CN103529220A - Applications of CTx to preparation of medicaments for diagnosing colorectal cancer - Google Patents
Applications of CTx to preparation of medicaments for diagnosing colorectal cancer Download PDFInfo
- Publication number
- CN103529220A CN103529220A CN201310476530.9A CN201310476530A CN103529220A CN 103529220 A CN103529220 A CN 103529220A CN 201310476530 A CN201310476530 A CN 201310476530A CN 103529220 A CN103529220 A CN 103529220A
- Authority
- CN
- China
- Prior art keywords
- colorectal cancer
- ctx
- diagnosis
- medicine
- collagen
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Images
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/94—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving narcotics or drugs or pharmaceuticals, neurotransmitters or associated receptors
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
- G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/06—Gastro-intestinal diseases
- G01N2800/065—Bowel diseases, e.g. Crohn, ulcerative colitis, IBS
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/52—Predicting or monitoring the response to treatment, e.g. for selection of therapy based on assay results in personalised medicine; Prognosis
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/56—Staging of a disease; Further complications associated with the disease
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Molecular Biology (AREA)
- Chemical & Material Sciences (AREA)
- Biomedical Technology (AREA)
- Urology & Nephrology (AREA)
- Hematology (AREA)
- Immunology (AREA)
- Cell Biology (AREA)
- General Health & Medical Sciences (AREA)
- Biotechnology (AREA)
- Pathology (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- Microbiology (AREA)
- General Physics & Mathematics (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Pharmacology & Pharmacy (AREA)
- Peptides Or Proteins (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention relates to the technical field of biology, and especially relates to applications of CTx to preparation of medicaments for diagnosing colorectal cancer, wherein CTx is a fragment at the carboxyl telopeptide of an extracellular matrix component type I collagen. The invention provides applications of CTx to prepare and screen medicaments for diagnosing colorectal cancer and screen medicaments for treating colorectal cancer. Colorectal cancer tissues and serum clinic specimens are selected for researching the clinic value of type I collagen on generation, development, staging prognosis and the like of colorectal cancer. Through a large number of experiments, inventors of the invention discover that the expression level change of CTx in serums of colorectal cancer patients at different stages is in positive correlation with pathology staging, and compared with conventional alimentary canal tumour oncofetal antigens, CTx is capable of better differencing the metastatic colorectal cancer patients and the non-metastatic colorectal cancer patients. Additionally, the inventors also discover that CTx is correlated with 3-year tumor-free survival of the colorectal cancer patients, and the high expression of CTx indicates poor tumor-free survival and prognosis of the colorectal cancer patients.
Description
Technical field
The present invention relates to biological technical field, the purposes of the c-terminal peptides CTx that particularly relates to a kind of extracellular matrix components type i collagen in preparing Colorectal Cancer Diagnosis medicine.
Background technology
Collagen is as extracellular matrix (Extracellular matrix, ECM) one of main composition, in most of connective tissues, for the structural stability that maintains ECM, play an important role, to many fiber diseases as relevant (Lai such as liver fibrosis, K.K., Shang, S., Lohia, N., etc.Extracellular matrix dynamics in hepatocarcinogenesis:a comparative proteomics study of PDGFC transgenic and Pten null mouse models.PLoS Genet., 2011, 7 (6): e1002147.).Recent report also points out that collagen is for the invasion and attack and transfer also extremely important (Sund, M., Kalluri of tumour cell, R.Tumor stroma derived biomarkers in cancer.Cancer Metastasis Rev., Jun, 2009,28 (1-2): 177-83.).Be reported at present and in human body, had 28 kinds of collagens and exist or may exist, and wherein type i collagen is people's in-vivo content the widest a kind of collagens that distribute at most.
