CN107881240B - The diagnosis and treatment marker of osteosarcoma - Google Patents
The diagnosis and treatment marker of osteosarcoma Download PDFInfo
- Publication number
- CN107881240B CN107881240B CN201711445268.6A CN201711445268A CN107881240B CN 107881240 B CN107881240 B CN 107881240B CN 201711445268 A CN201711445268 A CN 201711445268A CN 107881240 B CN107881240 B CN 107881240B
- Authority
- CN
- China
- Prior art keywords
- fibcd1
- osteosarcoma
- inhibitor
- application according
- genes
- 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.)
- Active
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
- C12Q1/6886—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material for cancer
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/7088—Compounds having three or more nucleosides or nucleotides
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
-
- 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
-
- 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
- G01N33/57484—Immunoassay; Biospecific binding assay; Materials therefor for cancer involving compounds serving as markers for tumor, cancer, neoplasia, e.g. cellular determinants, receptors, heat shock/stress proteins, A-protein, oligosaccharides, metabolites
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/158—Expression markers
-
- 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
- G01N2333/745—Assays involving non-enzymic blood coagulation factors
- G01N2333/75—Fibrin; Fibrinogen
Abstract
The invention discloses the diagnosis and treatment marker of osteosarcoma, the marker is FIBCD1, it is demonstrated experimentally that FIBCD1 mRNA and its albumen up-regulated expression in osteosarcoma tissue, prompt to be applied to FIBCD1 in the early diagnosis of osteosarcoma；Further, In vitro cell experiment confirms that FIBCD1 is related with the proliferation of osteosarcoma cell, apoptosis, migration and invasion, prompts FIBCD1 that can be applied to the clinical treatment of osteosarcoma as drug targets.
Description
Technical field
The invention belongs to biomedicine field, the diagnosis and treatment marker of osteosarcoma, the specific marker is FIBCD1.
Background technology
Osteosarcoma (Osteosarcoma, OS) is also known as osteogenic sarcoma, to be mainly in 10-25 Sui teen-age most common original
Hair property malignant bone tumor, the rate of transform, recurrence rate and the death rate are higher, are often associated with the bad knot such as amputation, Lung metastases, death
Office.Currently, the method for the treatment of osteosarcoma is still mostly based on operation, appropriate combined radio chemotherapy (Kushnir I, KolanderY,
Bickels J,et al.[J].Med Oncol.,2014,31(5):936.).Though tumor resection tissue is in clinical treatment of osteosarcoma
Important step, but surgery alone cure rate is only 15%-20% (Archer NP, Napier TS, Villanacci JF.
[J].Cancer Causes Control,2016,27(7):863 1 868.).Being constantly progressive of iconography means, various guarantor's limbs
Technology step up and the fast development etc. of associated bone substitute so that limbs retain therapy increasingly in bone and flesh
It cuts a striking figure in the treatment of tumor (Wang TY, Dormans JP, Chang B. [J] .Ann Plast Surg., 2012,69 (5):
560-564.).On the other hand, it is to combine to consolidate after tumor resection tissue in due course to be in progress particularly critical for control osteosarcoma
The chemotherapy of solidity or radiotherapy.In recent years, though the basis such as gene therapy, immunization therapy, molecular targeted therapy and clinical research obtain
Certain progress, current curative effect are not affirmed still.
The occurrence and development of tumour are a complicated processes, are bodies under various factors effect, the cell of local organization
The normal regulation to growth is lost at the genetic level, leads to the paraplasm of cell and the neoformation that is formed.Tumour is
Genopathy, Basic of Biology are the exceptions of gene.Currently, molecular regulation mechanism about osteosarcoma occurrence and development is so far still
It does not get across.Along with molecular genetics, cell biology, molecular biology, cytogenetics, proteomics and transcription
The further investigation that group is learned etc., the influencing each other of living environment and genetic mutation be increasingly becoming osteosarcoma field research hotspot it
One (Zhang G, Bai R, Zhang T, et al. [J] .Genet Mol Res., 2015,14 (3):8283-8289).
Recent studies have found that in osteosarcoma some genes present differential expression, as patent 201510075917.2,
201510075920.4, WWP1, C8orf59, CCT γ disclosed in 201510075918.7,201510075919.1,
PLEKHA5 genes.With going deep into osteosarcoma molecular biosciences research, the targeted therapy of osteosarcoma, which also more and more becomes, to be ground
The emphasis studied carefully, and following developing direction is represent, find new effective biomarker, the early diagnosis for osteosarcoma
And targeted therapy has great importance.
Invention content
In order to make up for the deficiencies of the prior art, it is an object of the present invention to provide biomarkers in osteosarcoma
Application in diagnosing and treating.
The second object of the present invention is, provides a kind of pharmaceutical composition of the product and treatment osteosarcoma of diagnosis osteosarcoma
Object.
To achieve the goals above, present invention employs following technical solutions：
The first aspect of the present invention provide it is a kind of detection biomarker reagent prepare for diagnosing osteosarcoma
Application in product, including：
A) biomarker in the sample of identification subject, wherein the biomarker is FIBCD1；And
B) biomarker and reference are compared, wherein with the difference of the biomarker with reference to compared with
For detecting osteosarcoma.
Further, the sample is tissue.
Further, with reference to compared with, biomarker expression level raises.
The second aspect of the present invention provides a kind of product, and the product includes the reagent for detecting FIBCD1 levels.Wherein,
FIBCD1 up-regulated expressions in Patients with Osteosarcoma, the product include but is not limited to preparation, chip, kit.
Further, the reagent is selected from：
The probe of specific recognition FIBCD1 genes；Or
The primer of specific amplification FIBCD1 genes；Or
Specifically bind the antibody or ligand of the albumen of FIBCD1 codings.
Further, the primer sequence of specific amplification FIBCD1 genes is as shown in NO.1~2 SEQ ID.
The third aspect of the present invention provides a kind of pharmaceutical composition, and described pharmaceutical composition includes the inhibition of FIBCD1
Agent, and/or other medicine classes and pharmaceutically acceptable carrier and/or auxiliary material with the inhibitor compatibility.
Further, the inhibitor is selected from：Nucleic acid inhibitor, protein inhibitor, proteolytic enzyme, protein binding molecule.
Further, the inhibitor is nucleic acid inhibitor siRNA, it is preferred that the sequence of siRNA such as IDNO.9~10 SEQ
It is shown.
The fourth aspect of the present invention provides following any one of them application：
1) application of the product described in second aspect of the present invention in the tool for preparing diagnosis osteosarcoma.
