CN101113179B - Human antibody sample molecule TEM8-Fc based on tumour endocytosis sign 8 and its application in tumour treatment - Google Patents
Human antibody sample molecule TEM8-Fc based on tumour endocytosis sign 8 and its application in tumour treatment Download PDFInfo
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Abstract
The invention discloses a human antibody sample molecular TEM8-Fc based on tumor endothelium marker 8 (TEM8) and the application thereof in tumor therapy and pertains to the biological pharmaceutical field. TEM8-Fc fusion protein is recombinant dimer fusion protein by the linkage of the 1-227 amino acid at extracellular domain of the TEM8, fc fragment, heavy chain constant region 2 and heavy chain constant region 3 of human immunoglobulin IgG1; mature single strand TEM8-Fc has 432 amino acid residues and the dimer has 864 amino acid residues. The invention has the advantages of small side effect, high specificity and wide indication. Animal experiments show that TEM8-Fc can not only effectively inhibit growth of tumor cells such as breast cancer, colorectal cancer, liver cancer, etc. in nude mice, but also can inhibit metastasis of cancer cells such as breast cancer, colorectal cancer, liver cancer, etc., and the angiogenesis in tumor tissues and reverse tumor cells phenotype.
Description
Technical field
The present invention relates to a kind of human antibody sample molecule TEM8-Fc and application in oncotherapy thereof, belong to field of biological pharmacy based on tumor endothelial cell mark 8 (TEM8).
Background technology
It is all processes that must take place that malignant solid tumor took place and developed late period that vasculogenesis and tumour cell shift.The key molecule that shifts with vasculogenesis and tumour cell becomes the desirable target of developing anti-tumor medicine naturally, and the medicine of development antagonism or blocking-up vasculogenesis and tumor cell invasion transferring path should have the activity of broad-spectrum anti-tumor.
It is the main promoting factor of malignancy of tumor growth and transfer that angiogenic growth is regulated out of control.Therefore, suppress tumor vascular growth, promptly the antitumor strategy of " tumour hungry to death " has caused tumor research person and clinician's very big interest and extensive concern.Theoretically, this antitumor strategy has following many-sided advantage: 1. the malignancy of most of solid tumor and transfer all need competent blood supply, and therefore suppressing angiogenic growth has the broad-spectrum anti-tumor characteristic; 2. different with the anti-tumor method of routine, suppress tumor vascular growth at be the metastable endotheliocyte of genotype, therefore avoided the generation of drug resistance of tumor to a certain extent; 3. the antineoplastic vascular of venoclysis growth medicine can arrive and act on the vascular endothelial cell of tumor tissues rapidly, has so just avoided the pharmacokinetics problem that conventional antitumor drug faced; 4. each endotheliocyte is all supported the growth of the tumour cell of some amount, and inhibition of endothelial cell proliferation is also with the growth of indirect inhibition tumour cell, promptly so-called " bystander effect ".Developed multiple antitumor drug at present at vascular endothelial growth factor (VEGF) and acceptor (VEGFR) thereof, especially seal the monoclonal antibody Avastin of VEGF, obtained widespread use clinically, this has confirmed to suppress the validity of the antitumor strategy of angiogenic growth actually.
Though the target spot that present inhibition angiogenic growth class medicine is acted on is woven with more abundant expression in tumor group, is not in the specific expression of tumor vascular endothelium.This has limited the clinical application of this class medicine to a certain extent.Therefore, bring into play the potential that suppresses the antitumor strategy of angiogenic growth fully, need to explore and find the specific expressed molecular marker of tumor vascular endothelium.Fortunately, a nearest research has found that by comparing healthy tissues and tumor tissues medium vessels endothelial cell gene expression difference a class has higher specific expressed molecular marker at tumor vascular endothelial cell.This class mark be named as the tumor endothelial cell mark (tumor endothelial markers, TEMs).Meaningfully, in nine kinds of such molecular markers, the multiple epicyte protein that belongs to is arranged, molecular structure has tangible acceptor sample feature.On evolving, this quasi-molecule has very high conservative property.Because TEMs is specific expressed in some tissue and the cell of tumour cell and embryonic development period, the normal cell expression level is not seldom even expressed, even if some TEMs does not express when wound repair yet, and VEGF expression, therefore, can infer, be that its specificity of medicine of the inhibition tumor vessel formation of target exploitation is higher than the VEGF target possibly with TEMs, and Side effects of pharmaceutical drugs may be less.High conservative, tumour expression specificity and acceptor sample feature three big characteristics have determined that this quasi-molecule may be an exploitation antibody class medicine, suppresses the highly desirable target of angiogenic growth and treatment tumour.