Type i collagen is by two α 1 chains and a triple helix structure that α 2 chains form, and generally by fibroblast, is synthesized, and as a part for tumor microenvironment, in tumor cell proliferation, transfer process, plays a significant role.In colorectal cancer research, find, type i collagen high expressed is (Oku Y in presenting the tumour cell that infiltrates growth, Shimoji T, Takifuji K, etc.Identification of the molecular mechanisms for dedifferentiation at the invasion front of colorectal cancer by a gene expression analysis.Clin Cancer Res, 2008,14:7215 – 7222.).In clone, can reduce in vitro E-cadherin and β-catenin expression declines cell and intercellular adhesion, impel cell to occur stem-like cell phenotype (SC Kirkland.Type I collagen inhibits differentiation and promotes a stem cell-like phenotype in human colorectal carcinoma cells.Br J Cancer.2009, 101 (2): 320-6.), thereby and can reduce Cdx2 and express inhibition Cell Differentiation (Brabletz, T., Spaderna, S., Kolb, J., etc.Down-regulation of the homeodomain factor Cdx2in colorectal cancer by collagen type I:an active role for the tumor environment in malignant tumor progression.Cancer Res., 2004, 64 (19): 6973-7.).In addition, type i collagen can be by pernicious transformation (the Grzesiak J with a2 β 1integrin effect promotion pancreatic cancer cell, Bouvet M.The a2 β 1integrin mediates the malignant phenotype on type I collagen in pancreatic cancer cell lines.Br J Cancer, 2006, 94:1311 – 1319), be also simultaneously 17 with one of the closely-related gene of metastases (Ramaswamy S, Ross KN, Lander ES, etc.A molecular signature of metastasis in primary solid tumors.Nat Genet.2003, 33 (1): 49-54.).Although above-mentioned in cell model type i collagen the effect of tumour is widely studied, at present less from the expression study in malignant tumor tissue for type i collagen.
When there is matrix reconstruction (ECM remodeling), type i collagen can be degraded, and the end peptide (N-telopeptides and C-telopeptides) of its N end and C end can be discharged in blood.CTx is exactly type i collagen C end end peptide 8 amino acid whose fragments of the preceding paragraph (EKAHDGGR), can in serum, by ELISA, detect (Bonde as the index of type i collagen degraded, M., Garnero, P., Fledelius, C., etc.Measurement of bone degradation products in serum using antibodies reactive with an isomerized form of an8amino acid sequence of the C-telopeptide of type I collagen.J Bone Miner Res, 1997, 12 (7): 1028-34.).Research for CTx is studied more in osteopathology, and CTx absorbs (Bone resorption) reasonable biomarker (Rosen as judgement bone at present, H.N., Moses, A.C., Garber, J., etc.Serum CTX:a new marker of bone resorption that shows treatment effect more often than other markers because of low coefficient of variability and large changes with bisphosphonate therapy.Calcif Tissue Int, 2000, 66 (2): 100-3.Marx, R.E., Cillo, J.E., Jr., Ulloa, J.J.Oral bisphosphonate-induced osteonecrosis:risk factors, prediction of risk using serum CTX testing, prevention, and treatment.J Oral Maxillofac Surg, 2007, 65 (12): 2397-410.).At present in the report about itself and cancer-related, in blood, the rise of CTx is at breast cancer (Cloos, P.A., Christgau, S., Lyubimova, N., etc.Breast cancer patients with bone metastases are characterised by increased levels of nonisomerised type I collagen fragments.Breast Cancer Res, 2003, 5 (4): R103-9.), prostate cancer (Garnero, P., Buchs, N., Zekri, J., etc.Markers of bone turnover for the management of patients with bone metastases from prostate cancer.Br J Cancer, 2000, 82 (4): 858-64.) and lung cancer (Leeming, D.J., Koizumi, M., Byrjalsen, I., etc.The relative use of eight collagenous and noncollagenous markers for diagnosis of skeletal metastases in breast, prostate, or lung cancer patients.Cancer Epidemiol Biomarkers Prev, 2006, 15 (1): 32-8.) in bone transporting patient, have report.But except the standard shifting as judgement neoplastic bone, CTx in other tumor development process and the clinical value of the aspect such as diagnosis of colorectal carcinoma prognosis do not appear in the newspapers.
Summary of the invention
The shortcoming of prior art in view of the above, the purposes of the c-terminal peptides CTx that the object of the present invention is to provide a kind of extracellular matrix components type i collagen in preparation or Screening Diagnosis colorectal cancer medicine, for solving the problems of the prior art.
For achieving the above object and other relevant objects, first aspect present invention provides the purposes of CTx in preparing Colorectal Cancer Diagnosis medicine, Screening Diagnosis colorectal cancer medicine and screening treatment colorectal cancer medicine.
Described Screening Diagnosis colorectal cancer medicine specifically refers to use CTx, to carrying out as the material of Colorectal Cancer Diagnosis drug use the assessment of biologically active, pharmacological action and medical value.
Described screening treatment colorectal cancer medicine specifically refers to use CTx, to carrying out the assessment of biologically active, pharmacological action and medical value as the material for the treatment of colorectal cancer drug use.
