CN110172448A - A kind of synovial sarcoma cells system hSS-005R and its progeny cell system - Google Patents
A kind of synovial sarcoma cells system hSS-005R and its progeny cell system Download PDFInfo
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- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; CARE OF BIRDS, FISHES, INSECTS; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; CARE OF BIRDS, FISHES, INSECTS; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; CARE OF BIRDS, FISHES, INSECTS; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
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Abstract
The present invention provides a kind of synovial sarcoma cells system hSS-005R, is deposited in China typical culture collection center, and deposit number is CCTCC NO:C201911.The present invention also provides a kind of progeny cell of synovial sarcoma cells system hSS-005R as described above systems.The present invention establishes a kind of new people's synovial sarcoma cells system, and character is stablized, and can stablize multiple passage.Under the premise of people's synovial sarcoma cells that the present invention establishes tie up to reservation Major Clinical biological property, with tumor formation rate height, incubation period is short, the features such as homogeneity is good, enrich synovial sarcoma cells library, synovial sarcoma animal model can successfully be prepared, obtained animal model can be used for basic research and drug screening, to provide strong scientific research data based on the research that Chinese population genetic background is carried out, also the test for new drug preclinical study experiment in vivo for clinical anti-cancer drug susceptibility and drug resistance provides new test material.
Description
Technical field
The invention belongs to tumour cell research fields, and in particular to a kind of synovial sarcoma cells system hSS-005R and its filial generation
Cell line.
Background technique
Synovial sarcoma (synovial sarcoma) is that a kind of invasion is strong, and the high tumour of grade malignancy, synovial sarcoma accounts for
The 5-10% of all soft tissue sarcomas accounts for 10-20% in the soft tissue neoplasm of teenager and young man.Nineteen eighty-three is to 2012
The disease incidence of Nian Jian, synovial sarcoma continue to increase from 0.906% to 1.548%.Synovial sarcoma is apt to occur in four limbs, accounts for owning
The 70% of synovial sarcoma case.The common metastasis site of synovial sarcoma is lung, followed by lymph node and marrow.Synovial sarcoma is most
Just name is that have phase on heterogeneous microstructure with the synovial tissue of surrounding because it is found in the tissue of juxtra-articular
Like property.However, more in recent years research shows that: synovial sarcoma may alternatively appear in oesophagus, lung, and heart etc. lacks the tissue of synovial membrane,
Fail to agree with such title.The definite origin of cell of synovial sarcoma is still a unsolved mystery, and the scholar of part thinks that it may come
From in mescenchymal stem cell.In addition, synovial sarcoma can behave as the tumour point of epithelial tissue source tumour and mesenchymal cell origin
The marker of change, although synovial sarcoma and the organization type similitude of above two source tumour are lower.Some scholars incline at present
To in the tumour that synovial sarcoma is defined as to " uncertain differentiation ".Synovial sarcoma is defined as follows by the World Health Organization: with one
Determine the Spindle Cell Tumors in the mesenchymal tissue source of degree differentiation (including body of gland ingredient).Synovial sarcoma can in Histopathology
To be classified as: biphasic or bipolar type has different degrees of epithelial cell and spindle cell ingredient；(2) unidirectional fiber type, only by shuttle shape
Cell composition；(3) single-phase epitheliated type, is mainly made of epithelial cell, and there is only a small amount of spindle cell groups；(4) low differentiated,
Can be similar to has the characteristics that advanced nucleus, height mitotic activity and the small round cell neoplasm containing necrotic zone, greatly
Cell/Epithelial Tumors or Spindle Cell Tumors.Wherein in multiple groups data, it is most common it is all in report be unidirectional fiber
Type synovial sarcoma, grade malignancy is very high, grows extremely rapid.And biphasic or bipolar type synovial sarcoma is most special in synovial sarcoma tissue
One kind of sign property, containing spindle cell and fibrocyte, the tumour cell that spindle cell and epithelioid cell are combined into is mutual
It migrates.There is research to think, not instead of jumping characteristic when synovial sarcoma changes from unipolar type to biphasic or bipolar type synovial sarcoma is presented one
The form of a gradual change conversion.