Also do not develop the report that the TEM8-Fc fusion rotein is used for oncotherapy at present both at home and abroad.
Summary of the invention
The technical problem to be solved in the present invention is: a kind of safe and effective human antibody class artificial protein TEM8-Fc fusion rotein based on TEM8 is provided.
Another technical problem that the present invention will solve provides the TEM8-Fc fusion rotein and expresses the purposes of CH0 clone in the medicine of preparation treatment cancer of this fusion rotein.
For achieving the above object, the present invention is by the following technical solutions:
A kind of human antibody sample molecule (TEM8-Fc) fusion rotein of tumor endothelial cell mark 8 is Fc section (dimer fusion proteins that hinge area (Hinge region), CH2 (CH2) and CH3 (CH3) are connected to form of tumor endothelial cell mark 8 (TEM8) extracellular region 1-227 amino acids and human normal immunoglobulin IgG1; Strand TEM8-Fc molecule has the aminoacid sequence shown in the sequence table SEQ ID No.1, totally 459 amino-acid residues; Wherein preceding 27 signal peptides that amino acid is ATR; Sophisticated strand ATR-Fc has 432 amino-acid residues, and dimer is 864 amino-acid residues.
Aminoacid sequence shown in the sequence table SEQ ID No.1 is identical with anthrax toxin acceptor ATR-Fc fusion protein in the Chinese patent 200510084233.5.
The encode gene of human antibody sampling molecule fusion protein strand of above-mentioned tumor endothelial cell mark 8.
The gene of described strand has the base sequence shown in the sequence table SEQ ID No.2.
The expression carrier that contains the human antibody sampling molecule fusion protein of codes for tumor endotheliocyte mark 8.
This carrier is a carrier for expression of eukaryon.
The clone that contains described expression vector.Described cell is a Chinese hamster ovary celI.
The TEM8-Fc fusion rotein is used to prepare the purposes of the medicine for the treatment of tumour.
Described tumour is mammary cancer, colorectal cancer, liver cancer and other tumours.
Described treatment is meant the phenotype that suppresses growth of tumor and transfer, the interior vasculogenesis of inhibition tumor tissues and reversing tumor cell.
The TEM8-Fc genetic engineering fusion protein, be according to Chinese patent 200510084233.5 described technological methods, to express the gene of TEM8-Fc fusion rotein, to the CHO-K1 cell, be 10-15 μ g/ (10 with G418 screening and acquisition TEM8-Fc expression level by the liposome method transfection
6Cellsd) genetically engineered Chinese hamster ovary celI is that ATR-Fc-ID5 (preserves at China Committee for Culture Collection of Microorganisms common micro-organisms center, register on the books and be numbered: CGMCCNo.1399, the cell strain name of preserving is called ATR-Fc fusion rotein CHO engineering cell strain, address: No. 13, North No.1 Row, Zhongguancun, Haidian District, Beijing City, Institute of Microorganism, Academia Sinica, 100080; Phone: 010-62542758; Fax: 010-62537796; E-mail: cgmcc@sun.im.ac.cn).According to Chinese patent ZL200310124257.X (Hu Xianwen etc., zooblast porous microcarrier immobilization high-efficient culture method and substratum thereof, Chinese patent ZL200310124257.X) described cultural method and substratum, serum-free culture ATR-Fc-1D5 genetically engineered Chinese hamster ovary celI (rCHO) adopts albumin A affinity chromatography method purification of recombinant proteins (Chinese patent 200510084233.5).
