Genetic medicine for preventing and treating cancer of colon and rectum, preparation process and use thereof

A technology of gene medicine and colorectal cancer, applied in gene therapy, drug combination, pharmaceutical formula, etc., can solve the problems that the treatment methods are difficult to treat effectively, and achieve the effect of reducing toxic side effects and enhancing curative effect

Inactive Publication Date: 2007-05-23
许瑞安 +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0012] For patients with advanced colorectal cancer, the tumor has invaded the muscularis mucosa, the whole thickness of the intestinal wall, and even lymph node metastasis has occurred, and the existing treatment methods are difficult to treat effectively
So far, no rAAV hybrid vector has been used to mediate kallistatin or vasostatin for the treatment of colorectal cancer. Our study provides a new method for the treatment of colorectal cancer

Method used

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  • Genetic medicine for preventing and treating cancer of colon and rectum, preparation process and use thereof
  • Genetic medicine for preventing and treating cancer of colon and rectum, preparation process and use thereof
  • Genetic medicine for preventing and treating cancer of colon and rectum, preparation process and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Preparation of rAAV7 / 8-kallistatin

[0047] Human kallistatin full-length cDNA was amplified from human liver first-strand cDNA by PCR. The exclusive primer for amplifying kallistatin is Kalli-F(5’-AA GAATTC GAGGATGCATCTTATCGAC) and Kalli-R(5’-AA GGTA CCAAGCTT CTATGGTTTCGTGGGGTC). Restriction sites (underlined) were introduced to facilitate subcloning. The PCR conditions were 94°C, 50°C and 68°C for 45 seconds each, with a total of 36 cycles. The PCR product was sequenced to verify its correctness, and subcloned into the AAV-2 expression vector to obtain the rAAV2-kallistatin expression plasmid. The promoter of the expression plasmid is CMV enhancer / chicken β-actin promoter. To enhance expression, after inserting the gene, WPRE elements were also introduced.

[0048] Recombinant virus was prepared by calcium phosphate helper-free virus multi-plasmid method: the expression plasmid, AV helper plasmid, and AAV7 helper plasmid and AAV8 helper plasmid were co-transfected int...

Embodiment 2

[0050] Preparation of rAAV7 / 8-vasostatin

[0051] Human vasostatin full-length cDNA was amplified from human liver first-strand cDNA by PCR. The exclusive primer for amplifying vasostatin is Vas-F(5’-AA CTCGAG CC CGCCATGCTG CTATCC) and Vas-R(5’-AA AAGCTT CTAGTTGTCTGG CCGCACAAT). Restriction sites (underlined) were introduced to facilitate subcloning, and a stop codon (in bold) was introduced in the primer Vas-R. The PCR conditions were 94°C, 50°C and 68°C for 45 seconds each, with a total of 36 cycles. The PCR product was sequenced to verify its correctness, and subcloned into the AAV-2 expression vector to obtain the rAAV2-kallistatin expression plasmid. The promoter of the expression plasmid is CMV enhancer / chicken β-actin promoter. To enhance expression, after inserting the gene, WPRE elements were also introduced.

[0052] Recombinant virus was prepared by calcium phosphate helper-free virus multi-plasmid method: the expression plasmid, AV helper plasmid, and AAV7 helper plas...

Embodiment 3

[0054] Preparation of recombinant virus injection

[0055] Take the recombinant virus rAAV7 / 8-kallistatin 2×10 prepared by the method in Example 1 15The virus titer is obtained by adding normal saline for injection to 1000ml, sterile filtration, and aliquoting into 2ml vials.

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Abstract

The invention relates to a gene drug based on genes as kallistatin and vasostatin, to treat carcinoma of colon, wherein it also provides relative application to improve treatmene effect. The carrier of genes is adenovirus (AAV), while the best one is AAV7 / 8. The inventive drug can be made into injection or dried powder, and it can be injected into muscle.

Description

Technical field [0001] The present invention relates to the fields of molecular pharmacology, molecular medicine and disease prevention and treatment, and in particular to two gene drugs for the treatment of colorectal cancer based on genes kallistatin and vasostatin, a preparation process, and the use of the drug for the treatment of colorectal cancer and the enhancement of colorectal cancer Application of cancer chemotherapy or radiotherapy curative effect. Background technique [0002] Colorectal cancer is one of the common malignant tumors, and its incidence is the third most common tumor in the world. In Europe and the United States, the incidence of colorectal cancer ranks second, second only to lung cancer (male) and breast cancer (female). At present, the number of new cases of colorectal cancer exceeds 900,000 per year, and the number of deaths is 500,000 per year. [0003] The cause of colorectal cancer is a combination of genetic factors and environmental factors. Gene...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61P35/00
Inventor 许瑞安冯戬云朱健华
Owner 许瑞安
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