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MS measurement of glycosaminoglycans in amniotic fluid of a MPS VII fetus
Abstracts / Molecular Genetics and Metabolism 120 (2016) S17–S145
Newark, DE, United States, bNemours/ Alfred I. duPont Children Hospital, Wilmington, DE, United States, cGifu University, Gifu, Japan, d UFRGS, Porto Alegre, Brazil, eHCPA, Porto Alegre, Brazil, fSt Mary's Hospital Manchester, Manchester, United Kingdom, gVietnam National Children's Hospital, Hanoi, Viet Nam, hShimane University, Izumo, Japan, iManipal University, Manipal, India Glycosaminoglycan (GAG) analyses in serum (or plasma) of patients with mucopolysaccharidoses (MPS) and mucolipidoses (ML) by tandem mass spectrometry have been established; however, there has been a limited study of GAG levels in dried blood spots (DBS) in these patients. The aim of this study was to compare levels of GAG in DBS from 124 MPS and ML patients with levels in DBS from 115 age-matched controls. Disaccharides were produced from polymer GAG by digestion with chondroitinase B, heparitinase, and keratanase II. Subsequently, dermatan sulfate (DS), heparan sulfate (HS-0S, HS-NS), and keratan sulfate (mono-sulfated KS, disulfated KS, and ratio di-KS in total KS) were measured by liquid chromatography/tandem mass spectrometry (LC/MS/MS). Untreated patients with MPS I, II, VI, and ML had higher levels of DS compared to control samples, with MPS I patients having the highest level. Untreated patients with MPS I, II, III, VI, and ML had higher levels of HS-0S than controls, with ML having the highest level. Untreated MPS I, II, III, and ML patients showed higher levels than controls, with MPS II having the highest. For KS, only MPS II and MPS IV showed higher levels than controls. For treated MPS I patients, DS and HS were significantly lower after ERT and even lower after HSCT. For MPS III, HSCT did not decrease HS (0S and NS, one MPS IIIA and two MPS IIIB patients). DS was reduced in an HSCT-treated MPS VI patient. Although HS levels in a HSCT treated MPS VII patient were similar to control, untreated MPS VII samples were not available to determine whether this represented a decrease in levels. Overall, it appears that GAG levels in DBS measured by LC/MS/MS may be a useful tool for diagnosis and treatment efficacy monitoring in MPS and ML.
doi:10.1016/j.ymgme.2016.11.183
175 Hematopoietic stem cell transplantation for patients with mucopolysaccharidosis type II Francyne Kubaskia,b, Yasuyuki Suzukic, Hiromasa Yabec, Robert W. Masona,d, Li Xied, Tor G.H. Onstene,f, Sandra Leistner-Segale,f, Roberto Giuglianie,f, Vũ C. Dũngg, Can T.B. Ngocg, Can T.B. Ngocg, Seiji Yamaguchih, Adriana M. Montañoi, Kenji E. Oriic, Toshiyuki Fukaoc, Haruo Shinatakuj, Tadao Oriic, Shunji Tomatsua,d, aUniversity of Delaware, Newark, DE, United States, bNemours/Alfred I. duPont Children Hospital, Wilmington, DE, United States, cGifu University, Gifu, Japan, dNemours/ Alfred I. duPont Children Hospital, Wilmington, DE, United States, eHCPA, Porto Alegre, Brazil, fUFRGS, Porto Alegre, Brazil, g Vietnam National Children's Hospital, Hanoi, Viet Nam, hShimane University, Izumo, Japan, iSaint Louis University, Saint Louis, MO, United States, jOsaka City University Graduate School of Medicine, Osaka, Japan Little has been reported about the long-term outcomes of hematopoietic stem cell transplantation (HSCT) for mucopolysaccharidosis type II (MPS II), except sporadic cases. In this study, clinical and biochemical findings were compared between patients who underwent HSCT and/or enzyme replacement therapy (ERT). Demographic data for 130 HSCT patients were derived from 107 published cases and from 23 new cases from Japan, were compared with 54 ERT-treated and 11 untreated cases. Activity of daily living (ADL) was also assessed.
