CN101626759A

CN101626759A - The method, compositions and the preparation that are used for the treatment of thyroid eye diseas

Info

Publication number: CN101626759A
Application number: CN200780046741.1A
Authority: CN
Inventors: J·D·多巴克
Original assignee: LIPOTHERA Inc
Current assignee: Niort Tikkes company
Priority date: 2006-10-17
Filing date: 2007-09-27
Publication date: 2010-01-13
Anticipated expiration: 2027-09-27
Also published as: CN101594873A; CN101626759B

Abstract

The invention provides the compositions, preparation, the method and system that are used for the treatment of thyroid eye diseas and relevant disease (for example graves' ophthalmopathy).The inventive method comprises that whole body or topical administration have patient's Beta-3 adrenergic agonist (for example controlled-release crystallized microgranule suspensoid) of needs.This method also can be included in and give to be used to reduce the chemical compound (for example corticosteroid) that Beta-3 adrenergic receptor desensitizes jointly before the Beta-3 adrenergic agonist or with the Beta-3 adrenergic agonist.The inventive method also can be included in and give Beta-3 adrenergic agonist topical administration eye immunosuppressant (for example rapamycin) before.The present composition comprises the eye medicinal preparation of Beta-3 adrenergic agonist of controlled-release crystallized microgranule suspensoid form or the mixture of crystal fine grain suspensoid and Beta-3 adrenergic agonist solution.Described compositions also comprises the ophthalmic preparation of chemical compound that is used to reduce Beta-3 adrenergic receptor desensitization of controlled-release crystallized microgranule suspensoid form.

Description

The method, compositions and the preparation that are used for the treatment of thyroid eye diseas

Cross reference

[0001] the application requires the U.S. Provisional Patent Application serial number 60/852 of application on October 17th, 2006,221,60/898 of application on January 29th, 2007,60/919,011 rights and interests of on April 13rd, 009 and 2007 application, described application all is attached to herein by reference.

Background of invention

[0002] (Graves ' disease) is a kind of commonly encountered diseases to Graves disease, and women's sickness rate was 1/1000 people/year.Beyond dethyroidize was hyperfunction, 25-50% suffered from the individual PD of Graves disease to involving eyes, i.e. thyroid eye diseas clinically.(Graves ' Ophthalmopathy GO) is the canonic form of thyroid eye diseas to graves' ophthalmopathy.Though some GO patients only suffer from slight eyes discomfort, 3-5% suffer from have an intense pain and inflammation with diplopia and even visual loss.

[0003] clinical symptoms of GO and sign can mechanically be interpreted as bone eye socket (bonyorbit) inner tissue volume obviously increases.The socket of the eye organization grows causes eyeball reach, is obstructed from the effluent of socket of the eye vein and socket of the eye lymph.These change in conjunction with the local cytokine that produces and other inflammatory mediator, cause exophthalmos, periorbital edema, conjunctiva erythema and chemosis (Fig. 1).

Summary of the invention

[0004] disclosed herein is to contact by the compound compositions of Beta-3 adrenergic receptor in making the fixed eye lipidosis of target (fat deposit) and comprising long-acting beta-2 adrenergic receptor agonists and reduce target tissue to long-acting beta-2 adrenergic receptor agonists desensitization, thereby is used for the treatment of compositions, preparation, the method and system of thyroid eye diseas.For example inject and/or give the compositions of each embodiment through eye through (behind the eyes) behind the eyeball.Glucocorticoid has the inflammatory cell that exists in minimizing socket of the eye and the socket of the eye fatty tissue and reduces the adjection that inflammatory cytokine discharges.

[0005] therefore, on the one hand, this paper is provided for reducing the lipopectic method of socket of the eye that the patient (curee who for example suffers from thyroid eye diseas) who needs is arranged, and this method is by at least a Beta-3 adrenergic agonist that gives the patient treatment effective dose and at least a chemical compound that is used to reduce the B-adrenergic receptor desensitization for the treatment of effective dose.In some embodiments, the patient who had described just now treatment to need suffers from graves' ophthalmopathy.In some embodiments, need the patient of treatment to suffer from the extraocular muscles hypertrophy.In some embodiments, the patient suffers from exophthalmos.

[0006] in some embodiments, give at least a Beta-3 adrenergic agonist or at least a chemical compound that is used to reduce desensitization in parenteral, oral, ophthalmic, socket of the eye, in the cone (intraconal), behind eye (ophthalmic), eyeball, socket of the eye week, part, intramuscular, percutaneous, Sublingual, intranasal or respiratory tract carry out.

[0007] in some embodiments, at least a chemical compound (before for example about 3 days to about 7 days) before giving at least a Beta-3 adrenergic agonist that is used to reduce the B-adrenergic receptor desensitization gives.In some embodiments, at least a chemical compound gives by route of administration in ophthalmic, the socket of the eye, behind eye, socket of the eye week, eyeball or in the cone.In some embodiments, at least a chemical compound gives with the form of crystal fine grain suspensoid (crystalline microparticle suspension).

[0008] in some embodiments, at least a chemical compound oral administration gives, and at least a Beta-3 adrenergic agonist gives through eye.At least a Beta-3 adrenergic agonist that gives through eye gives with the crystal fine grain preparation.

[0009] in some embodiments, wait that at least a Beta-3 adrenergic agonist that gives the patient comprises the long-acting beta 2-adrenergic agonist components.In some embodiments, at least a chemical compound is a glucocorticoid, and the long-acting beta 2-adrenergic agonist components is salmaterol (salmeterol), formoterol (formoterol) or its combination.In some embodiments, Beta-3 adrenergic agonist to be given is to beta-2 adrenergic receptor Beta-3 adrenergic agonist selectively.In some embodiments, at least a Beta-3 adrenergic agonist comprises salmaterol, formoterol or its any combination.In some embodiments, at least a Beta-3 adrenergic agonist comprises salmaterol, and the salmaterol of treatment effective dose is the salmaterol of about 0.01 μ g/ days to about 100 μ g/ days (for example about 1 μ g/ days to about 100 μ g/ days, about 10 μ g/ days extremely about 100 μ g/ days or about 50 μ g/ days to about 100 μ g/ days).In other embodiments, at least a Beta-3 adrenergic agonist comprises formoterol, and the formoterol of treatment effective dose is about 0.001 μ g/ days to about 50 μ g/ days (for example 0.01 μ g/ days to about 1.0 μ g/ days, about 0.1 μ g/ days extremely about 10 μ g/ days, about 1 μ g/ days extremely about 20 μ g/ days or about 5 μ g/ days to about 40 μ g/ days).

[0010] in some embodiments, at least a chemical compound that is used to reduce the Beta-3 adrenergic receptor desensitization is glucocorticoid, antihistaminic or its any combination.In some embodiments, at least a chemical compound that is used to reduce the Beta-3 adrenergic receptor desensitization comprises dexamethasone (dexamethasone), prednisolone (prednisolone), methylprednisolone (methylprednisolone), fluticasone propionate (fluticasone propionate), budesonide (budesonide), ketotifen (ketotifen) or its any combination.

[0011] in some embodiments, before giving at least a Beta-3 adrenergic agonist of patient and at least a chemical compound that is used to reduce the Beta-3 adrenergic receptor desensitization, treat the immunosuppressant of effective dose by approach in ophthalmic, the socket of the eye, behind eye, socket of the eye week, eyeball or in the cone.In some embodiments, immunosuppressant gives with the form of crystal fine grain suspensoid.

[0012] on the other hand, provided herein is to be used for the treatment of pop-eyed method, and this method comprises the compositions of at least a Beta-3 adrenergic agonist for the treatment of effective dose by the patient who needs treatment.