The present invention is based on tumor endothelial cell mark---TEM8, developed a kind of antibody molecule TEM8-Fc of full-length human, preliminary result of study shows, this antibody has the multiple tissue-derived human tumor of being transplanted to nude mice and surpasses 90% inhibiting rate, has demonstrated good broad-spectrum anti-tumor growth and transfer character.
The application that gene recombination TEM8-Fc fusion rotein of the present invention is used to prepare the medicine for the treatment of tumour.Find first with the ELISA method, the native ligand of TEM8 is uPA (urokinase type plasminogen activator), and much studies show that, nearly all malignant tumour, especially late period, all overexpression uPA and acceptor (uPAR) thereof on its tumor cell membrane, uPA plays keying action in multiple malignant tumour Invasion and Metastasis such as colorectal carcinoma, ovarian cancer, mammary cancer, cancer of the stomach, prostate cancer and lung cancer, participated in tumor growth, vasculogenesis and transfer.Identify the affinity of TEM8-Fc and uPA, provide explanation TEM8-Fc to suppress growth and Transfer Mechanism in the tumour cell body, be that TEM8-Fc can catch uPA in vivo, thereby stop the uPA to combine with TEM8 on the cytolemma, blocked that TEM8 mediates with relevant signal paths such as growth of tumour cell, migration.Experimentation on animals confirms: TEM8-Fc can obviously suppress the growth and the transfer of breast cancer cell, colon cancer cell and liver cancer cell, and this fusion rotein is worth further studying and being developed to the antibody class medicine of the tumour for the treatment of mammary cancer, colorectal carcinoma, liver cancer and other uPA or TEM8 overexpression.
Select the target of tumor-blood-vessel growth as oncotherapy, have the following advantages: (1) side effect is little, and vasculogenesis seldom takes place the healthy tissues of generally being grown up, so the antagonizing vessel generation can not cause serious toxic side effect; (2) specificity height, because the unstable of the heredity of tumour cell, the expression pattern of the specificity marker thing on its film surface has very big difference, for example, even if all be mammary cancer, patient's ratio of overexpression EGFR II is only less than 30%, therefore the humanized antibody Herceptin (He Saiting) that treats a line medication anti-EGFR II of advanced breast cancer needs to use just effectively behind the screening patient, and the vasculogenesis of tumour almost all plays a role in all solid tumor growths and transfer process, and do not have the variation problem, so the specificity of angiogenesis inhibitor treatment is higher; (3) indication is wide, and market in the future is big.Antineoplastic vascular generates, and not only is confined to one type cancer, may be all effective in the treatment of many solid tumors, and indication is wide.In fact, the humanized antibody Avastin (A Wasiting) of the anti-VEGF that is used for the treatment of colorectal cancer of FDA approval in 2004, finished the III clinical trial phase that is used for nonsmall-cell lung cancer and breast cancer treatment with chemotherapy drugs in combination in the first half of the year in 2005, obtain clinical efficacy preferably, be expected to obtain in the recent period the treatment that the FDA approval is used for nonsmall-cell lung cancer and mammary cancer.(4) antineoplastic vascular of venoclysis growth medicine can arrive and act on the vascular endothelial cell of tumor tissues rapidly, has so just avoided the pharmacokinetics problem that conventional antitumor drug faced.(5) clinical proof, the angiogenesis inhibitor treatment of tumour has the characteristic of better curative effect and broad-spectrum anti-tumor really.In addition, because TEM8 is specific expressed in tumour cell, therefore use TEM8-Fc as the interaction of disturbing between TEM8 and other albumen, antitumor action is more special, can not damage healthy tissues.