Most of research is thought: new adjuvant chemotherapy or the extensive tumorectomy with radiotherapy are still synovial membrane meat
The prefered method of tumor treatment.Even if receiving the clinical treatment of specification, effective survival rate is still less than 10% in 10 years, only
8.9%.So improving the therapeutic scheme of synovial sarcoma to improve patient's prognosis as an intractable problem.
Currently, the main problems in the progress of numerous limitation synovial sarcomas treatments are how construction one rationally and has
The tumor model of effect.
Cell line (cell line) refers to the cell colony bred after primary cell culture is passed on successfully for the first time.Especially
It is the cell for tiding over second of dead crisis divided by 40-50 times, referred to as cell line.Because of the cell one of originally culture
As reach or so 10 generations and be just not easy to relay again, the growth of cell just will appear stagnation, most cells aging death.But
" crisis " can be spent and continue to relay by having the cell of only a few, and the cell of these survivals is generally possible to pass to 40--50 generation,
This passage cell is called cell strain.No change has taken place for the inhereditary material of cell strain.It can go out again after cell reached for 50 generations
Existing " crisis ", cannot relay again.But there is the inhereditary material of part cell to be changed, and have canceration the characteristics of,
It is possible that unlimitedly passage is gone down under culture conditions, this passage cell is known as cell line.
Energy infinite multiplication and the cell line for stablizing passage can provide stable character and abundance for the research of synovial sarcoma
Raw material, but the synovial sarcoma cells system that can at home and abroad buy at present is extremely limited, wherein cell bank can be directly at home
The synovial sarcoma cells system of purchase only has one kind, SW-982.And the biology of existing synovial sarcoma cells system and clinical synovial sarcoma
It is larger to learn character difference.Therefore, for the relevant research of synovial sarcoma cells, there is a need in the field to provide more kinds of new synovial membrane meat
Oncocyte system with and its preparation method and application.
Summary of the invention
The technical problem to be solved in the present invention is to low for existing people's synovial sarcoma cells system bio-diversity, especially
It is that Establishment of Cell Line success rate is low, while existing cell line differs the deficiencies of larger with the biological character of clinical synovial sarcoma,
A kind of new people's synovial sarcoma cells system and its method for building up and application are provided.
The present invention provides a kind of synovial sarcoma cells system hSS-005R, is deposited in China typical culture collection center,
Deposit number is CCTCC NO:C201911.
The present invention also provides a kind of progeny cell of synovial sarcoma cells system hSS-005R as described above systems.
In a kind of specific embodiment, the cell line c-myc oncogene height expression.
In a kind of specific embodiment, SS18-SSX fusion is not present in the cell line.
The present invention also provides a kind of construction methods of the mouse in vivo models of cell line as described above, including take hSS-005R
Single side Mice Inoculated under cell skin, after inoculation 1~3 week, tumour is initially formed and is grown, and 30~50 days mouse are doctor after inoculation
Mouse model is used in medicine research.
In a kind of specific embodiment, the mouse is immunodeficient mouse, and the preferably described immunodeficient mouse is
Nude mice or SCID mouse.
The present invention also provides the preparation method of synovial sarcoma cells system hSS-005R a kind of, the synovial sarcoma cells system is protected
China typical culture collection center is ensconced, deposit number is CCTCC NO:C201911；The preparation method includes primary training
Feeding and secondary culture.
Skilled person will appreciate that, originally culture is cell monolayer directly to be grown by tissue block or with enzyme or machinery
Tissue dispersion is started to cultivate by method at individual cells, and the culture before passing on for the first time is regarded as originally culture.Originally culture
Biggest advantage is that tissue and cell are just in vitro, and not yet great changes will take place for biological character, to a certain extent can antimer
Interior state.And secondary culture is defined as: after the cell monolayer that originally culture is formed converges, it is separately cultured, it is no
Then cell can cause nutrition exhausted because of generation insufficient space or since cell density is excessive, all will affect the growth of cell, this
Program is frequently referred to passage or secondary culture.In general, secondary culture refers to expansion culture, that is, a cell is divided into two or
Person's one dividing into three is cultivated etc..But narrowly, cell is shifted or is transplanted to separately from a culture bottle no matter dilute
One culture bottle is known as passage or secondary culture.It is appreciated that at any time, cell is inoculated into another from a bottle
A part is always lost when bottle, therefore, must be diluted in objectively cell.
