hematopoietic stem cell transplantation in...

Hematopoietic Stem Cell Transplantationin Thalassemia and Sickle Cell Anemia

Guido Lucarelli, Antonella Isgro, Pietro Sodani, and Javid Gaziev

International Center for Transplantation in Thalassemia and Sickle Cell Anemia–MediterraneanInstitute of Hematology, Policlinic of the University of Rome Tor Vergata, Tor Vergata, Italy

Correspondence: [email protected]

The globally widespread single-gene disorders b-thalassemia and sickle cell anemia (SCA)can only be cured by allogeneic hematopoietic stem cell transplantation (HSCT). HSCTtreatment of thalassemia has substantially improved over the last two decades, with advance-ments in preventive strategies, control of transplant-related complications, and preparativeregimens. A risk class–based transplantation approach results in disease-free survival prob-abilities of 90%, 84%, and 78% for class 1, 2, and 3 thalassemia patients, respectively.Because of disease advancement, adult thalassemia patients have a higher risk for trans-plant-related toxicity and a 65% cure rate. Patients without matched donors could benefitfrom haploidentical mother-to-child transplantation. There is a high cure rate for childrenwith SCA who receive HSCT following myeloablative conditioning protocols. Novel non-myeloablative transplantation protocols could make HSCT available to adult SCA patientswho were previously excluded from allogeneic stem cell transplantation.

Thalassemia and sickle cell anemia (SCA) arethe two most widely diffused hereditary he-

moglobinopathies in the world. Thalassemiaoriginated in Mediterranean, Middle Eastern,and Asian countries, and sickle cell anemia orig-inated throughout African countries; both soonbecame globally spread because of spontaneousmigration and migration imposed through slav-ery. Modern migration is currently causing bothdiseases, particularly SCA, to further invade Eu-ropean countries (see Williams 2012).

b-Thalassemia involves deficient or absentsynthesis of the b-globin chains that constituteadult hemoglobin molecules. This genetic de-fect affects erythropoiesis through the entireprocess of maturation and proliferation of the

erythron, with severe apoptosis due to accumu-lation of a2 chains in the erythroid precursors.Early death of the red blood cells continues inthe blood circulation. Patients without transfu-sions experience an enormous expansion of theerythroid and severe hemolytic anemia.

In SCA, a single nucleotide substitution—valine replacing glutamic acid at the sixth posi-tion of the b-globin chain of hemoglobin A—forms a pathological hemoglobin called hemo-globin S (HbS). The propensity of HbS poly-merization causes the red blood cell structure tochange to a stable sickle shape. Resulting epi-sodic anemia and acute and chronic vaso-occlu-sion of small and large vessels in virtually allorgans cause the polymorphic clinical features

Editors: David Weatherall, Alan N. Schechter, and David G. Nathan

Additional Perspectives on Hemoglobin and Its Diseases available at www.perspectivesinmedicine.org

Copyright # 2012 Cold Spring Harbor Laboratory Press; all rights reserved; doi: 10.1101/cshperspect.a011825

Cite this article as Cold Spring Harb Perspect Med 2012;2:a011825

1

ww

w.p

ersp

ecti

vesi

nm

edic

ine.

org

Press on February 13, 2020 - Published by Cold Spring Harbor Laboratoryhttp://perspectivesinmedicine.cshlp.org/Downloaded from

http://perspectivesinmedicine.cshlp.org/

of the sickle cell disease. Intramedullary apop-tosis is also present but moderate, never at thelevels seen in thalassemia.

Many thalassemic patients need continuousred blood cell replacement through a sequentialprogram of red blood cell transfusions; the SCApatient does not. Without transfusions, thalas-semic patients have tremendous skeleton de-formities, as well as hepatomegaly and spleno-megaly due to expansion of the hematopoieticsystem with extramedullary hematopoiesis. TheSCA patient can suffer from severe organ dam-age due to the sequence of HbS polymerization,formation of microemboli, adhesion to the in-tima of the vessels, and organ infarction. Thepathogeneses and clinical aspects of thalassemiaand sickle cell anemia are described in greaterdetail in other articles in this collection.

Because both thalassemia and SCA are ge-netic diseases in which the genetic defect is ex-pressed in the hematopoietic system, they areboth curable by allogeneic cellular gene therapythrough hematopoietic stem cell transplanta-tion. On the other hand, the separate pathogen-eses of these two disorders explain why theremust also be some differences between the trans-plantation procedures.

HEMATOPOIETIC STEM CELLTRANSPLANTATION FOR THALASSEMIA

HSCT for Thalassemia: Risk Class Approach

Allogeneic HSCT is used to treat hemoglobin-opathies; afterconditioning to overcome the im-munological barrier, allogeneic stem cells areused as vectors to correct the basic genetic defectby replanting genes that are essential for normalhematopoiesis. Essentially, allogeneic HSCTtreatment of these diseases can be considered asallogeneic stem cell gene therapy. It may someday be possible to use autologous stem cellstransformed by the insertion of normal genesas vectors, but there is no indication that thisapproach will be a clinical option in the fore-seeable future.

For treating diseases other than aplasticanemia, preparatory regimens for HSCT mustachieve two objectives: elimination of the disor-

dered marrow and establishment of a tolerantenvironment that will permit transplanted mar-row to survive and thrive. There is a consider-able body of experience with the use of busulfan(BU) and its derivatives for ablating marrowin patients undergoing HSCT for the treatmentof non-malignant conditions (Parkman et al.1978; Kapoor et al. 1981; Hobbs et al. 1986).Cyclophosphamide (CY) is a well-establishedagent for providing adequate immunosuppres-sion for allogeneic engraftment (Thomas et al.1972; Storb et al. 1991). The combination of BUand CY can eradicate thalassemia and facilitatesustained allogeneic engraftment. The two fun-damental treatments in pretransplant prepara-tion include eradication of the hematopoieticsystem by administration of a total dose of14–16 mg/kg busulfan and suppression of theimmune system by administration of a totaldose of 200 mg/kg cyclophosphamide. Becauseof the overlap of the anti-proliferative activity ofbusulfan on the immune cell system and of cy-clophosphamide on the hematopoietic stem cellsystem, the summed toxicity of these dosagesof busulfan and cyclophosphamide reaches thetolerability threshold and cannot be increased.

