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Enzyme replacement therapy in mucopolysaccharidosis VI (Maroteaux-Lamy syndrome)
ENZYME REPLACEMENT THERAPY IN MUCOPOLYSACCHARIDOSIS VI (MAROTEAUX-LAMY SYNDROME) PAUL HARMATZ, MD, CHESTER B. WHITLEY, PHD, MD, LEWIS WABER, MD, PHD, RAY PAIS, MD, ROBERT STEINER, MD, BARBARA PLECKO, MD, PAIGE KAPLAN, MD, JULIE SIMON, RN, ELLEN BUTENSKY, PNP, AND JOHN J. HOPWOOD, PHD
Objectives To evaluate the safety and efficacy of weekly treatment with human recombinant N-acetylgalactosamine 4-sulfatase (rhASB) in humans with mucopolysaccharidosis type VI (MPS VI). Study design An ongoing Phase I/II, randomized, two-dose, double-blind study. Patients were randomized to weekly infusions of either high (1.0 mg/kg) or low (0.2 mg/kg) doses of rhASB. Six patients (3 male, 3 female; age 7-16 years) completed at least 24 weeks of treatment, five of this group have completed at least 48 weeks. Results No drug-related serious adverse events, significant laboratory abnormalities, or allergic reactions were observed in the study. The high-dose group experienced a more rapid and larger relative reduction in urinary glycosaminoglycan that was sustained through week 48. Improvements in the 6-minute walk test were observed in all patients with dramatic gains in those walking <100 meters at baseline. Shoulder range of motion improved in all patients at week 48 and joint pain improved in patients with significant pain at baseline. Conclusions
rhASB treatment was well-tolerated and reduced lysosomal storage as evidenced by a dose-dependent reduction in urinary glycosaminoglycan. Clinical responses were present in all patients, but the largest gains occurred in patients with advanced disease receiving high-dose rhASB. (J Pediatr 2004;144:574-80)
ucopolysaccharidosis type VI (MPS VI; Maroteaux-Lamy syndrome) is a lysosomal storage disease in which affected persons lack the enzyme N-acetylgalactosamine 4-sulfatase (ASB).1 In the absence of this enzyme, the stepwise degradation of the glycosaminoglycan (GAG) dermatan sulfate (DS) is blocked. This results in the intracellular accumulation of partially degraded GAG in lysosomes of a wide range of tissues. The accumulation causes a chronic progressive disorder involving multiple organs that often results in death in the second decade of life. The estimated birth incidence of MPS VI ranges from 1:100,000 to 1:1,300,000 in various populations.2-5 The diagnosis of MPS VI is usually made at 6 to 24 months of age when children show progressive deceleration of growth, skeletal deformities, coarse facial features, and upper airway obstruction. Clouding of the cornea, communicating hydrocephalus, blindness, or heart disease may develop. MPS VI is not typically associated with progressive impairment of mental status, although physical limitations may impact learning and development. There is no satisfactory treatment for MPS VI. Most patients receive symptomatic treatment for specific problems as their only form of therapy. Although a few patients have benefited from bone marrow transplantation (BMT),6,7 it is not universally performed due to lack of a suitable donor and is associated with significant morbidity and mortality. The European Group for Bone Marrow Transplantation reported transplant-related mortality of 10% (HLA identical) to 20% to 25% (HLA mismatched) for 63 transplantation cases of lysosomal disorders.8
M
AHI ASB AUC0-t
CHAQ
574
Apnea-hypopnia index N-acetylgalactosamine 4-sulfatase Area under the plasma concentration-time curve from 0 to the final sample with a concentration equal to or greater than the qualified limit of detection of the assay Childhood Health Assessment Questionnaire
DMB ERT GAG MPS rhASB T½
1,9-dimethylmethylene blue Enzyme replacement therapy Glycosaminoglycan(s) Mucopolysaccharidosis Recombinant human N-acetylgalactosamine 4-sulfatase Elimination half-life
See editorial, p 561, and related articles, p 569 and p 581. From Pediatric Clinical Research Center, Children’s Hospital & Research Center at Oakland, California; Department of Pediatrics, University of Minnesota Medical School, Minneapolis, Minnesota; Pediatric Genetics and Metabolism, University of Texas Southwest Medical Center, Dallas, Texas; Pediatric Hematology/Oncology, East Tennessee Children’s Hospital, Knoxville, Tennessee; Division of Metabolism, Oregon Health & Science University, Portland, Oregon; U Klinik fur Kinder-und Jugendheilkunde, Graz, Austria; Section of Biochemical Genetics, Children’s Hospital Philadelphia, Philadelphia, Pennsylvania; and Department of Chemical Pathology, Women’s and Children’s Hospital Adelaide, Adelaide, Australia. This study was sponsored by BioMarin Pharmaceutical Inc. It was supported in part by PHS, National Center for Research, Resources grants: MO1_RR01271, MO1_RR00334, MO1_RR00240. Submitted for publication Aug 13, 2003; last revision received Feb 25, 2004; accepted Mar 4, 2004. Reprint requests: Paul Harmatz, MD, Children’s Hospital & Research Center at Oakland, 747 52nd St, Oakland, CA 94609. E-mail: Pharmatz@mail. cho.org. 0022-3476/$ - see front matter Copyright ª 2004 Elsevier Inc. All rights reserved. 10.1016/j.jpeds.2004.03.018
Table I. Patient demographics, disease severity (at baseline) Patient #
Age
Sex
Ethnicity
High dose: 1.0 mg/kg 42 43
7 16
M F
Caucasian Caucasian
44
12
F
African American
Low dose: 0.2 mg/kg 41 45 50
7 11 13
F M M
Caucasian Caucasian Caucasian
Ventilatory assistance
Major surgical procedures
None Nasal oxygen for 10 h at night Tracheostomy, CPAP/ BiPAP only with upper respiratory infections
None Ventriculoperitoneal shunt
None Tracheostomy Tracheostomy, CPAP/BiPAP for 8 h at night
None Tracheostomy Ventriculoperitoneal shunt, tracheostomy, spinal surgery
Ventriculoperitoneal shunt, tracheostomy
BiPAP, Bilevel positive air pressure; CPAP, continuous positive air pressure.
Enzyme replacement therapy (ERT) has been approved for use in three lysosomal storage disorders: Gaucher disease, Fabry disease, and MPS I.9 In each of these diseases, recombinant human enzyme synthesized in vitro is infused on a regular schedule. The infused enzymes utilize either mannose-6-phosphate (Fabry disease and MPS I) or mannose (Gaucher disease) receptor-mediated endocytosis to reach intracellular sites and facilitate degradation of material stored in lysosomes. Similarly, ERT with recombinant human ASB (rhASB) in a feline model of MPS VI was associated with a reduction of storage vacuoles in Kupffer cells and connective tissue, an increase in bone mineral volume, and greater mobility.10,11 Onthebasisoftheseanimalstudies,aclinicalstudy of rhASB replacement was initiated in patients with MPS VI.
METHODS Patients Seven patients with MPS VI were enrolled. All patients had clinical manifestations of the disease, and diagnosis was confirmed by leukocyte ASB enzyme activity of <20% of the normal range of the measuring laboratory. There were 3 females and 4 males, ages 7 to 16 years. Patient demographics and representation of disease severity for those completing the initial 24-week double-blind stage are presented in Table I. The protocol was approved by the institutional review board at each participating clinical site. Written consent was obtained from all parents or guardians before enrollment and written assent was obtained from all patients.
Study Design Patients were randomized to receive either 0.2 or 1.0 mg/kg of rhASB as weekly intravenous infusions in a doubleblind design. rhASB doses were selected for the current study based on previous studies in the feline model of MPS VI using human (1 and 5 mg/kg) and feline ASB (1 mg/kg) ERT.