[0013] in some embodiments, at least a Beta-3 adrenergic agonist comprises the long-acting beta 2-adrenergic agonist components.In some embodiments, at least a Beta-3 adrenergic agonist comprises beta-2 adrenergic receptor Beta-3 adrenergic agonist selectively.In some embodiments, at least a Beta-3 adrenergic agonist comprises salmaterol, formoterol, bambuterol (bambuterol), eformoterol, isoproterenol (isoproterenol), albuterol (albuterol) or fenoterol (fenoterol).In some embodiments, compositions comprises the mixture of at least a long-acting beta 2-adrenergic agonist components and at least a fugitive Beta-3 adrenergic agonist.In some embodiments, compositions also comprises the hyaluronidase for the treatment of effective dose.

[0014] in some embodiments, compositions comprises salmaterol, and the salmaterol that gives the patient treatment effective dose is about 0.01 μ g/ days to about 100 μ g/ days.In other embodiments, compositions comprises formoterol, and the formoterol that gives the patient treatment effective dose is about 0.001 μ g/ days to about 50 μ g/ days.

[0015] in some embodiments, give compositions by in parenteral, oral, ophthalmic, the socket of the eye, socket of the eye week, behind eye, eyeball, in the cone, part, intramuscular, percutaneous, Sublingual, intranasal or respiratory tract carry out.

[0016] on the other hand, provided herein is the lipopectic method of socket of the eye that is used to reduce the patient of needs treatment, this method by giving the patient treatment effective dose one or more adrenoreceptor pathway stimulation chemical compounds and at least a chemical compound that is used to reduce the Beta-3 adrenergic receptor desensitization of treatment effective dose.In some embodiments, one or more adrenoreceptor pathway stimulation chemical compounds comprise catecholamine, alpha-1 adrenergic antagonists, forskolin, aminophylline or its analog.

[0017] another aspect, provided herein is medical composite for eye, described compositions comprises the methylprednisolone acetate or the fluticasone propionate of the crystal fine grain suspensoid form of acceptable excipient of eye and treatment effective dose.In some embodiments, medical composite for eye also comprises solubilising methylprednisolone acetate or solubilising fluticasone propionate.In some embodiments, medical composite for eye also comprises at least a long-acting beta-2 agonist that is crystal fine grain suspensoid form for the treatment of effective dose.

[0018] on the other hand, provided herein is medical composite for eye, and described compositions comprises at least a long-acting beta-2 agonist that is crystal fine grain suspensoid form of acceptable excipient of eye and treatment effective dose.In some embodiments, at least a long-acting beta-2 agonist comprises salmaterol or formoterol.In some embodiments, medical composite for eye also comprises at least a solubilising long-acting beta-2 agonist for the treatment of effective dose.In some embodiments, medical composite for eye also comprises at least a chemical compound that is used to reduce the crystal fine grain suspensoid form of Beta-3 adrenergic receptor desensitization of treatment effective dose.

[0019] another aspect, provided herein is at least a Beta-3 adrenergic agonist and at least a chemical compound that is used for reducing the Beta-3 adrenergic receptor desensitization are used for the treatment of the medicine of the disease that comprises the socket of the eye lipopexia in preparation purposes.

[0020] on the other hand, provided herein is at least a Beta-3 adrenergic agonist and at least a purposes that is used for reducing the chemical compound of Beta-3 adrenergic receptor desensitization in the method that is used for the treatment of the disease that comprises the socket of the eye lipopexia.

Quoting of list of references

[0021] mentioned all publications and patent application all is attached to herein by reference in this description, if the concrete and indivedual dated combinations by reference of each publication or patent application, it is identical with it then to quote degree.

The accompanying drawing summary

[0022] new feature of the present invention is referring to the concrete feature that is provided in the appended claims.Can understand feature of the present invention and advantage better by the exemplary that provides with reference to following utilization principle of the present invention and the detailed description of accompanying drawing:

[0023] Fig. 1 is steatolysis (adipocyte liposysis) sketch map of explanation adipose cell.

[0024] fat-splitting dose dependent was induced after Fig. 2 block diagram explanation was being hatched 3 hours by long-acting beta-2 agonist formoterol in the adipose cell of cultivating.

[0025] Fig. 3 block diagram illustrates that hatching the fat-splitting dose dependent in back at 3 hours by long-acting beta-2 agonist salmaterol in the adipose cell of cultivating induces.

[0026] Fig. 4 block diagram illustrates that hatching the fat-splitting dose dependent in back by the glucocorticoid budesonide in the short time (3 hours) in the adipose cell of cultivating induces, and hatches the fat-splitting inhibitory action in back in the long period (18 hours).

[0027] Fig. 5 block diagram explanation reaches 18 hours to fat-splitting dose-dependent inhibition by giving long-acting beta-2 agonist salmaterol separately in the adipose cell of cultivating, and when with glucocorticoid budesonide drug combination after 18 hours salmaterol fat-splitting dose dependent is induced.

[0028] Fig. 6 block diagram explanation is during treatment in 3 days, injects solvent solutions (2%PEG) in fat pad, when only formoterol or formoterol add budesonide, the mean difference (left fat pad and right fat pad) of epididymis fat pad quality in the animal body.

[0029] explanation of Fig. 7 block diagram is during treatment in 3 days, and β-2 agonist formoterol of two kinds of various dose combinations and glucocorticoid budesonide alleviate the dose dependent of fat pad quality.

Description of Preferred Embodiments

[0030] as if computed tomography scan shows most of GO patient's socket of the eye fat and extraocular muscles both hypertrophys, and other patient only involves fatty tissue or extraocular muscles.Extraocular muscles cell itself is not impaired in the active disease in early days, and this just shows that they itself are not the targets that autoimmune is attacked.Yet accumulating of hydrophilic mucopolysaccharide (especially comprising hyaluronan) caused extraocular muscles body hypertrophy in the perimysium connective tissue.In the disease, intramuscular decomposability inflammatory process makes it fibre modification and displacement late.

[0031] as if capsula adiposa bulbi/connective tissue volume increase impels the overall expansion of socket of the eye tissue volume more more remarkable than extraocular muscles hypertrophy.Computed tomography studies show that the dependency of these patients' exophthalmos size and fat deposit (fat compartment) volume is the closest.As if the expansion of this adipose tissue volume is accumulated by the hyaluronan of the edema that occurs together and the interior new adipose cell group's who breaks up of these tissues appearance causes.

[0032] Microscopic examination showed of GO socket of the eye tissue, this characteristic change mainly by the hyaluronan of the edema that occurs together accumulate, the expansion of fat deposit and produce by the tissue infiltration that the T lymphocyte causes.As if this sick socket of the eye adipose cell differentiation occurs, although littler than other adipose cell in the body (for example subcutaneous or nethike embrane).The socket of the eye adipose cell of GO is expressed PPAR-γ, adiponectin and the leptin mRNA transcript of higher level.In addition, the socket of the eye adipose cell can contain 1 less type 11-beta-hydroxysteroid dehydrogenase, and this is the enzyme that a kind of participation changes into cortisone the hydrocortisone of activity form.

[0033] uses and studies show that available from the cell of these tissues the socket of the eye fibroblast is the socket of the eye cell that can participate in the different cell processes of this class.These cells are to responsive especially by cytokine and the caused stimulation of other immune mediator, and the reaction of appearance is that increase CD40 expresses, synthetic a large amount of hyaluronan justacrine goes out inflammatory cytokine.In addition, fibroblastic preceding adipose cell subgroup can be divided into the mature fat cell with high-level TSHR.Research shows that also fibroblast has IGF-1R.When combining with the IgG that is derived from the Robert Graves patient, these receptor promoter downstream signal transductions cause producing RANTES and IL-16, and cause regional nodes's cellular infiltration.The Graves disease mesoconch get involved the part relative site specific can explain that as if these positions be squeezed in fibroblast in low pressure lymphatic vessel and the duct of Arantius by these fibroblasts to the relative sensitivity of immune mediator and unique anatomical features at these positions.

[0034] fatty tissue is a main energy storage tissue in the body.Adipose cell (Fatcell/adipocyte) is stored this energy with the form of triglyceride.Transfer triglyceride by fat depot, thereby provide heat energy to whole body by the triglyceride hydrolysis of hormone induction.This process will be dissociated or non-esterified fatty acid and glycerol is discharged in the blood for other bodily tissue utilization.The decomposition of triglyceride is called as steatolysis (lipolysis) in the fat depot.Also can carry out new adipose cell growth, this is called as lipogenesis.

