Transplantation (2005) 35, S17–S21 & 2005 Nature Publishing Group All rights reserved 0268-3369/05 $30.00 www.nature.com/bmt

Hematopoietic transplantation for bone marrow failure syndromes and

J Sevilla, S Ferna´ ndez-Plaza, MA Diaz and L Madero, on behalf of the Paediatric Disease Working Party of the EBMT

Department of Pediatric Oncohematology, Hospital Infantil Universitario Nin˜o Jesu´s, Madrid,

Summary: and hypersensitivity to alkylating agents. It is an autosomal recessive disease and diagnosis of FA homozygotes is Several genetic diseases, generally considered as con- confirmed by the detection of chromosome aberrations in genital diseases, are characterized by bone marrow failure lymphocytes (or fibroblasts) following culture with cross- during early childhood. Hematopoietic trans- linking agents (diepoxybutane or mitomycin C). To date, plantation is the only curative treatment for syndromes 11 complementation groups have been identified and eight involving bone marrow failure and thalassemia. In this FA genes have already been cloned (Table 2).3 slate-of-the-art review, we wish to focus on the results of Hematopoietic cell transplantation is the only treatment hematopoietic transplantation in treating some of these that can cure the bone marrow aplasia in patients with FA. diseases, with a special emphasis on congenital bone Nevertheless, while this problem is the earliest and most marrow failure and thalassemia. The results of this frequent life-threatening complication, severe birth defects procedure have improved over the previous years, mainly and predisposition must also be taken into account when performed by experienced teams. New conditioning in children with FA. Hematopoietic progenitor cell regimes based on fludarabine and the use of HLA- transplantation in patients with FA has been limited in identical donors have been related with better survivals. the past by high rates of graft failure, regimen-related In the previous years, donors other than HLA-identical toxicity, and acute and chronic graft-versus-host disease. siblings have been increasingly used in patients not Furthermore, recent studies have found that the risk of responding to conventional measures, but this approach some malignancies appearing is higher in patients after needs to be evaluated in larger studies. transplantation than in patients who did not undergo Bone Marrow Transplantation (2005) 35, S17–S21. transplantation.4 doi:10.1038/sj.bmt.1704838 It is well known that patients with FA are highly Keywords: congenital bone marrow failure; Fanconi susceptible to alkylating agents and irradiation. Therefore, ; Diamond–Blackfan anemia; severe congenital it has been suggested that the dose of cyclophosphamide ; INCLUIRIMAGEN thalassemia; hemato- could be decreased in order to improve survival, as well poietic transplantation avoiding the use of irradiation in the conditioning regime. Low-toxicity regimes (nonmyeloablative) with fludarabine have achieved very good results with HLA-identical 5 Several genetic diseases, generally considered as congenital donors, and indeed this regime achieved 100% event-free diseases, are characterized by bone marrow failure during survival in one series of patients when combined with 6 early childhood (Table 1). Although they have recently T-cell-depleted grafting. When alternative donors are used, been addressed in different reviews,1,2 here we wish to focus some authors consider that higher myeloablative regimens on the results of hematopoietic transplantation in treating are necessary. In this scenario, irradiation and fludarabine some of these diseases, with a special emphasis on lead to 76% overall survival in a standard risk group (less congenital bone marrow failure and thalassemia. than 18 years old, absence of acute or myelodys- plastic syndrome (MDS), and HLA-matched bone marrow or cord ).6 However, there is as yet no consensus Congenital bone marrow failure regarding the use of irradiation with these patients and some groups have avoided its use with good results even when unrelated donors are involved. Indeed, the severe Fanconi anemia (FA) is a heterogeneous disorder char- working party of the EBMT has recently acterized by bone marrow failure, susceptibility to cancer, reported 85% overall survival for children under 8 years of age after hematopoietic transplantation with alternate donors. In this protocol, the conditioning regime involved fludarabine 30 mg/m2, cyclophosphamide 300 mg/m2, and Correspondence: Dr J Sevilla, Hospital Infantil Universitario Nin˜ o 7 Jesu´ s, Avd Mene´ ndez Pelayo 65, Madrid 28009, Spain; ATG 3.75 mg/kg for 4 days. A similar regime was also E-mail: [email protected] described by de la Fuente et al8 in four children diagnosed Hematopoietic transplantation for bone marrow failure and thalassemia J Sevilla et al S18 Table 1 Congenital bone marrow failure syndromes