2) applications of the FIBCD1 in the drug for preparing treatment tumour；
3) applications of the FIBCD1 in the pharmaceutical composition for preparing treatment bone and flesh tumor metastasis；
4) applications of the FIBCD1 in the drug candidate of screening treatment osteosarcoma.
Further, 1) product include with RT-PCR, real-time quantitative PCR, new-generation sequencing, in situ hybridization, chip or
Immunoassay detects the reagent of FIBCD1；
Or 3) 2) pharmaceutical composition includes FIBCD1 inhibitor described in, and/or other medicines with the inhibitor compatibility
Class and pharmaceutically acceptable carrier and/or auxiliary material；
Wherein, the inhibitor is selected from：As target sequence and FIBCD1 gene tables can be inhibited using FIBCD1 or its transcript
It reaches or the disturbing molecule of genetic transcription, including：ShRNA (children purpura nephritis), siRNA (siRNA), dsRNA, Microrna,
Antisense nucleic acid, or can express or be formed the construction of the shRNA, siRNA, dsRNA, Microrna, antisense nucleic acid；Or
The protein bound binding molecule (antibody or ligand that can such as inhibit FIBCD1 protein actives) of specificity and FIBCD1 codings.
4) the step of drug candidate of screening treatment osteosarcoma, includes in：
The system for the albumen expressed or containing FIBCD1 genes or its coding is handled with candidate substances；With
Detect the expression of the albumen of FIBCD1 genes or its coding or activity in the system.
Description of the drawings
Fig. 1 is the expression figure in osteosarcoma tissue using QPCR detection FIBCD1 genes；
Fig. 2 is the expression figure in osteosarcoma tissue using Western blot detection FIBCD1 albumen；
Fig. 3 is transfected condition figures of the FIBCD1 in osteosarcoma cell；Wherein, figure A is using QPCR detection transfections to bone
The influence diagram that FIBCD1mRNA is expressed in sarcoma cell；Figure B is using Western blot detection transfections in osteosarcoma cell
The influence diagram of FIBCD1 albumen；
Fig. 4 is the influence diagram for detecting FIBCD1 gene pairs human osteosarcoma cell proliferations；
Fig. 5 is the influence diagram for detecting FIBCD1 and being migrated to osteosarcoma cell；
Fig. 6 is the influence diagram for detecting FIBCD1 and being invaded to osteosarcoma cell.
Specific implementation mode
The present invention after extensive and in-depth study, by high-flux sequence method, detects gene in osteosarcoma samples and exists
The expression of tumor tissues and cancer beside organism, the gene for the expression that finds differences inquire into its relationship between the generation of osteosarcoma, from
And it is that the early detection of osteosarcoma and targeted therapy find better approaches and methods.By screening, present invention firstly discovers that
FIBCD1 conspicuousnesses raise in osteosarcoma.It is demonstrated experimentally that the expression by reducing FIBCD1, can effectively inhibit bone and flesh
The growth and invasion of oncocyte prompt the expression of detection FIBCD1 genes that can become the auxiliary diagnosis of osteosarcoma early diagnosis
One of index, interference FIBCD1 gene expressions, which can become, prevents or treats osteosarcoma or the new way of bone and flesh tumor metastasis.
(biology) marker
" biomarker " refers to the molecule with specific biological characteristic, biochemical characteristics or aspect and indicates
Object can be used for determining the severity presence or absence of specified disease or situation and/or specified disease or situation.
In the present invention, " marker " refer to one or more relevant parameters of biomolecule (i.e. " biomarker "),
Such as natural or artificial synthesized generation nucleic acid (i.e. genes of individuals, and coding and noncoding DNA and RNA) and albumen (such as
Peptide, polypeptide)." marker " in the present invention further includes that finger can be by considering the expression number from two or more unlike signal objects
According to the single parameter for calculating or otherwise obtaining.
" protein ", " peptide " and " polypeptide " may be used interchangeably and broadly refer to have two or more amino acid sub-
The compound of base, amino acid analogue or peptidomimetic.The subunit can be connected by peptide bond.Protein or peptide must include at least
Two amino acid and amino acid maximum number is unlimited can include the sequence of protein or peptide.Terms used herein " ammonia
Base acid " refers to natural and/or non-natural or the amino acid of synthesis, including both glycine and D and L optical isomers, amino
Acid-like substance and peptidomimetic.
FIBCD1 genes
FIBCD1 in the present invention includes wild type, saltant type or its segment.A kind of representative FIBCD1 gene orders
As shown in FIBCD1 genes (NC_000009.12) in current international public nucleic acid database GeneBank, people of the invention
FIBCD1 nucleotide full length sequences or its segment can usually use PCR amplification method, recombination method or artificial synthesized method to obtain.
Detection technique (method)
The expression of the gene of the present invention is examined using a variety of detection techniques known to persons of ordinary skill in the art
It surveys, these technologies include but not limited to detect the technology of gene or protein expression level.
The expression of gene " detection " refer to determining biological sample marker gene mRNA exist and its expression with
Just it predicts the process of gastric cancer prognosis and the amount by measuring mRNA can be achieved.Analysis method is but not limited to for this purpose
RT-PCR, competitive RT-PCR (competitiveRT-PCR), real-time RT-PCR (Real-timeRTPCR), RNA enzyme protection are surveyed
Determine method (RPA；RNaseprotectionassay), northern blottings (northernblotting), DNA microarray chip
Deng.
" expression of detection albumen " refers to the protein presence expressed in determining biological sample marker gene and its table
Up to level to predict the process of osteosarcoma, and can be combined by using with the protein specific expressed in said gene
Antibody determine the amount of protein.Analysis method is but not limited to western blottings for this purpose
(westernblotting), ELISA (enzyme linked immunosorbent assay (ELISA)), radioimmunoassay (Radioimmunoassay), put
Penetrating property immunodiffusion (Radioimmunodiffusion), Auchterlonie (Ouchterlony) immunodiffusion, rocket
(Rocket) electrophoresis, histogenic immunity decoration method, immunoprecipitation assay (immunoprecipitationassay), complement knot
Close measuring method (completefixationassay), FACS, protein-chip (proteinchip) etc..
The nucleic acid or albumen of non-amplification or amplification can be detected by any conventional means in the present invention.