The present invention will be further described below in conjunction with accompanying drawing and better embodiment, so that the public has whole to summary of the invention and understand fully, and is not qualification to protection domain of the present invention.Aforementioned part fully discloses the protection domain that the present invention can implement, and therefore allly any well known in the artly is equal to replacement according to what the disclosure of invention was carried out, all belongs to infringement of the present invention.
Description of drawings
Fig. 1 is the structural representation of TEM8-Fc.
Fig. 2 (a) detects the glycosylation TEM8-Fc fusion rotein of purifying for SDS-PAGE.
Fig. 2 (b) detects the TEM8-Fc fusion rotein of the removal sugar chain of purifying for SDS-PAGE.
Fig. 3 combines with the uPA specificity for ELISA detects TEM8-Fc.
Fig. 4 (a) is the surperficial overexpression TEM8 with flow cytometry analysis liver cancer cell HepG2.
Fig. 4 (b) is the migration of TEM8-Fc vitro inhibition LMW-UK inductive HepG2 cell.
Fig. 5 (a) and Fig. 5 (b) are TEM8-Fc suppresses tumor growth to nude inoculation colorectal cancer cell LS180 exercising result (effect that suppresses tumor growth in the body).
Fig. 6 (a) and Fig. 6 (b) are TEM8-Fc suppresses tumor growth to nude inoculation breast cancer cell MCF-7 exercising result (effect that suppresses tumor growth in the body).
Fig. 7 is in the experiment of MCF-7 mammary cancer nude mice lotus knurl, and the TEM8-Fc fusion rotein is to suppressing the result of angiogenic action in the tumor tissues.
Fig. 8 is that the TEM8-Fc fusion rotein is the result of normal phenotype to tumour cell reversing in the experiment of LS180 colorectal cancer nude mice lotus knurl.
Embodiment
Embodiment 1.SDS-PAGE analyzing gene recombinant products
Press Chinese patent 200510084233.5 described methods, expressing gene reorganization TEM8-Fc fusion rotein in the rCHO cell, and cultivate the rCHO cell with serum free medium (Chinese patent ZL 200310124257.X), with the TEM8-Fc in nProtein A Sepharose 4 Fast Flow affinity column (Amersham Biosciences) the purifying cells culture supernatant.The schematic arrangement of TEM8-Fc fusion rotein as shown in Figure 1.If TEM8-Fc can correctly express in Chinese hamster ovary celI, it should be the homodimer antibody molecule that expressed proteins is estimated, promptly the TEM8-Fc strand connects into dimer (Fig. 1) at hinge area by interchain disulfide bond.Strand TEM8-Fc molecule has 459 amino-acid residues, wherein preceding 27 signal peptides that amino acid is TEM8, and exocytosis is cut in cells and supernatant the time, so actual strand TEM8-Fc has 432aa, dimer is 864aa.Because TEM8 and Fc section all have glycosylation site, so the molecular weight of strand TEM8-Fc is expected at 60-65kD.The TEM8-Fc fusion rotein that affinitive layer purification obtains carries out SDS-PAGE respectively and analyzes (concentrating gum concentration is 5%, and resolving gel concentration is 10%), coomassie brilliant blue staining under reduction and non-reduced condition.Under non-reduced condition, a protein band is arranged at the 120-130kD place, and under reductive condition, a tangible protein band is arranged at the 60-65kD place, and do not have other protein band (Fig. 2 (a)).Because all disulfide linkage comprise that interchain disulfide bond is all interrupted under the reductive condition, the molecular weight that records is a strand TEM8-Fc molecule, but not different peptide chains still connect by disulfide linkage under the reductive condition, at the molecular weight under the non-reduced condition just in time is the twice of molecular weight under the reductive condition, show that the TEM8-Fc fusion rotein that we obtain is the homodimer antibody molecule, conform to expected result.