In a kind of specific embodiment, the originally culture is adhere-wall culture or enzyme digestion culture.In the present invention,
Enzyme digestion includes that digestive ferment is added to make histolysis and discharge cell.The enzyme digestion culture is those skilled in the art
Routine operation known to member.
In a kind of specific embodiment, the originally culture is adhere-wall culture, fresh the method includes that will obtain
Clinical people's synovial sarcoma patients surgery tumor resection sample be cut into 3-5mm3Thin piece, it is adherent in cell culture bottom of bottle by thin piece
Adhere-wall culture after adhere-wall culture 1-2 weeks, climbs out of tumour cell successively in tumor tissue, then thin to the tumour of originally culture
Born of the same parents carry out secondary culture.
In a kind of specific embodiment, the secondary culture includes the training discarded in the culture bottle for carrying out originally culture
Fresh trypsase cell dissociation buffer is added into culture bottle for nutrient solution, after cell detachment, terminates digestion, is added fresh
Complete medium concussion or beating are allowed to be detached from bottle wall formation cell suspension, and centrifugation resuspension is inoculated in new culture bottle and continues to train
It supports；Every time when cell cover with entire culture bottle or it is long to 80% or more when then sub-bottle carry out secondary culture, every bottle of cell is carrying out
The generation tumour cell is divided into 2~5 bottles when secondary culture, continues to cultivate its filial generation tumour cell in these new culture bottles.
The present invention also provides a kind of synovial sarcoma cells system hSS-005R as described above to screen or prepare anti-synovial sarcoma
Application in cell drug.
The present invention also provides a kind of vivo experiment method of screening treatment synovial sarcoma drug candidate, the experiment in vivo sides
Method is led after application the following steps are included: be applied to the mammal tumor model of hSS-005R cell tumor formation for compound is tested
The test compound for causing the gross tumor volume of synovial sarcoma to reduce or disappear is exactly to treat synovial sarcoma candidate compound.
In the present invention, the hSS-005R cell each means the cell in synovial sarcoma cells system hSS-005R.
The present invention also provides a kind of ill vitro test method of screening treatment synovial sarcoma drug candidate, the experiment in vitro sides
Method is the following steps are included: be directly applied to hSS-005R for the combination for testing compound or different test compounds with various concentration
Cell judges to test compound according to test compound of various concentration or combinations thereof to the inhibitory effect of tumor cell proliferation
To the anti-tumor capacity of synovial sarcoma.
In a kind of specific embodiment, the outer experimental method is the following steps are included: 1) by the hSS-005R
Cell is inoculated in 96 porocyte culture plates holes with the density in 3000/ hole, is cultivated 24 hours；2) diluted chemical compound will be tested at not
Same concentration is applied to hSS-005R cell, by the vigor of measurement cell after drug effect 24 hours, calculates the change of various concentration
Object is closed to the Proliferation Ability ability of cell, to judge the anti-tumor capacity of test compound.
In a kind of specific embodiment, the test compound includes cis-platinum, Doxorubicin and c-myc inhibitor
One of Kj-pyr-9 and 10058-F4 or a variety of.
The present invention also provides a kind of synovial sarcoma cells system hSS-005R as described above to prepare synovial sarcoma cells model
Or the application in animal model, the animal in the preferably described animal model are immunodeficient mouse.
Under the premise of people's synovial sarcoma cells that the present invention establishes tie up to reservation Major Clinical biological property, there is tumor formation
The features such as rate is high, and incubation period is short, and homogeneity is good, enriches synovial sarcoma cells library, to be carried out based on Chinese population genetic background
Research provide strong scientific research data, also for new drug preclinical study experiment in vivo be directed to clinical antitumor medicine sensibility and
The test of drug resistance provides new test material；And the people's synovial sarcoma cells system is suitable to synovial sarcoma clinic first-line drug-
Platinum drug resistance is the good test material for studying synovial sarcoma resistance mechanism；It is also one group excellently to cisplatin resistance new mechanism
Exploratory development material has very high scientific research value and development significance.
