NZ741060B2 - Novel insulin analogs and use thereof - Google Patents
Novel insulin analogs and use thereof Download PDFInfo
- Publication number
- NZ741060B2 NZ741060B2 NZ741060A NZ74106016A NZ741060B2 NZ 741060 B2 NZ741060 B2 NZ 741060B2 NZ 741060 A NZ741060 A NZ 741060A NZ 74106016 A NZ74106016 A NZ 74106016A NZ 741060 B2 NZ741060 B2 NZ 741060B2
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- Prior art keywords
- seq
- insulin analog
- chain
- tyrosine
- analog peptide
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- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical class N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 title claims abstract 23
- 239000004026 insulin derivative Substances 0.000 claims abstract 20
- 108020004707 nucleic acids Proteins 0.000 claims abstract 8
- 150000007523 nucleic acids Chemical class 0.000 claims abstract 8
- 102000039446 nucleic acids Human genes 0.000 claims abstract 8
- 239000013604 expression vector Substances 0.000 claims abstract 6
- 238000000034 method Methods 0.000 claims abstract 6
- 206010012601 diabetes mellitus Diseases 0.000 claims abstract 3
- 239000008194 pharmaceutical composition Substances 0.000 claims abstract 3
- 108090000765 processed proteins & peptides Proteins 0.000 claims 14
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 claims 9
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 claims 9
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 claims 8
- DCXYFEDJOCDNAF-UHFFFAOYSA-N Asparagine Natural products OC(=O)C(N)CC(N)=O DCXYFEDJOCDNAF-UHFFFAOYSA-N 0.000 claims 6
- DCXYFEDJOCDNAF-REOHCLBHSA-N L-asparagine Chemical compound OC(=O)[C@@H](N)CC(N)=O DCXYFEDJOCDNAF-REOHCLBHSA-N 0.000 claims 6
- ONIBWKKTOPOVIA-UHFFFAOYSA-N Proline Natural products OC(=O)C1CCCN1 ONIBWKKTOPOVIA-UHFFFAOYSA-N 0.000 claims 6
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 claims 6
- 235000009582 asparagine Nutrition 0.000 claims 6
- 229960001230 asparagine Drugs 0.000 claims 6
- ROHFNLRQFUQHCH-YFKPBYRVSA-N L-leucine Chemical compound CC(C)C[C@H](N)C(O)=O ROHFNLRQFUQHCH-YFKPBYRVSA-N 0.000 claims 5
- ROHFNLRQFUQHCH-UHFFFAOYSA-N Leucine Natural products CC(C)CC(N)C(O)=O ROHFNLRQFUQHCH-UHFFFAOYSA-N 0.000 claims 5
- AYFVYJQAPQTCCC-UHFFFAOYSA-N Threonine Natural products CC(O)C(N)C(O)=O AYFVYJQAPQTCCC-UHFFFAOYSA-N 0.000 claims 5
- 239000004473 Threonine Substances 0.000 claims 5
- ZDXPYRJPNDTMRX-UHFFFAOYSA-N glutamine Natural products OC(=O)C(N)CCC(N)=O ZDXPYRJPNDTMRX-UHFFFAOYSA-N 0.000 claims 5
- 238000003259 recombinant expression Methods 0.000 claims 4
- WHUUTDBJXJRKMK-UHFFFAOYSA-N Glutamic acid Natural products OC(=O)C(N)CCC(O)=O WHUUTDBJXJRKMK-UHFFFAOYSA-N 0.000 claims 3
- 239000004471 Glycine Substances 0.000 claims 3
- 235000013922 glutamic acid Nutrition 0.000 claims 3
- 239000004220 glutamic acid Substances 0.000 claims 3
- 241000588724 Escherichia coli Species 0.000 claims 2
- CKLJMWTZIZZHCS-REOHCLBHSA-N L-aspartic acid Chemical compound OC(=O)[C@@H](N)CC(O)=O CKLJMWTZIZZHCS-REOHCLBHSA-N 0.000 claims 2
- COLNVLDHVKWLRT-QMMMGPOBSA-N L-phenylalanine Chemical compound OC(=O)[C@@H](N)CC1=CC=CC=C1 COLNVLDHVKWLRT-QMMMGPOBSA-N 0.000 claims 2
- 150000001413 amino acids Chemical group 0.000 claims 2
- 235000003704 aspartic acid Nutrition 0.000 claims 2
- OQFSQFPPLPISGP-UHFFFAOYSA-N beta-carboxyaspartic acid Natural products OC(=O)C(N)C(C(O)=O)C(O)=O OQFSQFPPLPISGP-UHFFFAOYSA-N 0.000 claims 2
- 238000005277 cation exchange chromatography Methods 0.000 claims 2
- 239000002773 nucleotide Substances 0.000 claims 2
- 125000003729 nucleotide group Chemical group 0.000 claims 2
- COLNVLDHVKWLRT-UHFFFAOYSA-N phenylalanine Natural products OC(=O)C(N)CC1=CC=CC=C1 COLNVLDHVKWLRT-UHFFFAOYSA-N 0.000 claims 2
- 102000003670 Carboxypeptidase B Human genes 0.000 claims 1
- 108090000087 Carboxypeptidase B Proteins 0.000 claims 1
