title: Characterization and pharmacokinetic study of recombinant human N-acetylgalactosamine-6-sulfate sulfatase.
abstract: Mucopolysaccharidosis IVA (MPS IVA) is an autosomal recessive disorder caused by a deficiency of N-acetylgalactosamine-6-sulfate sulfatase (GALNS). The aims of this study were to establish Chinese hamster ovary (CHO) cells overexpressing recombinant human GALNS (rhGALNS) and to assess pharmacokinetics and tissue distribution of purified enzymes by using MPS IVA knock-out mouse (Galns(-/-)). The CHO-cell derived rhGALNS was purified from the media by a two-step affinity chromatography procedure. The rhGALNS was administered intravenously to 3-month-old Galns(-/-) mice at a single dose of 250U/g of body weight. The treated mice were examined by assaying the GALNS activity at baseline and up to 240min to assess clearance of the enzyme from blood circulation. The mice were sacrificed 4h after infusion of the enzyme to study the enzyme distribution in tissues. The rhGALNS was purified 1317-fold with 71% yield. The enzyme was taken up by Galns(-/-) chondrocytes (150U/mg/15h). The uptake was inhibited by mannose-6-phosphate. The enzyme activity disappeared from circulation with a half-life of 2.9min. After enzyme infusion, the enzyme was taken up and detected in multiple tissues (40.7% of total infused enzymes in liver). Twenty-four hours after a single infusion of the fluorescence-labeled enzymes into MPS IVA mice, biodistribution pattern showed the amount of tagged enzyme retained in bone, bone marrow, liver, spleen, kidney, and heart. In conclusion, we have shown that the phosphorylated rhGALNS is delivered to multiple tissues, including bone, and that it functions bioactively in Galns(-/-) chondrocytes implying a potential enzyme replacement treatment.
authors.affiliation: Department of Pediatrics, Saint Louis University, Pediatric Research Institute, St Louis, MO 63110-2586, USA. tomatsus@slu.edu
authors.forename: Shunji
authors.initials: S
authors.lastname: Tomatsu
authors.forename: Adriana M
authors.initials: AM
authors.lastname: Montaño
authors.forename: Monica
authors.initials: M
authors.lastname: Gutierrez
authors.forename: Jeffrey H
authors.initials: JH
authors.lastname: Grubb
authors.forename: Hirotaka
authors.initials: H
authors.lastname: Oikawa
authors.forename: Vu Chi
authors.initials: VC
authors.lastname: Dung
authors.forename: Amiko
authors.initials: A
authors.lastname: Ohashi
authors.forename: Tatsuo
authors.initials: T
authors.lastname: Nishioka
authors.forename: Masamichi
authors.initials: M
authors.lastname: Yamada
authors.forename: Mana
authors.initials: M
authors.lastname: Yamada
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Tosaka
authors.forename: Georgeta G
authors.initials: GG
authors.lastname: Trandafirescu
authors.forename: Tadao
authors.initials: T
authors.lastname: Orii
mesh_terms: D000818:Animals; D016466:CHO Cells; D002811:Chondroitinsulfatases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000493:pharmacokinetics*; D006224:Cricetinae; D003412:Cricetulus; D004195:Disease Models, Animal; D004795:Enzyme Stability; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D008822:Mice, Transgenic; D009085:Mucopolysaccharidosis IV / Q000188:drug therapy / Q000201:enzymology; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000493:pharmacokinetics*; D013997:Time Factors; D014018:Tissue Distribution
chemical_list: D011994:Recombinant Proteins; D002811:Chondroitinsulfatases; C102295:GALNS protein, human; C491158:Galns protein, mouse