Formulations comprising recombinant acid alpha-glucosidase
Provided are pharmaceutical formulations comprising a recombinant acid α-glucosidase, wherein the recombinant acid α-glucosidase is expressed in Chinese hamster ovary (CHO) cells and comprises an increased content of N-glycan units bearing one or two mannose-6-phosphate residues when compared to a content of N-glycan units bearing one or two mannose-6-phosphate residues of alglucosidase alfa; at least one buffer selected from the group consisting of a citrate, a phosphate and combinations thereof; and at least one excipient selected from the group consisting of mannitol, polysorbate 80, and combinations thereof, wherein the formulation has a pH of from about 5.0 to about 7.0. Also provided are methods of treating Pompe disease using these pharmaceutical formulations.
1. A pharmaceutical formulation comprising:
(a) a population of recombinant human acid α-glucosidase (rhGAA) molecules, wherein the rhGAA molecules are expressed in Chinese hamster ovary (CHO) cells and comprise a first, second, third, fourth, fifth, sixth, and seventh potential N-glycosylation site at amino acids corresponding to N84, N177, N334, N414, N596, N826, and N869 of SEQ ID NO:2, respectively, wherein 40%-60% of the N-glycans on the rhGAA molecules are complex type N-glycans, and wherein at least 50% of the rhGAA molecules comprise a glycan bearing bis-mannose-6-phosphate (bis-M6P) at the first potential N-glycosylation site;
(b) a citrate buffer, and
(c) at least one excipient selected from the group consisting of mannitol, polysorbate 80, and combinations thereof,
wherein the formulation has a pH of from about 5.0 to about 7.0.
2. The pharmaceutical formulation of claim 1 , wherein the rhGAA molecules are present in a concentration of about 5 mg/mL to about 50 mg/mL.
3. The pharmaceutical formulation of claim 2 , wherein the rhGAA molecules are present in a concentration of about 15 mg/mL.
4. The pharmaceutical formulation of claim 1 , wherein the formulation has a pH of from about 5.5 to about 7.0.
5. The pharmaceutical formulation of claim 4 , wherein the formulation has a pH of about 6.0.
6. The pharmaceutical formulation of claim 1 , wherein the citrate buffer comprises a potassium, sodium, or ammonium salt.
7. The pharmaceutical formulation of claim 1 , wherein the citrate buffer comprises sodium citrate.
8. The pharmaceutical formulation of claim 1 , wherein the citrate buffer is present in a concentration of about 10 mM to about 100 mM.
9. The pharmaceutical formulation of claim 8 , wherein the citrate buffer is present in a concentration of about 25 mM.
10. The pharmaceutical formulation of claim 1 , wherein trehalose, sucrose, glycine, or combinations thereof is excluded.
11. The pharmaceutical formulation of claim 1 , wherein the at least one excipient is mannitol present in a concentration of about 10 mg/mL to about 50 mg/mL.
12. The pharmaceutical formulation of claim 1 , wherein the at least one excipient is polysorbate 80 present in a concentration of about 0.2 mg/mL to about 0.5 mg/mL.
13. The pharmaceutical formulation of claim 1 , wherein the at least one excipient comprises mannitol and polysorbate 80, and wherein the mannitol is present at a concentration of about 20 mg/mL and the polysorbate 80 is present at a concentration of about 0.5 mg/mL.
14. The pharmaceutical formulation of claim 1 , further comprising:
(a) an alkalizing agent; and/or
(b) an acidifying agent,
wherein the alkalizing agent and/or acidifying agent are present in amounts to maintain the pharmaceutical formulation at a pH of from about 5.0 to about 6.0.
15. The pharmaceutical formulation of claim 1 , wherein at least 75% of the rhGAA molecules comprise a glycan bearing bis-M6P at the first potential N-glycosylation site.
16. The pharmaceutical formulation of claim 1 , wherein the rhGAA molecules comprise from about 3.0 mol to about 5.0 mol mannose-6-phosphate (M6P) residues per mol rhGAA.
17. The pharmaceutical formulation of claim 1 , wherein the rhGAA molecules comprise at least 3 mol M6P residues per mole of and at least 4 of mol sialic acid residues per mole rhGAA.
18. The pharmaceutical formulation of claim 1 , wherein 75% to about 80% of the rhGAA molecules comprise a glycan bearing bis-M6P at the first potential N-glycosylation site, about 40% to about 60% of the rhGAA molecules comprise a glycan bearing mono-mannose-6-phosphate (mono-M6P) at the second potential N-glycosylation site, about 40% to about 60% of the rhGAA molecules comprise a glycan bearing bis-M6P at the fourth potential N-glycosylation site, and about 25% to about 40% of the rhGAA molecules comprise a glycan bearing mono-M6P at the fourth potential N-glycosylation site.
19. The pharmaceutical formulation of claim 1 , wherein the pharmaceutical formulation consists essentially of:
(a) the population of rhGAA molecules;
(b) sodium citrate;
(c) citric acid monohydrate;
(d) mannitol;
(e) polysorbate 80;
(f) water;
(g) optionally, an acidifying agent; and
(h) optionally, an alkalizing agent,
wherein the formulation has a pH of from about 5.0 to about 6.0.
20. The pharmaceutical formulation of claim 1 , wherein the pharmaceutical formulation consists essentially of:
(a) the population of rhGAA molecules, present at a concentration of about 15 mg/mL;
(b) sodium citrate buffer, present at a concentration of about 25 mM;
(c) mannitol, present at a concentration of about 20 mg/mL;
(d) polysorbate 80, present at a concentration of about 0.5 mg/mL; and
(e) water;
(f) optionally, an acidifying agent; and
(g) optionally, an alkalizing agent,
wherein the formulation has a pH of from about 5.0 to about 6.0.
21. A pharmaceutical composition comprising the formulation of claim 1 after lyophilization.
22. A method of treating Pompe disease comprising administering to a patient in need thereof the pharmaceutical formulation of claim 1 .
23. The method of claim 22 , further comprising diluting the pharmaceutical formulation prior to administration to the patient.