Same-species feline rfASB (1 mg/kg) appeared to be at least equivalent to high-dose human rhASB (5 mg/kg) in the feline model.10 Thus, same species human rhASB at 0.2 and 1.0 mg/ kg were selected for the current study such that both doses were likely to show some efficacy. Safety and efficacy evaluations were completed at baseline and then at 1 to 6 week intervals. After the 24 week safety and efficacy evaluations, the study blind was removed, but all patients remained on their assigned dose until after the 48-week evaluation.
Study Drug rhASB was produced in a suspension bioreactor by genetically engineered Chinese hamster ovary cells. The enzyme was purified through a traditional column chromatography procedure and formulated in phosphate buffered saline at pH 5.8 containing 0.005% polysorbate 80. The specific activity of the formulated enzyme averaged 56 units/ mg. The purified enzyme contains a high level of bismannose-6-phosphate oligomannose oligosaccharide as confirmed by analytical assays and by the saturable robust ‘‘uptake’’ of enzyme by MPS VI fibroblasts in cell culture. Addition of 50 mM mannose-6-phosphate in the cell culture media reduced the cell uptake by more than 90%.
rhASB Administration All patients were premedicated with diphenhydramine (0.5 mg/kg body weight). rhASB was diluted in 0.9% saline and administered at 0.2 or 1.0 mg/kg over 4 hours once weekly. The infusion rate was adjusted so that approximately 2.5% of the total enzyme dose was infused during the first hour and the remaining enzyme dose (; 97.5%) was infused over the next three hours.
Biochemical Studies Studies to monitor toxicity were performed every one to six weeks and included complete blood count, chemistry panel,
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urinalysis, serum IgG anti-rhASB antibody by enzyme-linked immunosorbent assay, and measurement of serum complement before and after infusion. Urine was obtained every one to six weeks to determine total GAG, a surrogate for the extent of clearance of these compounds from lysosomal storage. Total GAG concentrations in urine samples were determined with a method based on spectrophotometric detection of metachromatic changes to the dye 1,9dimethylmethylene blue (DMB) resulting from GAG binding.12 GAG concentrations were subsequently normalized to urinary creatinine concentrations, which were determined separately. Total GAG was quantitated by measuring DMB binding, using dermatan sulfate as a standard. GAG concentrations (lg/mL) were divided by creatinine concentrations (converted to mg/mL) to yield a final reported value for GAG concentration in units of ‘‘lg of total GAG per mg of creatinine.’’ Pharmacokinetic studies were performed at 1, 2, 12, and 24 weeks. One-milliliter blood samples were collected from a second intravenous line placed in the arm opposite to that used for the enzyme infusion. rhASB levels were measured by enzyme linked immunosorbent assay. Pharmacokinetic parameters determined included area under the plasma concentration-time curve from 0 to the final sample with concentration equal to or greater than the limit of detection (AUC0-t ), maximum observed concentration, and elimination half-life (T½).
Clinical Evaluations Evaluation of mobility and physical function was performed at baseline and every 6 weeks, including a 6-minute walk test, joint range of motion, spirometry, and assessment of functional status. For the 6-minute walk test,13 patients were instructed to walk as far as possible in 6 minutes, but were allowed to rest when needed. Oxyhemoglobin saturation and heart rate were monitored by pulse oximetry (Nellcor 395). Range of motion (ROM) of the shoulders, elbows and knees was measured with a goniometer by occupational and physical therapists. Forced vital capacity (FVC) and forced expiratory volume at 1 minute (FEV1) were evaluated by standard spirometry technique according to American Thoracic Society guidelines. Functional status was assessed by an age-appropriate questionnaire, the Childhood Health Assessment Questionnaire (CHAQ),14 that was completed by a patient’s parent or guardian. Finally, each patient was videotaped while performing a standard set of physical movements based on activities in the Denver Developmental examination,15 including standing unassisted, walking forward, walking backward, throwing a ball, kicking a ball, jumping in place, balancing on each foot, hopping on each foot, catching a bounced ball, reaching for and grasping a ball above eye level, climbing stairs, building a tower of blocks, copying a square, pulling on a sweatshirt over the head, picking up and writing with a pen, pouring a cup of water, buttoning buttons, zipping a jacket, putting on and lacing shoes, and picking up coins and placing them into a cup. 576
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An ophthalmology evaluation including fundoscopic and slit-lamp examinations, assessment of glaucoma, and determination of visual acuity, was performed at baseline and every 12 weeks. Standard 12-lead electrocardiogram (EKG), 2-dimensional Doppler echocardiogram and chest radiography were performed at baseline and every 12 weeks. Liver volume and lumbar vertebral trabecular bone density were assessed by CT scan at baseline, week 12 and week 24. The liver volume was calculated from the axial image dataset using the postprocessing graphic workstation (Picker Omnipro/ Algotec Pro Vision software, Raanana, Israel), and bone density was determined using QCT Pro (Mindways Software, San Francisco, Calif ). Polysomnography was performed every 24 weeks according to the guidelines of the American Thoracic Society. The number of episodes of apnea and hypopnea, the number of minutes during sleep in which oxygen saturation fell below 89 percent, and the total sleeping time were recorded. An apnea-hypopnea index (AHI) was calculated by dividing the total number of episodes of apnea and hypopnea by the number of hours of sleep.
Statistics Descriptive statistics, including means, standard deviations (SD), and percent change over time, were calculated using Systat 10.2 (Systat Software, Inc, Richmond, Calif). Change in parameter between baseline and at 48 weeks was compared using the Wilcoxon signed rank test for small groups (Systat 10.2).
RESULTS Six of the 7 patients completed 24 weeks of treatment (Table I), 5 patients completed 48 weeks. One patient (#40) withdrew from the study at week 3 due to personal reasons unrelated to the study drug treatment, and another (#50) withdrew at week 32 because of a lack of perceived benefit. This patient was on low-dose rhASB at the time of study exit, and was unwilling to wait for institutional review board approval to be placed on the higher dose.
Safety There were 7 serious adverse events (SAEs) reported during the first 48 weeks of treatment. One event, mild desaturation during sleep (Patient #43), was subsequently determined not to be an adverse event, because the findings did not represent a change from baseline. Another event, brain glioma and colonic adenocarcinoma reported for Patient #40 approximately 10 months after the patient withdrew from the trial, was determined to be due to an unrelated genetic mutation. This patient withdrew from the trial after receiving only three infusions. The remaining five SAEs included one episode of vasovagal syncope at the time of intravenous line placement (Patient #40), hypoxia at the time of a routine tracheostomy tube change (Patient #44), paratracheal skin hypertrophy (Patient # 44), worsening eustachian tube dysfunction (Patient # 41), and bilateral carpal tunnel The Journal of Pediatrics May 2004
Fig 1. Urinary GAG excretion in patients during ERT by individual patients (A) and in patients grouped by dose (B). Urinary GAG excretion was measured in terms of relative reduction in excretion and expressed as the percent of the baseline excretion. Patient #50 (low dose) dropped out of the study after week 30, therefore, week 36-48 mean values for the low-dose group are calculated from one less data point. The difference between urinary GAG at baseline and that at week 48 (relative reduction) for the total population was significant (P = .04).
syndrome (Patient #41). The last three events were considered serious because the resulting surgical interventions required in-patient hospitalization. No SAEs occurred during study drug infusion, and none were considered related to study drug. There were no reports of allergic reactions (urticaria, angioedema) during infusion. Thirty-one adverse events occurred during the infusion. All were assessed as mild, and all resolved. Events of hypotension, tachypnea, skin irritation on neck and chest, pain at IV insertion site, and a 3-second run of premature ventricular contractions were assessed as possibly related to study drug, whereas the other events were considered to be unrelated. Adverse events (n = 126) were reported to occur during the first 48 weeks of treatment, 50 in the low-dose group and 76 in the high-dose group. The following adverse events were assessed as possibly related to study drug (number in parentheses is the number of discrete events): lethargy (2), hypotension (1), itching (4), rash/skin irritation (4), and intermittent joint popping (1). The majority of the adverse events were attributed to manifestations of the underlying disease.
