Drug for diagnosing colon cancer and/or colon polyp, observing postoperative course and monitoring reoccurrence

A technology for colorectal polyps and colorectal cancer, applied in the field of diagnosing drugs and monitoring drugs, can solve the problems of increasing medical expenses and the like

Inactive Publication Date: 2007-12-26
PERSEUS PROTEOMICS INC +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

That is, there are problems such as tissue resection that burdens the patient, complex operations such as extraction of cysteine ​​protease inhibitor SN from tissue and electrophoresis, and increased medical expenses.

Method used

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  • Drug for diagnosing colon cancer and/or colon polyp, observing postoperative course and monitoring reoccurrence
  • Drug for diagnosing colon cancer and/or colon polyp, observing postoperative course and monitoring reoccurrence
  • Drug for diagnosing colon cancer and/or colon polyp, observing postoperative course and monitoring reoccurrence

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0108] Example 1 (cloning of cDNA of Cystatin SN)

[0109] In order to produce a monoclonal antibody against cystatin SN, its antigen was produced. First, cloning of the sequence containing the full-length ORF region of cystatin SN was attempted. The salivary gland single-stranded cDNA library, which is a cystatin SN-expressing tissue, was prepared in the same manner as above, and used as a template, using primers Cystatin SN-f (SEQ ID NO: 3) and Cystatin SN designed according to GenBank number (NM 001898) -r (SEQ ID NO: 4), isolate the full-length ORF gene by PCR method. Then, using the cystatin SN full-length sequence as a template, Cystatin SN-F (Hind) primers (SEQ ID NO. 5) and Cystatin SN-R (His-BamHI) primer (SEQ ID NO: 6), the target fragment was amplified again by PCR method, and inserted into phCMV vector (Stratagene). After performing base sequence analysis by a conventional method, the fragment cut at the HindIII and BamHI sites was inserted into the phCMV vector...

Embodiment 2

[0110] Embodiment 2 (preparation of cysteine ​​protease inhibitor SN antigen)

[0111] According to the experimental design of TransIT (TaKaRa Corporation), 1 × 10 5 Cell CHO cells were seeded in 6-well culture dishes and cultured overnight. On the next day, 8 μg of the expression vector phCMV-Cystatin SN-His and 16 μL of TransITreagent were mixed in 100 μL of serum-free DMEM medium, and incubated at room temperature for 20 minutes, and then added to cell culture medium for transfection. In addition, when using FUGENE6, according to the experimental design of Roche Diagnostics, 8×10 5 The CHO cells of the cell were inoculated in a 10cm culture dish and cultured overnight. On the next day, 8 μg of the expression vector phCMV-Cystat was used when using FUGENE6. According to the experimental design of Roche Diagnostics, 8×10 5 Cells in SN-His and 16 μL of FUGENE6 reagent were mixed in 400 μL of serum-free DMEM medium in a 10 cm dish, incubated at room temperature for 20 minutes...

Embodiment 3

[0113] Example 3 (production of cysteine ​​protease inhibitor SN monoclonal antibody)

[0114] Cystatin SN-His equivalent to 100 μg of protein suspended in PBS was mixed with Freund's complete adjuvant, and injected intraperitoneally into BALB / c mice to perform primary immunization. Similarly, the prepared Cystatin SN-His equivalent to 50 μg of protein and Freund's complete adjuvant were mixed and injected intraperitoneally, thereby implementing the second immunization. Cystatin SN-His equivalent to 50 μg of protein was injected intravenously for final immunization. Splenocytes were prepared from this mouse, and cell fusion with mouse P3U1 cells was carried out by a usual method using polyethylene glycol. Screening is performed by ELISA using Cystatin SN-His to produce specific binding antibodies.

[0115] Through this screening, monoclonal antibodies PPMX0201 and PPMX0202 with high specificity to the cysteine ​​protease inhibitor SN were successfully produced. Since PPMX020...

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Abstract

A method of diagnosing colon cancer and / or colon polyp and a method of observing the postoperative course and monitoring the reoccurrence thereof characterized by comprising detecting cystatin SN protein by using anti-cystatin SN antibody. Thus, a kit for assaying cystatin SN conveniently applicable to the diagnosis to be performed before the barium examination and the endoscopic examination, which have been performed hitherto and impose burdens to patients, the indications for metastasis and reoccurrence and the evaluation of therapeutic effects. As a result, colon cancer and / or colon polyp can be diagnosed and monitored by a convenient method and a new therapeutic program can be quickly designed thereby.

Description

technical field [0001] The present invention relates to the diagnosis of colorectal cancer and / or colorectal polyp, the postoperative follow-up and the monitoring method of recurrence, as well as the diagnostic medicine and monitoring medicine. Background technique [0002] In Japan, due to the Westernization of dietary life, colorectal cancer has a tendency to increase significantly. About 60,000 people suffer from it every year. It is estimated that it will surpass gastric cancer in 2015 and become the number one cancer in women, second only to lung cancer, lung cancer, and cancer in men. Liver cancer ranks third. It is speculated from epidemiology that, compared with genetic factors, the cause of colorectal cancer is diet, especially excessive intake of animal fat and protein. As the part of the large intestine, it is easy to develop in the sigmoid colon and rectum. [0003] There are (i) blood test, (ii) fecal occult blood test, (iii) barium meal test, (iv) endoscopy, (...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/574
CPCG01N33/57419
Inventor 油谷浩幸岛村隆浩渡边清高浅野岳晴大西真浜窪隆雄杉山晓细见直树岩成宏子石井敬介
Owner PERSEUS PROTEOMICS INC
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