Patent Application
20150368317
This disclosure provides compositions and methods related to CTLA4-Fc fusion proteins (e.g., abatacept).
Abatacept (marketed under the trade name ORENCIA® in both the United States and Europe) is a genetically engineered fusion protein composed of a modified Fc region of the immunoglobulin IgG1 fused to the extracellular domain of cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4). Abatacept has an approximate molecular weight of 92,300 Daltons, as determined by mass spectrometry, and has a binding affinity for CD80 of approximately 6.5 nM and for CD86 of approximately 14 nM (see, Larsen et al., American Journal of Transplantation, 5:443-453 (2005)).
Abatacept is produced by mammalian cell (Chinese Hamster Ovary) culture in a nutrient medium. For intravenous administration, abatacept is supplied as a 250 mg lyophilized powder in single-use vial for reconstitution with 10 mL of sterile water. The reconstituted abatacept solution is further diluted to 100 mL by replacing a volume of 0.9% Sodium Chloride Injection, USP, with an equal volume of reconstituted abatacept solution, as required for a patient dose (see ORENCIA® Product Label). For subcutaneous administration, abatacept is supplied as a 125 mg/mL single-dose prefilled glass syringe.
The present disclosure provides, in part, methods for evaluating, identifying, and/or producing (e.g., manufacturing) a CTLA4-Fc fusion protein, e.g., abatacept. In some instances, methods herein allow highly resolved evaluation of abatacept useful for, inter alia, manufacturing abatacept, characterizing abatacept, identifying and/or confirming abatacept, monitoring the structure of abatacept, comparing abatacept preparations made over time or made under different conditions, and/or controlling the structure of abatacept.
In the first aspect, the invention features a method of manufacturing a pharmaceutical product. This method includes a CTLA4-Fc fusion protein, including: providing a sample of a test biologic preparation including a CTLA4-Fc fusion protein having an amino acid sequence with at least 95%, 98%, 99%, or 100% identity to SEQ ID NO:1, wherein amino acid 29 is alanine and amino acid 104 is leucine; acquiring an input value for each of one or more parameters in the test biologic preparation listed in Table 1; and processing the test biologic preparation into a pharmaceutical product including the CTLA4-Fc fusion protein if the input value(s) for the one or more parameters in the test biologic preparation listed in Table 1 meets the corresponding reference criterion for the parameter, thereby manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein.
In a further aspect, the invention features another method of manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein. This method includes: providing a host cell that is genetically engineered to express a CTLA4-Fc fusion protein having an amino acid sequence with at least about 95% identity to SEQ ID NO:1 (e.g., 95%, 98%, 99%, or 100% identity to SEQ ID NO:1), wherein amino acid 29 is alanine and amino acid 104 is leucine; culturing the host cell under conditions whereby the cell expresses the CTLA4-Fc fusion protein; harvesting the CTLA4-Fc fusion protein from the host cell culture to produce a test biologic preparation; acquiring a value for one or more parameters listed in Table 1 for the test biologic preparation; and processing at least a portion of the test biologic preparation into a pharmaceutical product including the CTLA4-Fc fusion protein if the input values for the one or more parameters in the test biologic preparation listed in Table 1 meets the corresponding reference criterion for the parameter, thereby manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein.
In another aspect, the invention features yet another method of manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein. This method includes: providing a host cell that is genetically engineered to express a CTLA4-Fc fusion protein having an amino acid sequence with 100% identity to SEQ ID NO: 1; culturing the host cell under conditions whereby the cell expresses the CTLA4-Fc fusion protein; harvesting the CTLA4-Fc fusion protein from the host cell culture to produce a test biologic preparation; and processing or directing the processing of at least a portion of the test biologic preparation as a pharmaceutical product including the CTLA4-Fc fusion protein if the input values for the one or more parameters in the test biologic preparation listed in Table 1 meets the reference criterion for the parameter, thereby manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein.
In another aspect, the invention features a method of manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein. This method includes: providing a sample of a test biologic preparation including a CTLA4-Fc fusion protein having an amino acid sequence with at least 95%, 98%, 99%, or 100% identity to SEQ ID NO:1, wherein amino acid 29 is alanine and amino acid 104 is leucine; acquiring an input value for each of one or more parameters in the test biologic preparation listed in Table 1; and processing the test biologic preparation into a pharmaceutical product including the CTLA4-Fc fusion protein if the input value(s) for at least one parameter in the test biologic preparation listed in Table 1 meets the corresponding reference criterion for the parameter, thereby manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein.
In a further aspect, the invention features a method of manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein. This method includes: providing a sample of a test biologic preparation including a CTLA4-Fc fusion protein having an amino acid sequence with at least 95%, 98%, 99%, or 100% identity to SEQ ID NO:1; confirming that amino acid 29 is alanine and amino acid 104 is leucine; acquiring an input value for each of one or more parameters in the test biologic preparation listed in Table 1; and processing the test biologic preparation into a pharmaceutical product including the a CTLA4-Fc fusion protein if the input value(s) for at least one parameter in the test biologic preparation listed in Table 1 meets the corresponding reference criterion for the parameter, thereby manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein.
In another aspect, the invention features a method of manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein. This method includes: providing a sample of a test biologic preparation including a CTLA4-Fc fusion protein having an amino acid sequence with at least 95%, 98%, 99%, or 100% identity to SEQ ID NO:1; confirming that amino acid 29 is not tyrosine and amino acid 104 is not glutamic acid; acquiring an input value for each of one or more parameters in the test biologic preparation listed in Table 1; and processing the test biologic preparation into a pharmaceutical product including the CTLA4-Fc fusion protein if the input value(s) for at least one parameter in the test biologic preparation listed in Table 1 meets the corresponding reference criterion for the parameter, thereby manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein.
In a further aspect, the invention features a method of manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein. This method includes: providing a sample of a test biologic preparation including a CTLA4-Fc fusion protein having an amino acid sequence with at least 95%, 98%, 99%, or 100% identity to SEQ ID NO:1; confirming that, in the CTLA4-Fc fusion protein, amino acid 29 is tyrosine and amino acid 104 is glutamic acid; acquiring an input value for each of one or more parameters in the test biologic preparation listed in Table 1; and processing the test biologic preparation into a pharmaceutical product including CTLA4-Fc fusion protein if the input value(s) for at least one parameter in the test biologic preparation listed in Table 1 meets the corresponding reference criterion for the parameter, thereby manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein.
In another aspect, the invention features a method of manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein. This method including: providing a sample of a test biologic preparation including a CTLA4-Fc fusion protein having an amino acid sequence with at least 95%, 98%, 99%, or 100% identity to SEQ ID NO:1; confirming that amino acid 29 is alanine and amino acid 104 is leucine; acquiring an input value for each of one or more parameters in the test biologic preparation listed in Table 1; and processing the test biologic preparation into a pharmaceutical product including the a CTLA4-Fc fusion protein if the input value(s) for the one or more parameters in the test biologic preparation listed in Table 1 meets the corresponding reference criterion for the parameter, thereby manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein.
In a further aspect, the invention features a method of manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein. This method includes: providing a sample of a test biologic preparation including a CTLA4-Fc fusion protein having an amino acid sequence with at least 95%, 98%, 99%, or 100% identity to SEQ ID NO:1; confirming that amino acid 29 is not tyrosine and amino acid 104 is not glutamic acid; acquiring an input value for each of one or more parameters in the test biologic preparation listed in Table 1; and processing the test biologic preparation into a pharmaceutical product including the CTLA4-Fc fusion protein if the input value(s) for the one or more parameters in the test biologic preparation listed in Table 1 meets the corresponding reference criterion for the parameter, thereby manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein.
In another aspect, the invention features a method of manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein. This method includes: providing a sample of a test biologic preparation including a CTLA4-Fc fusion protein having an amino acid sequence with at least 95%, 98%, 99%, or 100% identity to SEQ ID NO:1; confirming that, in the CTLA4-Fc fusion protein, amino acid 29 is tyrosine and amino acid 104 is glutamic acid; acquiring an input value for each of one or more parameters in the test biologic preparation listed in Table 1; and processing the test biologic preparation into a pharmaceutical product including CTLA4-Fc fusion protein if the input value(s) for the one or more parameters in the test biologic preparation listed in Table 1 meets the corresponding reference criterion for the parameter, thereby manufacturing a pharmaceutical product including a CTLA4-Fc fusion protein.
In certain embodiments, any of the foregoing methods also include after the step of acquiring the input value(s) and before the step of processing, acquiring one or more assessments made by comparing the input value with one or more target values for a target biologic having a first amino acid sequence with 100% identity to SEQ ID NO:1, wherein the target biologic is approved under a BLA or a supplemental BLA.
In other embodiments of any of the foregoing methods, the processed biologic is approved under Section 351(k) of the Public Health Service (PHS) Act.
In further embodiments of any of the foregoing methods, the processed biologic is not approved under a BLA under Section 351(a) of the PHS Act.
In further embodiments of any of the foregoing methods, the processed biologic is approved under a BLA under Section 351(a) of the PHS Act.
In some embodiments of any of the foregoing methods, the input value is acquired for one, two, or more samples or batches.
In still further embodiments of any of the foregoing methods, the value for the test biologic preparation comprises an average (e.g., mean) of a range of values for the parameter for multiple (e.g., 2, 3, 4, 5, 10, 15, 20, or more) batches or samples of the test biologic.
In some embodiments of any of the foregoing methods, one or more, including all, of the reference criteria shown in Table 1 is/are a specification for commercial release of a drug product under Section 351(k) of the PHS Act.
In another aspect, the invention features a method of manufacturing abatacept, including: providing or obtaining a test biologic preparation; acquiring an input value for one or more parameters listed in Table 1 for the test biologic preparation; and processing at least a portion of the test biologic preparation into a pharmaceutical product if the input values for the one or more parameters in the test biologic preparation listed in Table 1 meets the corresponding reference criterion for the parameter, thereby manufacturing abatacept.
In some embodiments, any of the foregoing methods include acquiring input values for any combination of two or more parameters listed in Table 1.
In a further embodiment, any of the foregoing methods include acquiring an input value for a plurality of parameters listed in Table 1.
In a still further embodiment, the test biologic preparation has an amino acid sequence with 100% identity to SEQ ID NO:1.
In yet other embodiments of any of the foregoing methods, the method includes, after the step of acquiring the value(s) and before the step of processing, obtaining a plurality of assessments made by comparing the value(s) with a corresponding reference criterion shown in Table 1.
In further embodiments of any of the foregoing methods, at least one value is directly obtained by performing an analytical test on the test biologic preparation.
In other embodiments of any of the foregoing methods, the value is directly obtained using a method provided in Table 7.
In still other embodiments of any of the foregoing methods, the processing step comprises combining the test biologic preparation with an excipient or buffer.
In some embodiments of any of the foregoing methods, the processing step includes one or more of: formulating the test biologic preparation; processing the test biologic preparation into a drug product; combining the test biologic preparation with a second component, e.g., an excipient or buffer; changing the concentration of the biologic in the preparation; lyophilizing the test biologic preparation; combining a first and second aliquot of the biologic to provide a third, larger, aliquot; dividing the test biologic preparation into smaller aliquots; disposing the test biologic preparation into a container, e.g., a gas or liquid tight container; packaging the test biologic preparation; associating a container including the test biologic preparation with a label (e.g., labeling); and shipping or moving the test biologic to a different location.
In another aspect, the disclosure features methods for high resolution and site-specific deconvolution of glycan profiles for glycoproteins having multiple (e.g., two or more) glycosylation sites. In some instances, methods includes the steps of: (a) providing a test biologic preparation that includes one or more N- and/or O-glycans; (b) digesting the test biologic preparation with one or more protease enzymes (e.g., including, but not limited to, Arg-C endoproteinase, trypsin, chymo-trypsin, Asp-N endopeptidase, Glu-C endoproteinase, Lys-C endoproteinase) in a digestion buffer that is compatible with mass spectrometry, wherein the digestion is optionally performed using pressure cycling technology under conditions sufficient to allow at least 75% digestion of the protein into constituent peptides in a time of under 5 hours; in some instances so as to minimize sample prep induced modifications of the amino acid backbone (e.g. deamidation, methionine oxidation); (c) separating the digested test biologic preparation to produce two or more separated components of the test biologic preparation; and (d) determining the identity, location, and/or abundance of one or more N- and/or O-linked glycans in one or more of the separated components.
In one embodiment, the test biologic preparation includes a biologic having an amino acid sequence with at least 95% identity (e.g., 95%, 96%, 97%, 98%, 99%, wherein amino acid 29 is alanine and amino acid 104 is leucine or 100% identity) to SEQ ID NO:1. In another embodiment, the test biologic preparation includes a biologic having an amino acid sequence with 100% identity to SEQ ID NO:1.
In one embodiment, the digesting and separating steps permit the separation of a first component containing Ser129 from a second component containing Ser139. In another embodiment, the digesting and separating steps permit the separation of the following into distinct components: a component containing O-linked site Ser129; a component containing Asn207; a component containing major non-glycopeptides; a component containing Ser139; a component containing Asn108; and a component containing Asn76.