CTx in the present invention is 8 amino acid peptide sections of type i collagen α 1 chain carboxy-terminal, and its amino acid sequence is EKAHDGGR(Seq ID No.1).When cell generation matrix reconstruction under some physiology or pathological state, type i collagen in extracellular matrix can be degraded, the end peptide of its N end and C end can be discharged in blood, one of type i collagen C end end fragments of peptides that described CTx discharges while being type i collagen degraded.
Preferably, described Colorectal Cancer Diagnosis specifically refers to colorectal cancer preoperative diagnosis to judge by stages or to colorectal cancer prognosis.(mainly contain help not need before operation can be to proactive judgement by stages by pathology detection)
According to and according to International Union Against Cancer (UICC) and U.S. tumour federation (AJCC), combine the TNM(tumor-node-metastasis of formulation) allotment method, colorectal cancer can be divided into I-IV phase four-stage, described staging of colorectal cancer specifically refer to distinguish colorectal cancer according to described TNM allotment method in which in stage.
Described to colorectal cancer prognosis judgement specifically finger the course of disease of Patients with Colorectal Cancer and final result are carried out to prognosis judgement, more specifically for the disease free survival of Patients with Colorectal Cancer is carried out to prognosis judgement.
Preferably, described colorectal cancer is Human colorectal carcinoma.
Preferably, described CTx is biomarker.
Preferred, described CTx is serum biomarker.
The type i collagen C end end fragments of peptides discharging when type i collagen is degraded when preferably, described CTx is matrix reconstruction.
Preferred, described type i collagen derives from people.
Because I in clinical diagnostics, II phase belong to the commitment of colorectal cancer, if now find, tumour can be cured by excision mostly, III phase patient has started to find lymphatic metastasis, there is far-end and shift in IV phase patient, four period five-year survival rate reduce successively, recurrence rate after healing increases progressively.Because CTx just can be used as early stage special biomarker, its discovery is significant for the prediction prognosis of colorectal cancer.
Second aspect present invention provides a kind of Colorectal Cancer Diagnosis biomarker, and described biomarker is CTx, and the amino acid sequence of described CTx is as shown in Seq ID No.1.
Preferably, described biomarker is serum biomarker.
Preferably, described Colorectal Cancer Diagnosis specifically refers to colorectal cancer preoperative diagnosis to judge by stages or to colorectal cancer prognosis.
Preferably, described colorectal cancer is Human colorectal carcinoma, the type i collagen C end end fragments of peptides discharging during type i collagen when described biomarker CTx is matrix reconstruction (people source) degraded.
The present invention chooses Colorectal Carcinoma, serum clinical samples, and research type i collagen and CTx, in colorectal cancer, development and the clinical value of the aspect such as prognosis by stages occur.Pass through great many of experiments, inventor finds that CTx is in difference by stages in Patients with Colorectal Cancer serum, variation and the pathological staging of expression present positive correlation, and CTx and existing tumor in digestive tract carcinomebryonic antigen (Carcinoembryonic antigen, CEA) compare, can better distinguish and shift and non-transfer Patients with Colorectal Cancer.In addition, inventor also finds that 3 years disease free survivals of CTx and Patients with Colorectal Cancer are relevant, and its high expressed is indicating that Patients with Colorectal Cancer 3 years is without knurl existence prognosis mala.
In addition, in the present invention, inventor further provides the protein I Collagen Type VI that generates this end peptide of CTx in Expression in Colorectal Cancer situation, for understanding the effect of extracellular matrix in colorectal cancer generation evolution in depth, provide reference frame, and find thus, variation and the pathological staging of the expression of the protein I Collagen Type VI of this end peptide of generation CTx there is no obvious correlativity, have further confirmed thus the using value of CTx in preparing Colorectal Cancer Diagnosis medicine.
Accompanying drawing explanation
Fig. 1 represents that type i collagen is in the difference expression of Colorectal Carcinoma mRNA level by stages.
Fig. 2 represents that type i collagen is in the difference expression of Colorectal Carcinoma protein level by stages.
Fig. 3 represents that type i collagen is in difference Colorectal Carcinoma immunohistochemical staining result by stages.
Fig. 4 represent type i collagen degraded index CTx and degrade at random small fragment COL1 normal and difference by stages in Patients with Colorectal Cancer serum ELISA detect.
Fig. 5 represents that CTx compares with CEA and can better distinguish shifts and non-transfer Patients with Colorectal Cancer.
