Paul L. Martin, 1 Shelly L. Carter, 2 Nancy A. Kernan, 3 Indira Sahdev, 4 Donna Wall, 5 Daniel Pietryga, 6 John E. Wagner, 7 Joanne Kurtzberg 1

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1 Biology of Blood and Marrow Transplantation 12: (2006) 2006 American Society for Blood and Marrow Transplantation /06/ $32.00/0 doi: /j.bbmt Results of the Cord Blood Transplantation Study (COBLT): Outcomes of Unrelated Donor Umbilical Cord Blood Transplantation in Pediatric Patients with Lysosomal and Peroxisomal Storage Diseases Paul L. Martin, 1 Shelly L. Carter, 2 Nancy A. Kernan, 3 Indira Sahdev, 4 Donna Wall, 5 Daniel Pietryga, 6 John E. Wagner, 7 Joanne Kurtzberg 1 1 Duke University Medical Center, Durham, North Carolina; 2 The EMMES Corporation, Rockville, Maryland; 3 Memorial Sloan-Kettering Cancer Center, New York, New York; 4 Schneider Children s Hospital, New Hyde Park, New York; 5 Texas Transplant Institute, San Antonio, Texas; 6 DeVos Children s Hospital, Grand Rapids, Michigan; 7 University of Minnesota, Minneapolis, Minnesota Correspondence and reprint requests: Joanne Kurtzberg, MD, Duke University Medical Center, Box 3350, Durham, NC ( kurtz001@mc.duke.edu). Received May 19, 2005; accepted September 23, 2005 ABSTRACT The Cord Blood Transplantation Study (COBLT), sponsored by the National Heart, Lung, and Blood Institute, is a phase II multicenter study designed to evaluate the use of cord blood in allogeneic transplantation. In this report, we evaluated the outcomes of cord blood transplantation in 69 patients with lysosomal and peroxisomal storage diseases. Patients with mucopolysaccharidoses I to III, mucolipidoses (ML) II (n 36), adrenoleukodystrophy (n 8), metachromatic leukodystrophy (n 6), Krabbe disease (n 16), and Tay-Sachs disease (n 3) were enrolled between August 1999 and June All patients received the same preparative regimen, graft-versus-host disease (GVHD) prophylaxis, and supportive care. End points included survival, engraftment, GVHD, and toxicity. Sixty-nine patients (64% men; 81% white) with a median age of 1.8 years underwent transplantation with a median cell dose of /kg. One-year survival was 72% (95% confidence interval, 61%-83%). The cumulative incidence of neutrophil engraftment by day 42 was 78% (95% confidence interval, 67%-87%) at a median of 25 days. Grade II to IV acute GVHD occurred in 36% of patients. Cord blood donors are readily available for rapid transplantation. Cord blood transplantation should be considered as frontline therapy for young patients with lysosomal and peroxisomal storage diseases American Society for Blood and Marrow Transplantation KEY WORDS Cord blood transplantation Inborn errors of metabolism Lysosomal storage diseases Peroxisomal storage diseases INTRODUCTION Inherited metabolic storage diseases are a heterogeneous group of rare disorders due to mutations that cause deficiencies in various enzymes, thus resulting in an accumulation of toxic metabolites in various tissues. A subset of these conditions, lysosomal and peroxisomal storage diseases (LSDs), is characterized by severe neurologic deterioration and eventual death in the first or second decade of life. LSDs can be divided into the mucopolysaccharidoses, which are characterized by an accumulation of glycosaminoglycans, causing damage to the brain, heart, cornea, cartilage, liver, and other organs, and sphingolipidoses or leukodystrophies, which are characterized by defects in lysosomal acid hydrolases involved in sphingolipid catabolism, resulting in demyelination in the central and peripheral nervous systems. Allogeneic stem cell transplantation can prevent the progression of LSD symptoms by providing a continuous source of normal enzymes from engrafted donor cells, provided that the enzyme is produced by normal leukocytes [1]. Over the past 2 decades, allogeneic bone marrow transplantation from matched 184

2 CBT and Lysosomal Storage Diseases related carrier and noncarrier donors has been shown to be beneficial in 200 patients with Hurler syndrome [1-13] and others with adrenoleukodystrophy, metachromatic leukodystrophy [14-19], and Krabbe disease (globoid leukodystrophy) [20-22]. Many children with LSDs who could benefit from allogeneic transplantation therapy lack an appropriately matched donor. Partially HLA-matched, banked, unrelated donor umbilical cord blood can provide donor stem cells for transplantation of children with either malignancies or inherited diseases who lack suitable bone marrow donors [23-26]. Between 2000 and 2004, the Cord Blood Transplantation Study (COBLT), sponsored by the National Heart, Lung, and Blood Institute, National Institutes of Health, investigated the safety and feasibility of unrelated donor umbilical cord blood transplantation (CBT) in children and adults with lymphohematologic disorders and in 69 young children with LSDs. We describe in this article the results for the patients with LSD. MATERIALS AND METHODS Patients BB&MT Between August 1999 and June 2004, 69 children with LSDs who lacked HLA-matched related donors were treated with unrelated donor umbilical CBT. Although