- Email: [email protected]
6. Post-surgical deaths among MPS I registry patients
Abstracts/Molecular Genetics and Metabolism 99 (2010) S8–S41 Discussion: Each MPS disorder appears to have its own profile of quantitative MRI and neuropsychological test results. Grey matter brain volumes are consistently larger in MPS II than in MPS I or controls, even controlling for gender. Although IQ and memory appear to be normal in MPS II, attention and information processing appear to be a deficit area. Cross sectional data is subject to confounding effects such as ascertainment/disease severity in MPS II. Only a longitudinal study such as that in the Lysosomal Disease Network will provide definitive results.
S9
ance of respiratory-related deaths point to the risk of surgery itself in this patient population. doi:10.1016/j.ymgme.2009.10.023
7. A urinary glycosaminoglycan analysis by tandem mass spectrometry doi:10.1016/j.ymgme.2009.10.021
5. Reliability of manual and automated tracing of hippocampal volumes in MPS patients and normal controls: A report of the neuroimaging core of the lysosomal disease network Alia Ahmed, Igor Nestrasil, Kyle Rudser, Elsa Shapiro, University of Minnesota, Minneapolis, MN, USA Background: Quantitative methods in neuroimaging such as diffusion tensor imaging (DTI) and volumetric analysis (VA) hold great promise to examine brain changes with disease and treatment in longitudinal studies of lysosomal diseases. Clinical scoring systems correlate with major functional abnormality but are not sensitive enough to examine treatment changes over time. Goals: Determine the reliability and validity of manual tracings and automated tracings of the hippocampus; automated systems may not provide accurate volumes. This work is necessary to set the standards for the Neuroimaging Core of the Lysosomal Disease Network to accurately quantify change in multiple studies. Methods: Both automated and manual tracings were obtained on 20 MPS I, 1 MPS II, and 2 MPS VII patients and 9 control subjects. Three-dimensional T 1 weighted gradient echo sequence (fast low angle shot – FLASH) was performed on the whole brain with a slice thickness of 1.0 mm. Automated volumes were calculated using FreeSurfer (run on the University of Minnesota Supercomputer) to determine feasibility in patients with abnormal brains. Brains2 manual tracings of the hippocampus were accomplished by two experienced raters. To obtain interrater and intrarater reliability, both raters (A.A. and I.N.) traced the same images and also traced 6 randomly chosen images twice. Results: Intrarater reliability determined by intraclass correlation was excellent at 0.99 indicating consistency within rater. Across two experienced raters, overall reliability for left and right, with and without white matter, was 0.93 (95% CI: 0.90, 0.95) with the lowest observed at 0.81. Comparing the Brains2 with FreeSurfer indicated that FreeSurfer overestimated hippocampal volumes by 60%. For example, while mean left hippocampus volume including white matter was 2.6 and 2.7 for the two raters, mean automated volumes were 4.2 cm3. Discussion: Brains2 manual tracing of hippocampal size was found to be reliable comparing within and across raters. FreeSurfer volumes were larger and probably overinclusive providing an overestimation of hippocampal plus hippocampus-adjacent white matter size. Other data indicates that the volumes of other structures are accurately determined by FreeSurfer but for the hippocampus, accurate manual tracings will be necessary. doi:10.1016/j.ymgme.2009.10.022
6. Post-surgical deaths among MPS I registry patients Pamela Arna,b,c,d, H. Warner Webba, Gerald Coxb, aThe Nemours Childrens Clinic, Jacksonville, FL, USA, bGenzyme Corporation, Cambridge, MA, USA, cDivision of Genetics, Childrens Hospital, Boston, USA, dDepartment of Pediatrics, Harvard Medical School, Boston, MA, USA Mucopolysaccharidosis I (MPS I) is a rare lysosomal storage disorder caused by deficiency of a L-iduronidase resulting in progressive multisystemic disease. Patients with MPS I often require multiple surgical interventions. Although the surgical risks are assumed to be high, there are no systematic reviews of deaths around the time of surgery among large MPS I populations. We queried the MPS I Registry, a volunteer observational database, for deaths occurring within 1 month of a surgical procedure among the 544 patients enrolled as of October 2006. For this cohort, surgical information from free-text fields was previously captured and analyzed. Among the 85 patients who reported