In certain embodiments of any of the above methods, the one or more parameters in the test biologic preparation include parameter 1. In other embodiments, the one or more parameters in the test biologic preparation include parameter 2. In other embodiments, the one or more parameters in the test biologic preparation include parameter 3. In other embodiments, the one or more parameters in the test biologic preparation include parameter 4. In other embodiments, the one or more parameters in the test biologic preparation include parameter 5. In other embodiments, the one or more parameters in the test biologic preparation include parameter 6. In other embodiments, the one or more parameters in the test biologic preparation include parameter 7. In other embodiments, the one or more parameters in the test biologic preparation include parameter 8. In other embodiments, the one or more parameters in the test biologic preparation include parameter 9. In other embodiments, the one or more parameters in the test biologic preparation include parameter 10. In other embodiments, the one or more parameters in the test biologic preparation include parameter 11. In other embodiments, the one or more parameters in the test biologic preparation include parameter 12. In other embodiments, the one or more parameters in the test biologic preparation include parameter 13. In other embodiments, the one or more parameters in the test biologic preparation include parameter 14. In other embodiments, the one or more parameters in the test biologic preparation include parameter 15. In other embodiments, the one or more parameters in the test biologic preparation include parameter 16. In other embodiments, the one or more parameters in the test biologic preparation include parameter 17. In other embodiments, the one or more parameters in the test biologic preparation include parameter 18. In other embodiments, the one or more parameters in the test biologic preparation include parameter 19. In other embodiments, the one or more parameters in the test biologic preparation include parameter 20. In other embodiments, the one or more parameters in the test biologic preparation include parameter 21. In other embodiments, the one or more parameters in the test biologic preparation include parameter 22. In other embodiments, the one or more parameters in the test biologic preparation include parameter 23. In other embodiments, the one or more parameters in the test biologic preparation include parameter 24. In other embodiments, the one or more parameters in the test biologic preparation include parameter 25. In other embodiments, the one or more parameters in the test biologic preparation include parameter 26. In other embodiments, the one or more parameters in the test biologic preparation include parameter 27. In other embodiments, the one or more parameters in the test biologic preparation include parameter 28. In other embodiments, the one or more parameters in the test biologic preparation include parameter 29. In other embodiments, the one or more parameters in the test biologic preparation include parameter 30. In other embodiments, the one or more parameters in the test biologic preparation include parameter 31. In other embodiments, the one or more parameters in the test biologic preparation include parameter 32. In other embodiments, the one or more parameters in the test biologic preparation include parameter 33. In other embodiments, the one or more parameters in the test biologic preparation include parameter 34. In other embodiments, the one or more parameters in the test biologic preparation include parameter 35. In other embodiments, the one or more parameters in the test biologic preparation include parameter 36. In other embodiments, the one or more parameters in the test biologic preparation include parameter 37. In other embodiments, the one or more parameters in the test biologic preparation include parameter 38. In other embodiments, the one or more parameters in the test biologic preparation include parameter 39. In other embodiments, the one or more parameters in the test biologic preparation include parameter 40. In other embodiments, the one or more parameters in the test biologic preparation include parameter 41. In other embodiments, the one or more parameters in the test biologic preparation include parameter 42. In other embodiments, the one or more parameters in the test biologic preparation include parameter 43. In other embodiments, the one or more parameters in the test biologic preparation include parameter 44. In other embodiments, the one or more parameters in the test biologic preparation include parameter 45. In other embodiments, the one or more parameters in the test biologic preparation include parameter 46. In other embodiments, the one or more parameters in the test biologic preparation include parameter 47. In other embodiments, the one or more parameters in the test biologic preparation include parameter 48. In other embodiments, the one or more parameters in the test biologic preparation include parameter 49. In other embodiments, the one or more parameters in the test biologic preparation include parameter 50. In other embodiments, the one or more parameters in the test biologic preparation include parameter 51. In other embodiments, the one or more parameters in the test biologic preparation include parameter 52. In other embodiments, the one or more parameters in the test biologic preparation include parameter 53. In other embodiments, the one or more parameters in the test biologic preparation include parameter 54.
In other embodiments of any of the above methods, the one or more parameters in the test biologic preparation do not include parameter 1. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 2. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 3. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 4. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 5. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 6. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 7. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 8. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 9. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 10. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 11. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 12. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 13. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 14. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 15. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 16. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 17. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 18. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 19. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 20. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 21. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 22. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 23. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 24. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 25. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 26. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 27. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 28. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 29. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 30. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 31. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 32. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 33. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 34. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 35. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 36. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 37. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 38. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 39. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 40. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 41. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 42. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 43. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 44. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 45. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 46. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 47. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 48. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 49. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 50. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 51. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 52. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 53. In other embodiments, the one or more parameters in the test biologic preparation do not include parameter 54.
In still other embodiments of any of the above methods, the one or more parameters in the test biologic preparation include a parameter present at N108 (parameters 2, 7, 17, 20, 22, 23, 25, 33, 34, 37, and 40). In other embodiments, the one or more parameters in the test biologic preparation include a parameter present at N76 (parameters 1, 6, 16, 19, 21, 27, 32, 34, 36, and 39). In other embodiments, the one or more parameters in the test biologic preparation include a parameter present at N207 (parameters 5, 10, 15, 18, 24, 26, 29-31, and 38). In other embodiments, the one or more parameters in the test biologic preparation include a parameter at present O129 (parameters 3, 8, 41, and 42). In other embodiments, the one or more parameters in the test biologic preparation include a parameter at present O139 (parameters 4, 9, and 44-46). In other embodiments, the one or more parameters in the test biologic preparation include a parameter pertaining to the structure of one or more glycans (parameters 5-10, 15-46, and 52-54). In other embodiments, the one or more parameters in the test biologic preparation include a parameter pertaining to the location of one or more glycans (parameters 1-10 and 15-46). In other embodiments, the one or more parameters in the test biologic preparation include a parameter pertaining to the abundance of one or more glycans (parameters 15-46, and 52-54). In other embodiments, the one or more parameters in the test biologic preparation include a parameter pertaining to the amount of lysine at the C-terminus (parameter 11). In other embodiments, the one or more parameters in the test biologic preparation include a parameter pertaining to the charge distribution of the glycans (parameters 49-51). In other embodiments, the one or more parameters in the test biologic preparation include a parameter pertaining to the number of different glycans (parameters 1-5). In other embodiments, the one or more parameters in the test biologic preparation include a parameter pertaining to the amount of post-translational glycation (parameters 12-14). In other embodiments, the one or more parameters in the test biologic preparation include a parameter pertaining to the abundance of NeuGC (parameters 15-20). In other embodiments, the one or more parameters in the test biologic preparation include a parameter pertaining to the amount of post-translational sulfation (parameters 21-22). In other embodiments, the one or more parameters in the test biologic preparation include a parameter pertaining to the amount of post-translational acylation (parameters 23-27). In certain embodiments, the one or more parameters in the test biologic preparation include a parameter pertaining to the structure, location, or abundance of a glycan (parameters 1-10, 15-46, and 52-54).
In still other embodiments of any of the above methods, the one or more parameters in the test biologic preparation do not include a parameter present at N108 (parameters 2, 7, 17, 20, 22, 23, 25, 33, 34, 37, and 40). In other embodiments, the one or more parameters in the test biologic preparation do not include a parameter present at N76 (parameters 1, 6, 16, 19, 21, 27, 32, 34, 36, and 39). In other embodiments, the one or more parameters in the test biologic preparation do not include a parameter present at N207 (parameters 5, 10, 15, 18, 24, 26, 29-31, and 38). In other embodiments, the one or more parameters in the test biologic preparation do not include a parameter at present O129 (parameters 3, 8, 41, and 42). In other embodiments, the one or more parameters in the test biologic preparation do not include a parameter at present O139 (parameters 4, 9, and 44-46). In other embodiments, the one or more parameters in the test biologic preparation do not include a parameter pertaining to the structure of one or more glycans (parameters 5-10, 15-46, and 52-54). In other embodiments, the one or more parameters in the test biologic preparation do not include a parameter pertaining to the location of one or more glycans (parameters 1-10 and 15-46). In other embodiments, the one or more parameters in the test biologic preparation do not include a parameter pertaining to the abundance of one or more glycans (parameters 15-46, and 52-54). In other embodiments, the one or more parameters in the test biologic preparation do not include a parameter pertaining to the amount of lysine at the C-terminus (parameter 11). In other embodiments, the one or more parameters in the test biologic preparation do not include a parameter pertaining to the charge distribution of the glycans (parameters 49-51). In other embodiments, the one or more parameters in the test biologic preparation do not include a parameter pertaining to the number of different glycans (parameters 1-5). In other embodiments, the one or more parameters in the test biologic preparation do not include a parameter pertaining to the amount of post-translational glycation (parameters 12-14). In other embodiments, the one or more parameters in the test biologic preparation do not include a parameter pertaining to the abundance of NeuGC (parameters 15-20). In other embodiments, the one or more parameters in the test biologic preparation do not include a parameter pertaining to the amount of post-translational sulfation (parameters 21-22). In other embodiments, the one or more parameters in the test biologic preparation do not include a parameter pertaining to the amount of post-translational acylation (parameters 23-27). In certain embodiments, the one or more parameters in the test biologic preparation do not include a parameter pertaining to the structure, location, or abundance of a glycan (parameters 1-10, 15-46, and 52-54).
