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2 study of intrathecal heparan-N-sulfatase in patients with mucopolysaccharidosis IIIA
Molecular Genetics and Metabolism 118 (2016) 198–205
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A phase 1/2 study of intrathecal heparan-N-sulfatase in patients with mucopolysaccharidosis IIIA Simon A. Jones a, Catherine Breen a, Fiona Heap a, Stewart Rust b, Jessica de Ruijter c, Evelien Tump c, Jan Pieter Marchal c, Luying Pan d, Yongchang Qiu d, Jou-Ku Chung d, Nitin Nair d,1, Patrick A.J. Haslett d,2, Ann J. Barbier d,3, Frits A. Wijburg c,⁎ a
Willink Unit, Manchester Centre for Genomic Medicine, St Mary's Hospital, Central Manchester University Hospitals NHS Foundation Trust (CMFT), University of Manchester, United Kingdom Paediatric Psychosocial Department, Royal Manchester Children's Hospital, Manchester, United Kingdom c Department of Paediatrics, Academic Medical Center, Amsterdam, The Netherlands d Shire, Lexington, MA, USA b
a r t i c l e
i n f o
Article history: Received 9 May 2016 Accepted 9 May 2016 Available online 10 May 2016 Keywords: Enzyme replacement therapy Heparan sulfate Intrathecal drug delivery device Lysosomal storage disease Mucopolysaccharidosis IIIA (MPS IIIA) Sanfilippo syndrome A
a b s t r a c t Objective: This was an open-label, phase 1/2 dose-escalation, safety trial of intrathecal recombinant human heparan-N-sulfatase (rhHNS) administered via intrathecal drug delivery device (IDDD) for treating mucopolysaccharidosis IIIA (NCT01155778). Study design: Twelve patients received 10, 45, or 90 mg of rhHNS via IDDD once monthly for a total of 6 doses. Primary endpoints included adverse events (AEs) and anti-rhHNS antibodies. Secondary endpoints included standardized neurocognitive assessments, cortical gray matter volume, and pharmacokinetic/pharmacodynamic analyses. Results: All patients experienced treatment-emergent AEs; most of mild-to-moderate severity. Seven patients reported a total of 10 serious AEs (SAEs), all but one due to hospitalization to revise a nonfunctioning IDDD. No SAEs were considered related to rhHNS. Anti-rhHNS antibodies were detected in the serum of 6 patients and in the cerebrospinal fluid (CSF) of 2 of these. CSF heparan sulfate levels were elevated at baseline and there were sustained declines in all tested patients following the first rhHNS dose. No impact of anti-rhHNS antibodies on any pharmacodynamic or safety parameters was evident. 4 of 12 patients showed a decline in developmental quotient, 6 were stable, and 2 patients had only a single data point. No dose group showed a clearly different response pattern. Conclusions: rhHNS administration via IDDD appeared generally safe and well tolerated. Treatment resulted in consistent declines in CSF heparan sulfate, suggesting in vivo activity in the relevant anatomical compartment. Results of this small study should be interpreted with caution. Future studies are required to assess the potential clinical benefits of rhHNS and to test improved IDDD models. © 2016 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
1. Introduction Abbreviations: AE, adverse event; BSID-III, Bayley Scales of Infant and Toddler Development, Third Edition; CSF, cerebrospinal fluid; CNS, central nervous system; DQ, developmental quotient; EC, Enzyme Commission; ELISA, enzyme-linked immunosorbent assay; GAG, glycosaminoglycan; IDDD, intrathecal drug delivery device; IT, intrathecal; KABC-II, Kaufman Assessment Battery for Children, Second Edition; LC-MS/MS, liquid chromatography-tandem mass spectrometry; MPS IIIA, mucopolysaccharidosis IIIA/ Sanfilippo syndrome type A; MRI, magnetic resonance imaging; rhHNS, recombinant human heparan-N-sulfatase (EC 3.10.1.1); SAE, serious adverse event; TEAE, treatmentemergent adverse event; Tmax, time to maximum serum concentration; uGAG, urinary glycosaminoglycan; VABS-II, Vineland Adaptive Behavior Scales, Second Edition. ⁎ Corresponding author at: Department of Paediatrics (H7-270), Academic Medical Center, Meibergdreef 9, 1105 AZ Amsterdam, The Netherlands. E-mail address: [email protected] (F.A. Wijburg). 1 Current address: Vertex Pharmaceuticals, Inc., 50 Northern Ave, Boston, MA 02210. 2 Current address: Alnylam Pharmaceuticals, Inc., 300 Third Street, Cambridge, MA 02142. 3 Current address: Agios Pharmaceuticals, 88 Sidney Street, Cambridge, MA 02139.
Mucopolysaccharidosis IIIA (MPS IIIA, Sanfilippo syndrome type A; OMIM 252900) is a lysosomal storage disorder caused by a deficiency of the enzyme heparan-N-sulfatase (EC 3.10.1.1), leading to accumulation of the glycosaminoglycan (GAG), heparan sulfate, in the lysosomes [1]. The clinical manifestations of MPS IIIA are primarily neurologic, with rapid and progressive disease manifestation, which are in contrast to the relatively mild somatic symptoms compared with other types of MPS disorders [1–3]. Neurologic signs include developmental delay, behavioral abnormalities, and sleep disturbances [2,3]. Life span is usually severely shortened [3]. There are no approved disease-modifying therapies available for MPS IIIA. Preclinical safety of recombinant human heparan-N-sulfatase (rhHNS) formulated for intrathecal (IT) delivery, has been demonstrated in toxicology studies performed in juvenile cynomolgus monkeys,
http://dx.doi.org/10.1016/j.ymgme.2016.05.006 1096-7192/© 2016 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
S.A. Jones et al. / Molecular Genetics and Metabolism 118 (2016) 198–205
where the drug was administered via a surgically implanted intrathecal drug delivery device (IDDD) [4]. Here we report the results from the first-in-human, phase 1/2 clinical trial (ClinicalTrials.gov identifier: NCT01155778) of rhHNS. The primary objective of the study was to determine the safety and tolerability of rhHNS in patients with MPS IIIA via ascending doses administered intrathecally through a surgically implanted IDDD once monthly for 6 months. 2. Patients and methods 2.1. Patients Patients were enrolled from the Academic Medical Center in Amsterdam, the Netherlands, and St Mary's Hospital in Manchester, United Kingdom. Included patients were required to have heparan-N-sulfatase enzyme activity ≤10% of the lower limit of normal as measured in fibroblasts or leukocytes, plus either (1) a normal enzyme activity level of at least one other sulfatase (to rule out multiple sulfatase deficiency) as measured in fibroblasts or leukocytes, or (2) two documented mutations in the SGSH gene. Included patients were ≥3 years of age, with a developmental age ≥ 1 year, based on the results of a developmental questionnaire at the time of screening (see Endpoints and assessments). The exclusion criteria were significant central nervous system (CNS) impairment or behavioral disturbances with causes other than MPS IIIA, previous implantation of a CNS shunt, poorly controlled seizure disorder, prior hematopoietic stem cell or bone marrow transplantation, and treatment with any investigational drug or a device intended as a treatment for MPS IIIA within the 30 days prior to the study. The study was conducted in compliance with the Declaration of Helsinki, the relevant Institutional Review Board regulations, and the International Conference on Harmonisation Good Clinical Practice guidelines. Written informed consent was obtained from patients or their parents/legal guardians. 2.2. Study design This was a phase 1/2 multicenter, open-label, multiple-dose, doseescalation study. Patients were enrolled sequentially into each of 3 treatment arms: 10, 45, or 90 mg rhHNS via IDDD dosed every 4 weeks (28 days ± 7 days). A total of 6 doses were planned. Enrollment was staggered so that initial safety could be assessed on the first 2 patients within each dose group. Moreover, dose escalations were only allowed after a review of safety data from 4 patients at the preceding dose level by an independent monitoring board. The study had no control arm. All patients were implanted with the IDDD (PORT-A-CATH® II Low Profile™; Smiths Medical ASD, Inc., St. Paul, Minnesota, USA) on week 1, day 1, followed by a recovery period of at least 7 days. Patients were admitted to the study center 24 h prior to their first rhHNS dose for safety and baseline assessments and were not discharged until approximately 24 h after the injection. Patients continued to receive safety assessments before and following each subsequent dose of rhHNS. In the event of IDDD malfunction, the dose of rhHNS could be administered via lumbar puncture. 2.3. Endpoints and assessments The primary objective was to assess the safety and tolerability of intrathecal administration of rhHNS in patients with MPS IIIA. Primary endpoints included observations of adverse events (AEs) and changes in clinical laboratory values, including cerebrospinal fluid (CSF) cell counts and chemistries, electrocardiograms, and anti-rhHNS antibodies in the CSF and serum. The secondary objectives of the study were to determine by dose group the effects of intrathecal administration of rhHNS on standardized neurocognitive assessments, cortical gray