Efficacy A significant relative decrease in urinary GAG was observed for the total population (P = .04) (Fig 1). The high dose produced a more rapid and larger sustained relative reduction in urinary GAG (63% vs 51% at week 48). Functional capacity of the patients improved during the study period. Five of the 6 patients (the sixth patient withdrew after week 30) improved in the 6-minute walk test at the 48week time point (Table II), with a significant improvement noted for the total population (P = .04). Dramatic gains were observed at week 48 for the two patients walking <100 meters at baseline (238% and 43% increase in meters walked).
Improvement in ability to walk up stairs was also demonstrated on the videotape of functional activities. Five of the six patients demonstrated improvement in shoulder range of motion at week 48 (the sixth patient withdrew after week 30). Restriction in shoulder flexion decreased significantly from baseline to week 48 for the total population (right shoulder: 84 to 68 degrees, P = .04; left shoulder: 83 to 67 degrees, P = .04). Smaller improvements were observed in the knees and elbows. Improvements in functional status were evidenced by investigator observation (including picking up coins, tying shoelaces, pulling shirt overhead) and by responses to the pain and arthritis questions on the CHAQ (P = .04, change from baseline to 48 weeks). Liver volumes were assessed at baseline and at the 24week time point. At baseline, only the two oldest patients’ livers were significantly enlarged for age and body weight, and these patients showed a 14% percent reduction in liver volume after 24 weeks of treatment. Spirometry examinations (FVC and FEV1) were limited by severe restrictive lung disease in several patients (#43, #44, #41, #50), difficulty producing the required 6-second expiration, and also the presence of tracheostomy in 3 patients (#44, #45, #50). Improvement in spirometry (FVC increased 11%) was observed in one patient (#42). Attempts to define upper airway obstruction by polysomnography were impeded by tracheostomies present in patients #44, #45, and #50. Of the other three patients, patients #41 and #42 showed improvement at week 48 with a 7.5- and 4-fold decrease, respectively, in AHI. Patient #43 had a decrease in the amount of nasal oxygen required during the sleep study at week 48, but no change in the AHI. There were no appreciable gains in height for any patient between baseline and week 48. Patients #44 and #45 experienced approximately 17% and 14% weight gains from baseline, respectively, whereas the other 4 patients experienced little or no weight gains.
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Fig 2. Circulating concentrations of rhASB in the 6 hours after initiation of rhASB infusion at week 1 (A) and changes in area under the curve (AUC0-t) values of rhASB in patients grouped by dose followed over 24-week study period (B). rhASB plasma concentrations during and 2 hours following the first infusion are shown for individual patients. Plasma rhASB concentrations confirmed two doses and the rhASB cleared rapidly from the circulation at the end of the infusion. The AUC0-t was calculated from the plasma concentrations of rhASB to the last time point with a concentration above the limit of quantitation (33.75 ng/mL). AUC0-t values confirmed two doses. Mean values for weeks 12 and 24 for the low-dose group are calculated from one less data point.
Table II. Six-minute walk test: Change in distance walked Patient # 42 43 44 High-dose mean 41 45 50 Low-dose mean
Dose (mg/kg) 1 1 1 0.2 0.2 0.2
Baseline meters (mean of 3 ± SD)
Week 6 meters (%)
Week 12 meters (%)
388 ± 60 53 ± 15 88 ± 12 176 ± 29 197 ± 21 283 ± 20 133 ± 19 204 ± 20
ÿ62 (ÿ16) 18 (34) 46 (52) 2 (23) ÿ97 (ÿ49) 29 (10) ÿ5 (ÿ4) ÿ24 (ÿ14)
ÿ43 (ÿ11) 53 (100) 65 (74) 25 (54) ÿ60 (ÿ30) 53 (19) ÿ22 (ÿ17) ÿ10 (ÿ9)
Week 24 meters (%) ÿ33 99 63 43 16 42 10 23
(ÿ9) (187) (72) (83) (8) (15) (8) (10)
Week 48 meters (%) 20 (5) 126 (238) 38 (43) 61 (95) 7 (4) 125 (44) DC 66 (24)
DC, Discontinued.
No appreciable changes from baseline to week 48 were observed for echocardiogram, bone density examinations, visual examinations, grip tests, or pinch strength testing.
Pharmacokinetics rhASB plasma concentrations during and 2 hours after the first infusion are shown for individual patients (Fig 2, A). Maximum observed concentration was 572 ± 60 ng/mL and 61 ± 11 ng/mL for the high- and low-dose groups, respectively. In general, the antigen was not measurable in the plasma within 10 minutes after the completion of the enzyme infusion in all patients (t½ = 4.31 minutes calculated for one patient in the high-dose group; unable to calculate for the low-dose group). rhASB pharmacokinetics appeared to be nonlinear, as reflected by greater than dose-proportionate increases in AUC0-t (Fig 2, B). AUC0-t increased relative to week 1 values in the high dose patients, but remained unchanged in the low dose patients. 578
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Antibody Formation All 6 patients seroconverted, developing IgG antirhASB antibodies, by 30 weeks of treatment. The higher dose was not associated with higher antibody levels. Antibody development was not associated with significant complement depletion (maximum 21%) during infusions or persistent levels outside the normal range, and did not appear to have any significant influence on safety or efficacy.
DISCUSSION The current study assessed the safety and efficacy of ERT with rhASB for the treatment of MPS VI in humans. The bioactivity of the enzyme was confirmed by the observation of decreases in total GAG that were reduced in a dose-dependent fashion. Functional status also improved as seen by an increase in distance walked, stair-climbing ability, shoulder range of motion, and decrease in pain and arthritis The Journal of Pediatrics May 2004
severity scores on the CHAQ. These improvements were most evident in the patients with more clinically advanced disease receiving high-dose ERT, and were achieved without serious adverse events, laboratory abnormalities or allergic reactions. Dose-dependent responses were not seen in the functional status parameters, possibly related to the small number of patients and large range of ages and disease severity within each group. The results of this study provide promising evidence that ERT may serve as an effective treatment for MPS VI disease. Significant gains have not been observed for pulmonary function in this study. Three patients had tracheostomies and could not be effectively tested. Of the 3 patients without tracheostomies, absence of improvement may relate to low enzyme dose in one patient (#41) and the presence of severe restrictive lung disease in an older patient with advanced disease (#43). The one patient showing a significant improvement in pulmonary function received high-dose enzyme and had less clinically advanced disease at baseline, particularly in the skeletal system. This may indicate that short-term improvements in FVC related to the clearance of stored GAG in the soft tissue surrounding the airway may not be appreciated once advanced changes in the skeletal system and trachea have occurred, or that longer treatment may be required to effect a change. Additional patients will need to be studied to confirm this supposition. Although no improvement in pulmonary function was observed in the patients with clinically advanced disease, it is possible that these patients, in the absence of ERT, would show disease-related deterioration over the one-year study period, a possibility that could not be assessed with the current study design. Improvements in secluded sites of GAG accumulation, including joint cartilage and corneas, were not observed after intravenous administration of rhASB in the feline study.11 More recently, Byers et al16 have reported in preliminary studies that ASB injection in the large weight-bearing joints in cats preserves cartilage. More notable was the finding that injections could be limited to every few months, giving this therapy a potential role in humans. Simonaro et al17 described increased release of nitric oxide and tumor necrosis factor-a in cultured human MPS VI articular chondrocytes, suggesting that ERT may act by reducing inflammation around the joint. In the current study, improvement in joint symptoms with decreased pain and improved range of motion was observed. We did not assess intra-articular structure or damage with MRI and, therefore, we cannot state whether joint/cartilage structure improved with ERT. Development of antibodies in the setting of ERT is common. In a large study of patients with Gaucher disease receiving ERT, 12.8% developed antibodies; however, 90% of individuals became tolerant over time.18 Similarly, our patients developed antibodies to ASB but no impact on clinical efficacy or biochemical activity was noted. It is possible that the high level of antibody in a high-dose patient (#43) at week 24 explains the prolonged survival of rhASB in the circulation and increased AUC0-t noted at 24 weeks. Although all patients in this study were older than five years and had significant disease, it is anticipated that, in the
future, ERT will be targeted for use in individuals at very early disease stages or even before the onset of clinical symptoms. It has been shown that earlier treatment has a more profound effect on skeletal pathology and, because fetal storage has been demonstrated in MPS VI kittens,19 it is anticipated that early treatment before the onset of irreversible pathology will maximize the benefits of therapy. Early treatment clearly depends on effective newborn screening and identification of patients at risk of severe disease. Programs for newborn screening are currently being proposed20-22 but are still in development and not in widespread use. In conclusion, our findings demonstrate that ERT with rhASB at 1 mg/kg results in clinical and biochemical improvements without significant side effects. The total urine GAG excretion measurements provide strong biochemical evidence for a stable, dose-dependent effect of ERT in these patients. The higher dose of 1.0 mg/kg appears to be superior to the lower dose of 0.2 mg/kg based both on the rapidity with which urinary GAG declined and on the relative reductions in urine GAG. The higher dose also resulted in larger gains in absolute levels of circulating enzyme. Because of the great clinical heterogeneity reported with MPS disease,23 it is difficult to design trials based on clinically relevant end-points. Clearly, the largest and most rapid improvements were seen in patients with advanced disease. Future studies to confirm efficacy and safety will depend on identification of clinical measures to capture and quantify the additional benefits observed in this Phase I/II Study. We appreciate the Austrian MPS Society’s support of one of the participating families. We would like to thank Dr Jerry Thompson (University of Alabama at Birmingham) for providing quantitative measurements of leukocyte ASB, Karen Hildebrand for her expertise in assessments of joint function, and Drs Hector Gutierrez, Richard Rowe, Howard Rosenfeld, Selim Koseoglu, Ronald Cohen, John R. Waterson, Edward Lammer, Kumar G. Belani, and David S. Beebe for their clinical expertise. We would like to acknowledge the efforts of the following study coordinators: Sandra A. Banta-Wright, RNC, MN, NNP (Oregon Health & Science University), Maggie Wolf (U Klinik fur Kinder-und Jugendheilkunde, Graz, Austria), Sheri McCaffery (Children’s Hospital of Philadelphia), Laurie Cariveau, RN, and Amy Deyle (University of Minnesota).
REFERENCES 1. Neufeld EF, Muenzer J. The mucopolysaccharidoses. In: Scriver C, Beaudet AL, Sly WS, et al, editors. The metabolic basis of inherited disease. New York: McGraw-Hill; 1995. p. 2465-93. 2. Lowry RB, Applegarth DA, Toone JR, MacDonald E, Thunem NY. An update on the frequency of mucopolysaccharide syndromes in British Columbia. Hum Genet 1990;85:389-90. 3. Meikle PJ, Hopwood JJ, Clague AE, Carey WF. Prevalence of lysosomal storage disorders. JAMA 1999;281:249-54. 4. Nelson J. Incidence of the mucopolysaccharidoses in Northern Ireland. Hum Genet 1997;101:355-8. 5. Poorthuis BJ, Wevers RA, Kleijer WJ, Groener JE, doJong JG, van Weely S, et al. The frequency of lysosomal storage diseases in the Netherlands. Hum Genet 1999;105:151-6. 6. Krivit W, Pierpont ME, Ayaz K, Tsai M, Ramsay NK, Kersey JH, et al. Bone-marrow transplantation in the Maroteaux-Lamy syndrome
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(mucopolysaccharidosis Type VI). Biochemical and clinical status 24 months after transplantation. N Engl J Med 1984;311:1606-11. 7. Krivit W. Maroteaux-Lamy syndrome (mucopolysaccharidosis type VI). Treatment by allogeneic bone marrow transplantation in 6 patients and potential for autotransplantation bone marrow gene insertion. Int Pediatr 1992;7:47-52. 8. Hoogerbrugge PM, Brouwer OF, Bordigoni P, The European Group for Transplantation. Allogeneic bone marrow transplantation for lysosomal storage diseases. Lancet 1995;246:1398-402. 9. Wraith JE. Advances in the treatment of lysosomal storage disease. Dev Med Child Neurol 2001;43:639-46. 10. Bielicki J, Crawley AC, Davey RCA, Varnai JC, Hopwood JJ. Advantages of using same species enzyme for replacement therapy in a feline model of mucopolysaccharidosis type VI. J Biol Chem 1999;274: 36335-43. 11. Crawley AC, Brooks DA, Muller VJ, Petersen BA, Isaac EL, Bielicki J, et al. Enzyme replacement therapy in a feline model of Maroteaux-Lamy syndrome. J Clin Invest 1996;97:1864-73. 12. Whitley CB, Ridnour MD, Draper KA, Dutton CM, Neglia JP. Diagnostic test for mucopolysaccharidosis I. Direct method for quantifying excessive urinary glycosaminoglycan excretion. Clin Chem 1989;35:374-9. 13. Nixon PA, Joswiak ML, Fricker FJ. A six-minute walk test for assessing exercise tolerance in severely ill children. J Pediatr 1996;129:362-6. 14. Singh G, Athreya BH, Fries JF, Goldsmith DP. Measurement of health status in children with juvenile rheumatoid arthritis. Arth Rheum 1994;37:1761-9. 15. Sperhac AM, Salzer JL. A new developmental screening test. The Denver II. J Am Acad Nurse Pract 1991;3:152-7.
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16. Byers S, Auclair D, Brumfield LK, Robinson A, Hopwood JJ. Localised therapy for joint disease in MPS VI. 2002. Paris, 7th International Symposium on MPS and Related Diseases. 6-20-2002. 17. Simonaro CM, Haskins ME, Schuchman EH. Articular chondrocytes from animals with a dermatan sulfate storage disease undergo a high rate of apoptosis and release nitric oxide and inflammatory cytokines: a possible mechanism underlying degenerative joint disease in the mucopolysaccharidoses. Lab Invest 2001;81:1319-28. 18. Rosenberg M, Kingma W, Fitzpatrick MA, Richards SM. Immunosurveillance of alglucerase enzyme therapy for gaucher patients: induction of humoral tolerance in seroconverted patients after repeat administration. Blood 1999;93:2081-8. 19. Crawley AC, Niedzielski KH, Isaac EL, Davey RCA, Byers S, Hopwood JJ. Enzyme replacement therapy from birth in a feline model of mucopolysaccharidosis type VI. J Clin Invest 1997;99:651-62. 20. Meikle PJ, Ranieri E, Ravenscroft EM, Hua CT, Brooks DA, Hopwood JJ. Newborn screening for lysosomal storage disorders. Southeast Asian J Trop Med Public Health 1999;30:104-10. 21. Whitley CB, Draper KA, Dutton CM, Brown PA, Severson SL, France LA. Diagnostic test for mucopolysaccharidosis II. Rapid quantification of glycosaminoglycan in urine samples collected on a paper matrix. Clin Chem 1989;35:2074-81. 22. Whitley CB, Spielman RC, Herro G, Severson ST. Urinary glycosaminoglycan excretion quantified by an automated semimicro method in specimens conveniently transported from around the globe. Mol Genet Metabol 2002;75:56-64. 23. Wraith JE. Enzyme replacement therapy in mucopolysaccharidosis type I: progress and emerging difficulties. J Inherit Metab Dis 2001;24:245-50.
The Journal of Pediatrics May 2004
Objectives To evaluate the safety and efficacy of weekly treatment with human recombinant N-acetylgalactosamine 4-sulfatase (rhASB) in humans with mucopolysaccharidosis type VI (MPS VI). Study design An ongoing Phase I/II, randomized, two-dose, double-blind study. Patients were randomized to weekly infusions of either high (1.0 mg/kg) or low (0.2 mg/kg) doses of rhASB. Six patients (3 male, 3 female; age 7-16 years) completed at least 24 weeks of treatment, five of this group have completed at least 48 weeks. Results No drug-related serious adverse events, significant laboratory abnormalities, or allergic reactions were observed in the study. The high-dose group experienced a more rapid and larger relative reduction in urinary glycosaminoglycan that was sustained through week 48. Improvements in the 6-minute walk test were observed in all patients with dramatic gains in those walking <100 meters at baseline. Shoulder range of motion improved in all patients at week 48 and joint pain improved in patients with significant pain at baseline. Conclusions
rhASB treatment was well-tolerated and reduced lysosomal storage as evidenced by a dose-dependent reduction in urinary glycosaminoglycan. Clinical responses were present in all patients, but the largest gains occurred in patients with advanced disease receiving high-dose rhASB. (J Pediatr 2004;144:574-80)
ucopolysaccharidosis type VI (MPS VI; Maroteaux-Lamy syndrome) is a lysosomal storage disease in which affected persons lack the enzyme N-acetylgalactosamine 4-sulfatase (ASB).1 In the absence of this enzyme, the stepwise degradation of the glycosaminoglycan (GAG) dermatan sulfate (DS) is blocked. This results in the intracellular accumulation of partially degraded GAG in lysosomes of a wide range of tissues. The accumulation causes a chronic progressive disorder involving multiple organs that often results in death in the second decade of life. The estimated birth incidence of MPS VI ranges from 1:100,000 to 1:1,300,000 in various populations.2-5 The diagnosis of MPS VI is usually made at 6 to 24 months of age when children show progressive deceleration of growth, skeletal deformities, coarse facial features, and upper airway obstruction. Clouding of the cornea, communicating hydrocephalus, blindness, or heart disease may develop. MPS VI is not typically associated with progressive impairment of mental status, although physical limitations may impact learning and development. There is no satisfactory treatment for MPS VI. Most patients receive symptomatic treatment for specific problems as their only form of therapy. Although a few patients have benefited from bone marrow transplantation (BMT),6,7 it is not universally performed due to lack of a suitable donor and is associated with significant morbidity and mortality. The European Group for Bone Marrow Transplantation reported transplant-related mortality of 10% (HLA identical) to 20% to 25% (HLA mismatched) for 63 transplantation cases of lysosomal disorders.8
M
AHI ASB AUC0-t
CHAQ
574
Apnea-hypopnia index N-acetylgalactosamine 4-sulfatase Area under the plasma concentration-time curve from 0 to the final sample with a concentration equal to or greater than the qualified limit of detection of the assay Childhood Health Assessment Questionnaire
DMB ERT GAG MPS rhASB T½
1,9-dimethylmethylene blue Enzyme replacement therapy Glycosaminoglycan(s) Mucopolysaccharidosis Recombinant human N-acetylgalactosamine 4-sulfatase Elimination half-life
See editorial, p 561, and related articles, p 569 and p 581. From Pediatric Clinical Research Center, Children’s Hospital & Research Center at Oakland, California; Department of Pediatrics, University of Minnesota Medical School, Minneapolis, Minnesota; Pediatric Genetics and Metabolism, University of Texas Southwest Medical Center, Dallas, Texas; Pediatric Hematology/Oncology, East Tennessee Children’s Hospital, Knoxville, Tennessee; Division of Metabolism, Oregon Health & Science University, Portland, Oregon; U Klinik fur Kinder-und Jugendheilkunde, Graz, Austria; Section of Biochemical Genetics, Children’s Hospital Philadelphia, Philadelphia, Pennsylvania; and Department of Chemical Pathology, Women’s and Children’s Hospital Adelaide, Adelaide, Australia. This study was sponsored by BioMarin Pharmaceutical Inc. It was supported in part by PHS, National Center for Research, Resources grants: MO1_RR01271, MO1_RR00334, MO1_RR00240. Submitted for publication Aug 13, 2003; last revision received Feb 25, 2004; accepted Mar 4, 2004. Reprint requests: Paul Harmatz, MD, Children’s Hospital & Research Center at Oakland, 747 52nd St, Oakland, CA 94609. E-mail: Pharmatz@mail. cho.org. 0022-3476/$ - see front matter Copyright ª 2004 Elsevier Inc. All rights reserved. 10.1016/j.jpeds.2004.03.018
Table I. Patient demographics, disease severity (at baseline) Patient #
Age
Sex
Ethnicity
High dose: 1.0 mg/kg 42 43
7 16
M F
Caucasian Caucasian
44
12
F
African American
Low dose: 0.2 mg/kg 41 45 50
7 11 13
F M M
Caucasian Caucasian Caucasian
Ventilatory assistance
Major surgical procedures
None Nasal oxygen for 10 h at night Tracheostomy, CPAP/ BiPAP only with upper respiratory infections
None Ventriculoperitoneal shunt
None Tracheostomy Tracheostomy, CPAP/BiPAP for 8 h at night
None Tracheostomy Ventriculoperitoneal shunt, tracheostomy, spinal surgery
Ventriculoperitoneal shunt, tracheostomy
BiPAP, Bilevel positive air pressure; CPAP, continuous positive air pressure.
Enzyme replacement therapy (ERT) has been approved for use in three lysosomal storage disorders: Gaucher disease, Fabry disease, and MPS I.9 In each of these diseases, recombinant human enzyme synthesized in vitro is infused on a regular schedule. The infused enzymes utilize either mannose-6-phosphate (Fabry disease and MPS I) or mannose (Gaucher disease) receptor-mediated endocytosis to reach intracellular sites and facilitate degradation of material stored in lysosomes. Similarly, ERT with recombinant human ASB (rhASB) in a feline model of MPS VI was associated with a reduction of storage vacuoles in Kupffer cells and connective tissue, an increase in bone mineral volume, and greater mobility.10,11 Onthebasisoftheseanimalstudies,aclinicalstudy of rhASB replacement was initiated in patients with MPS VI.
METHODS Patients Seven patients with MPS VI were enrolled. All patients had clinical manifestations of the disease, and diagnosis was confirmed by leukocyte ASB enzyme activity of <20% of the normal range of the measuring laboratory. There were 3 females and 4 males, ages 7 to 16 years. Patient demographics and representation of disease severity for those completing the initial 24-week double-blind stage are presented in Table I. The protocol was approved by the institutional review board at each participating clinical site. Written consent was obtained from all parents or guardians before enrollment and written assent was obtained from all patients.
Study Design Patients were randomized to receive either 0.2 or 1.0 mg/kg of rhASB as weekly intravenous infusions in a doubleblind design. rhASB doses were selected for the current study based on previous studies in the feline model of MPS VI using human (1 and 5 mg/kg) and feline ASB (1 mg/kg) ERT.
Same-species feline rfASB (1 mg/kg) appeared to be at least equivalent to high-dose human rhASB (5 mg/kg) in the feline model.10 Thus, same species human rhASB at 0.2 and 1.0 mg/ kg were selected for the current study such that both doses were likely to show some efficacy. Safety and efficacy evaluations were completed at baseline and then at 1 to 6 week intervals. After the 24 week safety and efficacy evaluations, the study blind was removed, but all patients remained on their assigned dose until after the 48-week evaluation.
Study Drug rhASB was produced in a suspension bioreactor by genetically engineered Chinese hamster ovary cells. The enzyme was purified through a traditional column chromatography procedure and formulated in phosphate buffered saline at pH 5.8 containing 0.005% polysorbate 80. The specific activity of the formulated enzyme averaged 56 units/ mg. The purified enzyme contains a high level of bismannose-6-phosphate oligomannose oligosaccharide as confirmed by analytical assays and by the saturable robust ‘‘uptake’’ of enzyme by MPS VI fibroblasts in cell culture. Addition of 50 mM mannose-6-phosphate in the cell culture media reduced the cell uptake by more than 90%.
rhASB Administration All patients were premedicated with diphenhydramine (0.5 mg/kg body weight). rhASB was diluted in 0.9% saline and administered at 0.2 or 1.0 mg/kg over 4 hours once weekly. The infusion rate was adjusted so that approximately 2.5% of the total enzyme dose was infused during the first hour and the remaining enzyme dose (; 97.5%) was infused over the next three hours.
Biochemical Studies Studies to monitor toxicity were performed every one to six weeks and included complete blood count, chemistry panel,
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urinalysis, serum IgG anti-rhASB antibody by enzyme-linked immunosorbent assay, and measurement of serum complement before and after infusion. Urine was obtained every one to six weeks to determine total GAG, a surrogate for the extent of clearance of these compounds from lysosomal storage. Total GAG concentrations in urine samples were determined with a method based on spectrophotometric detection of metachromatic changes to the dye 1,9dimethylmethylene blue (DMB) resulting from GAG binding.12 GAG concentrations were subsequently normalized to urinary creatinine concentrations, which were determined separately. Total GAG was quantitated by measuring DMB binding, using dermatan sulfate as a standard. GAG concentrations (lg/mL) were divided by creatinine concentrations (converted to mg/mL) to yield a final reported value for GAG concentration in units of ‘‘lg of total GAG per mg of creatinine.’’ Pharmacokinetic studies were performed at 1, 2, 12, and 24 weeks. One-milliliter blood samples were collected from a second intravenous line placed in the arm opposite to that used for the enzyme infusion. rhASB levels were measured by enzyme linked immunosorbent assay. Pharmacokinetic parameters determined included area under the plasma concentration-time curve from 0 to the final sample with concentration equal to or greater than the limit of detection (AUC0-t ), maximum observed concentration, and elimination half-life (T½).
Clinical Evaluations Evaluation of mobility and physical function was performed at baseline and every 6 weeks, including a 6-minute walk test, joint range of motion, spirometry, and assessment of functional status. For the 6-minute walk test,13 patients were instructed to walk as far as possible in 6 minutes, but were allowed to rest when needed. Oxyhemoglobin saturation and heart rate were monitored by pulse oximetry (Nellcor 395). Range of motion (ROM) of the shoulders, elbows and knees was measured with a goniometer by occupational and physical therapists. Forced vital capacity (FVC) and forced expiratory volume at 1 minute (FEV1) were evaluated by standard spirometry technique according to American Thoracic Society guidelines. Functional status was assessed by an age-appropriate questionnaire, the Childhood Health Assessment Questionnaire (CHAQ),14 that was completed by a patient’s parent or guardian. Finally, each patient was videotaped while performing a standard set of physical movements based on activities in the Denver Developmental examination,15 including standing unassisted, walking forward, walking backward, throwing a ball, kicking a ball, jumping in place, balancing on each foot, hopping on each foot, catching a bounced ball, reaching for and grasping a ball above eye level, climbing stairs, building a tower of blocks, copying a square, pulling on a sweatshirt over the head, picking up and writing with a pen, pouring a cup of water, buttoning buttons, zipping a jacket, putting on and lacing shoes, and picking up coins and placing them into a cup. 576
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An ophthalmology evaluation including fundoscopic and slit-lamp examinations, assessment of glaucoma, and determination of visual acuity, was performed at baseline and every 12 weeks. Standard 12-lead electrocardiogram (EKG), 2-dimensional Doppler echocardiogram and chest radiography were performed at baseline and every 12 weeks. Liver volume and lumbar vertebral trabecular bone density were assessed by CT scan at baseline, week 12 and week 24. The liver volume was calculated from the axial image dataset using the postprocessing graphic workstation (Picker Omnipro/ Algotec Pro Vision software, Raanana, Israel), and bone density was determined using QCT Pro (Mindways Software, San Francisco, Calif ). Polysomnography was performed every 24 weeks according to the guidelines of the American Thoracic Society. The number of episodes of apnea and hypopnea, the number of minutes during sleep in which oxygen saturation fell below 89 percent, and the total sleeping time were recorded. An apnea-hypopnea index (AHI) was calculated by dividing the total number of episodes of apnea and hypopnea by the number of hours of sleep.
Statistics Descriptive statistics, including means, standard deviations (SD), and percent change over time, were calculated using Systat 10.2 (Systat Software, Inc, Richmond, Calif). Change in parameter between baseline and at 48 weeks was compared using the Wilcoxon signed rank test for small groups (Systat 10.2).
RESULTS Six of the 7 patients completed 24 weeks of treatment (Table I), 5 patients completed 48 weeks. One patient (#40) withdrew from the study at week 3 due to personal reasons unrelated to the study drug treatment, and another (#50) withdrew at week 32 because of a lack of perceived benefit. This patient was on low-dose rhASB at the time of study exit, and was unwilling to wait for institutional review board approval to be placed on the higher dose.
Safety There were 7 serious adverse events (SAEs) reported during the first 48 weeks of treatment. One event, mild desaturation during sleep (Patient #43), was subsequently determined not to be an adverse event, because the findings did not represent a change from baseline. Another event, brain glioma and colonic adenocarcinoma reported for Patient #40 approximately 10 months after the patient withdrew from the trial, was determined to be due to an unrelated genetic mutation. This patient withdrew from the trial after receiving only three infusions. The remaining five SAEs included one episode of vasovagal syncope at the time of intravenous line placement (Patient #40), hypoxia at the time of a routine tracheostomy tube change (Patient #44), paratracheal skin hypertrophy (Patient # 44), worsening eustachian tube dysfunction (Patient # 41), and bilateral carpal tunnel The Journal of Pediatrics May 2004
Fig 1. Urinary GAG excretion in patients during ERT by individual patients (A) and in patients grouped by dose (B). Urinary GAG excretion was measured in terms of relative reduction in excretion and expressed as the percent of the baseline excretion. Patient #50 (low dose) dropped out of the study after week 30, therefore, week 36-48 mean values for the low-dose group are calculated from one less data point. The difference between urinary GAG at baseline and that at week 48 (relative reduction) for the total population was significant (P = .04).
syndrome (Patient #41). The last three events were considered serious because the resulting surgical interventions required in-patient hospitalization. No SAEs occurred during study drug infusion, and none were considered related to study drug. There were no reports of allergic reactions (urticaria, angioedema) during infusion. Thirty-one adverse events occurred during the infusion. All were assessed as mild, and all resolved. Events of hypotension, tachypnea, skin irritation on neck and chest, pain at IV insertion site, and a 3-second run of premature ventricular contractions were assessed as possibly related to study drug, whereas the other events were considered to be unrelated. Adverse events (n = 126) were reported to occur during the first 48 weeks of treatment, 50 in the low-dose group and 76 in the high-dose group. The following adverse events were assessed as possibly related to study drug (number in parentheses is the number of discrete events): lethargy (2), hypotension (1), itching (4), rash/skin irritation (4), and intermittent joint popping (1). The majority of the adverse events were attributed to manifestations of the underlying disease.
Efficacy A significant relative decrease in urinary GAG was observed for the total population (P = .04) (Fig 1). The high dose produced a more rapid and larger sustained relative reduction in urinary GAG (63% vs 51% at week 48). Functional capacity of the patients improved during the study period. Five of the 6 patients (the sixth patient withdrew after week 30) improved in the 6-minute walk test at the 48week time point (Table II), with a significant improvement noted for the total population (P = .04). Dramatic gains were observed at week 48 for the two patients walking <100 meters at baseline (238% and 43% increase in meters walked).
Improvement in ability to walk up stairs was also demonstrated on the videotape of functional activities. Five of the six patients demonstrated improvement in shoulder range of motion at week 48 (the sixth patient withdrew after week 30). Restriction in shoulder flexion decreased significantly from baseline to week 48 for the total population (right shoulder: 84 to 68 degrees, P = .04; left shoulder: 83 to 67 degrees, P = .04). Smaller improvements were observed in the knees and elbows. Improvements in functional status were evidenced by investigator observation (including picking up coins, tying shoelaces, pulling shirt overhead) and by responses to the pain and arthritis questions on the CHAQ (P = .04, change from baseline to 48 weeks). Liver volumes were assessed at baseline and at the 24week time point. At baseline, only the two oldest patients’ livers were significantly enlarged for age and body weight, and these patients showed a 14% percent reduction in liver volume after 24 weeks of treatment. Spirometry examinations (FVC and FEV1) were limited by severe restrictive lung disease in several patients (#43, #44, #41, #50), difficulty producing the required 6-second expiration, and also the presence of tracheostomy in 3 patients (#44, #45, #50). Improvement in spirometry (FVC increased 11%) was observed in one patient (#42). Attempts to define upper airway obstruction by polysomnography were impeded by tracheostomies present in patients #44, #45, and #50. Of the other three patients, patients #41 and #42 showed improvement at week 48 with a 7.5- and 4-fold decrease, respectively, in AHI. Patient #43 had a decrease in the amount of nasal oxygen required during the sleep study at week 48, but no change in the AHI. There were no appreciable gains in height for any patient between baseline and week 48. Patients #44 and #45 experienced approximately 17% and 14% weight gains from baseline, respectively, whereas the other 4 patients experienced little or no weight gains.
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Fig 2. Circulating concentrations of rhASB in the 6 hours after initiation of rhASB infusion at week 1 (A) and changes in area under the curve (AUC0-t) values of rhASB in patients grouped by dose followed over 24-week study period (B). rhASB plasma concentrations during and 2 hours following the first infusion are shown for individual patients. Plasma rhASB concentrations confirmed two doses and the rhASB cleared rapidly from the circulation at the end of the infusion. The AUC0-t was calculated from the plasma concentrations of rhASB to the last time point with a concentration above the limit of quantitation (33.75 ng/mL). AUC0-t values confirmed two doses. Mean values for weeks 12 and 24 for the low-dose group are calculated from one less data point.
Table II. Six-minute walk test: Change in distance walked Patient # 42 43 44 High-dose mean 41 45 50 Low-dose mean
Dose (mg/kg) 1 1 1 0.2 0.2 0.2
Baseline meters (mean of 3 ± SD)
Week 6 meters (%)
Week 12 meters (%)
388 ± 60 53 ± 15 88 ± 12 176 ± 29 197 ± 21 283 ± 20 133 ± 19 204 ± 20
ÿ62 (ÿ16) 18 (34) 46 (52) 2 (23) ÿ97 (ÿ49) 29 (10) ÿ5 (ÿ4) ÿ24 (ÿ14)
ÿ43 (ÿ11) 53 (100) 65 (74) 25 (54) ÿ60 (ÿ30) 53 (19) ÿ22 (ÿ17) ÿ10 (ÿ9)
Week 24 meters (%) ÿ33 99 63 43 16 42 10 23
(ÿ9) (187) (72) (83) (8) (15) (8) (10)
Week 48 meters (%) 20 (5) 126 (238) 38 (43) 61 (95) 7 (4) 125 (44) DC 66 (24)
DC, Discontinued.
No appreciable changes from baseline to week 48 were observed for echocardiogram, bone density examinations, visual examinations, grip tests, or pinch strength testing.
Pharmacokinetics rhASB plasma concentrations during and 2 hours after the first infusion are shown for individual patients (Fig 2, A). Maximum observed concentration was 572 ± 60 ng/mL and 61 ± 11 ng/mL for the high- and low-dose groups, respectively. In general, the antigen was not measurable in the plasma within 10 minutes after the completion of the enzyme infusion in all patients (t½ = 4.31 minutes calculated for one patient in the high-dose group; unable to calculate for the low-dose group). rhASB pharmacokinetics appeared to be nonlinear, as reflected by greater than dose-proportionate increases in AUC0-t (Fig 2, B). AUC0-t increased relative to week 1 values in the high dose patients, but remained unchanged in the low dose patients. 578
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Antibody Formation All 6 patients seroconverted, developing IgG antirhASB antibodies, by 30 weeks of treatment. The higher dose was not associated with higher antibody levels. Antibody development was not associated with significant complement depletion (maximum 21%) during infusions or persistent levels outside the normal range, and did not appear to have any significant influence on safety or efficacy.
DISCUSSION The current study assessed the safety and efficacy of ERT with rhASB for the treatment of MPS VI in humans. The bioactivity of the enzyme was confirmed by the observation of decreases in total GAG that were reduced in a dose-dependent fashion. Functional status also improved as seen by an increase in distance walked, stair-climbing ability, shoulder range of motion, and decrease in pain and arthritis The Journal of Pediatrics May 2004
severity scores on the CHAQ. These improvements were most evident in the patients with more clinically advanced disease receiving high-dose ERT, and were achieved without serious adverse events, laboratory abnormalities or allergic reactions. Dose-dependent responses were not seen in the functional status parameters, possibly related to the small number of patients and large range of ages and disease severity within each group. The results of this study provide promising evidence that ERT may serve as an effective treatment for MPS VI disease. Significant gains have not been observed for pulmonary function in this study. Three patients had tracheostomies and could not be effectively tested. Of the 3 patients without tracheostomies, absence of improvement may relate to low enzyme dose in one patient (#41) and the presence of severe restrictive lung disease in an older patient with advanced disease (#43). The one patient showing a significant improvement in pulmonary function received high-dose enzyme and had less clinically advanced disease at baseline, particularly in the skeletal system. This may indicate that short-term improvements in FVC related to the clearance of stored GAG in the soft tissue surrounding the airway may not be appreciated once advanced changes in the skeletal system and trachea have occurred, or that longer treatment may be required to effect a change. Additional patients will need to be studied to confirm this supposition. Although no improvement in pulmonary function was observed in the patients with clinically advanced disease, it is possible that these patients, in the absence of ERT, would show disease-related deterioration over the one-year study period, a possibility that could not be assessed with the current study design. Improvements in secluded sites of GAG accumulation, including joint cartilage and corneas, were not observed after intravenous administration of rhASB in the feline study.11 More recently, Byers et al16 have reported in preliminary studies that ASB injection in the large weight-bearing joints in cats preserves cartilage. More notable was the finding that injections could be limited to every few months, giving this therapy a potential role in humans. Simonaro et al17 described increased release of nitric oxide and tumor necrosis factor-a in cultured human MPS VI articular chondrocytes, suggesting that ERT may act by reducing inflammation around the joint. In the current study, improvement in joint symptoms with decreased pain and improved range of motion was observed. We did not assess intra-articular structure or damage with MRI and, therefore, we cannot state whether joint/cartilage structure improved with ERT. Development of antibodies in the setting of ERT is common. In a large study of patients with Gaucher disease receiving ERT, 12.8% developed antibodies; however, 90% of individuals became tolerant over time.18 Similarly, our patients developed antibodies to ASB but no impact on clinical efficacy or biochemical activity was noted. It is possible that the high level of antibody in a high-dose patient (#43) at week 24 explains the prolonged survival of rhASB in the circulation and increased AUC0-t noted at 24 weeks. Although all patients in this study were older than five years and had significant disease, it is anticipated that, in the
future, ERT will be targeted for use in individuals at very early disease stages or even before the onset of clinical symptoms. It has been shown that earlier treatment has a more profound effect on skeletal pathology and, because fetal storage has been demonstrated in MPS VI kittens,19 it is anticipated that early treatment before the onset of irreversible pathology will maximize the benefits of therapy. Early treatment clearly depends on effective newborn screening and identification of patients at risk of severe disease. Programs for newborn screening are currently being proposed20-22 but are still in development and not in widespread use. In conclusion, our findings demonstrate that ERT with rhASB at 1 mg/kg results in clinical and biochemical improvements without significant side effects. The total urine GAG excretion measurements provide strong biochemical evidence for a stable, dose-dependent effect of ERT in these patients. The higher dose of 1.0 mg/kg appears to be superior to the lower dose of 0.2 mg/kg based both on the rapidity with which urinary GAG declined and on the relative reductions in urine GAG. The higher dose also resulted in larger gains in absolute levels of circulating enzyme. Because of the great clinical heterogeneity reported with MPS disease,23 it is difficult to design trials based on clinically relevant end-points. Clearly, the largest and most rapid improvements were seen in patients with advanced disease. Future studies to confirm efficacy and safety will depend on identification of clinical measures to capture and quantify the additional benefits observed in this Phase I/II Study. We appreciate the Austrian MPS Society’s support of one of the participating families. We would like to thank Dr Jerry Thompson (University of Alabama at Birmingham) for providing quantitative measurements of leukocyte ASB, Karen Hildebrand for her expertise in assessments of joint function, and Drs Hector Gutierrez, Richard Rowe, Howard Rosenfeld, Selim Koseoglu, Ronald Cohen, John R. Waterson, Edward Lammer, Kumar G. Belani, and David S. Beebe for their clinical expertise. We would like to acknowledge the efforts of the following study coordinators: Sandra A. Banta-Wright, RNC, MN, NNP (Oregon Health & Science University), Maggie Wolf (U Klinik fur Kinder-und Jugendheilkunde, Graz, Austria), Sheri McCaffery (Children’s Hospital of Philadelphia), Laurie Cariveau, RN, and Amy Deyle (University of Minnesota).
REFERENCES 1. Neufeld EF, Muenzer J. The mucopolysaccharidoses. In: Scriver C, Beaudet AL, Sly WS, et al, editors. The metabolic basis of inherited disease. New York: McGraw-Hill; 1995. p. 2465-93. 2. Lowry RB, Applegarth DA, Toone JR, MacDonald E, Thunem NY. An update on the frequency of mucopolysaccharide syndromes in British Columbia. Hum Genet 1990;85:389-90. 3. Meikle PJ, Hopwood JJ, Clague AE, Carey WF. Prevalence of lysosomal storage disorders. JAMA 1999;281:249-54. 4. Nelson J. Incidence of the mucopolysaccharidoses in Northern Ireland. Hum Genet 1997;101:355-8. 5. Poorthuis BJ, Wevers RA, Kleijer WJ, Groener JE, doJong JG, van Weely S, et al. The frequency of lysosomal storage diseases in the Netherlands. Hum Genet 1999;105:151-6. 6. Krivit W, Pierpont ME, Ayaz K, Tsai M, Ramsay NK, Kersey JH, et al. Bone-marrow transplantation in the Maroteaux-Lamy syndrome
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(mucopolysaccharidosis Type VI). Biochemical and clinical status 24 months after transplantation. N Engl J Med 1984;311:1606-11. 7. Krivit W. Maroteaux-Lamy syndrome (mucopolysaccharidosis type VI). Treatment by allogeneic bone marrow transplantation in 6 patients and potential for autotransplantation bone marrow gene insertion. Int Pediatr 1992;7:47-52. 8. Hoogerbrugge PM, Brouwer OF, Bordigoni P, The European Group for Transplantation. Allogeneic bone marrow transplantation for lysosomal storage diseases. Lancet 1995;246:1398-402. 9. Wraith JE. Advances in the treatment of lysosomal storage disease. Dev Med Child Neurol 2001;43:639-46. 10. Bielicki J, Crawley AC, Davey RCA, Varnai JC, Hopwood JJ. Advantages of using same species enzyme for replacement therapy in a feline model of mucopolysaccharidosis type VI. J Biol Chem 1999;274: 36335-43. 11. Crawley AC, Brooks DA, Muller VJ, Petersen BA, Isaac EL, Bielicki J, et al. Enzyme replacement therapy in a feline model of Maroteaux-Lamy syndrome. J Clin Invest 1996;97:1864-73. 12. Whitley CB, Ridnour MD, Draper KA, Dutton CM, Neglia JP. Diagnostic test for mucopolysaccharidosis I. Direct method for quantifying excessive urinary glycosaminoglycan excretion. Clin Chem 1989;35:374-9. 13. Nixon PA, Joswiak ML, Fricker FJ. A six-minute walk test for assessing exercise tolerance in severely ill children. J Pediatr 1996;129:362-6. 14. Singh G, Athreya BH, Fries JF, Goldsmith DP. Measurement of health status in children with juvenile rheumatoid arthritis. Arth Rheum 1994;37:1761-9. 15. Sperhac AM, Salzer JL. A new developmental screening test. The Denver II. J Am Acad Nurse Pract 1991;3:152-7.
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16. Byers S, Auclair D, Brumfield LK, Robinson A, Hopwood JJ. Localised therapy for joint disease in MPS VI. 2002. Paris, 7th International Symposium on MPS and Related Diseases. 6-20-2002. 17. Simonaro CM, Haskins ME, Schuchman EH. Articular chondrocytes from animals with a dermatan sulfate storage disease undergo a high rate of apoptosis and release nitric oxide and inflammatory cytokines: a possible mechanism underlying degenerative joint disease in the mucopolysaccharidoses. Lab Invest 2001;81:1319-28. 18. Rosenberg M, Kingma W, Fitzpatrick MA, Richards SM. Immunosurveillance of alglucerase enzyme therapy for gaucher patients: induction of humoral tolerance in seroconverted patients after repeat administration. Blood 1999;93:2081-8. 19. Crawley AC, Niedzielski KH, Isaac EL, Davey RCA, Byers S, Hopwood JJ. Enzyme replacement therapy from birth in a feline model of mucopolysaccharidosis type VI. J Clin Invest 1997;99:651-62. 20. Meikle PJ, Ranieri E, Ravenscroft EM, Hua CT, Brooks DA, Hopwood JJ. Newborn screening for lysosomal storage disorders. Southeast Asian J Trop Med Public Health 1999;30:104-10. 21. Whitley CB, Draper KA, Dutton CM, Brown PA, Severson SL, France LA. Diagnostic test for mucopolysaccharidosis II. Rapid quantification of glycosaminoglycan in urine samples collected on a paper matrix. Clin Chem 1989;35:2074-81. 22. Whitley CB, Spielman RC, Herro G, Severson ST. Urinary glycosaminoglycan excretion quantified by an automated semimicro method in specimens conveniently transported from around the globe. Mol Genet Metabol 2002;75:56-64. 23. Wraith JE. Enzyme replacement therapy in mucopolysaccharidosis type I: progress and emerging difficulties. J Inherit Metab Dis 2001;24:245-50.
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