title: Hydrophobic Man-1-P derivatives correct abnormal glycosylation in Type I congenital disorder of glycosylation fibroblasts.
abstract: Patients with Type I congenital disorders of glycosylation (CDG-I) make incomplete lipid-linked oligosaccharides (LLO). These glycans are poorly transferred to proteins resulting in unoccupied glycosylation sequons. Mutations in phosphomannomutase (PMM2) cause CDG-Ia by reducing the activity of PMM, which converts mannose (Man)-6-P to Man-1-P before formation of GDP-Man. These patients have reduced Man-1-P and GDP-Man. To replenish intracellular Man-1-P pools in CDG-Ia cells, we synthesized two hydrophobic, membrane permeable acylated versions of Man-1-P and determined their ability to normalize LLO size and N-glycosylation in CDG-Ia fibroblasts. Both compounds, compound I (diacetoxymethyl 2,3,4,6-tetra-O-acetyl-alpha-D-mannopyranosyl phosphate) (C-I) and compound II (diacetoxymethyl 2,3,4,6-tetra-O-ethyloxycarbonyl-alpha-D-mannopyranosyl phosphate) (C-II), contain two acetoxymethyl (CH2OAc) groups O-linked to phosphorous. C-I contains acetyl esters and C-II contains ethylcarbonate (CO2Et) esters on the Man residue. Both C-I and C-II normalized truncated LLO, but C-II was about 2-fold more efficient than C-I. C-II replenished the GDP-Man pool in CDG-Ia cells and was more efficiently incorporated into glycoproteins than exogenous Man at low concentrations (25-75 mM). In a glycosylation assay of DNaseI in CDG-Ia cells, C-II restored glycosylation to control cell levels. C-II also corrected impaired LLO biosynthesis in cells from a Dolichol (Dol)-P-Man deficient patient (CDG-Ie) and partially corrected LLO in cells from an ALG12 mannosyltransferase-deficient patient (CDG-Ig), whereas cells from an ALG3-deficient patient (CDG-Id) and from an MPDU1-deficient patient (CDG-If) were not corrected. These results validate the general concept of using pro-Man-1-P substrates as potential therapeutics for CDG-I patients.
authors.affiliation: The Burnham Institute, La Jolla, CA 92037, USA.
authors.forename: Erik A
authors.initials: EA
authors.lastname: Eklund
authors.forename: Nabyl
authors.initials: N
authors.lastname: Merbouh
authors.forename: Mie
authors.initials: M
authors.lastname: Ichikawa
authors.forename: Atsushi
authors.initials: A
authors.lastname: Nishikawa
authors.forename: Jessica M
authors.initials: JM
authors.lastname: Clima
authors.forename: James A
authors.initials: JA
authors.lastname: Dorman
authors.forename: Thomas
authors.initials: T
authors.lastname: Norberg
authors.forename: Hudson H
authors.initials: HH
authors.lastname: Freeze
mesh_terms: D002236:Carbohydrate Conformation; D050260:Carbohydrate Metabolism; D049109:Cell Proliferation / Q000187:drug effects; D018981:Congenital Disorders of Glycosylation / Q000378:metabolism*; D003470:Culture Media / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D005347:Fibroblasts / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D006031:Glycosylation / Q000187:drug effects; D006801:Humans; D008360:Mannosephosphates / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D009154:Mutation; D017875:Phosphotransferases (Phosphomutases) / Q000235:genetics / Q000378:metabolism; D013403:Sugar Phosphates / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D013997:Time Factors
chemical_list: D003470:Culture Media; D008360:Mannosephosphates; D013403:Sugar Phosphates; C508474:diacetoxymethyl 2,3,4,6-tetra-O-acetylmannopyranosyl phosphate; C508475:diacetoxymethyl 2,3,4,6-tetra-O-ethyloxycarbonylmannopyranosyl phoshate; C047217:mannose 1-phosphate; D017875:Phosphotransferases (Phosphomutases); C052799:phosphomannomutase