Comparative study on mannose 6-phosphate residue contents of recombinant lysosomal enzymes.
As most recombinant lysosomal enzymes are incorporated into cells via mannose 6-phosphate (M6P) receptors, the M6P content is important for effective enzyme replacement therapy (ERT) for lysosomal diseases. However, there have been no comprehensive reports of the M6P contents of lysosomal enzymes. We developed an M6P assay method comprising three steps, i.e., acid hydrolysis of glycoproteins, derivatization of M6P, and high-performance liquid chromatography, and determined the M6P contents of six recombinant lysosomal enzymes now available for ERT and one in the process of development. The assay is easy, specific, and reproducible. The results of the comparative study revealed that the M6P contents of agalsidase alfa, agalsidase beta, modified α-N-acetylgalactosaminidase, alglucosidase alfa, laronidase, idursulfase, and imiglucerase are 2.1, 2.9, 5.9, 0.7, 2.5, 3.2, and <0.3 mol/mol enzyme, respectively. The results were correlated with those of the biochemical analyses previously performed and that of the binding assay of exposed M6P of the enzymes with the domain 9 of the cation-independent M6P receptor. This assay method is useful for comparison of the M6P contents of recombinant lysosomal enzymes for ERT.
Department of Functional Bioanalysis, Meiji Pharmaceutical University, Tokyo 204-8588, Japan.
Tadayasu
T
Togawa
Department of Analytical Biochemistry, Meiji Pharmaceutical University, Tokyo 204-8588, Japan.
Masaru
M
Takada
Department of Analytical Biochemistry, Meiji Pharmaceutical University, Tokyo 204-8588, Japan.
Yoshiaki
Y
Aizawa
Department of Functional Bioanalysis, Meiji Pharmaceutical University, Tokyo 204-8588, Japan.
Takahiro
T
Tsukimura
Bioprocess Research Institute, National Institute of Advanced Industrial Science and Technology, Tsukuba 305-8566, Japan.
Yasunori
Y
Chiba
Department of Clinical Genetics, Meiji Pharmaceutical University, Tokyo 204-8588, Japan. Electronic address: sakuraba@my-pharm.ac.jp.
Hitoshi
H
Sakuraba
D056947:Enzyme Replacement Therapy; D006801:Humans; D006867:Hydrolases / Q000737:chemistry; D007527:Isoenzymes / Q000737:chemistry; D008247:Lysosomes / Q000737:chemistry / Q000201:enzymology*; D008360:Mannosephosphates / Q000737:chemistry* / Q000302:isolation & purification; D017527:Receptor, IGF Type 2 / Q000737:chemistry* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry; D000519:alpha-Galactosidase / Q000737:chemistry
D007527:Isoenzymes; D008360:Mannosephosphates; D017527:Receptor, IGF Type 2; D011994:Recombinant Proteins; C000627036:agalsidase alfa; C027693:mannose-6-phosphate; D006867:Hydrolases; D000519:alpha-Galactosidase; C459420:agalsidase beta
Enzyme replacement therapy; Lysosomal disease; Lysosomal enzyme; Mannose 6-phosphate