Chip, kit
In the present invention, " chip ", " microarray ", " array " can be with equivalent substitutes, including but not limited to：DNA microarray
(for example, cDNA microarrays and oligonucleotide microarray), protein microarray, micro-array tissue, transfection or cell microarray, change
Chemical combination object microarray and Antibody microarray.The DNA microarray of commonly referred to as genetic chip, DNA chip or biochip is micro-
The set of DNA points is seen, these points are connected on the surface of solids (for example, glass, plastics or silicon chip), are formed for thousands of kinds
Gene is carried out at the same time expression pattern analysis or the array of expression monitoring.Fixed DNA fragmentation is known as probe, thousands of available
In single DNA microarray.Microarray can be used for identifying disease base by comparing the gene expression in disease and normal cell
Cause or transcript (for example, ncRNA).Multiple technologies can be used to be manufactured for microarray, including but not limited to：It is printed with apicule needle
Photoetching is carried out on to glass slide, using prefabricated mask, carries out photoetching, ink jet printing or microelectrode battle array using dynamic micro mirror element
Electrochemical method on row.
Kit in the present invention can be used for detecting the expression of FIBCD1, it is preferred that the kit, which includes detection, to be had
The reagent of the albumen of the detection FIBCD1 genes of effect amount or its coding, one or more substances selected from the group below：Container, using saying
Bright book, positive control, negative control object, buffer, auxiliary agent or solvent.Such as the solution for being suspended or fixing cell, it can
The label or tag of detection makes nucleic acid be easy to the solution of hybridization, is used for the solution of lytic cell, or for the molten of nucleic acid purification
Liquid.
The present invention kit in can also have kit operation instructions, be described how using kit into
Row detection, and how tumor development to be judged using testing result, therapeutic scheme is selected.
Screening prevents or the compound for the treatment of osteosarcoma
The present invention can utilize biomarker FIBCD1 screenings to prevent or treat the compound of osteosarcoma, the present invention's
In a kind of embodiment, including step：In test group, the addition test compound in cultivating system, and observe the test group
Cell in FIBCD1 expression quantity and/or activity；In control group, test chemical combination is not added in identical cultivating system
Object, and observe the expression quantity and/or activity of FIBCD1 in the cell of control group；
Wherein, if the expression quantity of the FIBCD1 of cell and/or activity are less than control group in test group, the test is indicated that
Compound is expression to FIBCD1 and/or activity have inhibiting effect treating cancer drug candidate.
As one embodiment of the present invention, the step further includes：To the candidate compound of acquisition into advancing one
The cell experiment and/or animal experiment of step, further to select and determine for prevention, alleviation or treatment from candidate compound
The useful substance of osteosarcoma.
As embodiments of the present invention, screening prevents or the system of the candidate compound for the treatment of osteosarcoma is not limited to cell
System further includes cell system, subcellular system, solution system, organizational framework, organ systems or animal system etc., the body
System is not limited to above-mentioned form, as long as the system can detect test compound and can reduce expression and the/activity of ASPRV1
.
Inhibitor and pharmaceutical composition
Discovery based on inventor, the present invention provides a kind of purposes of the inhibitor of FIBCD1, are used to prepare inhibition bone
The pharmaceutical composition of sarcoma.As used herein, the inhibitor of the FIBCD1 includes but not limited to inhibitor, antagonist, resistance
Stagnant dose, blocking agent, nucleic acid inhibitor etc..
The inhibitor of the FIBCD1 genes or albumen refers to any activity for reducing FIBCD1 albumen, reduces
The stability of FIBCD1 genes or albumen, the expression for lowering FIBCD1 albumen reduce FIBCD1 albumen effective acting times or suppression
The substance of the transcription and translation of FIBCD1 genes processed, these substances are used equally for the present invention, as useful for lowering FIBCD1
Substance, so as to be used to prevent or treat osteosarcoma.For example, the inhibitor is：Nucleic acid inhibitor, protein inhibitor,
Antibody, ligand, proteolytic enzyme, protein binding molecule, as long as it can lower FIBCD1 albumen on albumen or gene level
Or the expression of its encoding gene.
As a kind of selection mode of the present invention, the inhibitor of the FIBCD1 is that a species specificity is combined with FIBCD1
Antibody.The specific antibody includes monoclonal antibody, polyclonal antibody；The present invention includes not only complete antibody molecule,
Also include any segment or the modification of antibody, for example, chimeric antibody, scFv, Fab, F (ab ') 2, Fv etc..As long as the segment energy
Enough binding abilities retained with FIBCD1 albumen.Those skilled in the art are public when preparation for the antibody of protein level
Know, and the present invention may use any method to prepare the antibody.
As a kind of preferred embodiment of the present invention, the inhibitor of the FIBCD1 is a kind of small interference of FIBCD1 specificity
RNA molecule.As used herein, " siRNA " refers to a kind of short-movie section double stranded rna molecule, can be with homologous complementary
The mRNA of sequence is the target specific mRNA of degradation, this process is exactly RNA interference (RNA interference) processes.It is small
RNA interfering can be prepared into the form of double-strandednucleic acid, it contains there are one positive-sense strand and an antisense strand, this two chains are only hybridizing
Under conditions of form double-strand.One double-stranded RNA compound can be prepared by the positive-sense strand that is separated from each other and antisense strand.Therefore,
For example, complementary positive-sense strand and antisense strand are chemical synthesis, and can generate the double-strand of synthesis by anneal thereafter
RNA compounds.
When screening effective siRNA sequence, the present inventor is best effective to find out by largely comparing analysis
Segment.The present inventor's design has synthesized a variety of siRNA sequences, and they are transfected osteosarcoma cell line by transfection reagent respectively
It is verified, selects the best siRNA of interference effect, they are respectively provided with sequence shown in SEQ ID NO.9, SEQ ID NO.10
Row are further tested in cellular level, as a result prove to inhibit efficiency very high for test cell line.
The nucleic acid inhibitor such as siRNA of the present invention can be with chemical synthesis, can also be by a recombinant nucleic acid structure
Expression cassette is prepared after being transcribed into single stranded RNA.The nucleic acid inhibitors such as siRNA, can be by using transfection reagent quilt appropriate
It is transported into the cell, or multiple technologies known in the art also can be used and be transported into the cell.
As a kind of optional mode of the present invention, the inhibitor of the FIBCD1 can also be a kind of " children purpura nephritis
(Small hairpin RNA, shRNA) " is the non-coding small RNA molecular that can form hairpin structure, children purpura nephritis energy
Enough by RNA interference channels come the expression of suppressor.As above-mentioned, shRNA can be expressed by double-stranded DNA template.Double-stranded DNA
Template is inserted into a carrier, such as plasmid or viral vectors, is then connected to a promoter carry out table in vitro or in vivo
It reaches.ShRN under the action of DICER enzymes, can be cut into siRNA molecule in eukaryocyte, hence into RNAi approach.