When the allogeneic graft starts to prolifer-ate in the recipient, an immunological graft-versus-host reaction (graft-versus-host disease[GVHD]) may occur. GVHD can affect one ormore organs to varying degrees, with the mostfrequently affected being the skin, gastrointesti-nal tract, and liver. GVHD is a serious compli-cation of bone marrow transplantation and canbe fatal; therefore, the prophylactic administra-tion of cyclosporine (an immunosuppressivedrug) is an important part of the pretransplantand posttransplant treatment. Three or fourdoses of methotrexate may also be given in ad-dition to cyclosporine during the first 15 d aftertransplant.

The influence of pretransplant characteris-tics on the outcome of transplantation was ana-lyzed in 161 patients ,17 yr of age, who were alltreated with exactly the same regimen (Storbet al. 1977; Lucarelli et al. 1990). Multivariateanalysis showed that hepatomegaly of morethan 2 cm, portal fibrosis, and irregular chela-tion history were associated with a significantly

G. Lucarelli et al.

2 Cite this article as Cold Spring Harb Perspect Med 2012;2:a011825

ww

w.p

ersp

ecti

vesi

nm

edic

ine.

org



reduced probability of survival. The quality ofchelation was characterized as regular when de-feroxamine therapy was initiated no later than18 mo after the first transfusion and was admin-istered subcutaneously for 8-10 h continuouslyfor at least 5 d/wk. Any deviation from this reg-imen was defined as irregular chelation. On thebasis of these risk factors, patients were catego-rized into three risk classes: Class 1 patients hadnone of these adverse risk factors, class 2 patientshad one or two adverse risk factors, and class 3patients had all three.

HSCT from HLA-Matched Related Donors

The results of transplants from HLA-matchedrelated donors to class 1, class 2, and class 3patients, obtained by our group, are reportedin Table 1 (Lucarelli et al. 2008; Isgro et al. 2010).

Adult Patients

Adult thalassemia patients have more advanceddisease, with both disease- and treatment-relat-ed organ complications that are mainly dueto prolonged exposure to iron overload. FromNovember 1988 through September 1996, 107.16-yr-old patients received transplants from

matched donors at the Hospital of Pesaro. Themedian age for this population was 20 yr, with arange of 17–35 yr. The probabilities of survival,thalassemia-free survival, rejection mortality,and non-rejection mortality for this entiregroup were 66%, 62%, 4%, and 37%, respective-ly (Lucarelli et al. 1992, 1999).

From April 1997, 15 high-risk adult patientswere prepared for transplantationwith a reducedtotal dose of 90 mg/kg cyclophosphamide.The probabilities of survival, thalassemia-freesurvival, rejection mortality, and non-rejectionmortality were 65%, 65%, 7%, and 28%, respec-tively (Gaziev et al. 2005).

HSCT FROM ALTERNATIVE DONORS FORTHALASSEMIA

A major obstacle to successful transplantationis the limited number of HLA-matched relateddonors within families. Approximately 60%of patients lack a suitable family donor. Someof these patients could benefit from HSCT frommatched unrelated donors. However, the chanceof finding a matched unrelated donor is stronglydependent on the patient’s ethnic background.

One study reported the outcome of bonemarrow transplant (BMT) from matched unre-lated donors who were prospectively selected

Table 1. Outcomes of hematopoietic cell transplantation from HLA-matched donors in casesof b-thalassemia

Patients Stem cell source OS (%) TFS (%) Reference

,17 yr: 515 (class 1–2)73 (class 3)Adult: 107

MSD/MRD Class 1–2: 88%Class 3: 87%Adult: 66%

Class 1–2: 85%Class 3: 82%Adult: 62%

Lucarelli et al. 2008

68 MUD 79.3% 40% La Nasa et al. 2005a33 Cord blood 100% 79% Locatelli et al. 200370 MSD/MRD 83% 73% Ghavamzadeh et al. 199749 Related: 28

Unrelated: 2189% (all patients) Related: 82%

Unrelated: 71%Hongeng et al. 2006

37 (class 1–2)35 (class 3)

MSD Class 1–2: 97%Class 3: 87%

Class 1–2: 89%Class 3: 80%

Isgro et al. 2010

3 (class 1)75 (class 2)64 (class 3)37 (NA)

MSD Class 1: NAClass 2: 91%Class 3: 64%

Class 1: NAClass 2: 88%Class 3: 62%

Sabloff et al. 2011

Abbreviations: OS, overall survival; TFR, thalassemia-free survival; NA, not available; MSD, matched sibling donor; MRD,

matched-related donor; MUD, matched unrelated donor; SCD, sickle cell disease.

HSCT in Thalassemia and Sickle Cell Anemia

Cite this article as Cold Spring Harb Perspect Med 2012;2:a011825 3

ww

w.p

ersp

ecti

vesi

nm

edic

ine.

org



using high-resolution molecular typing for HLAclass I and class II loci; this study included 68patients with thalassemia major who receivedBU/CYor BU/FLU (fludarabine) and/or thio-tepa (TT) as a conditioning regimen (La Nasaet al. 2005a). The study included 14 class 1, 16class 2, and 38 class 3 patients. Overall rates ofsurvival, thalassemia-free survival, rejection,and transplant-related mortality (TRM) for theentire cohort were 79.3%, 65.8%, 14.4%, and20.7%, respectively. The incidences of acutegrade II–IV or grade III–IV GVHD were 40%and 17%, respectively. Ten of 56 evaluable pa-tients (18%) developed chronic GVHD. Class 1and class 2 patients had much better overall anddisease-free survival (96.7% and 80%, respec-tively) than class 3 patients (65.5% and 54.5%,respectively). These investigators have also re-cently reported encouraging results inadult thal-assemia patients who received bone marrowfrom matched unrelated donors, with rates ofsurvival, thalassemia-free survival, TRM, andrejection of 70%, 70%, 30%, and 4%, respective-ly (La Nasa et al. 2005b). Similar results werereported in another study from Asia that includ-ed 21 children who received transplants frommatched unrelated donors, as well as 28 patientswho received transplants from matched relateddonors (Hongeng et al. 2006). The 2-yr thalas-semia-free survival for patients who receivedtransplants from matched unrelated donorswas 71%, compared with 82% for patients whoreceived transplants from matched related do-nors.