In certain embodiments, the combination of two or more parameters in the test biologic preparation include a parameter pertaining to the structure, location, or abundance of a glycan (parameters 1-10, 12-46, and 52-54) and a parameter pertaining to the amount of lysine at the C-terminus (parameter 11). In certain embodiments, the combination of two or more parameters in the test biologic preparation include a parameter pertaining to the structure, location, or abundance of a glycan (parameters 1-10, 12-46, and 52-54) and a parameter pertaining to the charge distribution of the glycans (parameters 49-51). In certain embodiments, the combination of two or more parameters in the test biologic preparation include a parameter pertaining to the structure, location, or abundance of a glycan (parameters 1-10, 12-46, and 52-54) and a parameter pertaining to the amount of post-translational glycation (parameters 12-14). In certain embodiments, the combination of two or more parameters in the test biologic preparation include a parameter pertaining to the charge distribution of the glycans (parameters 47-51) and a parameter pertaining to the amount of lysine at the C-terminus (parameter 11). In certain embodiments, the combination of two or more parameters in the test biologic preparation include a parameter pertaining to the charge distribution of the glycans (parameters 47-51) and a parameter pertaining to the amount of post-translational glycation (parameters 12-14). In certain embodiments, the combination of two or more parameters in the test biologic preparation include a parameter pertaining to the amount of lysine at the C-terminus (parameter 11) and a parameter pertaining to the amount of post-translational glycation (parameters 12-14). In certain embodiments, the combination of two or more parameters in the test biologic preparation include a parameter pertaining to the structure, location, or abundance of a glycan (parameters 1-10, 12-46, and 52-54), a parameter pertaining to the charge distribution of the glycans (parameters 47-51), and a parameter pertaining to the amount of lysine at the C-terminus (parameter 11). In certain embodiments, the combination of two or more parameters in the test biologic preparation include a parameter pertaining to the structure, location, or abundance of a glycan (parameters 1-10, 12-46, and 52-54), a parameter pertaining to the charge distribution of the glycans (parameters 47-51), and a parameter pertaining to the amount of post-translational glycation (parameters 12-14). In certain embodiments, the combination of two or more parameters in the test biologic preparation include a parameter pertaining to the structure, location, or abundance of a glycan (parameters 1-10, 12-46, and 52-54), a parameter pertaining to the amount of post-translational glycation (parameters 12-14), and a parameter pertaining to the amount of lysine at the C-terminus (parameter 11). In certain embodiments, the combination of two or more parameters in the test biologic preparation include a parameter pertaining to the charge distribution of the glycans (parameters 47-51), a parameter pertaining to the amount of post-translational glycation (parameters 12-14), and a parameter pertaining to the amount of lysine at the C-terminus (parameter 11). In certain embodiments, the combination of two or more parameters in the test biologic preparation include a parameter pertaining to the structure, location, or abundance of a glycan (parameters 1-10, 12-46, and 52-54), a parameter pertaining to the charge distribution of the glycans (parameters 47-51), a parameter pertaining to the amount of post-translational glycation (parameters 12-14), and a parameter pertaining to the amount of lysine at the C-terminus (parameter 11).
In still other embodiments of any of the above methods, a combination of two parameters are compared to the reference criteria. In other embodiments, a combination of three parameters are compared to the reference criteria. In other embodiments, a combination of four parameters are compared to the reference criteria. In other embodiments a combination of five parameters are compared to the reference criteria. In other embodiments, a combination of six parameters are compared to the reference criteria. In other embodiments, a combination of seven parameters are compared to the reference criteria. In other embodiments, a combination of eight parameters are compared to the reference criteria. In other embodiments, a combination of nine parameters are compared to the reference criteria. In other embodiments, a combination of ten parameters are compared to the reference criteria. In other embodiments, a combination of 11 parameters are compared to the reference criteria. In other embodiments, a combination of 12 parameters are compared to the reference criteria. In other embodiments, a combination of 13 parameters are compared to the reference criteria. In other embodiments, a combination of 14 parameters are compared to the reference criteria. In other embodiments, a combination of 15 parameters are compared to the reference criteria. In other embodiments, a combination of 16 parameters are compared to the reference criteria. In other embodiments, a combination of 17 parameters are compared to the reference criteria. In other embodiments, a combination of 18 parameters are compared to the reference criteria. In other embodiments, a combination of 19 parameters are compared to the reference criteria. In other embodiments, a combination of 20 parameters are compared to the reference criteria. In other embodiments, a combination of 21 parameters are compared to the reference criteria. In other embodiments, a combination of 22 parameters are compared to the reference criteria. In other embodiments, a combination of 23 parameters are compared to the reference criteria. In other embodiments, a combination of 24 parameters are compared to the reference criteria. In other embodiments, a combination of 25 parameters are compared to the reference criteria. In other embodiments, a combination of 26 parameters are compared to the reference criteria. In other embodiments, a combination of 27 parameters are compared to the reference criteria. In other embodiments, a combination of 28 parameters are compared to the reference criteria. In other embodiments, a combination of 29 parameters are compared to the reference criteria. In other embodiments, a combination of 30 parameters are compared to the reference criteria. In other embodiments, a combination of 31 parameters are compared to the reference criteria. In other embodiments, a combination of 32 parameters are compared to the reference criteria. In other embodiments, a combination of 33 parameters are compared to the reference criteria. In other embodiments, a combination of 34 parameters are compared to the reference criteria. In other embodiments, a combination of 35 parameters are compared to the reference criteria. In other embodiments, a combination of 36 parameters are compared to the reference criteria. In other embodiments, a combination of 37 parameters are compared to the reference criteria. In other embodiments, a combination of 38 parameters are compared to the reference criteria. In other embodiments, a combination of 39 parameters are compared to the reference criteria. In other embodiments, a combination of 40 parameters are compared to the reference criteria. In other embodiments, a combination of 41 parameters are compared to the reference criteria. In other embodiments, a combination of 42 parameters are compared to the reference criteria. In other embodiments, a combination of 43 parameters are compared to the reference criteria. In other embodiments, a combination of 44 parameters are compared to the reference criteria. In other embodiments, a combination of 45 parameters are compared to the reference criteria. In other embodiments, a combination of 46 parameters are compared to the reference criteria. In other embodiments, a combination of 47 parameters are compared to the reference criteria. In other embodiments, a combination of 48 parameters are compared to the reference criteria. In other embodiments, a combination of 49 parameters are compared to the reference criteria. In other embodiments, a combination of 50 parameters are compared to the reference criteria. In other embodiments, a combination of 51 parameters are compared to the reference criteria. In other embodiments, a combination of 52 parameters are compared to the reference criteria. In other embodiments, a combination of 53 parameters are compared to the reference criteria. In other embodiments, a combination of 54 parameters are compared to the reference criteria.
As used herein, an abatacept signature or fingerprint comprises a plurality of reference criteria or rules for a plurality of parameters that define abatacept. In some instances, an abatacept signature can be contained in or part of a pharmaceutical specification, a commercial product release specification, a product acceptance criterion, a pharmacopeial standard, or a product labeling description. In some instances, the abatacept signature comprises a plurality of reference criteria or rules for a plurality of parameters shown in Table 1:
For parameters 15-27, 30-40, 42, and 44-46, percent (%) refers to the number of moles of peptide containing glycan X relative to total moles of that peptide (with or without a glycan) detected as disclosed herein, wherein X represents the parameter of interest.
For parameter 11, percent (%) refers to the level of C-terminal-lysine-containing peptide relative to the sum of the levels of C-terminal-lysine-containing and C-terminal-lysine-free peptides detected as disclosed in Table 7.
For parameters 12-14, percent (%) refers to the level of glycated peptide relative to the sum of the glycated peptide and unmodified peptides detected as disclosed in Table 7.
For parameters 28-29, 41, and 43, percent (%) refers to the level of Aglyco peptide Y relative to the sum of the levels of Aglyco peptide Y and glycan-containing peptide Y, detected as disclosed in Table 7, wherein Y represents the parameter of interest.
For parameters 47-51, percent (%) refers to the number of moles of PNGase F-released glycan(s) X relative to total moles of PNGase F-released glycans detected as disclosed in Table 7, wherein X represents the parameter of interest.
For parameter 52, percent (%) refers to the number of moles of sialic acid that is NeuGc relative to total moles of sialic acid, detected as disclosed in Table 7.
For parameter 53, percent (%) refers to the number of moles of sialic acid residues containing two or more acetyle groups relative to total moles of sialic acid residues, detected as disclosed in Table 7.
For parameter 54, percent (%) refers to the number of moles of PNGase F-released glycans that contain one or more sulfate moieties relative to total moles of PNGase F-released glycans detected as disclosed in Table 7.
By “detectable” is meant that the recited parameter can be detected by one of ordinary skill using a method disclosed in Table 7 or as disclosed herein.
While the present disclosure provides exemplary units and methods for the evaluation, identification, and production methods disclosed herein (see, e.g., Tables 1-7), a person of ordinary skill in the art will appreciate that performance of the evaluation, identification, and production methods herein is not limited to use of those units and/or methods. For example, abatacept signatures provided herein are generally described, for certain parameters, as a value for a glycan or structure relative to total glycan or structure on a mol/mol basis (see, e.g., Table 1). A person of skill in the art understands that although the use of other metrics or units (e.g., mass/mass, mole percent vs. weight percent) to measure a described parameter might give rise to different absolute values than those described herein, e.g., in Table 1, a test biologic preparation meets a disclosed abatacept reference criterion or signature even if other units or metrics are used, as long as the test biologic preparation meets the herein disclosed reference criterion or signature when the herein disclosed units and metrics are used, e.g., allowing for the sensitivity (e.g., analytical variability) of the method being used to measure the value.
Abatacept parameters shown in Table 1 are parameters that, alone, in any combination, or together, distinguish abatacept from non-abatacept biologic (see below). In some instances, an abatacept parameter is part of the biologic, e.g., connected with the rest of the biologic by a covalent bond, i.e., an intrinsic parameter. Intrinsic parameters include the presence, absence, level, ratio (with another entity), or distribution of a physical moiety, e.g., a moiety arising from or associated with a post-translational event. Parameters of this type include the presence (or absence), abundance, absolute or relative amount, ratio (with another entity), or distribution of a glycan, a linkage, a glycoform, or post-translationally added components of the preparation. In some instances, a parameter is not part of the biologic but is present in the preparation with the biologic (i.e., in a biologic preparation), i.e., an extrinsic parameter. Exemplary parameters of this type include the presence (or absence), abundance, ratio (with another entity), or distribution of, e.g., impurities, e.g., host cell proteins, residue from purification processes, viral impurities, and enclosure components.
In some instances, an abatacept signature includes reference criteria or rules for 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13,14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, or 54 parameters shown in Table 1. In some instances, an abatacept signature comprises reference criteria or rules for two or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13,14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53 or 54) of abatacept parameter(s) 1 to 54. In some instances, an abatacept signature comprises predetermined reference criteria or rule(s) for 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13,14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, or 54 parameters shown in Table 1. In some instances, an abatacept signature comprises reference criteria or rules for one or more, including any combination, or all, of parameter numbers 1 to 54.
In some instances, methods (i.e., evaluation, identification, and production methods) can further include, e.g., one or more of: providing or obtaining a biologic preparation (e.g., such as a glycoprotein drug substance or a precursor thereof); memorializing confirmation or identification of the biologic preparation as abatacept using a recordable medium (e.g., on paper or in a computer readable medium, e.g., in a Certificate of Testing, Certificate of Analysis, Material Safety Data Sheet (MSDS), batch record, or Certificate of Analysis (CofA)); informing a party or entity (e.g., a contractual or manufacturing partner, a care giver or other end-user, a regulatory entity, e.g., the FDA or other U.S., European, Japanese, Chinese or other governmental agency, or another entity, e.g., a compendial entity (e.g., U.S. Pharmacopoeia (USP)) or insurance company) that a biologic preparation is abatacept; selecting the biologic preparation for further processing (e.g., processing (e.g., formulating) the biologic preparation as a drug product (e.g., a pharmaceutical product) if the biologic preparation is identified as abatacept; reprocessing or disposing of the biologic preparation if the biologic preparation is not identified as abatacept.
In some instances, methods (i.e., evaluation, identification, and production methods) include taking action (e.g., physical action) in response to the methods disclosed herein. For example, the biologic preparation is classified, selected, accepted or discarded, released or withheld, processed into a drug product, shipped, moved to a different location, formulated, labeled, packaged, released into commerce, or sold or offered for sale, depending on whether the preselected relationship is met.
In some instances, processing may include formulating, packaging (e.g., in a syringe or vial), labeling, or shipping at least a portion of the glycoprotein preparation. In some instances, processing includes formulating, packaging (e.g., in a syringe or vial), and labeling at least a portion of the biologic as abatacept drug product. Processing can include directing and/or contracting another party to process as described herein.
As used herein, a biologic refers to amino acid sequences that include one or more oligosaccharide chains (e.g., glycans) covalently attached thereto. Exemplary amino acid sequences include peptides, polypeptides and proteins. Exemplary biologics include glycosylated antibodies and antibody-like molecules (e.g., Fc fusion proteins). Exemplary antibodies include monoclonal antibodies and/or fragments thereof, polyclonal antibodies and/or fragments thereof, and Fc domain containing fusion proteins (e.g., fusion proteins containing the Fc region of IgG1, or a glycosylated portion thereof). A biologic preparation is a composition or mixture that includes at least one biologic.
As used herein, a CTLA4-Fc fusion protein refers to a genetically engineered fusion protein composed of a modified Fc region of the immunoglobulin IgG1 fused to the extracellular domain of cytotoxic T-lymphocyte-associated antigen 4 (CTLA4) that has at least 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1.
A biologic preparation (e.g., such as a biologic drug substance or a precursor thereof) included herein is or includes a biologic (e.g., a recombinant fusion protein) that has an amino acid sequence with at least 85% identity to SEQ ID NO:1. In certain embodiments of the biologic, amino acid 29 is alanine and amino acid 104 is leucine. In other embodiments of the biologic, amino acid 29 is not alanine and amino acid 104 is not leucine. In other embodiments of the biologic, amino acid 29 is tyrosine and amino acid 104 is glutamic acid. In other embodiments of the biologic, amino acid 29 is not tyrosine and amino acid 104 is not glutamic acid. In some instances, the amino acid sequence has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identity to SEQ ID NO:1.
In some instances, a biologic preparation (e.g., such as a biologic drug substance or a precursor thereof) can be a sample from a proposed or test batch of a drug substance or drug product. As used herein, a batch of a biologic preparation refers to a single production run of the biologic. Evaluation of different batches thus means evaluation of different production runs or batches. As used herein sample(s) refer to separately procured samples. For example, evaluation of separate samples could mean evaluation of different commercially available containers or vials of the same batch or from different batches. A batch can include a drug product or drug substance.
As used herein, abatacept is the generic, compendial, nonproprietary, or official FDA name for the product marketed in the United States as ORENCIA®.
As used herein, evaluating, e.g., in the evaluation/evaluating, identifying, and/or producing aspects disclosed herein means reviewing, considering, determining, assessing, analyzing, measuring, and/or detecting the presence, absence, level, and/or ratio of one or more abatacept-specific parameters in a biologic preparation to provide information pertaining to the one or more abatacept-specific parameters. In some instances, evaluating can include performing a process that involves a physical change in a sample or another substance, e.g., a starting material. Exemplary changes include making a physical entity from two or more starting materials, shearing or fragmenting a substance, separating or purifying a substance, combining two or more separate entities into a mixture, performing a chemical reaction that includes breaking or forming a covalent or non-covalent bond. Evaluating can include performing an analytical process which includes a physical change in a substance, e.g., a sample, analyte, or reagent (sometimes referred to herein as “physical analysis”), performing an analytical method, e.g., a method which includes one or more of the following: separating or purifying a substance, e.g., an analyte, or a fragment or other derivative thereof, from another substance; combining an analyte, or fragment or other derivative thereof, with another substance, e.g., a buffer, solvent, or reactant; or changing the structure of an analyte, or a fragment or other derivative thereof, e.g., by breaking or forming a covalent or non-covalent bond, between a first and a second atom of the analyte; or by changing the structure of a reagent, or a fragment or other derivative thereof, e.g., by breaking or forming a covalent or non-covalent bond, between a first and a second atom of the reagent. In some instances, evaluating a biologic preparation includes detecting the presence, absence, level, or ratio of one or more (e.g., two or more when working with ratios) disclosed in Table 1 using methods disclosed in Table 7.
Information (e.g., value(s)) pertaining to an abatacept-specific parameter or an abatacept parameter means information, regardless of form, that describes the presence, absence, abundance, absolute or relative amount, ratio (with another entity), or distribution of a moiety associated with the biologic preparation and/or abatacept. Information is evaluated in a biologic preparation as disclosed herein. Information is also conveyed in an abatacept signature. Information can be qualitative, e.g., present, absent, intermediate, or quantitative, e.g., a numerical value such as a single number, or a range, for a parameter. In some instances, information is from a single sample or batch or a plurality of samples or batches. In some instances, information can be a range or average (or other measure of central tendency), e.g., based on the values from any X samples or batches, e.g., wherein at least X of the samples or batches is being evaluated for commercial release, wherein X is equal to, at least, or no more than, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, or 24. In some instances, information can be, for example: a statistical function, e.g., an average, of a number of values; a function of another value, e.g., of the presence, distribution or amount of a second entity present in the sample, e.g., an internal standard; a statistical function, e.g., an average, of a number of values; a function of another value, e.g., of the presence, distribution or amount of a second entity present in the sample, e.g., an internal standard; a value, e.g., a qualitative value, e.g., present, absent, “below limit of detection,” “within normal limits,” or intermediate. In some instances, information can be a quantitative value, e.g., a numerical value such as a single number, a range of values, a “no less than x amount” value, a “no more than x amount” value. In some instances, information can be abundance. Abundance can be expressed in relative terms, e.g., abundance can be expressed in terms of the abundance of a structure in relation to another component in the preparation. E.g., abundance can be expressed as: the abundance of a structure (or a first group of structures) in Table 1 relative to the amount of protein; the abundance of a structure (or a first group of structures) in Table 1 relative to the abundance of a second structure (or second group of structures) in Table 1. Abundance, e.g., abundance of a first structure relative to another structure, can be with regard to the preparation as a whole, a single molecule, or a selected site on the protein backbone. E.g., the parameter can be the relative proportion of a first structure from Table 1 and a second structure from Table 1 at a selected site and the value can be expressed as, e.g., a proportion, ratio or percentage. Information can be expressed in any useful term or unit, e.g., in terms of weight/weight, number/number, number/weight, and weight/number. In many cases, the reference criterion is defined by a range of values.
As used herein, acquire or acquiring (e.g., acquiring information) means obtaining possession of a physical entity, or a value, e.g., a numerical value, by “directly acquiring” or “indirectly acquiring” the physical entity or value. Directly acquiring means performing a process (e.g., performing an assay or test on a sample or “analyzing a sample” as that term is defined herein) to obtain the physical entity or value. Indirectly acquiring refers to receiving the physical entity or value from another party or source (e.g., a third party laboratory that directly acquired the physical entity or value). Directly acquiring a physical entity includes performing a process, e.g., analyzing a sample, that includes a physical change in a physical substance, e.g., a starting material. Exemplary changes include making a physical entity from two or more starting materials, shearing or fragmenting a substance, separating or purifying a substance, combining two or more separate entities into a mixture, performing a chemical reaction that includes breaking or forming a covalent or non-covalent bond. Directly acquiring a value includes performing a process that includes a physical change in a sample or another substance, e.g., performing an analytical process which includes a physical change in a substance, e.g., a sample, analyte, or reagent (sometimes referred to herein as “physical analysis”), performing an analytical method, e.g., a method which includes one or more of the following: separating or purifying a substance, e.g., an analyte, or a fragment or other derivative thereof, from another substance; combining an analyte, or fragment or other derivative thereof, with another substance, e.g., a buffer, solvent, or reactant; or changing the structure of an analyte, or a fragment or other derivative thereof, e.g., by breaking or forming a covalent or non-covalent bond, between a first and a second atom of the analyte; or by changing the structure of a reagent, or a fragment or other derivative thereof, e.g., by breaking or forming a covalent or non-covalent bond, between a first and a second atom of the reagent. Exemplary analytical methods are shown in Table 7.
All literature and similar material cited in this application, including, but not limited to, patents, patent applications, articles, books, treatises, and web pages, regardless of the format of such literature and similar materials, are expressly incorporated by reference in their entirety. In the event that one or more of the incorporated literature and similar materials differs from or contradicts this application, including but not limited to defined terms, term usage, described techniques, or the like, this application controls. The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described in any way.
These and other aspects of the invention, are described in more detail below and in the claims.
Detailed, high resolution, structural information about ORENCIA® (e.g., related to the presence of signature glycan species or quantitative analyses ascribing site-specificity for backbone modifications) is useful to be able to make and test products that qualify as abatacept, e.g., that are interchangeable versions of ORENCIA®. Such information is also useful in monitoring product changes and controlling structural drift that may occur as a result of manufacturing changes. The art supports, however, that information necessary to be able to make and test products that qualify as abatacept, e.g., that are interchangeable versions of ORENCIA®, or any other branded biologic, is unavailable (see, e.g., Nowicki, “Basic Facts about Biosimilars,” Kidney Blood Press. Res., 30:267-272 (2007); Hincal “An Introduction To Safety Issues In Biosimilars/Follow-On Biopharmaceuticals”, J. Med. CBR Def., 7:1-18, (2009); Roger, “Biosimilars: current status and future directions,” Expert Opin. Biol. Ther., 10(7):1011-1018 (2010); Schellekens et al., Nat. Biotechnol. 28:28-31 (2010); Sekhon et al., Biosimilars, 1:1-11 (2011)). One exemplary report states that “[t]he size and complexity of . . . therapeutic proteins make the production of an exact replica almost impossible; therefore, there are no true generic forms of these proteins . . . Verification of the similarity of biosimilars to innovator medicines remains a key challenge” (Hincal, supra). This disclosure provides, in part, methods and compositions sufficient to make and test products that qualify as abatacept, e.g., that are interchangeable versions of ORENCIA®.
Biologic preparations useful herein can be obtained from any source. In some instances, providing or obtaining a biologic preparation (e.g., such as a biologic drug substance or a precursor thereof), e.g., that is or includes a biologic, can include providing a host cell, e.g., a mammalian host cell (e.g., a CHO cell) that is genetically engineered to express a biologic having an amino acid sequence at least 85% identical to SEQ ID NO:1 (e.g., a genetically engineered cell), wherein amino acid 29 is alanine and amino acid 104 is leucine; culturing the host cell under conditions suitable to express the biologic (e.g., mRNA and/or protein); and, optionally, purifying the expressed biologic, e.g., in the form of a recombinant fusion protein) from the cultured cell, thereby producing a biologic preparation. In some instances, the host cell is genetically engineered to express a biologic having an amino acid sequence at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO:1, wherein the expressed protein forms a recombinant fusion protein composition.
As used herein percent (%) sequence identity with respect to a sequence is defined as the percentage of amino acid residues or nucleotides in a candidate sequence that are identical with the amino acid residues or nucleotides in the reference sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity. (E.g., gaps can be introduced in one or both of a first and a second amino acid or nucleic acid sequence for optimal alignment and non-homologous sequences can be disregarded for comparison purposes). Alignment for purposes of determining percent sequence identity can be achieved in various ways that are within the skill in the art, for instance, using publicly available computer software such as BLAST, ALIGN or Megalign (DNASTAR) software. Those skilled in the art can determine appropriate parameters for measuring alignment, including any algorithms needed to achieve maximal alignment over the full length of the sequences being compared. In one embodiment, the length of a reference sequence aligned for comparison purposes is at least 30%, e.g., at least 40%, e.g., at least 50%, 60%, 70%, 80%, 90%, or 100% of the length of the reference sequence. The amino acid residues or nucleotides at corresponding amino acid positions or nucleotide positions are then compared. When a position in the first sequence is occupied by the same amino acid residue or nucleotide as the corresponding position in the second sequence, then the molecules are identical at that position. In some instances a product will include amino acid variants, e.g., species that differ at terminal residues, e.g., at one, two, three, or four N-terminal residues and/or one C-terminal residue (see Table 1). In instances of such cases the sequence identity which is compared is the identity between the primary amino acid sequences of the most abundant active species in each of the products being compared. In some instances sequence identity refers to the amino acid sequence encoded by a nucleic acid that can be used to make the product.
In some instances, an abatacept signature disclosed herein can include 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13,14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, or 54 of the abatacept parameters (e.g., the reference criterion therefore) shown in Table 1 (e.g., including any combination of two or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13,14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, or 54) of parameter numbers 1 to 54 shown in Table 1).
In some instances, an abatacept signature disclosed herein can include structures or characteristics (whether intrinsic or extrinsic) of abatacept, e.g., that distinguish abatacept from non-abatacept glycoprotein. Examples of structures or characteristics include: the amount of GalNAc in the preparation (e.g., relative to total glycans of the preparation); the amount of truncated core glycans; the amount of aglycosylated glycans; the amount of each species of high mannose glycans; the amount of sialylated glycans or particular species of sialylated glycans; the ratio of monosialylated:disialylated glycans, the amount of diacetylated sialic acids (NeuXAc2), and the amount of one or more of: NeuGc; NeuAc; Neu5,7,Ac2; Neu5Gc,9Ac; Neu5,8Ac2; Neu5,9Ac2; Neu4,5Ac2. Examples of parameters related to the glycan linkage composition of a biologic preparation can be: the presence or amount of one or more of terminal fucose; terminal mannose; terminal galactose; 2 linked mannose; 3,6 linked mannose; terminal GlcNAc; terminal GalNAc; 4 linked GlcNAc; 4,6 linked GlcNAc. A parameter may also be the ratio of one of these to another or to another property. Examples of parameters related to the glycoform composition of a biologic preparation include: the absence or presence of one or more specific glycoforms (e.g., one or more glycoforms described in Table 1); the amount or abundance of a specific glycoform in the preparation relative to total glycoforms (e.g., in a w/w basis); the ratio of one particular glycoform to another. Examples of parameters related to post-translational modification in the preparation include: the absence or presence of one or more specific post-translational modification; the abundance or distribution of one or more specific post-translational modification.
In some instances, the present disclosure includes determining whether information evaluated for a biologic preparation meets an abatacept signature, e.g., by comparing the information with the abatacept signature and/or confirming that the information has a defined (e.g., predefined) relationship with the abatacept signature.
In some instances, methods disclosed herein can be used to confirm the identity and/or quality of abatacept preparations. For example, methods can include assessing preparations (e.g., samples, lots, and/or batches) of a test biologic to confirm whether the test biologic qualifies as abatacept, and, optionally, qualifying the test biologic as abatacept if qualifying criteria (e.g. predefined qualifying criteria) are met; thereby evaluating, identifying, and/or producing (e.g., manufacturing) abatacept.
Methods of the disclosure have a variety of applications and include, e.g., quality control at different stages of manufacture, analysis of abatacept preparations prior to or after completion of manufacture (e.g., prior to or after distribution to a fill/finish environment or facility), prior to or after release into commerce (e.g., before distribution to a pharmacy, a caregiver, a patient, or other end-user). Thus, the preparation can be any preparation that potentially comprises abatacept. In an embodiment the abatacept preparation is a drug substance (an active pharmaceutical ingredient or “API”) or a drug product (an API formulated for use in a subject such as a human patient). In an embodiment the preparation is from a stage of manufacture or use that is prior to release to care givers or other end-users; prior to packaging into individual dosage forms, such as syringes, pens, vials, or multi-dose vials; prior to determination that the batch can be commercially released, prior to production of a Certificate of Testing, Material Safety Data Sheet (MSDS) or Certificate of Analysis (CofA) of the preparation. In an embodiment, the biologic preparation is from an intermediate step in production, e.g., it is after secretion of the biologic from a cell but prior to purification of drug substance.
Evaluations from methods of the invention are useful for guiding, controlling, or implementing a number of activities or steps in the process of making, distributing, and monitoring and providing for the safe and efficacious use of abatacept. Thus, in an embodiment, e.g., responsive to the evaluation, e.g., depending on whether a criterion is met, a decision or step is taken. The method can further include one or both of the decision to take the step and/or carrying out the step itself. E.g., the step can include one in which the preparation (or another preparation for which the preparation is representative) is: classified; selected; accepted or discarded; released or processed into a drug product; rendered unusable for commercial release, e.g., by labeling it, sequestering it, or destroying it; passed on to a subsequent step in manufacture; reprocessed (e.g., the preparation may undergo a repetition of a previous process step or subjected to a corrective process); formulated, e.g., into drug substance or drug product; combined with another component, e.g., an excipient, buffer or diluent; disposed into a container; divided into smaller aliquots, e.g., unit doses, or multi-dose containers; combined with another preparation of abatacept; packaged; shipped; moved to a different location; combined with another element to form a kit; combined, e.g., placed into a package with a delivery device, diluent, or package insert; released into commerce; sold or offered for sale; delivered to a care giver or other end-user; or administered to a subject. E.g., based on the result of the determination or whether one or more subject entities is present, or upon comparison to a reference standard, the batch from which the preparation is taken can be processed, e.g., as just described.
Methods described herein may include making a decision: (a) as to whether a preparation may be formulated into drug substance or drug product; (b) as to whether a preparation may be reprocessed (e.g., the preparation may undergo a repetition of a previous process step); or (c) that the preparation is not suitable for formulation into drug substance or drug product. In instances the method comprises: formulating as referred to in step (a), reprocessing as referred to in step (b), or rendering the preparation unusable for commercial release, e.g., by labeling it or destroying it, as referred to in step (c).
Parameter Evaluation
The amino acid sequence of abatacept (ORENCIA®) is disclosed herein as SEQ ID NO:1.