[0035] catecholamine is the main fatty tissue regulator by adrenoreceptor.Fatty tissue has β-1 adrenoreceptor, beta-2 adrenergic receptor and β-3 adrenoreceptor and alpha-2 adrenergic receptor.In the fatty tissue, beta-agonists combines the steatolysis that can cause adipose cell with beta receptor, and can suppress steatolysis with combining of α receptor stimulating agent.The beta receptor activation also can suppress lipogenesis.In the human body, the β on the fat cell surfaces-2 receptor is the abundantest usually, and β-2 receptor is the fat-splitting main medium of beta receptor irriate.The steatolysis that is stimulated by beta-agonists (is encircled AMP, cAMP) forms increase and mediate by adenyl cyclase and 3'5'-AMP.Alpha-2 receptor reduces the steatolysis of mature fat cell.Alpha-2 adrenergic receptor can participate in preceding lipocyte proliferation.Glucocorticoid has permission effect (permissive effect) to fatty tissue, and strengthens the reaction (for example steatolysis) that adipose cell stimulates catecholamine.Due to this permission effect may be raised by other composition of second message,second messenger in B-adrenergic receptor and the participation born of the same parents.

[0036] treatment GO is necessary that targeting reduces the volume that the socket of the eye fatty tissue enlarges.Therefore, can confirm that the preparation that reduces the adipose cell volume by steatolysis can be used for this disease.In addition, slow down with inflammatory process in fatty tissue enlarges relevant socket of the eye and to reduce and to enlarge with fatty tissue that inflammatory cell can further reduce the socket of the eye tissue volume in the relevant socket of the eye.In addition, hyaluronan is accumulated and can additionally be reduced volume again in the treatment socket of the eye, and alleviates described oculopathy.At last, suppress lipogenesis and can improve this disease.

[0037] once researched and proposed adrenergic active component (beta-agonists and α-2 antagonisies) was delivered to subcutaneous tissue, and confirmed that this can cause local fat loss and local lipopectic the appearance to improve.For example, studies show that isoproterenol 11 and Yohimbine (yohimbine) 8 reduces women's thigh circumference.Because these fat-splitting agents (especially beta-agonists) are fugitive, can dispose fast from fatty tissue, may only in the short time after injection steatolysis take place, therefore, though multiple injection, potential effect value still reduced.In addition, the adipose cell long term exposure causes receptor desensitization, downward modulation and forfeiture lipolysis activity in beta-agonists.Reduce or prevent and also can improve therapy the method for these effects of receptor.Yet the strategy that uses decomposition of adrenergic and glucocorticoid induced lipolysis and inhibition lipogenesis to treat the GO adipose cell can effectively alleviate the liver mass that causes clinical sign and symptom.

[0038] this paper has described by alleviating the embodiment of pharmaceutical composition that the socket of the eye liver mass is used for the treatment of thyroid eye diseas (for example graves' ophthalmopathy or " GO "), preparation, method and system.Alleviate the socket of the eye liver mass and can alleviate exophthalmos, recover or prevent visual deterioration and diplopia, and ease the pain.Alleviating of this liver mass can realize by comprising at least one following aspect: alleviate the socket of the eye fat mass, reduce inflammation (for example inflammatory cell and cytokine) and reduce glycosaminoglycans (GAG) and accumulate.Fat mass alleviates to regulate by adrenergic system to be realized.Term " adjusting " used herein and/or citation generally uses its its ordinary meaning, is meant that more particularly adrenergic receptor kinase 1 action usefulness, adrenoreceptor antagonism and/or receptor signal transduction pathway change.The example that the receptor signal transduction pathway changes comprises that ring AMP increases, for example referring to Fig. 1 sketch map.In some embodiments, adjusting is meant that receptor raises or adrenoreceptor is counted increase, acceptor passivation or chelating minimizing, receptor active changes (for example active increase) and/or receptor affinity changes.The use of glucocorticoid or antihistaminic can preferably produce the adjusting to adrenoreceptor, and it also can be used to reduce inflammation.Use glucocorticoid can reduce glycosaminoglycans equally and accumulate, use the hyaluronic enzyme of degraded (for example recombined human hyaluronidase) further to reduce glycosaminoglycans and accumulate.

[0039] preferably in non-excision mode, by starting steatolysis, inhibition lipogenesis or reducing accumulation of lipid and realize alleviating of socket of the eye tissue fat quality.The cutting method that is used for reducing fat (for example phosphatldylcholine or deoxycholic acid) may have problems because when behind eye, using behind eye non-selectively disorganize can cause nerve or muscle injury, perhaps can cause incrustation and fibre modification.Stimulate steatolysis, inhibition lipogenesis and minimizing accumulation of lipid to realize by stimulating Beta-3 adrenergic receptor.Stimulate Beta-3 adrenergic receptor also can offset known some cell transcription things that in Robert Graves patient socket of the eye adipose cell, raise, for example PPAR-γ, adiponectin and leptin.For example, stimulate Beta-3 adrenergic receptor can in the adipose cell that has broken up, reduce the expression of PPAR-γ and adiponectin.We think that in the fatty tissue of some embodiments, the lasting adjusting of adrenoreceptor causes continuing some combination that lipid content reduces, the adipose cell size reduces, adipose tissue mass alleviates or lipopexia reduces of steatolysis, adipose cell.Some embodiments are by continuing adrenergic modulation, and the socket of the eye that provides selectivity to reduce fatty tissue and adipose cell is accumulated.In some embodiments, the adrenergic modulation of Chi Xuing causes lipocyte proliferation (lipogenesis) to be subjected to continuing to suppress.

[0040] the disclosed pharmaceutical composition of each embodiment comprises at least a selectivity beta-2 adrenergic receptor agonists (for example long-acting selectivity β-2 agonist) and at least a chemical compound combination that reduces B-adrenergic receptor desensitization (for example moving agent target tissue desensitization of beta-adrenergic receptor kinase 1, for example glucocorticoid or ketotifen or its analog) and reduce inflammation.Term desensitizes and comprises short term desensitization (desensitization fast) and long-term desensitization and interior At All Other Times desensitization.Beta-2 adrenergic receptor agonists this paper also claims " β-2 agonist " and " β-2 receptor stimulating agent ".Except as otherwise noted, otherwise also comprise its analog, physiologically acceptable salt and/or solvate when mentioning beta-2 adrenergic receptor agonists.Long-acting selectivity β-2 agonist that the compositions of some embodiments comprises is about 100: 1 to about 1: 100 with the ratio of glucocorticoid.

[0041] as mentioned above, we think that lipolysis activity, lipocyte proliferation suppress and accumulation of lipid reduces by regulating in the fatty tissue and/or the adrenoreceptor on the adipose cell mediates.In some embodiments, promote relieve therapies (reduction therapy) by prolongation exposure or continuous activity in one or more 3 adrenergic receptor agonists and/or receptor pathway stimulus compound (for example catecholamine, beta-agonists, alpha-2 antagonists, forskolin, aminophylline, its analog or its combination).

[0042] some embodiments comprise one or more by use the fully optionally pharmaceutical composition of long-acting beta-2 receptor stimulating agent, and lasting adrenergic modulation is provided.The continuous activity pharmaceutical composition of some embodiments comprises one or more suitable long lasting selectivity β-2 agonist, for example salmaterol 1, formoterol 2, bambuterol 3, the physiologically acceptable salt of eformoterol, or its solvate, or its combination.

[0043] can't observe lasting adrenergic modulation with typical adrenergic compositions, because the general part of adrenergic compounds is disposed by blood and/or lymph fast because of its hydrophilic from fatty tissue.And we think that with the fatty tissue long term exposure in adrenergic, particularly the beta receptor agonist will cause receptor desensitization by receptor phosphorylation and chelation.We think, these effects have limited the ability of adrenergic modulation combination treatment fatty tissue, and cause quick desensitization, and this is to give initial dosage with after reaching desired fats decomposition and lipogenesis resisting effect, health is to the quick a kind of state that reduces of reaction experience of agonist.Therefore, therapeutic effect is of short duration.

