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Polypeptide compound, pharmaceutical composition, its preparation method and application thereof

A technology of polypeptide complexes and compositions, applied in the directions of drug combinations, pharmaceutical formulations, inactive components of polymer compounds, etc., can solve the problems of short half-life and increase the pain of patients, and achieve the effect of prolonging the half-life and facilitating clinical promotion and application.

Active Publication Date: 2014-02-19
TIANJIN INSTITUTE OF PHARMA RESEARCH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] However, the Exendin-4 drug on the market still has the problem of short half-life, and the medicinal Exenatide (Exenatide) is injected twice a day, which greatly increases the suffering of patients.
Therefore, there is currently a need for methods to address the short in vivo half-life of Exendin-4 drugs

Method used

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  • Polypeptide compound, pharmaceutical composition, its preparation method and application thereof
  • Polypeptide compound, pharmaceutical composition, its preparation method and application thereof
  • Polypeptide compound, pharmaceutical composition, its preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0057] Embodiment 1: Preparation of carrier polypeptide A

[0058] According to the preferred codon (but not limited to the preferred codon) of Escherichia coli, the base sequence containing polypeptide A is synthesized by the method of total gene synthesis, and the DNA sequence is as follows:

[0059] SEQ ID NO 3:

[0060] 5 TATG ATTGAAGGCCGT GGCCTGTGGTGGAAAGCGTGGTGGAAAGCGTGGTGGAAATCCCTGTGGTGGCGTAAACGTAAACGTAAAGCGTAATAAG 3

[0061] SEQ ID NO 4:

[0062] 5 GATCCTTATTACGCTTTACGTTTACGTTTACGCCACCACAGGGATTTCCACCACGCTTTCCACCACGCTTTCCACCACAGGCC ACGGCCTTCAAT

[0063] To bind DNA sequence 3 and sequence 4, add equal molar amounts of SEQ ID NO3 and SEQ ID NO 4 respectively, incubate in a water bath at 95°C for 5 minutes, and then slowly cool down to room temperature. This double-stranded DNA can be connected with the Escherichia coli expression vector pET15b after being digested by NdeI / BamHI. As shown below, the double-stranded DNA synthesized by the above method contains...

Embodiment 2

[0067] Embodiment 2: Preparation of carrier polypeptide B

[0068] According to the preferred codon (but not limited to the preferred codon) of Escherichia coli, the DNA sequence of Polypeptide B synthesized by the method of total gene synthesis is as follows:

[0069] SEQ ID NO 5:

[0070] 5 TATG ATTGAAGGCCGT GGCCTGTGGTGGAAAGTGTGGTGGAAACTGTGGTGGAAAAGCCTGTGGTGGCGCAAACGCCTGCGCAAAGCGTAATAAG 3

[0071] SEQ ID NO 6:

[0072] 5 GATCCTTATTACGCTTTGCGCAGGCGTTTGCGCCACCACAGGCTTTTCCACCACAGTTTCCACCACACTTTTCCACCACAGGCC ACGGCCTTCAAT

[0073] To bond DNA sequence 5 and sequence 6, add equal molar amounts of SEQ ID NO3 and SEQ ID NO 4 respectively, place in a water bath at 95°C for 5 minutes, and then slowly cool to room temperature. This double-stranded DNA can be connected with the Escherichia coli expression vector pET15b after being digested by NdeI / BamHI. As shown below, the double-stranded DNA synthesized by the above method contains the sharp ends of 5'NdeI and 3'BamHI, an...

Embodiment 3

[0076] Embodiment 3: Preparation and purification technology of recombinant Exendin-4

[0077] Exendin-4 is a natural product without patent restrictions. In the present invention, biological methods are used to prepare Exendin-4, and this patent is also applicable to the use of other polypeptide preparation methods, such as polypeptide chemical synthesis.

[0078] SEQ ID NO 9:

[0079] HGEGTFTSDLSKQMEEEAVRLFIEWLKNGGPS SGAPPPS

[0080] The coding DNA was synthesized and prepared according to the amino acid sequence of Exendin-4 (SEQ ID NO 9). Two complementary DNA single strands were synthesized by Shanghai Sangong, see SEQ ID NO: 7, SEQ ID NO: 8. And referring to the description in Example 1, a double-stranded DNA encoding Exendin-4 was prepared, and the double-stranded DNA had a restriction site (NdeI / BamHI) connected to the vector (pET15b). Referring to the content of Example 1, the expression plasmid of Exendin-4 was constructed.

[0081] Recombinant Exendin-4 was pr...

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PUM

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Abstract

The invention discloses a polypeptide compound, which contains Exendin-4 and a carrier polypeptide. The carrier polypeptide can effectively prolong blood half-life time of Exendin-4, overcomes the present situation that Exendin-4 cannot be used in clinical practice due to its short half-life time, and has an application prospect in the field of drugs for treating diabetes and obesity. The invention also discloses a preparation method of the polypeptide compound and an application thereof. In addition, the invention further discloses a pharmaceutical composition containing the polypeptide compound.

Description

technical field [0001] The present invention relates to the field of medicines related to diabetes, in particular, the present invention relates to a polypeptide complex, which has prolonged exendin-4 half-life in vivo. The invention also relates to the preparation method and application of the polypeptide complex. Background technique [0002] Diabetes mellitus is a metabolic disorder syndrome characterized by chronic hyperglycemia associated with genetic factors and multiple environmental factors. Because diabetes is also accompanied by many complications, it has become the third largest health killer after malignant tumors and cardiovascular diseases. In 1984, glucagon peptide-1 (GLP-1) drugs were discovered, which is an incretin with 30 amino acids. The main reason why GLP-1 is limited as a hypoglycemic drug is that the half-life of GLP-1 in vivo is only 3-5 minutes. [0003] Exendin-4 is a straight-chain polypeptide composed of 39 amino acids, which was originally is...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K38/22A61K47/42A61P3/04A61P3/10
Inventor 龚珉王玉丽徐为人汤立达任晓文付刚郑学敏
Owner TIANJIN INSTITUTE OF PHARMA RESEARCH
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