Parameters disclosed herein can be analyzed by any available suitable method. In some instances, glycan structure and composition as described herein are analyzed, for example, by one or more, enzymatic, chromatographic, mass spectrometry (MS), chromatographic followed by MS, electrophoretic methods, electrophoretic methods followed by MS, nuclear magnetic resonance (NMR) methods, and combinations thereof. Exemplary enzymatic methods include contacting a glycoprotein preparation with one or more enzymes under conditions and for a time sufficient to release one or more glycans (e.g., one or more exposed glycans). In some instances, the one or more enzymes include PNGase F. Exemplary chromatographic methods include, but are not limited to, Strong Anion Exchange chromatography using Pulsed Amperometric Detection (SAX-PAD), liquid chromatography (LC), high performance liquid chromatography (HPLC), ultra performance liquid chromatography (UPLC), thin layer chromatography (TLC), amide column chromatography, and combinations thereof. Exemplary mass spectrometry (MS) include, but are not limited to, tandem MS, LC-MS, LC-MS/MS, matrix assisted laser desorption ionisation mass spectrometry (MALDI-MS), Fourier transform mass spectrometry (FTMS), ion mobility separation with mass spectrometry (IMS-MS), electron transfer dissociation (ETD-MS), and combinations thereof. Exemplary electrophoretic methods include, but are not limited to, capillary electrophoresis (CE), CE-MS, gel electrophoresis, agarose gel electrophoresis, acrylamide gel electrophoresis, SDS-polyacrylamide gel electrophoresis (SDS-PAGE) followed by Western blotting using antibodies that recognize specific glycan structures, and combinations thereof. Exemplary nuclear magnetic resonance (NMR) include, but are not limited to, one-dimensional NMR (1D-NMR), two-dimensional NMR (2D-NMR), correlation spectroscopy magnetic-angle spinning NMR (COSY-NMR), total correlated spectroscopy NMR (TOCSY-NMR), heteronuclear single-quantum coherence NMR (HSQC-NMR), heteronuclear multiple quantum coherence (HMQC-NMR), rotational nuclear overhauser effect spectroscopy NMR (ROESY-NMR), nuclear overhauser effect spectroscopy (NOESY-NMR), and combinations thereof.
In some instances, techniques described herein may be combined with one or more other technologies for the detection, analysis, and or isolation of glycans or glycoproteins. For example, in certain instances, glycans are analyzed in accordance with the present disclosure using one or more available methods (to give but a few examples, see Anumula, Anal. Biochem. 350(1):1, 2006; Klein et al., Anal. Biochem., 179:162, 1989; and/or Townsend, R. R. Carbohydrate Analysis” High Performance Liquid Chromatography and Capillary Electrophoresis., Ed. Z. El Rassi, pp 181-209, 1995, each of which is incorporated herein by reference in its entirety). For example, in some instances, glycans are characterized using one or more of chromatographic methods, electrophoretic methods, nuclear magnetic resonance methods, and combinations thereof.
In some instances, methods for evaluating one or more abatacept-specific parameters, e.g., in a biologic preparation, e.g., one or more of abatacept parameters disclosed in Table 1 in a biologic preparation are known in the art and/or are disclosed in Table 7:
References listed in Table 7 are hereby incorporated by reference in their entirety or, in the alternative, to the extent that they pertain to one or more of the methods disclosed in Table 7. Other methods for evaluating one or more abatacept specific parameters are disclosed in the examples.
An ORENCIA® sample was analyzed to determine the amino acid sequence of the fusion protein. The sequence of the protein is shown as SEQ ID NO:1 (
Characterization of abatacept was performed by orthogonal methods. Samples of abatacept were analyzed and measurements were made including use of glycan profiling, glycoform analysis, post-translational modification analysis, and analysis of other intrinsic and extrinsic structures or features. In one method, described herein, abatacept samples were processed by digesting with trypsin in mass spectrometry compatible digestion buffer compatible, performed using pressure cycling technology. Digestion was performed using a BAROCYCLER® NEP 2320 (Pressure Biosciences) with the Barocycler settings: Temperature: 37° C.; high pressure: 20,000 PSI; time 1 (high pressure): 90 sec; Time 2 (ambient pressure) 20 sec, with 40 cycles, and total digestion time of about 75 minutes. Digests were then analyzed by C18 reversed phase HPLC-MS peptide mapping run utilizing an Orbitrap XL, with multiple sections for the LC gradient to separate each cluster of glycopeptides within each glycosylation site and multiple tune methods to coordinate with the multiple sections of the LC gradient. Of the abatacept structures or features that were measured or determined, 54 were determined to be abatacept parameters, i.e., parameters of abatacept that distinguish abatacept from non-abatacept drug products. These 54 abatacept parameters and values are listed in Table 1.
As disclosed herein, one or more, including a subset, class, multiple classes or all, of the reference criteria shown in Table 1, may be included in a specification for commercial release of abatacept under Section 351(k) of the PHS Act. This concept is exemplified in Table 8, in which parameters A-J are reference criteria defining the abatacept fingerprint, wherein each of parameters A-J is one of parameters 1 to 54 in Table 1. Table 8 also illustrates comparison of these reference criteria with input values corresponding thereto for two hypothetical test biologic preparations (Test Biologic Preparations A and B) having amino acid sequence with identity to SEQ ID NO:1.
Illustrates that a value meets the reference criterion/rule. Blank illustrates that a value does not meet the reference criterion/rule.
According to Table 8, Test Biologic Preparation A would not qualify as abatacept and thus would not be processed as abatacept because input values for each of one or more parameters for the test biologic preparation do not meet the corresponding reference criterion for that parameter. In contrast, Test Biologic Preparation B would qualify as abatacept, as it meets the abatacept fingerprint, and thus would be processed as abatacept because input values for each of one or more parameters for the test biologic preparation meet the corresponding reference criterion for that parameter.
As disclosed herein, one or more, including a subset, class, multiple classes or all, of the reference criteria shown in Table 1, may be included in a specification for commercial release of a pharmaceutical product comprising a CTLA4-Fc fusion. This concept is exemplified in Table 9, in which parameters A-J represent reference criteria for a subset or class of parameters 1 to 54 in Table 1. Table 9 also illustrates comparison of the reference criteria (the signature or fingerprint representing the specification for commercial release of a pharmaceutical product comprising a CTLA4-Fc fusion) with input values corresponding thereto for two hypothetical test CTLA4-Fc fusion preparations (Test Biologic Preparations A and B) having amino acid sequence with identity to SEQ ID NO:1. In this example, only a subset of input values corresponding to A-J meets the criterion.
Illustrates that a value meets the reference criterion/rule. Blank illustrates that a value does not meet the reference criterion/rule.
According to Table 9, Test Biologic Preparation A and B would qualify as CTLA4-Fc fusion pharmaceutical product and thus could be processed into a CTLA4-Fc fusion pharmaceutical product because input values for a subset of reference criterion A-J meet the reference criteria therefor. Test Biologic Preparation A and B shown here would not, however, qualify as abatacept as they do not meet the abatacept fingerprint.
While the methods have been described in conjunction with various instances and examples, it is not intended that the methods be limited to such instances or examples. On the contrary, the methods encompass various alternatives, modifications, and equivalents, as will be appreciated by those of skill in the art.
This application claims benefit of U.S. Provisional Application No. 61/799,682, filed Mar. 15, 2013, the entire contents of which are hereby incorporated by reference.
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/US13/43786 | 5/31/2013 | WO | 00 |
| Number | Date | Country | |
|---|---|---|---|
| 61799682 | Mar 2013 | US |
