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Application of deubiquitinating enzyme in preparation of medicine for preventing or treating acute graft versus host disease and graft anti-leukemia

A technique of graft-versus-host and deubiquitinating enzymes, which is applied in the field of biomedicine to achieve the effects of inhibiting proliferation, inhibiting development, and promoting secretion

Pending Publication Date: 2022-03-18
SUZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, its role in aGVHD has not been reported so far

Method used

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  • Application of deubiquitinating enzyme in preparation of medicine for preventing or treating acute graft versus host disease and graft anti-leukemia
  • Application of deubiquitinating enzyme in preparation of medicine for preventing or treating acute graft versus host disease and graft anti-leukemia
  • Application of deubiquitinating enzyme in preparation of medicine for preventing or treating acute graft versus host disease and graft anti-leukemia

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1CaCl 2 Preparation of DH5α Competent

[0039] Prepare 200ml 0.1M CaCl in a sterile ultra-clean bench 2 2 bottles each, pre-cooled on ice before use after autoclaving; take the DH5α monoclonal strains stored at -80°C, dip and smear on the sterilized solid LB medium without antibiotics, and store in a water-proof constant temperature incubator at 37°C Cultivate for 12-16 hours until colonies are formed; pick a single colony on solid LB medium, inoculate into 5ml of fresh LB medium after sterilization without antibiotics, place on a horizontal shaker at 37°C, shake at 250rpm overnight (12-16h); take 50μl of the above Put the bacterial liquid into 100ml of fresh LB medium after sterilization without antibiotics, place on a horizontal shaker at 37°C, shake at 250rpm for 3h; put the above bacterial liquid in an ice bath for 30min, pour it into a pre-cooled 50ml sterile centrifuge tube, and place it in an ice bath for 10min , 3500rpm, centrifuge at 4°C for 15min; di...

Embodiment 2

[0040] The extraction of embodiment 2 plasmid

[0041]Plasmid transformation: Take 50 μl of DH5α competent bacteria and immediately put it on ice to melt for 6-8 minutes; absorb 1 μl of the target plasmid and add it to the competent bacteria, blow it gently, and put it in an ice bath for 15-30 minutes; put it in a water bath / metal bath at 42°C , heat shock for 90s, put it on ice immediately, let it stand for 2-3min; add 200μl sterilized fresh LB medium, place it on a horizontal shaker at 37°C, shake at 250rpm for 30-45min; centrifuge at 3000rpm for 10min, suck off 100μl Clear, gently blow evenly and apply to after sterilization A + On a solid LB medium plate; after the bacterial solution is absorbed, place the LB culture plate in a constant temperature and humidity incubator at 37°C and incubate overnight until colonies are formed (12-16h).

[0042] Monoclonal amplification: Pick a single transformed colony and add it to 3ml of sterilized fresh LB medium containing ampicillin...

Embodiment 3

[0044] The extraction of embodiment 3 microcircle plasmids

[0045] The eukaryotic expression plasmid can be transformed into a microcircle plasmid after transformation and induction, which can be stably expressed in vivo without being degraded. It is injected into mice by the HGT method, resulting in stable and high expression of the target gene in the mouse, and because it does not contain bacterial DNA The backbone will not cause immunogenic reactions, and is suitable for in vivo experiments to construct mouse models for overexpression systems.

[0046] 3.1 Transformation of microcircle plasmids: Take 50 μl of ZYCY10P3S2T E.coli competent bacteria and immediately put them on ice to thaw for 6-8 minutes; absorb 1 μl of microcircle plasmids and add them to the competent bacteria, blow them gently, and put them in ice bath for 15-30 minutes; In a water bath / metal bath at 42°C, heat shock for 90s, put it on ice immediately, and let it stand for 2-3min; add 200μl sterilized fres...

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Abstract

The invention belongs to the field of biological medicines, and particularly relates to application of deubiquitinating enzyme BRCC3 in preparation of medicines for preventing or treating acute graft versus host disease and graft anti-leukemia. The invention discloses application of a deubiquitinating enzyme BRCC3 in preparation of a medicine for preventing and treating graft versus host disease through animal experiment research. The result shows that the BRCC3 has the effect of inhibiting the acute graft versus host disease after allogenic hematopoietic stem cell transplantation; the activation of donor T cells and the function of the T cells can be inhibited, and the proliferation of macrophages can be inhibited; through a proteasome way, ubiquitination modification of Trim25 is lowered, and the protein level of Trim25 is stabilized, so that secretion of IFN beta is promoted, and generation and development of aGVHD are inhibited; and meanwhile, the GVL effect is reserved, and the method has a relatively great application prospect.

Description

technical field [0001] The invention belongs to the field of biomedicine, and specifically relates to the application of deubiquitinating enzyme BRCC3 in the preparation of drugs for preventing or treating acute graft-versus-host disease and graft-versus-leukemia. Background technique [0002] Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is an important method for the treatment of malignant hematological diseases, some genetic diseases and bone marrow failure diseases ( figure 1 ), mainly through the donor-derived graft-mediated anti-leukemia response (graft-versusleukemia, GVL) plays a key role in the treatment of malignant hematological diseases and the prevention of relapse after transplantation. However, donor-derived T cells in these grafts can also induce graft-versus-host disease, especially acute graft-versus host disease (aGVHD), which is currently one of the most serious complications restricting allo-HSCT , is also a major cause of post-transpla...

Claims

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Application Information

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IPC IPC(8): A61K38/48A61P37/06A61P35/02
CPCA61K38/4813C12Y304/19012A61P37/06A61P35/02
Inventor 徐杨程巧吴德沛刘吟郑慧朱婷婷
Owner SUZHOU UNIV
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