" shRNA expression vectors " refers to plasmid of some this fields conventionally used for building shRNA structures, exist on the usual plasmid "
Every sequence " and positioned at " intervening sequence " both sides multiple cloning sites or for replace sequence, to people can by shRNA (or
Analog) corresponding DNA sequence dna be inserted by way of forward and reverse multiple cloning sites or replace thereon for replacing sequence,
RNA after DNA sequence dna transcription can form shRNA (Short Hairpin) structure." the shRNA expression vectors " is current
It can be bought and be obtained by commercially available approach completely, such as some viral vectors.
The present invention also provides a kind of pharmaceutical compositions, it contains the inhibitor of a effective amount of FIBCD1, and
Pharmaceutically acceptable carrier.The composition can be used for inhibiting osteosarcoma.The inhibitor of any FIBCD1 above-mentioned
Preparation for composition.The carrier includes but is not limited to diluent, excipient, adhesive, disintegrant, absorption enhancement
Agent, surfactant, Humectant, absorption carrier, lubricant, buffer, stabilizer, bacteriostatic agent, isotonic agent, chelating agent, pH controls
Preparation.
Pharmaceutical composition can be various oral or parenteral dosage forms.Using including filler, filler, adhesive,
Conventional thinner including wetting agent, disintegrant and surfactant or excipient pharmaceutical composition.Solid orally ingestible
Including tablet, pill, pulvis, granule, capsule etc..These solid pharmaceutical preparations can by by least one compound with it is a kind of or more
Kind of excipient, for example, starch, calcium carbonate, sucrose, lactose, prepared by the mixing such as gelatin.In addition to simple excipient, can also make
With lubricator such as magnesium stearate or talcum.In addition, liquid oral medicine includes suspension, and solution, emulsion and syrup etc..In addition to logical
It is commonly used for outside the water and atoleine of simple diluent, may also include various excipient, for example, wetting agent, sweetener, fragrance,
Preservative etc..The preparation of parenteral administration includes sterile water solution, and nonaqueous solvents, suspending agent, emulsion is freeze-dried, suppository etc..Third
Glycol, polyethylene glycol, vegetable oil such as olive oil, injectable esters such as ethyl oleate etc. may be used as nonaqueous solvents and suspending agent.Bolt
Agent main component may include witepsol, polyethylene glycol, Tween61, cocoa butter, laurel tallow, glycerin gelatine etc..
Pharmaceutical composition can have any one preparation selected from the group below：Tablet, pill, powder, granule, capsule suspend
Liquid, solution, emulsion, syrup, sterile water solution, non-aqueous solution, suspension, lotion, lyophilized preparation and suppository.
As used in the present invention, " effective quantity " refers to that its dosage is enough to treat disease, with the conjunction suitable for any therapeutic treatment
Interests/risk-ratio of reason.The effective dose level of composition can according to the type of subject, the severity of disease, by
The age of examination person and gender, pharmaceutical activity, the sensibility to drug, administration time, administration route, excretion rate, treatment time, with
Other known facts determine in drug associated with composition and medical field.The present invention pharmaceutical composition can be used alone or
It is administered in combination, and can be sequentially or simultaneously administered with conventional therapeutic agent with other therapeutic agents.It can be used one or more doses
Composition is applied in type.All above-mentioned factors are considered, in the case where minimum of the maximum efficiency without causing side effect can be shown
Most important using composition, which can be readily determined by those skilled in the art.
The pharmaceutical composition of the present invention can also be with the drug combination of other treatment osteosarcoma, and other therapeutic compound can
To be administered simultaneously with main active constituent, or even it is administered simultaneously in same composition.Can also with individual composition or
The dosage form different from main active constituent individually gives other therapeutic compounds.
Preferably, the means that gene therapy can be used carry out.For example, can be directly by the inhibitor of FIBCD1 by such as noting
It the methods of penetrates and to deliver medicine to subject；Alternatively, can will be carried by certain approach the inhibitor of FIBCD1 ceneme (such as
Expression vector or virus etc. or siRNA or shRNA) it is delivered on target spot, and it is allowed to the FIBCD1 inhibitor of expression activity, have
Body situation need to be depending on the type of the inhibitor, these are well-known to those skilled in the art.
Experiment in the present invention is all at least completed according to being repeated 3 times, and result data is all with average value ± standard
The mode of difference indicates, using SPSS18.0 statistical softwares come for statistical analysis, the paired comparisons of cancer and cancer beside organism adopt
It is examined with t, it is believed that work as P<There is statistical significance when 0.05.
It being further illustrated the present invention with reference to specific embodiment, the embodiment of the present invention is only used for explaining the present invention,
It is not intended to limit protection scope of the present invention.
Experimental method used in following embodiments is conventional method unless otherwise specified.
The materials, reagents and the like used in the following examples is commercially available unless otherwise specified.
The screening of embodiment 1 and the relevant gene marker of osteosarcoma
1, sample collection
6 osteosarcoma tissues and cancer beside organism's sample are collected respectively, the acquirement of tissue samples obtains the informed consent of patient,
And obtain the agreement by the committee of organizational ethics.
2, the preparation of RNA sample
RNA is extracted using the tissue RNA extracts kits of Invitrogen companies, concrete operations are with reference to specification.
3, the quality analysis of RNA sample
The RNA concentration and purity extracted are detected using Nanodrop2000, agarose gel electrophoresis detects RNA
Integrality, Agilent2100 measure RIN values.Concentration >=200ng/ μ l, OD260/280 is between 1.8~2.2.
4, rRNA is removed
The rRNA in total serum IgE is removed using Ribo-Zero kits.
5, construction cDNA library
It is specific to grasp using the structure for carrying out cDNA library using the Truseq RNA sample Prep Kit of Illumina
Make by specification progress.
6, upper machine sequencing
CDNA library is sequenced using Hiseq4000 microarray datasets, concrete operations by specification carries out.
7, high-throughput transcript profile sequencing data analysis
Bioinformatic analysis and processing are carried out to sequencing result, the screening criteria of differential gene is fdr<0.05, two groups
Fpkm average values difference be more than 5.
8, result
RNA-seq is poor the results show that differential expression is presented in FIBCD1 in Patients with Osteosarcoma cancerous tissue and cancer beside organism
It is different that there is statistical significance (P<0.05).
