CN1342077A - Inhibition of formation of vascular hyperpermeability - Google Patents

Abstract

Vascular hyperpermeability and the subsequent events such as macular edema, retinoblastoma, ocular ischemia, ocular inflammatory disease or infection, choroidal melanoma, edematous side-effects induced by iron chelation therapy, pulmonary edema, myocardial infarction, rheumatoid diseases, anaphylaxis, allergies, hypersensitive reactions, cerebral edema, brain tumor fluid-filled cysts, communicating hydrocephalus, carpal tunnel syndrome, organ damage resulting from a burn, irritation or infection, erythema multiforme, edematous macules and other disorders, brain tumors, tumor effusions, lung or breast carcinomas, ascites, pleural effusions, pericardial effusions, high altitude ''sickness'', radioanaphylaxis, radiodermatitis, glaucoma, conjunctivitis, choroidal melanoma, adult respiratory distress syndrome, asthma, bronchitis, ovarian hyperstimulation syndrome, polycystic ovary syndrome, menstrual swelling, menstrual cramps, stroke, head trauma, cerebral infarct or occlusion, hyotension, ulcerations, sprains, fractures, effusions associated with synovitis, diabetic complications, hyperviscosity syndrome, liver cirrhosis, microalbuminuria, proteinuria, oliguria, electrolyte imbalance, nephrotic syndrome, exudates, fibroses, keloid, can be inhibited by the administration of a compound that inhibits the enzyme activity of the VEGF tyrosine kinase receptor known as KDR tyrosine kinase. The preferred compound 4,5-dihydro-3-pyridin-4-yl-1(2)H-benzo[g]indazole selectively inhibits the function of KDR tyrosine kinase but do not block the activity of Flt-1 tyrosine kinase which is another VEGE tyrosine kinase receptor.

Description

Suppress the too high formation of vascular permeability

Background of the present invention

Edema can be described as the increase of interstitial fluid volume.This is normally unusual, typically need seek to alleviate the patient's condition.This patient's condition is usually because the endothelium permeability is too high makes liquid leave the blood vascular system to cause, and exosmoses with macromole usually and finds new dwelling relevant in a matter space.

There are multiple physiology and biochemical mechanism to explain edema and the formation of edema state in the individuality.In one or more these mechanism, a kind of important media is " a vascular endothelial cell growth.This factor raises the transportation in the vascular endothelial cell, and causes multiple vascular lamina such as skin, subcutaneous tissue, and peritoneal wall, mesentery, diaphragm, trachea, bronchus, the permeability in duodenum and uterus etc. increases.Significant blood cell oozes out, and passes the change of the exchange of endothelium, and macromole oozes out and deposits and the hypopiesia that prolongs may be followed the permeability effect of this increase these positions.Think that these processes are the preludes that promote neovascularization.VEGF is by inflammatory T-cell, and macrophage is bitten neutrophilic granulocyte and bitten eosinophile etc. and express in inflamed sites.This factor can be by histanoxia, some blood vessel pressurization hormone, and somatomedin, reproductive hormone and multiple inflammatory cytokine raise.Hinted the vascular permeability of VEGF-mediation with such as tumor ascites, endometriosis, adult respiratory distress syndrome (ARDS), back cardiopulmonary bypass-relevant hypopiesia and permeability crossed bleb occurred frequently, edema reaction to burn and wound, endothelial function disorder in the diabetes, ovary is high to stimulate the syndrome complication, and disease such as ocular edema is relevant.

Like this, it is useful obviously suppressing VEGF generation or activity, particularly blocks the performance of above-named disease.Especially, the permeability medicament too high and edema and related syndromes that can suppress VEGF mediation is useful for relaxing these diseases.

Protein tyrosine kinase.Protein tyrosine kinase (PTK) comprises a large amount of inhomogeneous protein that enzymatic activity is arranged.PTKs plays an important role in growth of control cell and differentiation, and (summary is referring to Schlessinger ﹠amp; Ullrich, 1992, Neuron 9:383-391).

The abnormal stimulation that has shown PTK is expressed or sudden change can cause uncontrolled cell proliferation (for example malignant growth) or crucial growth, the defective of adjusting or repair process.Therefore, the suitable financial resources of biomedical group's cost are found PTK family member's single-minded biological agent, their functions in atomization, they are to the participation of tumor generation and other disease, the biochemical foundation of their the activatory signal transduction pathway of ligand stimulation and the exploitation of novel drugs.

Tyrosine kinase can be receptor type (have outside the born of the same parents, wear film and born of the same parents' intracellular domain) or non-receptor type (being in the born of the same parents fully).

Receptor tyrosine kinase (RTK).RTKs comprises the different bioactive membrane receptors of wearing of having of extended familys.At present, 19 different RTK subfamilies have been identified at least.Receptor tyrosine kinase (RTK) family comprises the growth of various kinds of cell type and breaks up vital receptor (Yarden and Ullrich, Ann.Rev.Biochem.57:433-478,1998; Ullrich and Schlessinger, Cell 61:243-254,1990).The inherent function in conjunction with activation RTK of part causes the phosphorylation of receptor and various kinds of cell the 5th, and causes various kinds of cell reaction (Ullrich ﹠amp subsequently; Schlessinger, 1990, Cell 61:203-212).Like this, interacted the receptor tyrosine kinase Mediated Signal Transduction initial, typically followed, stimulated inherent protein hydroxyphenylaminopropionic acid kinase activity and receptor transphosphorylation by receptor dimerizationization with the born of the same parents of single-minded somatomedin (part) are outer.So just produced the binding site of intracellular signal transduction molecule and cause and promote a series of kytoplasm signal transduction molecules of cell effect to form complexs.(for example cell division breaks up, metabolic effect, born of the same parents are the change of microenvironment outward) and referring to Schessinger and Ullrich, 1992, Neuron 9:1-20.

There are SH2 (src congener-2) or phosphotyrosine to combine with activatory tyrosine kinase receptor and substrate thereof, signal is transmitted in the cell in conjunction with the high affinity of the protein ground of (PTB) domain.Two kinds of domains are all discerned phosphotyrosine.(SH2:Fantle etc., 1992, Cell69:413-423; Songyant etc., 1994, Mol.Cell.Biol.14:2777-2785; Songyang etc., 1993, Cell 72:767-778; With Koch etc., 1991, Science252:668-678; Schoelson, Curr.Opin.Chem.Biol. (1997), 1 (2), 227-234; Cowburn, Curr.Opi.Struct.Biol. (1997), 7 (6), 835-838).Identified several iuntercellular substrates (intercellular substrate) protein relevant with receptor tyrosine kinase (RTK).They can be divided into two big classes: (1) has the substrate of catalyst structure domain; (2) lack this domain but as adapter and the substrate (Songyang etc., 1993, Cell 72:767-778) relevant with the catalytic activity molecule.The SH2 of receptor or protein and their substrate or the interactional specificity of PTB domain are to be determined by the amino acid residue around the phosphorylated tyrosine residue next-door neighbour.For example, the difference in conjunction with affinity between the aminoacid sequence around the phosphotyrosine is and observed their difference relevant (Songyang etc., 1993, Cell 72:767-778) of substrate phosphorylation characteristic on SH2 domain and the special receptor.Observe hint, the function of each receptor tyrosine kinase is not only determined by its expression way and part availability, and is determined by the arrangement of the activated downstream signal transduction pathway of special receptor and the selection of time and the persistent period of these stimulations.Like this, phosphorylation provides important regulating step, is replenished by specific growth factor receptors and differentiation factor receptors, and it has determined the selectivity of signal transduction path.

Pointed out multiple receptor tyrosine kinase and in vascularization, worked, though be work indirectly (Mustonen and Alitalo, J.Cell Biol.129:895-898,1995) sometimes in conjunction with the somatomedin on it.A kind of being called of this receptor tyrosine kinase " tire liver kinases 1 " is a member in the III type subclass of RTKs (FLK-1).The another kind name of people FLK-1 is the " receptor (KDR) (Terman etc., Oncogene 6:1677-83,1991) that contains kinases insert structure territory.The name of the another kind of FLK-1/KDR be " vascular endothelial growth factor receptor 2 " (VEGFR-2) because its can high affinity in conjunction with VEGF.The FLK-1/VEGFR-2 of Mus is also referred to as NYK (Oelrichs etc., Oncogene 8 (1): 11-15,1993).Separated encoding murine, the DNA of rat and people FLK-1 has also reported nucleotide and amino acid sequence coded (Matthews etc., Proc.Natl.Acad.Sci.USA, 88:9026-30,1991; Terman etc., 1991, supra; Terman etc., Biochem.Biophysic.Res.Comm.187:1579-86,1992; Sarzani etc., supra; With Millauer etc., Cell 72:835-846,1993).Such as Millauer etc., hint VEGF and FLK-1/KDR/VEGFR-2 are that ligand-receptor is right in the multiple researchs such as ibid literary composition, in vascular endothelial cell proliferation and vascularization and growth, be called that blood vessel takes place and angiogenesis in play an important role.

Another kind of called after " fms sample tyrosine kinase-1 " III type subclass RTK (Flt-1) relevant with FLK-1/KDR (DeVries etc., Science 255; 989-991,1992; Shibuya etc., Oncogene 5:519-524,1990).Another name of Flt-1 be " vascular endothelial growth factor receptor 1 " (VEGFR-1).Up to now, find that FLK-1/KDR/VEGFR-2 and flt-1/VEGFR-1 subfamily member mainly express on endotheliocyte.The single-minded member by vascular endothelial cell growth factor (VEGF) ligand family of these subclass member stimulates (Klagsburn and D ' Amore, Cytokine ﹠amp; Growth FactorReviews 7:259-270,1996).Vascular endothelial cell growth factor (VEGF), in conjunction with Flt-1 than in conjunction with FLK-1/KDR have higher affinity and to the vascular endothelial cell mitogenesis (Terman etc., 1992, ibid; Mustonen etc., ibid; DeVries etc., ibid).Believe that Flt-1 is important for endothelial tissue in vascular development.The early stage vascular development and the neovascularization in the wound healing relevant (Mustonen and Alitalo, ibid) of the expression of Flt-1 and mice embryonic.Flt-1 is becoming human organs such as the expression in the glomerule to hint this receptor and irrelevant another function (Mustonen and Alitalo, ibid) of cell growth.

