CN107102141A - Molecular marked compound CD71 and Endometrial Carcinomas diagnosis, the application by stages and in prognosis - Google Patents
Molecular marked compound CD71 and Endometrial Carcinomas diagnosis, the application by stages and in prognosis Download PDFInfo
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- CN107102141A CN107102141A CN201610099982.3A CN201610099982A CN107102141A CN 107102141 A CN107102141 A CN 107102141A CN 201610099982 A CN201610099982 A CN 201610099982A CN 107102141 A CN107102141 A CN 107102141A
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- 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
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
- G01N33/57407—Specifically defined cancers
- G01N33/57442—Specifically defined cancers of the uterus and endometrial
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- 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
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/577—Immunoassay; Biospecific binding assay; Materials therefor involving monoclonal antibodies binding reaction mechanisms characterised by the use of monoclonal antibodies; monoclonal antibodies per se are classified with their corresponding antigens
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
- G01N2333/705—Assays involving receptors, cell surface antigens or cell surface determinants
- G01N2333/70582—CD71
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Abstract
The present invention relates to molecular marked compound CD71 and Endometrial Carcinomas diagnosis, the application by stages and in prognosis, the molecular marked compound is human TfR 1 (CD71), belongs to biology field.The present invention utilizes the antibody of anti-human transferrin receptor 1 (CD71), it was found that human TfR 1 (CD71) is not expressed or extremely low expression in normal endometrial tissue and Precancerous Lesion, the specificity overexpression in people's endometrial, it is relevant with stages of endometrial cancer and prognosis.Based on this, CD71 can be applied to diagnosis, clinical stages and the prognosis of endometrial.
Description
Technical field
The present invention relates to a kind of molecular marked compound, anti-human transferrin receptor (transferrin receptor 1,
TfR1, also known as CD71) diagnosis of antibody Endometrial Carcinomas, in clinical stages and prognosis evaluation should
With belonging to biology field.
Background technology
Carcinoma of endometrium is one of three common big malignant tumours of female reproductive system, accounts for female cancer
7%, account for the 20-30% of female reproductive system malignant tumour.In recent years, worldwide, it is sub
The morbidity of endometrial carcinoma is on the rise, and in rejuvenation (Parkin DM, Bray F, Ferlay J, et al.
Global cancer statistics,2002[J].CA Cancer J Clin,2005,55(2):74-108)。
With the continuous discovery for the deep and novel marker that conventional tag thing is studied, SABC is in uterus
Play the part of in the pathological diagnosis and antidiastole of endometrial carcinomas more and more important role (Wu Huanwen, Liang Zhiyong,
The progress Diagnosis pathologies of Zhang Hui, Liu Tong China carcinoma of endometrium immunohistochemical markers things,
2013, Vol.20, No.3180-183).Carcinoma of endometrium diagnosis immunohistochemical markers conventional at present
Thing includes hormone receptor ER, PR, CEA, P53, P16, PTEN etc..These current tumor markers
Deficiency in terms of there is sensitivity, broad spectrum activity and specificity in terms of the diagnosis.As P53 is mainly used in
The diagnosis or antidiastole of II type carcinomas of endometrium, low differentiation endometrial carcinomas is high there is also part in I types
Phenomenon is expressed, some cases reach in complete radiolucent table；P16 is mainly used in the diagnosis of cervical carcinoma, in son
Express relatively low in endometrial carcinoma, be mainly used in the antidiastole of carcinoma of endometrium and cervical carcinoma.