The reagents and materials used in the present invention are commercially available.
Beneficial effects of the present invention at least that:
1. the present invention establishes a kind of new people's synovial sarcoma cells system, character is stablized, and can stablize multiple passage, for cunning
Film sarcoma research provides the new experimental material closer to clinical tumor biological characteristics.
2. people's synovial sarcoma cells that the present invention establishes tie up to retain Major Clinical biological property under the premise of, have at
The features such as ratio of outflow is high, and incubation period is short, and homogeneity is good, enriches synovial sarcoma cells library, can successfully prepare synovial sarcoma animal
Model, obtained animal model can be used for basic research and drug screening, for what is carried out based on Chinese population genetic background
Research provides strong scientific research data, is also new drug preclinical study experiment in vivo for clinical anti-cancer drug susceptibility and resistance to
The test of pharmacological property provides new test material.
3. people's synovial sarcoma cells system that the present invention establishes is acquired to synovial sarcoma clinic front-line chemotherapeutic agents-cis-platinum
Drug resistance, in vivo with stable cisplatin resistance characteristic is showed in experiment in vitro, this exploration for cisplatin resistance new mechanism is
One group of excellent research material has very high scientific research value and development significance, is the good of research synovial sarcoma resistance mechanism
Test material.
4. people's synovial sarcoma cells system that the present invention establishes is by can be used to compared with nude mice interior generation parental tumor
The correlation of external, internal drug susceptibility and drug resistance is analyzed, and then can establish external, internal two associated drugs
Screening Platform is the ideal cell line of the basic research of people's synovial sarcoma and preclinical phase application.
The preservation of biomaterial
People's synovial sarcoma cells of the invention, which are lain in, is deposited in China typical culture collection center on January 15th, 2019
(CCCTCC) (address: Wuhan, China Wuhan University postcode 430072), culture title are as follows: people's synovial sarcoma cells hSS-
005R, deposit number are as follows: CCTCC NO:C201911.
Detailed description of the invention
Fig. 1 is the morphological observation of hSS-005R cell under an optical microscope.A. (100 times) B. (200 times).
Fig. 2 is short-movie section repetitive sequence (CSTR) qualification result of hSS-005R cell.
Fig. 3 is the flow cytometry analysis hSS-005R cell cycle.
Fig. 4 is inhibiting effect of the multiple anticancer drugs of testing in vitro to hSS-005R cell Proliferation.
Fig. 5 is the Tumor formation of hSS-005R cell, animal tumor tumor growth curve.Wherein A is tumour in nude mouse
Growth curve, B be tumour in the intracorporal growing state photograph taking of nude mice.
Fig. 6 is the pathological tissue of primary tumor tissue and hSS-005R the cell tumor formation in nude mouse.Wherein, A: primary swollen
Tumor tissue；B:hSS-005R cell.
Specific embodiment
The present invention is further illustrated below by the mode of embodiment, but does not therefore limit the present invention to the reality
It applies among a range.In the following examples, the experimental methods for specific conditions are not specified, according to conventional methods and conditions, or according to quotient
The selection of product specification.
In the present invention, the hSS-005R cell, which is each meant, is deposited in China typical culture collection center, and preservation is compiled
Number be CCTCC NO:C201911, the cell in synovial sarcoma cells system hSS-005R.
Embodiment 1
Embodiment 1 is the preparation of cell line hSS-005R cell of the present invention.