The differential expression of 2 QPCR sequence verification FIBCD1 genes of embodiment
1, large sample QPCR verifications are carried out to FIBCD1 gene differential expressions.According to the sample collection mode in embodiment 1
Select Patients with Osteosarcoma cancer beside organism and each 50 of osteosarcoma tissue.
2, it is as described in Example 1 to extract specific steps by RNA.
3, reverse transcription
Using FastQuant cDNA the first chain synthetic agent box (article No.s：KR106 mRNA reverse transcriptions) are carried out.Specific steps
It is as follows：
(1) 5 × gDNA Buffer 2.0 μ l, 1 μ g of total serum IgE is added and adds Rnase Free ddH2O makes total volume to 10 μ
L, 42 DEG C of heating 3min in water-bath；
(2) 20 μ l reaction systems are built：10 × Fast RT Buffer, 2.0 μ L, RT Enzyme Mix 1.0 μ l, FQ-
2.0 μ l, RNase Free ddH of RT Primer Mix2Mixing in the mixed liquor in (1) is added after 5.0 μ l mixing of O；
(3) 42 DEG C of heating 15min in water-bath, 95 DEG C of heating 3min, -20 DEG C store for future use.
4, QPCR is expanded
(1) design of primers
QPCR amplifications are designed according to the coded sequence of FIBCD1 genes in Genebank and house-keeping gene GAPDH genes to draw
Object is synthesized by Bo Maide companies.
The primer sequence of FIBCD1 genes：
Forward primer sequence is 5 '-GACCATTCAGAGAACAACT-3 ' (SEQ ID NO.1)；
Reverse primer sequences are 5 '-GTACTGCCCATTGAGGTT-3 ' (SEQ ID NO.2).
The primer sequence of GAPDH genes：
Forward primer sequence is 5 '-GGAGCGAGATCCCTCCAAAAT-3 ' (SEQ ID NO.3)；
Reverse primer sequences are 5 '-GGCTGTTGTCATACTTCTCATGG-3 ' (SEQ ID NO.4).
(2) PCR reaction systems：Forward primer and each 0.6 μ l, 2 × SuperReal PreMix Plus, 10 μ of reverse primer
L, DNA profiling 2 μ l, ddH27.4 μ l, 50 × ROX Reference Dye of O△2 μ l, 4.8 μ l of sterile purified water.
(3) PCR reaction conditions：95 DEG C of 15min, (95 DEG C of 10s, 55 DEG C of 30s, 72 DEG C of 32s) × 40 cycles, 95 DEG C of 15s,
60 DEG C of 60s, 95 DEG C of 15s.PCR reactions are carried out on 7300 type fluorescence quantitative PCR instruments of ABI, pass through melt curve analysis analysis and electrophoresis
Determine that purpose band, Δ Δ CT methods carry out relative quantification.
5, result
The results are shown in Figure 1, and compared with cancer beside organism, FIBCD1 up-regulated expressions in osteosarcoma tissue, difference has system
Meter learns meaning (P<0.05), consistent with high-flux sequence result.
The differential expression of 3 protein immunization imprinting of embodiment experiment detection FIBCD1 albumen
1, the extraction of total protein is organized
It puts it into and is placed in the glass homogenizer in ice after shredding tissue with scissors, RIPA lysates and PMSF are with 100:
The RIPA lysates of corresponding amount, glass is added in 1 ratio mixing, the ratio that 100 μ l lysates are added according to every 20mg tissue specimens
Glass homogenizer pulverize tissue until its fully crack, the liquid after cracking is drawn in EP pipes, at 4 DEG C 14000rpm centrifuge
5min collects supernatant.
2, total protein concentration measures
The measurement of albumen concentration is carried out according to the specification of BCA determination of protein concentration kits.
3, SDS-PAGE electrophoresis
8% separation gel and 5% concentration glue are prepared according to the specification of PAGE gel reagent preparation box and are carried out
Electrophoresis.
4, western is detected
1) electrotransfer
Pvdf membrane is put into methanol solution and activates 5min, is put into transferring film buffer solution and balances 20min.PAGE glue is taken out to put
Enter in transferring film buffer solution, cut corresponding PAGE glue, according to be followed successively by from down to up filter paper, pvdf membrane, PAGE glue, filter paper it is suitable
Sequence is put into half-dried transferring film instrument, constant pressure 25V transferring films 1.5h；
2) immuning hybridization
Pvdf membrane is taken out, PBS flushings are placed in 5%BSA solution shakes closing 2h at room temperature, and pvdf membrane is put into hybridization
In bag, primary antibody is added and stays overnight, washs pvdf membrane with TBST buffer solutions, adds corresponding secondary antibody, be incubated 2h at room temperature, TBST is slow
Fliud flushing is washed.
3) DAB develops the color
The slightly dry rear DAB developing solutions that Fresh is added dropwise of pvdf membrane, record is scanned after pvdf membrane colour developing.Made with β-actin
For internal reference, sxemiquantitative gray analysis is carried out to band using Quantity One Labworks image acquisition and analysis softwares, experiment repeats 3
It is secondary, as a result take average gray value.
5, result
The results are shown in Figure 2, and compared with cancer beside organism, expression of the FIBCD1 albumen in osteosarcoma tissue significantly rises
Height, difference have statistical significance.
The silence of 4 FIBCD1 genes of embodiment
1, cell culture
Cell line of human osteosarcoma U-2OS, with the DMEM culture mediums containing 10% fetal calf serum and 1%P/S 37 DEG C, 5%
CO2, relative humidity be 90% incubator in cultivate.Change within 2-3 days liquid 1 time, cell growth is good, is grown in monolayer adherence.Make
With the 0.25% trypsase conventional digestion passage containing EDTA.
2, it transfects
1) precellular processing is transfected
The day before transfection plants 3~5 × 10 on 6 well culture plates5A cells/well cultivates one in antibiotic-free culture medium
It, cell density is 30~50% when transfection, changes serum free medium into before transfection.