With 20 μ g TEM8-Fc with 20 μ l sex change damping fluid (0.5%SDS, the 1%2-mercaptoethanol) under 100 ℃ of conditions, boils sex change in 10 minutes, add 2 μ l, 10 * reaction buffer (500mM sodium phosphate then, pH7.5,10%NP-40) and the Glycosylase PNGase F (Sigma) of 200 activity units.Under 37 ℃ of conditions, hatched 1 hour, remove the N-sugar chain in the TEM8-Fc molecule.Under reductive condition, analyze with SDS-PAGE (concentration of separation gel is 12%).The result shows that the molecular weight of deglycosylated TEM8-Fc single chain molecule is about 50kD (Fig. 2 (b)), and is consistent with theoretical value, shows that the gene recombination TEM8-Fc fusion rotein that we obtain is the glycoprotein of homodimer.
Embodiment 2.ELISA confirms that TEM8 combines with the specificity of uPA or tPA
With gene recombination uPA (Pro-UK, source: Hu Xianwen, Xiao Chengzu, Li Zuohu.Pilotproduction of u-PA with porous microcarrier cell culture.Cytotechnology.2000.33 (1～3): 13-19), gene recombination low molecular weight urokinase (LMW-UK, Chinese patent: low molecular-weight urokinase mutant and expression vector thereof, application number: 200610065813.4, publication number CN1880449) and (tPA of gene recombination tissue-type plasminogen activator, source: Roche company) wrap by the NUNCTM elisa plate by 0.8 μ g/ hole, 3%BSA sealing back adds the TEM8-Fc fusion rotein, hatch the back thorough washing, how anti-add the anti-human IgG of HRP mark goat (H+L) enzyme mark again, hatch the back and add the TMB reagent colour development, 405nm surveys the OD value.With the negative contrast of Humanized monoclonal antibodies Herceptin (Roche).The result as shown in Figure 3, we find first, but the TEM8-Fc specificity is in conjunction with Pro-UK and UK (being collectively referred to as uPA), and promptly uPA is the native ligand of TEM8, and almost all overexpressions in all malignant tumours of uPA, stimulate the growth and the migration of tumour cell, and the vasculogenesis of promotion tumor tissues, therefore, TEM8-Fc can catch uPA, blocking-up uPA promotes the propagation and the transporting action of tumour cell, thereby suppresses growth of tumor and transfer.In contrast, being used for the humanized antibody Herceptin of the anti-Her2 of mammary cancer (the Her2 positive) treatment then can not be in conjunction with uPA or tPA.
Embodiment 3.TEM8-Fc fusion rotein can suppress the migration of uPA inductive tumour cell
Because it is very important that uPA plays a part in the migration of tumour cell, sealing uPA may suppress the migration of tumour cell.With the surperficial overexpression TEM8 (Fig. 4 (a)) of flow cytometry analysis liver cancer cell HepG2 (ATCC No.HB8065), how anti-available from Santa Cruz company the anti-TEM8 rabbit of using in this experiment is.Adopt Transwell cell in vitro migration experiment (source: Corning Costar Corporation, thereby http://www.tc.umn.edu/～shimi002/SOPtranswell.pdf) checking TEM8-Fc has the transporting action that sealing uPA suppresses the HepG2 tumour cell.
Method: the HepG2 cell is by every hole 1 * 10
5Individual cell inoculation is to the upper strata cell of Transwells, and the HepG2 cell acts on 4 hours with LMW-UK (10nmol/L) and moves to stimulate, and observes the sealing process of TEM8-Fc (25g/mL) to LMW-UK.Act on after 4 hours, the cell that migrates to lower floor's cell is also taken a picture with 0.5% violet staining.
The results are shown in Figure the migration that 4 (b): LMW-UK can significantly promote the HepG2 cell, behind the adding TEM8-Fc, can suppress the cell migration of LMW-UK inductive.This result shows that TEM8-Fc may stop the transfer of tumour in vivo.