As mentioned above, nearest evidence shows that VEGF is stimulating normally and all work in the vascularization of pathology (Jackeman etc., Endocrinology 133:848-859,1993; Kolch etc., Breast Cancer Research and Treatment 36:139-155,1995; Ferrara etc., Endocrine Reviews 18 (1); 4-25,1997; Ferrara etc., Regulation of Angiogenesis (ed.L.D.Goldberg and E.M.Rosen), 209-232,1997).In addition, the control of VEGF and vascular permeability with strengthen relevant (Connolly, etc., J, Biol.Chem.264; 20017-20024; Brown etc., Regulation of Angiogenesis (ed.L.D.Goldberg and E.M.Rosen), 233-269,1997).

Reported the multi-form VEGF that produces by the alternative splicing of mRNA, comprised four kinds (J.Cell.Biochem.47:211-218,1991) that Ferrara etc. describes.Ferrara etc. have identified the VEGF of excretory and main cell related specy as previously described, and known this albumen exists with the binary form that disulfide bond connects.

Identified the relevant homologue of a plurality of VEGF recently.But, do not illustrate their effects in physiology and pathological process.In addition, the common and VEGF coexpression of the member of VEGF family in multiple tissue, and can form heterodimer with VEGF usually.As described below, this specific character may change the receptor specificity and the biological effect of heterodimer, and further makes the complexity of illustrating change (Korpelainen and Alitalo, the Curr.Opin.Cell Biol. of their single-minded effect, 159-164,1988 and the document wherein quoted).

The aminoacid sequence of placental growth factor (PIGF) and VEGF sequence have significant homology (Park etc., J.Biol.Chem.269:25646-54,1994; Oncogene 8:925-31 such as Maglione, 1993).Similar VEGF, different types of PIGF is from the difference splicing of mRNA, and there are (Park etc., ibid) in protein with the binary form.PIGF-1 and PIGF-2 can high-affinity be attached to Flt-1, and PIGF-2 also be attached to strongly neuropilin-1 (Migdal etc., J.Biol.Chem.273 (35): 22272-22278), but all do not combine with FLK-1/KDR (Park etc., supra).Be reported that when the VEGF low concentration exists PIGF can strengthen vascular permeability and the mitogenesis effect (it is said it is because the formation of heterodimer) (Park etc., ibid) of VEGF to endotheliocyte.

It seems that VEGF-B produces with two kinds of isoform forms (167 and 185 residues), also combine with Flt-1/VEGFR-1.It may regulate the extracellular matrix degraded by the expression and the activity of regulating urokinase type plasminogen activator and plasminogen activation inhibitor 1, cell adhesion, with work in the migration (Pepper etc., Proc.Nstl.Acad.Sci.U.S.A. (1998), 95 (20): 11709-11714).

At first with VEGF-C as mainly cloning by the part of the VEGFR-3/Flt-4 of lymph endotheliocyte expression.The VEGF-C of full form processing also can and stimulate the propagation of external endotheliocyte and the vascularization in migration and the body inner model (Am.J.Pathol. (1998) such as Lymboussaki, 153 (2): 395-403 in conjunction with KDR/VEGFR-2; Am J.Pathol. such as Witzenbichler (1998) 153 (2), 381-394).The overexpression of VEGF-C only causes vasculolymphatic propagation and expansion, but blood vessel is unaffected.Different with VEGF, the expression of VEGF-C be not subjected to the inducing of histanoxia (Ristimake etc., J.Biol.Chem. (1998), 273 (14), 8413-8418).

Very similar on the VEGF-D structure of recent findings with VEGF-C.Report VEGF-D combination and activation at least two kinds of VEGFRs, VEGFR-3/Flt4 and KDR/VEGFR-2.It is the earliest as the derivable fibroblastic mitogen of c-fos, cloned, and main (Achen etc., the Proc.Natl.Acad.Sci.U.S.A. (1998) of in the mesenchymal cell of lung and skin, expressing, 95 (2), 548-553 and list of references wherein).

When being expelled to skin histology, Miles detects and shows induction of vascular permeability (PCT/US97/14696 in VEGF-C and the VEGF-D body; WO98/07832, Witzenbichler etc., ibid).Physiological action and importance that these parts are regulated in the tissue that their are expressed in the too high and endothelium reaction of vascular permeability are also uncertain.

Based on other VEGF of continuous discovery and the precedent of VEGFRs homologue and part and the different dimerization of receptor, the effect of these VEGF homologues may relate to formation VEGF part dimer, and/or receptor heterodimer, or be attached to also undiscovered VEGFR (Witzenbichler etc., ibid).Nearest report also hints, neuropilin-1 (Migdal etc., ibid) or VEGFR-3/Flt-4 (Witzenbichler etc., ibid) and KDR/VEGFR-2 outside receptor may participate in causing induction of vascular permeability (Stacker, SA, Domagala, T., Nice, E., and Wilks, A.F., " Angiogenesis and Cancer " Conference, Amer.Assoc.Cancer Res., Jan.1998, Orlando, FL; Williams, Diabetelogia 40:S118-120 (1997)).

The chemical compound of PTKs is regulated in exploitation.Consider the control of the PTKs on cell proliferation of supposition, regulate and modulation, and disease relevant and disorderly importance with the cell proliferation of abnormality, use several different methods to carry out a large amount of trials and identified receptor and nonreceptor tyrosine kinase " inhibitor ", comprise that (U.S. applies for No.4 to use mutant part, 966,849), soluble recepter and antibody (application No.WO 94/10202; Kendall ﹠amp; Thomas, 1994, Proc.Natl.Acad.Sci 90:10705-09; Kim etc., 1993, Nature 362:841-844), RNA part (Jellinek etc., Biochemistry 33:10450-56; Takano etc., 1993, Mol.Bio.Cell 4:358A; Kinsella waits 192, Exp.CellRes.199:56-62; Wright is etc., 1992, J.Cellular Phys.152:448-57) and tyrosine kinase inhibitor (WO 94/03427; WO 92/21660; WO 91/15495; WO94/14808; U.S. patent No.5,330,992; Mariani, etc., 1994, Proc.Am.Assoc.Cancer Res.35:2268).

Recently attempted identifying micromolecule as tyrosine kinase inhibitor.For example, described two monocycles, dicyclo or heterocyclic aryl chemical compound (PCT WO 92/20642) and 1,2 ethenylidene-7-azaindole derivatives (PCT WO 94/14808) are usually as tyrosine kinase inhibitor.The pyridinyl compounds (U.S. patent No.5,302,606) that styryl replaces, some quinazoline derivant (EP application No.0 566 266 A1) have been described compound of styryl (U.S. patent No.5,217,999); Expert Opin.Ther.Pat. (1998), 8 (4); 475-478, seleno indole (seleoindoles) and selenides (PCTWO 94/03427), three ring polyols (PCT WO 92/21660) and benzylphosphonic acid chemical compound (PCT WO 91/15495) are as the compounds for treating cancer of tyrosine kinase inhibitor.Described anilino-cinnolines (PCT WO 97/34876) and quinazoline derivant chemical compound (PCT WO 97/22596; PCT WO 97/42187) as the inhibitor of vascularization and vascular permeability.

In addition, attempted identifying micromolecule as the serine/threonine kinase inhibitor.Particularly described and two (Indolylmaleimide) chemical compounds can have been suppressed specific PKC serine/threonine kinase isoform, the dysfunction of this enzyme and relevant (the PCT WO 97/40830 of the infiltrative change of VEGF-relevant disease medium vessels; PCT WO 97/40831).

Therefore, need to identify by modulating effective macromole and the little organic molecule that the active specificity of receptor or nonreceptor tyrosine kinase suppresses the tyrosine signal transduction, in order to regulate and modulation abnormality or unsuitable cell function, cell proliferation or differentiation.Especially, identify that the method for single-minded inhibition tyrosine kinase function and chemical compound are useful, this function causes edema for formation, seepage, ooze out and macromole to blend deposition and the vascular permeability of relevant disease too high outward be necessary.

The present invention's general introduction

It is too high to the present invention relates to suppress vascular permeability by the cellular signal transduction function that suppresses the KDR tyrosine kinase.Also the not obvious Flt-1/VEGFR1 of influence or other tyrosine kinase activity provide a kind of too high method of vascular permeability that suppresses by the catalytic kinase reaction of selective destruction KDR/VEGF-2 in the present invention.Compare with present comprising such as the Therapeutic Method of the materials such as steroid that tend to produce multiple unwanted side effect, the medicament that works according to the present invention has tangible pharmacology's advantage.These methods of the present invention also are preferable over uses low narrow spectrum inhibitors of kinases, comprises that those suppress multiple vegf receptor, because these methods can directly not upset other kinase whose important normal physiological function.As suppressing the too high result of blood vessel endothelium permeability, also suppressed edema subsequently by the tyrosine kinase activity that suppresses KDR, relevant hemocyte oozes out, and passes the change of the molecule exchange of endothelium, exosmoses seepage, the formation of oozing out.Because the incident of back usually causes excessive apposition, unusual substrate propagation and organ dysfunction suppress the KDR tyrosine kinase and also can be used for treating the multiple non-Cancerous disease that these etiology features are arranged.In addition, by suppressing the activity of KDR tyrosine kinase, also will be attached to the blood vessel hypotension that the relevant activation of the VEGF-part of kdr tyrosine kinase receptor causes and minimize.

By the chemical compound medication with single-minded inhibition KDR tyrosine kinase activity, the present invention also provides and has suppressed the Therapeutic Method that individual medium vessels permeability is too high and edema forms.Detailed description of the present invention

The cellular signal transduction function that the invention discloses by suppressing the KDR tyrosine kinase suppresses the too high a kind of method of vascular permeability.The medicament that the invention also discloses by using selectivity to suppress the cellular signal transduction function of KDR suppresses the too high a kind of method of vascular permeability.Be tested and appraised and use the high selectivity KDR inhibitor of the too high foundation method of the present invention of effective obstruction KDR cellular signal transduction and vascular permeability thereupon, set up KDR and mediated necessity effect in the vascular permeability reaction of VEGF.Proved the effect of this high selectivity KDR inhibitor, and need not suppress high affinity receptor VEGFR-1/Flt-1 function at the modulation vascular permeability.This specific character is than using the low existing Therapeutic Method that optionally destroys the medicament of other non--kinase whose function of KDR that better treatment toleration should be provided.

