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ITGB4BP and derivates thereof used for preventing and/or treating hypertrophic scar and fibrosis lesion

A technology for hypertrophic scars and derivatives, which can be used in gene therapy, skin diseases, drug combinations, etc., and can solve problems such as unreported effects and functions.

Active Publication Date: 2009-12-09
SOUTHWEST HOSPITAL OF CHONGQING
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

ITGB4BP can be detected in normal mucosal cells, but it is overexpressed in cancer cells, such as the upregulation of ITGB4BP in colorectal cancer (19), and it is especially obvious in lymph node metastases (29), so it is suggested that the high expression of ITGB4BP may Involved in tumor proliferation and metastasis
[0019] It can be seen from the above that ITGB4BP plays an important role in ribosome synthesis, cytoskeleton, and tumor proliferation, differentiation, and metastasis. However, there are few related literatures on ITGB4BP protein. It has not been reported yet, so its function and role in fibrosis need further in-depth research

Method used

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  • ITGB4BP and derivates thereof used for preventing and/or treating hypertrophic scar and fibrosis lesion
  • ITGB4BP and derivates thereof used for preventing and/or treating hypertrophic scar and fibrosis lesion
  • ITGB4BP and derivates thereof used for preventing and/or treating hypertrophic scar and fibrosis lesion

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] Example 1. Construction of Shuttle Vectors

[0054] 1.1 Acquisition of target gene sequence

[0055] This experiment was carried out using pEGFP-N2 and pITGB4BP-EGFP plasmids, among which pEGFP-N2 was purchased from clontech company (product number: 6081-1), and pITGB4BP-EGFP was a plasmid constructed in our laboratory. The construction method of pITGB4BP-EGFP is: design primers according to the sequence of the recombinant vector pACT2-ITGB4BP (liver cDNA library, BD Clontech, USA):

[0056] Upstream primer pITGB4BP-EGFP-EcoRI (inserted into the EcoRI restriction site):

[0057] 5'-CAGAATTCATGGCGGTCCGAGCTTCGTT-3';

[0058] Downstream primer pITGB4BP-EGFP-BamHI (inserted into BamHI restriction site):

[0059] 5'-CAGGATCCCGGTGAGGCTGTCAATGAGGGAAT-3'.

[0060] The PCR reaction was carried out with pACT2-ITGB4BP as the template, and the reagents were products of TaKaRa Company. The reaction system is as follows:

[0061] pACT2-ITGB4BP 1μl (about 1μg)

[0062] 10×PCR b...

Embodiment 2

[0109] Example 2. Homologous recombination of adenovirus

[0110] The successfully constructed shuttle vector in Example 1 was recovered after digested with Pme I, and simultaneously transformed into competent Escherichia coli BJ5183 containing the pAdEasy-1 plasmid (purchased from Qbiogene, USA) prepared by the calcium chloride method , complete the homologous recombination of adenovirus. The specific operation content is as follows:

[0111] 2.1 Linearization of the shuttle plasmid:

[0112]The successfully constructed shuttle plasmids pShuttle-CMV-ITGB4BP-EGFP and pShuttle-CMV-EGFP were digested and recovered with Pme I (purchased from NEB, UK). The respective enzyme digestion systems of the two are as follows:

[0113] pShuttle-CMV-ITGB4BP-EGFP or pShuttle-CMV-EGFP 18μl (about 0.5μg)

[0114] NEB buffer 4 5 μl

[0115] 100×BSA 0.5 μl

[0116] wxya 2 O 25.5 μl

[0117] Pme I 1 μl

[0118] Final volume 50μl

[0119] Reaction conditions: after 2 hours of reaction at...

Embodiment 3

[0129] Example 3. Packaging and amplification of adenovirus

[0130] 3.1 Adenovirus vector transfection HEK293 cells packaging adenovirus

[0131] 3.1.1 Linearization of adenoviral vectors:

[0132] Recombinant adenoviral vectors pAdEasy-ITGB4BP-EGFP and pAdEasy-EGFP were digested with Pac I and recovered. The enzyme digestion system is as follows:

[0133] pAdEasy-ITGB4BP-EGFP or pAdEasy-EGFP 48μl (about 1.0μg)

[0134] NEB buffer 1 8 μl

[0135] 100×BSA 0.8μl

[0136] wxya 2 O 21.7 μl

[0137] Pac I 1.5μl

[0138] Final volume 80μl

[0139] Reaction conditions: After reacting at 37°C for 1 hour, electrophoresis on 0.6% agarose gel, staining with Goldview nucleic acid dye showed bands of 23kb and 3.0kb or 23kb and 4.5kb respectively. Genes pAdEasy-ITGB4BP-EGFP and pAdEasy-EGFP) were used for gel recovery (the method is the same as 1.1 in Example 1).

[0140] 3.1.2 Packaging of adenovirus:

[0141] will be 3×10 5 Cells HEK293 cells (purchased from the Cell Bank of ...

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Abstract

The invention relates to the new applications of integrin beta 4 binding protein (ITGB4BP) and derivates thereof, or gene recombinant expression vector expressing the ITGB4BP or derivates thereof in preventing and / or treating hypertrophic scar or fibrosis lesion; wherein, the amino acid sequence of the ITGB4GB is SEQ ID NO 1; the derivates of the ITGB4BP are protein derivates retaining the combining site and the functional site of the ITGB4BP (SEQ ID NO 1). The application is realized by applying the ITGB4BP or derivates thereof, or gene recombinant expression vector expressing the ITGB4BP orderivates thereof, or the drug combination or the kit of the invention on subjects with an effective dose in treatment. The drug combination or the kit contains the ITGB4BP or derivates thereof, or gene recombinant expression vector expressing the ITGB4BP or derivates thereof and an officinal excipient or vectors and the like with an effective dose in treatment. Preferably, the subjects are humanbeings or animals.

Description

technical field [0001] The present invention relates to new pharmaceutical uses of known substances. Specifically, the present invention relates to β4 integrin binding protein (Integrin beta 4 binding protein, ITGB4BP) and its derivatives, gene recombinant expression vectors expressing ITGB4BP and its derivatives, in the prevention and / or treatment of hypertrophic scars and fibrotic lesions New apps in . Background technique [0002] Fibrotic diseases are a type of disease in which tissue hyperplasia, hardening and scar formation are caused by excessive deposition of extracellular matrix (ECM) such as collagen. Long-term stimuli of chronic inflammation such as persistent infection, autoimmune disease, allergic reaction, chemical injury, radiation, and tissue damage lead to the differentiation of fibroblasts into myofibroblasts, resulting in massive connective tissue deposition and fibrosis, resulting in loss of organ function until Death (1-3). Hypertrophic scar is a kind...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K38/17A61K48/00A61P17/02A61P11/00A61P13/12
CPCC07K14/4702A61K38/1709A61K48/00A61P11/00A61P13/12A61P17/02
Inventor 吴军谭江琳罗高兴贺伟峰彭旭
Owner SOUTHWEST HOSPITAL OF CHONGQING
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