[0044] as mentioned above, fugitive β-2 agonist usually causes quick desensitization.Yet, because long-acting selectivity β-2 agonist of embodiment preferred has fully optionally β-2 receptor active and highly lipophilic, so compare with fugitive β-2 agonist, the activity of long-acting beta-2 agonist continues long period of time in fatty tissue.Part β-2 receptor antagonist activity that occurs in when using salmaterol can prevent some desensitization, and this class desensitization can occur in when adipose cell continues to be exposed to complete 2-adrenergic agonist components.In addition, salmaterol can not activate the Profilin signal transduction that causes receptor internalization and degraded and cause long-term receptor down-regulated fully.Compare with fugitive β-2 agonist, the steatolysis of longer a period of time also takes place after the administration, because long-acting selectivity β-2 agonist half life is longer.Half life, can reduce the frequency that gives pharmaceutical composition than long with active combination.Therefore, in some embodiments, once a day or to give compositions once a day be not essential.And long-acting selectivity β-2 agonist of embodiment preferred also has bigger selectivity to β-2 receptor, and causing just can reach than low dosage to the chemical compound of the similar substantially therapeutical effect of fugitive β-2 agonist becomes possibility.In addition, also can limit side effect to heart than high selectivity β-2 activity, these side effect usually are inductive by β in the heart-1 receptor for stimulating institute.

[0045] as mentioned above, the inhibition of steatolysis and/or lipogenesis and accumulation of lipid is stimulated by β-1, β-2 or beta-3 receptor hypotype.The agonist of therefore, a kind, 2 kinds and/or all 3 kinds of receptors can stimulate steatolysis and/or suppress lipogenesis.We think in human body, and β-2 receptor active is even more important for stimulating steatolysis, particularly when anti-inflammatory steroids or glucocorticoid exist.

[0046] long-acting selectivity β-2 agonist, for example salmaterol 1 (± 2-(methylol)-4-[1-hydroxyl-2-[6-(4-phenyl butoxy) hexyl amino] ethyl]-phenol, CAS number of registration 94749-08-3) and formoterol 2 (± N-[2-hydroxyl-5-[1-hydroxyl-2-[1-(4-methoxyphenyl) third-2-base is amino] ethyl]-phenyl] Methanamide, CAS number of registration 73573-87-2) be preferred in some embodiments.The compositions of some embodiments comprises one or more long-acting selectivity β-2 agonist as physiologically acceptable salt or solvate, for example former times naphthoic acid salmaterol and/or formoterol fumarate.In many cases, the salt of β-2 agonist and/or solvate can have needed activity.Therefore, except as otherwise noted, otherwise mention active component, for example when salmaterol 1, formoterol 2, isoproterenol 4, albuterol 5, fenoterol and forskolin, comprise chemical compound itself and physiologically acceptable analog, salt and/or solvate or its combination.

[0047] some preferred long-acting beta agonist have higher inherent adenylate cyclase activity, and it is synthetic that this has just increased cAMP.For example, some embodiments comprise formoterol 2 as long-acting beta-2 selective agonist, and it has some combination of following aspect: effect is higher, general action reduces, the adenyl cyclase height is inherent activates and/or ring AMP (a kind of steatolysis medium) increase.

[0048] in some preferred embodiments, formoterol 2 exists with its physiologically acceptable salt and/or solvate.Formoterol 2 suitable physiologically acceptable salt comprise and for example derive from mineral acid and organic acid acid-addition salts, for example hydrochlorate, hydrobromate, sulfate, phosphate, maleate, fumarate, tartrate, citrate, benzoate, 4-methoxybenzoic acid salt, 2 hydroxybenzoic acid salt, the 4-hydroxy benzoate, the 4-chloro benzoate, tosilate, mesylate, Ascorbate, Salicylate, acetate, succinate, lactate, glutarate, gluconate, tricarballylic acid salt, hydroxyl naphthalene-carboxylic acid salt, oleate, and combination etc.Embodiment preferred comprises that formoterol 2 is its fumarate and/or is dihydrate.For adipose tissue treatment, formoterol 2 suitable tissue concentrations comprise that about 1pM is to about 100 μ M, more preferably from about 0.1nM is to about 10 μ M, for example about 1nM to about 1 μ M, about 40nM extremely about 3 μ M, about 0.1 μ M extremely about 1 μ M or from about 0.1nM to any other formoterol tissue concentration about 10 μ M.

[0049] in some embodiments, salmaterol is used in compositions as herein described and the method.Salmaterol 1 has partial agonist activity, and we think that this can reduce receptor desensitization, and can limit the Profilin signal transduction that causes less receptor down-regulated.In some embodiments, salmaterol 1 exists with physiologically acceptable salt and/or its solvate.Salmaterol 1 suitable physiologically acceptable salt includes but not limited to derive from mineral acid and organic acid acid-addition salts, for example hydrochlorate, hydrobromate, sulfate, phosphate, maleate, tartrate, citrate, benzoate, 4-methoxybenzoic acid salt, 2 hydroxybenzoic acid salt, the 4-hydroxy benzoate, the 4-chloro benzoate, tosilate, mesylate, Ascorbate, Salicylate, acetate, fumarate, succinate, lactate, glutarate, gluconate, tricarballylic acid salt, hydroxyl naphthalene-carboxylic acid salt, 1-hydroxyl-2-naphthalene-carboxylic acid salt, 3-hydroxyl-2-naphthalene-carboxylic acid salt, oleate, and combination etc.In some embodiments, salmaterol 1 provides as 1-hydroxyl-2-naphthalene-carboxylic acid salt (Hydroxynaphthoate).

[0050] in some embodiments, for adipose tissue treatment, salmaterol 1 suitable tissue concentration scope is that about 1pM is to about 100 μ M, preferred about 1.0nM is to about 1 μ M, for example about 10nM to about 1 μ M, about 40nM extremely about 3 μ M, about 0.1 μ M extremely about 1 μ M or from about 1.0nM to any other salmaterol tissue concentration about 10 μ M.

[0051] in some embodiments, long-acting selectivity β-2 agonist to be given is a formoterol, the formoterol of treatment effective dose is about 0.001 μ g/ days to about 100 μ g/ days, for example about 0.001 μ g/ days to about 50 μ g/ days, 0.01 μ g/ days to about 1.0 μ g/ days, about 0.1 μ g/ days to about 10 μ g/ days, about 1 μ g/ days to about 20 μ g/ days, about 5 μ g/ days to about 40 μ g/ days, about 25 μ g/ days to about 75 μ g/ days, about 50 μ g/ days to about 100 μ g/ days formoterol or from about 0.001 μ g/ days to about 100 μ g/ days the formoterol of any other dosage.

[0052] in some embodiments, long-acting selectivity β-2 agonist to be given is a salmaterol, the treatment effective dose of salmaterol to be given is about 0.01 μ g/ days to about 1000 μ g/ days, for example about 0.1 μ g/ days to about 100 μ g/ days, about 1 μ g/ days to about 100 μ g/ days, about 10 μ g/ days to about 100 μ g/ days, about 50 μ g/ days to about 100 μ g/ days or from about 0.01 μ g/ days to about 1000 μ g/ days the salmaterol of any other dosage.