Pancytopenia aplasia Neutropenia Thrombopenia

Fanconi anemia Diamond–Blackfan anemia Kostmann Syndrome Amegakaryocytic with absent radii Schwachman–Diamond syndrome Schwachman–Diamond Congenital amegakaryocytic syndrome thrombocytopenia Others: Pearson syndrome, dyskeratosis congenital, reticular dysgenesia, cartilage hair hypoplasia

Table 2 Fanconi anemia complementation groups

Complementation group Identified gene Chromosomal location Reference

FA-A FANCA 16q24–3 Lo Ten Foe et al. Nat Genet 1996; 14: 320 FABC consortium.a Nat Genet 1996; 14: 324 FA-B — — Strathdee et al. Nat Genet 1992; 1: 196–198 FA-C FANCC 9q22.3 Strathdee et al. Nat Genet 1992; 1: 196–198 FA-D1 BRCA2 13q12–13 Howlett et al. Science 2002; 297: 606–609 FA-D2 FANCD2 3p25.3 Timmers et al. Mol Cell 2001; 7: 241–248 FA-EFANCE6p21.3 De Winter et al. Am J Hum Genet 2000; 67: 1306–1308 FA-F FANCF 11p15 De Winter et al. Nat Genet 2000; 24: 15–16 FA-G FANCG 9p13 De Winter et al. Nat Genet 1998; 20: 281–283 FA-I — — Levitus et al. Blood 2004; 103: 2498–2503 FA-J — — Levitus et al. Blood 2004; 103: 2498–2503 FA-L FANCL 2p16 Meetei et al. Nat Genet 2003; 35: 165–170

aThe Fanconi Anaemia/Breast cancer consortium.

with FA. Progenitor cells from unrelated donors were used Other congenital bone marrow failures in two patients, and HLA-matched sibling donors in the other two children. Full donor grafting was achieved in the Recently, encouraging results have been reported when first month after transplantation with minimal toxicity in hematopoietic transplants have been performed in patients all of these cases, and at 11, 23, 30, and 51 months after diagnosed with Severe Congenital Neutropenia12,13 and transplantation they were alive and well (Karnofsky Diamond–Blackfan Anemia14 (DBA). Indeed, the ‘Inter- 100%).8 In other studies, encouraging results have also national Bone Marrow Transplant Registry’ recently been reported with other fludarabine-based conditioning presented their experiences with DBA14 in which they regimes.5,9 reviewed the outcome of 61 patients registered after While progenitor cells obtained from bone marrow are receiving allogeneic bone marrow transplants for DBA.14 the most frequently used for patients diagnosed with FA, in The probability of overall survival was 67% (54–77%) after some cases transplants have been performed with stem cells 1 year and remained at 64% (50–74%) after 3 years. In a collected from peripheral blood. However, to date, no in- univariant analysis, a Karnofsky score X90 was associated depth comparison of these two stem cell sources has been with better overall survival for 100 days (93 vs 63%, performed. The experiences of the Eurocord registry Po0.01) and for 3 years (75 vs 42%, P ¼ 0.01).14 Similarly, involving 44 patients diagnosed with FA have recently transplants from a HLA-identical sibling were associated been reported,10 seven of which were transplanted from with higher survival than those using an alternative donor, HLA-identical donors and 37 from nonidentical donors. the survival being 78 vs 45% after 1 year (P ¼ 0.01) and 75 The overall survival for these patients was 36% after 2 vs 39% after 3 years (P ¼ 0.01).14 years, and this appeared to be related to the number of The availability of G-CSF dramatically changes the nucleated cells at freezing (45.5 Â 107/kg, 55 vs 16%) and prognosis of severe congenital neutropenia and the quality the use of fludarabine in the preparative regime (67 vs of life of these patients. However, almost 10% of patients 25%). Indeed, the overall survival in this study is similar to are refractory to this therapy and, for them, the only that reported earlier by Guardiola et al11 with bone marrow treatment available is hematopoietic transplantation.12 This stem cells from unrelated donors in a study on behalf of the is also the only curative treatment for those patients who European group for blood and marrow transplantation. develop MDS or acute leukemia (AL), and those who have Therefore, this source of stem cells may expand the mutations in the G-CSF receptor, since this seems to play possibility of finding an unrelated donor for children that an important role in the development of leukemia.12 All would otherwise be unable to undergo transplantation. patients reported in the International Severe Chronic Moreover, with a better selection of the and the Neutropenia register who received a transplant from a use of fludarabine in the conditioning regime, these results sibling donor without evidence of leukemic transformation may improve in the near future.10 were still alive at the time the study was published (8/8).12