Fig. 6 represents that 3 years disease free survivals of CTx and Patients with Colorectal Cancer are relevant, and its high expressed is indicating that Patients with Colorectal Cancer 3 years is without knurl existence prognosis mala.
Fig. 7 represents that 3 years disease free survivals of CTx and Patients with Colorectal Cancer are relevant, and 3 years disease free survivals of its expression and Patients with Colorectal Cancer are negative correlativing relation.
Embodiment
Below, by specific instantiation explanation embodiments of the present invention, those skilled in the art can understand other advantages of the present invention and effect easily by the disclosed content of this instructions.The present invention can also be implemented or be applied by other different embodiment, and the every details in this instructions also can be based on different viewpoints and application, carries out various modifications or change not deviating under spirit of the present invention.
Before further describing the specific embodiment of the invention, should be understood that protection scope of the present invention is not limited to following specific specific embodiments; It is also understood that the term using in the embodiment of the present invention is in order to describe specific specific embodiments, rather than in order to limit the scope of the invention; In instructions of the present invention and claims, unless explicitly pointed out in addition in literary composition, singulative " ", " one " and " this " comprise plural form.
When embodiment provides numerical range, unless should be understood that the present invention is otherwise noted, between two end points of each numerical range and two end points, any one numerical value all can be selected.Unless otherwise defined, all technology of using in the present invention are identical with the meaning that those skilled in the art of the present technique understand conventionally with scientific terminology.The concrete grammar using in embodiment, equipment, material, according to those skilled in the art to the grasp of prior art and record of the present invention, can also with to the method described in the embodiment of the present invention, equipment, material is similar or any method, equipment and the material of the prior art that is equal to are realized the present invention.
Unless otherwise indicated, in the present invention, disclosed experimental technique, detection method, preparation method all adopt the routine techniques of molecular biology, biological chemistry, chromatin Structure and analysis, analytical chemistry, cell cultivation, recombinant DNA technology and the association area of the art routine.These technology are existing perfect explanation in existing document, specifically can be referring to MOLECULAR CLONING:A LABORATORY MANUAL such as Sambrook, Second edition, Cold Spring Harbor Laboratory Press, 1989and Third edition, 2001; Ausubel etc., CURRENT PROTOCOLS IN MOLECULAR BIOLOGY, John Wiley& Sons, New York, 1987and periodic updates; The series METHODS IN ENZYMOLOGY, Academic Press, San Diego; Wolffe, CHROMATIN STRUCTURE AND FUNCTION, Third edition, Academic Press, San Diego, 1998; METHODS IN ENZYMOLOGY, Vol.304, Chromatin (P.M.Wassarman and A.P.Wolffe, eds.), Academic Press, San Diego, 1999; With METHODS IN MOLECULAR BIOLOGY, Vol.119, Chromatin Protocols (P.B.Becker, ed.) Humana Press, Totowa, 1999 etc.
Inventor finds in the work of colorectal cancer proteomics in early stage, and type i collagen, at difference high expressed in Patients with Colorectal Cancer serum by stages, shows that type i collagen may be closely related with colorectal cancer.Inventor has found the high expressed of type i collagen in Colorectal Carcinoma in difference by stages, and further in serum, detect type i collagen different fragments in the difference expression in Patients with Colorectal Cancer by stages, find its c-terminus degraded end PEPC Tx and Patients with Colorectal Cancer by stages and prognosis relevant, completed on this basis the present invention.One, the detection of type i collagen in tissue
Embodiment 1: the collection of different Patients with Colorectal Cancer samples by stages and normal sample
Year May in May, 2009 to 2010, we collect the tissue of 91 routine Patients with Colorectal Cancers and serum sample altogether, and (patient has clarified a diagnosis by histopathology, and it suffers from colorectal cancer, and combine the TNM(tumor-node-metastasis of formulation according to International Union Against Cancer (UICC) and U.S. tumour federation (AJCC)) allotment method determined pathological staging, I phases 21 people wherein, II phases 41 people, III phases 22 people, IV phases 7 people, all patients have all carried out laparoscopic surgical procedure), collected the healthy human body serum sample of 33 examples without disease of digestive tract and diseases associated with inflammation simultaneously.All patients follow the tracks of return visit for every 3 months in after surgery first 2 years, within postoperative the 3rd year every 6 months, follow the tracks of return visit, within postoperative the 41st month, stopping, recording patient's recurrence or death time point.Patients clinical information is as shown in table 1.
Table 1 patients clinical information
aaC, the colon ascendens; TC, transverse colon; DC, colon descendens; SC, sigmoid colon; RC, rectum;
Embodiment 2:I Collagen Type VI is in the difference expression of Colorectal Carcinoma RNA and protein level by stages
1, organize RNA extraction and real-time fluorescence quantitative PCR (Realtime PCR) to detect:
We have extracted total mRNA of tumor tissues and corresponding normal colorectal carcinoma, and have detected respectively the mrna expression level of its type i collagen.With the Trizol kit of Invitrogen company, extract total mRNA of tissue, use the PrimeScript RT reagent Kit kit of Takara company from the synthetic 20ml cDNA of RNA reverse transcription of 2mg, use 7500 real-time fluorescence quantitative PCR instrument and the corresponding SYBR Green PCR Master Mix kit of Applied Biosystems company to carry out real-time fluorescence quantitative PCR (adopt 2-△ △ Ct analytic approach to analyze gene expression abundance, GAPDH is as reference gene).The primer sequence that PCR reaction is used is as shown in table 2.(result is as shown in Fig. 1 and table 3)
The primer sequence that table 2PCR reaction is used
2, histone extracts and Western blotting (Western Blot) detection
For carrying out the protein expression analysis of tumor tissues and normal colorectal carcinoma, freezing tissue samples 100mg is added to RIPA lysate (50mM Tris-HCl (pH7.4), 0.4wt%sodium deoxycholate, 150mM NaCl, 1wt%Triton, 1mM PMSF) after, on ice, carry out homogenate fragmentation and cracking in approximately 30 minutes, afterwards at 4 ℃, 16, under 000 * g condition centrifugal 30 minutes, by BCA method, carry out the protein quantification of solution.Use β-actin as the quantitative internal reference of loading; the equal protein of each sample (50mg/ band) is carried out to 8wt%SDS-PAGE electrophoretic separation; be transferred to afterwards on the nitrocellulose filter of GE company; after sealing, use 4 ℃ of overnight incubation of primary antibodie (anti-collagen I (1:5000, ab34710; Abcam).The positive band of immune response is used the Odyssey Infrared Imaging System of Li-COR company observe and take pictures, and uses Quantity One to carry out quantitatively.(result is as Fig. 2)
3, histogenic immunity groupization detects
The tissue paraffin section de of 3 – 5mm is used 4wt% pepsin in 37 ℃ of reaction 20min, carries out antigen retrieval, reacts spend the night (anti-collagen I, 1:500, ab34710 afterwards with 4 ℃ of primary antibodies; Abcam), use the Envision detection system of DAKO company observe and take pictures.(result is as Fig. 3)
4, interpretation of result
Table 3I Collagen Type VI is in the difference real-time fluorescence quantitative PCR testing result raw data of Colorectal Carcinoma by stages
| Sample number into spectrum | Pathological staging | COL1A1(tumour/normal) | COL1A2(tumour/normal) |
| 88 | I | 17.09 | 5.99 |
| 51 | I | 3.04 | 1.81 |
| 74 | I | 3.01 | 1.57 |
| 101 | I | 1.81 | 1.12 |
| 72 | I | 1.69 | 0.91 |
| 116 | I | 266.35 | 30.92 |
| 67 | II | 124.16 | 33.06 |
| 48 | II | 113.29 | 32.96 |
| 79 | II | 16.42 | 10.02 |
| 59 | II | 15.11 | 7.22 |
| 84 | II | 11.31 | 7.03 |
| 75 | II | 7.61 | 4.91 |
| 47 | II | 7.11 | 1.04 |
| 55 | III | 18.33 | 14.49 |
| 17 | III | 4.81 | 3.22 |
| 68 | III | 4.36 | 2.93 |
| 82 | III | 3.18 | 2.00 |
| 62 | III | 1.91 | 1.86 |
| 39 | III | 1.34 | 1.74 |
| 81 | III | 1.14 | 0.98 |
| 94 | IV | 3.43 | 3.34 |
| 61 | IV | 1.21 | 1.58 |
| 53 | IV | 1.27 | 1.04 |
| 103 | IV | 0.94 | 0.59 |
| 44 | IV | 1.41 | 0.44 |
From table 3 and Fig. 1, the result of Fig. 2 is visible, and no matter at mRNA level or protein level, in each phase colon cancer tumor tissues, type i collagen is expressed all highly than normal structure, illustrates that type i collagen is up-regulated expression really in tumor tissues when cancer occurs.But, from the result of Fig. 1 and Fig. 2, can find out, type i collagen the non-transfer phase (I phase and II phase) than transfer phase (III phase and IV phase) tumor tissues in high expressed, especially, on protein level, there is significant difference (p < 0.05).
Type i collagen expression is expressed the highest in II phase tumor tissues, and on protein level, its tumor tissues of COL1A1 and COL1A2 and normal structure ratio reach respectively 22.54 and 7.19 times.In clinical diagnostics, I, II phase belong to commitment, if now find, tumour can be cured by excision mostly, III phase patient has started to find lymphatic metastasis, and IV phase patient far-end occurs and shifts, four period five-year survival rate reduce successively, recurrence rate after healing increases progressively.Therefore, at clinical middle commitment, especially the II phase is critical stage in the evolution of colorectal cancer, clinical stages the finding that there is and help predict prognosis of diagnostic biomarkers, in important significance in clinical application.In addition, the albumen of some II phase high expresseds is probably relevant with the aggressive migration of tumour cell.
Two, the detection of CTx in serum
Embodiment 3:I Collagen Type VI degraded index CTx and degrade at random small fragment COL1 normal and difference by stages in Patients with Colorectal Cancer serum ELISA detect:
1, ELISA detection kit and method
COL1 in serum and CTx level can be used commercial reagents box (Uscn Life Science& Technology Co., Houston, TX, USA) carry out ELISA detection, change of serum C Tx is used the being at war with property of specific monoclonal antibody of linear EKAHDGGR peptide section (fragment of the α 1 chain carboxy-terminal peptide of type i collagen) to suppress ELISA and detects, in brief, unlabelled CTx and special being at war with property of the CTx antibody inhibitory reaction being coated with in advance in biotin labeled exactly synthetic CTx peptide section and serum, in each reaction micropore, add horseradish peroxidase Avidin, hatch concrete detection method reference reagent box instructions.Add after substrate solution, in the color of reaction mixture and serum, the concentration of CTx is negative correlation.In serum, CEA level is used commercial reagents box to detect (Roche Diagnostics, Mannheim, Germany), concrete detection method reference reagent box instructions.
2, Analysis of test results
The average serum content of CTx from the I phase to the IV phase is respectively 2.19,3.16, and 3.58 and 4.58ng/ml, the variation of its expression and the pathological staging of colorectal cancer present positive correlation.In terminal cancer (III+IV phase), the level of CTx is significantly higher than the respective horizontal (p=0.003, Mann – Whitney U test) of CTx in early-stage cancer (I+II phase).(result is as Fig. 4)
3, CTx and staging of colorectal cancer and Analysis of relationship between prognosis
Between the pathological staging due to CTx and colorectal cancer, there is good positive correlation, and it is by stages all closely related with early diagnosis, result for the treatment of and the prognosis information of cancer patient, therefore we so that CTx has been carried out to experimenter's operating characteristic curve (Receiver operating characteristic, ROC) and 3 years disease free survival analysis (Disease-free survival, DFS), in order to the potential clinical value of CTx is made to assessment.Result can find out, CTx is better than the differentiation effect (AUC:0.584) of CEA for differentiation effects (AUC:0.696) early stage, end-stage patients.(result is as Fig. 5)
In addition we have assessed CTx and colorectal cancer without contacting between knurl existence, and the mean value 4.08ng/ml of all patient CTx of take is boundary, and patient is divided into two groups.Kaplan-Meier survival analysis result shows, 3 years disease free survivals of high expressed group of CTx are starkly lower than another group (log rank test, p=0.006), CTx is carried out to the analysis of Cox proportional hazard model and also illustrate that CTx can provide important information for the prognosis judgement of Patients with Colorectal Cancer.(result is as Fig. 6)
Further, experimenter is divided into 3 groups by the serum content of CTx, is respectively 0 < CTx≤2,2 < CTx≤4,4 < CTx; (log rank test, p=0.022), Kaplan-Meier survival analysis result shows, lifting gradually along with CTx expression concentration, 3 years disease free survivals of Patients with Colorectal Cancer reduce gradually, both present negative correlativing relation, have further confirmed the important value that CTx can judge for the prognosis of Patients with Colorectal Cancer.(result as shown in Figure 7)
In sum, the present invention has effectively overcome various shortcoming of the prior art and tool high industrial utilization.
Above-described embodiment is illustrative principle of the present invention and effect thereof only, but not for limiting the present invention.Any person skilled in the art scholar all can, under spirit of the present invention and category, modify or change above-described embodiment.Therefore, such as in affiliated technical field, have and conventionally know that the knowledgeable, not departing from all equivalence modifications that complete under disclosed spirit and technological thought or changing, must be contained by claim of the present invention.
Claims (11)
- The purposes of 1.CTx in preparing Colorectal Cancer Diagnosis medicine, Screening Diagnosis colorectal cancer medicine and screening treatment colorectal cancer medicine.
- 2. CTx as claimed in claim 1 is preparing Colorectal Cancer Diagnosis medicine, Screening Diagnosis colorectal cancer medicine and is screening the purposes in treatment colorectal cancer medicine, it is characterized in that, described Colorectal Cancer Diagnosis specifically refers to colorectal cancer preoperative diagnosis to judge by stages or to colorectal cancer prognosis.
- 3. the purposes of CTx as claimed in claim 1 in preparing Colorectal Cancer Diagnosis medicine, Screening Diagnosis colorectal cancer medicine and screening treatment colorectal cancer medicine, is characterized in that, described colorectal cancer is Human colorectal carcinoma.
- 4. the purposes of CTx as claimed in claim 1 in preparing Colorectal Cancer Diagnosis medicine, Screening Diagnosis colorectal cancer medicine and screening treatment colorectal cancer medicine, is characterized in that, described CTx is biomarker.
- 5. the purposes of CTx as claimed in claim 4 in preparing Colorectal Cancer Diagnosis medicine, Screening Diagnosis colorectal cancer medicine and screening treatment colorectal cancer medicine, is characterized in that, described CTx is serum biomarker.
- 6. CTx as claimed in claim 1 is preparing Colorectal Cancer Diagnosis medicine, Screening Diagnosis colorectal cancer medicine and is screening the purposes in treatment colorectal cancer medicine, it is characterized in that the type i collagen C end end fragments of peptides that described CTx discharges while being type i collagen degraded.
- 7. CTx as claimed in claim 6 is preparing Colorectal Cancer Diagnosis medicine, Screening Diagnosis colorectal cancer medicine and is screening the purposes in treatment colorectal cancer medicine, it is characterized in that the type i collagen C end end fragments of peptides discharging when type i collagen is degraded when described CTx is matrix reconstruction.
- 8. a Colorectal Cancer Diagnosis biomarker, described biomarker is CTx, the amino acid sequence of described CTx is as shown in Seq ID No.1.
- 9. a kind of Colorectal Cancer Diagnosis biomarker as claimed in claim 8, is characterized in that, described biomarker is serum biomarker.
- 10. a kind of Colorectal Cancer Diagnosis biomarker as claimed in claim 8, is characterized in that, described Colorectal Cancer Diagnosis specifically refers to colorectal cancer preoperative diagnosis to judge by stages or to colorectal cancer prognosis.
- 11. a kind of Colorectal Cancer Diagnosis biomarkers as claimed in claim 8, is characterized in that, described colorectal cancer is Human colorectal carcinoma, the type i collagen C end end fragments of peptides discharging when type i collagen is degraded when described biomarker CTx is matrix reconstruction.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310476530.9A CN103529220B (en) | 2013-10-12 | 2013-10-12 | CTx is preparing the purposes in Colorectal Cancer Diagnosis medicine |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310476530.9A CN103529220B (en) | 2013-10-12 | 2013-10-12 | CTx is preparing the purposes in Colorectal Cancer Diagnosis medicine |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN103529220A true CN103529220A (en) | 2014-01-22 |
| CN103529220B CN103529220B (en) | 2015-09-23 |
Family
ID=49931397
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310476530.9A Active CN103529220B (en) | 2013-10-12 | 2013-10-12 | CTx is preparing the purposes in Colorectal Cancer Diagnosis medicine |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103529220B (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104651522A (en) * | 2015-03-06 | 2015-05-27 | 河北医科大学第四医院 | Novel application of I-type alpha 1 chain |
| CN107102141A (en) * | 2016-02-23 | 2017-08-29 | 中国科学院生物物理研究所 | Molecular marked compound CD71 and Endometrial Carcinomas diagnosis, the application by stages and in prognosis |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2002014103A (en) * | 2000-06-30 | 2002-01-18 | Kyokuho Yagita | Method of diagnosing cancer at early stage thereof |
| WO2013033629A2 (en) * | 2011-08-31 | 2013-03-07 | Oncocyte Corporation | Methods and compositions for the treatment and diagnosis of colorectal cancer |
-
2013
- 2013-10-12 CN CN201310476530.9A patent/CN103529220B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2002014103A (en) * | 2000-06-30 | 2002-01-18 | Kyokuho Yagita | Method of diagnosing cancer at early stage thereof |
| WO2013033629A2 (en) * | 2011-08-31 | 2013-03-07 | Oncocyte Corporation | Methods and compositions for the treatment and diagnosis of colorectal cancer |
Non-Patent Citations (2)
| Title |
|---|
| C. DE LA PIEDRA ET AL.: "Urinary α and β C-Telopeptides of Collagen I: Clinical Implications in Bone Remodeling in Patients with Anorexia Nervosa", 《OSTEOPOROS INT》 * |
| 宋斌斌等: "上海地区人群血清骨转换标志物参考区间的建立", 《中华检验医学杂志》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104651522A (en) * | 2015-03-06 | 2015-05-27 | 河北医科大学第四医院 | Novel application of I-type alpha 1 chain |
| CN107102141A (en) * | 2016-02-23 | 2017-08-29 | 中国科学院生物物理研究所 | Molecular marked compound CD71 and Endometrial Carcinomas diagnosis, the application by stages and in prognosis |
Also Published As
| Publication number | Publication date |
|---|---|
| CN103529220B (en) | 2015-09-23 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Rivera et al. | Prognostic biomarkers in oral squamous cell carcinoma: a systematic review | |
| Schmitt et al. | Salivary duct carcinoma: an aggressive salivary gland malignancy with opportunities for targeted therapy | |
| Nakagawa et al. | Thymoma: a clinicopathologic study based on the new World Health Organization classification | |
| Ahn et al. | PD-L1 expression in gastric cancer: interchangeability of 22C3 and 28-8 pharmDx assays for responses to immunotherapy | |
| Waldmann et al. | Surgery for multiple endocrine neoplasia type 1-associated primary hyperparathyroidism | |
| Takikita et al. | Fascin and CK4 as biomarkers for esophageal squamous cell carcinoma | |
| Cho-Chung | Autoantibody biomarkers in the detection of cancer | |
| CN110007083B (en) | Hyaluronic acid as biomarker for metastatic breast cancer | |
| Helm et al. | Dedifferentiation precedes invasion in the progression from Barrett's metaplasia to esophageal adenocarcinoma | |
| Oloomi et al. | Comparing blood versus tissue-based biomarkers expression in breast cancer patients | |
| Ding et al. | Expression of focal adhesion kinase and phosphorylated focal adhesion kinase in human gliomas is associated with unfavorable overall survival | |
| Gong et al. | Overexpression of Cripto and its prognostic significance in breast cancer: a study with long-term survival | |
| Tissier | Classification of adrenal cortical tumors: what limits for the pathological approach? | |
| Yamac et al. | Serum YKL-40 levels as a prognostic factor in patients with locally advanced breast cancer | |
| Okabayashi et al. | Invasive carcinoma derived from branch duct-type IPMN may be a more aggressive neoplasm than that derived from main duct-type IPMN | |
| Moris et al. | Risk factors for malignant progression of intraductal papillary mucinous neoplasms | |
| Siar et al. | Epithelial-to-mesenchymal transition in ameloblastoma: focus on morphologically evident mesenchymal phenotypic transition | |
| Li et al. | Screening and validating the core biomarkers in patients with pancreatic ductal adenocarcinoma | |
| Nicolini et al. | “Tumour marker guided” salvage treatment prolongs survival of breast cancer patients: final report of a 7-year study | |
| Sato et al. | Bone marrow adipocytes induce cancer‐associated fibroblasts and immune evasion, enhancing invasion and drug resistance | |
| Ortega et al. | Prognostic role of IRS-4 in the survival of patients with pancreatic cancer | |
| CN103529220B (en) | CTx is preparing the purposes in Colorectal Cancer Diagnosis medicine | |
| Park et al. | Relationships between metastasis-associated protein (MTA) 1 and lymphatic metastasis in tonsil cancer | |
| Zhao et al. | Integrative analysis of cancer-associated fibroblast signature in gastric cancer | |
| US20220170105A1 (en) | Methods for diagnosis and prognosis of prostate cancer |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant |