period for the more severe forms. Six patients received ECP with IFN-alpha. ECP was done using the photopheresis UVAR system and UVAR XTS (Therakos, West Chester, Pa) and always with 8-MOP liquid formulation injected directly into the buffy coat bag. Lymphocytes in peripheral blood were immunophenotypically characterized for each patient and every ECP session. All patients tolerated ECP well, without significant side effects. Thirty patients are clinically evaluable (at least three ECP cycles). A favourable clinical response was obtained in 80.9% (16/21) of MF patients (complete response 33%, partial response 47.6%) and in 66% (6/9) of patients in the Sézary's syndrome phase (complete response 33.3%, partial response 33.3%). Five of the six patients given IFN-alpha as adjunctive therapy had a PR and one a CR. Four patients are in CR without therapy at follow-ups of 46, 20, 10 and 8 months. There have been no changes in the peripheral lymphocyte immunophenotype during the follow-up. In 19/30 patients the CD95 antigen, correlated with cellular apoptosis, was expressed and was frequently associated with a good clinical response. In our experience ECP achieved favourable clinical responses in 73% of patients, in monotherapy or in combination with IFN-alpha, without significant side effects.

Evidence for the cofactor role of human T-cell lymphotropic virus type 1 in mycosis fungoides and Sézary syndrome.

The aetiology of mycosis fungoides (MF) and Sézary syndrome (SS) is unknown. A pathogenic role for the human T-cell lymphotropic virus type 1 (HTLV-1) has been suggested but remains controversial. We used an animal model to test the possibility that peripheral blood mononuclear cells (PBMC) obtained from MF patients harbour the HTLV-1 virus which may be infective. The polymerase chain reaction (PCR) was used to detect HTLV-1 proviral DNA sequences in PBMC of 27 MF patients and one SS patient of non-Iranian origin. Positive results were found in six of the patients. Twelve of the 28 patients tested by Western blot showed HTLV-1 antibodies. Twenty-eight immunosuppressed inbred Fisher F344 rats were inoculated intravenously with cultures of PBMC obtained from the 28 patients. Eight of these 28 rats showed antibodies to HTLV-1 while the proviral genome was demonstrated in the blood of only two of the rats. PBMC from two MF patients, in spite of showing negative results for the proviral genome by PCR, still induced HTLV-1 antibody formation in the F344 rat model. None of 10 control rats inoculated with normal donor PBMC showed antibodies to HTLV-1, nor the proviral genome. The present study suggests that HTLV-1 plays a cofactor role in MF/SS patients.

The epidermotropic mycosis fungoides associated alpha1beta1 integrin (VLA-1, CD49a/CD29) is primarily a collagen IV receptor on malignant T cells.

Several of the beta1 integrin receptors [very late antigen (VLA) molecules] for extracellular matrix (ECM) proteins are expressed by malignant T cells in cutaneous T-cell lymphoma (CTCL). We evaluated the function of VLA-1, a beta1 integrin specifically expressed in epidermotropic mycosis fungoides (MF), in CD4+ leukemic T cells Jurkat line). We found that Jurkat cells adhere significantly to collagens only after their activation with phorbol 12-myristate 13-acetate (PMA). However, the adhesion to collagen IV (but not to collagen I) of Jurkat cells selected for expressing increased levels of VLA-1 (with unchanged levels of VLA-2, the second collagen integrin receptor) was significantly enhanced relative to that of "VLA-1 low" cells. Monoclonal antibody (mAb) 1B3.1, directed against the collagen binding domain of VLA-1, inhibited adhesion to collagen IV and to collagen I by 36.67%+/-5.25% and 18%+/-4.32%, respectively (p<0.05), whereas the inhibition by anti-VLA-2 mAb PIE6 was comparable on both collagens (25%+/-7.48% and 36.3%+/-0.94%, respectively; p<0.09). Immuno-histochemical studies of skin biopsies from 10 untreated MF patients showed that in all cases at least 10% of the lymphocytes residing in the epidermis are VLA-1+VLA-2-. While not directly applicable to MF, the demonstrated functions of VLA-1 in leukemic Jurkat cells, together with its expression in MF skin, suggest a role for VLA-1 integrins in epidermotropism in a small proportion of leukemic MF cells.

