Mycosis Fungoides. Treatment of advanced mycosis fungoides by allogeneic stem-cell transplantation with a nonmyeloablative regimen. D Soligo1, A Ibatici1, ...
Bone Marrow Transplantation (2003) 31, 663–666 & 2003 Nature Publishing Group All rights reserved 0268-3369/03 $25.00
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Mycosis Fungoides Treatment of advanced mycosis fungoides by allogeneic stem-cell transplantation with a nonmyeloablative regimen D Soligo1, A Ibatici1, E Berti2, P Morandi1, E Longhi3, L Venegoni2, M Corbellino4, C Annaloro1, L Robbiolo1, A Della Volpe1, E Alessi2 and G Lambertenghi Deliliers1 1 Department of Hematology, IRCCS Ospedale Maggiore Milano, Italy; 2Department of Dermatology, IRCCS Ospedale Maggiore Milano, Italy; 3Transplantation Immunology and Blood Transfusion Services, IRCCS Ospedale Maggiore Milano, Italy; and 4 Institute of Infectious and Tropical Diseases, University of Milano, Milano, Italy
Summary: Given the poor prognosis of patients with advanced cutaneous T-cell lymphoma and the high transplantrelated mortality associated with conventional allogeneic bone marrow transplantation, we performed nonmyeloablative transplantation of allogeneic stem cells (ASCT) from HLA-identical siblings in three patients with this disease. All patients achieved full donor engraftment, clearance of clonal T cells leading to durable complete remissions but experienced high incidence of infections, which proved fatal in one case. These results suggest that nonmyeloablative ASCT is a novel and potentially curative therapy for patients with advanced T-cell lymphomas who have a histocompatible sibling. Bone Marrow Transplantation (2003) 31, 663–666. doi:10.1038/sj.bmt.1703872 Keywords: mycosis fungoides; nonmyeloablative regimen; allogeneic stem cell transplantation
aged 58 years or more,3 who are generally not eligible for more aggressive therapies. For patients with advanced MF, single-agent or combination chemotherapy does not show any survival advantage. Autologous bone marrow transplantation (BMT) has been used in a few patients with advanced MF, but the short duration of clinical response induced investigators to delay larger perspective studies.4–6 In contrast, conventional allogeneic stem cells (ASCT) has been described thus far in few patients with MF with evidence of a graft-versustumor (GVT) effect.7–9 The poor prognosis of elderly patients with advanced or refractory MF and the high transplant-related mortality after conventional ASCT led us to investigate the role of HLA-identical sibling transplantation with a nonmyeloablative regimen.
Patients and methods Patient selection
Introduction Mycosis fungoides (MF) is the most common primary lymphoma involving the skin. Histopathological features are characterized by marked epidermotropism of T lymphocytes with convoluted pleomorphic nuclei. The malignant cells express a mature memory T-helper phenotype (CD45RO+CD4+) and their clonality emerges from Tcell receptor TCR b or g gene rearrangements by molecular analyses.1,2 The natural history of MF is usually indolent, but some patients eventually progress to more advanced stages with an aggressive clinical course and shortened survival.1,2 In a large series of patients, the median survival of all stage IV patients was 13 months from the date of first treatment;3 even shorter survivals (1 year) were observed in patients
Correspondence: Dr D Soligo, Centro Trapianti di Midollo, IRCCS Ospedale Maggiore Milano, Via F. Sforza 35, 20122 Milano, Italy Received 20 May 2002; accepted 14 October 2002
All patients had a diagnosis compatible with MF, as confirmed by clinical evaluation, histopathology, immunohistochemistry and molecular biology. Major eligibility criteria for this study were: (1) An HLA-identical sibling donor; (2) MF from stage IIIB to IVB according to the Mycosis Fungoides Cooperative Group staging criteria; (3) relapse after at least two lines of therapy; and (4) Karnovsky score more than 70.
Transplantation procedure The conditioning regimen consisted of two cycles of fludarabine 30 mg/m2/day for 3 days and cyclophosphamide 300 mg/m2/day for 3 days (F–C) repeated every 28 days, followed by 200 cGy TBI (day 0) delivered at 7 cGy/ min 14 days after the second F–C cycle. One case (case 1) received fludarabine 30 mg/m2 on day 4, 3, 2 as the preparative regimen, instead of the F–C cycles. GVHD prophylaxis included oral cyclosporine-A (3 mg/kg/day, twice daily) from day 0 to +100 (targeted serum level of 400–600 ng/ml) and oral mycophenolate mofetil acid (MMF) (5 mg/kg/day, twice daily) from day 0 to +27, then tapered within 2 weeks. Patients were infused with
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G-CSF-mobilized donor peripheral blood stem cells (targeted dose of 5 106 CD34+ cells).
Immunohistochemical and TCR-g rearrangement studies Frozen sections of each biopsy specimen were labeled with a panel of monoclonal antibodies including CD3,7,4,8,30 and immunoreactivity revealed using a sensitive alkaline phosphatase antialkalinephosphatase method.10 The immunostaining of the frozen sections and the peripheral blood confirmed in all cases the ‘classical’ CTCL immunophenotype CD3+, CD4+, CD45RO+, CD29+ and CD8 . Clonality of skin infiltrating T cells was assessed by means of heteroduplex analysis of the TCRg VJ junctions, separated in nondenaturating polyacrylamide gel as previously described.11
Assessment of engraftment Donor–recipient chimerism was monitored on genomic DNA isolated (QIAamp 96 Spin Blood Kit, QIAGEN) from immunomagnetically sorted peripheral blood CD8+ T-cells (Mini-MACS, Miltenyi Biotec) since the recommended selection for CD3+T-cells in nonmyeloablative allografts might have included a high number of circulating MF cells. DNA samples were prepared for PCR according to AmpF_Profiler Plus kit instructions (Applied Biosystems, Foster City, CA, USA). Nine STR loci (D3S1358, vWA, FGA, D8S1179, D21S11, D18S51, D5S818, D13S317, D7S820) and a segment of the X-Y homologous gene coding for amelogenin was amplified and electrophoresed on a capillary gel (ABI PRISM 310, Applied Biosystems). Each electrophoretic run was analyzed with GeneScan Analysis software (Applied Biosystems). Alleles peaks detected in the post-transplant samples were compared with the patient pretransplant and donor samples to determine if the amplified alleles contained DNA from the recipient, donor or both. Calculation of mixed chimerism was performed according to Pindolia et al.12
2 months following ASCT cyclosporine-A was gradually tapered: at this time, the patient was still erythrodermic (Figure 1a) with enlarged lymph nodes. Skin biopsy showed a clonal infiltrate (TCR-g +), with the same characteristics of the rearrangements observed before BMT both in the skin, lymph node and in the peripheral blood, compatible with the persistence of MF. (Figures 1e and 2). Acute graftversus-host-disease (GVDH) (skin grade III) ensued, which was accompanied by rapid regression of lymphomatous infiltrates in all involved sites (Figure 1b). A few days after reintroduction of cyclosporine-A, the skin GVHD improved and completely resolved. By day +180 immunosuppression was again suspended, with no further evidence of both GVHD and MF (Figures 1b and f) and PCR analysis was unable to detect clonal T cells both in peripheral blood and skin. At 24 months of follow-up, the patient is in complete remission (TCRg-), with donor-type full chimerism, and very well (performance status of 100%). Case 2: A 47-year-old man was diagnosed with erythrodermic MF (stage IIIA) in 1996. He received EP and
Case reports Patients Case 1: A 56-year-old male was diagnosed with patchstage MF in 1996, which progressed to erythrodermic MF in 1997 and was treated with PUVA, a-interferon, steroids, extracorporeal photophoresis (EP), methotrexate (MTX) and alkylating agents. In May 2000, the patient developed overt leukemic progression and diffuse lymph-node enlargement (diagnosed as SS)13 and he was started on monthly fludarabine at conventional doses. Following the second fludarabine dose, he developed aggressive Epstein– Barr virus (EBV)-related lymphoproliferative disorder (LPD) that resolved after 3 weekly doses of rituximab (375 mg/m2). As the signs of progressive MF persisted, the patient underwent ASCT following nonmyeloablative conditioning. DNA-microsatellite studies showed full donor chimerism in CD8+/CD3+ PBL by day +30. At Bone Marrow Transplantation
Figure 1 Extensive erythematous patch lesions persist on day +60 posttransplant (a) but completely disappear upon discontinuation of immunosuppression (b) (case 1) Extensive hyperkeratotic scaling and fissuring lesions involving both hands (c) are completely resolved on day +90 after the transplant (d). (case 2). Day +60 skin biopsy showing a typical bandlike epidermotropic infiltrate with plaques invading the superficial dermis (e); on day +180 the biopsy shows an apparently normal skin: acantotic and hyperkeratotic epidermis with an irregular outline and absence of lymphomatous infiltrate (f) (case 1).
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obtained a short remission after six cycles of CHOP chemotherapy. In March 2001, upon progression of MF to stage IVA, he entered our transplant protocol. During the pretransplant phase, he had an episode of pulmonary embolism and SA sepsis, both of which resolved after treatment. By day +28 the patient developed grade III acute GVHD that improved with corticosteroid therapy (starting dose of 1 mg/kg/day). Skin biopsy confirmed the clinical diagnosis of GVHD, but demonstrated disappearance of MF (Figure 2). Human cytomegalovirus (HCMV) reactivation and BK-virus-related hemorrhagic cystitis complicated the post-transplant clinical course. The patient died on day +73 with SA sepsis and acute myocarditis. At that time there were no signs of liver or intestinal GVHD but only grade II cutaneous GVHD; the underlying disease was in CR, with full donor-type chimerism.
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Discussion
Figure 2 PCR analysis of the patient’s T-cell receptor g-chain gene rearrangement in the involved skin (or in peripheral blood in case 1) and on day +90 after the transplant. Lane M: Molecular Weight Marker V from Boehringer Manheim; lanes 4, 8 and 12: patients with known DNA TVCRg-chain gene rearrangement used as positive controls; lanes 5, 9 and 13: skin biopsies from healthy donors used as negative controls; lanes 1, 2, 3: PCR products obtained with primers for V9-J1-2 junctional sequences (pt1); lanes 6,7: V2a-J1-2 junctional sequences (pt2); lanes 10 and 11: V10/ 11-J1-2 junctional sequences (pt3); lane B: water blank.
corticosteroids until September 2000, when he presented with diffuse skin erythroderma, scaling, hyperkeratosis and fissuring with prominent involvement of palms and soles (Figures 1c and 2), lymphadenopathy and circulating Se´zary cells.13 A combination of MTX and steroids failed to control the disease which was complicated by Herpes zoster infection and Staphyococcus aureus (SA) bacteremia. He was enrolled into our protocol in January 2001. On day +16 he developed hallucinations, generalized seizures and loss of consciousness that did not improve after switching from cyclosporine-A to FK506. Polymerase chain reaction (PCR) analysis was consistent with an EBV-related LPD, as demonstrated by the presence of high levels of EBVDNA and clonal rearrangement of the immunoglobulin k-chains in the cerebrospinal fluid (data not shown). The clinical picture completely resolved over a period of 3 weeks secondary to reduction of the immunosuppressive regimen. On day +58, the patient developed grade III acute skin GVHD, bilateral pulmonary infiltrates and eosinophilia. The symptoms abated after administration of prednisone at a starting dose of 1 mg/kg/day. A total of 18 months after ASCT, the patient is in complete clinical and molecular (both in the skin and peripheral blood) remission (Figures 1d and 2) with limited chronic GVHD that responded to resumption of FK506. Case 3: In 1986, a 37-year-old male was diagnosed with MF (stage IIB). He had failed multiple prior therapies, and
Advanced MF is associated with a poor outcome3 and both combination chemotherapy and autologous BMT have failed to have an impact on survival.4–6 The role of conventional ASCT has been hypothesized considering: (1) the anti-MF activity through immune mechanisms observed in phase I/II trials using systemic cytokines or fusion toxins;14,15 (2) the skin as a target of both MF and GVHD, which one associates with the GVT effect. In addition, there has been evidence of a long-lasting CR and GVT effect after conventional ASCT in a few patients with advanced MF or Se´zary syndrome.7–9 However, this therapy has been limited by the advanced median age of the patients at diagnosis, the indolent clinical course, the increased susceptibility to systemic infection and sustained associated comorbidity. In this regard, the recent advent of ASCT that employs nonmyeloablative, but heavily immunosuppressive, conditioning regimens in order to facilitate donor engraftment with reduced toxicity16,17 may represent a potential therapeutic strategy in advanced MF/SS. This approach, which is obviously limited to a minority of patients with a histocompatible donor, could extend the indications of ASCT to patients aged up to 70 years. All patients engrafted fully, achieving 100% donor chimerism on PB CD8+ T-cells from day +28, and 100% donor chimerism on unfractionated bone marrow on day +100. The severe host immunodeficiency underlying both disorders probably facilitated these accelerated engraftment kinetics. Between days +30 and +60, all patients had completely cleared the clonal T cells from the skin, lymph nodes, bone marrow and peripheral blood. Clinical, histological and molecular remissions were durable (ie, up to 16 months following ASCT). The clinical responses we observed cannot be solely attributable to the conditioning and/or immunosuppressive regimens. TBI of 200 cGy is unlikely to be effective in controlling advanced MF/SS, since conventional autologous BMT with high-dose TBI is associated with short-lived responses in these disorders.4,5 Similarly, purine analogs seldom give rise to complete remission in MF/SS,18 while the use of cyclosporine-A is uniformly Bone Marrow Transplantation
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associated with rapid tumor recurrence upon withdrawal of the drug.19 We documented a close temporal association between onset of skin GVHD in all patients, modulation of immunosuppression and disappearance of the tumor, as proven by histology. These findings confirm the existence of a strong graft-versus-tumor effect and indicate that ASCT may be an effective curative option for patients with MF. Severe, life-threatening infections during the transplant course were common to all our patients. Two out of three developed SA bacteremia which is reportedly the most common cause of death in MF/SS.1,20,21 Profoundly compromised T-cell function is also emphasized by the HCMV reactivation events and EBV-related LPD that we frequently observed. However, since neither subject with EBV-LPD experienced a recurrence despite ongoing immunosuppressive therapy, one may hypothesize that ASCT was ultimately associated with restoration of immune functions. Although patient selection, preparative regimens and identification and treatment of infectious complications should be further improved, nonmyeloablative ASCT may represent a novel therapeutic strategy for patients with MF and deserves further exploration in larger studies.
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