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matter volume as assessed by magnetic resonance imaging (MRI), pharmacokinetic parameters, and the pharmacodynamic responses. The primary pharmacodynamic variable was the level of heparan sulfate in the CSF. The level of total heparan sulfate in the CSF was determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Briefly, heparan sulfate in the CSF was first extracted using an anion-exchange resin and then digested by a combination of enzymes including heparinase I, II, and III. The resultant heparan sulfate disaccharides were labeled with 12C-4-N-butylaniline by reductive amination and then analyzed by LC-MS/MS. The disaccharides were quantified based on a calibration curve generated using 6 commercially available disaccharide standards that are the most abundant in human CSF heparan sulfate. The secondary pharmacodynamic measurement was urinary glycosaminoglycan (uGAG) concentration, determined by a 1,9dimethylmethylene blue dye-binding assay using the Blyscan™ GAG assay kit (Biocolor Ltd., Carrickfergus, Northern Ireland, United Kingdom), and was reported relative to creatinine concentration (mg GAG/ mmol creatinine) [5]. Serum samples for pharmacokinetic testing were collected at the time of the first and last dose of rhHNS. Samples were collected immediately prior to the intrathecal injection and then at 0.5, 1, 1.5, 2, 3, 4, 6, 8, 12, 24, 48, and 72 h following completion of the intrathecal injection. The rhHNS concentrations were measured using an enzyme-linked immunosorbent assay (ELISA) with paired anti-rhHNS antibodies. Testing for serum and CSF antibodies against rhHNS was conducted at week 2 and at weeks 6, 10, 14, 18, 22, and 26. Antibody assessment was performed via a tiered approach by a bridging electrochemiluminescent immunoassay using paired biotin- and sulfo-tagged rhHNS. This assay detects total antidrug antibodies and does not distinguish immunoglobulin isotypes. Samples that screened positive for anti-rhHNS antibodies were confirmed by a ligand-competition assay, and then further tested for antibody titer with the same assay platform. Antibodies were not tested for their ability to neutralize rhHNS. CSF samples were collected from patients through the implanted IDDD or, in the event of a nonfunctioning IDDD, via lumbar puncture immediately prior to administration of rhHNS. Urine samples were collected at weeks 2, 6, 10, 14, 18, 22, and 26. To determine study eligibility, the patient's level of functioning was estimated at screening using the Vineland Adaptive Behavior Scales, Second Edition (VABS-II) [6]. The VABS-II is a caregiver-reported outcome that measures adaptive behaviors, including the ability to cope with environmental changes, to learn new everyday skills, and to demonstrate independence. The test measures various key domains (communication, daily living skills, socialization, and motor skills) and provides an adaptive behavior composite (a composite of the other 4 domains). Formal neurodevelopmental testing of enrolled patients was performed at weeks 2 and 22 using either the Bayley Scales of Infant and Toddler Development, Third Edition (BSID-III) [7] or the Kaufman Assessment Battery for Children, Second Edition (KABC-II) [8]. The BSID-III is designed for the neurodevelopmental assessment of children 0 to 42 months of age, whereas the KABC-II is designed to assess children from 3 to 18 years old. All children who were ≤ 42 months of age at enrollment, or who at screening had an average developmental age b36 months across all domains of the VABS-II, were assessed using the BSID-III. Children aged N42 months and with a developmental age N36 months were administered the KABC-II, unless they were unable to cooperate, in which case the BSID-III was administered. The test used at baseline was also used at week 22. Both tests served to ascertain the mental age equivalent in months of each child, using data obtained from the cognitive domain of the BSID-III and the mean age equivalent of the nonverbal domains of the KABC-II. The developmental quotient (DQ) was calculated by dividing the mental age equivalent by the calendar age in months, multiplied by 100, as described by Delaney et al. [9]. Cortical gray matter volumes were obtained by automated volumetric analysis of MRI scan data, using FreeSurfer software (Athinoula A. Martinos Center for Biomedical Imaging, Charlestown, MA). Total
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Table 1 Patient demographics at screening. rhHNS dose group, mg
10
45
90
Patient no.
Sex
Age, year
BMI
1 2 3 4 5 6 7 8 9 10 11 12
F M M M M F M F M M M F
13 5 13 5 3 4 6 24 4 4 12 22
22.9 20.4 22.8 17.8 17.5a 19.5 15.0 24.2 20.1 18.4 18.7 23.4
F, female; M, male; rhHNS, recombinant human heparan-N-sulfatase. a Patient was noncompliant at screening; these measurements were taken at week 1, day 7.
brain cortical gray matter volume was determined by automated analysis of brain MRIs performed at weeks 2 and 22, and results were read centrally. All calculations were performed using SAS v9.1 or later (SAS Institute Inc., Cary, NC). 3. Results 3.1. Study population Twelve patients were enrolled (4 females) with a median age of 5.5 years (range, = 3–24 years; Table 1). Four patients (2 pairs of siblings) had a relatively attenuated disease phenotype by clinical impression at baseline: patients 8 and 12, and patients 3 and 11. A fifth patient (patient 7) had an intermediate clinical phenotype at baseline. The remaining 7 patients had a severe phenotype. All the patients' SGSH mutations were compound heterozygous and varied across the 3 dose groups. In the 10 mg IT dose group one patient had a missense/unclassifiable (SGSH allele 1/allele 2) mutation, one had a missense/missense, and 2 had missense/frameshift mutations. In the 45-mg IT dose group 2 patients had a missense/unclassifiable mutation, one had a nonsense/nonsense, and one had a missense/missense mutation. In the 90-mg IT dose group one patient had an unclassifiable/missense mutation, one had a frameshift/frameshift, and 2 had missense/ missense mutations. 3.2. Safety results Treatment with rhHNS appeared generally safe and well tolerated. No clinically meaningful trends were observed in laboratory values, vital signs, physical examinations, or electrocardiographic assessments. There were no clinical signs or symptoms of meningeal irritation following enzyme infusion, and no change in the predose CSF clinical laboratory parameters to suggest meningeal inflammation.
All patients experienced at least 1 treatment-emergent AE (TEAE). The majority of these were mild to moderate in severity, and all resolved. There was no evidence of a higher frequency of TEAEs with higher doses of rhHNS. Four patients (33%) reported rhHNS-related AEs. These included fatigue, pain, pyrexia, cognitive disorder, memory impairment, motor dysfunction, malaise, pallor, and transient symptoms suggesting saddle paresthesia. Six patients (50%) reported IDDDrelated AEs. These included device failure, device breakage, and device component issues; device change; postoperative wound infections; and urinary incontinence. Three patients (25%) reported AEs that were related to the intrathecal administration process, including malaise; nausea and vomiting; and transient symptoms suggesting saddle paresthesia. Two patients (17%) reported AEs that began within 24 h after the start of injection and were judged possibly/probably related to rhHNS: 1 patient experienced irritability, vomiting, and increased CSF protein and white blood cell count; and 1 patient experienced pyrexia. Ten patients (83%) reported surgery-related AEs. Seven (58%) patients reported a total of 10 serious AEs (SAEs). This included 3 patients in the 10-mg dose group who reported 5 SAEs, 3 patients in the 45-mg dose group who reported 3 SAEs, and 1 patient in the 90-mg dose group who reported 2 SAEs. No SAEs were considered to be related to rhHNS; all were designated SAEs because of hospitalization, and 9 of 10 SAEs were related to the IDDD. The exception was a hospitalization for a suspected intercurrent viral infection unrelated to the study drug or procedures. A summary of IDDD failures is presented in Table 2, revealing that all failures were mechanical in nature owing to disconnection, breakage, or dislocation of the device from the intrathecal space. Patient 12 experienced 2 IDDD failures and received the remaining rhHNS doses via lumbar puncture. No patients discontinued from the study, and no patients died during the study. In total, 9 of 12 patients (75%) received all 6 of their scheduled doses, and 3 of 12 patients (25%) received 5 of their 6 scheduled doses. 3.3. Immunogenicity Serum antibodies to rhHNS were detected in 6 of 12 (50%) patients (Fig. 1). The response did not appear to be dose-dependent. Two patients (16.7%) were positive for low-titer antibodies in serum at baseline and demonstrated an increase in titer over the course of the study. In the other 4 patients, seroconversion occurred within the first 10 weeks of the study. One patient with low-serum antibody titer reverted to antibody-negative status at the week 26 visit. Low-titer antibodies in the CSF were detected for 1 patient in the 10-mg dose group beginning at week 18 and in 1 patient in the 90-mg dose group at week 22. Both patients had high titers of serum antibodies. One of the patients with pre-existing antibodies who also developed the highest titer of serum rhHNS antibodies and was positive for CSF antibodies had frame-shift mutations for both alleles. The other patient with pre-existing antibodies, and who developed a moderate serum antibody titer, was negative for CSF antibodies and had nonsense mutations for both alleles.