Primary culture method: by the Fresh synovial sarcoma sample cut off in art be placed under aseptic conditions DMEM high sugar+
In 10%FBS complete medium, save on ice.It is rinsed with fresh PBS buffer solution (containing 1% dual anti-- penicillin, streptomysin)
Afterwards, it thinly slices, the tissue shredded is adherent in Tissue Culture Flask bottom of bottle, it is complete that suitable DMEM high sugar+10%FBS is added
Culture is uprightly placed 1 hour based on 37 DEG C of 5% carbon dioxide conditions of cell incubator, and tissue block is made to paste jail in bottom of bottle.Then will
Upright culture bottle, which softly slowly fell, makes complete medium submerge adherent tissue block.Without changing liquid, can be seen after general 4-5 days
Cell is observed to climb out of from tissue block.The culture medium more renewed when culture medium turns yellow when cells being waited to cover with substantially, is passed as normal
Secondary culture is carried out for mode.
Secondary culture: the culture solution in the culture bottle for carrying out originally culture is discarded, fresh pancreas egg is added into culture bottle
White enzyme cell dissociation buffer terminates digestion after cell detachment, and fresh complete medium concussion or beating is added and is allowed to be detached from bottle
Wall forms cell suspension, and centrifugation is inoculated in new culture bottle and continues to cultivate；Culture bottle of every replacement is then equivalent to secondary culture
Once；Every time when cell cover with entire culture bottle or it is long to 80% or more when then sub-bottle carry out secondary culture, every bottle of cell into
The generation tumour cell is divided into 2~5 bottles when row secondary culture, continues to cultivate its filial generation tumour in these new culture bottles thin
Born of the same parents.
In the present invention, by after 50 generation of primary cell secondary culture of tumor tissues, cell line is obtained, is named as
HSS-005R, submits preservation, and deposit number is CCTCC NO:C201911.
Embodiment 2
Embodiment 2 is biological characteristics and the application of hSS-005R cell.
The present invention, can external long term growth and stabilization using the sugared culture solution culture purified hSS-005R cell of DMEM high
Passage.When it is more than generation to reach 30 for cell, cell quality is gradually stable, carries out relevant biology, science of heredity and tissue-derived mirror
It is fixed, until the 50th generation stable character all having the same.The hSS-005R cell can form tumour in nude mouse, have
Oncogenicity.The clinical synovial sarcoma tumor specimen of the hSS-005R cell and its source, nude mice interior generation parental tumor are formed
Corresponding relationship can provide new test material to study the correlation of external, internal and clinical anti-cancer drug susceptibility and drug resistance
Material.It is specific as follows:
A. morphological observation
The culture bottle for cultivating hSS-005R cell is placed under inverted microscope, is observed under bright-field, it is seen that
HSS-005R cell bodies are in paving stone sample, and it to be in Clonal growth that there are a small amount of irregular cells, be can express when density is larger viscous
Attached property aggregation, the result is shown in Figure 1 (A100 times, B200 times).
B. short-movie section repetitive sequence CSTR) identification
Short tandem repeat (short tandem repeat, STR) is also known as microsatellite DNA, refers on chromosome,
A kind of DNA sequence dna (the number of repetition 10- formed by several base-pairs as core unit (2-6 base-pair), tandem sequence repeats
More than 60 times, genetic fragment is below 400 base-pairs)；Each duplicate number of core unit will appear individual difference, to be formed
The different allele of fragment length.Therefore, the number of repetition of one group of STR sequence is almost unique in Different Individual, is
The main method that the gene identities feature and cell biology of individual identify cell identity and source.
The hSS-005R cell for collecting fresh cultured, uses the Animal genome extraction agent box (article No. of Tsingke
TSP201-200 the genomic DNA for) extracting cell carries out PCR amplification with the primer of 5 ' end fluorescent markers, carries out to products therefrom
Sequencing, analysis include Amelogenin, THO1, TPOX, D13S317, vWA, D16S539, D5S818, CSF1PO and D7S820
Etc. the sequence repeat number of each STR bit point, wherein the primer sequence of STR bit point is as shown in table 1.Above-mentioned sequence and ATCC, DSM2Z
The database that equal cells save library carries out inquiry comparison, identical genetic map is not returned to, in American Type Tissue Culture
STR sequence retrieval is carried out in the heart (CATCC) database, does not find identical STR testing result.It is possible thereby to prove that its is unique
Property, and the cross contamination with other cells does not occur during originally culture (testing result is shown in Fig. 2).