S77
Glycosaminoglycan (GAG) levels were analyzed by liquid chromatography tandem mass spectrometry in blood samples from HSCT, ERT, and untreated patients, as well as age-matched controls. Mean age at HSCT was as follow: 5.5 years (10m to 19.8 years) in published cases, and 4.7 years (1.7 to 9 years) in the new cases, respectively. Graftversus-host disease occurred in 8 (16%) out of 51 published cases, and 9 (18%) of those cases died due to transplant-associated complications (none of the new cases died). Most HSCT patients presented improvement in somatic features and joint movements and had better ADL related with “Movement” and “Movement with Cognition” than ERT patients. Both therapies provided a significant reduction of GAG, but in average HSCT had lower GAG levels. Two patients with attenuated phenotype had improvement in MRI findings post-HSCT. Although controlled studies are needed, this report indicates that HSCT seems to be effective for patients with MPS II and could be considered as a treatment option. doi:10.1016/j.ymgme.2016.11.184
176 LC/MS/MS measurement of glycosaminoglycans in amniotic fluid of a MPS VII fetus Francyne Kubaskia,b, Robert W. Masona,b, Maira G. Burinc, Kristiane Michelin-Tirellic, Rejane G. Kesslerc, Fernanda Benderc, Ana C. Brusius-Facchinc, Sandra Leistner-Segalc, Carolina A. Morenod, Denise P. Cavalcantid, Roberto Giuglianic,e, Shunji Tomatsub,f, aUniversity of Delaware, Newark, DE, United States, bNemours/Alfred I. duPont Hospital for Children, Wilmington, DE, United States, cHCPA, Porto Alegre, Brazil, dUNICAMP, Campinas, Brazil, eUFRGS, Porto Alegre, Brazil, f UDEL, Newark, DE, United States Mucopolysaccharidosis VII or Sly syndrome is an autosomal recessive disorder caused by deficiency of β-glucuronidase leading to accumulation of dermatan sulfate (DS), chondroitin sulfate (C4S & C6S), and heparan sulfate (HS). MPS VII has a broad clinical spectrum, from the most severe lethal hydrops fetalis to attenuated forms with survival into adulthood despite somatic and cognitive impairment. The aim of this study was to quantify glycosaminoglycans in amniotic fluid from a MPS VII fetus and to compare levels with 5 age-matched AF obtained from normal pregnancies (19 to 21 weeks of gestation). Disaccharides were produced from polymer GAG by digestion with chondroitinase B, chondroitinase ACII, heparitinase, and keratanase II. DS, CS (6S & 4S), HS (NS, 0S, and diS1) and keratan sulfate (mono & di-sulfated) levels were measured by liquid chromatography tandem mass spectrometry (LC/MS/MS). Prenatal studies were initiated due to fetal hydrops at 21 weeks of gestation. No activity of β-glucuronidase was detected in fetal cells. The pregnancy spontaneously terminated in the third trimester, the fetus was stillborn. Genetic studies identified a mutation in homozygosis at the GUSB gene. All GAG were increased in the MPS VII AF compared to the age-matched controls AF (mean +SD). HS-0S (890 ng/mg protein) (controls: 59 ± 25 ng/mg protein); C6S, DS and HS-NS (621, 712, and 142 ng/mg protein, respectively) in the MPS VII (controls: 57 ± 19, 61 ± 44, 12 ± 5 ng/mg protein, respectively), mono-sulfated KS (361 ng/mg protein) (controls: 131 ± 91 ng/mg protein), C4S (543 ng/mg protein) (controls: 142 ± 56 ng/mg protein), and di-sulfated KS (25 ng/mg protein) (controls: 6 ± 2 ng/mg protein). No elevation of HS-diS1 was observed. This is the first report of GAG analysis in AF from MPS VII fetus quantified by LC/MS/MS and shows GAG elevation at 21 weeks of gestation. doi:10.1016/j.ymgme.2016.11.185
Newark, DE, United States, bNemours/ Alfred I. duPont Children Hospital, Wilmington, DE, United States, cGifu University, Gifu, Japan, d UFRGS, Porto Alegre, Brazil, eHCPA, Porto Alegre, Brazil, fSt Mary's Hospital Manchester, Manchester, United Kingdom, gVietnam National Children's Hospital, Hanoi, Viet Nam, hShimane University, Izumo, Japan, iManipal University, Manipal, India Glycosaminoglycan (GAG) analyses in serum (or plasma) of patients with mucopolysaccharidoses (MPS) and mucolipidoses (ML) by tandem mass spectrometry have been established; however, there has been a limited study of GAG levels in dried blood spots (DBS) in these patients. The aim of this study was to compare levels of GAG in DBS from 124 MPS and ML patients with levels in DBS from 115 age-matched controls. Disaccharides were produced from polymer GAG by digestion with chondroitinase B, heparitinase, and keratanase II. Subsequently, dermatan sulfate (DS), heparan sulfate (HS-0S, HS-NS), and keratan sulfate (mono-sulfated KS, disulfated KS, and ratio di-KS in total KS) were measured by liquid chromatography/tandem mass spectrometry (LC/MS/MS). Untreated patients with MPS I, II, VI, and ML had higher levels of DS compared to control samples, with MPS I patients having the highest level. Untreated patients with MPS I, II, III, VI, and ML had higher levels of HS-0S than controls, with ML having the highest level. Untreated MPS I, II, III, and ML patients showed higher levels than controls, with MPS II having the highest. For KS, only MPS II and MPS IV showed higher levels than controls. For treated MPS I patients, DS and HS were significantly lower after ERT and even lower after HSCT. For MPS III, HSCT did not decrease HS (0S and NS, one MPS IIIA and two MPS IIIB patients). DS was reduced in an HSCT-treated MPS VI patient. Although HS levels in a HSCT treated MPS VII patient were similar to control, untreated MPS VII samples were not available to determine whether this represented a decrease in levels. Overall, it appears that GAG levels in DBS measured by LC/MS/MS may be a useful tool for diagnosis and treatment efficacy monitoring in MPS and ML.
doi:10.1016/j.ymgme.2016.11.183
175 Hematopoietic stem cell transplantation for patients with mucopolysaccharidosis type II Francyne Kubaskia,b, Yasuyuki Suzukic, Hiromasa Yabec, Robert W. Masona,d, Li Xied, Tor G.H. Onstene,f, Sandra Leistner-Segale,f, Roberto Giuglianie,f, Vũ C. Dũngg, Can T.B. Ngocg, Can T.B. Ngocg, Seiji Yamaguchih, Adriana M. Montañoi, Kenji E. Oriic, Toshiyuki Fukaoc, Haruo Shinatakuj, Tadao Oriic, Shunji Tomatsua,d, aUniversity of Delaware, Newark, DE, United States, bNemours/Alfred I. duPont Children Hospital, Wilmington, DE, United States, cGifu University, Gifu, Japan, dNemours/ Alfred I. duPont Children Hospital, Wilmington, DE, United States, eHCPA, Porto Alegre, Brazil, fUFRGS, Porto Alegre, Brazil, g Vietnam National Children's Hospital, Hanoi, Viet Nam, hShimane University, Izumo, Japan, iSaint Louis University, Saint Louis, MO, United States, jOsaka City University Graduate School of Medicine, Osaka, Japan Little has been reported about the long-term outcomes of hematopoietic stem cell transplantation (HSCT) for mucopolysaccharidosis type II (MPS II), except sporadic cases. In this study, clinical and biochemical findings were compared between patients who underwent HSCT and/or enzyme replacement therapy (ERT). Demographic data for 130 HSCT patients were derived from 107 published cases and from 23 new cases from Japan, were compared with 54 ERT-treated and 11 untreated cases. Activity of daily living (ADL) was also assessed.