When vascular endothelial cell growth factor (VEGF) or other activation part (such as HIV Tat protein, VEGF-C or VEGF-D) is attached to the kdr tyrosine kinase receptor that is distributed in the vascular endothelial cell surface, just activated the KDR tyrosine kinase.Though do not find kinases-activated mutant body and truncate of naturally occurring KDR, existing report in EGFR and Tie-2 receptor kinase.Thereby also can expect the situation of constitutively activate of KDR.The KDR tyrosine kinase may also be referred to as the FLK-1 tyrosine kinase, NYK tyrosine kinase or VEGFR-2 tyrosine kinase.

Except stimulating vascularization, and endothelial cell migration and propagation, the VEGF also permeability of induction of vascular is too high.The result is that liquid passes blood vessel wall and enters a matter space from blood flow, thereby forms the edema zone.Hemocyte oozes out also usually follows this reaction.Similarly, excessive vascular permeability is too high can to destroy the normal molecule exchange that (for example lung and kidney) passes endothelium in the important tissue and organ, thereby cause organ dysfunction, macromole oozes out, and substrate propagation apposition together normal and that promote.When betiding the lacuna of restriction, edema (for example cerebral edema) may cause the organ dysfunction and the damage that damage.

Can use several different methods to suppress the cellular signal transduction function of KDR: the generation of blocking-up activation part, blocking-up activation part is attached to kdr tyrosine kinase receptor, prevent receptor dimerization and transphosphorylation, suppress the enzymatic activity (the phosphorylation ability of inhibitory enzyme) of KDR tyrosine kinase or conduct (D.Mukhopedhyay etc., Cancer Res.58:1278-1284 (1998) and list of references thereof) by other machine-processed signal that destroys its downstream.According to method disclosed herein, it is too high that these class methods of selective destruction KDR cellular signal transduction function will reduce vascular permeability, and relevant oozing out, and edema thereupon forms and the substrate precipitation.

Multiple chemical compound has required KDR tyrosine kinase rejection characteristic.Comprise in conjunction with the enzyme part of outer KDR receptor domain of born of the same parents or cell kinase in the chemical compound or alternately, in conjunction with the antibody (after this looking like is to comprise the single-chain antibody construct) of VEGF itself.These antibody interfere VEGF to be attached to kdr tyrosine kinase receptor and/or importantly, interfere KDR tyrosine kinase cellular signal transduction ground function.Being attached to KDR tyrosine kinase ground antibody may be as the antagonist of VEGF or more generally, as the antagonist of VEGF activator.Perhaps, these antibody may block function receptor dimerizations or they are KDR tyrosine kinase inhibitors.With VEGF or activation part bonded antibody be the neutralizing antibody of VEGF or activation part.Must know that this class VEGF neutralizing antibody may react by KDR and F1t-1 receptor blocking VEGF, and typically to single activation ligand specific.As a rule, not essential or expectation by Flt-1 receptor blocking VEGF reaction.Owing to reported the different epi-positions of VEGFR identification VEGF, expectation can be to the activated single-minded blocking-up of KDR by using single-minded combination also " to shelter " VEGF or other KDR-that activates part finishes in conjunction with the antibody of epi-position.

Can suppress the KDR tyrosine kinase activity, thus vascular permeability is too high and form minimized other chemical compound of edema and comprise peptide and organic molecule.Peptide comprises the solubility ectodomain of KDR and KDR binding fragment.Other useful peptide is the mutant (VEGF-C for example of the relevant somatomedin of VEGF or VEGF, VEGF-D or HIV Tat protein and their fusion rotein), their in conjunction with and the blocking-up further therewith the part of receptor in conjunction with but do not stimulate Dimerized, the activation or KDR tyrosine kinase transphosphorylation.This class mutant can be monomer or non-functional heterodimer, thereby blocks the combination of natural dimer VEGF or activation part.Similarly, other blocking-up receptor dimerization and/or activated peptide or micromolecule of use that can be successful.These chemical compounds are also as the antagonist of activation part or the inhibitor of KDR tyrosine kinase activity.Preferred chemical compound is little organic molecule.

In addition, suppress the biosynthesis of active functional KDR tyrosine kinase or correctly present such as KDR-specificity ribozyme, single-chain antibody (S in antisense polynucleotide (such as antisense mRNA) or the cell _cF _v) equimolecular can effectively block the reaction to VEGF of KDR-mediation.Molecule can be introduced ready-formed cell or can in cell, induce their generation (for example by using suitable adenovirus, retrovirus retrovirus or baculovirus vector).

Preferred compound of the present invention has the characteristic that suppresses KDR cellular signal transduction function but the cellular signal transduction function (the Flt-1 tyrosine kinase is also referred to as the VEGFR-1 tyrosine kinase) that can significantly not suppress Flt-1.KDR tyrosine kinase and Flt-1 tyrosine kinase all are to be attached to kdr tyrosine kinase receptor respectively and the flt-1 tyrosine kinase receptor is activated by VEGF.Because the Flt-1 tyrosine kinase activity may mediate endothelium and keep critical event with vascular function, suppresses the active or relevant transduction signal of this kind of enzyme and may cause toxicity or disadvantageous side reaction.At least, this inhibition for the blocking-up vascular permeability too high induce and to form edema optional, be to waste and unworthy therefore for individual speech.It is unique to preferred compounds of the invention are, and does not suppress other receptor tyrosine kinase because their suppress by the active of the activation activated VEGF-receptor tyrosine kinase of part (KDR), such as also being by the activated Flt-1 of certain activation part.Therefore most preferred of the present invention is selectively on its tyrosine-kinase enzyme inhibition activity.