Bone Marrow Transplantation Hematopoietic transplantation for bone marrow failure and thalassemia J Sevilla et al S19 This is the best scenario, since the prognosis was not so overall survival of 66%.17 An update of these data showed good for those patients who developed MDS/AL. Recently, a nonrejection-related mortality rate for older patients of Zeidler et al13 reviewed this topic at the ASH meeting and 36%, while in patients below 17 years of age it was only suggested that hematopoietic transplantation should be 18%.18 Hence, it is clear from both studies that the main performed as early as possible. The main objective is to problem for younger patients is the rate of rejection. Thus, prevent before transplantation and therefore while the results were optimal for younger patients intensive AML-type chemotherapy should be avoided.13 belonging to risk groups 1 and 2, the challenge then was Indeed, these authors suggested using low-dose chemother- to improve the results in older patients and those with liver apy to control the neoplastic clone until hematopoietic disease. transplantation could be performed.13 This improves the Based on prior experiences, the regime for patients results from the 31% reported by the international classified in risk group 3 was modified in 1997,19 involving database up to the end of 2000, to the 59% reported in a hypertransfusion and intensive iron chelation, and hydro- European cohort of patients.13 xyurea 30 mg/kg daily and azathioprine 3 mg/kg daily For ‘Severe Congenital Neutropenia’ and DBA, the between days À45 and À11. The aim was to reduce the problem of the optimal timing of transplantation is expansion of the immature red blood cells and eradicate the constantly under debate since some of these patients can marrow. Subsequently, fludarabine was added (20 mg/m2 be controlled for years with supportive therapies. However, from day À17 through to day À13) because of its it seems clear that for those cases that are nonresponsive to immunosuppressive and bone-marrow-suppressive activ- medical therapies and where a matched sibling donor is ities. Additionally, busulfan 1 mg/kg/dose in three doses per available, stem cell transplantation should be performed day was administered for 5 days (À10 through to À6, total rapidly in order to decrease transplant-related mortality. dose 14 mg/kg), and cyclophosphamide 40 mg/kg daily Transplantation from donors other than a matched sibling (À5 through to À2, total dose 160 mg/kg).16 For graft- donor should be considered on a case-by-case basis when versus-host disease prophylaxis, the following regime was individual circumstances justify the risk. followed: cyclophosphamide 7.5 mg/kg on day 1; metho- trexate 10 mg/m2 intravenously on days 3 and 6; cyclos- porine 5 mg/kg from day À2 through to day 5, and then Stem cell transplantation for thalassemia reduced to 3 mg/kg intravenously until oral administration could be tolerated; and prednisolone 0.5 mg/kg from day The EBMT registry included 2312 patients from 112 À1 until day 25. With this new regime, the probability of different centers diagnosed with thalassemia, who had overall survival was 93% and thalassemia-free survival undergone hematopoietic transplantation by April 2004. improved to 85%.16 Moreover, the probability of rejection Most transplants were performed using only bone marrow was reduced to 8% and nonrejection mortality to 6%,16 as the stem cell source (90%) and most of them had a results similar to those obtained for risk groups 1 and 2 matched related donor (90%; Dr Angelucci, EBMT, 2004). with the initial protocols. For adult patients in this group, a In Pesaro, Dr Guido Lucarelli and his group first began to new conditioning regime was also developed that was use bone marrow transplantation in selected patients with similar to that used for younger patients, but with a total this disease in 1981, and, over the last 20 years, 1003 dose of cyclophosphamide of 90 mg/kg. However, the patients have been transplanted in their unit from a results of using such a regime have yet to be reported. matched related donor. As a result, most of our current Thus, it seems that we have overcome the challenge to knowledge regarding the use of this therapy for thalassemia improve the treatment for group 3 patients, such that the has been generated by this group. Initially, all patients new challenges will focus on those patients lacking a received a conditioning regime involving total doses of matched related donor. cyclophosphamide of 200 mg/kg and 14–16 mg/kg of busulfan. In patients younger than 17 years of age, the Unrelated transplants results of this protocol led to their classification into three different risk groups according to the appearance of hepatic The first protocol using unrelated donors to treat thalasse- disease secondary to iron deposition and hepatomegaly. mia was reported by Contu et al in 1994,20 although the The thalassemia-free survival in these groups varied from largest published study on this subject, which includes 32 94% for patients in group 1, to 77% for those in group 2, patients, has only just recently been published.21 In this and 53% for patients in group 3.15 In order to reduce the study, La Nasa et al reported the results of using unrelated toxicity in patients in group 3 associated with the donors for bone marrow transplants after a careful conditioning regime, the total dose of cyclophosphamide immunogenetic selection of the donor/recipient pairs. was reduced to 120–160 mg. This modification in the Among these patients, four were assigned to risk class 1, protocol increased the overall survival to 57% with the 11 to risk class 2, and 17 to risk class 3, and not all patients higher doses of cyclophosphamide or 74% with the lower received the same conditioning regime. Initially, total doses doses. However, the probability of rejection also increased of cyclophosphamide 200–160 mg/kg and busulfan 14 mg/ from 13% to 35% and, as a result, there was no kg were used in four patients, but since graft failure improvement in thalassemia-free survival (53 vs 49%).16 occurred in two of them, thiotepa 10 mg/kg was added to Furthermore, older patients treated with lower doses of this protocol. Finally, cyclophosphamide was reduced to a cyclophosphamide, who were included in risk groups 2 or total dose of 120 mg/kg in those patients over 16 years of 3, only achieved 62% thalassemia-free survival and an age. In this study, the Kaplan–Meier probability of survival