[0053] " treatment effective dose " used herein is meant that the amount for the treatment of administered agents (for example long-acting β2Ji Dongji) or chemical compound is enough to alleviate to a certain extent one or more symptoms of disease to be treated or disease.Its result can be sign, symptom or the cause of disease that alleviates and/or alleviate disease, or the variation of any other needed biosystem.For example, " effective dose " that is used for the treatment of purposes is that needed confession significantly palliates a disease symptom clinically and do not have the amount that comprises compound compositions disclosed herein of excessive toxic and side effects.In any indivedual cases,, application dose determines suitable " effective dose " but increasing progressively technology such as research.Term " treatment effective dose " comprises and for example prevents effective dose.Chemical compound disclosed herein for example selectivity β-2 agonist list with or with " effective dose " of other chemical compound (for example being used to reduce the chemical compound of beta-2 adrenergic receptor desensitization) coupling, be to be enough to reach required pharmacological action or to improve treatment and do not have the amount of excessive toxic and side effects.Be appreciated that, since β-2 agonist with the order of severity of age of the metabolism of the chemical compound (for example glucocorticoid) of β-2 agonist coupling, curee, body weight, general situation, disease to be treated, disease to be treated and prescriber judge different, so " effective dose " or " treatment effective dose " can be different between curee and curee.

[0054] some embodiments comprise the optically-active pure isomer of Beta-3 adrenergic agonist, and this can improve steatolysis and lipogenesis suppresses, and reduces potential side effect.In some embodiments, these optically-active pure isomers for example by removing one or more no physiological effecies, low physiological effect, negative effect and/or the destroyed isomer of physiological effect, make preparation comprise the active component than heavy load.Remove unwanted bonded racemic mixture, isolate active isomer or excellent enantiomer (eutomer), thereby in giving customization agent, make it the more excellent enantiomer of load by the composition of removing non-activity.

[0055] two diastereomers of the general generation in two three-dimensional centers in the molecule, this paper is called (R ^*, R ^*) and (R ^*, S ^*) and enantiomer.Diastereomer that is to say for being not the stereoisomer of enantiomer, the mirror image of a diastereomer and another diastereomer can not be overlapping.Enantiomer is the stereoisomer of mirror image each other.Racemic modification is 1: 1 mixture of enantiomer.(R ^*, R ^*) enantiomer of diastereomer is called (R, R) enantiomer and (they are mirror image each other for S, S) enantiomer, and therefore some common physicochemical property, for example fusing point are arranged.Similarly, (R is S) with (S, R) isomer is (R ^*, S ^*) enantiomer of enantiomer.For example, can obtain (R, R)-isomer with (S, S)-ratio of isomer is the formoterol 2 of 1: 1 racemic modification, is generally the dihydrate of its fumarate.Some embodiments comprise (R, R) enantiomer, promptly (R, R)-formoterol, it during as long-acting beta-2 agonist activity bigger.Some embodiments comprise the optically-active pure isomer of other β-2 agonist, for example (R)-salmaterol.

[0056] in addition, in some embodiments, at least a long-acting selectivity β-2 agonist is highly lipophilic, and therefore the pharmaceutical composition with continuous activity is provided in fatty tissue.We think that fat-solubility (high lipid solubility) has prolonged the time that β-2 agonist exists in fatty tissue, therefore in some embodiments, eliminate or have reduced needs to slow release and/or controlled release carrier.In comprising the preparation of slow-released carrier, release polymer for example, the highly lipophilic of β-2 agonist helps mixing in the slow-released carrier, and more details vide infra.

[0057] salmaterol 1 and formoterol 2 have fat-solubility, and this has prolonged their life periods in fatty tissue and/or one or more adipose cells.The compositions of some embodiments comprises the beta-agonists of highly lipophilic since be assigned in the fatty tissue and in fatty tissue chelating, thereby reduce or eliminated needs slow release or controlled release carrier, and then therapeutic effect is prolonged.In some embodiments, use oil-water partition coefficient at least about 1000 or at least about 10,000 to 1 beta-agonists.For example, the lipotropy of salmaterol 1 is than at least 10,000 times of albuterol 5 (a kind of fugitive hydrophilic beta-agonists) height.In addition, salmaterol 1 and formoterol 2 have antiinflammatory property, the GO that equally is used for the treatment of as mentioned below.In some embodiments, they also promote favourable extracellular matrix to change, and reduce body fluid and accumulate, and this has improved GO and the lipopectic treatment of socket of the eye.

[0058] continues the Beta-3 adrenergic activity and be further improved, when aforesaid adipose cell is exposed to 2-adrenergic agonist components continuously, just can desensitize by preventing desensitization (desensitization fast).(for example reducing the chemical compound of beta-agonists target tissue desensitization) comprises the chemical compound of all suitable reduction target tissues to the toleration of the moving agent of beta-adrenergic receptor kinase 1 " to reduce the chemical compound of B-adrenergic receptor desensitization ", comprise glucocorticoid and suitable antihistaminic, for example thyroxins such as ketotifen and T3 and T4.Glucocorticoid this paper also claims " anti-inflammatory steroids ", " glucocorticoid " and/or " corticosteroid ".We think that glucocorticoid makes the sensitization of socket of the eye lipopexia by increasing β-2 acceptor number, therefore helps steatolysis or fat minimizing during depot fat.Glucocorticoid also can reduce α-2 acceptor number.Glucocorticoid also can make the stable or reduction receptor down-regulated of receptor, especially when with beta-agonists administration simultaneously.It should be noted that women's Graves disease and GO than the male generally many.Estrogen can induce alpha-2 adrenergic receptor to express in women's subcutaneus adipose tissue, and the ratio that causes β-2 receptor and α-2 receptors is less than 1.It is generally acknowledged when the ratio about 1 of β-2 receptor and α-2 receptors is above, cause that in adipose cell fat reduces rather than lipopexia.The compositions that comprises one or more glucocorticoid of some embodiments is effective to treat fat sites, comprise and reduce β-2 acceptor number and/or increase α-2 acceptor number, steatolysis or lipogenesis that this can resist Beta-3 adrenergic stimulates suppress, for example subcutaneus adipose tissue, especially women.

[0059] therefore, we think between the beta-agonists exposure period, are used to reduce glucocorticoid or other chemical compound improvement steatolysis, lipogenesis inhibition and/or the local fat minimizing of Beta-3 adrenergic receptor desensitization.In some embodiments,, promptly keep and/or improve lipolysis activity in the target tissue, keep again and/or increased the beta-receptor number with the glucocorticoid treatment adipose cell that improves lipolysis activity.The example of suitable corticosteroid comprises dexamethasone 6, fluticasone propionate 7, budesonide 8, prednisolone 9, methylprednisolone 10 and analog thereof.In some embodiments, glucocorticoid is a dexamethasone.In some embodiments, corticosteroid is a methylprednisolone, 6 (9-fluoro-11,17-dihydroxy-17-(2-hydroxyacetyl)-10,13; 16-trimethyl-6,7,8,11,12; 14,15,16-octahydro cyclopenta [a] phenanthrene-3-ketone, CAS number of registration 50-02-2) and/or fluticasone propionate 7.

[0060] as mentioned above, in some embodiments, the suitable compound that is used to reduce the beta receptor desensitization is a ketotifen 11, and it also can be used as antihistaminic.The compositions of some embodiments comprises a kind of chemical compound of minimizing fatty tissue to the desensitization of β-2 agonist.

[0061] in some embodiments, use the multiple chemical compound that reduces the beta receptor desensitization, for example multiple glucocorticoid.Some embodiment preferred comprise the analog of at least a glucocorticoid and antihistaminic ketotifen or ketotifen.

[0062] in some embodiments, in people's socket of the eye adipose cell, when response anti-inflammatory steroids or ketotifen administration, particularly when beta-agonists existed, at least a β-2 receptor active or density increased.In some embodiments, increasing β-2 receptor active and/or density strengthens the effect of long-acting and fugitive β-2 agonist.Therefore, in some embodiments, glucocorticoid can make the sensitization of socket of the eye fat, to realize that β-2 receptor for stimulating, steatolysis, lipogenesis suppress and/or apoptosis, and/or the ratio of raising beta-2 adrenergic receptor and alpha-2 adrenergic receptor, thereby change lipopexia and fatty balance of losing in the fatty tissue.In some embodiments, especially use glucocorticoid, ketotifen or thyroxin, especially when with beta-2 adrenergic agonist co-administered, β-2 acceptor number increases or remains unchanged.

[0063] adds anti-inflammatory agent (for example glucocorticoid or antihistaminic) inflammatory cell and inflammatory reaction in the minimizing socket of the eye are had extra effect.The socket of the eye of GO and socket of the eye fat have the dispersivity lymphocytic infiltration, comprise that lymph is assembled or lymph is gone back to the nest (lymphoid nest).Other tissue of socket of the eye (for example extraocular muscles) has similar leukocyte infiltration.Also there is a large amount of mastocytes.These lymphocytic cytokine secretions are consistent with replying of cell and body fluid mediation.Glucocorticoid reduces leukocyte recruitment, cytokine secretion, and can induce the leukocyte apoptosis.Can strengthen these effects by long-acting selectivity β-2 agonist.In addition, long-acting selectivity β-2 agonist can make the glycocorticosteroid receptor in the leukocyte (even adipose cell) raise and make it stable, and can promote it to migrate in the nuclear, further strengthens antiinflammatory action.Glucocorticoid and long-acting beta agonist can make mastocyte stable, and can bring into play addition or potentiation.Ketotifen also can make mastocyte stable, also can suppress TNF-α, and TNF-α is a kind of important cytokine during the cellular inflammation of cell mediated is reacted.

[0064] glucocorticoid suitable that is used for therapy described herein organizes the range of concentrations can be between about 0.001 μ M between about 10mM, for example about 1.0 μ M to about 5mM, about 40 μ M extremely about 3mM, about 100 μ M extremely between about 1mM or from about 10 μ M to any other glucocorticoid tissue concentration about 10mM.

[0065] in some embodiments, glucocorticoid to be given is a budesonide, the budesonide of medicinal effective dose is about 1.0 μ g/ days to about 320 μ g/ days, for example about 80 μ g/ days to about 300 μ g/ days, about 100 μ g/ days to about 280 μ g/ days, about 120 μ g/ days to about 260 μ g/ days, about 140 μ g/ days to about 240 μ g/ days, about 160 μ g/ days to about 220 μ g/ days, about 180 μ g/ days to about 200 μ g/ days, about 185 μ g/ days to about 195 μ g/ days budesonide or from about 60 μ g/ days to about 320 μ g/ days the budesonide of any other dosage.

[0066] in some embodiments, glucocorticoid to be given is a fluticasone, the fluticasone of treatment effective dose is about 1.0 μ g/ days to about 500 μ g/ days, for example about 120 μ g/ days to about 480 μ g/ days, about 140 μ g/ days to about 460 μ g/ days, about 160 μ g/ days to about 440 μ g/ days, about 180 μ g/ days to about 420 μ g/ days, about 200 μ g/ days to about 400 μ g/ days, about 220 μ g/ days to about 380 μ g/ days, about 240 μ g/ days to about 360 μ g/ days, about 260 μ g/ days to about 340 μ g/ days, the fluticasone of about 275 μ g/ days to about 310 μ g/ days or about 290 μ g/ days to about 300 μ g/ days perhaps is the fluticasone of any other dosage from about 100 μ g/ days to about 500 μ g/ days.

[0067] in some embodiments, glucocorticoid to be given be about 1.0 μ g/ days to 10,000 μ g/ days above methylprednisolones, 50-5 for example, 000 μ g/ days, 100-5,000 μ g/ days, 500-5000 μ g/ days, 700-3,000 μ g/ days, 800-2500 μ g/ days, 1000-2000 μ g/ days or be about 1.0-10, any other dosage of 000 μ g/ days.In some embodiments, the succinic acid methylprednisolone can be solubilized in the crystal fine grain methylprednisolone acetate suspensoid, perhaps can with crystal fine grain methylprednisolone acetate suspensoid co-administered, so that instant dosage and lasting dosage to be provided.

[0068] compositions of some embodiments comprises extra optional ingredients.For example particularly under the GO situation, contain a large amount of glycosaminoglycans in eye socket and the socket of the eye lipopexia, glycosaminoglycans is made up of hyaluronic acid basically.In some cases, it is favourable making this hyaluronic acid degradation, for example improves Beta-3 adrenergic agonist and the diffusion of glucocorticoid preparation in whole socket of the eye.In addition, make hyaluronic acid degradation also further alleviate the socket of the eye liver mass, alleviate orbital edema, thereby improve exophthalmos and GO disease.The compositions of some embodiments comprises enzyme, for example make hyaluronic acid degradation hyaluronidase (recombined human hyaluronidase for example, i.e. recombined human hyaluronic acid injections (Hylenex), Halozyme Therapeutics, San Diego, CA).

[0069] compositions of some embodiments comprises one or more lipotropism fat and decomposes blocade (anti-lipolytic blocking agent), for example in accumulating, blocks local fat selectivity α-2 receptor antagonist of lipotropism fat decomposition, for example phentolamine (phentolamine) 12 (CAS number of registration 73-05-2) or Yohimbine 13 (CAS number of registration 146-48-5).Usually observe the lipotropism fat decomposition of adipose cell and fatty tissue at subcutaneous and lipopectic regional area.For example, when being exposed to beta-agonists, compared with interior fat, it is slow that the steatolysis speed of subcutaneous fat is wanted.In some embodiments, socket of the eye fat is exposed to lipotropism fat decomposition blocade and can improves lipolysis activity.

[0070] compositions of some embodiments comprises other adrenergic that improves the effect of long-acting selectivity β-2 agonist.For example, aminophylline (aminophylline) 14 (1,3-dimethyl-7H-purine-2,6-diketone, diethylamine, CAS number of registration 317-34-0) and theophylline (theophylline) 15 (CAS number of registration 58-55-9) all be the steatolysis medicine (lipolytic agent) that blocking-up ring AMP decomposes.

[0071] other optional ingredients strengthens by beta-agonists in conjunction with the secondary signal that produces.For example, in some embodiments, compositions comprises the forskolin 16 (CAS number of registration 66575-29-9) that stimulates adenyl cyclase, therefore makes the synthetic increase of the ring AMP that is started by the long-acting beta agonist.The increase of ring AMP concentration helps the maintenance lipolysis activity.

[0072] as if the compositions of some embodiments comprises the combination of growth hormone and long-acting beta agonist and glucocorticoid, and this can stimulate steatolysis.

[0073] compositions of other embodiment also comprises one or more non-selective beta-agonists (for example isoproterenol 4) and/or fugitive selectivity β-2 agonist (for example terbutaline (terbutaline)).Some compositions comprises at least a α-2 antagonist or its physiologically acceptable salt or solvate.

[0074] compositions of embodiment can be used for administration by the preparation of any suitable method, for example referring to Remington:The Science And Practice Of Pharmacy (the 21st edition, Lippincott Williams ﹠amp; Wilkins).The exemplary approach of administration includes but not limited in parenteral, oral, ophthalmic, the socket of the eye, socket of the eye is all, behind eye, eyeball, part, intramuscular, percutaneous, Sublingual, intranasal or respiratory tract.In some embodiments, compositions is formulated into the position that is used to be expelled to the needs treatment, for example in socket of the eye or the socket of the eye lipidosis.

[0075] in some embodiments, with beta-agonists, be used to prevent that the chemical compound of beta receptor desensitization or both are mixed with the crystal fine grain suspensoid and discharge to prolong, thereby further keep adrenergic modulation.

[0076] can use the excipient that is suitable for injection.Excipient normally ophthalmology is acceptable.In other words, excipient does not have long-term or persistent illeffects basically for the eyes for the treatment of administration.The example of ophthalmology acceptable carrier comprises water (distilled water or deionized water), saline and other aqueous medium.Chemical compound of the present invention preferably dissolves in the carrier that is applicable to administration, and therefore, chemical compound gives eyes with the solution form.Perhaps, also can use the suspensoid (for example suspensoid of crystal fine grain) of one or more reactive compounds in the suitable carrier.In some embodiments, one or more β-2 receptor stimulating agent or glucocorticoid are prepared in liquid-carrier, for example be mixed with solution, suspensoid, gel and/or Emulsion.Some embodiments comprise any suitable low-polarity component, and for example modified oil (for example

BASF), soybean oil, propylene glycol, Polyethylene Glycol, derivatization polyethers, its combination etc.Some embodiments comprise microparticle and/or the nano-particle carrier that is used at least a β-2 receptor stimulating agent and/or glucocorticoid, and more detailed the argumentation sees below.Some embodiments comprise one or more slow-released carriers or composition or controlled release carrier or composition, for example polymer microballoon.Some embodiments comprise the excipient of the stable suspensoid that is applicable to β-2 receptor stimulating agent or glucocorticoid micronized particle.

[0077] gives injection by any way, comprise and use single needle injection device, spininess injection device and/or use Needleless injection device.In some embodiments, the active component of in suitable carrier, preparing by injected delivery of organizing load capacity.In some embodiments, send and comprise single needle injection.In some embodiments, pass the medicine method and comprise use spininess array (multi-needle array) injection, in some embodiments, this can supply preparation long range diffusion in target tissue.In some embodiments, the mode of preparation injection allows to be diffused in the suitable socket of the eye fat deposit.In some embodiments, inject a spot of aliquot preparation to reduce to join the additional volumes in the socket of the eye, the scope of injection volume is that about 0.1ml is to about 0.5ml.In some embodiments, used injection device has looper and is beneficial to be delivered to behind the socket of the eye (retro-orbital) or bores intravital gap.

[0078] in some embodiments, give β-2 agonist and reduce the chemical compound that desensitizes, for example as independent preparation injection, perhaps, through independent administration (for example injecting long-acting beta-2 agonist behind the orally give glucocorticoid again).In some embodiments, before giving β-2 agonist, reduce the chemical compound of desensitization.In other embodiments, before the chemical compound that reduces desensitization, give β-2 agonist.

[0079] reduce the chemical compound of desensitization and the interval that gives between β-2 agonist can be about 5 minutes to reaching 7 days, for example 30 minutes, 1 hour, 6 hours, 12 hours, 1 day, 2 days, 3 days, 4 days, 5 days, 6 days or 7 days or from about 5 minutes to about 7 days any At All Other Times at interval.In a preferred embodiment, reach about 7 days before giving β-2 agonist (for example the ophthalmic preparation part being used for eyes), for example 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days or 10 days, orally give reduced the chemical compound (for example corticosteroid) of desensitization.

[0080] in other embodiments, the β-2 agonist ingredient of same preparation (for example as) is with chemical compound (for example glucocorticoid) co-administered that reduces the beta receptor desensitization.

[0081] in some embodiments, preparation provides depot formulation for curee to be treated, and depot formulation comprises one or more slow release compositions or controlled release composition and obtains slow release or controlled release so that β-2 agonist or be used to suppresses the chemical compound (for example glucocorticoid) of beta receptor desensitization.In this class preparation, β-2 agonist, be used for reducing the chemical compound of beta receptor desensitization or both and be encapsulated in slow release or controlled release composition or carrier, combine and/or put together with slow release or controlled release composition or carrier.In some embodiments, biodegradable biocompatibility slow release or controlled release preparation provide local tissue activity to reach several weeks to the several months.Suitable slow release or controlled release composition or carrier comprise polymer, macromole, active component conjugate, hydrogel, its pollutant etc.The slow-released carrier targeting fat of some embodiments, for example liposome.Preferred select to help time per unit, particularly at least about 3 days the course of treatment, more especially at least about 4 days to reaching the slow-release material of sending the active substance of basic equivalent more than 1 year in the course of treatment.Can change carry out slow releasing preparation some in time and take turns injection with the treatment single area.In some embodiments, with β-2 agonist or be used to reduce the chemical compound of beta receptor desensitization or both are mixed with crystalline drug microgranule suspensoid and produce slow release.

[0082] in some embodiments, the slow release composition comprises polymer, for example admixture, mixture or the copolymer of polylactide, poly-Acetic acid, hydroxy-, bimol. cyclic ester, poly-(lactide glycolide) polylactic acid, polyglycolic acid, polyanhydride, poe, polyethers, polycaprolactone, polyesteramide, Merlon, polybutylcyanoacrylate, polyurethanes, polyacrylate and above-mentioned polymer, these are used to encapsulated active ingredients (for example beta-agonists and/or glucocorticoid), combine with active component (for example beta-agonists and/or glucocorticoid) or put together.The release polymer of certain preferred embodiments comprises the polyethylene group that one or more active component are puted together with it.In some preferred embodiments, the slow release composition comprises poly-(lactide glycolide) (PLGA, lactic acid-ethanol copolymer) copolymer 17.

[0083] the slow release composition of some embodiments comprises one or more hydrogels, comprises the modification alginate.The example of suitable modification alginate comprises disclosed modification alginate among the WO 98/12228.The slow release composition of some embodiments comprises based on albuminous nanoparticulate carriers or excipient.

[0084] in some embodiments, the preparation that comprises prepolymer solution is injected into target tissue site, polymerization in body is carried out at this position then (for example by photopolymerization) or curing (for example by the responsive Binder Materials of serviceability temperature).

[0085] in some embodiments, controlled-release material has and is designed to be exclusively used in the release characteristics that dwindles tissue.In some embodiments, make slow release composition or controlled release composition form microgranule (for example microsphere), it is mixed with injection solution agent and/or gel.In some embodiments, the scope of mean particle dia size is extremely about 100 μ m of about 10 μ m, and size generally is uniform.In some embodiments, agent provides as injected gel to comprise the preparation of alginate and/or lactic acid-ethanol copolymer 17, perhaps is processed to microsphere.In other embodiments, make β-2 agonist or corticosteroid (or other is used to reduce the chemical compound of beta receptor desensitization) form crystal fine grain.Other example that is suitable for forming the suitable biodegradable biocompatible materials of injection of microgranule comprises chitosan, glucosan, hydroxyapatite and silicon.

[0086] adopts any method, comprise solvent evaporation and/or emulsion polymerisation, make microsphere and/or microgranule.In some embodiments, the average diameter of microsphere is extremely about 60 μ m of about 5 μ m, preferred about 20 μ m.In some embodiments, prepare PLGA with the lactide and the Acetic acid, hydroxy-, bimol. cyclic ester of different ratios, this ratio depends on the rate of release of required active component.Because the ratio of Acetic acid, hydroxy-, bimol. cyclic ester is proportional in the degradation rate of this copolymer and its degree of crystallinity and the preparation, the non-racemic mixture of lactide and/or Acetic acid, hydroxy-, bimol. cyclic ester improves degree of crystallinity and slows down degradation rate.The Acetic acid, hydroxy-, bimol. cyclic ester of higher proportion improves degradation rate.In some embodiments, the ratio of about 65%-75% lactide and about 25%-35% Acetic acid, hydroxy-, bimol. cyclic ester for active component in about 2 releases during thoughtful about 45 days.In other embodiments, the ratio of lactide and Acetic acid, hydroxy-, bimol. cyclic ester is about 0: 100 to about 100: 0, and therefore other rate of release is provided.

[0087] microsphere of some embodiments or microgranule have hollow and/or porous inside.In some embodiments, microsphere has closely knit or porous shell.

[0088] in some embodiments, the preparation that comprises porous shell and/or porous microsphere has the two-phase release characteristics of active component, and the initial disintegrate of active component disengages, and then is and the relevant lasting release of polymerizing microballoons degraded.Initial disintegrate is loaded into the steatolysis/lipogenesis inhibition and the antiphlogistic valid density of active component in the tissue, and needed concentration is then kept in slower subsequently release.In some embodiments, different micro-sphere structure and the active component release characteristics therapeutic effect that makes socket of the eye fatty tissue and adipose cell and by adrenergic receptor modulation with to the effect optimization of leukocyte infiltration to reduce inflammation.In some preferred embodiments, the lasting local organization concentration of long-acting selectivity beta-2 adrenergic medicine, for example the concentration of salmaterol 1 and/or formoterol 2 for about 1.0pM to about 10 μ M, for example about 0.01 μ M to about 10 μ M, about 0.1 μ M to about 5 μ M, 0.5 μ M extremely about 4 μ M or from about 0.001 μ M to any other concentration about 10 μ M.The lasting local organization concentration range of glucocorticoid can be about 0.01 μ M-10mM.