2) design of siRNA
Negative control siRNA sequence (siRNA-NC)：
Positive-sense strand is 5 '-UUCUCCGAACGUGUCACGU-3 ' (SEQ ID NO.5)
Antisense strand is 5 '-ACGUGACACGUUCGGAGAA-3 ' (SEQ ID NO.6)
siRNA1：
Positive-sense strand is 5 '-UUUAUUGUUUUUUAAGCACAA-3 ' (SEQ ID NO.7)
Antisense strand is 5 '-GUGCUUAAAAAACAAUAAAUU-3 ' (SEQ ID NO.8)
siRNA2：
Positive-sense strand is 5 '-UUUUAAGCACAAACUUCAGUG-3 ' (SEQ ID NO.9)
Antisense strand is 5 '-CUGAAGUUUGUGCUUAAAAAA-3 ' (SEQ ID NO.10)
Experiment is divided into three groups：Control group (U-2OS), negative control group (siRNA-NC) and experimental group (siRNA1,
SiRNA2), the sequence of wherein negative control group siRNA and FIBCD1 genes is without homology.
3) it transfects
It is transfected using the liposome Lipofectamine 3000 of Invitrogen companies, by specification is grasped
Make.
5, QPCR detects the transcriptional level of FIBCD1 genes
The extraction of 5.1 cell total rnas
The RNA in cell is extracted using the cell RNA extracts kit of Qiagen, experimental implementation is to specifications
It carries out.
5.2 reverse transcription steps are the same as embodiment 2.
5.3QPCR amplification steps are the same as embodiment 2.
6, Western detects the expression of FIBCD1 albumen
The cell for collecting the different disposal group in logarithmic phase washs cell with the PBS of precooling, RIPA lysates is added,
30min is placed on ice, is scraped off the cell of cracking using cell scraper, and the liquid after cracking, which is drawn to EP, using pipettor manages
In, 14000rpm centrifuges 5min at 4 DEG C, collects the supernatant after centrifugation, remaining step is the same as embodiment 3.
7, result
As a result such as Fig. 3 is shown, with non-transfection group compared with transfecting siRNA-NC groups, experimental group siRNA2 expressions reduce
Significantly, difference has statistical significance (P<0.05), therefore selection siRNA2 carries out subsequent experimental.
The influence of 5 FIBCD1 gene pairs human osteosarcoma cell proliferations of embodiment
FIBCD1 gene pairs human osteosarcoma cell proliferation capacities are detected using MTT experiment
1, the good cell of upgrowth situation is taken, after conventional digestion is at single cell suspension, cell is counted, cell is diluted to conjunction
The cell suspension of suitable concentration.
2, in 96 well culture plates, the different disposal group cell per well after dilution is inoculated with 2000 cells, 3 is at least set and puts down
Row hole and cell-free medium control, 37 DEG C, 5%CO2Culture is for 24 hours.
3, it 1 after inoculation, 2,3,4,5 days daily OD values taken out 3 hole cells and detect its 490nm with mtt assay, is counted
Number calculates average value.
4, liquid is discarded supernatant before detecting, culture solution is washed 3 times, and MTT free serum cultures based sols (5mg/ml) 10 μ is added per hole
L continues in 37 DEG C of incubators to cultivate 4h, terminates culture.
5,100 μ l Formanzan lysates are added per hole, shaking table shakes 1min slowly.With wavelength it is 490nm in microplate reader
Optical density (OD) value is measured, using the time as horizontal axis, OD value is that the longitudinal axis draws cell growth curve.
6, result
The results are shown in Figure 4, and compared with the control, for experimental group after transfecting siRNA2, the proliferation of cell obviously receives suppression
System, difference have statistical significance (P<0.05), illustrate that the proliferation of FIBCD1 and osteosarcoma cell is closely related, drop can be passed through
The expression of low FIBCD1 inhibits the proliferation of osteosarcoma cell.
The influence of 6 FIBCD1 gene pairs apoptosis in osteosarcoma cells of embodiment
Use the influence of flow cytomery FIBCD1 gene pairs Apoptosis.
1, cell culture and transfection procedure are the same as embodiment 3.
2, flow cytomery
1) cell of the different disposal group in exponential phase is broken into cell suspension through pancreatin digestion after-blow and counted.
Take 106The cell suspension of amount, 1000rpm centrifuge 5min；
2) supernatant is abandoned, 195 μ l Annexin V-FITC combination liquid is added, cell is gently resuspended；
3) the Annexin V-FITC of 5 μ l are added, soft mixing is protected from light is incubated 10min at room temperature；
4) 1000rpm centrifuges 5min, abandons supernatant, and cell is gently resuspended in the Annexin V-FITC combination liquid that 190 μ l are added；
5) 10 μ l propidium iodides (PI) dyeing liquors, soft mixing are added, ice bath avoid light place carries out flow cytomery
Apoptosis situation, all experiments are repeated 3 times, and results are averaged.
3, result：
The results show that compared with the control group, the apoptosis rate of the cell of experimental group significantly increases, the cell of siRNA2 groups is transfected
Apoptosis rate is (20.17 ± 0.021) %, and the apoptosis rate of transfection siRNA-NC groups is (4.89 ± 0.23) %, above-mentioned difference
With statistical significance (P<0.05), the above results show that FIBCD1 is related with the apoptosis of osteosarcoma cell, which crosses table
Up to the apoptosis for inhibiting osteosarcoma cell.
Influences of 7 FIBCD1 of embodiment to cell migration
1, the fibronectin of 50 μ g/ml of 1ml is added per hole into 6 orifice plates, is placed in 4 DEG C of refrigerator overnights.
2, remaining Fibronectin solution is discarded, is cleaned with serum free medium, by not existing together in exponential phase
Reason group cell is inoculated in after pancreatin digestion is resuspended in the 6 orifice plates for being covered with fibronectin, and every group of cell sets 2 multiple holes, per hole 5
×105A cell, 37 DEG C, 5%CO2Overnight incubation in incubator.
3, it is grown to when about 90% fusion when cell, marks an acellular cut with the Tip heads of 10 μ l, PBS solution is washed
The cell to fall off is removed, serum free medium is added and continues to cultivate.
4,0h, 48h observe the healing state at cell cut and take pictures after cut.Experiment is repeated 3 times, and is as a result taken
Average value.
5, result
The results are shown in Figure 5, compared to the control group for, transfect the cell migration distance after siRNA2 and be substantially reduced, and it is right
According to, without significant difference, illustrating that FIBCD1 is related with the migration of osteosarcoma cell between group.
Influences of 8 FIBCD1 of embodiment to cell invasion
1, prepared by the cells Transwell
1) serum free medium that the Matrigel glue of 50mg/L is pre-chilled with 4 DEG C is with 1:8 dilution proportion, mixing；
2) the 60 diluted Matrigel glue of the μ of μ l~80 l (3.9 μ g/ μ l) is taken to be placed in the Transwell upper chambers that aperture is 8 μm
Polycarbonate membrane on, so that all micropores on film is covered by Matrigel, being placed in 37 DEG C of 30min makes Matrigel polymerize
At gel.