Together, these data strongly suggest thatimprovements in donor selection and trans-plantation preparation have improved the safetyof unrelated donor HSCT for thalassemia treat-ment. This appears to be a viable treatment op-tion for selected patients when there is no suit-able sibling donor.

Results of Transplant from HaploidenticalDonors

Haploidentical transplantation may extend theuse of this treatment option to the 50%–60% ofpatients who lack a suitably matched familialdonor or an HLA-identical unrelated donor.

The presence of fetal cells in maternal bloodand of maternal cells in fetal blood (fetomater-nal microchimerism) suggests that immunolog-ical tolerance may exist between mother andoffspring. The combination of a megadose ofpurified CD34þ cells and a highly immuno-myeloablative conditioning regimen is crucialfor overcoming the barrier of residual anti-do-nor cytotoxic T-lymphocyte precursors in T-cell-depleted mismatched transplants, and theaddition of BMMCs (including NK cells, mes-enchymal stem cells, and T-cells) to a T-cell-depleted allograft may help engraftment andcontrol GVHD. Recently, we reported the out-comes of 31 children with thalassemia who re-ceived transplants from haploidentical donors;27 were from mothers, two from brothers, andtwo from fathers (Sodani et al. 2011). Eightpatients received T-cell-depleted peripheralblood progenitor cells (CD34þ immunoselec-tion) and CD3þ- and CD19þ-depleted bonemarrow stem cells. Twenty-three patients re-ceived CD34þ mobilized peripheral and bonemarrow progenitor cells. Positive selection wasperformed using the CliniMACS procedure.The CD34þ grafts contained a median of 14.2� 106/kg CD34þ cells (range, 5.4 � 106/kg to39 � 106/kg), 2�105/kg CD3þ cells, and 0.19� 106/kg CD19þ cells.

Between Day 259 and Day 211 before thetransplantation, 60 mg/kg daily hydroxyureaand 3 mg/kg daily azathioprine were adminis-tered to eradicate marrow; and growth factors,granulocyte colony-stimulating factor, anderythropoietin were given twice weekly to main-tain stem cell proliferation in the face of hyper-transfusion, thereby facilitating the effect of thehydroxyurea. Fludarabine was administered at adosage of 30 mg/m2 per day from Day 217through Day 213. Starting on Day 210, thefirst 17 patients received 14 doses of 1 mg/kgoral busulfan (BU), three times daily over 4 d(total dose of 14 mg/kg over 4 d). The following14 patients received a corresponding dose of BU,given intravenously, followed by intravenous CYat a dose of 50 mg/kg daily on each of the next4 d (total dose of 200 mg/kg); 10 mg/kg thio-tepa and 12.5 mg/kg anti-thymocyte globulin(ATG-Fresenius S) were given daily from Days

G. Lucarelli et al.


ww

w.p

ersp

ecti

vesi

nm

edic

ine.

org



25 to 22. All patients received cyclosporine forGVHD prophylaxis for the first 2 mo posttrans-plantation.

All patients showed donor chimerism byDay 14 after HSCT. A granulocyte count of.500/mL occurred after a median time of 13d (range of 11–17). Seven patients rejectedtheir grafts and survived with thalassemia; threepatients showed early mixed chimerism, whichwas deemed persistent when observed at 14, 38,and 42 mo after the transplants. In 19 cases, thetransplantation was successful with complete al-logeneic reconstitution. Two patients died fromtransplantation-related causes; one died on Dayþ114 of Epstein-Barr virus cerebral lymphoma,and the other died on Day þ92 from CMVpneumonia. All of the 22 cured children are nolonger transfusion-dependent, with hemoglo-bin levels ranging from 10.3 g/dL to 13.8 g/dL(Sodani et al. 2010, 2011).

Mixed Chimerism following HSCTfor Thalassemia

Mixed hematopoietic chimerism (MC) is aninteresting phenomenon that sometimes occursafter HSCT for thalassemia. The incidenceof MC was evaluated in 335 patients who re-ceived BMT from HLA-matched family donorsfor thalassemia; it was found to be 32.2% at 2mo after transplant (Lucarelli et al. 2002). Ofthe 227 patients with complete donor chime-rism, none rejected their grafts, whereas graftloss occurred in 35 of 108 patients (32.4%)with MC, indicating that MC is a risk factorfor graft rejection in thalassemia patients. Thepercentage of residual host hematopoietic cells(RHCs) determined at 2 mo after transplantwas predictive for graft rejection, with nearlyall patients experiencing graft rejection whenRHCs exceeded 25%. The risk of graft rejectionwas only 13% in patients with ,10% RHCs andwas 41% in patients with 10%–25% RHCs (An-dreani et al. 1996, 2000). Unlike hematologicalmalignancies in which residual host cells arepredictive of relapse, patients with thalassemiacan have lifelong stable mixed chimerism with-out rejection following transplantation. Of pa-tients receiving BMT for thalassemia following

myeloablative conditioning, 10% became per-sistent mixed chimeras and became transfusionindependent, suggesting that once donor-hosttolerance is established, a limited number ofengrafted donor cells might be sufficient to pro-vide significant improvement of disease pheno-type in patients with thalassemia major.

We recently published a paper describingfour long-term transplanted patients affectedby hemoglobinopathies (Andreani et al. 2011a).These patients are characterized by the presenceof few donor-engrafted nucleated cells, bothin the peripheral blood and in the bone mar-row; the majority of the erythrocytes were ofdonor origin. Moreover, by analyzing singularlypicked-up burst-forming unit erythroid colo-nies, we showed that the proportion of donor-derived erythroid precursors was equivalent tothat observed in the mature nucleated cells, rath-er than that of the red blood cells. These resultssuggest that in patients characterized by the pres-ence of PMC after HSCT, a selective advantage ofthe maturation of donor erythroid precursorsmight successfully contrast the problems associ-ated with the recipient ineffective erythropoiesis(Andreani et al. 2011b).