As can be seen from Figure 2, synovial sarcoma cells system hSS-005R provided by the invention belongs to completely new people's synovial sarcoma cells
System.
The primer sequence in the site table 1:STR
C. cell cycle distribution
Collect about 106For a hSS-005R cell in 1.5ml centrifuge tube, supernatant is abandoned in centrifugation.Cell precipitation is with 1 milliliter -20
DEG C 75% ethyl alcohol is resuspended, and room temperature fixes 1 hour.Supernatant is abandoned in centrifugation, and 500 microlitres of PI dyeing liquors are added.It mixes, incubation at room temperature 30
Minute.With the total DNA content of the every hole cell number of flow cytomery and each cell (the total DNA content of each cell and should
Total PI fluorescence intensity of cell is proportional)；And when according to the different cell cycles, the variation of cell total DNA content is calculated each thin
The total number of cells in born of the same parents' period.Detection obtains period profile and each period profile ratio is as shown in Figure 3.
Skilled person will appreciate that, the interphase of cell cycle was divided into for three phases, i.e. DNA pre-synthesis phase (G1 phase), DNA is closed
At phase (S phase) and DNA post-synthesis phase (G2 phase).The 1.G1 phase (first gap) be from mitosis to DNA replication dna before one section when
Phase, also known as pre-synthesis phase, this phase mainly synthesize RNA and ribosomes.The phase feature is that metabolism is active, synthesize rapidly RNA and
Protein, cell volume significantly increase.The major significance of this phase is that the DNA replication dna for phase next stage S performs substance and energy
The preparation of amount.2.S phase (synthesis) the i.e. DNA synthesis phase other than synthetic DNA, while will also synthesize histone in this phase.
Enzyme required for DNA replication dna all synthesizes in this period.The 3.G2 phase (second gap) is DNA post-synthesis phase, is mitosis
Preparatory stage.In this period, DNA synthesis is terminated, a large amount of to synthesize RNA and protein, including tubulin and maturation-promoting factor
Deng.And Proliferating antigen Ki67 refers to that the sum of S phase and G2 phase cell account for the ratio of total cell number, reflects the proliferation speed of this group of cells
Degree.The S phase increases, and indicates that cell division is active.
As can be seen from Figure 3, the proliferative capacity of synovial sarcoma cells system hSS-005R cell is active in the present invention.The cell meets
The proliferative properties of tumour cell.
D. cell in vitro poison is tested
External test synovial sarcoma clinic Common Chemotherapy drug-cis-platinum (Cislatin), Doxorubicin (doxorubicin
That is adriamycin) and Kj-pyr-9 (c-myc inhibitor), 10058-F4 (c-myc inhibitor) hSS-005R cell line is resisted
Proliferation function.Subject cell is seeded in 96 orifice plates with the density in 3000/ hole, drug and drug combination 24 to various concentration
After hour, the cell viability under each drug concentration is detected with CCK8.Wherein, the drug concentration of two kinds of drugs of drug combination respectively accounts for
Half, total dosage are identical with the dosage of the other drugs compared with concentration.Wherein, Doxorubicin combines Kj-pyr-9 (c-myc
Inhibitor) medication effect is best, as a result see Fig. 4.
E. the Tumor formation of cell
In vitro culture and collection hSS-005R cell, inoculating nude mice (has purchased from Hunan Si Laike scape up to experimental animal
Limit company), every animal inoculation pvaccination 0.2ml contains 1.0 × 107A cell, is inoculated with 6 mouse, unilateral side inoculation, and N=6 is monitored weekly
The weight of animals and tumor size.After inoculation about 2 weeks, tumour is initially formed and is grown, and tumor formation rate is essentially 100%.Draw tumour
Growth curve, wherein gross tumor volume=(long × wide × wide) ÷ 2；Its result is as shown in Figure 5.From figure 5 it can be seen that hSS-005R cell
Tumour can be formed in immunodeficient mouse body and is grown uniform quick.