the COBLT study is a multicenter study, 67 of the 69 patients enrolled were treated at a single institution (Duke University). Thirty-one patients were consecutively entered on the LSD stratum of the COBLT protocol between August 1999 and July Once accrual was reached in the LSD stratum, the stratum was closed. Subsequent patients who met entry criteria could obtain a COBLT unit and be treated under the expanded access protocol (EAP). An additional 38 were consecutively entered under the EAP between July 2002 and June The criteria for study enrollment and the treatment plans were identical for both groups. The diagnosis of LSD was confirmed by the lack of specific enzyme activity in peripheral blood leukocytes or fibroblasts. Institutional review boards approved the protocols at each participating center, and the parents of each patient provided written informed consent before study enrollment. Results in children with Hurler syndrome (14 of whom were also included in this article) and Krabbe disease (4 patients in this article) have been previously reported [26,27]. Patients diagnosed with LSD whose developmental quotient, IQ, or clinical neurodevelopment examination demonstrated a level of functioning at which continuous life support would not be predicted to be required in the year after transplantation were eligible for enrollment. Patients with a Lansky performance score 50%, primary myelofibrosis, or suitable related donors were ineligible. Patients with prior allogeneic stem cell transplantation within 12 months or autologous transplantation within 6 months were excluded, as were individuals with uncontrolled bacterial, viral, or fungal infections. Multiorgan assessment performed before enrollment was required to demonstrate the following: serum creatinine normal for age or a creatinine clearance 50% of the lower limits of normal for age; aspartate aminotransferase 5 times the upper limits of normal and total serum bilirubin 2.5 mg/dl; cardiac status asymptomatic or left ventricular ejection fraction 40% that improved with exercise; and asymptomatic pulmonary function or carbon monoxide diffusion in the lung, forced expiratory volume in 1 second, and diffusion capacity 45% of predicted (corrected for hemoglobin) or oxygen saturation 85% on room air. The selected cord blood unit (CBU) was required to provide a minimum of total nucleated cells (NCs; before cryopreservation) per kilogram of recipient weight. Selection of Donors Searches for CBUs from unrelated donors were conducted by using low-/intermediate-resolution molecular typing for HLA matching class I (A and B) and high-resolution molecular typing for HLA-DRB1. The CBU was selected that provided the highest number of NCs and matched at a minimum of 4 of 6 HLA loci. A unit matched at 3 of 6 alleles could be selected if the match was based on high-resolution typing of HLA-A, -B, and -DRB1 and if there was at least 1 match per locus. In most cases (n 61), enzyme testing on a sample of the cord blood donor cells or plasma was tested for the enzyme lacking in the intended recipient to ensure that affected or carrier donors were not selected. Transplantation Procedure COBLT CBUs were thawed as described in the COBLT manual of procedures ( emmes.com/study/cord/). Cryopreserved CBUs from other banks were thawed and processed according to the thawing instructions provided by the cord blood bank. NCs, clonal hematopoietic progenitor cells, and CD3 and CD34 cells were counted, ABO and Rh typing were performed, cell viability was assessed, and bacterial and fungal cultures were obtained at the time of banking and again at the time of thawing and infusion of the cord blood donor cells. Cells were infused intravenously (IV) over 10 to 30 minutes after premedication with diphenhydramine, acetaminophen, and steroids. Conditioning Regimen All patients were prepared for transplantation with oral busulfan (20-40 mg/m 2 per dose, with dosing 185

3 P. L. Martin et al. based on patient age) for 16 doses on days 9 through 6, cyclophosphamide 50 mg/kg per dose IV on days 5 through 2, and antithymocyte globulin (ATG; Pharmacia & Upjohn, Kalamazoo, MI) 30 mg/kg per dose IV on days 3 through 1. First-dose pharmacokinetics were measured for busulfan, and a concentration steady state of 600 to 900 ng/ml was targeted. Phenytoin was given during busulfan therapy to prevent seizures, mesna was given during cyclophosphamide therapy to prevent hemorrhagic cystitis, and methylprednisolone, acetaminophen, and diphenhydramine were given before and during ATG therapy to prevent hypersensitivity reactions. The cord blood was infused on day 0. Prophylaxis against Graft-versus-Host Disease All patients received cyclosporine for 9 months and methylprednisolone for 2 to 3 months as prophylaxis against graft-versus-host disease (GVHD). Cyclosporine was dosed to maintain levels of 200 to 400 ng/ml. Corticosteroids were given at 1 mg/kg/d on days 0 to 4 and 2 mg/kg/d on day 5 until the minimum of day 21 or engraftment and were then tapered at 0.2 mg/kg/wk. Patients with grade I acute GVHD were treated with topical