FUSION PROTEIN OF EXENDIN-4 AND ITS ANALOG, PREPARATION METHOD AND USE THEREOF

• US 20130142795A1
• Filed: 06/10/2011
• Published: 06/06/2013
• Est. Priority Date: 06/11/2010
• Status: Active Grant

• Current Assignee: Beijing Dongfang Biotech Co., Ltd.
• Original Assignee: Beijing Dongfang Biotech Co., Ltd.
• Inventors: Li, Xianzhong, Bai, Xianhong, Tu, Hua
• Granted Patent: US 8,889,619 B2
• Time in Patent Office: Days
• US Class Current: 424/134.1
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61P 3/00 Drugs for disorders of the metabolism of the blood or the extracellular fluid A61P7/00
• CPC Class Codes: A61P 3/04 Anorexiants; Antiobesity agents
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: C07K 14/46 from vertebrates
• CPC Class Codes: C07K 14/575 Hormones derived from pro-opiomelanocortin, pro-enkephalin or pro-dynorphin C07K14/665, e.g. corticotropin C07K14/695
• CPC Class Codes: C07K 14/57563 Vasoactive intestinal peptide [VIP]; Related peptides
• CPC Class Codes: C07K 14/605 Glucagons
• CPC Class Codes: C07K 16/00 Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies antibodies with enzymatic activity, e.g. abzymes C12N9/0002
• CPC Class Codes: C07K 2319/30 Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto

Abstract

Provided are a fusion protein of Exendin-4 and its analog, the preparation method and use thereof. The fusion protein is obtained by fusing of Exendin-4 or its analog to Fc region of human IgG2 via a linking peptide, which has the better stability and prolonged serum half-life, and can be used for treating diabetes and obesity.

Claims

1. A fusion protein, which is obtained by fusing peptide hormone to transport protein via linker, wherein, the said peptide hormone is Exendin-4 or analogue of Exendin-4, and the said peptide hormone is capable of lowering the blood glucose;

the said transport protein is the Fc fragment of the immunoglobulin IgG2;

the said fusion protein is capable of lowering the blood glucose;

the said peptide hormone comprises the sequence shown by Formula I;

His-Xaa²-Glu-Gly-Thr-Phe-Thr-Ser-Asp-Xaa¹⁰-Ser-Xaa¹²-Xaa¹³-Xaa¹⁴-Glu-Glu-Glu-Ala-Xaa¹⁹-Xaa²⁰-Xaa²¹-Phe-Ile-Xaa²⁴-Trp-Leu-Xaa²⁷-Xaa²⁸-Gly-Xaa³⁰-Xaa³¹-Xaa³²-Xaa³³-Xaa³⁴-Xaa³⁵-Xaa³⁶-Xaa³⁷-Xaa³⁸-Xaa³⁹ 

 

Formula I

Wherein;

Xaa² is Gly, Thr, Ala, Ser, Leu, Ile or Lys;

Xaa¹⁰ is Leu, Ala, Ser, Leu, Ile, Glu or Lys;

Xaa¹² is Lys, Leu, Thr, Ser, Leu, Ile or Cys;

Xaa¹³ is Gln, Thr, Ala, Val, Leu, Ile or Lys;

Xaa¹⁴ is Met, Tyr, Thr, Ala, Ser, Ile or Lys;

Xaa¹⁹ is Val, Cys, Ala, Ser, Leu, Ile or Lys;

Xaa²⁰ is Arg, Thr, Tyr, Ser, Leu, Ile or Lys;

Xaa²¹ is Leu, Thr, Ala, Asp, Glu, His or Lys;

Xaa²⁴ is Glu, Leu, Thr, Ala, Ser, Lys or Ile;

Xaa²⁷ is Lys, Ala, Ser, Leu, Thr, Ile or Lys;

Xaa²⁸ is Asp, Thr, Ala, Ser, Leu, Ile or Lys;

Xaa³⁰ is Gly, Thr, Ala, Ser, Leu, Ile or Arg;

Xaa³¹ is Pro, Val, Ser, Ala, Leu, Ile or Lys;

Xaa³² is Ser, Thr, Glu, Ser, Asp, Lys or Ile;

Xaa³³ is Thr, Ser, Ala, Met, Leu, Ile or Lys;

Xaa³⁴ is Gly, Thr, Met, Ser, Ile, Leu or Lys;

Xaa³⁵ is Ala, Thr, Ala, Glu, Leu, Ile or Phe;

Xaa³⁶ is Pro, Ala, Thr, Ser, Leu, Ile or Cys;

Xaa³⁷ is Pro, Thr, Ser, Ala, His, Lys or Ile;

Xaa³⁸ is Pro, Thr, Val, Ser, Leu, Lys or Ile;

Xaa³⁹ is Ser, Tyr, Ala, Leu, Ser, Ile or Lys.

2. The fusion protein according to claim 1, wherein the peptide hormone is exendin-4, which has the amino acid sequence shown by SEQ ID NO:

1.

3. The fusion protein according to claim 1, wherein the linker is the peptide which has the sequence shown by (Gly)_m-Xaa-(Gly)_n, in which m is an integer between 3 and 8, n is an integer between 3 and 8, and Xaa is any one selected from the group consisting of Gly, Ser, Ala and Thr.

4. The fusion protein according to claim 3, wherein m is an integer between 4 and 6;

and n is an integer between 4 and 6.

5. The fusion protein according to claim 4, wherein the linker has the amino acid sequence shown by SEQ ID NO:

3.

6. The fusion protein according to claim 1, wherein the Fc fragment of IgG2 originates from human.

7. The fusion protein according to claim 1, wherein the said fusion protein has the amino acid sequence shown by the amino acid sequence shown by SEQ ID NO:

4.

8. A polynucleotide sequence encoding the fusion protein according to claim 1.

10. A host cell, wherein, the said host cell is generated by transfecting the recipient cell by the vector comprising the polynucleotide sequence according to claim 8, the said recipient cell is CHO cell.

14. A pharmaceutical composition, wherein, the said pharmaceutical composition comprises the fusion protein according to claim 1 and pharmacologically acceptable excipient.

15. The pharmaceutical composition according to claim 14, wherein the pharmaceutical composition is administered as local administration, aerosol or injection.

16. The pharmaceutical composition according to claim 15, wherein the injection is administered through at least one of intraperitoneal injection, subcutaneous injection, intramuscular injection or intravenous injection.

17. A method of treating diabetes or obesity, wherein the said method comprises the step of administering the fusion protein according to claim 1 to subjects.

9. (canceled)

11-13. -13. (canceled)