Embodiment 4.TEM8-Fc fusion rotein suppresses the effect of tumor growth to the cancer cells in nude inoculation human body source
Method: human colon cancer cell LS180 in the vegetative period of taking the logarithm (ATCC No.CL-187) or human breast cancer cell MCF-7 (ATCC No.HTB-22) are mixed with 5 * 10 with fresh medium DMEM (Hyclone)
6The cell suspension of individual/mL, every nude mice (Beijing Vital River Experimental Animals Technology Co., Ltd., www.vitalriver.com.cn) armpit subcutaneous vaccination 0.2ml, inoculation back mouse is divided into three dosage groups of 10,4 and 0.8mg/kg of physiological saline group, endoxan (10mg/kg), TEM8-Fc, totally 5 groups at random.Inoculate beginning administration in the 2nd day, administering mode: the next day subcutaneous injection, altogether administration is 10 times.The administration volume is the 0.2ml/20g body weight, takes off neck execution animal in 2 days after the last administration, dissects and gets the knurl piece, claims knurl heavy, calculates the tumor growth tumour inhibiting rate according to following formula:
Result: TEM8-Fc shows the very strong inhibition tumor growth and the effect of vasculogenesis in the experiment of nude mice lotus knurl.The effect that TEM8-Fc treatment group suppresses tumour has tangible dose-dependently, increase with antibody administration dosage, tumor killing effect improves, and colorectal carcinoma lotus knurl experimental result shows that the tumour inhibiting rate of TEM8-Fc 10mg/kg, 4mg/kg, 0.8mg/kg is respectively 91%, 85%, 77% (Fig. 5).The tumour inhibiting rate of chemotherapeutics endoxan (10mg/kg CTX) group is about 90%, and high dose group TEM8-Fc tumor killing effect is suitable with anticarcinogen CTX, and does not almost have significant side effects, and that CTX organizes the mouse body weight is light, the obvious atrophy of spleen.Mammary cancer lotus knurl experimental result shows that the tumour inhibiting rate of TEM8-Fc10mg/kg, 4mg/kg, 0.8mg/kg is respectively 92%, 67%, 54% (Fig. 6).Show that TEM8-Fc can be used for treating colorectal cancer, mammary cancer etc.
Embodiment 5.TEM8-Fc fusion rotein can suppress the transfer of tumour cell
In the experiment of nude mice lotus knurl, can obviously reduce the incidence of cancer metastasis with the TEM8-Fc treatment, and obviously reduce metastasis quantity, and be significant dose-dependently relation (table 1).With the experiment of mammary cancer MCF-7 nude mice lotus knurl is example, TEM8-Fc treatment group metastasis of cancer incidence is significantly less than control group, and, dosage is 10.0mg/kgTEM8-Fc treatment group, there is not the cancer metastasis incident to take place, therefore, TEM8-Fc can not only suppress growth of tumor, also can be used to prevent the transfer of tumour cell.
Table 1.TEM8-Fc presses down the hepatic metastases that suppresses tumour cell in the knurl experiment at nude inoculation breast cancer cell MCF-7
aMean+SD,
bP＜0.05vs control group.
Embodiment 6.TEM8-Fc fusion rotein can suppress the vasculogenesis in the tumor tissues
In the experiment of nude mice lotus knurl, can obviously reduce the generation of tumor tissues medium vessels with the TEM8-Fc treatment, and be significant dose-dependently relation (Fig. 7).With the experiment of colorectal cancer LS180 cell nude mice lotus knurl is example, carries out the experiment of nude mice lotus knurl according to the method for embodiment 4, and the human colon cancer cell LS180 in the vegetative period of promptly taking the logarithm is mixed with 5 * 10 with fresh medium DMEM (Hyclone)
6The cell suspension of individual/mL, every mouse armpit subcutaneous vaccination 0.2ml, inoculation back mouse is divided into three dosage groups of 10,4 and 0.8mg/kg of physiological saline group, endoxan (10mg/kg), TEM8-Fc, totally 5 groups at random.Inoculate beginning administration in the 2nd day, administering mode: the next day subcutaneous injection, altogether administration is 10 times.The administration volume is the 0.2ml/20g body weight, takes off neck execution animal in 2 days after the last administration, dissects and gets the knurl piece.With paraffin embedding knurl piece, section.The immunohistochemical methods method please refer to document: Nanda A., et al.TEM8 interacts with the cleavedC5domain of collagen α 3 (VI) .Cancer Research.64,817-820,2004.Simply, paraffin section is hatched dewaxing back horseradish peroxidase confining liquid (DAKO) processing in 20 minutes in 95 ℃ citrate buffer solution (pH6.0).Rabbit with anti-vWF (vWF ELISA) resists (Sigma) to hatch more afterwards, and the goat-anti rabbit two with horseradish peroxidase-labeled resists (Santa Cruz Biotechnogies) combinations again, with diaminobenzidine (DAB) dyeing.Haematoxylin redyeing is used in section at last again.The result shows that TEM8-Fc treatment group can significantly reduce the vessel density (arrow shows vascular endothelial cell among Fig. 7) in the tumor tissues, and blood vessel increases and obviously reduces with TEM8-Fc dosage.Therefore, TEM8-Fc can suppress the vasculogenesis in the tumor tissues.
The phenotype of embodiment 7.TEM8-Fc fusion rotein energy reversing tumor cell
Carcinomebryonic antigen (CEA) is the specific expressed membranin of embryonic development period and malignant tumor tissue, seldom expresses in adult's healthy tissues.TEM8 is the tumor endothelial cell mark, and is specific expressed in the tumor endothelial cell surface, and its concrete physiological function it be unclear that, and it is generally acknowledged that TEM8 has brought into play vital role in tumor vessel formation and tumor cell invasion.According to embodiment 6 described immunohistochemical methods methods, use the anti-CEA rabbit how anti-(Santa CruzBiotechnogies) expression of detecting the CEA in the tissue slice of tumour in the experiment of colorectal cancer LS180 cell nude mice lotus knurl find, colorectal carcinoma through the TEM8-Fc treatment, its expression of tumor tissue carcinomebryonic antigen (CEA) reduce along with the increase of TEM8-Fc dosage, colorectal carcinoma tumor tissues of organizing at 4mg/kg group and 10mg/kg even the expression of not finding CEA, especially 100mg/kg organizes, the form (Fig. 8) of noble cells appears in colon cancer cell, be that TEM8-Fc can not only directly control growth of tumor, and may promote that tumour cell transforms to normal cell, TEM8 and CEA are the oncofetal proteins of just expressing malignant tumour and embryonic development period, and the relation between the two it be not immediately clear.
We find that first TEM8-Fc can significantly suppress mammary cancer, colorectal carcinoma, liver cancer etc. in the intravital growth of nude mice, also can suppress tumor vascular formation, and have the promotion tumour cell to the function that the normal cell phenotype transforms, and have the broad-spectrum anti-tumor characteristic.The invention provides a kind of treat late malignant tumour, safe and effective based on TEM8 human antibody class TEM8-Fc fusion rotein.
Sequence table
＜110〉Biologic Engineering Inst., Academy of Millitary Medical Sciences of P.L.A
＜120〉a kind of based on the human antibody sample molecule TEM8-Fc of tumor endothelial cell mark 8 and in oncotherapy
Use
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＜213〉synthetic
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Claims (2)
1. the TEM8-Fc fusion rotein of the aminoacid sequence shown in the sequence table SEQ ID No.1 is used to prepare the purposes of the medicine for the treatment of mammary cancer, colorectal cancer, liver cancer.
2. TEM8-Fc fusion rotein according to claim 1 is used to prepare the purposes of treatment mammary cancer, colorectal cancer, liver-cancer medicine, it is characterized in that: described treatment is meant the phenotype that suppresses growth of tumor and transfer, the interior vasculogenesis of inhibition tumor tissues and reversing tumor cell.
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