creams, whereas patients with grades II and above received a high-dose methylprednisolone pulse (500 mg/m 2 per dose IV every 12 hours four 4 doses). Most patients whose GVHD did not respond or whose disease recurred after the steroid pulse were changed to tacrolimus with or without daclizumab. Supportive Care All patients were hospitalized and maintained in reverse isolation under high-energy particulate air filtration and positive-pressure ventilation. Prophylaxis against Pneumocystis carinii, viral, and fungal infections was administered to all patients while they remained on immunosuppressive therapy. Empiric treatment with broad-spectrum IV antibiotics was started with the first episode of neutropenic fever and continued until neutropenia resolved. For immunoprophylaxis, IV immunoglobulin was administered weekly at a dose of 500 mg/kg per dose through posttransplantation day 100 and then monthly for most of the first posttransplantation year. Low-dose continuous infusion heparin was administered IV from the initiation of the preparative regimen through posttransplantation day 28 for prophylaxis against veno-occlusive disease. All patients were supported as needed with transfusions of leukocyte-depleted, irradiated packed red blood cells and platelets. Filgastrim (Amgen, Thousand Oaks, CA) was administered IV from day 0 until engraftment of donor cells. Study End Points and Statistical Analysis The primary end point of the study was survival at 180 days after transplantation. The secondary endpoints included engraftment (neutrophil and platelet), acute and chronic GVHD, and regimen-related morbidity and mortality. Neutrophil engraftment was defined as achieving an absolute neutrophil count (ANC) of at least 500/ L for 3 consecutive measurements on different days and demonstrated donor chimerism of 90%. Primary graft failure was defined as failure to attain neutrophil engraftment by day 42. Patients who survived to day 14 and died before neutrophil engraftment were classified as primary graft failures. Hematopoietic recovery was defined as an ANC of at least 500/ L for 3 consecutive measurements on different days without demonstrated donor chimerism 90%. Secondary graft failure was defined as a loss of neutrophil engraftment. The platelet engraftment date was defined as the first of 3 consecutive days with a platelet count / L unsupported by platelet transfusion for a minimum of 7 days. The time to neutrophil or platelet engraftment was defined as the time from transplantation to the first day of engraftment. Clinically significant infections were reported after transplantation by the site of infection, organism, and severity of infection. Readmissions after the initial discharge for the transplantation were reported by date and primary and secondary reasons for admission. The Bearman toxicity scale was used to report maximum regimen-related toxicity by day 42 after transplantation [28]. The staging of acute GVHD followed National Marrow Donor Program guidelines and included weekly capture of symptoms through day 100 and at days 120 and 150 with characterization of alternative causes. The grading of acute GVHD followed the GVHD consensus grading scheme [29]. An algorithm calculated the maximum GVHD clinical grade on the basis of the weekly organ staging in skin, upper and lower gastrointestinal tract, and liver. This calculated organ stage was decreased by 1 stage if a listed specific differential diagnosis was reported for either the gastrointestinal tract or liver. An independent panel reviewed weekly records and assigned each patient a final maximum grade; this is similar to the methods described by Weisdorf et al. [30]. Primary causes of death were reported by following the hierarchy developed for the Unrelated Donor Marrow Transplantation Trial ( com/study/tcd/). The hierarchy was developed by an expert panel and was based on refinements to the National Marrow Donor Program hierarchy for reporting causes of death. The hierarchy requires that graft failure or GVHD be reported as the primary cause of death, rather than another condition s leading to death, such as infection, organ failure, or hemor- 186

4 CBT and Lysosomal Storage Diseases rhage. It also provides rules for reporting secondary causes of death. Survival estimates were calculated by using the Kaplan-Meier method [31]. Testing for differences in survival between groups in the univariate analysis used the log-rank test. Variables considered in the univariate analysis included age, sex, ethnicity, donor ethnicity, primary disease, performance status, HLA matching, pretransplantation cytomegalovirus serostatus, weight, NC dose, CD34 dose, CD3 dose, and protocol. Cox proportional hazard models were used for multivariate analysis of survival [32]. Neutrophil and platelet recoveries, acute GVHD, and chronic GVHD were analyzed by using the cause-specific failure probability method (or cumulative incidence), in which death and second infusion were treated as competing risks [33]. The complement of the Kaplan-Meier (1 KM) estimate for these end points was used to calculate the