US4421685A
Expired / Lapsed
US
⏱️ 0.0 years remaining
Process for producing an insulin
Assignee: Eli Lilly and Co
•
Filing: 1981-05-11
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Granted: 1983-12-20
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Expires: 2001-05-11
Abstract
Abstract Production of insulin or an insulin analog is provided by combination of an A-chain and a B-chain, which comprises bringing the S-sulfonated form of the A-chain, the S-sulfonated form of the B-chain, and a thiol reducing agent together in an aqueous medium under conditions which produce a mixture having (1) a pH of from about 8 to about 12, (2) a total protein concentration of from about 0.1 to about 50 milligrams per milliliter, and (3) an amount of thiol reducing agent which affords a total of from about 0.4 to about 2.5 --SH groups per each --SSO 3 - group present in the total amount of A- and B-chain S-sulfonates, and allowing formation of insulin or an insulin analog to occur by maintaining the mixture at a temperature of from about 0° C. to about 25° C. and in an environment which provides a source of oxygen.
Inventors
- 👤 Ronald E. Chance
- 👤 James A. Hoffmann
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-23 19:26
CPC Classifications
C; C07; C07K; C07K1/00; C07K1/12; C07K1/06; C07K1/061; C07K1/067
Claims Analysis
Total Claims: 19
Claim 1
Independent Claim
161 words
A process for combining an A-chain of an insulin or an insulin analog and B-chain of an insulin or an insulin analog to produce an insulin or an insulin analog, which comprises bringing the S-sulfonated form of the A-chain, the S-sulfonated form of the B-chain, and a thiol reducing agent together in an aqueous medium under conditions which produce a mixture having (1) a pH of from about 8 to about 12, (2) a total protein concentration of from about 0.1 to about 50 milligrams per milliliter, and (3) an amount of thiol reducing agent which affords a total of from about 0.4 to about 2.5 --SH groups per each --SSO 3 - group present in the total amount of A- and B-chain S-sulfonates, and allowing formation of insulin or an insulin analog to occur by maintaining the mixture at a temperature of from about 0° C. to about 25° C. and in an environment which provides a source of oxygen.
Claim 2
Dependent Claim
161 words
A process for combining an A-chain of an insulin or an insulin analog and B-chain of an insulin or an insulin analog to produce an insulin or an insulin analog, which comprises bringing the S-sulfonated form of the A-chain, the S-sulfonated form of the B-chain, and a thiol reducing agent together in an aqueous medium under conditions which produce a mixture having (1) a pH of from about 8 to about 12, (2) a total protein concentration of from about 0.1 to about 50 milligrams per milliliter, and (3) an amount of thiol reducing agent which affords a total of from about 0.4 to about 2.5 --SH groups per each --SSO 3 - group present in the total amount of A- and B-chain S-sulfonates, and allowing formation of insulin or an insulin analog to occur by maintaining the mixture at a temperature of from about 0° C. to about 25° C. and in an environment which provides a source of oxygen.
Claim 3
Dependent Claim
24 words
Process of claim 1, in which the A-chain S-sulfonate and the B-chain S-sulfonate each has the amino acid sequence represented by a naturally-occurring insulin.
Claim 4
Dependent Claim
22 words
Process of claim 2, in which the weight ratio of A-chain S-sulfonate to B-chain S-sulfonate is from about 0.1:1 to about 10:1.
Claim 5
Dependent Claim
21 words
Process of claim 3, in which the weight ratio A-chain S-sulfonate to B-chain S-sulfonate is from about 1:1 to about 3:1.
Claim 6
Dependent Claim
19 words
Process of claim 4, in which the protein concentration is from about 2 to about 20 milligrams per milliliter.
Claim 7
Dependent Claim
19 words
Process of claim 5, in which the pH of the reaction mixture is from about 9.5 to about 11.0.
Claim 8
Dependent Claim
19 words
Process of claim 6, in which the pH of the reaction mixture is from about 10.4 to about 10.6.
Claim 9
Dependent Claim
44 words
Process of claim 7, in which the thiol reducing agent is present in an amount which affords a total of from about 0.9 to about 1.1 --SH group per each --SSO 3 - group present in the total amount of A- and B-chain S-sulfonates.
Claim 10
Dependent Claim
20 words
Process of claim 8, in which the thiol reducing agent is selected from the group consisting of dithiothreitol and dithioerythritol.
Claim 11
Dependent Claim
12 words
Process of claim 9, in which the thiol reducing agent is dithiothreitol.
Claim 12
Dependent Claim
20 words
Process of claim 8, in which the thiol reducing agent is selected from the group consisting of cysteine and dithiothreitol.
Claim 13
Dependent Claim
12 words
Process of claim 11, in which the thiol reducing agent is cysteine.
Claim 14
Dependent Claim
23 words
Process of claim 11, in which the reaction mixture is maintained at a temperature of from about 2° C. to about 10° C.
Claim 15
Dependent Claim
17 words
Process of claim 13, in which the insulin that is produced is bovine, porcine, or human insulin.
Claim 16
Dependent Claim
44 words
Process of claim 14, in which the insulin that is produced is bovine insulin, the weight ratio of A-chain S-sulfonate to B-chain S-sulfonate is from about 1.4:1.0 to about 1.8:1.0, and the protein concentration is from about 3 to about 8 milligrams per milliliter.
Claim 17
Dependent Claim
44 words
Process of claim 14, in which the insulin that is produced is porcine insulin, the weight ratio of A-chain S-sulfonate to B-chain S-sulfonate is from about 1.0:1.0 to about 1.4:1.0, and the protein concentration is from about 8 to about 16 milligrams per milliliter.
Claim 18
Dependent Claim
14 words
Process of claim 14, in which the insulin that is produced is human insulin.
Claim 19
Dependent Claim
44 words
Process of claim 17, in which the insulin that is produced is human insulin, the weight ratio of A-chain S-sulfonate to B-chain S-sulfonate is from about 1.8:1.0 to about 2.2:1.0, and the protein concentration is from about 3 to about 8 milligrams per milliliter.
References Cited (Backward Citations)
US3420810A
Cited
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1986-12-19 | 1987-06-20 | 1987-12-20 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1990-12-19 | 1991-06-20 | 1991-12-20 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1994-12-19 | 1995-06-20 | 1995-12-20 | $7700.00 | Window Closed / Lapsed |