CN109797218B - Application of integrin beta 4 in preparation of reagent or medicine for distinguishing colon cancer from rectal cancer - Google Patents
Application of integrin beta 4 in preparation of reagent or medicine for distinguishing colon cancer from rectal cancer Download PDFInfo
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- CN109797218B CN109797218B CN201711148747.1A CN201711148747A CN109797218B CN 109797218 B CN109797218 B CN 109797218B CN 201711148747 A CN201711148747 A CN 201711148747A CN 109797218 B CN109797218 B CN 109797218B
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Abstract
The invention discloses an application of integrin beta 4 in distinguishing colon cancer from rectal cancer, which comprises the following steps: 1) screening for proteins whose expression is altered in colorectal cancer tissue of the patient compared to normal tissue; 2) screening integrin beta 4 by the step 1); 3) comparing the amount of integrin beta 4 expression in the colon cancer tissue to the amount of integrin beta 4 expression in the rectal cancer tissue; and 4) determining a threshold range of integrin beta 4 expression that distinguishes between rectal and colon cancer; thereby filling the blank of distinguishing the fields of colon cancer and rectal cancer by a molecular biological detection means, and further better distinguishing the tumor sources of the colon cancer and the rectal cancer.
Description
Technical Field
The invention relates to the field of biotechnology, in particular to application of integrin beta 4 in distinguishing colon cancer from rectal cancer.
Background
Large bowel cancer is one of the common digestive system malignancies, which includes colon and rectal cancers. The 3 rd place and the 2 nd place of the malignant tumor incidence spectrum of men and women all over the world. In recent years, under the influence of dietary structure and life style, the incidence of large intestine cancer in China, especially in big cities, is obviously increased, and the incidence of malignant tumor is leaped 2 nd, so that the prevention and treatment work is severe. Early colorectal cancer often has no obvious symptoms, and is often overlooked to delay early diagnosis and treatment opportunities. The world health organization proposes that cancer screening and early diagnosis and early treatment are the most effective ways for cancer prevention and control. Therefore, screening colorectal cancer of urban resident high-risk people and realizing early diagnosis and early treatment are effective measures for reducing the death rate of colorectal cancer and improving the survival rate of patients, and are hot problems which need to be solved urgently in the current colorectal cancer prevention and treatment work.
At present, the diagnosis of colon cancer or rectal cancer is limited to enteroscopy or visual observation of the tumor occurrence part for differentiation, and no clear molecular biological detection differentiation method is available. There is no effective way to clearly distinguish the origin of a tumor that occurs at the junction of the rectum and sigmoid colon. Because the advanced colon cancer patient and the advanced rectal cancer patient have different treatment scheme choices, namely the advanced rectal cancer patient or the patient with surgical contraindication can be selected for radiotherapy, and a better effect can be obtained; however, patients with colon cancer are more likely to be treated with chemotherapy, and thus have better therapeutic effects. Therefore, the tumor sources (rectum and colon) can be better distinguished, and especially the tumor sources at the junction of the rectum and the B are significant for the selection of the treatment scheme.
Integrins belong to the family of transmembrane protein cell adhesion molecules, usually heterodimeric structures consisting of two alpha and beta subunits, with a total of 18 alpha and 8 beta subunits in mammals, which mediate cell adhesion primarily through binding to ligands (collagen, laminin, fibronectin). Integrin beta 4(ITGB4) is a subtype of the beta subunit of the integrin family.
The information disclosed in this background section is only for enhancement of understanding of the general background of the invention and should not be taken as an acknowledgement or any form of suggestion that this information forms the prior art already known to a person skilled in the art.
Disclosure of Invention
The invention aims to provide application of integrin beta 4 in preparation of a reagent or a medicine for distinguishing colon cancer from rectal cancer, so that the blank of distinguishing the fields of the colon cancer and the rectal cancer by a molecular biological detection means is filled, and further, the tumor sources of the colon cancer and the rectal cancer are better distinguished.
To achieve the above objects, the present invention provides use of integrin beta 4 for the preparation of a reagent or a medicament for differentiating colon cancer and rectal cancer, comprising the steps of:
1) screening for proteins whose expression is altered in colorectal cancer tissue of the patient compared to normal tissue;
2) screening integrin beta 4 by the step 1);
3) comparing the amount of integrin beta 4 expression in the colon cancer tissue to the amount of integrin beta 4 expression in the rectal cancer tissue; and
4) a threshold range of integrin beta 4 expression was determined to distinguish between rectal and colon cancer.
In another embodiment of the above application, the expression level of integrin beta 4 is the relative expression level of mRNA measured by real-time fluorescent quantitative PCR.