cumulative probability. No imputation was used for any missing data. Frequency data in cross-tabulation tables were compared by using the 2 or Kruskal- Wallis test. All analyses were performed with SAS software, version 8.2 (SAS Institute Inc., Cary, NC). Retrospective HLA Typing High-resolution molecular typing for HLA-A and -B became available after the study was initiated. To determine the effect of this typing information on patient outcome, high-resolution typing for HLA-A, HLA-B, and HLA-DRB1 alleles was performed for 60 donor/recipient pairs (subsequently referred to as final HLA typing). The results of statistical analyses by using this information was compared with the results in which HLA-A and -B typing was performed with low-/intermediate-resolution HLA typing. RESULTS Patient Characteristics Sixty-nine patients were analyzed in this study. Table 1 shows the baseline characteristics of all patients. The age at the time of transplantation ranged from 0.1 to 11.7 years, with a median of 1.8 years. Two thirds of the patients were men, 19% were white, and one third had a Lansky performance score of 80. Twenty-five percent of patients were cytomegalovirus seropositive before transplantation. Using original HLA criteria (low- to intermediate-resolution molecular typing at class I A and B and high-resolution typing at HLA class II DRB1), 90% of patients received an HLA 5/6- or 4/6-matched graft. Forty-one percent of the HLA 4/6- matched and 26% of the HLA 5/6-matched grafts were downgraded by retrospective high-resolution molecular typing (Table 2). The median NC dose was , the CD3 dose was , and the CD34 cell dose was BB&MT As seen in Table 1, the characteristics of the patients on the LSD stratum were very similar to those of the patients on the EAP stratum. The LSD stratum showed slightly better performance status. The EAP cohort also had more men, more white patients, and slightly worse HLA matches. However, none of these differences was statistically significant. No patient with metachromatic leukodystrophy was treated in the LSD stratum, whereas 6 of the 38 patients in the EAP group had this diagnosis; similarly, only 2 patients in the LSD stratum had Sanfilippo syndrome, and 8 patients in the EAP group had this diagnosis (P.003). Overall survival at 180 days was 80% (95% confidence interval [CI], 71%-90%) and 72% (95% CI, 61%-83%) at 1 year. Long-term survival was 68% with a median follow-up time of 24.5 months (range, months; Figure 1A). Nonwhite ethnicity was significantly associated with lower survival (P.03; Figure 1B). It made little difference whether recipient or donor ethnicity was considered, because at the broad level of white versus nonwhite, there was a very high correlation between donor and recipient ethnicity. Patients on the EAP stratum had significantly worse survival than LSD stratum patients (Figure 1C; P.024). One-year survival was 81% (95% CI, 67%- 95%) for LSD stratum patients but only 64% (95% CI, 47%-80%) for EAP patients (Figure 1C). The explanation seems to be related to the lower performance status of patients on the EAP stratum (Table 1). The proportions with a Lansky score 80 were 16 (42%) of 38 for the EAP and 8 (26%) of 31 for the LSD stratum, and more patients with Sanfilippo and metachromatic leukodystrophy (MLD) were on the EAP (n 14 versus n 2 on the LSD stratum). Other factors, such as age, cell dose, CD34 cell dose, and performance status, did not exhibit a statistically significant association with survival. In contrast to what was observed in young children with malignancy who also received nonlimiting CD34 cell doses, the final HLA typing did not affect overall survival (Figure 1D). In a Cox proportional hazards regression model, only ethnicity and study cohort (LSD stratum versus EAP) had significant effects on survival. The primary cause of death (Table 3) was reported as acute GVHD (n 5), graft failure or autologous recovery (n 8), infection (bacterial, n 1; polyorganism, n 2), pulmonary failure (n 1), and other causes (progressive disease, n 2; refractory hemolytic anemia, n 1). Infection was a contributing cause of death for 10 patients whose primary cause of death was acute GVHD, graft failure, or autologous recovery. Eighteen of these 20 deaths occurred in the first 8 months after transplantation, and 2 additional deaths occurred at 13 and 21 months as a result of bacterial infection and autologous recovery followed by an aspergillus infection. 187