Table 2 Summary of IDDD failures. Patient No.
No. of doses via IDDD before First Failure Cause of first failure
No. of doses via IDDD before second failure
Cause of second failure
1 2 3
0 1 3
N/A N/A N/A
– – –
7
1
N/A
–
8 12
4 0
N/A 1
– Catheter migrated out of spinal canal
IDDD, intrathecal drug delivery device; N/A, not available.
Port/catheter disconnection Port outlet pinbreak Catheter migrated out of spinal canal Catheter migrated out of spinal canal Port outlet pinbreak Catheter migrated out of spinal canal
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patients tested (3/4) showed a decline in CSF levels that ranged from 58% to 75%. Mean heparan sulfate levels in CSF for the different dose groups over time are shown in Fig. 4. There was no apparent effect of antibody status on CSF levels of heparan sulfate (data not shown). At baseline, mean uGAG levels were elevated for all 3 study groups: 10-mg dose-group, 48.8 mg GAG/mmol creatinine (range, 38.2–63.9 mg GAG/mmol creatinine) (n = 4); 45-mg dose-group, 41.1 mg GAG/mmol creatinine (range, 13.3–74.8 mg GAG/mmol creatinine) (n = 4); 90-mg dose-group, 45.1 mg GAG/mmol creatinine (range, 23.7–70.0 mg GAG/mmol creatinine) (n = 4). By week 22, persistent uGAG reduction was observed in 3/4 patients in the 10-mg dosegroup, 3/4 patients in the 45-mg dose-group, and all patients (4/4) in the 90-mg dose-group. The decline was generally evident in urine collected 1 month later (week 6) after the first dose of intrathecal administration of rhHNS in 9/12 patients (Figs. 5 and 6). Normal reference range for uGAG levels are b10.5 mg GAG/mmol creatinine in children aged 4–5 years, and b 5.5 mg GAG/mmol creatinine in those aged N13 years [10]. Overall, there was no clear dose–response relationship for uGAG levels. Fig. 1. Semilogarithmic plot of serum anti-rhHNS antibody titers over time in the 6 patients exhibiting seropositivity. One patient developed very low titer serum antibodies since week 10, and reverted to serum antibody negative at week 26; two patients were positive for anti-rhHNS antibodies at baseline. For plotting purposes, an artificial titer of 1 was assigned to samples with negative antidrug results. rhHNS, recombinant human heparan-N-sulfatase.
3.4. Serum rhHNS pharmacokinetics At week 2 of intrathecal administration, rhHNS exhibited biphasic serum concentration-time profiles across the 10-, 45-, and 90-mg dose groups. Following the first intrathecal dose, the time to maximum serum concentration (Tmax) occurred between approximately 3 and 6 h, indicating a gradual transfer of rhHNS from the CNS to systemic compartments (Fig. 2A and B). Systemic exposure of rhHNS was doseproportional following the first dose of rhHNS (week 2; Fig. 2A), but not following the sixth dose (week 22; Fig. 2B). A high degree of variability was observed across pharmacokinetic parameters, which may have been due to the small number of patients within each dose group.
3.5. Pharmacodynamic response: levels of CSF heparan sulfate and uGAGs To establish a reference range for levels of heparan sulfate in the CSF, we analyzed 156 CSF samples from children without MPS from whom CSF had been obtained for other clinical reasons. These analyses indicated CSF levels of heparan sulfate ranged from below the limit of quantification (0.300 μM) to 0.501 μM (data not shown). In the current study, CSF levels of heparan sulfate were elevated at baseline for all 3 treatment groups relative to the non-MPS pediatric controls. Following intrathecal administration of rhHNS, CSF levels of heparan sulfate exhibited marked declines in all tested patients (11/12) from the 3 treatment groups. The CSF samples from patient 10 were not collected at weeks 6, 10, 14, and 18 for bioanalysis, and the week 22 CSF sample volume was insufficient to allow for HS testing. Four other patients (4/12) did not have sufficient sample CSF volume for HS quantification at week 6. Comparing the individual patient CSF heparan sulfate level data (Fig. 3), all patients tested (11/12) showed declines in their CSF heparan sulfate levels compared to their baseline (2 weeks) levels by week 22 or the end of the study, and a reduction in their CSF heparan sulfate level after the first IT dose was observed in all patients (7/12) who had week 6 samples available for testing. Those in the 10-mg dose group showed the least decline (11–52%) compared to baseline, whereas the 45-mg and 90-mg patients showed greater declines versus baseline (50–78%). In the 90-mg dose group all
Fig. 2. Mean serum rhHNS concentration-time profiles of patient samples following intrathecal administration of rhHNS by dose group at (A) week 2 and (B) week 22. Error bars are standard error of mean. rhHNS, recombinant human heparan-N-sulfatase.
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Fig. 4. Mean heparan sulfate levels in CSF over time by rhHNS dose group. CSF, cerebrospinal fluid. Error bars are standard error of mean.
each from the 10- and 45-mg dose groups and 2 patients from the 90mg dose group underwent the BSID-III. Two patients in the 10-mg dose group did not have baseline cognitive assessment data due to their inability to cooperate, but in these cases, cognitive data were successfully obtained by the BSID-III at week 22. Of the 12 patients, 4 showed a decline in DQ, 6 were essentially stable, showing a small DQ increase or decrease of similar magnitude that is not clinically significant, and 2 patients had only a single data point. No dose group showed a clearly different pattern of response (Fig. 7). 3.7. Cortical gray matter volume Total brain cortical gray matter volume was determined by automated analysis of brain MRIs performed at weeks 2 and 22. Declines in total cortical gray matter volume were observed in all dose groups and in most patients between weeks 2 and 22 (Fig. 8). However, 2 patients, siblings with a highly attenuated phenotype and relatively preserved cognitive function, did not show reductions in this parameter. 4. Discussion
Fig. 3. Heparan sulfate levels in CSF for individual patients. Week 2 levels were used as baseline. CSF, cerebrospinal fluid.
3.6. Neurodevelopmental analyses Cognitive status was assessed at screening and at week 22. Only one site performed the VABS-II assessment at both screening and at week 22; thus, 6 patients have VABS-II data from 2 time points (Table 3). Cognitive status was also assessed at screening and at week 22 by either the BSID-III or KABC-II (see Patients and methods). Three patients
The primary objective of the trial was to assess the safety and tolerability of rhHNS administered via IDDD in children with MPS IIIA. There was no significant clinical safety signal associated with the drug itself, which appeared to be generally well tolerated. All but one of the SAEs, which occurred in more than half of the patients, were related to malfunctions of the IDDD. None were related to the study drug. No patients died during the study, and none discontinued. IDDD failures occurred at an unexpectedly high rate, and were due to mechanical problems in the form of disconnection or breakage of the device or anatomical displacement of the catheter with migration out of the intrathecal space. Failures of similar devices have been reported in the literature, primarily due to mechanical malfunction [11,12]. These have been reported as occurring in up to 24% of patients, with disconnection of the catheter from the pump/port and migration of the catheter from the spinal canal accounting for most cases [13]. We found a similar pattern of failure in this study, but the overall frequency was higher than those reported in other clinical settings. It is possible that the hyperactivity and restlessness that frequently complicate progressive MPS IIIA may have increased the rate of IDDD complications. A next-generation IDDD has been developed to address these issues for the extension of the study and potential future studies
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Fig. 6. Normalized mean uGAG levels over time by rhHNS dose group. uGAG, urinary glycosaminoglycan.
Fig. 5. Normalized uGAG levels over time in individual patients. Week 2 levels were used as baseline. uGAG, urinary glycosaminoglycan.