2, cell suspension is configured
The cell of different disposal group in exponential phase is digested through pancreatin, after serum free medium is resuspended, adjustment
Cell concentration is 5 × 104A/ml.
3, cell inoculation
The cell suspension of 2ml is added in Transwell upper chambers, the complete training containing 10% fetal calf serum of 1ml is added in lower room
Base is supported, is positioned in mating 6 orifice plates, 37 DEG C, 5%CO2Under the conditions of cultivate 20-24h；The cells Transwell are taken out, cotton swab is wiped
The Matrigel glue in most upper chamber face and the cell for not penetrating film.
4, it dyes
After cell culture, the cells Transwell are taken out, cotton swab wipes the Matrigel glue in upper chamber face to the greatest extent and do not penetrate film
Cell, lower room face is with after 95% alcohol fixation 15min, haematoxylin dyeing 2min, and 5 high power lenses are taken at random under inverted microscope
Visual field observation is counted and is taken pictures.The cell number for counting the cell faces Xia Shi is to penetrate the cell number of Matrigel glue, is averaged
As experimental result, and the invasiveness of tumour cell is represented with the cell number, experiment is repeated 3 times, and every group of cell sets 3 multiple holes.
5, result
The results are shown in Figure 6, and compared with the control group, the cell for transfecting the experimental group of siRNA2 passes through the cells Transwell
The cell number of polycarbonate membrane significantly reduces, and no significant difference between control group U-2OS and transfection siRNA-NC, explanation
The overexpression of FIBCD1 promotes the invasion of osteosarcoma cell, prompt can be by targeting FIBCD1 genes, its expression of silence inhibits bone
The invasion of sarcoma.
The explanation of above-described embodiment is only intended to understand the method and its core concept of the present invention.It should be pointed out that for this
For the those of ordinary skill in field, without departing from the principle of the present invention, several improvement can also be carried out to the present invention
And modification, these improvement and modification will also be fallen into the protection domain of the claims in the present invention.
Sequence table
<110>Beijing Yang Shen biology information technologies Co., Ltd
<120>The diagnosis and treatment marker of osteosarcoma
<160> 10
<170> SIPOSequenceListing 1.0
<210> 1
<211> 19
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 1
gaccattcag agaacaact 19
<210> 2
<211> 18
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 2
gtactgccca ttgaggtt 18
<210> 3
<211> 21
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 3
ggagcgagat ccctccaaaa t 21
<210> 4
<211> 23
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 4
ggctgttgtc atacttctca tgg 23
<210> 5
<211> 19
<212> RNA
<213>Artificial sequence (Artificial Sequence)
<400> 5
uucuccgaac gugucacgu 19
<210> 6
<211> 19
<212> RNA
<213>Artificial sequence (Artificial Sequence)
<400> 6
acgugacacg uucggagaa 19
<210> 7
<211> 21
<212> RNA
<213>Artificial sequence (Artificial Sequence)
<400> 7
uuuauuguuu uuuaagcaca a 21
<210> 8
<211> 21
<212> RNA
<213>Artificial sequence (Artificial Sequence)
<400> 8
gugcuuaaaa aacaauaaau u 21
<210> 9
<211> 21
<212> RNA
<213>Artificial sequence (Artificial Sequence)
<400> 9
uuuuaagcac aaacuucagu g 21
<210> 10
<211> 21
<212> RNA
<213>Artificial sequence (Artificial Sequence)
<400> 10
cugaaguuug ugcuuaaaaa a 21
Claims (16)
1. detecting application of the reagent of biomarker in sample in preparing the product for diagnosing osteosarcoma, feature exists
In the biomarker is FIBCD1.
2. application according to claim 1, which is characterized in that the sample is tissue.
3. application according to claim 1, which is characterized in that with reference to compared with, biomarker expression level raises.
4. application according to claim 1, which is characterized in that the reagent is selected from：
The probe of specific recognition FIBCD1 genes；Or
The primer of specific amplification FIBCD1 genes；Or
Specifically bind the antibody or ligand of the albumen of FIBCD1 codings.
5. application according to claim 4, which is characterized in that the primer sequence such as SEQ of specific amplification FIBCD1 genes
Shown in NO.1 ~ 2 ID.
6. following any one of them application：
1）Application of the product in the tool for preparing diagnosis osteosarcoma, the product include the reagent for detecting FIBCD1 levels；
2）Application of the inhibitor of FIBCD1 in the pharmaceutical composition for preparing treatment osteosarcoma；
3）Application of the inhibitor of FIBCD1 in the pharmaceutical composition for preparing treatment bone and flesh tumor metastasis；
4）Applications of the FIBCD1 in the drug candidate of screening treatment osteosarcoma；
5）Application of the pharmaceutical composition in the product for preparing treatment osteosarcoma, described pharmaceutical composition includes the inhibition of FIBCD1
Agent, and/or other medicine classes and pharmaceutically acceptable carrier and/or auxiliary material with the inhibitor compatibility.
7. application according to claim 6, it is characterised in that 1）Described in reagent be selected from：
The probe of specific recognition FIBCD1 genes；Or
The primer of specific amplification FIBCD1 genes；Or
Specifically bind the antibody or ligand of the albumen of FIBCD1 codings.
8. application according to claim 7, which is characterized in that the primer sequence such as SEQ of specific amplification FIBCD1 genes
Shown in NO.1 ~ 2 ID.
9. application according to claim 6, which is characterized in that 2）Or 3）The pharmaceutical composition includes and the inhibition
Other medicine classes and pharmaceutically acceptable carrier and/or auxiliary material of agent compatibility.
10. application according to claim 9, which is characterized in that the inhibitor is selected from：Nucleic acid inhibitor, albumen inhibit
Agent, proteolytic enzyme, protein binding molecule.
11. application according to claim 10, which is characterized in that the inhibitor is nucleic acid inhibitor siRNA.
12. application according to claim 11, which is characterized in that the sequence of siRNA is as shown in NO.9 ~ 10 SEQ ID.
13. application according to claim 6, which is characterized in that 4）The step of drug candidate of middle screening treatment osteosarcoma
Including:
The system for the albumen expressed or containing FIBCD1 genes or its coding is handled with substance to be screened；With
Detect the expression of the albumen of FIBCD1 genes or its coding or activity in the system；
Wherein, if the substance to be screened can inhibit expression or the activity of the albumen of FIBCD1 genes or its coding, show
The substance to be screened is the drug candidate for treating osteosarcoma.
