Antineoplastic dibasic polypeptide and application and preparation method thereof

An anti-tumor and antigen technology, applied in anti-tumor drugs, chemical instruments and methods, medical preparations containing active ingredients, etc., can solve problems such as limiting the application value of antibodies, achieve specific targeting, and prevent poor prognosis.

Active Publication Date: 2011-06-08
姜荣锡
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This greatly limits the application value of these antibodies, causing drugs that kill tumor cells to also kill some normal cells

Method used

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  • Antineoplastic dibasic polypeptide and application and preparation method thereof
  • Antineoplastic dibasic polypeptide and application and preparation method thereof
  • Antineoplastic dibasic polypeptide and application and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1 Construction of plasmids expressing anti-tumor binary polypeptides and preparation of recombinant anti-tumor polypeptides

[0029] The original plasmid is the pET22b plasmid (plasmid size 7.6kb, provided by Harvard Univ.Dr.J.Collier) loaded with the wild-type Bacillus anthracis protein antigen gene SEQ ID NO. TM Kit, Strategene Company) after inserting the gene with six histidines-CACCACCACCACCACCAC- at the amino terminus, the genes encoding mutant Bacillus anthracis protein antigens SEQ ID NO.2, SEQ ID NO.4, SEQ ID NO.8 and wild type The B. anthracis protein antigen gene is operably linked to obtain the nucleotide sequences of SEQ ID NO. 6 and SEQ ID NO. 10 expressing the recombinant B. anthracis protein antigen gene. After respectively inserting the genes SEQ ID NO. 12 and SEQ ID NO. 18 encoding antibody mimics into the carboxyl-terminal G734 site of the recombinant Bacillus anthracis protein antigen gene, three recombinant plasmids of anti-tumor binary poly...

Embodiment 2

[0099] Example 2: Comparison of the toxic effects of the anti-tumor binary polypeptide of the present invention and wild-type Bacillus anthracis protein antigen on mice

[0100] The test mice were BALB / c immunodeficient nude mice inoculated with human non-small cell lung cancer cells (ATCC CCL-185) in the left armpit, and were divided into 5 experimental groups, each with 5 mice.

[0101] Test group 1: Intraperitoneal injection of wild-type Bacillus anthracis protein antigen / wild-type necrosis factor group, the injection amount was 2 μg / 2 μg / day.

[0102] Test group 2: Intraperitoneal injection of wild-type Bacillus anthracis protein antigen / wild-type necrosis factor containing lung cancer antibody mimic, the injection amount was 2 μg / 2 μg / day.

[0103] Test group 3: Intraperitoneal injection of mutant N682SD683K Bacillus anthracis protein antigen / wild-type necrosis factor containing lung cancer antibody mimic, the injection amount was 2 μg / 2 μg / day.

[0104] Test group 4: Th...

Embodiment 3

[0108] Example 3: Comparison of the anti-tumor binary polypeptide of the present invention and wild-type Bacillus anthracis protein antigen in vitro killing effects on human non-small cell lung cancer cells

[0109] Human non-small cell lung cancer cells: American ATCC standard cell line CCL-185.

[0110] Cell culture: Take out 0.1ml of the revived CCL-185 cell suspension, slowly add it to a petri dish containing 3ml of 1640 liquid medium (plus 10% serum) (dilution ratio is 1:30), mix well, put it at 37°C CO 2 Cultivated in an incubator. The test cells were divided into 3 groups, the first group was the blank group, that is, the blank preservation solution of recombinant anti-tumor polypeptide (10mMPB+0.2M NaCI phosphate buffer (pH8.0)) was added. The second group added 10 μg / ml of wild-type Bacillus anthracis protein antigen and 10 μg / ml of B. anthracis wild-type necrosis factor. The third group was the addition of 10 μg / ml of antitumor binary polypeptide 1 and 10 μg / ml of...

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Abstract

The invention provides a gene, recombinant plasmid and polypeptide for an anti-tumor binary polypeptide. The gene of the recombinant anti-tumor binary polypeptide is obtained by connecting in an operable way the gene of a coding antibody simulator with a recombinant bacillus anthraci protein antigen gene. The recombinant plasmid of the invention is formed by inserting the gene of the coding antibody simulator by double-chain oligomeric nucleotide directed mutagenesis method into the recombinant bacillus anthraci protein antigen gene. The obtained recombinant plasmid is infected into engineering bacillus coli BL-21 to get engineering bacillus coli cell of anti-tumor binary polypeptide; the anti-tumor binary polypeptide can be obtained by expanding the bacillus coli, settling in centrifugalway the bacillus coli body, crushing in altrasonic way, settling and crushing bacillus coli body by hi-speed centrifuging and treating the upper clean solution. The anti-tumor binary polypeptide is of special targeting characteristic, higher efficiency in killing special physical tumor than prior anti-tumor medicine, and will not attack normal cells, and has much lower toxicity and poor-reactionthan prior anti-tumor medicine.

Description

technical field [0001] The invention relates to a gene, recombinant plasmid and polypeptide of an anti-tumor binary polypeptide, and its application and preparation method. The tumor is human non-small cell lung cancer, small cell lung cancer or ovarian cancer. Background technique [0002] Malignant tumors such as non-small cell lung cancer, small cell lung cancer or ovarian cancer are a huge threat to human health. About seven million people die of malignant tumors in the world every year, of which China accounts for one-sixth. Existing anti-tumor drugs play an important role in tumor treatment. Although some tumors have achieved certain curative effects, they still have defects such as poor tumor cell selectivity, numerous and serious immunosuppression and adverse reactions, and drug resistance. [0003] In recent years, great progress has been made in basic and clinical research on tumors. One of the most important efforts to develop new anti-tumor drugs is to develop d...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/62C07K19/00A61K38/16A61P35/00
Inventor 丘小庆
Owner 姜荣锡
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