HEMATOPOIETIC STEM CELLTRANSPLANTATION FOR SICKLE CELLDISEASE

Sickle cell disease (SCD) is associated with con-siderable morbidity and premature mortality,and HSCT is the only available therapy with cu-rative intent. Outcomes of HSCT from matchedsibling donors in pediatric patients have beenexcellent; however, the applicability of HSCThas been limited by lack of matched siblingdonors and concerns regarding regimen-relatedtoxicity. A reduced-intensity conditioning reg-imen has the potential to decrease regimen-related morbidity and mortality. Improved un-derstanding of the typical sequence of compli-cations and the effects of treatments such ashydroxyurea and blood transfusions, as well asthe impact of transplantation on organ damage,are likely to influence the timing and indica-tions for transplantation.



ww

w.p

ersp

ecti

vesi

nm

edic

ine.

org



Following a demonstration of successful useof the combination of busulfan and cyclopho-sphamide in transplant for thalassemia, thisprotocol was applied to the transplant prepara-tion for treating the other genetic hemoglobin-opathy, sickle cell anemia, in 1986 (Vermylenet al. 1993). We first performed a transplant ina 16-yr-old African girl with SCA in 1993 (Giar-dini et al. 1993). Since then, there has been slowprogress in stem cell transplantation for SCA,with only approximately 250 transplants per-formed to date worldwide, mainly reported inmulticenter studies (Table 2). A major reasonfor this very conservative approach to usingtransplantation for SCA treatment is the ab-sence of universally accepted indications fortransplantation.

Transplantation has been offered primarilyto younger patients with overt symptomatology.Most patients have received an HSCT followinga preparative regimen with 14–16 mg/kg busul-fan and 200 mg/kg cyclophosphamide. Addi-tional immunosuppressive agents sometimesinclude equine antithymocyte globulin, rabbitantithymocyte globulin, antilymphocyte globu-lin, or total lymphoid irradiation. CyclosporineA, alone or in combination with methylprednis-olone or methotrexate, has been used for post-transplant graft-versus-host disease (GVHD)prophylaxis. The results of HSCT after myelo-ablative conditioning in children have beenvery encouraging, with disease-free survival inmost studies of �85% and a TRM rate of ,10%(Table 2). The results of transplantation mightbe even better if HSCT were performed in chil-dren before the development of irreversible sick-le vasculopathy-related complications (Vermy-len et al. 1998; Lucarelli et al. 2011).

The experience of cord blood transplanta-tion (CBT) has been limited in patients withSCD, with a total of 33 cases reported, two-thirds of whom were in the related donor set-ting (Hsieh et al. 2011). The majority of relatedCBT have been 6/6 HLA-matched. In unrelatedCBT, although more feasible for pediatric pa-tients on the basis of the availability of 4/6HLA-matched cord blood unit, it appears tobe associated with a great risk of graft rejectionand GVHD. The largest study consisting of sev-en patients reported one death, an EFS of 43%,five patients developed acute GVHD, and onepatient chronic GVHD (Adamkiewicz et al.2007). Moreover, CBT for adults is limited bythe necessary total nucleated cell count per kilo-gram of body weight for engraftment to occur.At this time, transplant should be safelyextendedto patients with SCD using HLA identical do-nors, minimizing morbidity and mortality. Theattainment of tolerance should allow extensionof this curative approach to alternative donorsources.

The Rome Experience in HSCT for SickleCell Disease

A total of 11 SCA patients (Lucarelli et al. 2011)with a median age of 12 yr (range of 2–16)received hematopoietic stem cell transplanta-tions from HLA-identical, related donors, fol-lowing a myeloablative conditioning regimen.Indications for transplantation were vaso-oc-clusive crisis, acute chest syndrome, avascularbone necrosis, chronic red blood cell transfu-sions, or hemorrhagic stroke. Five patients re-ceived preconditioning cytoreductive/immu-nosuppressive chemotherapy with hydroxyurea,

Table 2. Worldwide reported outcomes of myeloablative HSCT for sickle cell disease

n ¼ 50

(Vermylen

et al. 1998)

n ¼ 50

(Walters

et al. 2000 )

n ¼ 11

(Locatelli

et al. 2003)

n ¼ 67

(Panepinto

et al. 2007)

n ¼ 87

(Bernaudin

et al. 2007)

n ¼ 11

(Lucarelli

et al. 2011)

OS 93% 94% 100% 97% 92% 90%DFS 85% 84% 90% 85% 86% 90%Rejection 10% 10% 9% 13% 5% 10%

Abbreviations: OS, overall survival; DFS, disease-free survival.

G. Lucarelli et al.


ww

w.p

ersp

ecti

vesi

nm

edic

ine.

org



azathioprine, and fludarabine, as describedelsewhere (Lucarelli et al. 2011). The remainingsix patients were prepared for the transplantwith busulfan, cyclophosphamide, and anti-thymocyte globulin. They received cyclosporine(CSA), low-dose methylprednisolone, and ashort course of MTX as GVHD prophylaxis.All patients had sustained engraftment withdonor chimerism of 100%, except for one pa-tient, who showed a persistent MC with 25% ofdonor cells. Starting 2 mo after transplantation,this patient showed a progressive decrease indonor chimerism until 25%; she had stablemixed chimerism for more than 4 yr withoutSCD-related events and was transfusion inde-pendent (Andreani et al. 2011a). All of the re-maining patients had full donor chimerism inlymphoid and myeloid lineages. Seven patientsdeveloped grade 2 acute GVHD. The only pa-tient who developed grade 3 GVHD had re-ceived a peripheral blood stem cell transplanta-tion. All of the patients responded promptly tothe steroid treatment given to control the acuteGVHD. Four out of 11 patients had mild skinchronic GVHD that resolved completely follow-ing steroid use. Ten out of 11 patients survivedwithout sickle cell disease, with Lansky/Karnof-sky scores of 100. Median follow-up for survi-vors was 43 mo (range of 12–68 mo). One pa-tient died 1 yr after transplantation in her homecountry; she was on tapering immunosuppres-sive treatment for chronic skin GVHD and diedof pneumonia. The probabilities of survival,SCA-free survival, and transplant-related mor-tality 5 yr after transplant were 90%, 90%, and10%, respectively. After transplantation, noneof the patients had complications typical ofSCA, such as pains, stroke, or acute chest syn-drome.

In sickle cell trait carriers, the use of G-CSFfor peripheral stem cell mobilization may in-duce HbS polymerization and red blood cellsickling, and therefore transform a relativelystable steady state into a cascade of events re-sulting in a sickle cell crisis and, in severe cases,multi-organ dysfunction. We suggest the useof BM-derived HSCs versus G-CSF-mobilizedHSCs especially when the donor is an HbS traitcarrier.

Mixed Chimerism following MyeloablativeHSCT for SCA

A subgroup of patients who undergo HSCT forSCA develop lifelong stable MC once donor–host tolerance is established (Andreani et al.1996, 2000). Walters et al. (2001) observed sta-ble MC in 13 out of 50 patients (26%) whoshowed SCD-free survival for a median dura-tion of 6.9 yr (range of 4.2–13 yr). Of note,among these patients, five had mixed donor chi-merism of ,75% (range of 11%–74%), andnone of them developed sickle cell-related com-plications during a follow-up period that rangedfrom 22 to 70.2 mo (median of 36.3 mo).

Recently we reported the clinical course of a6-yr-old girl who received a bone marrow trans-plant for sickle cell anemia (M Marziali, A Isgro,D Fraboni, et al., in prep.). Four years after trans-plant, the patient showed stable mixed chime-rism, as evidenced by a 39% bone marrow (BM)level of donor-origin erythroid precursors, and80% donor-origin red blood cells (RBCs) in pe-ripheral blood (PB). The patient received BMfrom her HLA-matched sister (Hb AA) after aconditioning regimen of 14 mg/kg BU, 200 mg/kg CY, and 10 mg/kg anti-thymocyte globulin(ATG). For prophylaxis against graft-versus-hostdisease (GVHD), the patient received cyclospor-ine (starting on Day 21) and a short course ofmethotrexate (MTX; 10 mg/m2 on posttrans-plant Days 1, 3, and 6 with folinic acid rescue).The course after allogeneic hematopoietic stemcell transplantation was uneventful, with rapidhematologic engraftment and no signs of acuteor chronic GVHD. Molecular analysis of sortedcell subgroups revealed MC in nucleated cells,CD34þ progenitors, and RBCs in the PB andBM. Four years after transplantation, the levelof donor NCs was 39% in PB and 36% at theBM level, in parallel to a very high proportion ofdonor-derived RBCs (80%) in PB. The propor-tions of donor-derived RBCs and BFU-E at theBM level were 40% and 46%, respectively, indi-cating the presence of quantitatively different redcell/nucleated cell chimerism. A possible expla-nation for the presence of a greater proportion ofdonor-derived RBC may be found in the im-proved survival of donor erythroid precursors,



ww

w.p

ersp

ecti

vesi

nm

edic

ine.

org



compared with the host counterparts that mightbe destroyed during ineffective erythropoiesis.

New Approach to Overcome CurrentLimitations: Reduced-Intensity ConditioningRegimens and Non-Myeloablative Regimens

A frequently encountered barrier to marrowtransplantation for SCD treatment is lack ofan HLA-identical donor. Another major limi-tation of current approaches is the high regi-men-related toxicity that has stopped HSCTfrom being offered to patients older than 16 yrof age or with advanced organ damage. At-tempts to reduce the intensity of preparativeregimens for SCD patients have been based ontwo approaches. One is to produce myeloabla-tion, which requires donor marrow infusion forhematopoietic recovery. The second approach isto not eradicate host hematopoiesis, whichwould allow hematopoietic recovery even with-out donor stem cell infusion. Sustained engraft-ment and alleviation of clinical phenotype havebeen reported in SCD patients following a re-duced-intensity conditioning regimen; Howev-er, HSCTusing non-myeloablative regimens, al-though well tolerated, has generally not resultedin stable engraftment.

In a recent study, seven patients (18 yror younger) with high-risk SCD were given abone marrow graft from an HLA-matched sib-ling donor (Krishnamurti et al. 2008). The con-ditioning regimen consisted of a total dose of6.4 mg/kg intravenous or 8 mg/kg oral BU, atotal dose of 175 mg/m2 FLU, a total dose of130 mg/kg equine ATG, and TLI 500 cGy. Reg-imen-related toxicity was minimal. All but onepatient had sustained engraftment. None of thepatients developed greater than grade 2 acuteGVHD or extensive chronic GVHD. Five outof seven patients developed MC. At a follow-up of 2–8.5 yr after HSCT, all patients were alive,and six of the seven patients had no laboratoryorclinical evidence of disease. These data are en-couraging in terms of sustained engraftmentwith minimal toxicity. On the other hand, un-favorable outcomes have been reported from us-ing a reduced-intensity conditioning regimen totreat adults with SCD (Van Beisen et al. 2000).

The rationale for lower-intensity condition-ing regimens in hemoglobinopathies stemsfrom historical experience indicating that stableMC is sufficient for cure (Nesci et al. 1998; An-dreani et al. 2000). Thus, myeloablation maynot be necessary for a successful outcome. In aretrospective multicenter series, Iannone et al.(2003) described a non-myeloablative HSCTapproach in six pediatric patients with SCDand one with thalassemia. This very-low-inten-sity conditioning regimen consisted of minimalintensity treatment with FLU/2 Gy TBI. Twopatients also received ATG. Regimen-relatedtoxicity was minimal, but the treatment resultedin only transient donor engraftment in six ofseven patients, suggesting that more intensiveconditioning is required in individuals withpre-HSCT transfusion exposures.

More recently, a novel non-myeloablativeconditioning regimen was reported to be suc-cessful in adults with SCD (Hsieh et al. 2009).Ten adults with a median age of 26 yr (rangeof 16–45 yr) and severe SCD underwent non-myeloablative transplantation with PBSC fromHLA-matched siblings. The conditioning regi-men included a total dose of 1 mg/kg alemtuzu-mab on Days 7–3 and TBI of 300 cGy on Day2. As GVHD prophylaxis, recipients received si-rolimus initiated on Day 1 at a dose of 5 mg every4 h for three doses, then 5 mg daily. Patients re-ceived 5.51 � 106 to 23.8 � 106 CD34þ cells/kg. Nine of 10 patients had sustained engraft-ment, and one had rejection. No patient died.None of the patients had full donor chimerismin both lymphoid and myeloid lines. Acute orchronic GVHD did not develop in any patient. Ifstudies using this method can show sustainedmixed donor chimerism with low regimen-relat-ed toxicity, the paradigm that only severely af-fected patients with SCD be offered HSCT mayshift, thus allowing more low-risk patients theoption of cure without long-term morbidity.

Sustained and Full Fetal HemoglobinProduction after Failure of Bone MarrowTransplant

HbF increase has been reported in three trans-planted SCD patients after graft failure (Ferster

G. Lucarelli et al.


ww

w.p

ersp

ecti

vesi

nm

edic

ine.

org



et al. 1995). Recent clinical observations per-formed by our group strongly support suchnovel approaches. The reactivation of HbF syn-thesis has been documented after bone marrowtransplant (BMT) failure and autologous re-constitution in patients affected by b-hemoglo-bin disorders (Paciaroni et al. 2009).

It is possible that after myeloablative treat-ment, the vast majority of committed auto-logous erythroid progenitors were destroyed,and only autologous hematopoietic stem cellssurvived in marrow niches, restoring the onto-logical HbF regulation mechanisms (Bank 2006).Alternatively, the increased HbF may be due to anew status, induced by modulations and influ-ences operating in the setting of the BMT (mi-croenvironment and stage-specific cytokines),that reprograms globin-gene expression or se-lects “fetal clone” proliferation. These casesstrongly support further research efforts tofind ways to reverse the hemoglobin switchand induce HbF production in adults in orderto treat b-hemoglobin disorders.

CONCLUSIONS

The only radical cure for homozygous thalasse-mia is to transplant bone marrow from an HLA-identical donor who is normal or heterozygousfor thalassemia, which is capable of producingand maintaining a normal hemoglobin level inthe recipient. All thalassemic patients, togetherwith their parents and siblings, should be HLAtyped for this purpose. When an HLA donor isavailable, we believe that bone marrow trans-plantation is mandatory in thalassemic patientsof class 1 and 2, as well as for those of class 3who are aged ,17 yr; adult patients should alsobe offered this possibility of a cure, but 30% willdie of transplant-related mortality. Patientswithout matched family or unrelated donorscould benefit from haploidentical mother-to-child transplantation, which has shown encour-aging results, although it is still in the experi-mental phase.

A high cure rate has also been achieved byperforming HSCTon children with SCA follow-ing current myeloablative conditioning proto-cols. Again, we think that HLA typing should be

performed for all family members of an SCAchild. If a genotypically identical sibling or phe-notypically identical parent is identified, hema-topoietic stem cell transplantation should beperformed in all SCA patients aged ,17 yr,before the major complications of the diseaseaffect the child. The novel non-myeloablativetransplantation protocol for SCD patients couldserve as a platform for its wide application, notonly in adults but also in high-risk pediatricpatients with SCD.

REFERENCES�Reference is also in this collection.

Adamkiewicz TV, Szabolcs P, Haight A, Baker KS, Staba S,Kedar A, Chiange KY, Krishnamurti L, Boyer MW, Kurtz-berg J, et al. 2007. Unrelated cord blood transplantationin children with sickle cell disease: Review of four-centerexperience. Pediatr Transplant 11: 641–644.

Andreani M, Manna M, Lucarelli G, Tonucci P, Agostinelli F,Ripalti M, Rapa S, Talevi N, Galimberti M, Nesci S. 1996.Persistence of mixed chimerism in patients transplantedfor the treatment of thalassemia. Blood 87: 3494–3499.

Andreani M, Nesci S, Lucarelli G, Tonucci P, Rapa S, Ange-lucci E, Persini B, Agostinelli F, Donati M, Manna M.2000. Long-term survival of ex-thalassemic patientswith persistent mixed cjimerism after marrow transplan-tation. Bone Marrow Transplant 25: 401–404.

Andreani M, Testi M, Battarra M, Lucarelli G. 2011a. Splitchimerism between nucleated and red blood cells afterbone marrow transplantation for haemoglobinopathies.Chimerism 2: 21–22.

Andreani M, Testi M, Gaziev J, Condello R, Bontadini A,Tazzari PL, Ricci F, De Felice L, Agostini F, Fraboni D, etal. 2011b. Quantitatively different red cell/nucleated cellchimerism in patients with long-term, persistent hema-topoietic mixed chimerism after bone marrow transplan-tation for thalassemia major or sickle cell disease. Hae-matologica 96: 128–133.

Bank A. 2006. Regulation of human fetal haemoglobin: Newplayers, new complexities. Blood 107: 435–443.

Bernaudin F, Socie G, Kuentz M, Chevret S, Duval M, Ber-trand Y, Vannier JP, Yakouben K, Thuret I, Bordigoni P, etal. 2007. Long-term results of related myeloablative stem-cell transplantation to cure sickle cell disease. Blood 110:2749–2756.

Ferster A, Corazza F, Vertongen F, Bujan W, Devalck C, Fon-du P, Cochaux P, Lambermont M, Khaladji Z, Sariban E.1995. Transplanted sickle-cell disease patients with autol-ogous bone marrow recovery after graft failure developincreased levels of fetal haemoglobin which corrects dis-ease severity. Brit J Haematol 90: 804–808.

Gaziev J, Sodani P, Polchi P, Andreani M, Lucarelli G. 2005.Bone marrow transplantation in adults with thalassemia.Treatment and long-term follow-up. Ann NY Acad Sci1054: 196–205.



ww

w.p

ersp

ecti

vesi

nm

edic

ine.

org



Ghavamzadeh A, Nasseri P, Eshraghian MR, Jahani M, Bay-bordi I, Nateghi J, Khodabandeh A, Sadjadi AR, Mohyed-din M, Khademi Y. 1997. Prognostic factors in bone mar-row transplantation forb thalassemia major: Experiencesfrom Iran. Bone Marrow Transplant 22: 1167–1169.

Giardini C, Galimberti M, Lucarelli G, Polchi P, Angelucci E,Baronciani F, Agostinelli F, Giorni G, Muretto P. 1993.Bone marrow transplantation in sickle-cell anemia inPesaro. Bone Marrow Transplant 12: 122–123.

Hobbs JR, Hugh-Jones K, Shaw PJ, Downie CJC, William-son S. 1986. Engraftment rates related to busulphan andcyclophosphamide dosages for displacement bone mar-row transplants in fifty children. Bone Marrow Transplant1: 201–208.

Hongeng S, Pakakasama S, Chuansumrit A, Sirachainan N,Kitpoka P, Udomsubpayakul U, Ungkanont A, Jootar S.2006. Outcomes of transplantation with related and un-related-donor stem cells in children with severe thalasse-mia. Biol Blood Marrow Transplant 12: 683–687.

Hsieh MM, Kang EM, Fitzhugh CD, et al. 2009. Allogeneichematopoietic stem-cell transplantation for sickle celldisease. N Engl J Med 361: 2309–2317.

Hsieh MM, Fitzhugh CD, Tisdale JF. 2011. Allogeneic he-matopoietic stem cell transplantation for sickle cell dis-ease: The time is now. Blood 118: 1197–1207.

Iannone R, Casella JF, Fuchs EJ, Chen AR, Jones RJ, WoolfreyA, Amylon M, Sullivan KM, Storb RF, Walters MC. 2003.Results of minimally toxic nonmyeloablative transplan-tation in patients with sicule cell anemia and b-thalasse-mia. Biol Blood Marrow Transplant 9: 519–528.

Isgro A, Gaziev J, Sodani P, Lucarelli G. 2010. Progress inhematopoietic stem cell transplantation as allogeneic cel-lular gene therapy in thalassemia. Ann NY Acad Sci 1202:149–154.

Kapoor N, Kirkpatrick D, Blaese RM, Oleske J, HilgartnerMH, Changanti RS, Good RA, O’Reilly RJ. 1981. Recon-stitution of normal megakaryocytopoiesis and immuno-logic functions in Wiskott-Aldrich syndrome by marrowtransplantation following myeloablation and immuno-suppression with busulfan and cyclophosphamide. Blood57: 692–696.

Krishnamurti L, Kharbanda S, Biernacki M, Zhang W, BakerKS, Wagner JE, Wu CJ. 2008. Stable long-term donorengraftment following reduced-intensity hematopoieticcell transplantation for sickle cell disease. Biol Blood Mar-row Transplant 14: 1270–1278.

La Nasa G, Argiolu F, Giardini C, Pession A, Fagioli F, CaocciG, Vacca A, De Stefano P, Piras E, Ledda A, et al. 2005a.Unrelated bone marrow transplantation for b-thalasse-mia patients: The experience of the Italian Bone MarrowTranplant Group. Ann NY Acad Sci 1054: 186–195.

La Nasa G, Caocci G, Argiolu F, Giardini C, Locatelli F, VaccaA, Ororfino MG, Piras E, Addari MC, Ledda A, et al.2005b. Unrelated donor stem cell transplantation in adultpatients with thalassemia. Bone Marrow Transplant 36:971–975.

Locatelli F, Rocha V, Reed W, Bernaudin F, Ertem M, Grafa-kos S, Brichard B, Li X, Nagler A, Giorgiani G, et al. 2003.Related umbilical cord blood transplantation in patientswith thalassemia and sickle cell disease. Blood 101:2137–2143.

Lucarelli G, Gaziev J. 2008. Advances in the allogeneic trans-plantation for thalassemia. Blood Rev 22: 53–63.

Lucarelli G, Clift RA, Galimberti M, Angelucci E, GiardiniC, Baronciani D, Polchi P, Andreani M, Gazlev D, Erer B,et al. 1990. Bone marrow transplantation in patients withthalassemia. N Engl J Med 322: 417–421.

Lucarelli G, Galimberti M, Polchi P, Angelucci E, BaroncianiD, Durazzi SM, Giardini C, Albertini F, Clift RA. 1992.Bone marrow transplantation in adult thalassemia. Blood80: 1603–1607.

Lucarelli G, Clift RA, Galimberti M, Angelucci E, GiardiniC, Baronciani D, Polchi P, Andreani M, Gazlev D, Erer B,et al. 1999. Bone marrow transplantation in adult thalas-semic patients. Blood 93: 1164–1167.

Lucarelli G, Andreani M, Angelucci E. 2002. The cure ofthalassemia by bone marrow transplantation. Blood Rev16: 81–85.

Lucarelli G, Gaziev J, Isgro A, Sodani P, Paciaroni K, AlfieriC, De Angelis G, Marziali M, Simone MD, Gallucci C, etal. 2011. Allogeneic cellular gene therapy in hemoglobin-opathies—evaluation of hematopoietic SCT in sickle cellanemia. Bone Marrow Transplant doi: 10.1038/bmt.2011.79.

Nesci S, Manna M, Lucarelli G, Tonucci P, Donati M, BuffiO, Agostinelli F, Andreani M. 1998. Mixed chimerismafter bone marrow transplantation in thalassemia. AnnNY Acad Sci 850: 495–497.

Paciaroni K, Gallucci C, De Angelis G, Alfieri C, Roveda A,Lucarelli G. 2009. Sustained and full fetal hemoglobinproduction after failure of bone marrow transplant in apatient homozygous forb0-thalassemia: A clinical remis-sion despite genetic disease and transplant rejection. Am JHematol 84: 372–373.

Panepinto JA, Walters MC, Carreras J, Marsh J, Bredeson CN,Gale RP, Hale GA, Horan J, Hows JM, Klein JP, et al.2007. Matched-related donor transplantation for sicklecell disease: Report from the Center for InternationalBlood and Transplant Research. Br J Haematol 137:479–485.

Parkman R, Rappeport J, Geha R, Belli J, Cassady R, Levey R,Nathan DG, Rosen FS. 1978. Complete correction of theWiskott-Aldrich syndrome by allogeneic bone-marrowtransplantation. N Engl J Med 298: 921–927.

Sabloff M, Chandy M, Wang Z, Logan BR, Ghavamzadeh A,Li CK, Irfan SM, Bredeson CN, Cowan MJ, Gale RP, et al.2011. HLA-matched sibling bone marrow transplanta-tion for b-thalassemia major. Blood 117: 1745–1750.

Sodani P, Isgro A, Gaziev J, Polchi P, Paciaroni K, Marziali M,Simone MD, Roveda A, Montuoro A, Alfieri C, et al.2010. Purified T-depleted, CD34þ peripheral bloodand bone marrow cell transplantation from haploiden-tical mother to child with thalassemia. Blood 115:1296–1302 (Erratum in Blood 116: 1627).

Sodani P, Isgro A, Gaziev J, Paciaroni K, Marziali M, SimoneMD, Roveda A, De Angelis G, Gallucci C, Torelli F, et al.2011. T cell-depleted HLA-haploidentical stem cell trans-plantation in thalassemia young patients. Pediatr Rep 22:e13.

Storb R, Champlin RE. 1991. Bone marrow transplantationfor severe aplastic anemia. Bone Marrow Transplant 8:69–72.

G. Lucarelli et al.


ww

w.p

ersp

ecti

vesi

nm

edic

ine.

org



Storb R, Weiden PL, Graham TC, Lerner KG, Nelson N,Thomas ED. 1977. Hemopoietic grafts between DLA-identical canine littermates following dimethyl myleran.Evidence for resistance to grafts not associated with DLAand abrogated by antithymocyte serum. Transplantation24: 349–357.

Thomas ED, Storb R, Fefer A, Slichter SJ, Bryant JI, BucknerCD, Neiman PE, Clift RA, Funk DD, Lerner KE. 1972.Aplastic anaemia treated by marrow transplantation.Lancet 1: 284–289.

van Beisen K, Bartholomew A, Stock W, Peace D, Devine S,Sher D, Sosman J, Chen YH, Koshy M, Hoffman R. 2000.Fludarabine-based conditioning regimen for allogeneictransplantation in adult with sickle cell disease. BoneMarrow Transplant 26: 445–449.

Vermylen C, Cornu G. 1993. Bone marrow transplantationin sickle cell anemia. Blood Rev 7: 1–3.

Vermylen C, Cornu G, Ferster A, Brichard B, Ninane J, Fer-rant A, Zenebergh A, Maes P, Dhooge C, Benoit Y, et al.

1998. Haematopoietic stem cell transplantation for sicklecell anaemia: The first 50 patients transplanted in Bel-gium. Bone Marrow Transplant 22: 1–6.

Walters MC, Storb R, Patience M, Leisenring W, Taylor T,Sanders JE, Buchanan GE, Rogers ZR, Dinndorf P,Davies SC, et al. 2000. Impact of bone marrow trans-plantation for symptomatic sickle cell disease: An in-terim report. Multicenter investigation of bone marrowtransplantation for sickle cell disease. Blood 95: 1918–1924.

Walters MC, Patience M, Leisenring W, Rogers ZR, AquinoVM, Buchanan GR, Roberts IA, Yeager AM, Hsu L,Adamkiewicz T, et al. 2001. Stable mixed hematopoieticchimerism after bone marrow transplantation for sicklecell anemia. Biol Blood Marrow Transplant 7: 665–673.

� Williams T. 2012. World distribution, population genetics,and burden of thalassemic and sickle cell syndromes.Cold Spring Harb Perspect Med doi: 10.1101/cshper-spect.a011692.



ww

w.p

ersp

ecti

vesi

nm

edic

ine.

org



4, 20122012; doi: 10.1101/cshperspect.a011825 originally published online AprilCold Spring Harb Perspect Med

Guido Lucarelli, Antonella Isgrò, Pietro Sodani and Javid Gaziev Cell AnemiaHematopoietic Stem Cell Transplantation in Thalassemia and Sickle

Subject Collection Hemoglobin and Its Diseases

The Natural History of Sickle Cell DiseaseGraham R. Serjeant Hemoglobin Synthesis

Transcriptional Mechanisms Underlying

Nathaniel J. Pope, et al.Koichi R. Katsumura, Andrew W. DeVilbiss,

Current Management of Sickle Cell Anemia

WarePatrick T. McGann, Alecia C. Nero and Russell E. Disease

Iron Deficiency Anemia: A Common and Curable

Jeffery L. Miller

TherapiesTargetedNew Disease Models Leading the Way to

Cell-Free Hemoglobin and Its Scavenger Proteins:

Dominik J. Schaer and Paul W. Buehler

Management of the ThalassemiasNancy F. Olivieri and Gary M. Brittenham

-ThalassemiaαClinical Manifestations of Elliott P. Vichinsky

-ThalassemiaβThe Molecular Basis of Swee Lay Thein

Erythroid Heme Biosynthesis and Its DisordersHarry A. Dailey and Peter N. Meissner

Erythropoiesis: Development and DifferentiationElaine Dzierzak and Sjaak Philipsen

Clinical CorrelatesHemoglobin Variants: Biochemical Properties and

Gell, et al.Christopher S. Thom, Claire F. Dickson, David A.

ErythropoietinH. Franklin Bunn

The Prevention of ThalassemiaAntonio Cao and Yuet Wai Kan

Classification of the Disorders of HemoglobinBernard G. Forget and H. Franklin Bunn

The Switch from Fetal to Adult HemoglobinVijay G. Sankaran and Stuart H. Orkin

-ThalassemiaαThe Molecular Basis of Douglas R. Higgs

http://perspectivesinmedicine.cshlp.org/cgi/collection/ For additional articles in this collection, see

Copyright © 2012 Cold Spring Harbor Laboratory Press; all rights reserved


http://perspectivesinmedicine.cshlp.org/cgi/collection/

http://perspectivesinmedicine.cshlp.org/cgi/collection/


hematopoietic stem cell transplantation in...

Documents