S77
Glycosaminoglycan (GAG) levels were analyzed by liquid chromatography tandem mass spectrometry in blood samples from HSCT, ERT, and untreated patients, as well as age-matched controls. Mean age at HSCT was as follow: 5.5 years (10m to 19.8 years) in published cases, and 4.7 years (1.7 to 9 years) in the new cases, respectively. Graftversus-host disease occurred in 8 (16%) out of 51 published cases, and 9 (18%) of those cases died due to transplant-associated complications (none of the new cases died). Most HSCT patients presented improvement in somatic features and joint movements and had better ADL related with “Movement” and “Movement with Cognition” than ERT patients. Both therapies provided a significant reduction of GAG, but in average HSCT had lower GAG levels. Two patients with attenuated phenotype had improvement in MRI findings post-HSCT. Although controlled studies are needed, this report indicates that HSCT seems to be effective for patients with MPS II and could be considered as a treatment option. doi:10.1016/j.ymgme.2016.11.184
176 LC/MS/MS measurement of glycosaminoglycans in amniotic fluid of a MPS VII fetus Francyne Kubaskia,b, Robert W. Masona,b, Maira G. Burinc, Kristiane Michelin-Tirellic, Rejane G. Kesslerc, Fernanda Benderc, Ana C. Brusius-Facchinc, Sandra Leistner-Segalc, Carolina A. Morenod, Denise P. Cavalcantid, Roberto Giuglianic,e, Shunji Tomatsub,f, aUniversity of Delaware, Newark, DE, United States, bNemours/Alfred I. duPont Hospital for Children, Wilmington, DE, United States, cHCPA, Porto Alegre, Brazil, dUNICAMP, Campinas, Brazil, eUFRGS, Porto Alegre, Brazil, f UDEL, Newark, DE, United States Mucopolysaccharidosis VII or Sly syndrome is an autosomal recessive disorder caused by deficiency of β-glucuronidase leading to accumulation of dermatan sulfate (DS), chondroitin sulfate (C4S & C6S), and heparan sulfate (HS). MPS VII has a broad clinical spectrum, from the most severe lethal hydrops fetalis to attenuated forms with survival into adulthood despite somatic and cognitive impairment. The aim of this study was to quantify glycosaminoglycans in amniotic fluid from a MPS VII fetus and to compare levels with 5 age-matched AF obtained from normal pregnancies (19 to 21 weeks of gestation). Disaccharides were produced from polymer GAG by digestion with chondroitinase B, chondroitinase ACII, heparitinase, and keratanase II. DS, CS (6S & 4S), HS (NS, 0S, and diS1) and keratan sulfate (mono & di-sulfated) levels were measured by liquid chromatography tandem mass spectrometry (LC/MS/MS). Prenatal studies were initiated due to fetal hydrops at 21 weeks of gestation. No activity of β-glucuronidase was detected in fetal cells. The pregnancy spontaneously terminated in the third trimester, the fetus was stillborn. Genetic studies identified a mutation in homozygosis at the GUSB gene. All GAG were increased in the MPS VII AF compared to the age-matched controls AF (mean +SD). HS-0S (890 ng/mg protein) (controls: 59 ± 25 ng/mg protein); C6S, DS and HS-NS (621, 712, and 142 ng/mg protein, respectively) in the MPS VII (controls: 57 ± 19, 61 ± 44, 12 ± 5 ng/mg protein, respectively), mono-sulfated KS (361 ng/mg protein) (controls: 131 ± 91 ng/mg protein), C4S (543 ng/mg protein) (controls: 142 ± 56 ng/mg protein), and di-sulfated KS (25 ng/mg protein) (controls: 6 ± 2 ng/mg protein). No elevation of HS-diS1 was observed. This is the first report of GAG analysis in AF from MPS VII fetus quantified by LC/MS/MS and shows GAG elevation at 21 weeks of gestation. doi:10.1016/j.ymgme.2016.11.185