Known VEGF impels the too high and edema formation of vascular permeability.VEGF is by at the inflammatory T-of inflamed sites cell, and macrophage is bitten neutrophilic granulocyte and bitten that eosinophile etc. expresses.The generation of this factor is very fast by histanoxia, some blood vessel pressurization hormone, and somatomedin, reproductive hormone and multiple inflammatory cytokine raise.It seems that the too high and edema of in fact, relevant with the expression of many other somatomedin or administration vascular permeability be generation mediation by VEGF.Vascular permeability is too high, relevant edema, and the exchange of wearing endothelium and the macromole of change ooze out, and it usually follows hemocyte to ooze out, and can cause excessive apposition, unusual substrate propagation, fibrosis etc.Therefore, too high can the impelling significantly of permeability of VEGF-mediation has the disease with these etiology features.For example: (1) VEGF has significantly at the epidermis of damaging psoriatic skin and increases.This factor is powerful to stimulate dermal endothelial cell propagation and the blood capillary permeability relevant with psoriasis too high.(2) after burn and rats after severe scald burn, major organs has been damaged by regular meeting.This can be by by ischemia-weight perfusion injury, the expansion of viscera tissue and edema, and promptly endothelial cell damage uncontrollable " medium disease (the mediator disease) " that cause shows.For the burn victim, sucking injury is a main cause of death.The cast of casting off a skin and making up the trachea that the exudate of rich in proteins forms of tracheal bronchus epithelium may cause blocking these tracheas.Combination sucks burn and histanoxia and is exposed to high-concentration oxygen (attempting helping individual) thereupon and makes by the accumulated change that causes lung and be worse off, and forms hemorrhage edema variation to trachea and blood vessel wall such as the diffusion exudate.Victim's circulation Serum VEGF significantly increases (height is to 20 times) and may make the main media (Grad etc., Clin.Chem.Lab.Med.36:379-383,1998) of these complication after burn and multiple wound.(3) sunburn is also relevant with the formation of edema.Known after being exposed to ultraviolet the generation of VEGF also can raise.Other dermatosis that produces edema comprises blistering symptomaticerythedema (erythroderma), persistent acrodema and bulla disease (bullousdiseases) be such as erythema multiforme, pemphigus sample bleb (bullous pemphigoid) and dermatitis herpetiformis (dermatitis herpetiformis) (being acute or chronic inflammatory disease).The edema speckle is the further disease that shows edema with roseacea such as the disease relevant with telangiectasis.(4) enhanced blood capillary permeability and edema are the common traits of inflammation and tumor disease.Cerebroma such as nerve erosion matter tumor, form tumor or tumor around cerebral edema and topping up cyst, and meningiomas follows a large amount of cerebral edemas, is the example of this class disease.Partial high-caliber VEGF and these disease associations.Inducing of malignant ascite liquid and tumor exudate (particularly malignant pleural and pericardial effusion) is the further example that this class produces the disease of edema, and the known VEGF that relates to produces.In addition, the edema of head trauma generation can produce the sick infringement of concussion brain function.Similarly, confirmed that communicating hydrocephalus relates to cytokines such as the IGF-1 that produces such as known modulation VEGF and TGF-B1.(5) such as forming nasal polyp, in some chronic inflammatory diseases of cervical polyp stomach function regulating hypertrophy polyp edema can take place.In these cases, confirm that inflammatory cell plays an important role in the edema state takes place, to small part be by producing VEGF.(6) but the activated important source of biting eosinophile VEGF of cytokine, thereby promoted in allergic inflammation position formative tissue edema.Edema and exudate are the common complications that produces in irritated and delayed hypersensitivity; Sometimes also comprise anaphylaxis.VEGF is relevant with the reaction of non-confrontational histamine agents or aspirin response especially, and observes its rise in poison lacquer and contact dermatitis.In addition, pulmonary tuberculosis, some viral infection, angioedema, urticaria (hives) and exercise induced anaphylaxis also are possible relate to this class allergy of VEGF and the example of delayed hypersensitivity.As medicaments insensitive or anaphylaxis or use the result that VEGF-raises somatomedin or cytokine (for example IGF-1, FGF-2 or IL-2), usually also can form edema.Radioanaphylaxis is also too high relevant with vascular permeability with radiodermatitis.(7) VEGF relates to the eye neovascularization, causes diabetic retinopathy and microangiopathy, the age-that relevant degeneration of macula causes is blind and to be exposed to the neonate that too high oxygen produces blind.Under many circumstances, speckle or other ocular edema are all arranged before these diseases.Confirmed that VEGF is an iris in eye ischemia and the angioedema, the main amboceptor of cornea and retina neovascularization.In the multiple ocular disease that exudate and apposition conduct vascularization basis is subsequently arranged, the destruction of the too high promotion blood-retinal barrier of the inductive vascular permeability of VEGF.The cornea neovascularization is chemical burn, the main consequence after cornea inflammation and the edema.Nearest evidence shows that VEGF is relevant with the post-traumatic process of this class eye.Used the chelating agen deferoxamine clinical anticancer patient of ferrum.But the speckle edema is often brought out in this treatment.The concentration of this iron chelating agent that obtains in patient is induced VEGF mRNA, and the expression in the normal and tumor cell line has 3-5 increase doubly in all of research, and hint VEGF is likely the amboceptor of edema formation.The intraocular pressure of the increase that excessive generation of VEGF and edema cause can cause unsuitable apposition, visual distortion, and optic disk changes, visual field defective and may cause glaucoma.Vascular permeability is too high also usually relevant with conjunctivitis.(8) neonate and adult's chronic lung disease comes from injury of lung and inadequate repair process.Being reported in has VEGF to produce in the animal model of various pulmonary.The destruction of lung endotheliocyte also is the feature of the too high property of oxygen injury of lung.From the too high recovery of oxygen the time, II type vesicle cell raises VEGF and causes lung endothelial cell proliferation and regeneration subsequently.Yet these the possibility of result cause the destructive exchange of passing lung endotheliocyte and pulmonary edema.Asthma and bronchitis usually relate to bronchus vasodilation, the congestion of blood vessel, and bronchial wall edema and oozing out causes the air flue mucosa to thicken with the bronchus inner chamber and narrows down.Having albumen to ooze out with the edema of unusual substrate growth typical and these phenomenons entwines mutually.By correlated process, in adult respiratory distress syndrome, form pulmonary edema.The reason of adult respiratory distress syndrome comprises pneumonia, the suction of noxious substance, contusion of lung, approximate drowning and suction gastric content.(9) corticosteroid, such as cortisone, hydrocortisone belongs to the therapeutic agent of the treatment edema of normal use.They are strong inhibition agent of vegf expression.Believe that now this specific character has significantly promoted the known edema effect of these steroid.But their polyenergic biologic activity also can cause the side reaction of not expecting.Steroid hormone and their agonist and antagonist also can appreciable impact VEGF generation, particularly in germinal tissue.Endometritis and endometriosis can occur in pregnancy, in the middle of menstrual cycle or the gonadotherapy.The swelling of menstruation is too high relevant with angor and vascular permeability.Tamoxifen, a kind of medicine that reduces mammary cancer risk also increases uterine cell propagation and tumor incidence rate.This steroid analog, and estradiol cause uterus edema and cell proliferation, and they show the increase that relates to local VEGF generation.Ovary super thorn bowel syndrome (ovarian hyperstimulation syndrome) is the induced severe complication of influence ovulation.The most serious syndromic performance of this class is to form the ovary of bulk to increase and multiple cyst, ascites, and hemoconcentration and liquid are in the 3rd-spatial accumulation.Stimulate the back because the capillary permeability of the increase that excretory VEGF such as the luteinization granulosa cell of ovary causes is believed in these syndromes plays a crucial role with human chorionic gonadotropin.Verified in Si Tanli-Leventhal syndrome, VEGF is overexpression in the Hyperthecosis gamogenetic egg nest substrate of polycystic ovary (polycystic ovaries).(10) in the animal model of apoplexy, confirm after instantaneous mesencephalic arteries blocks, the quick induction of VEGF gene expression is arranged in neuron and mantle cell.VEGF has the brain cell of helping to be injured from ischemia, such as apoplexy, and the recovery in head trauma or the cerebral infarction, but cerebral lesion also can worsen owing to following of cerebral edema forms.Malaria also may be brought out edema as the ischemic result of the inductive brain of VEGF-.Because the tumor blood capillary lacks normal blood brain barrier function, can produce the cerebral tumor relevant cerebral edema and water-filling cyst.The VEGF most probable that neural erosion matter oncocyte original position discharges causes into the diagnostic histopathology's feature and the Clinical symptoms of erosion cell plastid struma tumor in patient, comprise the cerebral edema of increase.Carpal tunnel syndrome is followed the neural aquation of increase and is usually followed the extracellular matrix deposition (entrapment neuropathy) of increase.The VEGF level that increases in the perineural tissue is by the induction of vascular permeability, and liquid flows out and apposition causes the neural folder that falls into to organizing in neural week.(11) as the reaction to hypoxia, the generation of VEGF is raised significantly in the tissue.Thereby, observe in necrosis, ischemia, infarction, obturation, anemia, in blood circulation damage or other oxygen forfeiture zone, the VEGF level increases and usually has vascular permeability too high, edema and oozing out.Cause the low oxygen pressure of " altitude sickness " also can cause VEGF generation fast, it may be the reason that life-threatening brain and pulmonary edema (HACE and HAPE) take place unconformable people.(12) overexpression of VEGF equally also with as radiation injury, rheumatism, lupus, myocardial infarction, the too high pericardium that causes of wound or drug reaction result's vascular permeability oozes out relevant with pleura.In lung or breast carcinoma, usually observe in the obduction of lymphoma and leukemia patient VEGF overexpression and pericardium and pleura ooze out just strange.The amount of VEGF also has remarkable rising in the synovial fluid of the swollen joint of rheumatic arthritis individuality.Sprain and fracture, though with to promoting vascularization and healing useful some swelling and vascular permeability too high relevant, may follow pain and the excessive edema of not expecting.Similarly, expection VEGF with have the patient's condition (for example " water being arranged " in the knee) such as the synovitis of oozing out and meniscus injury relevant.(13) ulcer relevant with loop limit (for example decubital ulcer, gravity and varicose ulcer) is also usually followed edema and protein exudate.Diabetic complication usually produces the usually circulation of concomitant injury as the result of circulating-glucose levels (hyperglycemia) that improves and formation advanced glycosylation end product (AGE).These patient's condition, alone or in combination together, generation and the vascular permeability that can cause multiple diabetic complication thereupon of known stimulation VEGF are too high.(14) because the significant composing type of kidney podocyte produces the too high effect of vascular permeability of the VEGF level of endogenous VEGF and known raising, nephropathy is such as microalbuminuria, albuminuria, the oliguria electrolyte imbalance complication of diabetes (usually as) and nephrotic syndrome (burn particularly, after shock or the wound hypoxia inducible) may be treated according to the present invention.(15) protein exudate and blood cell ooze out, and follow edema usually and cause excessive apposition and substrate propagation, cause other advancing of disease.These diseases comprise HSV, hepatitis interstitialis chronica, fibrosis (fibroses), pimple cicatrix and form undesirable scar tissue.The too high meeting of permeability that suppresses the VEGF-mediation stops this class advancing of disease.(16) known have the VEGF isoform of significant quantity to be stored in platelet, in the middle of mastocyte etc. and the extracellular matrix.Under specific circumstances, thus these VEGF/VPF repertorys can discharge and cause the acute vascular permeability too high rapidly.

From these different examples, edema occurs under the multiple physiological condition and the intensive edema that involves of VEGF/VPF or related analogs forms and exosmoses significantly.Chemical compound of the present invention will with macular edema, aphakia/false aphakia capsule sample macular edema, retinoblastoma, ocular ischemia, ocular inflammatory disease or infection, choroidal melanoma, iron chelating agent is treated inductive edema side effect, pulmonary edema, leural effusion, PE, myocardial infarction, atrophic diseases, the tissue edema at wound or allergic inflammation position, the polyp edema at chronic inflammatory disease position, cerebral edema, cerebral tumor topping up cyst (brain tumor fluid-filled cysts), communicating hydrocephalus, the relevant edema state minimization of edema that edema of being correlated with the organ injury that burn causes and suction burn injury cause or the like.Chemical compound of the present invention also will with skin burn, vesicle, erythema multiforme, edema speckle and other dermatosis, the cerebral tumor, ascites relevant and multiple seepage with cancer, carpal tunnel syndrome, altitude sickness, irritated and anaphylaxis and allergy reaction, radioanaphylaxis, radiodermatitis, glaucoma, conjunctivitis, adult respiratory distress syndrome, asthma, bronchitis, the high thorn of ovary bowel syndrome (ovarianhyperstimulation syndrome), polycystic ovary syndrome (polycystic ovarysyndrome), menstruation swelling and angor, apoplexy, head trauma, cerebral infarction or obturation, ulcer is sprained, fracture, the seepage relevant with synovitis, diabetic complication, the edema state minimization that hepatitis interstitialis chronica and administration somatomedin or the like are relevant.Chemical compound of the present invention also can be used for the treatment of microalbuminuria disease, albuminuria, uropenia, electrolyte disturbance, nephrotic syndrome, HSV, exudate (exudates), fibroid (fibrose), the formation of keloid and undesirable scar tissue.

Chemical compound of the present invention can with one or more other medicines combination medicine-feedings, these medicines can suppress or prevent the generation of VEGF, reduction is to the cell internal reaction of VEGF, it is too high to suppress vascular permeability, reduces inflammation or inhibition or prevents to form edema.Process only to be administered is suitable, can with chemical compound of the present invention before or after other medicines or with other medicines administration simultaneously.Other medicine includes but not limited to resist-the edema steroid, NSAIDS, and the ras inhibitor, the anti-TNF medicine, anti--the IL1 medicine, hydryllin PAF-antagonist, COX-1 inhibitor, cox 2 inhibitor, NO synthase inhibitor, pkc inhibitor and PI ₃Inhibitors of kinases.The medicine of chemical compound of the present invention and other can adduction or is worked synergistically.Thereby it is too high and/or suppress the combination medicine-feeding of the medicine that edema forms that these are suppressed vascular permeabilities, than separately with the individually dosed ill-effect that more can alleviate the too high or edema of vascular permeability of any medicine.