PTEN loss of expression is the earliest events of I type endometrium carcinogenesis, further, since being missing from table
Reach, effective interpretation of PTEN coloration results limits its popularization and application in practical application.CEA exists
High expression, is mainly used for the antidiastole of carcinoma of endometrium in cervical carcinoma.ER, PR expression are more
Hormone therapy for carcinoma of endometrium is assessed.So far, the molecular marked compound of differential high efficient is not yet found
For carcinoma of endometrium diagnosis, the entirety in CD71 antibody Endometrial Carcinomas in this project is expressed
Rate is up to about 80%, it is believed that can as auxiliary diagnosis carcinoma of endometrium molecular marked compound.Current
Molecular marked compound predicts have research to think for clinical prognosis:In addition to PR, ER, PTEN, P53
And Ki-67 expression is to smaller (the carcinoma of endometrium molecule mark of prediction significance of patient clinical prognosis
Remember thing and clinical pathologic characteristic and relation Peking University journal (medicine) Vol.43NO.5 of prognosis
Oct.2011,743-748)。
TfR (TfR1, CD71) high expression in the active cell of multiple fission, especially
The malignant cell of a large amount of iron and some cell surface quickly divided overexpressions are being needed, these
Acceptor is not expressed or extremely low expression in the normal tissue, and research shows, TfR (CD71)
The high expression in Several Kinds of Malignancy, such as liver cancer, breast cancer, Malignant cerebral gliomas etc., do not have still at present
There is the research report that TfR (TfR1, CD71) is expressed in people's endometrial, more
CD71 the data that clinical prognosis is predicted is not used for.
The content of the invention
The present inventor is in research TfR (TfR1, CD71) table in people's endometrial
On the basis of reaching, it is found that CD71 not only can be for diagnosis of endometrial carcinoma, still can be to intrauterine
The clinical stages of film cancer and prognosis are analyzed.The antibody for CD71 is developed simultaneously, it is described anti-
Body can be used for diagnosis of endometrial carcinoma, and the clinical stages of carcinoma of endometrium and prognosis can be carried out
Analysis.In addition, it is used for diagnosis of endometrial carcinoma the invention further relates to one kind, and can be to endometrium
The detection kit that the clinical stages of cancer and prognosis are analyzed.
Specifically, the present invention includes following aspect：
On the one hand, the present invention relates to purposes of the CD71 as the molecular labeling of carcinoma of endometrium.
On the other hand, prepared the present invention relates to CD71 for diagnosis of endometrial carcinoma, and/or assessment
Purposes in carcinoma of endometrium clinical stages and the kit of prognosis.
The clinical stages, refer to according to tumor size, position whether there is the feelings such as infiltration and DISTANT METASTASES IN
Condition, carries out clinical stages, and the phase is not higher, and malignancy is higher.Wherein international association of gynemetrics
(FIGO) it is by stages in this area most common clinical stages.Specific FIGO staging scales：0
Phase：Carcinoma in situ (cancer before infiltration)；1 phase：Tumour is confined to corpus uteri；2 phases；Tumor-infiltrated palace
Neck but without departing from uterus；3 phases：Tumour is focal and/or regional diffusion；4 phases：Tumour involves bladder
Mucous membrane and/or intestinal mucosa, tumour is not with DISTANT METASTASES IN (not including being transferred to vagina, pelvic cavity serous coat or annex).
In preferred embodiments, the carcinoma of endometrium behaviour carcinoma of endometrium.
In preferred embodiments, the purposes of the reagent preparation box is by preparing specific recognition CD71
Antibody realize.
In the art, the preparation method of antibody be it is known, it is most common including hybridoma method,
Phage display method etc..Preparation method for antibody known in the art can be used for this hair for preparing
Bright antibody.In addition, CD71 can be obtained by following approach：First, from human tumor cell line
Middle extraction RNA, then, reverse transcription is cDNA library in reverse transcription system.Design CD71
CDNA primer, using PCR method, CD71 cDNA are cloned from cDNA library
Come.CD71 cDNA are building up on carrier for expression of eukaryon, the CD71 of selective expression total length
Sequence or part CD71 sequences, by the method for affinity chromatography, realize the purifying of CD71 antigens
Prepare.
On the other hand, the present invention is a kind of faces for diagnosis of endometrial carcinoma, and/or assessment carcinoma of endometrium
Bed by stages and prognosis kit, the kit includes specific recognition CD71 antibody.