Bone Marrow Transplantation Hematopoietic transplantation for bone marrow failure and thalassemia J Sevilla et al S20 was 79% and the thalassemia-free survival was 66%. Of the transplantation experienced autologous hematopoietic 22 patients that the authors considered had an optimal recovery.24 donor, 19 were still alive and 17 of them did not require transfusions. Only two patients developed a rejection of the graft after thiotepa was added to the protocol. In this Conclusion study, there were six transplant-related deaths, and, although not statistically significant, five of them belong transplantation is the only curative to risk class 3, and one to group 1. treatment for syndromes involving bone marrow failure More recently, the results from a series of 11 patients and thalassemia. Interestingly, new conditioning regimes with thalassemia who received transplants from unrelated based on fludarabine have achieved very good results in donors have been published.22 The patients underwent these syndromes in recent years. Moreover, donors other bone marrow transplantation following a conditioning than HLA-identical siblings are increasingly used, but this regime with cyclophosphamide (200 mg/kg total dose) and approach needs to be evaluated in larger studies. busulfan (16 mg/kg total dose), as well as the addition of antilymphocyte globulin (40 mg/kg total dose) to this regime. Most of the patients were classified in risk class 1 References (eight), while only two were assigned to risk class 3. In all, 1 Alter BP. Bone marrow failure syndromes in children. Pediatr 10 donors were identical at molecular level to the recipients Clin N Am 2002; 49: 973–988. for HLA-A, -B, and -DRB1, and only one patient was 2 Sieff CA, Nisbet-Brown E, Nathan DG. Congenital bone mismatched at for an HLA-DRB1 allele. The graft-versus- marrow failure syndromes. Br J Haematol 2000; 11: 30–42. host disease prophylaxis consisted of methotrexate and 3 Rosenberg PS, Huang Y, Alter BP. Individualized risks of cyclosporine. At the time of publication, all patients had adverse events in patients with Fanconi anemia. Blood 2004 full donor engraftment and were alive. However, three (DOI 10.1182/blood-2004-01-0083). patients (27%) developed chronic limited graft-versus-host 4 Socie G, Rosenberg P, Gluckman E, Alter B. How can we disease in this series.22 While this study needs to be quantify the risk of squamous cell cancer (SCC) and death in expanded in terms of the number of patients, it does transplanted versus non-transplanted patients with Fanconi’s anaemia. Bone Marrow Transplantation 2004; 33 (Suppl. 1): provide ‘additional evidence to support the view that is S27. quite reasonable to consider unrelated donor BMT as an 5 MacMillan ML, Auerbach AD, Champagne MA et al. High acceptable therapy to cure thalassemia, at least for patients probability of survival after related and alternate donor who are young and do not yet show irreversible severe hematopoietic cell transplantation for Fanconi anemia using complications of ’, as the authors point out.19 fludarabine based preparative therapy. Blood 2003; 102 (abstr. 1695). 6 Tischkowitz M, Dokal I. Fanconi anaemia and leukemia Other sources of stem cells – clinical and molecular aspects. Br J Haematol 2004; 126: 176–191. The experiences of ‘Eurocord’ were published in 2003,23 7 Bacigalupo A, Locatelli F, Socie´ G et al. Fludarabine, where 44 patients diagnosed with SCD or thalassemia cyclophosphamide and ATG for alternative donor transplants underwent transplantation using cord blood as the stem cell in aplastic anaemia: a report of the SAA Working Party. Bone source. In all, 33 thalassemic patients (20 risk class I, and 13 Marrow Transplant 2004; 33 (Suppl. 1): S29. class II) and 11 patients diagnosed with SCD received a 8 de la Fuente J, Reiss S, McCloy M et al. Non-TBI stem cell related cord blood transplant with a graft probability of transplantation protocol for Fanconi anaemia using HLA- compatible sibling and unrelated donors. Bone Marrow 89%, aGVHD 8%, and cGHVD 6%. The probability of Transplant 2003; 32: 653–656. event-free survival was 79% for the subjects with thalasse- 9 Slavin S, Bitan M, Aker M et al. Non-myeloablative stem mia and 90% for those with SCD. Survival was related to cell transplantation (NST) for the treatment of patients the use of methotrexate, since the event-free survival was with Fanconi’s anaemia. Bone Marrow Transplant 2004; 33 lower in those patients in whom it was used for GVHD (Suppl. 1): S141. prophylaxis than in those whom did not receive this drug. 10 Gluckman E, Pasquini R, Rocha V et al. Results of unrelated There was also a trend but not a significant association cord blood transplant in Fanconi’s anaemia. Bone Marrow between a higher event-free survival in risk group I Transplant 2004; 33 (Suppl. 1): S65. thalassemia patients, and in those whose conditioning 11 Guardiola PH, Pasquini R, Dokal I et al. Outcome of 69 regime did not involve busulfan–cyclophosphamide alone.23 allogeneic stem cell transplantation for Fanconi anemia using HLA-matched unrelated donors: a study on behalf of the European Group for Blood and Marrow Transplantation. Blood 2000; 95: 422–429. Lower-toxicity conditioning regimes 12 Zeidler C, Welte K, Barak Y et al. Stem cell transplantation in There is little information regarding nonmyeloablative patients with severe congenital neutropenia without evidence Blood regimes for hematopoietic transplantation in hemoglobi- of leukemic transformation. 2000; 95: 1195–1198. 13 Zeidler C, Schwinzer B, Pracht G et al. Hematopoietic stem nopathies. Conditioning regimes based on fludarabine, with cell transplantation in congenital neutropenias: recommenda- low-dose busulfan or total body irradiation (200 cGy), have tion for patients refractory to G-CSF treatment and secondary 24 been used in these diseases. Indeed, seven patients MDS or . Blood 2003; 102 (abstr. 971). diagnosed with who underwent a 14 Roy V, Perez WS, Marsh J et al. Bone marrow transplantation nonmyeloablative conditioning regime for hematopoietic for Diamond–Blackfan anemia: report of the International