- QNAYBMKLOCPYGJ-REOHCLBHSA-N L-alanine Chemical compound C[C@H](N)C(O)=O QNAYBMKLOCPYGJ-REOHCLBHSA-N 0.000 claims 1
- 102000004142 Trypsin Human genes 0.000 claims 1
- 108090000631 Trypsin Proteins 0.000 claims 1
- 239000002253 acid Substances 0.000 claims 1
- 235000004279 alanine Nutrition 0.000 claims 1
- 238000005571 anion exchange chromatography Methods 0.000 claims 1
- 239000013592 cell lysate Substances 0.000 claims 1
- 238000012258 culturing Methods 0.000 claims 1
- 238000010298 pulverizing process Methods 0.000 claims 1
- 230000001131 transforming effect Effects 0.000 claims 1
- 239000012588 trypsin Substances 0.000 claims 1
- 102000004877 Insulin Human genes 0.000 abstract 1
- 108090001061 Insulin Proteins 0.000 abstract 1
- 239000004480 active ingredient Substances 0.000 abstract 1
- 238000000338 in vitro Methods 0.000 abstract 1
- 229940125396 insulin Drugs 0.000 abstract 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- A61K38/22—Hormones
- A61K38/28—Insulins
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P3/00—Drugs for disorders of the metabolism
- A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
- A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/12—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length by hydrolysis, i.e. solvolysis in general
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/14—Extraction; Separation; Purification
- C07K1/16—Extraction; Separation; Purification by chromatography
- C07K1/18—Ion-exchange chromatography
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/14—Extraction; Separation; Purification
- C07K1/36—Extraction; Separation; Purification by a combination of two or more processes of different types
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/575—Hormones
- C07K14/62—Insulins
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/70—Vectors or expression systems specially adapted for E. coli
Abstract
The present invention relates to a novel insulin analog, and more specifically, to an insulin analog with an improved in vitro effect compared with native insulin, a nucleic acid encoding the same, an expression vector including the nucleic acid, a transformant introduced with the expression vector, a method of producing the insulin analog from the transformant, a pharmaceutical composition for treating diabetes containing the insulin analog as an active ingredient, and a method for treating diabetes using the insulin analog or the pharmaceutical composition.
Claims (16)
1. An insulin analog peptide comprising an A-chain of SEQ ID NO: 3 indicated in the following General Formula 1 and a B-chain of SEQ ID NO: 4 indicated in the following General Formula 2: (General Formula 1) Xaa1-Ile-Val-Glu-Xaa2-Cys-Cys-Thr-Ser-Ile-Cys-Xaa3-Leu-Xaa4-Gln-Xaa5-Glu- Asn-Xaa6-Cys-Xaa7(SEQ ID NO: 3), (General Formula 2) Phe-Val-Asn-Gln-His-Leu-Cys-Gly-Ser-His-Leu-Val-Glu-Ala-Leu-Xaa8-Leu-Val- y-Glu-Arg-Gly-Phe-Xaa9-Tyr-Xaa10-Xaa11-Lys-Thr (SEQ ID NO: 4) (i) wherein, in the n of SEQ ID NO: 3, Xaa1 is glycine; Xaa2 is glutamine; Xaa3 is serine; Xaa4 is aspartic acid; Xaa5 is leucine; Xaa6 is tyrosine; and Xaa7 is asparagine; and in the B-chain of SEQ ID NO: 4, Xaa8 is tyrosine; Xaa9 is phenylalanine; Xaa10 is threonine; and Xaa11 is proline; (ii) n, in the A-chain of SEQ ID NO: 3, Xaa1 is e, Xaa2 is glutamine, Xaa3 is serine, Xaa4 is ic acid, Xaa5 is leucine, Xaa6 is tyrosine, and Xaa7 is asparagine, and in the B-chain of SEQ ID NO: 4, Xaa8 is tyrosine, Xaa9 is absent, Xaa10 is threonine, and Xaa11 is proline; or (iii) wherein, in the A-chain of SEQ ID NO: 3, Xaa1 is glycine, Xaa2 is glutamine, Xaa3 is serine, Xaa4 is alanine, Xaa5 is e, Xaa6 is tyrosine, and Xaa7 is asparagine, in the B-chain of SEQ ID NO: 4, Xaa8 is glutamic acid, Xaa9 is absent, Xaa10 is threonine, and Xaa11 is proline.
2. The insulin analog peptide of claim 1, n, in the A-chain of SEQ ID NO: 3, Xaa1 is glycine; Xaa2 is glutamine; Xaa3 is serine; Xaa4 is aspartic acid; Xaa5 is leucine; Xaa6 is ne; and Xaa7 is asparagine; and in the B-chain of SEQ ID NO: 4, Xaa8 is tyrosine; Xaa9 is phenylalanine; Xaa10 is ine; and Xaa11 is proline.
3. The insulin analog peptide of claim 1, wherein, in the A-chain of SEQ ID NO: 3, Xaa1 is glycine, Xaa2 is glutamine, Xaa3 is serine, Xaa4 is glutamic acid, Xaa5 is leucine, Xaa6 is tyrosine, and Xaa7 is asparagine, in the B-chain of SEQ ID NO: 4, Xaa8 is tyrosine, Xaa9 is absent, Xaa10 is threonine, and Xaa11 is proline.
4. The insulin analog e of claim 3, wherein the peptide comprises an amino acid sequence represented by SEQ ID NO: 20.
5.The insulin analog peptide of claim 1, wherein, in the A-chain of SEQ ID NO: 3, Xaa1 is glycine, Xaa2 is ine, Xaa3 is serine, Xaa4 is e, Xaa5 is leucine, Xaa6 is tyrosine, and Xaa7 is asparagine, and in the B-chain of SEQ ID NO: 4, Xaa8 is glutamic acid, Xaa9 is absent, Xaa10 is threonine, and Xaa11 is proline.
6. The insulin analog peptide of claim 5, wherein the peptide comprises an amino acid sequence represented by SEQ ID NO: 22.
7. A nucleic acid encoding the n analog peptide according to any one of claims 1 to 6.
8. The nucleic acid of claim 7, wherein the nucleic acid comprises a nucleotide sequence selected from the group consisting of SEQ ID NOS: 19, and 21.
9. A recombinant expression vector comprising the nucleic acid according to claim 7.
10. An isolated transformant which is transformed with the recombinant expression vector according to claim 9.
11. The transformant of claim 10, wherein the transformant is E. coli.
12. A method of preparing the insulin analog peptide ing to claim 1, comprising: a) preparing a recombinant expression vector comprising a nucleic acid encoding the insulin analog peptide according to claim 1; b) transforming the recombinant expression vector into a host cell and obtaining an isolated transformant rom; c) culturing the transformant and expressing the insulin analog peptide; and d) isolating and purifying the expressed insulin analog e.
13. The method of claim 12, n the nucleic acid comprises a nucleotide sequence selected from the group consisting of SEQ ID NOS: 19, and 21.
14. The method of claim 12, wherein the transformant is E. coli.
15. The method of claim 12, wherein the isolating and purifying comprise: d-1) obtaining the ormant cells from the culture and pulverizing the same; d-2) recovering the expressed insulin analog e from the pulverized cell lysate followed by refolding the same; d-3) purifying the refolded insulin analog peptide by cation exchange chromatography; d-4) ng the purified insulin analog peptide with trypsin and carboxypeptidase B; d-5) sequentially ing the treated insulin analog e by cation exchange chromatography and anion exchange chromatography.
16. A pharmaceutical composition for treating diabetes comprising the insulin analog peptide according to any one of claims 1 to 6 and a pharmaceutically able carrier. None set by CAR MigrationNone set by CAR Unmarked set by CAR None set by CAR MigrationNone set by CAR Unmarked set by CAR [
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| KR20150121819 | 2015-08-28 | ||
| PCT/KR2016/009606 WO2017039267A1 (en) | 2015-08-28 | 2016-08-29 | Novel insulin analogs and use thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| NZ741060A NZ741060A (en) | 2024-05-31 |
| NZ741060B2 true NZ741060B2 (en) | 2024-09-03 |
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