Medicine for eliminating breast cancer cells with high efficiency and high specificity

A breast cancer cell and specific technology, applied in the field of medicine and biology, can solve the problem of not being able to kill breast cancer stem cells, and achieve the effect of no liver and kidney toxicity, exact drug effect, and low toxic and side effects

Active Publication Date: 2013-08-14
SUN YAT SEN UNIV CANCER CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Chemoradiation cannot kill breast cancer stem cells, leading to recurrence and metastasis

Method used

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  • Medicine for eliminating breast cancer cells with high efficiency and high specificity
  • Medicine for eliminating breast cancer cells with high efficiency and high specificity
  • Medicine for eliminating breast cancer cells with high efficiency and high specificity

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Example 1: Construction of T-VISA-BikDD therapeutic vector

[0018] (1) pCRII-TOPO-hTERT plasmid clone

[0019] 1. The DNA of breast cancer MCF-7 cells (purchased from ATCC) was extracted by conventional methods.

[0020] 2. Using the DNA as a template, design PCR primers for amplifying hTERT, hTERT PCR upstream (Forward) and downstream (Reverse) primers:

[0021] Forward: 5′-ata tct aga ggc ccc tcc ctc ggg tta ccc cac agc-3′(XbaI)

[0022] Reverse: 5'-ata gat ctt atg cgg ccg ccc acg tgc gca gca gga cgc agc gc-3'.

[0023] 3. Using breast cancer MCF-7 cell DNA as template, hTERT PCR primers (Forward: 5′-ata tct aga ggc ccc tcc ctc ggg tta ccc cac agc-3′; Reverse: 5′-ata gat ctt atg cgg ccg ccc acg tgc gca gca gga cgc agc gc-3′), Pfu DNA polymerase, dNTP and PCR reaction solution, amplify to obtain the hTERT promoter (-416 to+1) product, the sequence of which is shown in SEQ ID NO.2. The PCR reaction system: 10×PCR buffer 5μl, upstream (Forward) and downstream (Reverse) primers e...

Embodiment 2

[0079] Example 2: Preparation of liposomes:

[0080] Remove the lipid from the refrigerator (store DOTAP at -20°C, and store cholesterol at -4°C) and return to room temperature. Heat two rotary evaporators in a water bath to 30°C and 50°C respectively. Weigh 68.75mg of cholesterol and put it into a 1000ml round bottom flask. Add 100 mg DOTAP and 25 mg Choroform to the round bottom flask. Turn the flask to mix well. Rotate the round-bottom flask in a water bath at 30°C for 2 minutes to make it evenly mixed and form a film on the wall of the flask. Turn on the vacuum aspirator and leave it at 30°C for 30 minutes. Add 8.9ml of preheated 5% glucose solution to dissolve the dried film, and rotate it quickly at 105 rpm for 45 minutes at 50°C. Then reduce the temperature to 35°C and rotate for 10 minutes. Seal the flask with plastic wrap (or paraffin) and leave it at room temperature overnight to avoid light. Measure the volume and add double distilled water to 8.9ml. The flask ...

Embodiment 3

[0081] Example 3: Preparation of T-VISA-BiKDD liposomes

[0082] The T-VISA-BikDD treatment carrier was dissolved in a 5% glucose solution to make the final concentration 1ug / ul. At the same time, the liposomes of Example 2 at the storage concentration (20mM) were diluted with 5% glucose solution to the working concentration (8mM) . 1μg DNA / ul T-VISA-BikDD treatment carrier was slowly added to 8mM liposomes and allowed to stand at room temperature for 20 minutes to react to form T-VISA-BikDD liposomes.

[0083] Then check the quality control such as verification, biological characteristics test and endotoxin, aseptic packaging. The finished product is tested, packaged, and stored at 4-8°C; put it at room temperature for 20 minutes before use. It can be directly used for in vitro research or in vivo research, or it can be diluted with 5% glucose solution for use.

[0084] 2. Effect experiment:

[0085] 1. Biological characteristics of T-VISA-BiKDD liposomes:

[0086] The biological ch...

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Abstract

The invention discloses a medicine for eliminating breast cancer cells with high efficiency and high specificity. The medicine of the invention is capable of eliminating a T-VISA-BikDD therapy vector of breast cancer cells with high efficiency and high specificity, the sequence is shown as a SEQ ID NO.1. The T-VISA-BikDD liposome of the medicine for eliminating breast cancer cells with high efficiency and high specificity comprises liposome and the liposome-coated T-VISA-BikDD therapy vector, the nucleotide sequence of the T-VISA-BikDD therapy vector is shown as the SEQ ID NO.1. The T-VISA-BikDD therapy vector is coated by liposome and conveyed, and enriched on breast cancer position, is capable of targeting a breast cancer BCL-2 target site with high efficiency, and is combined with BCL-2 to release Bad and Bax for causing the apoptosis of the breast cancer cells, the medicine of the invention has no elimination effect to normal cells, enables systemic administration, has the advantages of high gene expression level, long period, high efficiency, exact drug effect, low toxic and side effect as well as no liver and kidney toxicity, and has wide prospect for treating breast cancer.

Description

Technical field: [0001] The invention belongs to the field of medical biology, and specifically relates to a therapeutic carrier and a medicine thereof that can efficiently and specifically kill breast cancer cells. Background technique: [0002] Breast cancer is the most common malignant tumor in women, and its incidence is increasing year by year, seriously endangering health. Surgery for breast cancer is mainly for early and mid-term breast cancer, and comprehensive treatment is generally required after surgery and late. Chemoradiation therapy cannot kill breast cancer stem cells, leading to recurrence and metastasis. After comprehensive treatment, the prognosis is still unsatisfactory, and the 5-year survival rate of patients with advanced stage is only 10%-30%. The prognosis of triple-negative breast cancer is worse. Breast cancer stem cells are the main cause of breast cancer recurrence, metastasis and drug resistance. Therefore, there is an urgent need to explore new d...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K48/00A61K9/127A61P35/00
Inventor 谢小明郭姣丽谢新华李来胜孔亚楠
Owner SUN YAT SEN UNIV CANCER CENT
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