14. application according to claim 6, which is characterized in that 5）Described in inhibitor be selected from：Nucleic acid inhibitor, albumen
Inhibitor, proteolytic enzyme, protein binding molecule.
15. application according to claim 14, which is characterized in that the inhibitor is nucleic acid inhibitor siRNA.
16. application according to claim 15, which is characterized in that the sequence of the siRNA is as shown in NO.9 ~ 10 SEQ ID.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201711445268.6A CN107881240B (en) | 2017-12-27 | 2017-12-27 | The diagnosis and treatment marker of osteosarcoma |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201711445268.6A CN107881240B (en) | 2017-12-27 | 2017-12-27 | The diagnosis and treatment marker of osteosarcoma |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN107881240A CN107881240A (en) | 2018-04-06 |
| CN107881240B true CN107881240B (en) | 2018-10-02 |
Family
ID=61771415
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201711445268.6A Active CN107881240B (en) | 2017-12-27 | 2017-12-27 | The diagnosis and treatment marker of osteosarcoma |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN107881240B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109655608B (en) * | 2018-12-13 | 2020-02-21 | 杭州多泰科技有限公司 | Exosome protein for osteosarcoma diagnosis and instant detection method thereof |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2008119356A1 (en) * | 2007-04-03 | 2008-10-09 | Syddansk Universitet | The use of a type ii transmembrane protein designated fibcd1 as an endocytosis receptor of acetylated compounds |
| WO2010075891A1 (en) * | 2008-12-30 | 2010-07-08 | Syddansk Universitet | Fibcd1 for the prevention and treatment of diseases |
| CN102448484A (en) * | 2009-04-01 | 2012-05-09 | 医学研究，基础设施和卫生服务基金的特拉维夫医疗中心 | A method of regulating proliferation and differentiation of keratinocyes |
| CN104651509A (en) * | 2015-02-13 | 2015-05-27 | 北京泱深生物信息技术有限公司 | New drug target for osteosarcoma |
| CN106834486A (en) * | 2017-02-28 | 2017-06-13 | 北京泱深生物信息技术有限公司 | Osteosarcoma molecule diagnosis and treatment mark and its application |
-
2017
- 2017-12-27 CN CN201711445268.6A patent/CN107881240B/en active Active
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2008119356A1 (en) * | 2007-04-03 | 2008-10-09 | Syddansk Universitet | The use of a type ii transmembrane protein designated fibcd1 as an endocytosis receptor of acetylated compounds |
| WO2010075891A1 (en) * | 2008-12-30 | 2010-07-08 | Syddansk Universitet | Fibcd1 for the prevention and treatment of diseases |
| CN102448484A (en) * | 2009-04-01 | 2012-05-09 | 医学研究，基础设施和卫生服务基金的特拉维夫医疗中心 | A method of regulating proliferation and differentiation of keratinocyes |
| CN104651509A (en) * | 2015-02-13 | 2015-05-27 | 北京泱深生物信息技术有限公司 | New drug target for osteosarcoma |
| CN106834486A (en) * | 2017-02-28 | 2017-06-13 | 北京泱深生物信息技术有限公司 | Osteosarcoma molecule diagnosis and treatment mark and its application |
Also Published As
| Publication number | Publication date |
|---|---|
| CN107881240A (en) | 2018-04-06 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN107435074A (en) | Application of the CES8 genes in clear cell carcinoma of kidney diagnosis and treatment | |
| CN109468382A (en) | Application of the lncRNA in adenocarcinoma of lung diagnosis and treatment | |
| CN107164554B (en) | Application of ASPRV1 as biomarker in diagnosis and treatment of laryngeal squamous cell carcinoma | |
| CN106834486A (en) | Osteosarcoma molecule diagnosis and treatment mark and its application | |
| CN107312855B (en) | Gene related to laryngeal squamous cell carcinoma and application thereof | |
| CN107881240B (en) | The diagnosis and treatment marker of osteosarcoma | |
| CN105483246B (en) | Application of the differential expression of gene in carcinoma of mouth diagnosis | |
| CN105288659B (en) | The application of TENM1 gene and its expression product in diagnosis and treatment papillary adenocarcinoma | |
| CN108165631B (en) | Osteosarcoma biomarker SYT12 and application thereof | |
| TW201204393A (en) | Diagnostic agent and therapeutic agent of cancer | |
| CN107779503A (en) | The related difference expression gene of Alzheimer and its application | |
| CN105233290B (en) | The application of C22orf26 genes and its expression product in Parkinson's diagnosis and treatment reagent is prepared | |
| CN107496923A (en) | A kind of biomarker related to clear cell carcinoma of kidney | |
| CN113293208B (en) | Molecular marker related to lung cancer proliferation and metastasis and application thereof | |
| CN107893119B (en) | Application of ZCCHC12 in osteosarcoma | |
| CN108165624A (en) | Application of the biomarker in osteosarcoma diagnosis and treatment | |
| CN107385081B (en) | Gene related to kidney cancer and application thereof | |
| CN106729756A (en) | Application of the biomarker as target in adenocarcinoma of lung diagnosis and treatment | |
| CN108220443A (en) | Applications of the CGREF1 as marker in clear cell carcinoma of kidney diagnosis and treatment | |
| CN107184983B (en) | Diagnosis and treatment target for lung adenocarcinoma | |
| CN108114283A (en) | Applications of the biomarker MUC21 in osteosarcoma diagnose and treat | |
| CN105385781B (en) | Application of the LCE3E in diagnosis and treatment carcinoma of mouth | |
| CN110042164A (en) | Lung cancer diagnosis and treatment lncRNA marker | |
| CN104740649B (en) | Applications of the PLEKHA5 in tumour diagnostic reagent is prepared | |
| CN109371136A (en) | One kind lncRNA relevant to adenocarcinoma of lung and its application |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| TR01 | Transfer of patent right | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20210402 Address after: 266000 room 2503, Qianshan building, D2, Qingdao International Innovation Park Phase II, No.1 Keyuan Weiyi Road, Laoshan District, Qingdao City, Shandong Province Patentee after: Qingdao Yangshen biomedical Co.,Ltd. Address before: 100080 room 3103, cube court building, 1 good luck street, Haidian District, Beijing. Patentee before: BEIJING MEDINTELL BIOMED INFORMATION TECHNOLOGY Co.,Ltd. |