The changing survival of patients with mycosis fungoides

The authors sought to evaluate trends in survival among a population-based group of patients with mycosis fungoides, in response to informal evidence of improved prognosis and concerns about the influence of detection bias on incidence data. Data used in this study were drawn from the Surveillance, Epidemiology, and End Results Program of the National Cancer Institute, which includes 9 population-based cancer registries. Together, these registries cover approximately 10% of the U.S. population and have tracked the survival of all mycosis fungoides patients who were registered during the years 1973-1992. Data included follow-up through 1994. There was a total of over 10,000 person-years of follow-up for the 1633 patients studied. Relative survival changed little after 11 years, at which point it was 66%. Advanced age and black race were associated with poorer survival. The prognoses of Asian and Hispanic patients were slightly but not significantly worse than those of whites, and there were no significant geographic differences related to prognosis. The survival of married women was superior to that of other gender and marital-status groups. Prognosis did improve substantially over the 20-year period of study. Mycosis fungoides is usually not a fatal disease, although the mortality risk is substantial. The authors quantified various prognostic factors and documented an improved prognosis over the 20-year period of study. The data from this study raise concern about possible detection bias in incidence data. These data are also consistent with the concept that beyond 11 years after diagnosis, this disorder has relatively little impact on the risk of death.

The changing survival of patients with mycosis fungoides

BACKGROUND The authors sought to evaluate trends in survival among a population-based group of patients with mycosis fungoides, in response to informal evidence of improved prognosis and concerns about the influence of detection bias on incidence data. METHODS Data used in this study were drawn from the Surveillance, Epidemiology, and End Results Program of the National Cancer Institute, which includes 9 population-based cancer registries. Together, these registries cover approximately 10% of the U.S. population and have tracked the survival of all mycosis fungoides patients who were registered during the years 1973–1992. Data included follow-up through 1994. RESULTS There was a total of over 10,000 person-years of follow-up for the 1633 patients studied. Relative survival changed little after 11 years, at which point it was 66%. Advanced age and black race were associated with poorer survival. The prognoses of Asian and Hispanic patients were slightly but not significantly worse than those of whites, and there were no significant geographic differences related to prognosis. The survival of married women was superior to that of other gender and marital-status groups. Prognosis did improve substantially over the 20-year period of study. CONCLUSIONS Mycosis fungoides is usually not a fatal disease, although the mortality risk is substantial. The authors quantified various prognostic factors and documented an improved prognosis over the 20-year period of study. The data from this study raise concern about possible detection bias in incidence data. These data are also consistent with the concept that beyond 11 years after diagnosis, this disorder has relatively little impact on the risk of death. Cancer 1999;85:208–12. © 1999 American Cancer Society.

Therapy with gemcitabine in pretreated peripheral T-cell lymphoma patients

Gemcitabine is a novel nucleoside analogue which has shown promising results in most solid tumors; like the arabinosylcytosine analogue, gemcitabine may be an active drug in lymphoproliferative malignancies. We tested it in pretreated peripheral T-cell lymphoma patients with isolated skin involvement. We performed a phase II study with the drug in 13 pretreated patients with peripheral T-cell lymphoma, five of whom had advanced-stage mycosis fungoides (MF), and eight peripheral T-cell lymphoma unspecified (PTCLU). Patients were treated on days 1, 8, and 15 of a 28-day schedule at the dosage of 1200 mg/m2 for a total of three courses. Of the 13 patients, one achieved complete response (CR) and eight achieved partial responses (PR); the remaining four showed no benefit from the treatment. Among the responders, one CR and four PR were documented in the PTCLU patients and four PR in MF patients. Treatment was well tolerated; hematologic toxicity was mild and no nausea/vomiting or organ toxicity was recorded. In view of its significant activity and its modest toxicity profile, the role of gemcitabine deserves further evaluation in the management of pretreated patients with peripheral T-cell lymphoma.

The small-cell variant of mycosis fungoides. A clinicopathological and quantitative electron microscopic study on 14 patients.

Small-cell variants of Sézary syndrome and mycosis fungoides (MF) have been described. However, in these studies the nuclear area of the small-cell variant of MF (SC-MF) as compared to histological classical MF (CL-MF) was not characterized objectively by quantitative electron microscopy. In a 14-year follow-up period, of a total of 76 patch/plaque stage MF patients seen in the Department of Dermatology of the University Hospital Utrecht, 14 (18%) had an infiltrate composed of atypical lymphocytes characterized by a distinctly smaller cell diameter and smaller, hyperchromatic, deeply indented nuclei as compared to the usual cell type of MF. The aim of the investigation was to confirm this observation objectively using quantitative electron microscopy (morphometry) and to define SC-MF as compared to CL-MF. The study was performed on the 14 patients with SC-MF, and 10 patients with clinical and histological CL-MF and 4 patients with chronic eczema. Electron micrographs of sections obtained from each biopsy were analysed by computer to produce the following data: a nuclear contour index (NCI), the mean nuclear area (MNA), the mean nuclear area of the cells above the 75th percentile (P75NA) and the percentage of cells larger than 30 microm2. The values of MNA differed significantly between patients with SC-MF and those with CL-MF (17.6 vs 23.2 microm2; P = 0.02), as did the values of P75NA (20.7 vs 27.9 microm2; P = 0.01). The NCI of the SC-MF and CL-MF patients were similar. These results are consistent with our observations that SC-MF does indeed exist.

Prognostic Significance of a Polymerase Chain Reaction–Detectable Dominant T-Lymphocyte Clone in Cutaneous Lesions of Patients With Mycosis Fungoides

Although mycosis fungoides (MF) is considered to be an indolent lymphoma, survival is highly influenced by TNM stage. At diagnosis, most MF patients present with early stage disease and a high probability of long-term survival. Treatment is generally directed towards skin lesions, and achievement and duration of complete responses are variable. A dominant T-cell clone is detectable in the cutaneous lesions of 60% of patients. The aim of this study was to determine whether the presence of a T-cell clonal population influences the clinical course of the disease after topical therapy. Cutaneous biopsies from 68 patients were histologically diagnosed as MF and T-cell clonality was analyzed by in vitro amplification of TCR-gamma chain gene rearrangements (polymerase chain reaction gamma [PCRgamma]). After a median follow-up of 48 months, response to treatment was clinically assessed. Age, sex, duration of symptoms before diagnosis, type of cutaneous lesions (T stage), TNM stage, and PCRgamma were evaluated as predictive factors of response to treatment in univariate and multivariate analyses. Univariate analysis demonstrated that T1 cutaneous lesions (P = .05) and PCRgamma negativity (P = .007) were associated with a higher complete remission rate. Using multivariate analysis, T stage (relative risk, 3.13; P = .06) and PCRgamma (relative risk, 4.4; P = .01) remained independent significant predictive parameters of response. In conclusion, T stage and cutaneous PCRgamma at diagnosis are the two predictive parameters of treatment response for MF. Therefore, the cutaneous PCRgamma findings should be considered in the analysis of future therapeutic trials.

Prognostic Significance of a Polymerase Chain Reaction–Detectable Dominant T-Lymphocyte Clone in Cutaneous Lesions of Patients With Mycosis Fungoides

AbstractAlthough mycosis fungoides (MF) is considered to be an indolent lymphoma, survival is highly influenced by TNM stage. At diagnosis, most MF patients present with early stage disease and a high probability of long-term survival. Treatment is generally directed towards skin lesions, and achievement and duration of complete responses are variable. A dominant T-cell clone is detectable in the cutaneous lesions of 60% of patients. The aim of this study was to determine whether the presence of a T-cell clonal population influences the clinical course of the disease after topical therapy. Cutaneous biopsies from 68 patients were histologically diagnosed as MF and T-cell clonality was analyzed by in vitro amplification of TCR-γ chain gene rearrangements (polymerase chain reaction γ [PCRγ]). After a median follow-up of 48 months, response to treatment was clinically assessed. Age, sex, duration of symptoms before diagnosis, type of cutaneous lesions (T stage), TNM stage, and PCRγ were evaluated as predictive factors of response to treatment in univariate and multivariate analyses. Univariate analysis demonstrated that T1 cutaneous lesions (P = .05) and PCRγ negativity (P = .007) were associated with a higher complete remission rate. Using multivariate analysis, T stage (relative risk, 3.13; P = .06) and PCRγ (relative risk, 4.4; P = .01) remained independent significant predictive parameters of response. In conclusion, T stage and cutaneous PCRγ at diagnosis are the two predictive parameters of treatment response for MF. Therefore, the cutaneous PCRγ findings should be considered in the analysis of future therapeutic trials.© 1998 by The American Society of Hematology.

Detection of monoclonal T-cells with TCR gamma PCR in mycosis fungoides

The monoclonal dominant malignant T cells in mycosis fungoides (MF) carry identical TCR gamma rearrangements. Their detection is a useful diagnostic tool. Thus, in routine diagnosis we investigated the occurrence of monoclonal T cells in skin biopsy samples of MF-patients by TCR gamma-PCR, followed by temperature gradient gel electrophoresis (TGGE). In 188 out of 208 MF patients, at least one skin sample with sufficient DNA quality for PCR analysis was obtained. Applying a consensus PCR for the TCR gamma genes V gamma I and J gamma 1/2, we detected monoclonal T cells in 122 cases (65%). In the remaining 66 cases, we performed two multiplex-PCRs, covering rearrangements of the other TCR gamma genes. Here we found in 11 cases (6%) predominant clonal rearrangements of V gamma II-IV and J gamma 1/2 and in 2 (1%) those of V gamma I-IV and J gamma P 1/2. In patients with MF, detecting rearrangements of only V gamma I and J gamma 1/2 is sufficient for PCR screening analysis. In 53 of the patients (28%) the applied methods revealed no monoclonal T cells. This may be due to a low cell number, oligoclonal nature, chromosomal abberations or remaining of TCR in germline configuration.

Interleukin-15 Is an Autocrine/Paracrine Viability Factor for Cutaneous T-Cell Lymphoma Cells

In this study we investigated the role of interleukin-15 (IL-15) in the immunobiology of cutaneous T-cell lymphoma (CTCL) cells. Using cell culture techniques, reverse transcriptase-polymerase chain reaction (RT-PCR), and immunhistochemistry we found that IL-15, like IL-7, is a growth factor for the Sézary cell line SeAx and that both cytokines prolonged the survival of malignant T cells directly isolated from Sézary syndrome (SS) patients. Both IL-15 and IL-7 were more potent than IL-2. IL-4 and IL-9, whose receptors share the same gamma chain with the receptors of IL-2, IL-7, and IL-15, did not sustain the growth of CTCL cells, indicating that signaling through the common gamma chain (gammac) is not sufficient for continuous growth. IL-13 and tumor necrosis factor-alpha (TNF-alpha) had no effect. IL-7 and IL-15 also supported the growth of SeAx cells in the presence of the apoptosis inducing agents dexamethasone and retinoic acid. The analysis of patient Sézary cells and three CTCL cell lines by RT-PCR showed that all these cells expressed IL-15 mRNA, but only a few (25%) produced IL-7 mRNA. Immunohistological analyses of skin biopsy samples of SS and Mycosis fungoides patients showed immunoreactivity for IL-15 in basal cell layer keratinocytes and in the infiltrating lymphocytes. We conclude that IL-15 is a growth or viability factor for CTCL-derived cell lines or shortly cultivated Sézary cells. The findings that IL-15 mRNA can be detected in Sézary syndrome peripheral blood mononuclear cells and that the IL-15 protein is detected in skin sections from CTCL patients suggest that IL-15 plays an important role in the biology of CTCL.

Spontaneous regression of granulomatous mycosis fungoides in an HIV positive patient

The development of mycosis fungoides in HIV-positive patients is uncommon. We describe the granulomatous type of mycosis fungoides in an HIV-positive patient whose cutaneous lesions regressed without treatment. In addition to HIV seropositivity and mycosis fungoides, he had a long history of pulmonary sarcoidosis. Immunophenotyping of the cutaneous lymphoid infiltrates revealed a predominance of CD4 + T cells, and monoclonality was demonstrated by T-cell gene rearrangement studies. The regression of mycosis fungoides in this patient was associated with a falling peripheral blood CD4 T-cell count. (J Am Acad Dermatol 1997:37:876-80.)

Treatment of cutaneous T‐cell lymphoma with extracorporeal photochemotherapy

AbstractBackground Cutaneous T‐cell lymphoma (CTCL) and its leukemic erythrodermic form (Sézary syndrome) are malignancies cies of CD4+ T lymphocytes. Extracorporeal photochemotherapy (ECP) selectively affects autoreactive as well as malignant T lymphocytes. The efficacy of ECP depends strongly upon adequate serum/buffy coat levels of the photosensitizer 8‐methoxypsoralen (8‐MOP). The resorption of orally applied 8‐MOP vanes inter‐ and intraindividually within a broad range, meaning that adequate therapeutic drug levels cannot always he achieved. Therefore, since July 1994 we have exclusively used a liquid 8‐MOP preparation which is added directly into the buffy coat fraction of the ECP circuit, resulting in constant high drug levels of approximately 190 ng/ml. Twelve CTCL‐patients (six with Sézary syndrome, six with mycosis fungoides received between six and 25 ECP treatments. Some of them had undergone previous therapy without success.Results All patients with Sézary syndrome declared a distinct reduction in intensity of pruritus. Three patients who received liquid 8‐MOP extracorporeally showed a partial remission on the basis of skin scores. Of the patients receiving 8‐MOP orally, two remained clinically unchanged and one showed a progression of the disease. In these cases, subtherapeutic 8‐MOP plasma levels were often found. Of the six mycosis fungoides patients one achieved complete and two partial remission: another two patients showed minor response. These five patients were treated with liquid 8‐MOP. One patient showed no change in skin lesions; he had received 8‐MOP orally and achieved subtherapeutic photosensitizer plasma levels.Conclusion Our treatment protocols confirm the beneficial effects of ECP on CTCL at any stage, but it seems that adequate ECP efficiency is ensured only when an 8‐MOP solution is applied extracorporeally.

Treatment of cutaneous T-cell lymphoma with extracorporeal photochemotherapy

Background Cutaneous T-cell lymphoma (CTCL) and its leukemic erythrodermic form (Sézary syndrome) are malignancies of CD4+ T lymphocytes. Extracorporeal photochemotherapy (ECP) selectively affects autoreactive as well as malignant T lymphocytes. The efficacy of ECP depends strongly upon adequate serum/buffy coat levels of the photosensitizer 8-methoxypsoralen (8-MOP). The resorption of orally applied 8-MOP varies inter- and intraindividually within a broad range, meaning that adequate therapeutic drug levels cannot always be achieved. Therefore, since July 1994 we have exclusively used a liquid 8-MOP preparation which is added directly into the buffy coat fraction of the ECP circuit, resulting in constant high drug levels of approximately 190 ng/ml. Twelve CTCL-patients (six with Sézary syndrome, six with mycosis fungoides) received between six and 25 ECP treatments. Some of them had undergone previous therapy without success.Results All patients with Sézary syndrome declared a distinct reduction in intensity of pruritus. Three patients who received liquid 8-MOP extracorporeally showed a partial remission on the basis of skin scores. Of the patients receiving 8-MOP orally, two remained clinically unchanged and one showed a progression of the disease. In these cases, subtherapeutic 8-MOP plasma levels were often found. Of the six mycosis fungoides patients one achieved complete and two partial remission; another two patients showed minor response. These five patients were treated with liquid 8-MOP. One patient showed no change in skin lesions; he had received 8-MOP orally and achieved subtherapeutic photosensitizer plasma levels.Conclusion Our treatment protocols confirm the beneficial effects of ECP on CTCL at any stage, but it seems that adequate ECP efficiency is ensured only when an 8-MOP solution is applied extracorporeally.

Immunohistochemical expression of IL-10 in mycosis fungoides.

Immunoreactivity of the cytokine IL-10 has been investigated in situ in mycosis fungoides (MF). Expression of IL-10 was detected using immunohistochemistry in skin biopsies (n = 8) and T-cell lines (n = 2) from mycosis fungoides patients. IL-10 positivity was seen in the dermal cell infiltrates and in T-cell lines in mycosis fungoides. The dermal IL-10 reaction in a skin biopsy from an active lesion in MF indicates the possibility for disease progression.

Constitutive activation of a slowly migrating isoform of Stat3 in mycosis fungoides: tyrphostin AG490 inhibits Stat3 activation and growth of mycosis fungoides tumor cell lines.

Mycosis fungoides (MF) is a low-grade cutaneous T cell lymphoma of unknown etiology. In this report, the Jak/Stat (Janus kinase/signal transducer and activator of transcription) signaling pathway was investigated in tumor cell lines established from skin biopsy specimens from a patient with MF. Jaks link cytokine receptors to Stats, and abnormal Jak/Stat signaling has been observed in some hemopoietic cancers. In MF tumor cells, a slowly migrating isoform of Stat3, Stat3(sm), was found to be constitutively activated, i.e., (i) Stat3(sm) was constitutively phosphorylated on tyrosine residues, and tyrosine phosphorylation was not enhanced by growth factor stimulation; (ii) band shift assays and immunoprecipitations of DNA/Stat complexes showed constitutive DNA-binding properties of Stat3(sm); and (iii) Stat3(sm) was constitutively associated with Jak3. The abnormal activation of Stat3(sm) was highly specific. Thus, neither the fast migrating isoform of Stat3 (Stat3(fm)) nor other Stats (Stat1, Stat2, and Stat4 through Stat6) were constitutively activated. The Jak kinase inhibitor, tyrphostin AG490, blocked the constitutive activation of Stat3(sm) and inhibited spontaneous as well as interleukin 2-induced growth of MF tumor cells. In conclusion, we have provided evidence for an abnormal Jak/Stat signaling and growth regulation in tumor cells obtained from affected skin of an MF patient.

Reexamination of human T cell lymphotropic virus (HTLV-I/II) prevalence.

In the United States, blood donors are being screened for infection with human T cell lymphotropic viruses I and II (HTLV-I/II) by serologic means, which detect antibodies to the structural proteins of these viruses. Because patients with mycosis fungoides (MF) usually do not have such antibodies even though their cells harbor HTLV-I Tax and/or pol proviral sequences, it was questioned whether the prevalence of HTLV infection among healthy blood donors may also be underestimated by current means of testing. To examine this possibility, a study on specimens of relatives of mycosis fungoides patients (MFR) was begun. In addition, to collect data more expeditiously, a cohort of former injection drug users (IDUs) was tested by routine serologic methods, as well as by PCR/Southern blot analysis for Tax, pol, and gag proviral sequences and Western blot analysis for antibodies to the Tax gene product. To date, 6/8 MFRs and 42/81 (51.8%) of HIV-negative IDUs proved to be positive for HTLV, whereas routine serology identified none of the MFR and only 18/81 (22.2%) of the IDUs. Among the latter test subjects, the incidence of HTLV-I also proved to be 10 times higher than expected. Therefore, it is likely that among healthy blood donors infection with HTLV-I/II is more prevalent than is currently assumed. Since Tax is the transforming sequence of HTLV-I/II, testing for Tax sequences and antibodies to its gene product may be desirable in blood transfusion and tissue donor facilities.

The difficulty of detecting HTLV-1 proviral sequences in patients with mycosis fungoides.

Although most patients with cutaneous T cell lymphomas, including mycosis fungoides (MF) and its leukemic variant, the Sézary syndrome, are seronegative for antibodies to the human T cell lymphotropic viruses (HTLV-I/II), it has recently been shown that > 95% of such patients harbor proviral DNA sequences related to the region of the HTLV genome that encodes the transregulatory/transforming gene, tax. However, the demonstration of HTLV sequences, even after amplification by polymerase chain reaction (PCR), has not been universally successful, and some investigators continue to question this observation. In an effort to resolve this controversy, we have compared published methodologies that have been less successful with techniques currently used in this laboratory. Major differences were found in (a) the nature of the cells used [freshly isolated versus cultured peripheral blood mononuclear cells (PBMC)] and (b) the methods used to prepare samples for PCR (whole cell lysates versus DNA extracts). PBMC from 10 different MF patients and the healthy daughter of 1 of the patients were subjected to comparative analyses. While all of the PBMC lysates were positive, the DNA extract from only one of these individuals revealed HTLV tax sequences. Studies were also conducted comparing cell lysates and DNA extracts of cultured cells derived from tax sequence-positive PBMC from seven different MF patients. The cells from four of the seven were shown to have retained tax sequences after varying times in culture, when whole-cell lysates were used as targets for PCR amplification and Southern analysis, whereas none of the DNA extracts were positive. It appears that the use of whole-cell lysates instead of DNA extracts and the use of fresh instead of cultured cells greatly enhance the ability to detect HTLV-1 tax sequences in specimens from MF patients.