Bone Marrow Transplantation Hematopoietic transplantation for bone marrow failure and thalassemia J Sevilla et al S21 Bone Marrow Transplant Registry experience. Blood 2003; 102 20 Contu L, La Nasa G, Arras M et al. Successful unrelated bone (abstr. 861). marrow transplantation in beta-thalassaemia. Bone Marrow 15 Lucarelli G, Galimberti M, Polchi P et al. Bone marrow Transplant 1994; 13: 329–331. transplantation in patients with thalassemia. N Engl J Med 21 La Nasa G, Giardini C, Argiolu F et al. Unrelated donor bone 1990; 322: 417–421. marrow transplantation for thalassemia: the effect of extended 16 Lucarelli G, Clift LA, Galimberti M et al. Marrow transplant- haplotypes. Blood 2002; 99: 4350–4356. ation for patients with thalassemia: results in class 3 patients. 22 Hongeng S, Pakakasama S, Chaisiripoomkere W et al. Out- Blood 1996; 87: 2082–2088. come of transplantation with unrelated donor bone marrow in 17 Lucarelli G, Clift LA, Galimberti M et al. Bone marrow children with severe thalassaemia. Bone Marrow Transplant transplantation in adult thalassemic patients. Blood 1999; 93: 2004; 33: 377–379. 1164–1167. 23 Locatelli F, Rocha V, Reed W et al. Related umbilical cord 18 Lucarelli G, Andreani M, Angelucci E. The cure of thalasse- blood transplantation in patients with thalassemia and sickle mia by bone marrow transplantation. Blood Rev 2002; 16: cell disease. Blood 2003; 101: 2137–2143. 81–85. 24 Iannone R, Casella JF, Fuchs EJ et al. Results of minimally 19 Sodani P, Gaziev D, Polchi P et al. New approach for bone toxic nonmyeloablative transplantation in patients with sickle marrow transplantation in class 3 thalassemic patients aged cell anemia and beta-thalassemia. Biol Blood Marrow Trans- less than 17 years. Blood 2004; 104: 1201–1203. plant 2003; 8: 519–528.

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