Ligand polypeptide and medicine transferring system combined with specificity of CD40L protein

A delivery system and specific technology, applied in the field of protein and peptides, can solve the problem of no further, blocking the CD40/CD40L signal in the same way, and achieve the effect of good targeting effect, strong tissue penetration, and small molecular weight.

Inactive Publication Date: 2014-11-19
SHANGHAI JIAOTONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there have been trimeric polypeptides targeting CD40, which effectively inhibit the CD40 / CD40L signaling pathway at the cellular level, but there is no further in vivo experiment report, and there is no ligand polypeptide targeting CD40L to achieve inhibition. A report on disconnection of CD40 / CD40L signaling

Method used

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  • Ligand polypeptide and medicine transferring system combined with specificity of CD40L protein
  • Ligand polypeptide and medicine transferring system combined with specificity of CD40L protein
  • Ligand polypeptide and medicine transferring system combined with specificity of CD40L protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Example 1, Derivative polypeptide of CD40L targeting polypeptide and preparation method thereof

[0049] This embodiment relates to the derivative polypeptide of the CD40L-targeting polypeptide, and its preparation method includes the following steps:

[0050] 1) Dissolve 1mg Cy5.5-NHS in 400μl DMSO (dimethyl sulfoxide), and dissolve 0.8mg peptide in 400μl DMSO;

[0051] 2) After mixing Cy5.5-NHS and peptide in equal volumes, add 15 μl triethylamine to the system, place in a constant temperature shaking reactor, and react overnight at room temperature in the dark;

[0052] 3) The reaction system was placed in a dialysis bag with a molecular weight cut-off of 1000, and dialyzed with PBS at room temperature in the dark, and the water was changed three times to remove DMSO, triethylamine and free Cy5.5-NHS;

[0053] 4) The purified CY5.5-polypeptide should be stored at -80°C in the dark.

[0054] For effect verification, the distribution of the polypeptide JD6 and its ...

Embodiment 2

[0055] Example 2, synthetic materials with CD40L targeting

[0056] This embodiment relates to a synthetic material with CD40L targeting, which is obtained by linking a derivative polypeptide specific to targeting polypeptide of CD40L molecule with Mal-PEG2000-DSPE. The sequence of the specific targeting polypeptide of the CD40L molecule is ESEEED, and GGGC is added to the C-terminus during synthesis to obtain a derivative polypeptide whose sequence is ESEEEDGGGC; The imine group is reacted and linked to DSPE-2000-Maleimide (distearoylglyceroylphosphatidylethanolamine polyethylene glycol 2000 maleimide, the polyethylene glycol can also choose PEG5000) to obtain a CD40L-targeted Further, the CD40L molecule-specific targeting polypeptide can also be directly linked with DSPE-2000Maleimide; another synthetic material with CD40L targeting can be obtained.

[0057] The preparation of the synthetic material with CD40L targeting (wherein the polypeptide is a CD40L molecule-specifi...

Embodiment 3

[0062] Example 3, CD40L molecular targeting polypeptide modified fluorescent liposome

[0063] This embodiment relates to a fluorescent liposome, which is CD40L molecule-targeting polypeptide-modified fluorescent liposome, and its preparation method includes the following steps:

[0064] 1) According to the molar ratio EPC (lecithin): CHOL (cholesterol): fluorescein DHPE (1,2-palmitoylphosphatidylethanolamine labeled with fluorescein) = 60:40:1, 70:30:1 or 80:20: 1 Weigh and fully dissolve in tert-butanol;

[0065] 2) In the lipid tert-butanol solution, add the polypeptide-PEG-DSPE aqueous solution (prepared in Example 2) dropwise to keep the system clarified until the molar ratios of polypeptide lipid to fluorescein DHPE are 0.1:1, 0.5:1, respectively, 1:1, 2:1, 3:1, 4:1, 5:1;

[0066] 3) Mix the peptide lipids and other lipid components, freeze them quickly in liquid nitrogen, and place them in a freeze dryer at -70 degrees Celsius to freeze-dry;

[0067] 4) Use PBS to ...

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Abstract

The invention discloses ligand polypeptide and a medicine transferring system combined with the specificity of CD40L protein. The ligand polypeptide combined with the specificity of the CD40L protein comprises an amino acid sequence showed in SEQ ID NO.1. The medicine transferring system provided by the invention comprises the ligand polypeptide, medicine carrying system and at least one kind of active substance, wherein the medicine carrying system is specifically liposome; the at least one kind of active substance is the substance which is required to be transferred to specified positions in the body; the ligand polypeptide provided by the invention has excellent targeting functions for CD40L; aiming at immune diseases in the body with high expression of CD40L, a reagent kit used for early diagnosis and therapeutic medicine with specificity can be developed; and meanwhile, the polypeptide can combined with the liposome to be used as targeting medicine carrier, so that the medicine transferring system is developed into a targeting medicine transferring system which is aiming at immune diseases in the body.

Description

technical field [0001] The invention relates to the technical field of protein polypeptides, in particular to a ligand polypeptide specifically binding to CD40L protein and a drug delivery system. Background technique [0002] CD40L is a member of the tumor necrosis factor (TNF) superfamily, a type II membrane protein, mainly expressed on the surface of CD4+ T cells, including Th0, Th1 and Th2 cell subsets. CD40L is also expressed on the surface of other lymphocytes, such as activated CD8+ T cells or B cells. Studies have shown that functional CD40L is also expressed on the surface of nonlymphoid cells of the hematopoietic lineage, such as activated platelet cells. Activated T cells can transiently express CD40L within ten minutes after activation, reach a peak at 6 hours, and then the expression level begins to decline. [0003] CD40 / CD40L is a pair of important co-stimulatory molecules in the specific immune response system, participating in the body's humoral and cellul...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K7/06A61K47/42A61K9/127A61K38/08A61K45/00A61K48/00A61K49/14A61K49/22A61K51/08A61P37/02
Inventor 徐宇虹丁茜刘丹
Owner SHANGHAI JIAOTONG UNIV
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