In preferred embodiments, the kit also includes the reagent for immune detection.
In preferred embodiments, the kit can also include the operation instruction of the kit.
In preferred embodiments, the immune detection includes SABC, immunofluorescence, is immunized
Precipitation, is immunized flow cyctometry and immuning hybridization.As long as can show that CD71 antibody and target group
The combination knitted and the immunologic detection method of respective strengths may serve to realize the present invention.
In preferred embodiments, the antibody includes monoclonal antibody or polyclonal antibody.
In preferred embodiments, the antibody is monoclonal antibody.
In preferred embodiments, the monoclonal antibody or polyclonal antibody are to turn iron egg for people
Prepared by all or part of amino acid sequence of polymeric immunoglobulin receptor.And the function with specific bond CD71
Antibody fragment be also contained in the scope of the present invention.
Anti-human transferrin receptor antibody mainly with the TfR knot in endometrial
Close.Anti-human transferrin receptor antibody is according to all or part of ammonia of human TfR in the present invention
Prepared by base acid sequence, can be combined with the TfR in people's endometrial.This
Invention is mainly used in Immunohistochemical detection, but be not limited to that SABC is detected.
Brief description of the drawings
Fig. 1 is the SABC figure of normal cycle endometrium：Wherein left figure (10 ×), right figure (20 ×)；
Fig. 2 is the SABC figure of Simple hyperplasia endometrium：Wherein left figure (10 ×), right figure (20 ×)；
Fig. 3 is the SABC figure of endometrial：Wherein left figure (10 ×), right figure (20 ×)；
Fig. 4 represents the endometrial and TfR of different clinical stages (FIGO is by stages)
The relation of (TfR1, CD71) expression intensity.Carcinoma of endometrium is divided into according to FIGO clinical stages
4 groups.Chi-square Test is shown：***p<0.001, NS represents no significant difference.
Embodiment
With reference to embodiment, the invention will be further described, and embodiments of the present invention are simultaneously
This is not limited to, it is every according to present disclosure or principle, any this area of implementation etc.
With replacing, protection scope of the present invention is belonged to.
Embodiment 1
SABC detects and compares normal endometrial tissue, Simple hyperplasia endometrial tissue and
The difference of Expression of Transferrin Receptor in endometrial.
1. experiment material
Organization chip (TMA) purchase is stood upright bio tech ltd from Shanghai, wherein including uterus
57, endometrial carcinomas sample, 23, normal endometrium sample, Simple hyperplasia endometrial tissue 7
Example.SABC reagent：The antibody (EC-1) of anti-human transferrin receptor passes through conventional murine hybridoma
Method prepares (Edward A.Greenfield.Generating Monoclonal Antibodies.
Antibodies:A Laboratory Manual,Second Edition,2014,Cold Spring Harbor
Laboratory Press), specifically it may refer to, its content is incorporated herein by reference.
1. experimental method
Routine immunization group SP methods are carried out.(De Jong AS,Van Kessel-van Vark M,Raap
AK.Histochem J.1985Oct；17(10):1119-30.Sensitivity of various
visualization methods for peroxidase and alkaline phosphatase activity in
Immunoenzyme histochemistry) to organization chip (TMA) paraffin section (5um is thick)
Processing is as follows：
1) section dewaxing aquation.
2) 5 minutes in distilled water；
3) fresh 3%H is configured with distilled water or PBS2O2Room temperature is closed 10 minutes, uses distilled water
Wash 1 time；
4) conventional antigen retrieval；
5) PBS is washed 5 minutes；
6) Normal Goat Serum confining liquid (Beijing Bioisystech Co., Ltd of Zhong Shan Golden Bridge), room is added dropwise
Temperature 30 minutes；
7) get rid of surplus liquid and primary antibody (CD71 antibody, Abcam is added dropwise：Ab49517), 37 DEG C 1
Hour or 4 DEG C overnight；
8) PBS is washed 2 times, each 5 minutes every time；
9) secondary antibody is added dropwise, and (horseradish enzyme marks mouse anti goat igg, and Beijing health is the century limited public affairs of biotechnology
Department) it is incubated 37 DEG C 45 minutes；
10) PBS is washed 2 times, each 5 minutes every time；
11) DAB develops the color 5 minutes；
12) distillation washing terminates dyeing, and haematoxylin is redyed, distilled water flushing.