Cause because the formation of edema is oozed out by the liquid of blood flow usually, taking place often in seepage, regular meeting produces hypotension.Hypotension also can be used as VEGF or VEGF activator and is attached to the result of the vegf receptor on the vascular endothelial cell and occurs.Seem that chemical compound of the present invention minimizes hypotensive development by the cellular signal transduction function of KDR that suppresses to be attached to as VEGF (or other activation part) result of this receptor.When chemical compound of the present invention being suppressed individual hypotension during to individual administration.Pharmaceutical preparation

Can with treatment or to improve vascular permeability too high, the dosage of edema and relevant disease with chemical compound of the present invention itself to patient's administration or with itself and appropriate carriers or the administration of mixed with excipients formation pharmaceutical composition.Can also be with the mixture of these chemical compounds as simple mixtures or appropriate formulations type pharmaceutical composition to patient's administration.The treatment effective dose is meant that further the amount of chemical compound is enough to prevent edema, the too high and/or relevant hypotensive development of VEGF-of the relevant permeability of VEGF-.Can be with the technology of compound formulationization of the present invention and administration at " Lei Mingdun pharmaceutical science (Remington ' s Pharmaceutical Sciences) " Mack Publishing Co., Easton, PA finds in the latest edition.Route of administration

Suitable route of administration can be, for example comprise oral, eye drop, rectum, saturating mucosa, the surface, or enteral administration; Parenteral, comprise intramuscular, subcutaneous, the injection in the marrow, and in the sheath, directly intraventricular, intravenous, endoperitoneal, intranasal, or the injection of ophthalmic.

In addition, can for example,, usually be to store or slow releasing preparation with chemical compound part rather than systemic administration by with edema position, chemical compound direct injection road.

In addition, can use this medicine, for example, use the liposome of endotheliocyte-specificity antibody sandwich by the targeted drug delivery system.Compositions/preparation

Pharmaceutical composition of the present invention can be made by known methods, and for example, by the mixing of routine, granule is made in dissolving, makes dragee, grind, and emulsifying, encapsulation wraps or the lyophilization program.

Like this, by conventional methods, use one or more physiology can accept carrier, comprise that promotion makes reactive compound the excipient and the auxiliary agent of the preparation of useful as drug, can be used for pharmaceutical composition of the present invention and form preparation.Appropriate formulation depends on the route of administration of selection.

For injection, medicament of the present invention can be made aqueous solution, preferred in the buffer of physiology compatibility such as Hanks solution, Ringer solution, or normal saline buffer solution.For the administration of passing mucosa, will be used for preparation for the suitable penetrating agent of the barrier that will permeate.This penetrating agent in the present technique field usually as can be known.

For oral administration, can be with active compound and the easy preparation that forms of pharmacological-acceptable carrier combination well known in the art.This class carrier makes compositions of the present invention can form tablet, pill, and dragee, capsule, liquid, gel, syrup, slurry, suspension or the like is used for the oral digestion of patient that will treat.By active compound and solid excipient combination are obtained to be used for oral pharmaceutical preparation, optionally grind the mixture that produces, and adding proper assistant post processing granulate mixture if desired, obtain tablet or dragee core.Appropriate excipients, in particular, filler comprises lactose such as sugar, sucrose, mannitol, or sorbitol; Cellulose preparation such as, corn starch for example, wheaten starch, rice starch, potato starch, gelatin, tragacanth (gum tragacanth), methylcellulose, hydroxypropyl emthylcellulose, hydroxy-methyl cellulose sodium, and/or polyvinylpyrrolidone (PVP).If desired, can add distintegrant, such as crospolyvinylpyrrolidone, agar, or alginic acid or its salt are such as sodium alginate.

Suitable bag quilt is provided for the dragee core.For this purpose, can use spissated sugar juice, it optionally comprises Radix Acaciae senegalis (gum arabic), Talcum, polyvinylpyrrolidone, carbopol gel, Polyethylene Glycol, and/or titanium dioxide, lacquer solution (lacquersolutions), and appropriate organic solvent or solvent mixture.Stain or pigment can be added tablet or dragee bag quilt, be used to differentiate or characterize the various combination of active compound doses.

Can comprise the sucking fit capsule made from gelatin as oral pharmaceutical preparation, and with gelatin and plasticizer such as glycerol or sorbitol make soft, the capsule of sealing.The sucking fit capsule may comprise with filler such as lactose, binding agent is such as starch, and/or lubricant such as Talcum or magnesium stearate and optionally and the blended active component of stabilizing agent.In soft capsule, reactive compound can be dissolved in or be suspended in suitable liquid, such as fatty oil, and liquid paraffin, or liquid macrogol.In addition, can add stabilizing agent.The oral preparation that is useful on should be the dosage that is fit to oral administration.

For the buccal administration, compositions may be taked the tablet or the lozenge of traditional approach preparation.

For inhalation, the pressure that passes through of the chemical compound routine that the present invention uses is packed with aerosol spray transmission or aerosol apparatus transmission, uses suitable propellant, dichlorodifluoromethane for example, Arcton 11, Dichlorotetrafluoromethane, carbon dioxide or other suitable gas.For the compression aerosol, may provide valve to discharge measured quantity and determine dosage unit.Capsule that be used to suck or for example gelatin of insufflator etc. makes or tube can be by containing chemical compound mixture of powders and suitable powder substrate such as lactose or starch make.

Chemical compound can be made parenteral formulations, by for example (bolus) injection or drug administration by injection fast continuously of injection.The preparation that is used to inject may be unit dosage form for example at ampoule or multi-dose container, add antiseptic.Compositions can be taked the suspension in oil or water excipient, solution or form of emulsion, and may contain the preparation chemical preparation such as suspension, stable and/or dispersant.

The pharmaceutical preparation of parenteral comprises the aqueous solution of reactive compound water-soluble form.In addition, the suspension of reactive compound can be prepared into suitable oily injection suspensions.Suitable lipophilic solvent or excipient comprise fatty oil such as Oleum sesami, or Acrawax, such as ethyl oleate salt or triglyceride, or liposome.The water injection suspension may comprise the material that strengthens suspension viscosity, such as sodium carboxy methyl cellulose, and sorbitol, or glucosan.Optionally, suspension also may contain suitable stabilizers or increase the solution of the medicament of compound dissolution with the preparation high concentration.

Perhaps, active component can be a powder type, makes up with for example aseptic apirogen water of appropriate excipients before use.

This chemical compound also can be made the compositions that rectum is used, and such as suppository or stagnation enema, for example, contains conventional suppository base such as cocoa butter or other glyceride.

Except previously described preparation, this chemical compound can also be made the storage preparation.This durative action preparation can be by transplanting administration (for example subcutaneous or intramuscular or by intramuscular injection).Like this, for example, can be with this chemical compound and polymer or hydrophobic substance (for example as at the emulsion that can accept in the oil) or ion exchange resin, or as soluble derivant slightly, for example soluble-salt forms preparation slightly.

The example that is used for the pharmaceutical carrier of hydrophobic compound of the present invention is the cosolvent system, comprises benzylalcohol, a kind of non-polar surfactant, a kind of easy and blended organic polymer of water, and a kind of water.The cosolvent system may be a VPD cosolvent system.VPD is a 3%w/v benzylalcohol, and 8%w/v non-polar surfactant Tween 80 and 65%w/v Liquid Macrogol are made volume required in anhydrous alcohol.VPD cosolvent system (VPD:5W) contains the VPD in the dilution in 1: 1 of 5% D/W.This cosolvent system is the solubilizing hydrophobic chemical compound well, and its is very low from toxicity when being administered systemically.Normally, the changes of contents of cosolvent system can be destroyed its dissolubility and toxic characteristic quite greatly and not.In addition, the concordance of co-solvent component can change: for example, can use other hypotoxicity non-polar surfactant to replace Spheron MD 30/70; The clip size of Polyethylene Glycol can change; Can use other bio-compatible polymer to replace Polyethylene Glycol, for example polyvinylpyrrolidone; And can use other sugar or polysaccharide to replace glucose.

Perhaps, can use other drug-supplying system with the hydrophobic pharmaceutical compounds administration.Liposome and Emulsion are known excipient or carriers with the hydrophobic drug administration.Though toxicity is bigger usually, also can use some organic solvent such as dimethyl sulfoxide.In addition, can use slow-released system, such as the solid hydrophobic polymer semipermeable membrane substrate that contains healing potion with this compound administration.Determined multiple slow-release material, they are known for present technique field quantity technical staff.Based on its chemical property, slow releasing capsule can extremely the time above 100 days discharges this chemical compound in several weeks.According to the chemical property and the biological stability of healing potion, can use other protein stabilized strategy.

Pharmaceutical composition also may comprise suitable solid or gel phase carrier or excipient.This class carrier or excipient include but not limited to calcium carbonate, calcium phosphate, and different sugar, starch, cellulose derivative, gelatin, and polymer is such as Polyethylene Glycol.

Many organic molecular compounds of the present invention can provide with the salt mensuration form of the counter ion compatible with materia medica.Can with acid, include but not limited to hydrochloric acid, sulphuric acid, acetic acid, lactic acid, tartaric acid, succinic acid etc. form the compatible salt of medicine.Salt is better than corresponding free alkali form dissolubility in water or other proton solvent.Effective dose

The pharmaceutical composition that is suitable among the present invention comprises and contains the effective dose composition of active components that can accomplish the end in view.More specifically, the treatment effective dose meaning is the amount that can effectively prevent or alleviate existing symptom to the object that will treat.The one skilled in the art has the ability to determine effective dose.

This chemical compound of effective dose can fully suppress the signal conduction function of KDR, can not cause tangible side reaction owing to suppressing Flt-1 or other tyrosine kinase function thereby the inhibition vascular permeability is too high.Can measure the dosage of KDR tyrosine kinase-dependency by vitro detection and suppress to determine to have the active chemical compound of this class.Under similar [ATP]/Km (ATP) and substrate condition, preferred chemical compound is to the IC of KDR ₅₀The IC of comparison Flt-1 or other PTK ' s ₅₀Low significantly (ideally, the KDR tyrosine kinase being had～100 times of selectivitys).

For any chemical compound that uses in the inventive method, begin and to estimate the treatment effective dose from cell detection.For example, can in cell and animal model, determine dosage, comprise the IC that determines in the cell detection to obtain the circulation composition scope ₅₀(promptly reach the compound concentrations of the test of maximum half that suppresses of pair cell signal conduction function KDR, this function normally reacts on VEGF or another activatory stimulation).The cell IC that in the presence of 3 to 5% serum albumin, measures ₅₀May be similar to plasma protein verify this chemical compound in conjunction with effect.Can utilize these information to measure effective dose accurately to the people.In addition, be used for the most preferred that is administered systemically and in intact cell, effectively suppress cellular signal transduction function KDR with the level of obtaining safely at blood plasma.