In another embodiment, the base sequences of the upstream and downstream primers used in the real-time fluorescent quantitative PCR are as follows:
f: 5'-TCTCTCAGAGTGAGCTGGCAG-3' is SEQ ID: 1;
r: 5'-TTCAGCAGCTGGTACTCCAC-3' is SEQ ID: 2.
In another embodiment, the internal reference used in real-time fluorescent quantitative PCR is GAPDH.
In another embodiment, the method for calculating the relative expression amount of mRNA used in real-time fluorescence quantitative PCR comprises
Method of, i
In another embodiment of the above application, the threshold range is 0.005-0.03.
In another embodiment of the above application, the threshold range is 0.008-0.02.
In another embodiment, the sensitivity and specificity of the ROC curve analysis threshold range is plotted by the SPSS software.
In another embodiment, if the expression level of integrin beta 4 is higher than the threshold range, colon cancer is determined, and if the expression level is lower than the threshold range, rectal cancer is determined.
Compared with the prior art, the invention has the following beneficial effects:
by using a gene chip database, a protein chip detection technology and an international advanced mass flow cytometry technology, 1 new tumor marker integrin beta 4(ITGB4) which has important value for early diagnosis and early treatment of colorectal cancer is successfully screened out from 60 ten thousand indexes, and by preliminary verification of small sample histology, the ROC curve analysis proves that ITGB4 has important value for distinguishing colorectal cancer tissues, so that the blank of distinguishing the colorectal cancer and the rectal cancer fields by a molecular biological detection means is filled, the colorectal cancer tumor sources at a direct junction are better distinguished, and the method has important significance for selecting a radiotherapy scheme; and the application and selection of anti-angiogenesis drugs such as integrin monoclonal antibody, cilengitide and the like can be directly influenced by determining whether the ITGB4 is expressed or not.
In addition, for the patients with advanced stage, the selection of integrin monoclonal antibody and the selection of the anti-vascular treatment drug are of great value.
Drawings
FIG. 1 is the relative mRNA expression levels of integrin beta 4 for rectal and colon cancers according to the present invention.
FIG. 2 is a ROC curve for sensitivity and specificity of assay thresholds according to the present invention.
Detailed Description
The following detailed description of the present invention is provided in conjunction with the accompanying drawings, but it should be understood that the scope of the present invention is not limited to the specific embodiments.
Example 1 protein screening
The distribution sites of Human proteins or polypeptides were analyzed by Human Genecards (http:// www.genecards.org /) and the Compertments database (https:// comppertments. jensenlab. org/Downloads) to obtain 4433 secreted proteins; through analysis of a BioGPS GeneAtlas U133A (http:// BioGPS. org/dataset/GSE 1133/geneeatlas-U133 a-gcrma /) database and an NC160on U133A (http:// BioGPS. org/dataset/BDS-00011/nci 60-on-U133a-gcrma /) database, it was found that 23 secreted proteins including integrin beta 4(ITGB4, CD104) were significantly changed in expression in colorectal cancer tissues (compared with normal colon tissues) and in colorectal cancer cell lines (compared with other tissue-derived cells).
Example 2 determination of threshold Range of integrin beta 4 expression
The ITGB4 mRNA expression of the colon cancer, the rectal cancer and the corresponding paracarcinoma tissues collected by the first hospital of Hebei medical university is analyzed by a real-time quantitative PCR method (ABI7500 model), and PCR conditions are as follows: 10 seconds at 95 ℃, 30 seconds at 60 ℃, 30 seconds at 72 ℃ and 42 cycles; and (3) primer F: TCTCTCAGAGTGAGCTGGCAG, R: TTCAGCAGCTGGTACTCCAC, respectively; the internal reference is GAPDH, and the relative expression calculation method comprises
The difference between normal tissue and cancer tissue was not significant, and when further analyzed, it was occasionally found that there was a significant difference in the expression of the normal tissue in colon and rectal cancers, p<0.05。
After the verification of 16 colorectal cancer tissues and 29 colon cancer tissues, when the threshold value is determined to be 0.01fold, the sensitivity and specificity of the threshold value for distinguishing the colon cancer from the rectal cancer are proved to be 0.759 and 0.750 in sequence by drawing an ROC curve through SPSS 21.0 software, the area under the curve is 0.763, and the result is detailed in the ROC curve in the attached figure 2.
The foregoing descriptions of specific exemplary embodiments of the present invention have been presented for purposes of illustration and description. It is not intended to limit the invention to the precise form disclosed, and obviously many modifications and variations are possible in light of the above teaching. The exemplary embodiments were chosen and described in order to explain certain principles of the invention and its practical application to enable one skilled in the art to make and use various exemplary embodiments of the invention and various alternatives and modifications as are suited to the particular use contemplated. It is intended that the scope of the invention be defined by the claims and their equivalents.
Sequence listing
<110> Hospital of Hebei medical university
<120> use of integrin beta 4 for differentiating between colon and rectal cancers
<130> P171597DD1F
<160> 2
<170> SIPOSequenceListing 1.0
<210> 1
<211> 21
<212> DNA
<213> Artificial sequence (SEQ ID:1)
<400> 1
tctctcagag tgagctggca g 21
<210> 2
<211> 20
<212> DNA
<213> Artificial sequence (SEQ ID:2)
<400> 2
ttcagcagct ggtactccac 20
Claims (6)
1. Use of integrin beta 4 in the manufacture of a reagent or medicament for distinguishing between colon and rectal cancer comprising the steps of:
1) analyzing distribution positions of Human proteins or polypeptides by using Human Genecards and components databases to obtain 4433 secreted proteins, and analyzing by using a BioGPS Geneatlas U133A database and an NC160on U133A database to obtain that the expression quantity of the 23 secreted proteins in colorectal cancer tissues is changed compared with normal colon tissues and in colorectal cancer cell strains compared with other tissue-derived cells; screening for proteins whose expression is altered in colorectal cancer tissue of the patient compared to normal tissue;
2) screening integrin beta 4 by the step 1);
3) comparing the amount of integrin beta 4 expression in the colon cancer tissue to the amount of integrin beta 4 expression in the rectal cancer tissue; and
4) determining a threshold range of integrin beta 4 expression that distinguishes between rectal and colon cancer, said threshold range being 0.005-0.03; the integrin beta 4 expression level is mRNA relative expression level measured by real-time fluorescence quantitative PCR; the base sequences of the upstream and downstream primers adopted by the real-time fluorescent quantitative PCR are as follows in sequence:
f: 5'-TCTCTCAGAGTGAGCTGGCAG-3' is SEQ ID: 1;
r: 5'-TTCAGCAGCTGGTACTCCAC-3' is SEQ ID: 2.
2. Use of integrin beta 4 in the preparation of a reagent or drug for the differentiation of colon and rectal cancer according to claim 1, characterized in that the internal reference used for real-time fluorescent quantitative PCR is GAPDH.
3. Use of integrin beta 4 for the preparation of a reagent or drug for the differentiation of colon and rectal cancer according to claim 1, characterized in that the relative mRNA expression calculation method used for real-time fluorescence quantitative PCR is 2－△C TA method.
4. Use of integrin beta 4 in the preparation of a reagent or drug for the differentiation of colon and rectal cancer according to claim 1, characterized in that the threshold range is 0.008-0.02.
5. Use of integrin beta 4 in the preparation of a reagent or drug for the differentiation of colon and rectal cancer according to claim 1, characterized by sensitivity and specificity for ROC curve analysis threshold range plotted by SPSS software.
6. Use of integrin beta 4 in the manufacture of a reagent or medicament for distinguishing between colon and rectal cancer according to claim 1, wherein integrin beta 4 is expressed in an amount above the threshold range for a determination of colon cancer and below the threshold range for a determination of rectal cancer.
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| CN110257524A (en) * | 2019-08-01 | 2019-09-20 | 浙江大学 | It is a kind of distinguish colorectal cancer cancerous tissue and Carcinoma side normal tissue colorectal cancer discrimination model and its construction method |
Non-Patent Citations (4)
| Title |
|---|
| Integrin α6β4 in colorectal cancer;Jean-Fran等;《World Journal of Gastrointestinal Pathophysiology》;20101231;第1卷(第1期);全文 * |
| 结直肠癌中Ets-1和整合素α6β4的表达及临床意义;赵承梅等;《中国肿瘤临床》;20111231;第38卷(第5期);全文 * |
| 结直肠癌组织中ITGβ-4表达及其临床意义;宦大为等;《山东医药》;20161231;第56卷(第24期);全文 * |
| 转录因子NFAT和整合素α6β4信号转导通路与大肠癌侵袭转移的关系;张坤;《中国优秀博硕士学位论文全文数据库(硕士)》;20060915(第09期);第8页第1段、第10-25页，四、实时定量PCR检测mRNA表达水平、第35-36页，六、大肠癌整合素β4亚基表达结果、第42页第3段第1-2行第14页，5.NFTA、Integrinα6β4 、GAPDH基因引物设计，表2 * |
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