Anti-tumor medicine

An anti-tumor drug and drug technology, applied in the field of anti-tumor drugs, can solve problems such as affecting the interaction between proteins and DNA, inhibiting the binding efficiency of transcription factors and DNA in the promoter region, etc.

Active Publication Date: 2006-08-23
SHEN ZHEN TSINGHUA YUANXING BIO PHARM SCI & TECHNOL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Methylation leads to changes in the conformation of DNA in some regions, which affects the interaction between proteins and DNA, and inhibits the binding efficiency of transcription factors to DNA in the promoter region

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Example 1: Survivin gene is expressed at a high level in NCI-H460 cells

[0041] Human non-small cell lung cancer (Non-small Cell Lung Cancer, NSCLC) cell line NCI- H460, grow to the logarithmic growth phase (the density is about 75% in the environment of 37°C and 5% CO2, the living cell rate is > 95% as detected by the trypan blue exclusion test), after the cells are digested with 0.25% trypsin, use Collect in sterile PBS (pH 7.4), extract nucleic acid, dissolve in 100 μ; 1 DEPC-H2O, measure its absorbance at 260nm, and calculate RNA concentration (A260×40 (μg / μl)).

[0042] Reverse transcription: use RNA extract (RNA concentration 0.025~0.125μg / μl) as a template, add buffer, DNase I (5U / μl), RNase inhibitor (40U / μl) and DEPC-H2O, then 37°C for 45 minutes and 75°C for 10 minutes, after cooling on ice, add buffer, DTT (100mM), dNTP mixture (10mM each), Oligo(dT)18 primer (0.1μg / μl), random primer ( 0.1 μg / μl), RNase inhibitor and MMuLV reverse transcriptase (200U / μl)....

Embodiment 2

[0045] Example 2: Surkex inhibits the expression of Survivin mRNA in NCI-H460

[0046]Transfection: NCI-H460 cells in exponential growth phase (density about 75%, tested by trypan blue exclusion test, viable cell rate > 95%) were digested with 0.25% trypsin, collected with culture medium, and placed in 24-well The plate was plated at a concentration of 105 cells / well, and incubated overnight at 37°C.

[0047] SurKex was dissolved in sterile PBS (pH 7.4) and filtered through a syringe filter. Add 100 μl of Opti-MEM medium at room temperature to Eppendorf tube A, then add 3 μl of GeneJammer transfection reagent, mix well and place at room temperature for 5 minutes. Add 135 μl of Opti-MEM medium at room temperature to Eppendorf tube B, add 15 μl of SurKex in PBS, and mix well. Mix tubes A and B, repeatedly pump and mix, and place at room temperature for 30 minutes. See Table 1 for SurKex transfection concentrations.

[0048] Well No.

drug

SurKex Conc.in...

Embodiment 3

[0073] Example 3: Surkex inhibits the expression of Survivin protein in NCI-H460

[0074] transfection

[0075] NCI-H460 cells in the exponential growth phase were plated in a 6-well plate at a concentration of 5105 cells / well.

[0076] Add 200 μl Opti-MEM medium and 6 μl GeneJammer transfection reagent to Eppendorf tube A, and add 270 μl Opti-MEM medium and 30 μl SurKex in PBS solution to Eppendorf tube B. For transfection, add 500 μl of tube A and tube B mixture and 500 μl of Opti-MEM medium to each well. After culturing at 37°C for 12 hours, 1 ml of RPMI 1640 medium containing 20% ​​FBS was added to each well, and the culture was continued at 37°C for 60 hours.

[0077] The SurKex transfection concentration is shown in Table 4, and the rest of the operations are the same as in Example 2.

[0078] Well No.

drug

SurKex Conc.in Tube B

(μM)

SurKex Conc.Final

(μM)

1

2

3

PBS (Control)

SurKex 1

SurKex 2

0...

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Abstract

The present invention provides a kind of medicine component capable of inhibiting the growth of lung cancer tumor through inducing the methylation of apoptosis inhibition factor. The medicine component includes oligonucleotides sequences Surkex 1 and Surkex 2 complementary with the partial promoter region sequence of apoptosis inhibition factor Survivin. The complementary combination of the sequences Surkex 1 and Surkex 2 with Survivin can reduce the expression of apoptosis inhibition factor Survivin to make tumor cell die and treat lung cancer. The action site is in the Survivin promoter region, and this is favorable to clinical medicine administration.

Description

technical field [0001] The invention relates to an antitumor drug, in particular to an antisense drug, the drug component acts on the promoter region of the Survivin gene, and silences the apoptosis inhibitor Survivin gene through methylation to achieve the effect of treating lung cancer. Background technique [0002] In recent years, in some countries in the world, especially industrially developed countries, the incidence of lung cancer has gradually increased, especially in women. Lung cancer kills more people each year than breast, prostate and bowel cancer combined. In 2002, more than 1.3 million people worldwide were diagnosed with lung cancer, and more than 1.1 million people died of lung cancer. In the United States, lung cancer ranks first among cancers, accounting for about 28.6% of all cancer deaths. In my country, lung cancer is also the number one cancer. The first China North-South Summit Forum on Lung Cancer held on September 24-26, 2004 predicted that from...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K31/7088A61P35/00
Inventor 周向军黄玮琳王永祥潘武宾
Owner SHEN ZHEN TSINGHUA YUANXING BIO PHARM SCI & TECHNOL
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