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Tubulin depolymerizing agent polypeptide and application thereof

A technology of tubulin and depolymerizing agent, which is applied in the field of tubulin depolymerizing agent polypeptide and its application, can solve the problem of lack of specificity of small molecule depolymerizing agent, and achieve the effect of increasing survival rate and improving survival rate

Active Publication Date: 2013-08-28
BEIJING SHENLANHAI BIO PHARM TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Many successful small-molecule depolymerization agents can inhibit the activity of tubulin at the nanomolar level, but small-molecule depolymerization agents lack specificity. If long-term use of tubulin depolymerization agents lacking specificity in chronic diseases may cause produce major side effects

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Effect of tubulin depolymerizer polypeptide 2 on tubulin polymerization and depolymerization in vitro.

[0018] Take fresh bovine brain tissue, peel off the meninges and large blood vessels, cut into pieces, wash 1-2 times with cold MES buffer, add MES buffer at a ratio of 0.5-1ml per gram of brain tissue, at 4°C, use electric Homogenize with a homogenizer; centrifuge at 105,000 g for 1 hour at 4°C, take the supernatant, add an equal volume of microtube polymerization buffer, and incubate in a water bath at 37°C for 30 minutes. Centrifuge at 105,000g at 26°C for 1 hour, take the precipitate, add about 1 / 10 of the homogenate volume of cold MES buffer, stir gently or crush the precipitate with a homogenizer; place the suspension in an ice bath for 30 minutes to dissolve the precipitate completely. Protein content was determined by Lowry's method (SDS polyamide gel electrophoresis). Tubulin was diluted to 4-5mg / ml with MES buffer and stored in liquid nitrogen. Take the f...

Embodiment 2

[0022] IC50 of tubulin depolymerizer polypeptide 2 on the growth and survival of tumor cells cultured in vitro.

[0023] The MTT colorimetric method was used. The logarithmic growth of U937 cells was added to a 96-well culture plate at 1.0×105, and cultured for 24 hours. Different concentrations of the experimental drug tubulin depolymerizer polypeptide 2 and the positive control drug Changchunxin were added to the experimental wells and positive drug control wells, respectively. base; add the same volume of solvent to the blank group. Set up five duplicate wells per well, culture for 48h, add MTT to each well at 0h, 2h, 8h, 14h, 20h, 24h, 36h, and 48h respectively, add DMSO after 4h of action, incubate for 30min, and place at 620nm in a microplate reader Measure the absorbance A value, according to the formula tumor cell growth inhibition rate=(1-absorbance value of the experimental group / absorbance value of the control group)×100%. The calculated IC50 of the test drug was ...

Embodiment 3

[0025] In vivo activity of tubulin depolymerizer polypeptide 2 was tested in a tumor model.

[0026] The U937 tumor model was established, and the positive control drug vincristine was added; the same volume of solvent was added to the blank group, and three doses were set in the experimental group: 0.75, 1.5 μM, and 3 μM mg / Kg. After 21 days, the number of surviving mice was observed, and the survival rate was calculated. The results showed that the tubulin depolymerizing agent polypeptide 2 could effectively protect mice and improve the survival rate of tumor-bearing mice, with the survival rate reaching 87.2%.

[0027]

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PUM

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Abstract

The invention discloses a tubulin depolymerizing agent polypeptide and an application thereof, relating to the field of medicines and particularly relating to a polypeptide which has a function of suppressing tubulin polymerization and is used for treating acute lymphocytic leukemia. The sequence of the tubulin depolymerizing agent polypeptide is MFSPSCVSLLRCTGCC which is a brand new sequence; and the tubulin depolymerizing agent polypeptide can suppress the tubulin polymerization in vitro at a 1mmol level and increase the survival rate of tumor-bearing mice during in vivo tests, and has potential values in new medicine development.

Description

technical field [0001] The invention relates to tubulin depolymerizing agent polypeptide 2 and its application, in particular to a polypeptide capable of inhibiting tubulin polymerization and treating acute lymphocytic leukemia. Background technique [0002] Acute lymphoblastic leukemia (ALL) is a progressive malignant disease characterized by large numbers of immature white blood cells that resemble lymphoblasts. These cells can be found in the blood, bone marrow, lymph nodes, spleen and other organs. Acute lymphoblastic leukemia accounts for 80% of acute leukemia in children, and the peak incidence is between 3 and 7 years old. ALL can also occur in adults and accounts for 20% of all adult leukemias. In recent years, with the deepening of medical research, great progress has been made in the understanding and treatment of acute lymphoblastic leukemia. Among them, tubulin plays an important role in acute lymphoblastic leukemia. [0003] Microtubules are the main compone...

Claims

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Application Information

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IPC IPC(8): C07K7/08A61K38/10A61P35/02
Inventor 罗瑞雪
Owner BEIJING SHENLANHAI BIO PHARM TECH
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