15) resinene (Beijing Bioisystech Co., Ltd of Zhong Shan Golden Bridge) is added, slide is capped.
2. experimental result
ImmunohistochemistryResults Results are shown in Fig. 1-3.Fig. 1 is the SABC figure of normal cycle endometrium；Fig. 2
It is the SABC figure of Simple hyperplasia endometrium；Fig. 3 is the SABC of endometrial
Figure.It can be seen that：In being in TfR endometrial tissues-severe positive expression,
Hardly expressed in normal cycle endometrium and Simple hyperplasia endometrium.Detailed data result is shown in
Table 1 below.
The TfR of table 1. (TfR1, CD71) Endometrial Carcinomas and the expression in non-cancer tissue
3. conclusion
High expression in TfR Endometrial Carcinomas, in normal cycle endometrium and pure
Do not expressed in Endometrial Hyperplasia or extremely low expression.
Embodiment 2
SABC detects the expression feelings of TfR in different clinical stages endometrials
Condition.
1. experiment material
Organization chip (TMA) is purchased from Shaanxi Ai Lina bio tech ltd, according to FIGO points
Phase, wherein 1 phases of FIGO sample 158,2 phase samples 32,3 phase samples 29,4 phases mark
This 4.SABC reagent：Anti-human transferrin receptor antibody be EC-1 as previously described.
2. experimental method：Be the same as Example 1.
3. experimental result：
The endometrial and anti-human transferrin receptor of different clinical stages (FIGO is by stages)
Expression power has correlation.Chi-square Test is shown：p<0.001, point out anti-human between different clinical stages
The expression of TfR has significant difference.As a result see below (accompanying drawing 4 in specification)
4. conclusion
Carcinoma of endometrium clinical stages, is higher, and prognosis is poorer, then anti-human transferrin receptor expression is stronger.
Thus, it is possible to which the intensity expressed according to anti-human transferrin receptor judges the clinic point of carcinoma of endometrium
Phase, so that judging prognosis.
Claims (10)
1.CD71 as the molecular labeling of carcinoma of endometrium purposes.
2.CD71 is being prepared for diagnosis of endometrial carcinoma, and/or is being assessed carcinoma of endometrium clinical stages
And the purposes in the kit of prognosis.
3. according to the purposes of claim 1 or 2, the carcinoma of endometrium behaviour carcinoma of endometrium.
4. purposes according to claim 2, the purposes is by preparing specific recognition CD71's
Antibody is realized.
For diagnosis of endometrial carcinoma, and/or carcinoma of endometrium clinical stages and prognosis are assessed 5. a kind of
Kit, the kit includes specific recognition CD71 antibody.
6. kit according to claim 5, the kit is also included for immune detection
Reagent.
7. kit according to claim 6, the immune detection includes SABC, is immunized
Fluorescence, immunoprecipitation, are immunized flow cyctometry and immuning hybridization.
8. the kit described in purposes according to claim 4 or claim 5, the antibody
Including monoclonal antibody or polyclonal antibody, preferably monoclonal antibody.
9. purposes according to claim 5 or kit, the monoclonal antibody or Anti-TNF-α
Body is prepared by all or part of amino acid sequence for being directed to human TfR.
10. kit according to claim 5, the clinical stages be FIGO by stages.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111304333A (en) * | 2020-03-13 | 2020-06-19 | 中国医学科学院肿瘤医院 | Novel application of SERINC4 gene |
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