The treatment effective dose is meant that the amount of chemical compound improves patient's symptom.Can learn program is measured this compounds in cell culture or laboratory animal toxicity and therapeutic effect by standard drug, for example measure maximal allowance dose (MTD) and ED ₅₀(effective dose of 50% maximum reaction).The dosage rate of toxicity and therapeutic effect is a therapeutic index, can use MTD and ED ₅₀Ratio express.The chemical compound that shows high therapeutic index is preferred.Can be used to determine dosage range from the data that these cell culture detect and zooscopy obtains in the human body use.The dosage of these chemical compounds preferably is in the scope of circulation composition, and it comprises few or avirulent ED ₅₀According to the dosage form that uses and the route of administration of use, dosage can change in this scope.The doctor can select preparation accurately according to patient status, route of administration and dosage.(referring to for example Fingle etc., 1975, " The Pharmacological Basis of Therapeutics ", Ch.1pl).When the treatment emergency situation, may need a large amount of fast or infusion administration, to obtain effect fast near MTD.

Can single adjusting dosage and at interval, so that the level of active component in blood plasma that is enough to keep KDR modulation effect or minimum effective drug concentration (MEC) to be provided.For each chemical compound MEC is different, but can estimate according to external data; The KDR tyrosine kinase 50-90% that reaches that for example uses detection method described herein to measure suppresses required concentration.The required dosage that reaches MEC depends on personal feature and route of administration.Yet can use HPLC to detect or bioassay affirmation plasma concentration.

Use the MEC value can also determine dosing interval.Should use such therapeutic scheme administration,, preferably keep blood plasma level on MEC, up to the improvement that reaches expection to symptom at 30-90% and in preferred time at 50-90% promptly at 10-90%.When topical or selectivity absorption, effective local concentration of medicine may be irrelevant with plasma concentration.

The amount of the compositions of administration can depend on the object that will treat certainly, depends on the body weight of object, painful seriousness, the administering mode and the doctor's that prescribes judgement.Packing

If desired, compositions can be packing or dispenser device form, wherein may comprise one or more unit dosage forms that contain active component.Packing can be metal or plastic foil for example, such as blister pack.Packing or dispenser device can have the administration explanation.Can also prepare and contain, place proper container, the patient's condition for the treatment of on the labelling the compositions of The compounds of this invention with the preparation of the pharmaceutical carrier formation of compatibility.The suitable patient's condition of labelling may comprise the treatment edema, and it is too high and ooze out to suppress vascular permeability, and apposition minimizes the relevant hypotension of VEGF-, and the similar patient's condition.

Example I. external PTK detects

Can use following vitro detection to determine activity and the effect level of different chemical compound of the present invention to one or more PTK.Use technology well known in the art can design similar detection to other tyrosine kinase according to identical method.A. use rhabdovirus system to produce the KDR tyrosine kinase:

Use has produced the coded sequence of people KDR born of the same parents intracellular domain (aminoacid 789-1354) by PCR from the cDNA of HUVEC cell separation.Simultaneously introduced poly-His at this proteinic N-end ₆Sequence.With Xba 1 and the Not1 site of this section fragment cloning to transfection carrier pVL1393.Use BaculoGold transfection reagent (PharMingen) to produce recombinant baculovirus (BV) by cotransfection.Plaque purification reorganization BV and detect by Western confirms.In order to produce protein, with the SF-9 cell in the SF-900-II culture medium with 2 * 10 ⁶/ ml grows, and infects with every cell 0.5 plaque forming unit (MOI).Infect back 48 hours harvestings.B. purification KDR

In the cell precipitation of 1 liter of cell culture, add 50ml Triton X-100 lysis buffer (20mM Tris, pH8.0,137mM NaCl, 10% glycerol, 1%Triton X-100,1mM PMSF, 10 μ g/ml aprotinins, 1 μ g/ml leupeptin) cracking expression (His) ₆The SF-9 cell of KDR (aminoacid 789-1354).In Sorval SS-34 rotary head under 4 ℃ with cell pyrolysis liquid 19, centrifugal 30 minutes of 000rpm.With 5mlNiCl on the cell pyrolysis liquid ₂The chelating agarose column is used 50mM HEPES, pH7.5,0.3M NaCl balance.Use contains the same buffer eluting KDR of 0.25M imidazoles.Use the ELISA of SDS-PAGE and mensuration kinase activity to detect (following) analytical column fraction.The KDR of purification is exchanged to 25mMHEPES, pH7.5,25mM NaCl, 5mM DTT buffer and-80 ℃ of preservations.C. kinase whose generation of people Tie-2 and purification

Use produces the coded sequence of people's Tie-2 cell intracellular domain (aminoacid 775-1124) by PCR as template from the mazolytic cDNA of people.Introduced the poly-His6 sequence and this construct has been cloned into Xbal and the Notl site of transfection carrier pVL1939 at the N-end.Use BaculoGold transfection reagent (PharMingen) to produce recombinant baculovirus (BV) by cotransfection.Plaque purification reorganization BV and detect by Western confirms.In order to produce protein, with the SF-9 cell in the SF-900-II culture medium with 2 * 10 ⁶/ ml grows, and infects at 0.5 MOI.The kinase whose purification of the His labelling that uses in the screening is similar to the KDR of description.D. people Flt-1 tyrosine kinase produces and purification

Used rhabdovirus expression vector pVL1393 (Phar Mingen, Los Angeles, CA).Poly-His will encode ₆Nucleotide sequence place the 5 ' end in the nucleotide district of the complete people Flt-1 intracellular kinase domain (aminoacid 786-1338) of coding.Use has produced the nucleotide sequence of coding kinase domain by PCR from the cDNA storehouse of HUVEC cell separation.Histidine residues makes can be with this protein of mode affinity purification that is similar to KDR (B part) and ZAP70 (F part).With 0.5 infection multiplicity infection SF-9 insect cell and in back 48 hours results of infection.The source of E.Lck and EGFR tyrosine kinase

The Lck of Lck or truncated-type can buy (Upstate BiotechnologyInc. for example, Saranac Lake, NY or Santa Cruz Biotechnology, Inc., SantaCruz, CA) or use conventional method from known natural or recombinant sources purification.EGFR is available from Sigma (Cat#E-3641;～500 units/50 μ l), the EGF part obtains from OncogeneResearch Products/Calbiochem (Cat#PF011-100).The generation of F.ZAP70 tyrosine kinase

Used rhabdovirus expression vector pVL1393 (Phar Mingen, Los Angeles, CA).Poly-His will encode ₆Nucleotide sequence place the 5 ' end in the nucleotide district of the complete ZAP70 (amino acid/11-619) of coding.Use has produced the nucleotide sequence of coding ZAP70 coding region by PCR from the cDNA storehouse of Jurkat immortalization T-cell separation.Histidine residues makes can affinity purification (referring to the B part) protein.The LVPRGS bridge has constituted the recognition sequence of thrombin proteolytic cleavage, thereby can remove affinity labeling from enzyme.With 0.5 infection multiplicity infection SF-9 insect cell and in back 48 hours results of infection.G. purification ZAP70

Containing 20mM Tris, pH8.0,137mM NaCl, 10% glycerol, 1%TritonX-100,1mM PMSF, 1 μ g/ml leupeptin, cracking SF-9 cell in the buffer of 10 μ g/ml aprotinins and 1mM sodium orthovanadate.With chelating agarose Hi Trap post (Pharmacia) on the solubility lysate, use 50mM HEPES, pH7.5,0.3M NaCl balance.With 250mM imidazoles eluting fusion rotein.The enzyme that reclaims is kept under-80 ℃ contains 50mM HEPES, pH7.5 is in the buffer of 50mM NaCl and 5mM DTT.The enzyme-linked immunosorbent assay of H.PTK (ELISA)

The existence of having used enzyme-linked immunosorbent assay (ELISA) to detect and measure tyrosine kinase activity.According to for example Voller etc., 1980, the clinical immunology handbook of writing at Rose and Friedman, second edition, Am.Soc.of Microbiology, Washington, the known method of describing in " enzyme-linked immunosorbent assay " among the D.C.pp359-371 carries out ELISA.

Improve the active method of disclosed mensuration for specific PTK.The method for optimizing of the ELISA experiment of carrying out KDR for example, is provided below.This method reorganization is used for measuring other member of RTK family, and the activity of the chemical compound of nonreceptor tyrosine kinase, all in those skilled in the art's limit of power.In order to measure the inhibitor selectivity, with a kind of general PTK substrate (poly (Glu for example ₄Tyr) randomcopolymer, molecular weight 20,000-50,000) with ATP (typical 5 μ M), the concentration with the apparent Km of about twice in mensuration is used.The ELISA of external KDR

Used following procedure to measure the depression effect of chemical compound of the present invention: buffer and solution: PGT: Poly (Glu, Tyr) 4: store powder for 1-20 ℃ to the KDR tyrosine kinase activity.Powder is dissolved in phosphate buffer (PBS) becomes 50mg/ml solution.Save as the 1ml equal portions at-20 ℃.When system is dull and stereotyped, in Gibco PBS, be diluted to 250 μ g/ml.Reaction buffer: 100mM Hepes, 20mM MgCl ₂, 4mM MnCl ₂, 5mM DTT, 0.02%BSA, 200 μ MNaVO ₄, pH7.10ATP: be stored as the 100mM equal portions at-20 ℃.In water, be diluted to 20 μ M lavation buffer solutions: the PBS antibody dilution buffer that contains 0.1%Tween 20: 0.1% bovine serum albumin (BSA) tmb substrate in PBS: before use tmb substrate is mixed with 9: 1 with peroxide solutions or use K-Blue substrate from Neogen.Stop bath: 1M phosphoric acid flow process 1. preparation plates

In PBS, PGT stock solution (50mg/ml, freezing) is diluted to 250 μ g/ml.Improve the affine elisa plate of flat height (Corning#25805-96) at Corning and upward add 125 μ l in every hole.The PBS that in blank well, adds 125 μ l.Be incubated overnight with the band covering and at 37 ℃.Clean 1 time and in 37 ℃ of dry incubators dry 2 hours with 250 μ l lavation buffer solution liquid.Use preceding with the plate 4 ℃ of storages in sealing bag that cover.The tyrosine-kinase enzyme reaction :-in the 20%DMSO aqueous solution, prepare the inhibitor solution of 4 * concentration.-preparation feedback buffer-preparation enzymatic solution makes it reach required unit in 50 μ l, for example for KDR, makes 1ng/ μ l, is every hole 50ng in reaction.Be stored on ice.

-make 4 * ATP solution to 20 μ M from 100mM storage of water solution.

Be stored on ice-every hole adds 50 μ l enzymatic solution (according to kinase whose than living, 5-50ng enzyme/hole) typically-add, 25 μ l, 4 * inhibitor.-adding 25 μ l 4 * ATP is used for inhibitor and measures.-room temperature incubation 10 minutes.-every hole adds the ultimate density of 50 μ l 0.05N HCl cessation reactions-wash plate * * reaction: ATP:5 μ M5%DMSO antibodies-be diluted to 50ng/ml by PY20-HRP (Pierce) antibody (a kind of phosphotyrosine antibody) of two steps dilution (100 *, then 200 *) with the 1mg/ml equal portions in containing the PBS of 0.1%BSA.-every hole adds 100 μ l Ab.At room temperature incubation is 1 hour.4 ℃ of incubations 1 hour.-wash 4 * plate.Color reaction-preparation tmb substrate and every hole add 100 μ l.-detect OD up to reaching 0.6 at 650nm.-stop with 1M phosphoric acid.On the plate readout instrument, shake.-read OD rapidly at 450nm.

The incubation time of the optimization of enzyme preparation and enzyme reaction condition can change slightly and each batch determined according to experience.

To Fltl, Tie-2, EGFR and ZAP70 have used similar condition determination.For Lck, the reaction buffer that uses under similar condition determination is 100mM MOPSO, pH6.5,4mMMnCl ₂, 20mM MaCl ₂, 5mM DTT, 0.2%BSA, 200mM NaVO ₄PKC kinases source

Can buy the catalytic subunit (Calbiochem) of PKC.The PKC kinase assays

Carry out radioactivity kinase assays (Yasuda according to the flow process of publishing, I., Kirshimoto, A., Tanaka, S., Tominaga, M., Sakurai.A., Nishizuka, Y.Biochemistry and Biophysical ResearchCommunication 3:166,1220-1227 (1990)).In brief, all pH7.5 by 50mM Tris-HCl, 10mM MgCl of being reflected at ₂, 2mM DTT, 1mM EGTA, 100 μ MATP, 8 μ M peptides, 5%DMSO and ³³Carry out in the kinase buffer liquid that P ATP (8Ci/mM) constitutes.Chemical compound and enzyme are blended in the reaction vessel, by adding the mixture initial action of ATP and substrate.After adding 10 μ l stop buffers (the 5mM ATP in 75mM phosphoric acid) cessation reaction, with partially mixed object point to cellulose phosphate filter paper.In 75mM phosphoric acid, the sample of point is washed 5 to 15 minutes 3 times under the room temperature.The radio-labeled that quantitatively mixes by liquid scintillation numeration.Estrogen receptor is in conjunction with mensuration

The reaction condition of Cancer Res.45:4192 (1985) (with list of references and in herein) such as use Shein after 20 hours, has been measured the combination of radiolabeled 17 beta estradiols of 1nM to the human estrogen acceptor in the cytosol of MCF-7 mammalian cancer cells at 4 ℃ of incubations.Behind the incubation, the Linesless charcoal suspension of cytosol fraction with glucosan-Bao quilt mixed, 4 ℃ kept 10 minutes, centrifugal, collected supernatant then.Use liquid scintillation cocktail (Formula989 Packard) with flicker numeration instrument (LS 6000, Beckman) measure keep in the Linesless charcoal supernatant in conjunction with radioactivity.8 kinds of concentration double detection compound, estimate the inhibition activity simultaneously to obtain competition curve.The single-minded radioligand that is attached to estrogen receptor is defined as the total binding of mensuration in the presence of excessive unlabelled 17 beta estradiols (6 μ M) and the difference of non-specific binding.The result

Obtained to have the following inhibition concentration of the representative compounds of following structural formula:

Detect	The IC of chemical compound 50
		????KDR	????0.2μM
????Flt-1	????＞50.0μM
		????Lck	????＞50.0μM
????TIE2	????＞50.0μM
		????ZAP70	????＞50.0μM
????EGFR	????＞50.0μM
		????PKC	????≥20μM
Estrogen receptor	??＞10.0μM(＜10％inh@10μM)

These results prove, the present invention and the chemical compound that herein exemplifies have pair remarkable inhibiting activity of KDR tyrosine kinase and special optionally as the inhibitor of KDR tyrosine kinase.II. cell RTK detects

Cell detection below using is measured activity and the effect of different chemical compound of the present invention to KDR.For other tyrosine kinase, use suitable antibody reagent and, detect like the design class in the same way such as technology well known in the art such as immunoprecipitation and Western traces.The inductive KDR phosphorylation of VEGF-in Human umbilical vein endothelial cells (HUVEC) that the A.Western trace is measured.1. (San Diego CA) buys HUVEC cell (from mixing donor) and instruct according to handbook and to cultivate from Clonetics.Only use go down to posterity in early days (3-8) to detect.Use complete EBM culture medium (Clonetics) at 100mm plate (Falcon for tissueculture; Becton Dickinson; Plymouth, England) middle cultured cell.2. for the inhibition activity of assessing compound, with the cell tryptase protease digestion and with 0.5-1.0 * 10 ⁵Cells/well is inoculated into the 6-hole flat board (Costar that clusters; Cambridge, MA).3. inoculation is after 3-4 days, and dull and stereotyped 90-100% is paved with.From all holes, remove culture medium, clean cell and do not contain EBM minimal medium incubation 18-24 hour of additive (being serum starvation) with 5ml with 5-10ml PBS.4. the inhibitor with serial dilution is added to cell in 1ml EBM culture medium, and final concentration is 25 μ M, 5 μ M, or 1 μ M and 37 ℃ of incubations 1 hour.VEGF then recombinates the people ₍₁₆₅₎(R﹠amp; DSystems) be added to institute porose in, the final concentration in the EBM of 2ml culture medium is 50ng/m1 and 37 ℃ of incubations 10 minutes.Control cells evaluation background phosphorous acidify or the inductive phosphorylation of VEGF that use is untreated or is only handled with VEGF.5. the cold PBS that contains 1mM sodium orthovanadate (Sigma) with 5-10ml then cleans all holes, and containing protease inhibitor (PMSF 1mM, aprotinin 1 μ g/ml, pepsin inhibitor 1 μ g/ml, leupeptin 1 μ g/ml, vanadic acid sodium 1mM, sodium fluoride 1mM) and 200 μ l RIPA buffer (50mM Tris-HCl pH7 of 1 μ g/ml DNA enzyme, 150mM NaCl, 1%NP-40,0.25% NaTDC, 1mM EDTA) (all chemical drugss are from Sigma Chemical Company, St Louis, MO) in cracking and scrape cell.14,000rpm removed enucleation in centrifugal 30 minutes with lysate.6. by adding cold (20 ℃) ethanol (2 times of volumes) at least 1 hour or spent the night at the most, the protein of precipitation equivalent.Containing 5% beta-mercaptoethanol (BioRad; Hercules rebuilds precipitate and boiled 5 minutes in Laemli sample buffer CA).Use polyacrylamide gel electrophoresis (6%, 1.5mm Novex, San Deigo, CA) isolated protein and use the Novex system to transfer to nitrocellulose filter.With after bovine serum albumin (3%) sealing, at 4 ℃ with anti--KDR polyclonal antibody (C20, Santa Cruz Biotechnology; Santa Cruz, CA) or anti--phosphotyrosine monoclonal antibody (4G10, Upstate Biotechnology, Lake Placid, NY) detection of spending the night.Goat-anti-the rabbit cleaning and put together with HRP-or F (ab) 2 incubations of sheep-anti--Mus IgG are after 1 hour, and (Amersham Life Sciences, Arlington Height IL) show band to use emission chemiluminescence (ECL) system.The result

Have the inhibition concentration of a kind of representative compounds I of following array structure to be:

????HUVEC	The cell IC of chemical compound 50
		The KDR phosphorylation	????5-10μM

Also confirmed the KDR tyrosine-kinase enzyme selectivity (referring to ground I part) of this chemical compound.

These results prove that suitable chemical compound of the present invention has remarkable inhibiting activity to the tyrosine phosphorylation of KDR tyrosine kinase in the inductive endotheliocyte of VEGF-.III. uterus edema model

The ability of the acute increase of the mouse uterine weight that the several hrs that this test determination chemical compound suppresses to begin after estrogen stimulates takes place.The early onset thereof that known uterus weight increases is because the edema that the permeability that the uterus vascular system increases causes causes.(Endocrinology (1993), 133:829-837) expression of rising that has proved the VEGF mRNA in the uterus edema of estrogen-stimulation and uterus has close temporary transient relation for Cullinan-Bove and Koss.Use the neutralizing monoclonal antibody of VEGF to confirm these results, this monoclonal antibody can significantly reduce the acute increase (WO 97/42187) that estrogen stimulates the back uterus weight.Therefore, this system can be used as the body inner model of the too high and edema of the permeability that suppresses the VEGF-mediation.Material: all hormones with lyophilized powder available from Sigma (St.Louis, MO) or Cal Biochem (La Jolla illustrates CA) and according to supplier to prepare.Excipient component (DMSO, Cremaphor EL) available from Sigma (St.Louis, MO).Mice (Balb/c, age in 8-12 week) (Germantown NY) and according to the institute animal takes care of and uses committee's guide (institutional Animal Care andUse Committee Guidelines) to feed in no cause of disease animal culture device available from Taconic.Method: first day: the pregnant mare serum gonadotrop(h)in (PMSG) (PMSG) of (i.p.) injection 12.5 units in the Balb/c mouse peritoneum.The 3rd day: mice was by i.p. acceptor chorionic-gonadotropin hormone (hCG).The 4th day: with the mice randomization and be divided into 5-10 each group only.According to dissolubility and excipient with the dosage range of 1-200mg/kg by intraperitoneal (i.p.), intravenous (i.v.) or oral (p.o.) approach are with the experimental compound administration.The excipient matched group is only accepted excipient, and two groups are untreated.Typically, after 30 minutes, to experimental group, vehicle group and a untreated fish group i.p. inject 17 beta estradiols (500 μ g/kg).After 2-3 hour, suck kill animals by carbon dioxide.Behind the midline incision, by separating with the coupling part between the fallopian tube and take out each uterus in tight below of cutting cervix uteri and uterus.Careful fat and the connective tissue removed do not influence the integrity in the preceding uterus of weighing.With the average weight of processed group and comparing of untreated or vehicle-treated group.Determine significance by the Student check.Use not stimulated control group monitoring estradiol reaction.The result

For three kinds of administrations of 100mg/kg dosage, obtained to have the inhibition percent of the representative compounds of following structural to uterus, estradiol stimulation back edema.

Route of administration	% suppresses	The P value
			????p.o.	????17	????ns
????i.v.	????57	????0.01
			????i.p.	????52	????0.04

Prove herein Compound I aspect the vitro inhibition kinase activity be KDR-optionally, and can effectively block the cell autophosphorylation of the KDR that stimulates in response to VEGF-.

These result's proofs optionally suppress the suitable compound of the present invention of KDR function such as the formation that Compound I is blocked edema effectively.The result also confirms the i.v. of this chemical compound and i.p. administration effective especially.Importantly, the selective depressant with the different KDR function of multiple structure also obtains similar edema effect.Equivalent

Though with reference to preferred embodiment particularly disclosure and description the present invention, the one skilled in the art will be understood that the spirit and scope of the present invention of claims definition that need not deviate from appendix can carry out the multiple change of form and details.Only use conventional method, the one skilled in the art will recognize that maybe can judge the equivalent of specifically described particular of the present invention herein.This class equivalent will be within the scope of the present invention.

Claims (30)

1. suppress the individual interior too high a kind of method of vascular permeability, comprise the cellular signal transduction function that suppresses KDR.

2. the process of claim 1 wherein that described inhibition to KDR cellular signal transduction function is that selectivity is at KDR signal conduction function.

3. the process of claim 1 wherein that described KDR cellular signal transduction function is attached to the KDR acceptor portion by activatory part and stimulates.

4. the method for claim 3, the inhibition of wherein said KDR cellular signal transduction function is that selectivity is at KDR signal conduction function.

5. the method for claim 1, the inhibition of wherein said KDR cellular signal transduction function is the process that is selected from following group: the generation of blocking-up activation part, modulation activation part is to the combination of kdr tyrosine kinase receptor, destroy the dimerization of receptor, blocking-up KDR changes phosphorylation, the activity that suppresses the KDR tyrosine kinase is destroyed raising of the interior substrate of KDR born of the same parents, and blocking-up is by the initial downstream signal conduction of KDR tyrosine kinase phosphorylation activity.

6. the method for claim 5, the inhibition of wherein said KDR cellular signal transduction function is that selectivity is at KDR signal conduction function.

7. the process of claim 1 wherein that described inhibition is by carrying out chemical compound to described individual administration.

8. the method for claim 7, wherein said chemical compound suppresses the catalysis kinase activity of described KDR.

9. the method for claim 7, wherein said chemical compound is the activatory antagonist of a kind of KDR tyrosine kinase.

10. the method for claim 7, wherein said compound selective suppresses the phosphorylation of KDR tyrosine kinase substrate.

11. the method for claim 7, wherein said chemical compound is optionally to described KDR tyrosine kinase.

12. the method for claim 11, wherein said chemical compound is selected from peptide, and antibody and organic molecule, wherein said chemical compound are attached to described KDR tyrosine kinase.

13. the method for claim 12 wherein can suppress described compound administration to be selected from the formation of the morbid state in following group: macular edema, aphakia/pseudophakia capsule sample speckle edema, retinoblastoma, eye ischemia, eye inflammation or infection, the edema side reaction that choroidal melanoma, iron chelating therapy cause, pulmonary edema, myocardial infarction, atrophic diseases, anaphylaxis, the tissue edema at wound and allergic inflammation position, allergy, allergy, the polyp edema at chronic inflammatory disease position, cerebral edema, cerebral tumor topping up cyst, communicating hydrocephalus, carpal tunnel syndrome, the organ injury that burn causes, suck burn injury, skin burn, the vesicle relevant with sun burns, stimulate or infection erythema multiforme, edema speckle and other dermatosis, the cerebral tumor, tumor is oozed out, pulmonary carcinoma or breast carcinoma, ascites, pleura oozes out, and pericardium oozes out, plateau " disease ", radioanaphylaxis, radiodermatitis, glaucoma, conjunctivitis, choroidal melanoma, adult respiratory distress syndrome, asthma, bronchitis, the super thorn of ovary bowel syndrome, polycystic ovarian syndrome, menstruation swelling, menstrual cramps, apoplexy, head trauma, cerebral infarction or obturation, hypotension, ulcer, sprain, fracture, what synovitis was relevant oozes out, diabetic complication, HSV, hepatitis interstitialis chronica, microalbuminuria disease, albuminuria, uropenia, electrolyte disturbance, nephrotic syndrome, exudate, fibrosis, keloid, and administration somatomedin.

14. the method for claim 11, during wherein with described compound administration, avoided with KDR outside the relevant side effect of change of cellular signal transduction function of other tyrosine kinase.

15. the method for claim 7, wherein said chemical compound is selected from single-chain antibody, and KDR-specificity ribozyme and antisense polynucleotide have suppressed suitably presenting of functional KDR tyrosine kinase thereby wherein introduce or produce said chemical compound in cell.

16. the method for claim 7, wherein with described chemical compound be selected from the edema steroid, Ras inhibitor, anti-TNF agent, anti--the IL1 agent, hydryllin, PAF-antagonist, COX-1 inhibitor, cox 2 inhibitor, NO synthase inhibitor, non-steroid antiinflammatory (NSAID), pkc inhibitor and PI ₃The medication combined administration of inhibitors of kinases.

17. in individuality, suppress the method for a kind of physiological process or state, described physiological process or state are selected from edema and form, hemocyte oozes out, exosmose, seepage is oozed out, and ascites forms, apposition and blood vessel hypotension, wherein said inhibition comprise and will suppress the compound administration of the cellular signal transduction function of KDR.

18. the method for claim 17, wherein said chemical compound is optionally to described KDR tyrosine kinase.

19. the method for claim 18, wherein said chemical compound is selected from peptide, and antibody and organic molecule, wherein said chemical compound are attached to described KDR tyrosine kinase.

20. the method for claim 19 wherein with described compound administration, suppresses to be selected from the formation of the morbid state in following group: macular edema, aphakia/pseudophakia capsule sample speckle edema, retinoblastoma, eye ischemia, eye inflammation or infection, the edema side reaction that choroidal melanoma, iron chelating therapy cause, pulmonary edema, myocardial infarction, atrophic diseases, anaphylaxis, the tissue edema at wound and allergic inflammation position, allergy, allergy, the polyp edema at chronic inflammatory disease position, cerebral edema, cerebral tumor topping up cyst, communicating hydrocephalus, carpal tunnel syndrome, the organ injury that burn causes sucks burn injury, skin burn, the vesicle relevant with sun burns stimulates or infection erythema multiforme, edema speckle and other dermatosis, the cerebral tumor, tumor is oozed out, pulmonary carcinoma or breast carcinoma, ascites, pleura oozes out, pericardium oozes out, plateau " disease ", radioanaphylaxis, radiodermatitis, glaucoma, conjunctivitis, choroidal melanoma, adult respiratory distress syndrome, asthma, bronchitis, the super thorn of ovary bowel syndrome, polycystic ovarian syndrome, menstruation swelling, menstrual cramps, apoplexy, head trauma, cerebral infarction or obturation, hypotension, ulcer is sprained, fracture, what synovitis was relevant oozes out diabetic complication, HSV, hepatitis interstitialis chronica, microalbuminuria disease, albuminuria, uropenia, electrolyte disturbance, nephrotic syndrome, exudate, fibroidization, keloid, and administration somatomedin.

21. the method for claim 17, wherein said chemical compound suppress the catalysis kinase activity of described KDR.

22. the method for claim 17, wherein said chemical compound are the activatory antagonisies of KDR tyrosine kinase.

23. the method for claim 17, wherein said compound selective suppresses the phosphorylation of KDR kinase substrate.

24. the method for claim 17, wherein said compound selective is at described KDR tyrosine kinase.

25. the method for claim 17, the cellular signal transduction function of wherein said KDR is attached to the KDR acceptor portion by the activation part to stimulate.

26. the method for claim 25, wherein said chemical compound are that selectivity is at described KDR tyrosine kinase.

27. the method for claim 17, wherein said chemical compound is selected from single-chain antibody, and KDR-specificity ribozyme and antisense polynucleotide have suppressed suitably presenting of functional KDR tyrosine kinase thereby wherein introduce or produce above-claimed cpd in cell.

28. the method for claim 17, the inhibition of wherein said KDR cellular signal transduction function is the process that is selected from following group: the generation of blocking-up activation part, modulation activation part is to the combination of kdr tyrosine kinase receptor, destroy the dimerization of receptor, blocking-up KDR changes phosphorylation, the activity that suppresses the KDR tyrosine kinase is damaged raising of the interior substrate of KDR born of the same parents, and blocking-up is by the initial downstream signal conduction of KDR tyrosine kinase phosphorylation activity.

29. the method for claim 17, during wherein with described compound administration, avoided with KDR outside the relevant side effect of change of cellular signal transduction function of other tyrosine kinase.

30. the method for claim 17, wherein with described chemical compound be selected from the edema steroid, Ras inhibitor, anti-TNF agent, anti--the IL1 agent, hydryllin, PAF-antagonist, COX-1 inhibitor, cox 2 inhibitor, NO synthase inhibitor, non-steroid antiinflammatory (NSAID), pkc inhibitor and PI ₃The medication combined administration of inhibitors of kinases.