5 P. L. Martin et al. Table 1. Baseline Characteristics of 69 Cord Blood Recipients with Lysosomal Diseases LSD* EAP Total Variable Median Range Median Range Median Range Patient age (y) Patient weight (kg) Total viable NCs 3 ( 10 7 /kg) Infused NCs ( 10 7 /kg) CD34 cell dose ( 10 5 /kg) CD3 cell dose ( 10 7 /kg) N % N % N % Recipient sex Male Female Recipient ethnicity White Black Asian Hispanic Mixed/other/unknown Primary disease Hurler syndrome (MPS I) Hurler-Scheie syndrome Hunter syndrome (MPS II) Sanfilippo syndrome (MPS III) I cell disease (ML II) Globoid cell leukodystrophy Adrenoleukodystrophy Metachromatic leukodystrophy Tay-Sachs syndrome Lansky performance status < Original HLA match 6/ / / / Final HLA match 6/ / / / / Missing Donor/recipient sex Male/male Male/female Female/male Female/female Pretransplantation CMV status Positive Negative NC indicates nucleated cell; CMV, cytomegalovirus. *Lysosomal and peroxisomal storage disease stratum. Expanded access protocol stratum. Neutrophil Engraftment The median time to neutrophil engraftment with documented donor cells 90% was 25 days (95% CI, days). Fifty-four patients achieved neutrophil engraftment by day 42, and an additional 4 patients engrafted on days 47, 48, 49, 51. Three (4%) of the patients with graft failure had ANC hematopoietic recovery without documented donor chimerism. The cumulative incidence estimate of engraftment by day 42 was 78% (95% CI, 67%-87%). By day 100, the probability of engraftment was 84% (95% CI, 75%-91%). As can be seen in Figure 2A, the cumula- 188

6 CBT and Lysosomal Storage Diseases Table 2. Final HLA Match by Original HLA Match Original Final HLA Match* HLA Match 2/6 3/6 4/6 5/6 6/6 All 3/6 1 (50) 1 (50) 0 (0) 0 (0) 0 (0) 2 (100) 4/6 3 (10) 9 (31) 17 (59) 0 (0) 0 (0) 29 (100) 5/6 0 (0) 1 (4) 6 (22) 20 (74) 0 (0) 27 (100) 6/6 0 (0) 0 (0) 0 (0) 0 (0) 2 (100) 2 (100) All 4 (7) 11 (18) 23 (38) 20 (33) 2 (3) 60 (100) Data are n (%). *Sixty of the 69 patients had high-resolution typing. tive incidence estimate was only marginally lower than the 1 KM estimate, because there was only 1 early death (at day 37). A higher CD3 dose (before cryopreservation) was significantly associated with the incidence of engraftment (P.006). The median time to engraftment was 22 days for patients with a CD3 dose exceeding /kg and 32.5 days for patients with a CD3 dose below this level. A higher CD34 (before cryopreservation) dose was suggestive (P.051) of faster engraftment; however, none of the other factors considered demonstrated a statistically significant association with neutrophil engraftment, including age, sex, race, primary disease, HLA match, weight, NC dose, or study cohort. Platelet Engraftment Forty-nine of 69 patients achieved a platelet engraftment of / L, and 42 of 69 achieved a platelet engraftment of (results were not known in 1 case). The median time to a platelet engraftment of was 88 days (95% CI, days) and was 103 days (95% CI, days) to a platelet engraftment of The cumulative incidence of platelet engraftment of was 47% (95% CI, 37%-59%) at day 100 and 62% (95% CI, 51%-74%) at day 180 (Figure 2B). The corresponding 1 KM estimates for a platelet engraftment of , in which competing risks were not taken into account, were 52% (95% CI, 40%-65%) and 71% (95% CI, 59%-83%) at day 100 and 180, respectively (Figure 2B). There was a weak indication of a statistical association of platelet engraftment of with donor ethnicity (P.035). For platelet engraftment at a level of , the association with ethnicity was not significant, but the association with CD3 dose was marginally significant (P.069). None of the other variables considered, including age, sex, race, primary disease, HLA match, weight, NC dose, or study cohort, had a statistically significant association with platelet engraftment. Acute and Chronic GVHD Six (9%) of 69 of patients experienced maximum acute GVHD grades of III or IV, whereas 25 (36%) of BB&MT 69 experienced grade II to IV acute GVHD. The cumulative incidence of acute GVHD grades II to IV was 44% (95% CI, 33%-55%), and that of grades III and IV was 10% (95% CI, 4%-17%) at day 100. As seen in Figure 2C, the cumulative incidence was only marginally lower than the 1 KM estimates because there were only a few early deaths. Lower weight was significantly associated with development of acute GVHD (P.034). The day 100 estimate among patients with lower weight was 54% (95% CI, 37%-71%), versus 33% (95% CI, 18%- 52%) for weight 12 kg. Higher CD3 (P.019) and CD34 (P.060) cell doses were also associated with increased GVHD. At day 100, the cumulative incidences were 30% (95% CI, 18%-45%) and 58% (95% CI, 42%-74%) for CD3 doses /kg and /kg, respectively. Similarly, at day 100, the cumulative incidences were 34% (95% CI, 19%- 51%) and 54% (95% CI, 38%-72%) for a CD34 dose /kg and /kg, respectively. Although not statistically significant, performance status was suggestive (P.078) of more frequent acute GVHD. Thirteen of 68 patients contracted chronic GVHD between 4 and 25 months after transplantation. Most of these cases (n 11) had a limited maximum grade (localized skin involvement, hepatic dysfunction, or both); 2 had extensive chronic GVHD. The cumulative incidence of chronic GVHD was 8% at 6 months and 18% (95% CI, 8%-27%) at 1 year after transplantation (Figure 2D). Toxicity By day 42 after transplantation, 8 (12%) patients out of 69 had severe pulmonary toxicity, 7 (10%) patients had severe central nervous system toxicity, and 4 (6%) had severe stomatitis. Moderate stomatitis toxicity was evident in 54 (78%) patients, and moderate renal toxicity was evident in 26 (38%) patients (Figure 3). There was 1 fatal pulmonary toxicity. DISCUSSION We describe the results of a prospective trial of unrelated CBT in 69 young children with a variety of LSDs. All patients received a common chemotherapybased conditioning regimen, GVHD prophylaxis, and supportive care. Most patients received HLA-mismatched grafts. Because of their young age and small size, relatively high doses of NCs, CD3 cells, and CD34 cells were administered to all patients. Overall, there was excellent survival, with a plateau approximately 2 years after transplantation. In a similarly aged cohort of children with childhood leukemia who also received nonlimiting doses of NCs, CD3 cells, and CD34 cells, we noted lower 189

7 P. L. Martin et al. Figure 1. Survival for patients with lysosomal diseases. A, Overall survival. B, Survival by recipient race. C, Survival by study cohort (LSD stratum versus EAP). D, Survival by final HLA match. 190

8 CBT and Lysosomal Storage Diseases Table 3. Causes of Death Contributing Total, Primary Cause of Death Infection No Infection n (%) Acute GVHD (25) Autologous recovery (20) Graft failure (20) Infection polyorganism 2 (10) Infection bacterial 1 (5) Organ failure pulmonary 1 (5) Other* 3 (15) Total (100) *Progressive disease (n 2) and refractory hemolytic anemia (n 1). estimates of neutrophil recovery, grade III and IV acute GVHD at day 100, and chronic GVHD [34]. Notably, in that cohort, a higher-level HLA match showed improved survival, and the difference was statistically significant when the final high-resolution HLA-A, -B, and -DRB1 match was considered. In this cohort, HLA matching by low- or high-resolution criteria did not seem to affect engraftment, GVHD, or survival; however, the small sample size may have limited the ability to detect the influence of HLA in this cohort. Children with congenital disorders have not been exposed to high doses of chemotherapy before cytoreduction for transplantation and, thus, might be considered to be at a higher risk for nonengraftment than children with malignancies. This was not the case in this study, in which patients received ATG, busulfan, and cyclophosphamide in the preparative regimen, as opposed to ATG, busulfan, and melphalan in the leukemic cohort of patients. Cyclophosphamide would be considered to be more immunosuppressive than melphalan, which was chosen for its antileukemic properties. Similar to what has been observed after unrelated marrow transplantation, children with congenital disorders in this study had a low incidence of severe acute and chronic GVHD as compared with young children with leukemia (N.A.K., unpublished data). Of note, in the cohort of young children with leukemia, ethnicity did not affect survival, whereas nonwhite ethnicity correlated with poorer survival in this study, although no obvious criteria emerged to explain this observation. The small sample size in this study prevents any definitive analysis on this issue. The effect of ethnicity on overall survival may reflect mismatching of minor HLA antigens that is not measurable with currently available techniques. Patients treated on the EAP protocol had poorer survival as compared with those enrolled on the LSD stratum. All patients were sequentially entered on the study with identical selection criteria. The EAP patients were treated more recently, after the LSD stratum had closed to accrual. Although patients were not BB&MT evaluated or treated any differently on the EAP, patients enrolled under the EAP had, by chance, a lower performance status, and more patients had a diagnosis of Sanfilippo syndrome and MLD. The difference in survival between the 2 cohorts of patients may be an artificial observation related to the small numbers of patients with varying diagnoses in both groups. The results of this study confirmed previously reported observations from trials that used CBT in patients with LSDs. Despite significant donor-recipient HLA mismatching, the incidence of acute grades II to IV GVHD was in an acceptable range. In this study, we learned that patients with higher CD3 cell doses were at an increased risk for acute GVHD. Chronic GVHD occurred at a significantly lower incidence than would be expected in unrelated marrow transplantation, and most patients had limited involvement without significant long-term sequelae. Overall, durable donor cell engraftment was achieved in 84% of patients. Because these patients were not exposed to immunosuppressive therapy before their conditioning therapy, the major reason for graft failure is host rejection caused by failure of the preparative regimen to fully ablate host immunity. The addition of fludarabine, Campath-1H (Berlex, Inc, Montville, NJ), or other non cross-reactive immune-suppressing therapy to the preparative regimen could be considered in future trials to address this obstacle to engraftment. More than half of the patients enrolled on this study had a diagnosis of mucopolysaccharidoses type 1 (MPS) syndrome. These patients have massive hepatosplenomegaly, which can interfere with engraftment by 2 mechanisms. First, immediately after infusion of donor cells, cells can be trapped in the reticuloendothelial system, thus resulting in a decrease in the administered cell dose. Second, the liver, spleen, or both can become a temporary site of extramedullary hematopoiesis, thus delaying neutrophil recovery. Strategies to induce endothelial blockade (eg, IV immunoglobulin before transplantation) could be considered to address this problem. It is important to note that no secondary graft failures have occurred. Donor availability is a major obstacle to bone marrow transplantation, with 50% of patients unable to identify a suitable adult stem cell donor in a timely fashion. Banked umbilical cord blood is prospectively HLA typed and screened for infections and other risk factors and is readily available for transplantation. In this report, the average time from the onset of a search to identification of a compatible umbilical CBU (including screening of enzyme activity, when applicable) was 15 days. The time from search onset to the beginning of the preparative regimen was 41 days (31 days in patients who underwent transplantation since 2000). In a disease for which the time from 191

9 P. L. Martin et al. Figure 2. Cumulative incidence (CINC) of engraftment and GVHD. A, CINC and 1 KM probability of neutrophil engraftment. B, CINC and 1 KM probability of a platelet engraftment of C, CINC and 1 KM probability of acute GVHD. D, CINC and 1 KM probability of chronic GVHD. 192

10 CBT and Lysosomal Storage Diseases diagnosis to definitive treatment may be crucial to prevent further disease progression, a source of stem cells that can be acquired rapidly is extremely desirable. Implementation of mandatory neonatal screening would allow for identification of neonates who would be candidates for stem cell therapies at a time when maximal benefit could be anticipated. We did not address the effects of CBT on the neurocognitive, language, and motor development of these patients. Results in children with Hurler syndrome (14 of whom were also included in this article) and Krabbe disease (4 patients in this article) have been previously reported and have shown considerable benefit on neurocognitive outcomes, overall quality of life, and overall survival [26,27]. Because this article represents a mixture of patients with various LSDs at different stages of progression, it was not feasible to summarize these outcomes. However, after engraftment and survival, the true efficacy of this procedure will be measured in CBT s ability to prolong life with improved quality of life. As more centers treat these children, standardization of testing and supportive therapies will be needed. Even with this short followup, several of the children are already exceeding their life expectancies without transplantation. In summary, we demonstrate that hematopoietic stem cell transplantation with partially HLA-matched unrelated donor banked umbilical cord is a viable approach to stem cell transplantation for patients with LSDs not amenable to other therapies. Engraftment and full donor chimerism were achieved and maintained without the use of total body irradiation. The incidence of acute and chronic GVHD was low. Donors were readily and rapidly available. The overall event-free survival was very good. Although longterm follow-up and extensive neurocognitive studies are needed to determine the full effect of umbilical CBT on the natural history of these syndromes, it is clear that many patients will derive benefit from this therapy and that it should be considered for all young patients diagnosed with LSDs amenable to transplantation therapy who lack a matched related noncarrier bone marrow donor. Figure 3. Transplant-related toxicity. Maximal toxicity by organ system is shown through day 42 as graded by the Bearman toxicity scale [28]. The maximum toxicity experienced in any organ system was 1 (1%) grade 0, 5 (7%) grade 1, 47 (68%) grade 2, 15 (22%) grade 3, and 1 (1%) grade 4. CNS, central nervous system; GI, gastrointestinal. BB&MT ACKNOWLEDGMENTS We greatly appreciate the dedication and outstanding care delivered to these patients by their nurses, nurse practitioners, nurse coordinators, social workers, physical, speech and occupational therapists, and other allied health care professionals involved in their care. We are indebted to Angela Norman for assistance in the preparation of the manuscript. This work was supported by a contract from the National Heart, Lung and Blood Institute (grant nos. N01-HB [P.L.M. and J.K.], N01-HB [S.L.C., N.A.K., I.S., D.W., and D.P.], and N01-HB [J.E.W.]). REFERENCES 1. Krivit W, Peters C, Shapiro EG. Bone marrow transplantation as effective treatment of central nervous system disease in globoid cell leukodystrophy, metachromatic leukodystrophy, adrenoleukodystrophy, mannosidosis, fucosidosis, aspartylglucosaminuria, Hurler, Maroteaux-Lamy, and Sly syndromes, and Gaucher disease type III. Curr Opin Neurol. 1999;12: Hobbs JR, Hugh-Jones K, Barrett AJ, et al. Reversal of clinical features of Hurler s disease and biochemical improvement after treatment by bone-marrow transplantation. Lancet. 