(NCT01299727, 02060526). Another consideration is that rhHNS could have leaked out into surrounding tissues due to catheter migration or disconnection and this leakage could potentially impact uGAG and serum anti-rhHNS antibody levels, We think that, if this did occur, then it would not have had significant effect as the serum pharmacokinetic data (Fig. 2) show that there is a considerable amount of enzyme released into the systemic circulation after IT delivery in patients who were not experiencing device failure. This systematic release is likely
to be the reason for the effect of IT administration on uGAG levels, and this far outweighs any contribution from potential leakage. The question of whether enzyme delivered into the subcutaneous tissue could increase anti-drug antibodies is an interesting one, for which we currently have no data available for evaluation. Systemic exposure of rhHNS was dose-proportional following the first dose of rhHNS, although the cause of disturbed dose proportionality following the sixth dose is unknown (Fig. 2B). There are a number of possible reasons for this. Anti-rhHNS antibodies could have interfered with the ELISA method used for drug quantification. Other possibilities are that a greater or lesser amount of the drug was administered due to IDDD malfunction, or the presence of antibodies alters the distribution and clearance of rhHNS. A combination of some or all of these factors could produce the observed effect. Intrathecal administration of rhHNS proved to be immunogenic, with half the patients exhibiting either de novo anti-rhHNS antibody formation (n = 4) or increases in anti-rhHNS titers from baseline levels (n = 2). The reason for anti-rhHNS reactivity at baseline is unknown; however, during assay development, anti-rhHNS antibodies were found in 2 of 25 untreated patients with MPS IIIA, but none were found in 54 normal healthy donors, suggesting that baseline serum cross-reactivity might be a feature of MPS IIIA disease (data not shown). In 5 of 6 patients with anti-rhHNS antibodies, including those with baseline antibodies, there was a clear increase in titer over time. However, 1 patient displayed only very low titers from weeks 10 to 22, and reverted to seronegativity at the end of study visit. There were 2 patients with transient, extremely low-titer anti-rhHNS antibodies in the CSF that were associated with high-titer serum antibodies, suggesting possible diffusion of serum antibodies into the CSF. Of note, immunogenicity of therapeutic enzymes present in the systemic circulation has been reported in other MPS diseases [14–16]. Fifty percent of patients (6/12) developed anti-rhHNS antibodies after receiving ERT including 2 who had pre-existing serum antirhHNS antibodies with low titers. One of these patients with preexisting antibodies was positive for CSF antibodies and had frameshift mutations for both alleles and the other, who was negative for CSF antibodies, had nonsense mutations for both alleles. In a recent natural history study of MPS IIIA patients, 2 patients (2/25) were positive for anti-rhHNS antibodies with low titers and both of these patients had missense mutations for both alleles [17]. It has been shown in MPS II patients receiving idursulfase ERT that higher antibody titers are correlated to patients with complete deletion/large rearrangement and frameshift/splice site mutations when compared to those with
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Table 3 Adaptive behavior composite scoresa as assessed by the VABS-II at screening (n = 12) and at week 22 (n = 6). Patient no.
Sex
rhHNS dose group, mg
Visit
Age at visit
Composite score
Age equivalent
1
F
10
2
M
10
3 4 5
M M M
10 10 45
6
F
45
7 8 9
M F M
45 45 90
10
M
90
11 12
M F
90 90
Screening week 22 Screening week 22 Screening Screening Screening week 22 Screening week 22 Screening Screening Screening week 22 Screening week 22 Screening Screening
13 years, 2 months 13 years, 7 months 5 years, 5 months 5 years, 10 months 13 years, 2 months 4 years, 9 months 3 years, 1 month 3 years, 7 months 3 years, 6 months 3 years, 11 months 6 years, 0 month 23 years, 5 months 4 years, 2 months 4 years, 9 months 3 years, 11 months 4 years, 5 months 11 years, 9 months 22 years, 4 months
50 45 59 46 83 63 76 63 80 69 62 20 56 56 72 61 90 20
2 years, 11 months 2 years, 5 months 2 years, 0 month 1 year, 1 month 10 years, 5 months 2 years, 2 months 2 years, 1 month 1 year, 6 months 2 years, 6 months 2 years, 1 month 2 years, 7 months 3 years, 2 months 1 year, 6 months 1 year, 8 months 2 years, 3 months 1 year, 10 months 10 years, 7 months 0 year, 3 months
Week 22 testing was only performed at one study site. F, female; M, male; rhHNS, recombinant human heparan-N-sulfatase; VABS-II, Vineland Adaptive Behavior Scales, Second Edition. a Composite adaptive behavior score is a composite of the 4 constituent domains of the VABS-II (normal = 100).
missense mutations [18]. Given the small number of patients in this study, no firm conclusions can be reached about any relation between genotype and immunogenicity. There were no clear safety signals associated with the presence of anti-rhHNS antibodies in the study. The clinical significance of the development of antidrug antibodies in this patient population is unclear. Serum pharmacokinetic analyses showed dose-proportional patterns in peripheral blood following the first intrathecal dose of rhHNS. Linear dose proportionality in serum pharmacokinetics was not observed at week 22 following the sixth dose, and this may have been related to the development of serum anti-rhHNS antibodies, and in one case, to IDDD failure. The primary pharmacodynamic parameter of biochemical efficacy, total heparan sulfate in the CSF, exhibited declines in response to therapy at all dose levels, with a greater impact observed in the higher dose groups. Remarkably, intrathecal administration of rhHNS also reduced uGAG concentrations at all dose levels, suggesting systemic activity of the drug despite the extremely low blood levels indicated by the pharmacokinetic analysis. Over the duration of this shortterm study, no impact of anti-rhHNS antibodies on any pharmacodynamic parameters was evident.
Fig. 7. Spaghetti plot of individual BSID-III/KABC-II DQ scores by chronological age (n = 12). Each patient's rhHNS dose group is indicated. Note: DQ was calculated from the mean developmental age. BSID-III, Bayley Scales of Infant and Toddler Development, Third Edition; DQ, developmental quotient; KABC-II, Kaufman Assessment Battery for Children, Second Edition.
Although the majority of patients exhibited declines in DQ during the study, the study was neither powered nor designed to adequately assess clinical efficacy. The wide range of patient ages, disease phenotypes, and disease stages renders interpretation of data very challenging. Additionally, the 6-month duration of this study is likely to be too short to reasonably expect a meaningful change in these parameters in response to therapy. Interpretation was further limited by the absence of week 2 cognitive data in 2 patients in the 10-mg dose group who were unable to cooperate with the assessment. In all but 2 patients, cortical gray matter volume, a potential biomarker of MPS IIIA brain pathology, showed declines between weeks 2 and 22. The exception was a sibling pair with particularly attenuated disease and unusually wellpreserved cognition. The clinical significance of small increases or decreases in DQ is probably limited. In practice, developmental tests always show minor variations in test results. Several patients may have a stable DQ, yet the chance of scoring the same DQ in repeated measurements is small, so patients may randomly show a slightly higher or lower DQ at different time points. Further, a somewhat higher DQ may indicate a practice effect, since these tests were only 22 weeks apart, and children (particularly those with higher age equivalents) may have remembered some of the tasks or specific solutions. So, for
Fig. 8. Spaghetti plot of gray matter volume by chronological age (n = 12). Each patient's rhHNS dose group is indicated.
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the 6 patients whose DQs showed small, similar magnitude, increases or decreases, they should probably be considered to have stable DQ. In conclusion, intrathecal administration of rhHNS was found to be generally safe and well tolerated, with no SAEs related to the drug itself. The high rate of complications with the IDDD in this patient population indicates that an improved administration system is needed for future studies. The administration of rhHNS resulted in consistent declines in CSF heparan sulfate levels, with trends suggesting dose responsiveness consistent with biological activity in the relevant anatomical compartment, while the decline in uGAG levels indicated systemic activity of the experimental therapy. There was no apparent effect of antibody development on these responses, and no safety signals were associated with antibody positivity. The lack of a clear clinical outcome signifying a positive effect of therapy in this relatively short trial is not unexpected, as the study was designed to provide an initial assessment of safety and tolerability. Patients who participated in this trial were offered the opportunity to participate in an extension trial (HGT-SAN-067, ClinicalTrials.gov identifier: NCT01299727) in which they may continue to receive treatment with rhHNS. This will permit a longer term assessment of the safety and potential impact on clinical parameters. Potential conflicts of interest S.A.J. has received honoraria, travel grants, or research grants from Shire. C.B. has received a travel grant from Shire. F.H. has received travel grants from Shire. S.R. has received a travel grant from Shire. J.d.R., E.T., and J.P.M. have nothing to disclose. L.P., Y.Q., and J-K.C. are employees of, and own stock in, Shire. N.N. and A.J.B. own stock in Shire and at the time of the study were employees of Shire. P.A.J.H. at the time of the study was an employee of Shire. F.A.W. has received honoraria, travel grants, or research grants from Shire. Funding The phase 1/2 study (HGT-SAN-055) was funded by Shire. Editorial assistance to the authors was provided by Jillian Lokere and Robin Smith of The Curry Rockefeller Group, LLC, Tarrytown, NY, and was funded by Shire. Acknowledgment The authors thank the National Institute for Health Research/ Wellcome Trust Manchester Clinical Research Facility.
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Contents lists available at ScienceDirect
Molecular Genetics and Metabolism journal homepage: www.elsevier.com/locate/ymgme
A phase 1/2 study of intrathecal heparan-N-sulfatase in patients with mucopolysaccharidosis IIIA Simon A. Jones a, Catherine Breen a, Fiona Heap a, Stewart Rust b, Jessica de Ruijter c, Evelien Tump c, Jan Pieter Marchal c, Luying Pan d, Yongchang Qiu d, Jou-Ku Chung d, Nitin Nair d,1, Patrick A.J. Haslett d,2, Ann J. Barbier d,3, Frits A. Wijburg c,⁎ a
Willink Unit, Manchester Centre for Genomic Medicine, St Mary's Hospital, Central Manchester University Hospitals NHS Foundation Trust (CMFT), University of Manchester, United Kingdom Paediatric Psychosocial Department, Royal Manchester Children's Hospital, Manchester, United Kingdom c Department of Paediatrics, Academic Medical Center, Amsterdam, The Netherlands d Shire, Lexington, MA, USA b
a r t i c l e
i n f o
Article history: Received 9 May 2016 Accepted 9 May 2016 Available online 10 May 2016 Keywords: Enzyme replacement therapy Heparan sulfate Intrathecal drug delivery device Lysosomal storage disease Mucopolysaccharidosis IIIA (MPS IIIA) Sanfilippo syndrome A
a b s t r a c t Objective: This was an open-label, phase 1/2 dose-escalation, safety trial of intrathecal recombinant human heparan-N-sulfatase (rhHNS) administered via intrathecal drug delivery device (IDDD) for treating mucopolysaccharidosis IIIA (NCT01155778). Study design: Twelve patients received 10, 45, or 90 mg of rhHNS via IDDD once monthly for a total of 6 doses. Primary endpoints included adverse events (AEs) and anti-rhHNS antibodies. Secondary endpoints included standardized neurocognitive assessments, cortical gray matter volume, and pharmacokinetic/pharmacodynamic analyses. Results: All patients experienced treatment-emergent AEs; most of mild-to-moderate severity. Seven patients reported a total of 10 serious AEs (SAEs), all but one due to hospitalization to revise a nonfunctioning IDDD. No SAEs were considered related to rhHNS. Anti-rhHNS antibodies were detected in the serum of 6 patients and in the cerebrospinal fluid (CSF) of 2 of these. CSF heparan sulfate levels were elevated at baseline and there were sustained declines in all tested patients following the first rhHNS dose. No impact of anti-rhHNS antibodies on any pharmacodynamic or safety parameters was evident. 4 of 12 patients showed a decline in developmental quotient, 6 were stable, and 2 patients had only a single data point. No dose group showed a clearly different response pattern. Conclusions: rhHNS administration via IDDD appeared generally safe and well tolerated. Treatment resulted in consistent declines in CSF heparan sulfate, suggesting in vivo activity in the relevant anatomical compartment. Results of this small study should be interpreted with caution. Future studies are required to assess the potential clinical benefits of rhHNS and to test improved IDDD models. © 2016 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
1. Introduction Abbreviations: AE, adverse event; BSID-III, Bayley Scales of Infant and Toddler Development, Third Edition; CSF, cerebrospinal fluid; CNS, central nervous system; DQ, developmental quotient; EC, Enzyme Commission; ELISA, enzyme-linked immunosorbent assay; GAG, glycosaminoglycan; IDDD, intrathecal drug delivery device; IT, intrathecal; KABC-II, Kaufman Assessment Battery for Children, Second Edition; LC-MS/MS, liquid chromatography-tandem mass spectrometry; MPS IIIA, mucopolysaccharidosis IIIA/ Sanfilippo syndrome type A; MRI, magnetic resonance imaging; rhHNS, recombinant human heparan-N-sulfatase (EC 3.10.1.1); SAE, serious adverse event; TEAE, treatmentemergent adverse event; Tmax, time to maximum serum concentration; uGAG, urinary glycosaminoglycan; VABS-II, Vineland Adaptive Behavior Scales, Second Edition. ⁎ Corresponding author at: Department of Paediatrics (H7-270), Academic Medical Center, Meibergdreef 9, 1105 AZ Amsterdam, The Netherlands. E-mail address: [email protected] (F.A. Wijburg). 1 Current address: Vertex Pharmaceuticals, Inc., 50 Northern Ave, Boston, MA 02210. 2 Current address: Alnylam Pharmaceuticals, Inc., 300 Third Street, Cambridge, MA 02142. 3 Current address: Agios Pharmaceuticals, 88 Sidney Street, Cambridge, MA 02139.
Mucopolysaccharidosis IIIA (MPS IIIA, Sanfilippo syndrome type A; OMIM 252900) is a lysosomal storage disorder caused by a deficiency of the enzyme heparan-N-sulfatase (EC 3.10.1.1), leading to accumulation of the glycosaminoglycan (GAG), heparan sulfate, in the lysosomes [1]. The clinical manifestations of MPS IIIA are primarily neurologic, with rapid and progressive disease manifestation, which are in contrast to the relatively mild somatic symptoms compared with other types of MPS disorders [1–3]. Neurologic signs include developmental delay, behavioral abnormalities, and sleep disturbances [2,3]. Life span is usually severely shortened [3]. There are no approved disease-modifying therapies available for MPS IIIA. Preclinical safety of recombinant human heparan-N-sulfatase (rhHNS) formulated for intrathecal (IT) delivery, has been demonstrated in toxicology studies performed in juvenile cynomolgus monkeys,
http://dx.doi.org/10.1016/j.ymgme.2016.05.006 1096-7192/© 2016 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
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where the drug was administered via a surgically implanted intrathecal drug delivery device (IDDD) [4]. Here we report the results from the first-in-human, phase 1/2 clinical trial (ClinicalTrials.gov identifier: NCT01155778) of rhHNS. The primary objective of the study was to determine the safety and tolerability of rhHNS in patients with MPS IIIA via ascending doses administered intrathecally through a surgically implanted IDDD once monthly for 6 months. 2. Patients and methods 2.1. Patients Patients were enrolled from the Academic Medical Center in Amsterdam, the Netherlands, and St Mary's Hospital in Manchester, United Kingdom. Included patients were required to have heparan-N-sulfatase enzyme activity ≤10% of the lower limit of normal as measured in fibroblasts or leukocytes, plus either (1) a normal enzyme activity level of at least one other sulfatase (to rule out multiple sulfatase deficiency) as measured in fibroblasts or leukocytes, or (2) two documented mutations in the SGSH gene. Included patients were ≥3 years of age, with a developmental age ≥ 1 year, based on the results of a developmental questionnaire at the time of screening (see Endpoints and assessments). The exclusion criteria were significant central nervous system (CNS) impairment or behavioral disturbances with causes other than MPS IIIA, previous implantation of a CNS shunt, poorly controlled seizure disorder, prior hematopoietic stem cell or bone marrow transplantation, and treatment with any investigational drug or a device intended as a treatment for MPS IIIA within the 30 days prior to the study. The study was conducted in compliance with the Declaration of Helsinki, the relevant Institutional Review Board regulations, and the International Conference on Harmonisation Good Clinical Practice guidelines. Written informed consent was obtained from patients or their parents/legal guardians. 2.2. Study design This was a phase 1/2 multicenter, open-label, multiple-dose, doseescalation study. Patients were enrolled sequentially into each of 3 treatment arms: 10, 45, or 90 mg rhHNS via IDDD dosed every 4 weeks (28 days ± 7 days). A total of 6 doses were planned. Enrollment was staggered so that initial safety could be assessed on the first 2 patients within each dose group. Moreover, dose escalations were only allowed after a review of safety data from 4 patients at the preceding dose level by an independent monitoring board. The study had no control arm. All patients were implanted with the IDDD (PORT-A-CATH® II Low Profile™; Smiths Medical ASD, Inc., St. Paul, Minnesota, USA) on week 1, day 1, followed by a recovery period of at least 7 days. Patients were admitted to the study center 24 h prior to their first rhHNS dose for safety and baseline assessments and were not discharged until approximately 24 h after the injection. Patients continued to receive safety assessments before and following each subsequent dose of rhHNS. In the event of IDDD malfunction, the dose of rhHNS could be administered via lumbar puncture. 2.3. Endpoints and assessments The primary objective was to assess the safety and tolerability of intrathecal administration of rhHNS in patients with MPS IIIA. Primary endpoints included observations of adverse events (AEs) and changes in clinical laboratory values, including cerebrospinal fluid (CSF) cell counts and chemistries, electrocardiograms, and anti-rhHNS antibodies in the CSF and serum. The secondary objectives of the study were to determine by dose group the effects of intrathecal administration of rhHNS on standardized neurocognitive assessments, cortical gray
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matter volume as assessed by magnetic resonance imaging (MRI), pharmacokinetic parameters, and the pharmacodynamic responses. The primary pharmacodynamic variable was the level of heparan sulfate in the CSF. The level of total heparan sulfate in the CSF was determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Briefly, heparan sulfate in the CSF was first extracted using an anion-exchange resin and then digested by a combination of enzymes including heparinase I, II, and III. The resultant heparan sulfate disaccharides were labeled with 12C-4-N-butylaniline by reductive amination and then analyzed by LC-MS/MS. The disaccharides were quantified based on a calibration curve generated using 6 commercially available disaccharide standards that are the most abundant in human CSF heparan sulfate. The secondary pharmacodynamic measurement was urinary glycosaminoglycan (uGAG) concentration, determined by a 1,9dimethylmethylene blue dye-binding assay using the Blyscan™ GAG assay kit (Biocolor Ltd., Carrickfergus, Northern Ireland, United Kingdom), and was reported relative to creatinine concentration (mg GAG/ mmol creatinine) [5]. Serum samples for pharmacokinetic testing were collected at the time of the first and last dose of rhHNS. Samples were collected immediately prior to the intrathecal injection and then at 0.5, 1, 1.5, 2, 3, 4, 6, 8, 12, 24, 48, and 72 h following completion of the intrathecal injection. The rhHNS concentrations were measured using an enzyme-linked immunosorbent assay (ELISA) with paired anti-rhHNS antibodies. Testing for serum and CSF antibodies against rhHNS was conducted at week 2 and at weeks 6, 10, 14, 18, 22, and 26. Antibody assessment was performed via a tiered approach by a bridging electrochemiluminescent immunoassay using paired biotin- and sulfo-tagged rhHNS. This assay detects total antidrug antibodies and does not distinguish immunoglobulin isotypes. Samples that screened positive for anti-rhHNS antibodies were confirmed by a ligand-competition assay, and then further tested for antibody titer with the same assay platform. Antibodies were not tested for their ability to neutralize rhHNS. CSF samples were collected from patients through the implanted IDDD or, in the event of a nonfunctioning IDDD, via lumbar puncture immediately prior to administration of rhHNS. Urine samples were collected at weeks 2, 6, 10, 14, 18, 22, and 26. To determine study eligibility, the patient's level of functioning was estimated at screening using the Vineland Adaptive Behavior Scales, Second Edition (VABS-II) [6]. The VABS-II is a caregiver-reported outcome that measures adaptive behaviors, including the ability to cope with environmental changes, to learn new everyday skills, and to demonstrate independence. The test measures various key domains (communication, daily living skills, socialization, and motor skills) and provides an adaptive behavior composite (a composite of the other 4 domains). Formal neurodevelopmental testing of enrolled patients was performed at weeks 2 and 22 using either the Bayley Scales of Infant and Toddler Development, Third Edition (BSID-III) [7] or the Kaufman Assessment Battery for Children, Second Edition (KABC-II) [8]. The BSID-III is designed for the neurodevelopmental assessment of children 0 to 42 months of age, whereas the KABC-II is designed to assess children from 3 to 18 years old. All children who were ≤ 42 months of age at enrollment, or who at screening had an average developmental age b36 months across all domains of the VABS-II, were assessed using the BSID-III. Children aged N42 months and with a developmental age N36 months were administered the KABC-II, unless they were unable to cooperate, in which case the BSID-III was administered. The test used at baseline was also used at week 22. Both tests served to ascertain the mental age equivalent in months of each child, using data obtained from the cognitive domain of the BSID-III and the mean age equivalent of the nonverbal domains of the KABC-II. The developmental quotient (DQ) was calculated by dividing the mental age equivalent by the calendar age in months, multiplied by 100, as described by Delaney et al. [9]. Cortical gray matter volumes were obtained by automated volumetric analysis of MRI scan data, using FreeSurfer software (Athinoula A. Martinos Center for Biomedical Imaging, Charlestown, MA). Total
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Table 1 Patient demographics at screening. rhHNS dose group, mg
10
45
90
Patient no.
Sex
Age, year
BMI
1 2 3 4 5 6 7 8 9 10 11 12
F M M M M F M F M M M F
13 5 13 5 3 4 6 24 4 4 12 22
22.9 20.4 22.8 17.8 17.5a 19.5 15.0 24.2 20.1 18.4 18.7 23.4
F, female; M, male; rhHNS, recombinant human heparan-N-sulfatase. a Patient was noncompliant at screening; these measurements were taken at week 1, day 7.
brain cortical gray matter volume was determined by automated analysis of brain MRIs performed at weeks 2 and 22, and results were read centrally. All calculations were performed using SAS v9.1 or later (SAS Institute Inc., Cary, NC). 3. Results 3.1. Study population Twelve patients were enrolled (4 females) with a median age of 5.5 years (range, = 3–24 years; Table 1). Four patients (2 pairs of siblings) had a relatively attenuated disease phenotype by clinical impression at baseline: patients 8 and 12, and patients 3 and 11. A fifth patient (patient 7) had an intermediate clinical phenotype at baseline. The remaining 7 patients had a severe phenotype. All the patients' SGSH mutations were compound heterozygous and varied across the 3 dose groups. In the 10 mg IT dose group one patient had a missense/unclassifiable (SGSH allele 1/allele 2) mutation, one had a missense/missense, and 2 had missense/frameshift mutations. In the 45-mg IT dose group 2 patients had a missense/unclassifiable mutation, one had a nonsense/nonsense, and one had a missense/missense mutation. In the 90-mg IT dose group one patient had an unclassifiable/missense mutation, one had a frameshift/frameshift, and 2 had missense/ missense mutations. 3.2. Safety results Treatment with rhHNS appeared generally safe and well tolerated. No clinically meaningful trends were observed in laboratory values, vital signs, physical examinations, or electrocardiographic assessments. There were no clinical signs or symptoms of meningeal irritation following enzyme infusion, and no change in the predose CSF clinical laboratory parameters to suggest meningeal inflammation.
All patients experienced at least 1 treatment-emergent AE (TEAE). The majority of these were mild to moderate in severity, and all resolved. There was no evidence of a higher frequency of TEAEs with higher doses of rhHNS. Four patients (33%) reported rhHNS-related AEs. These included fatigue, pain, pyrexia, cognitive disorder, memory impairment, motor dysfunction, malaise, pallor, and transient symptoms suggesting saddle paresthesia. Six patients (50%) reported IDDDrelated AEs. These included device failure, device breakage, and device component issues; device change; postoperative wound infections; and urinary incontinence. Three patients (25%) reported AEs that were related to the intrathecal administration process, including malaise; nausea and vomiting; and transient symptoms suggesting saddle paresthesia. Two patients (17%) reported AEs that began within 24 h after the start of injection and were judged possibly/probably related to rhHNS: 1 patient experienced irritability, vomiting, and increased CSF protein and white blood cell count; and 1 patient experienced pyrexia. Ten patients (83%) reported surgery-related AEs. Seven (58%) patients reported a total of 10 serious AEs (SAEs). This included 3 patients in the 10-mg dose group who reported 5 SAEs, 3 patients in the 45-mg dose group who reported 3 SAEs, and 1 patient in the 90-mg dose group who reported 2 SAEs. No SAEs were considered to be related to rhHNS; all were designated SAEs because of hospitalization, and 9 of 10 SAEs were related to the IDDD. The exception was a hospitalization for a suspected intercurrent viral infection unrelated to the study drug or procedures. A summary of IDDD failures is presented in Table 2, revealing that all failures were mechanical in nature owing to disconnection, breakage, or dislocation of the device from the intrathecal space. Patient 12 experienced 2 IDDD failures and received the remaining rhHNS doses via lumbar puncture. No patients discontinued from the study, and no patients died during the study. In total, 9 of 12 patients (75%) received all 6 of their scheduled doses, and 3 of 12 patients (25%) received 5 of their 6 scheduled doses. 3.3. Immunogenicity Serum antibodies to rhHNS were detected in 6 of 12 (50%) patients (Fig. 1). The response did not appear to be dose-dependent. Two patients (16.7%) were positive for low-titer antibodies in serum at baseline and demonstrated an increase in titer over the course of the study. In the other 4 patients, seroconversion occurred within the first 10 weeks of the study. One patient with low-serum antibody titer reverted to antibody-negative status at the week 26 visit. Low-titer antibodies in the CSF were detected for 1 patient in the 10-mg dose group beginning at week 18 and in 1 patient in the 90-mg dose group at week 22. Both patients had high titers of serum antibodies. One of the patients with pre-existing antibodies who also developed the highest titer of serum rhHNS antibodies and was positive for CSF antibodies had frame-shift mutations for both alleles. The other patient with pre-existing antibodies, and who developed a moderate serum antibody titer, was negative for CSF antibodies and had nonsense mutations for both alleles.
Table 2 Summary of IDDD failures. Patient No.
No. of doses via IDDD before First Failure Cause of first failure
No. of doses via IDDD before second failure
Cause of second failure
1 2 3
0 1 3
N/A N/A N/A
– – –
7
1
N/A
–
8 12
4 0
N/A 1
– Catheter migrated out of spinal canal
IDDD, intrathecal drug delivery device; N/A, not available.
Port/catheter disconnection Port outlet pinbreak Catheter migrated out of spinal canal Catheter migrated out of spinal canal Port outlet pinbreak Catheter migrated out of spinal canal
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patients tested (3/4) showed a decline in CSF levels that ranged from 58% to 75%. Mean heparan sulfate levels in CSF for the different dose groups over time are shown in Fig. 4. There was no apparent effect of antibody status on CSF levels of heparan sulfate (data not shown). At baseline, mean uGAG levels were elevated for all 3 study groups: 10-mg dose-group, 48.8 mg GAG/mmol creatinine (range, 38.2–63.9 mg GAG/mmol creatinine) (n = 4); 45-mg dose-group, 41.1 mg GAG/mmol creatinine (range, 13.3–74.8 mg GAG/mmol creatinine) (n = 4); 90-mg dose-group, 45.1 mg GAG/mmol creatinine (range, 23.7–70.0 mg GAG/mmol creatinine) (n = 4). By week 22, persistent uGAG reduction was observed in 3/4 patients in the 10-mg dosegroup, 3/4 patients in the 45-mg dose-group, and all patients (4/4) in the 90-mg dose-group. The decline was generally evident in urine collected 1 month later (week 6) after the first dose of intrathecal administration of rhHNS in 9/12 patients (Figs. 5 and 6). Normal reference range for uGAG levels are b10.5 mg GAG/mmol creatinine in children aged 4–5 years, and b 5.5 mg GAG/mmol creatinine in those aged N13 years [10]. Overall, there was no clear dose–response relationship for uGAG levels. Fig. 1. Semilogarithmic plot of serum anti-rhHNS antibody titers over time in the 6 patients exhibiting seropositivity. One patient developed very low titer serum antibodies since week 10, and reverted to serum antibody negative at week 26; two patients were positive for anti-rhHNS antibodies at baseline. For plotting purposes, an artificial titer of 1 was assigned to samples with negative antidrug results. rhHNS, recombinant human heparan-N-sulfatase.
3.4. Serum rhHNS pharmacokinetics At week 2 of intrathecal administration, rhHNS exhibited biphasic serum concentration-time profiles across the 10-, 45-, and 90-mg dose groups. Following the first intrathecal dose, the time to maximum serum concentration (Tmax) occurred between approximately 3 and 6 h, indicating a gradual transfer of rhHNS from the CNS to systemic compartments (Fig. 2A and B). Systemic exposure of rhHNS was doseproportional following the first dose of rhHNS (week 2; Fig. 2A), but not following the sixth dose (week 22; Fig. 2B). A high degree of variability was observed across pharmacokinetic parameters, which may have been due to the small number of patients within each dose group.
3.5. Pharmacodynamic response: levels of CSF heparan sulfate and uGAGs To establish a reference range for levels of heparan sulfate in the CSF, we analyzed 156 CSF samples from children without MPS from whom CSF had been obtained for other clinical reasons. These analyses indicated CSF levels of heparan sulfate ranged from below the limit of quantification (0.300 μM) to 0.501 μM (data not shown). In the current study, CSF levels of heparan sulfate were elevated at baseline for all 3 treatment groups relative to the non-MPS pediatric controls. Following intrathecal administration of rhHNS, CSF levels of heparan sulfate exhibited marked declines in all tested patients (11/12) from the 3 treatment groups. The CSF samples from patient 10 were not collected at weeks 6, 10, 14, and 18 for bioanalysis, and the week 22 CSF sample volume was insufficient to allow for HS testing. Four other patients (4/12) did not have sufficient sample CSF volume for HS quantification at week 6. Comparing the individual patient CSF heparan sulfate level data (Fig. 3), all patients tested (11/12) showed declines in their CSF heparan sulfate levels compared to their baseline (2 weeks) levels by week 22 or the end of the study, and a reduction in their CSF heparan sulfate level after the first IT dose was observed in all patients (7/12) who had week 6 samples available for testing. Those in the 10-mg dose group showed the least decline (11–52%) compared to baseline, whereas the 45-mg and 90-mg patients showed greater declines versus baseline (50–78%). In the 90-mg dose group all
Fig. 2. Mean serum rhHNS concentration-time profiles of patient samples following intrathecal administration of rhHNS by dose group at (A) week 2 and (B) week 22. Error bars are standard error of mean. rhHNS, recombinant human heparan-N-sulfatase.
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Fig. 4. Mean heparan sulfate levels in CSF over time by rhHNS dose group. CSF, cerebrospinal fluid. Error bars are standard error of mean.
each from the 10- and 45-mg dose groups and 2 patients from the 90mg dose group underwent the BSID-III. Two patients in the 10-mg dose group did not have baseline cognitive assessment data due to their inability to cooperate, but in these cases, cognitive data were successfully obtained by the BSID-III at week 22. Of the 12 patients, 4 showed a decline in DQ, 6 were essentially stable, showing a small DQ increase or decrease of similar magnitude that is not clinically significant, and 2 patients had only a single data point. No dose group showed a clearly different pattern of response (Fig. 7). 3.7. Cortical gray matter volume Total brain cortical gray matter volume was determined by automated analysis of brain MRIs performed at weeks 2 and 22. Declines in total cortical gray matter volume were observed in all dose groups and in most patients between weeks 2 and 22 (Fig. 8). However, 2 patients, siblings with a highly attenuated phenotype and relatively preserved cognitive function, did not show reductions in this parameter. 4. Discussion
Fig. 3. Heparan sulfate levels in CSF for individual patients. Week 2 levels were used as baseline. CSF, cerebrospinal fluid.
3.6. Neurodevelopmental analyses Cognitive status was assessed at screening and at week 22. Only one site performed the VABS-II assessment at both screening and at week 22; thus, 6 patients have VABS-II data from 2 time points (Table 3). Cognitive status was also assessed at screening and at week 22 by either the BSID-III or KABC-II (see Patients and methods). Three patients
The primary objective of the trial was to assess the safety and tolerability of rhHNS administered via IDDD in children with MPS IIIA. There was no significant clinical safety signal associated with the drug itself, which appeared to be generally well tolerated. All but one of the SAEs, which occurred in more than half of the patients, were related to malfunctions of the IDDD. None were related to the study drug. No patients died during the study, and none discontinued. IDDD failures occurred at an unexpectedly high rate, and were due to mechanical problems in the form of disconnection or breakage of the device or anatomical displacement of the catheter with migration out of the intrathecal space. Failures of similar devices have been reported in the literature, primarily due to mechanical malfunction [11,12]. These have been reported as occurring in up to 24% of patients, with disconnection of the catheter from the pump/port and migration of the catheter from the spinal canal accounting for most cases [13]. We found a similar pattern of failure in this study, but the overall frequency was higher than those reported in other clinical settings. It is possible that the hyperactivity and restlessness that frequently complicate progressive MPS IIIA may have increased the rate of IDDD complications. A next-generation IDDD has been developed to address these issues for the extension of the study and potential future studies
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Fig. 6. Normalized mean uGAG levels over time by rhHNS dose group. uGAG, urinary glycosaminoglycan.
Fig. 5. Normalized uGAG levels over time in individual patients. Week 2 levels were used as baseline. uGAG, urinary glycosaminoglycan.
(NCT01299727, 02060526). Another consideration is that rhHNS could have leaked out into surrounding tissues due to catheter migration or disconnection and this leakage could potentially impact uGAG and serum anti-rhHNS antibody levels, We think that, if this did occur, then it would not have had significant effect as the serum pharmacokinetic data (Fig. 2) show that there is a considerable amount of enzyme released into the systemic circulation after IT delivery in patients who were not experiencing device failure. This systematic release is likely
to be the reason for the effect of IT administration on uGAG levels, and this far outweighs any contribution from potential leakage. The question of whether enzyme delivered into the subcutaneous tissue could increase anti-drug antibodies is an interesting one, for which we currently have no data available for evaluation. Systemic exposure of rhHNS was dose-proportional following the first dose of rhHNS, although the cause of disturbed dose proportionality following the sixth dose is unknown (Fig. 2B). There are a number of possible reasons for this. Anti-rhHNS antibodies could have interfered with the ELISA method used for drug quantification. Other possibilities are that a greater or lesser amount of the drug was administered due to IDDD malfunction, or the presence of antibodies alters the distribution and clearance of rhHNS. A combination of some or all of these factors could produce the observed effect. Intrathecal administration of rhHNS proved to be immunogenic, with half the patients exhibiting either de novo anti-rhHNS antibody formation (n = 4) or increases in anti-rhHNS titers from baseline levels (n = 2). The reason for anti-rhHNS reactivity at baseline is unknown; however, during assay development, anti-rhHNS antibodies were found in 2 of 25 untreated patients with MPS IIIA, but none were found in 54 normal healthy donors, suggesting that baseline serum cross-reactivity might be a feature of MPS IIIA disease (data not shown). In 5 of 6 patients with anti-rhHNS antibodies, including those with baseline antibodies, there was a clear increase in titer over time. However, 1 patient displayed only very low titers from weeks 10 to 22, and reverted to seronegativity at the end of study visit. There were 2 patients with transient, extremely low-titer anti-rhHNS antibodies in the CSF that were associated with high-titer serum antibodies, suggesting possible diffusion of serum antibodies into the CSF. Of note, immunogenicity of therapeutic enzymes present in the systemic circulation has been reported in other MPS diseases [14–16]. Fifty percent of patients (6/12) developed anti-rhHNS antibodies after receiving ERT including 2 who had pre-existing serum antirhHNS antibodies with low titers. One of these patients with preexisting antibodies was positive for CSF antibodies and had frameshift mutations for both alleles and the other, who was negative for CSF antibodies, had nonsense mutations for both alleles. In a recent natural history study of MPS IIIA patients, 2 patients (2/25) were positive for anti-rhHNS antibodies with low titers and both of these patients had missense mutations for both alleles [17]. It has been shown in MPS II patients receiving idursulfase ERT that higher antibody titers are correlated to patients with complete deletion/large rearrangement and frameshift/splice site mutations when compared to those with
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Table 3 Adaptive behavior composite scoresa as assessed by the VABS-II at screening (n = 12) and at week 22 (n = 6). Patient no.
Sex
rhHNS dose group, mg
Visit
Age at visit
Composite score
Age equivalent
1
F
10
2
M
10
3 4 5
M M M
10 10 45
6
F
45
7 8 9
M F M
45 45 90
10
M
90
11 12
M F
90 90
Screening week 22 Screening week 22 Screening Screening Screening week 22 Screening week 22 Screening Screening Screening week 22 Screening week 22 Screening Screening
13 years, 2 months 13 years, 7 months 5 years, 5 months 5 years, 10 months 13 years, 2 months 4 years, 9 months 3 years, 1 month 3 years, 7 months 3 years, 6 months 3 years, 11 months 6 years, 0 month 23 years, 5 months 4 years, 2 months 4 years, 9 months 3 years, 11 months 4 years, 5 months 11 years, 9 months 22 years, 4 months
50 45 59 46 83 63 76 63 80 69 62 20 56 56 72 61 90 20
2 years, 11 months 2 years, 5 months 2 years, 0 month 1 year, 1 month 10 years, 5 months 2 years, 2 months 2 years, 1 month 1 year, 6 months 2 years, 6 months 2 years, 1 month 2 years, 7 months 3 years, 2 months 1 year, 6 months 1 year, 8 months 2 years, 3 months 1 year, 10 months 10 years, 7 months 0 year, 3 months
Week 22 testing was only performed at one study site. F, female; M, male; rhHNS, recombinant human heparan-N-sulfatase; VABS-II, Vineland Adaptive Behavior Scales, Second Edition. a Composite adaptive behavior score is a composite of the 4 constituent domains of the VABS-II (normal = 100).
missense mutations [18]. Given the small number of patients in this study, no firm conclusions can be reached about any relation between genotype and immunogenicity. There were no clear safety signals associated with the presence of anti-rhHNS antibodies in the study. The clinical significance of the development of antidrug antibodies in this patient population is unclear. Serum pharmacokinetic analyses showed dose-proportional patterns in peripheral blood following the first intrathecal dose of rhHNS. Linear dose proportionality in serum pharmacokinetics was not observed at week 22 following the sixth dose, and this may have been related to the development of serum anti-rhHNS antibodies, and in one case, to IDDD failure. The primary pharmacodynamic parameter of biochemical efficacy, total heparan sulfate in the CSF, exhibited declines in response to therapy at all dose levels, with a greater impact observed in the higher dose groups. Remarkably, intrathecal administration of rhHNS also reduced uGAG concentrations at all dose levels, suggesting systemic activity of the drug despite the extremely low blood levels indicated by the pharmacokinetic analysis. Over the duration of this shortterm study, no impact of anti-rhHNS antibodies on any pharmacodynamic parameters was evident.
Fig. 7. Spaghetti plot of individual BSID-III/KABC-II DQ scores by chronological age (n = 12). Each patient's rhHNS dose group is indicated. Note: DQ was calculated from the mean developmental age. BSID-III, Bayley Scales of Infant and Toddler Development, Third Edition; DQ, developmental quotient; KABC-II, Kaufman Assessment Battery for Children, Second Edition.
Although the majority of patients exhibited declines in DQ during the study, the study was neither powered nor designed to adequately assess clinical efficacy. The wide range of patient ages, disease phenotypes, and disease stages renders interpretation of data very challenging. Additionally, the 6-month duration of this study is likely to be too short to reasonably expect a meaningful change in these parameters in response to therapy. Interpretation was further limited by the absence of week 2 cognitive data in 2 patients in the 10-mg dose group who were unable to cooperate with the assessment. In all but 2 patients, cortical gray matter volume, a potential biomarker of MPS IIIA brain pathology, showed declines between weeks 2 and 22. The exception was a sibling pair with particularly attenuated disease and unusually wellpreserved cognition. The clinical significance of small increases or decreases in DQ is probably limited. In practice, developmental tests always show minor variations in test results. Several patients may have a stable DQ, yet the chance of scoring the same DQ in repeated measurements is small, so patients may randomly show a slightly higher or lower DQ at different time points. Further, a somewhat higher DQ may indicate a practice effect, since these tests were only 22 weeks apart, and children (particularly those with higher age equivalents) may have remembered some of the tasks or specific solutions. So, for
Fig. 8. Spaghetti plot of gray matter volume by chronological age (n = 12). Each patient's rhHNS dose group is indicated.
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the 6 patients whose DQs showed small, similar magnitude, increases or decreases, they should probably be considered to have stable DQ. In conclusion, intrathecal administration of rhHNS was found to be generally safe and well tolerated, with no SAEs related to the drug itself. The high rate of complications with the IDDD in this patient population indicates that an improved administration system is needed for future studies. The administration of rhHNS resulted in consistent declines in CSF heparan sulfate levels, with trends suggesting dose responsiveness consistent with biological activity in the relevant anatomical compartment, while the decline in uGAG levels indicated systemic activity of the experimental therapy. There was no apparent effect of antibody development on these responses, and no safety signals were associated with antibody positivity. The lack of a clear clinical outcome signifying a positive effect of therapy in this relatively short trial is not unexpected, as the study was designed to provide an initial assessment of safety and tolerability. Patients who participated in this trial were offered the opportunity to participate in an extension trial (HGT-SAN-067, ClinicalTrials.gov identifier: NCT01299727) in which they may continue to receive treatment with rhHNS. This will permit a longer term assessment of the safety and potential impact on clinical parameters. Potential conflicts of interest S.A.J. has received honoraria, travel grants, or research grants from Shire. C.B. has received a travel grant from Shire. F.H. has received travel grants from Shire. S.R. has received a travel grant from Shire. J.d.R., E.T., and J.P.M. have nothing to disclose. L.P., Y.Q., and J-K.C. are employees of, and own stock in, Shire. N.N. and A.J.B. own stock in Shire and at the time of the study were employees of Shire. P.A.J.H. at the time of the study was an employee of Shire. F.A.W. has received honoraria, travel grants, or research grants from Shire. Funding The phase 1/2 study (HGT-SAN-055) was funded by Shire. Editorial assistance to the authors was provided by Jillian Lokere and Robin Smith of The Curry Rockefeller Group, LLC, Tarrytown, NY, and was funded by Shire. Acknowledgment The authors thank the National Institute for Health Research/ Wellcome Trust Manchester Clinical Research Facility.
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