at least one surgery and were deceased, 71 patients had dates for all surgeries. Fourteen of these 71 patients (20%) died within 1 month of a surgical procedure – 7 within 7 days of surgery. Phenotype was Hurler in 13/14 patients (93%). Death was caused by respiratory failure or airway issues in 5/14 patients (36%) and 11/14 patients (79%) died at 3 years of age. Mortality is probably underestimated, since at least one surgery preceded diagnosis in at least one-third of patients and deaths are not accounted for among undiagnosed patients. Patients with MPS I often have surgical procedures at an advanced stages of disease with an inherently high risk of morbidity and mortality. However, the young age of our cohort and the preponder-
Christiane Auray-Blaisa, Patrick Bhrera, Ren Gagnona, Joe T.R. Clarkea, Haoyue Zhangb, Yan Anb, Sarah P. Youngb, David S. Millingtonb, aUniversit de Sherbrooke, Sherbrooke, Canada, bDuke University Medical Center, Durham, NC, USA Introduction: Mucopolysaccharidosis (MPS) disorders are the result of primary defects in lysosomal enzymes. Enzyme replacement therapy (ERT) is available for MPS I, MPS II and MPS VI. Mass spectrometry offers the possibility to detect even subtle alterations in biological fluids of MPS patients compared to controls by identifying disease-specific glycosaminoglycans (GAG), e.g. chondroitin sulfate (CS), dermatan sulfate (DS) and heparin sulfate (HS). Method: Urine samples of MPS patients and age-matched controls were derivatized and analysis performed with an Alliance 2795XE LC coupled to a Quattro micro MS/MS (Waters). Gradient chromatography (8 min) was carried out with methanol and 10 mM ammonium acetate buffer using an Atlantis T3 column (Waters). Ionization of these molecules was performed by positive electrospray using the following MRM transitions: 426 > 236 (CS), 426 > 236 (DS) and 384 > 162 (HS). Results: Linearity was excellent (r2 over 0.995) with residuals below 15%. Limit of detection (LOD) and limit of quantification (LOQ) were 0.1 and 0.5 g/mL for CS, 10 and 25 g/mL for DS and HS, respectively. Levels of DS and HS in controls were below LOQ. Excretion of DS and HS by a MPS I patient was 17.8 and 78.0 mg/mmol creatinine, respectively. Excretion of DS and HS by a MPS II patient was 17.1 and 84.1 mg/mmol creatinine, respectively. Conclusion: We devised a simple, rapid and efficient MS/MS methodology for the measurement of urinary GAG applicable to high-risk screening and long term followup of MPS patients. doi:10.1016/j.ymgme.2009.10.024
8. How useful is urinary lyso-Gb3 as a biomarker for Fabry disease? Christiane Auray-Blaisa, Ren Gagnona, David G. Warnockb, Sarah P. Youngc, David S. Millingtonc, Joe T.R. Clarkea, aUniversit de Sherbrooke, Sherbrooke, QC, Canada, b University of Alabama at Birmingham, Birmingham, AL, USA, cDuke University Medical Center, Durham, NC, USA Introduction: Fabry disease (FD), an X-linked inborn error of glycosphingolipid catabolism, results in globotriaosylceramine (Gb3) accumulation in various tissues and body fluids. It was recently reported that lyso-Gb3, a deacylated Gb3, is increased in plasma of male FD patients with the classic phenotype, but no urinary lyso-Gb3 studies have been reported. We present the characterization of the molecule by time-of-flight mass spectrometry and a simple, efficient lyso-Gb3 methodology using tandem mass spectrometry (MS/MS). Method: Urinary lyso-Gb3 + internal standard were extracted using solid phase extraction, evaporated and 25 L injected into the MS/MS (Waters Micromass Quattro micro LC–MS/MS). MS/MS analyses were carried out by positive electrospray. Quantification was performed with the QuanLynx software. Urinary lyso-Gb3 content was normalized to creatinine. Results: Methodology validation showed a coefficient of variation less than 15% for intra- and inter-day assays, a limit of quantification at 50 pM, a limit of detection at 5 pM and a 60% recovery. No lyso-Gb3 was found in either adult or pediatric control cohorts. Urinary lyso-Gb3 excretion levels varied from 12 to 368 pM/mmol creatinine in 27 untreated FD patients and from 16 to 234 in 32 treated patients, about 1000 times less than Gb3/creatinine urine excretion levels. In untreated FD patients, a male (17y) and 2 females (39 and 62y) with the R363H missense mutation did not excrete lyso-Gb3 and had normal urinary Gb3/creatinine levels. Conclusions: Urinary lyso-Gb3 may be a useful biomarker for the investigation and follow-up of FD patients. doi:10.1016/j.ymgme.2009.10.025
9. Novel enzyme replacement therapy for Gaucher disease: Phase III pivotal clinical trial with plant cell expressed recombinantglucocerebrosidase (prGCD) – taliglucerase alpha David Aviezera,b, Einat Almon-Brilla, Yoseph Shaaltiela, Raul Chertkoffa, Sharon Hashmuelia, Ari Zimranb, aProtalix Biotherapeutics, Carmiel, Israel, bGaucher Clinic, Shaare Zedek Medical Center, Jerusalem, Israel
S9
ance of respiratory-related deaths point to the risk of surgery itself in this patient population. doi:10.1016/j.ymgme.2009.10.023
7. A urinary glycosaminoglycan analysis by tandem mass spectrometry doi:10.1016/j.ymgme.2009.10.021
5. Reliability of manual and automated tracing of hippocampal volumes in MPS patients and normal controls: A report of the neuroimaging core of the lysosomal disease network Alia Ahmed, Igor Nestrasil, Kyle Rudser, Elsa Shapiro, University of Minnesota, Minneapolis, MN, USA Background: Quantitative methods in neuroimaging such as diffusion tensor imaging (DTI) and volumetric analysis (VA) hold great promise to examine brain changes with disease and treatment in longitudinal studies of lysosomal diseases. Clinical scoring systems correlate with major functional abnormality but are not sensitive enough to examine treatment changes over time. Goals: Determine the reliability and validity of manual tracings and automated tracings of the hippocampus; automated systems may not provide accurate volumes. This work is necessary to set the standards for the Neuroimaging Core of the Lysosomal Disease Network to accurately quantify change in multiple studies. Methods: Both automated and manual tracings were obtained on 20 MPS I, 1 MPS II, and 2 MPS VII patients and 9 control subjects. Three-dimensional T 1 weighted gradient echo sequence (fast low angle shot – FLASH) was performed on the whole brain with a slice thickness of 1.0 mm. Automated volumes were calculated using FreeSurfer (run on the University of Minnesota Supercomputer) to determine feasibility in patients with abnormal brains. Brains2 manual tracings of the hippocampus were accomplished by two experienced raters. To obtain interrater and intrarater reliability, both raters (A.A. and I.N.) traced the same images and also traced 6 randomly chosen images twice. Results: Intrarater reliability determined by intraclass correlation was excellent at 0.99 indicating consistency within rater. Across two experienced raters, overall reliability for left and right, with and without white matter, was 0.93 (95% CI: 0.90, 0.95) with the lowest observed at 0.81. Comparing the Brains2 with FreeSurfer indicated that FreeSurfer overestimated hippocampal volumes by 60%. For example, while mean left hippocampus volume including white matter was 2.6 and 2.7 for the two raters, mean automated volumes were 4.2 cm3. Discussion: Brains2 manual tracing of hippocampal size was found to be reliable comparing within and across raters. FreeSurfer volumes were larger and probably overinclusive providing an overestimation of hippocampal plus hippocampus-adjacent white matter size. Other data indicates that the volumes of other structures are accurately determined by FreeSurfer but for the hippocampus, accurate manual tracings will be necessary. doi:10.1016/j.ymgme.2009.10.022
6. Post-surgical deaths among MPS I registry patients Pamela Arna,b,c,d, H. Warner Webba, Gerald Coxb, aThe Nemours Childrens Clinic, Jacksonville, FL, USA, bGenzyme Corporation, Cambridge, MA, USA, cDivision of Genetics, Childrens Hospital, Boston, USA, dDepartment of Pediatrics, Harvard Medical School, Boston, MA, USA Mucopolysaccharidosis I (MPS I) is a rare lysosomal storage disorder caused by deficiency of a L-iduronidase resulting in progressive multisystemic disease. Patients with MPS I often require multiple surgical interventions. Although the surgical risks are assumed to be high, there are no systematic reviews of deaths around the time of surgery among large MPS I populations. We queried the MPS I Registry, a volunteer observational database, for deaths occurring within 1 month of a surgical procedure among the 544 patients enrolled as of October 2006. For this cohort, surgical information from free-text fields was previously captured and analyzed. Among the 85 patients who reported at least one surgery and were deceased, 71 patients had dates for all surgeries. Fourteen of these 71 patients (20%) died within 1 month of a surgical procedure – 7 within 7 days of surgery. Phenotype was Hurler in 13/14 patients (93%). Death was caused by respiratory failure or airway issues in 5/14 patients (36%) and 11/14 patients (79%) died at 3 years of age. Mortality is probably underestimated, since at least one surgery preceded diagnosis in at least one-third of patients and deaths are not accounted for among undiagnosed patients. Patients with MPS I often have surgical procedures at an advanced stages of disease with an inherently high risk of morbidity and mortality. However, the young age of our cohort and the preponder-
Christiane Auray-Blaisa, Patrick Bhrera, Ren Gagnona, Joe T.R. Clarkea, Haoyue Zhangb, Yan Anb, Sarah P. Youngb, David S. Millingtonb, aUniversit de Sherbrooke, Sherbrooke, Canada, bDuke University Medical Center, Durham, NC, USA Introduction: Mucopolysaccharidosis (MPS) disorders are the result of primary defects in lysosomal enzymes. Enzyme replacement therapy (ERT) is available for MPS I, MPS II and MPS VI. Mass spectrometry offers the possibility to detect even subtle alterations in biological fluids of MPS patients compared to controls by identifying disease-specific glycosaminoglycans (GAG), e.g. chondroitin sulfate (CS), dermatan sulfate (DS) and heparin sulfate (HS). Method: Urine samples of MPS patients and age-matched controls were derivatized and analysis performed with an Alliance 2795XE LC coupled to a Quattro micro MS/MS (Waters). Gradient chromatography (8 min) was carried out with methanol and 10 mM ammonium acetate buffer using an Atlantis T3 column (Waters). Ionization of these molecules was performed by positive electrospray using the following MRM transitions: 426 > 236 (CS), 426 > 236 (DS) and 384 > 162 (HS). Results: Linearity was excellent (r2 over 0.995) with residuals below 15%. Limit of detection (LOD) and limit of quantification (LOQ) were 0.1 and 0.5 g/mL for CS, 10 and 25 g/mL for DS and HS, respectively. Levels of DS and HS in controls were below LOQ. Excretion of DS and HS by a MPS I patient was 17.8 and 78.0 mg/mmol creatinine, respectively. Excretion of DS and HS by a MPS II patient was 17.1 and 84.1 mg/mmol creatinine, respectively. Conclusion: We devised a simple, rapid and efficient MS/MS methodology for the measurement of urinary GAG applicable to high-risk screening and long term followup of MPS patients. doi:10.1016/j.ymgme.2009.10.024
8. How useful is urinary lyso-Gb3 as a biomarker for Fabry disease? Christiane Auray-Blaisa, Ren Gagnona, David G. Warnockb, Sarah P. Youngc, David S. Millingtonc, Joe T.R. Clarkea, aUniversit de Sherbrooke, Sherbrooke, QC, Canada, b University of Alabama at Birmingham, Birmingham, AL, USA, cDuke University Medical Center, Durham, NC, USA Introduction: Fabry disease (FD), an X-linked inborn error of glycosphingolipid catabolism, results in globotriaosylceramine (Gb3) accumulation in various tissues and body fluids. It was recently reported that lyso-Gb3, a deacylated Gb3, is increased in plasma of male FD patients with the classic phenotype, but no urinary lyso-Gb3 studies have been reported. We present the characterization of the molecule by time-of-flight mass spectrometry and a simple, efficient lyso-Gb3 methodology using tandem mass spectrometry (MS/MS). Method: Urinary lyso-Gb3 + internal standard were extracted using solid phase extraction, evaporated and 25 L injected into the MS/MS (Waters Micromass Quattro micro LC–MS/MS). MS/MS analyses were carried out by positive electrospray. Quantification was performed with the QuanLynx software. Urinary lyso-Gb3 content was normalized to creatinine. Results: Methodology validation showed a coefficient of variation less than 15% for intra- and inter-day assays, a limit of quantification at 50 pM, a limit of detection at 5 pM and a 60% recovery. No lyso-Gb3 was found in either adult or pediatric control cohorts. Urinary lyso-Gb3 excretion levels varied from 12 to 368 pM/mmol creatinine in 27 untreated FD patients and from 16 to 234 in 32 treated patients, about 1000 times less than Gb3/creatinine urine excretion levels. In untreated FD patients, a male (17y) and 2 females (39 and 62y) with the R363H missense mutation did not excrete lyso-Gb3 and had normal urinary Gb3/creatinine levels. Conclusions: Urinary lyso-Gb3 may be a useful biomarker for the investigation and follow-up of FD patients. doi:10.1016/j.ymgme.2009.10.025
9. Novel enzyme replacement therapy for Gaucher disease: Phase III pivotal clinical trial with plant cell expressed recombinantglucocerebrosidase (prGCD) – taliglucerase alpha David Aviezera,b, Einat Almon-Brilla, Yoseph Shaaltiela, Raul Chertkoffa, Sharon Hashmuelia, Ari Zimranb, aProtalix Biotherapeutics, Carmiel, Israel, bGaucher Clinic, Shaare Zedek Medical Center, Jerusalem, Israel