After the subcutaneous tumor formation of f.hSS-005R cell mouse, paraffin embedding after tumour is fixed is taken out, preparation, which is sliced, simultaneously carries out HE
Dyeing.Fig. 6 A is basic stitch tumor mass, and Fig. 6 B is hSS-005R cell mouse tumor mass, and pathological diagnosis result is synovial sarcoma, group
Form height is knitted close to basic stitch tumor mass.As a result see Fig. 6.This illustrates that cell line hSS-005R cell of the present invention can be very well
Ground reflects the case where former tumor tissues.
G. full exon genes convergence analysis
From the point of view of the result of Gene Fusion, the phenomenon that hSS-005R cell does not all contain SSX Gene Fusion, and support gene
Readable (read) number of fusion is all seldom, can be determined as background noise substantially.So this example synovial sarcoma tumor model
For the negative synovial sarcoma of clinically rare SS18-SSX expression.It the results are shown in Table 2.
Table 2: fusion result
As known to those skilled in the art: most of synovial sarcoma there is t (X, 18；P11p, q11) transposition, thus
Synovial sarcoma one mostly important specific fusion gene SS18-SSX is generated, fusion translation forms fusion protein
SS18-SSX.And during the entire process of this chromosome translocation mutation is always present in synovial sarcoma cells occurrence and development.Cause
And thering are some scholars to think, fusion caused by synovial sarcoma chromosome translocation, SS18-SSX may be used as synovial sarcoma
Proto-oncogene, drive the occurrence and development of tumour.It is present in most of synovial sarcoma in view of SS18-SSX fusion,
Specificity with height, this provides one for the specific diagnosis of synovial sarcoma and important makes a definite diagnosis thinking: being examined by molecule
The presence of survey means detection fusion gene SS18-SSX diagnoses synovial sarcoma, specificity with higher.Although for synovial membrane meat
For tumor, there is the fusion of SS18-SSX in the patient of the synovial sarcoma more than 95%, occurs merging without containing SS18-SSX
The synovial sarcoma probability of gene is lower than 5%.
That is, being a kind of rare parting, Ke Nengqi in synovial sarcoma cells system hSS-005R provided by the invention
Grade malignancy is relatively higher, and the medicament categories that perhaps can treat the parting are less, and correlative study also accordingly can be less.
The variation of h.MYC copy number
There is the phenomenon that a large amount of copy number amplification and a small amount of copy number missing in No. 8 chromosomes of hSS-005R cell.In table 3
The section of leukorrhagia marking covers MYC gene, and the variation of CNV copy number reaches 7, ProbCall 0.999999998, accurately
Degree is high.It the results are shown in Table 3.
The variation of table 3:MYC copy number
As known to those skilled in the art: c-myc oncogene is related with kinds of tumors occurrence and development.Cell line of the present invention
By its c-myc oncogene height expression known to gene extron sequencing expression.The sequencing shows the synovial sarcoma cells system
Cell infinite multiplication characteristic.
The above content is combine specific preferred embodiment to the further description of the invention made, and it cannot be said that originally
The specific implementation of invention is only limited to these instructions.For those of ordinary skill in the art to which the present invention belongs, not
Under the premise of being detached from present inventive concept, several simple deductions and replacement can also be made, all shall be regarded as belonging to guarantor of the invention
Protect range.
Claims (6)
1. a kind of synovial sarcoma cells system hSS-005R, is deposited in China typical culture collection center, deposit number is
CCTCC NO:C201911.
2. the progeny cell of synovial sarcoma cells system hSS-005R as described in claim 1 a kind of is.
3. cell line according to claim 1 or claim 2, which is characterized in that the cell line c-myc oncogene height expression.
4. cell line according to claim 1 or claim 2, which is characterized in that there is no SS18-SSX to merge base in the cell line
Cause.
5. a kind of construction method of the mouse in vivo models of the cell line as described in any one of Claims 1 to 4, including take
Single side Mice Inoculated under hSS-005R cell skin, after inoculation 1~3 week, tumour is initially formed and is grown, 30~50 days after inoculation
Mouse is medical research mouse model.
6. construction method according to claim 5, which is characterized in that the mouse is immunodeficient mouse, preferably described
Immunodeficient mouse is nude mice or SCID mouse.
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