1981;2: Braunlin EA, Rose AG, Hopwood JJ, Candel RD, Krivit W. Coronary artery patency following long-term successful engraftment 14 years after bone marrow transplantation in the Hurler syndrome. Am J Cardiol. 2001;88: Shapiro EG, Lockman LA, Balthazor M, Krivit W. Neuropsychological outcomes of several storage diseases with and without bone marrow transplantation. J Inherit Metab Dis. 1995;18: Krivit W, Henslee-Downey J, Klemperer M, et al. Survival in Hurler s disease following bone marrow transplantation in 84 patients. Bone Marrow Transplant. 1995;15(suppl): Whitley CB, Belani KG, Chang PN, et al. Long-term outcome of Hurler syndrome following bone marrow transplantation. Am J Med Genet. 1993;46: Peters C, Balthazor M, Shapiro EG, et al. Outcome of unrelated donor bone marrow transplantation in 40 children with Hurler syndrome. Blood. 1996;87: Peters C, Shapiro EG, Krivit W. Hurler syndrome: past, present and future. J Pediatr. 1998;133: Guffon N, Souillet G, Maire I, Straczek J, Guibaud P. Follow-up of nine patients with Hurler syndrome after bone marrow transplantation. J Pediatr. 1998;133: Peters C, Shapiro EG, Anderson J, et al. Hurler syndrome: II. 193

11 P. L. Martin et al. Outcome of HLA-genotypically identical sibling and HLA-haploidentical related donor bone marrow transplantation in fifty-four children. Blood. 1998;91: Cowan M. Bone marrow transplantation for the treatment of genetic diseases. Clin Biochem. 1991;24: Whitley CB, Ramsay NK, Kersey JH, Krivit W. Bone marrow transplantation for Hurler syndrome: assessment of metabolic correction. Birth Defects Orig Artic Ser. 1986;22: Fleming DR, Henslee-Downey PJ, Ciocci G, et al. The use of partially HLA-mismatched donors for allogeneic transplantation in patients with mucopolysaccharidosis I. Pediatr Transplant. 1998;2: Kidd D, Nelson J, Jones F, et al. Long-term stabilization after bone marrow transplantation in juvenile metachromatic leukodystrophy. Arch Neurol. 1998;55: Krivit W, Shapiro E, Kennedy W, et al. Treatment of late infantile metachromatic leukodystrophy by bone marrow transplantation. N Engl J Med. 1990;322: Malm G, Ringden O, Winiarski J, et al. Clinical outcome in four children with metachromatic leukodystrophy treated by bone marrow transplantation. Bone Marrow Transplant. 1996; 17: Miano M, Porta F, Locatelli F, et al. Unrelated donor marrow transplantation for inborn errors. Bone Marrow Transplant. 1998;21(suppl 2): Krivit W, Shapiro EG, Peters C, et al. Hematopoietic stem-cell transplantation in globoid-cell leukodystrophy. N Engl J Med. 1998;338: Navarro C, Fernandez JM, Dominguez C, Fachal C, Alvarez M. Late juvenile metachromatic leukodystrophy treated with bone marrow transplantation: a 4-year follow-up study. Neurology. 1996;46: Shapiro E, Krivit W, Lockman L, et al. Long-term effect of bone-marrow transplantation for childhood-onset cerebral X- linked adrenoleukodystrophy. Lancet. 2000;356: Peters C, Charnas LR, Tan Y, et al. Cerebral X-linked adrenoleukodystrophy: the international hematopoietic cell transplantation experience from 1982 to Blood. 2004; 104: Aubourg P, Blanche S, Jambaque I, et al. Reversal of early neurologic and neuroradiologic manifestations of X-linked adrenoleukodystrophy by bone marrow transplantation. N Engl J Med. 1990;322: Kurtzberg J, Laughlin M, Graham ML, et al. Placental blood as a source of hematopoietic stem cells for transplantation into unrelated recipients. N Engl J Med. 1996;335: Gluckman E, Broxmeyer HE, Auerbach AD, et al. Hematopoietic reconstitution in a patient with Fanconi s anemia by means of umbilical cord blood from an HLA-identical sibling. N Engl J Med. 1989;321: Rubinstein P, Rosenfield RE, Adamson JW, Stevens CE. Stored placental blood for unrelated bone marrow reconstitution. Blood. 1993;81: Staba SL, Escolar ML, Poe M, et al. Cord-blood transplants from unrelated donors in patients with Hurler s syndrome. N Engl J Med. 2004;350: Escolar ML, Poe MD, Provenzale JM, et al. Transplantation of umbilical-cord blood in babies in with infantile Krabbe s disease. N Engl J Med. 2005;352: Bearman SI, Appelbaum FR, Buckner CD, et al. Regimenrelated toxicity in patients undergoing bone marrow transplantation. J Clin Oncol. 1988;6: Przepiorka D, Weisdorf D, Martin P, et al consensus conference on acute GVHD grading. Bone Marrow Transplant. 1995;15: Weisdorf DJ, Hurd D, Carter S, et al. Prospective grading of graft-versus-host disease after unrelated donor marrow transplantation: a grading algorithm versus blinded expert panel review. Biol Blood Marrow Transplant. 2003;9: Kaplan EL, Meier P. Nonparametric estimation from incomplete observations. J Am Stat Assoc. 1958;53: Cox DR. Regression models and life-tables. J R Stat Soc B. 1972;34: Gooley TA, Leisenring W, Crowley J, Storer BE. Estimation of failure probabilities in the presence of competing risks: new representations of old estimators. Stat Med. 1999;18: Wall DA, Carter SL, Kernan NA, et al. Busulfan/melphalan/antithymocyte globulin followed by unrelated donor cord blood transplantation for treatment of infant leukemia and leukemia in young children: the Cord Blood Transplantation study (COBLT) experience. Biol Blood Marrow Transplant. 2005;11: