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US 8,940,513 B2Granted
Manufacture of active highly phosphorylated human N-acetylgalactosamine-6-sulfatase and uses thereof
AssigneeBiomarin Pharmaceutical Inc.
InventorsVish Koppaka, Michel Claude Vellard, Augustus Okhamafe, Kidisti Araya
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Abstract
This invention provides compositions of active highly phosphorylated human N-acetylgalactosamine-6-sulfatase (GALNS), and pharmaceutical compositions and formulations thereof, methods of producing and purifying GALNS, and its use in the diagnosis, prophylaxis, or treatment of diseases and conditions, including particularly lysosomal storage diseases that are caused by, or associated with, a deficiency in the GALNS enzyme, e.g., Mucopolysaccharidosis IVa (MPS IVa or Morquio A syndrome).
Claims20
A formulation comprising
- (a) recombinant human N-acetylgalactosamine-6-sulfatase (GALNS) enzyme, said GALNS enzyme comprising an amino acid sequence at least 95% identical to amino acids 27 to 522 of SEQ ID NO:4, and
- having a purity of at least 95% as determined by Coomassie Blue staining when subjected to SDS-PAGE under non-reducing conditions,
- having at least 50% conversion of the cysteine residue at position 53 to Cα-formylglycine (FGly), and
- having between 0.5 to 0.8 bis-phosphorylated oligomannose chains per monomeric protein chain,
- wherein at least 98% of said GALNS enzyme is in the precursor form as determined by SDS-CGE, and
- (b) one or more pharmaceutically acceptable carriers, diluents or excipients comprising:
- (i) an amount of phosphate buffer effective to reduce dephosphorylation of said GALNS enzyme, wherein the phosphate buffer is NaH2PO4 at a concentration from about 25 mM to 75 mM;
- and
- (ii) a stabilizing amount of the following stabilizers:
- an arginine salt or buffer, optionally arginine hydrochloride, wherein the arginine salt or buffer is at a concentration from about 10 mM to 50 mM;
- a polysorbate, optionally polysorbate 20;
- and a trihydric or higher sugar alcohol, optionally sorbitol;
- wherein said formulation is at a pH of about 5.0-5.8.
10 more elements
A formulation comprising
- (a) recombinant human N-acetylgalactosamine-6-sulfatase (GALNS) enzyme, said GALNS enzyme comprising an amino acid sequence at least 95% identical to amino acids 27 to 522 of SEQ ID NO:4, and
- having a purity of at least 95% as determined by Coomassie Blue staining when subjected to SDS-PAGE under non-reducing conditions,
- having at least 50% conversion of the cysteine residue at position 53 to Cα-formylglycine (FGly), and
- having between 0.5 to 0.8 bis-phosphorylated oligomannose chains per monomeric protein chain,
- wherein at least 98% of said GALNS enzyme is in the precursor form as determined by Coomassie Blue staining when subjected to SDS-PAGE under reducing conditions, and
- (b) one or more pharmaceutically acceptable carriers, diluents or excipients comprising:
- (i) NaOAc/HOAc and NaH2PO4 as buffering agents, wherein the concentration of NaOAc/HOAc is about 20+/−10 mM and the concentration of NaH2PO4 is about 50+/−25 mM;
- (ii) Arginine HCl, Tween-20 and sorbitol as stabilizers, wherein the concentration of Arginine HCl is about 30+/−20 mM, the concentration of Tween-20 is about 0.01%+/−0.005% (w/v) and the concentration of sorbitol is about 2.0%+/−1.0% (w/v);
- and
- (iii) a pH of about 5.4+/−0.4.
7 more elements
Dependencies are inferred from the claim text.