title: Characterization of a chemically modified plant cell culture expressed human α-Galactosidase-A enzyme for treatment of Fabry disease.
abstract: Fabry disease is an X-linked recessive disorder caused by the loss of function of the lysosomal enzyme α-Galactosidase-A. Although two enzyme replacement therapies (ERTs) are commercially available, they may not effectively reverse some of the Fabry pathology. PRX-102 is a novel enzyme for the therapy of Fabry disease expressed in a BY2 Tobacco cell culture. PRX-102 is chemically modified, resulting in a cross-linked homo-dimer. We have characterized the in-vitro and in-vivo properties of PRX-102 and compared the results with the two commercially produced α-Galactosidase-A enzymes. Results show that PRX-102 has prolonged in-vitro stability in plasma, after 1h incubation it retains 30% activity compared with complete inactivation of the commercial enzymes. Under lysosomal-like conditions PRX-102 maintains over 80% activity following 10 days of incubation, while commercial enzymes become inactive after 2days. Pharmacokinetic profile of PRX-102 measured in male Fabry mice shows a 10 fold increase in t1/2 in mice (581min) compared to approved drugs. The enzyme has significantly different kinetic parameters to the alternative ERTs available (p-value<0.05, one way ANOVA), although these differences do not indicate any significant biochemical variations. PRX-102 is uptaken to primary human Fabry fibroblasts. The repeat administration of the enzyme to Fabry mice caused significant reduction (p-value<0.05) of Gb3 in various tissues (the measured residual content was 64% in kidney, liver was cleaned, 23% in heart, 5.7% in skin and 16.2% in spleen). PRX-102 has a relatively simple glycosylation pattern, characteristic to plants, having mainly tri-mannose structures with the addition of either α(1-3)-linked fucose or β(1-2)-linked xylose, or both, in addition to various high mannose structures, while agalsidase beta has a mixture of sialylated glycans in addition to high mannose structures. This study concludes that PRX-102 is equivalent in functionality to the current ERTs available, with superior stability and prolonged circulatory half-life. Therefore we propose that PRX-102 is a promising alternative for treatment of Fabry disease.
authors.affiliation: Protalix Biotherapeutics, 2 Snunit Street, P.O. Box 455, Carmiel 2161401, Israel.
authors.forename: Tali
authors.initials: T
authors.lastname: Kizhner
authors.affiliation: Protalix Biotherapeutics, 2 Snunit Street, P.O. Box 455, Carmiel 2161401, Israel.
authors.forename: Yaniv
authors.initials: Y
authors.lastname: Azulay
authors.affiliation: Protalix Biotherapeutics, 2 Snunit Street, P.O. Box 455, Carmiel 2161401, Israel.
authors.forename: Mariana
authors.initials: M
authors.lastname: Hainrichson
authors.affiliation: Protalix Biotherapeutics, 2 Snunit Street, P.O. Box 455, Carmiel 2161401, Israel. Electronic address: yoram.tekoah@protalix.com.
authors.forename: Yoram
authors.initials: Y
authors.lastname: Tekoah
authors.affiliation: Protalix Biotherapeutics, 2 Snunit Street, P.O. Box 455, Carmiel 2161401, Israel.
authors.forename: Gil
authors.initials: G
authors.lastname: Arvatz
authors.affiliation: Protalix Biotherapeutics, 2 Snunit Street, P.O. Box 455, Carmiel 2161401, Israel.
authors.forename: Avidor
authors.initials: A
authors.lastname: Shulman
authors.affiliation: Protalix Biotherapeutics, 2 Snunit Street, P.O. Box 455, Carmiel 2161401, Israel.
authors.forename: Ilya
authors.initials: I
authors.lastname: Ruderfer
authors.affiliation: Protalix Biotherapeutics, 2 Snunit Street, P.O. Box 455, Carmiel 2161401, Israel; Faculty of Life Sciences, Bar Ilan University, Ramat Gan, 5290002 Israel.
authors.forename: David
authors.initials: D
authors.lastname: Aviezer
authors.affiliation: Protalix Biotherapeutics, 2 Snunit Street, P.O. Box 455, Carmiel 2161401, Israel.
authors.forename: Yoseph
authors.initials: Y
authors.lastname: Shaaltiel
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D056947:Enzyme Replacement Therapy; D004795:Enzyme Stability; D000795:Fabry Disease / Q000188:drug therapy*; D006321:Heart; D007527:Isoenzymes / Q000627:therapeutic use; D007668:Kidney / Q000201:enzymology; D008099:Liver / Q000201:enzymology; D008297:Male; D051379:Mice; D011994:Recombinant Proteins / Q000627:therapeutic use; D012867:Skin / Q000201:enzymology; D013154:Spleen / Q000201:enzymology; D014026:Nicotiana / Q000235:genetics; D000519:alpha-Galactosidase / Q000235:genetics* / Q000493:pharmacokinetics / Q000627:therapeutic use*
chemical_list: D007527:Isoenzymes; D011994:Recombinant Proteins; D000519:alpha-Galactosidase; C459420:agalsidase beta
keywords: ERT; Enzyme stability; Fabry disease; Protein chemical modification; globotriaosylceramide (Gb(3)); α-Galactosidase-A