[0089] in some embodiments, one or more active component are aggregated thing and seal, combine with polymer and/or put together by being about 10-12% (quality) with the ratio of polymer microballoon.The amount this paper that accounts for the active component of carrier (for example microgranule or microsphere) mass percent is called " active component load (active ingredient loading) ".Term used herein " loading " and " load " are meant that active component is encapsulated in the carrier substantially, combines with carrier substantially and/or puts together.In some embodiments, the active component load is up to about 75%.Therefore, some preferred preparations comprise one or more beta-2 adrenergic active component, for example salmaterol 1, formoterol 2 and/or its physiologically acceptable salt and solvate are loaded on the polymer microballoon by every about 10 milligrams of active component to about 200 milligrams of about 1mg of polymer to about 20mg.In some embodiments, the preparation with this active component load is enough to by being suitable for producing steatolysis and/or the inhibiting concentration of lipogenesis provides active component to discharge about 15 days to about 45 days.Similarly, the glucocorticoid budesonide of pharmaceutically acceptable form and fluticasone can load to produce antiinflammatory action by the active component of every about 10mg to the about 1mg of about 200mg polymer to about 20mg.

[0090] in some embodiments, two or more active component are loaded in the same microgranule (for example liposome or PLGA).Therefore, some embodiments, the polymer of sealing glucocorticoid and adrenergic compounds is delivered to fatty tissue simultaneously.Perhaps, two kinds of active component are loaded on the independent microgranule.Then with two types microsphere mixing to obtain to have the preparation of required beta-receptor agonist and glucocorticoid ratio, while administration then.Perhaps, the sequential microgranule that gives two types.

[0091] microsphere that will comprise active component is suspended in the suitable physiologically acceptable liquid-carrier of about 0.5ml-10ml.In some embodiments, use independent active component microsphere, with this microsphere mixing in liquid-carrier.In other embodiments, every type microsphere mixes with liquid-carrier respectively.In some embodiments, liquid-carrier can contain the hyaluronidase of pharmaceutically acceptable form (1.0-15IU/ml) and pharmaceutically acceptable amount.The turbidity decline that the employed 1IU hyaluronidase of this paper produces is pressed identical that 1I.U. (iu) standard transparent matter acid enzyme preparation produces with hyaluronic acid and albumin mixture.Referring to (1966) such as for example Mathews, Methods Enzymol.8,654-662.In some embodiments, the microsphere suspensoid is carried out the socket of the eye injection or is expelled to retro-ocular space by the 0.1-0.5ml aliquot.Perhaps, prepare injection according to the method described above separately, and at same position two kinds of microball preparations of sealing each active component of sequential use.

[0092] in some embodiments, glucocorticoid, for example dexamethasone 6, budesonide 8 and/or fluticasone propionate 7 also are used as anti-inflammatory drug, therefore, alleviate by giving the inflammation that preparation polymer, polymerizing microballoons and/or the liposome of slow releasing preparation (for example by) is produced.

[0093] PLGA 15 microspheres are sealed hydrophobic compound and are more prone to than encapsulating hydrophilic chemical compound.In some embodiments, in order to increase the load of hydrophilic active composition, microsphere is with the unit modification of aforesaid Polyethylene Glycol.The microsphere of specific size does not absorb in the blood or not substantially and is eliminated by lymph, thereby supplies at needed position release of active ingredients.For example, in some embodiments, microsphere diameter is that about 20 μ m are to about 200 μ m.In some embodiments, the microsphere size also influences the release characteristics of active component in tissue.Generally speaking, bigger microsphere often provides long and uniform release characteristics.

[0094] in exemplary embodiment, slow releasing preparation comprise about 0.5 milligram to about 7.5mg (for example about 0.7mg, 1mg, 1.5mg, 2.0mg, 2.5mg, 3mg, 3.5mg, 4mg, 4.5mg, 5mg, 5.5mg, 6mg, 6.5mg, 7mg or from about 0.5mg to the 7.5mg any other the amount) salmaterol 1 and/or formoterol 2, and about 1.5mg is to about 7.5mg (for example about 2mg, 2.5mg, 3mg, 3.5mg, 4mg, 4.5mg, 5mg, 5.5mg, 6mg, 6.5mg, 7mg or about 1.5mg between about 7.5mg any other measure) dexamethasone 6, fluticasone propionate 7 and/or budesonide 8, described active component are by about 70 lactides: the ratio of 30 Acetic acid, hydroxy-, bimol. cyclic esters is encapsulated in about 100 milligrams of polylactide Acetic acid, hydroxy-, bimol. cyclic esters (PLGA) 15 copolymer microspheres.The amount of every kind of active component depends on the natural law (for example about 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days or about 10 days) of needed controlled release/slow release in the slow releasing preparation.In some embodiments, copolymer ratios and active component seal send every about 1mg copolymer up to about every days 1.0 μ g (for example every day about 0.02 μ g, 0.04 μ g, 0.06 μ g, 0.07 μ g, 0.1 μ g, 0.2 μ g, 0.4 μ g, 0.5 μ g, 0.6 μ g, 0.8 μ g or from about 0.02 μ g to any other amount about 100.0 μ g) salmaterol 1 and/or up to about every day 0.5 μ g (for example every day about 0.02 μ g, 0.04 μ g, 0.06 μ g, 0.07 μ g, 0.08 μ g, 0.09 μ g, 0.1 μ g, 0.2 μ g, 0.3 μ g, 0.4 μ g or from about 0.02 μ g to about 0.5 μ g any other the amount) formoterol and up to every days 5 μ g (for example every day about 0.2 μ g, 0.4 μ g, 0.5 μ g, 0.7 μ g, 0.9 μ g, 1.0 μ g, 1.5 μ g, 2 μ g, 2.5 μ g, 3 μ g, 3.5 μ g, 4 μ g, 4.5 μ g or from about 0.2 μ g to about 5 μ g any other the amount) fluticasone and/or budesonide 6, reach about 30 days.In another exemplary, slow releasing preparation to be given comprises as methylprednisolone acetate in the crystal fine grain suspensoid and beta-2 adrenergic agonist.The beta-2 adrenergic agonist can be mixed with solution, perhaps also can be mixed with the crystal fine grain suspensoid.In some embodiments, slow releasing preparation also comprises solubility succinic acid methylprednisolone, thereby except that the continuous action of the methylprednisolone acetate of crystal form, also provides instant effect (due to rapid release).

[0095] in some embodiments, preceding in administration (for example injection), one or more β-2 agonist and methylprednisolone mixed to be provided.In other embodiments, one or more β-2 agonist and methylprednisolone were mixed in administration (for example injection) time.

[0096] in some embodiments, before giving β-2 agonist, send selected corticosteroid slow releasing preparation (for example combination of methylprednisolone crystallization suspensoid, methylprednisolone solution or crystallization suspensoid and solution) separately and reach about 7 days (for example at least 12 hours, 1 day, 2 days, 3 days, 4 days, 5 days, 6 days or any scope At All Other Times from about 12 hours to about 7 days), so that raise beta receptor.

[0097] in some embodiments, provide non-slow releasing preparation for curee to be treated.In some embodiments, non-slow releasing preparation is after single agent, provide the activity of one or more long-acting selectivity β-2 agonist to continue about 4 hours to about 24 hours, β-2 agonist activity of for example 6 hours, 8 hours, 10 hours, 12 hours, 16 hours, 18 hours, 21 hours or any other persistent period from about 4 hours to about 24 hours.

[0098] in other embodiments, for curee to be treated provides the non-slow releasing preparation that comprises fugitive selectivity β-2 agonist, the activity of said preparation continues to be no more than about 4 hours (for example about 3.5 hours, 3 hours, 2.5 hours, 2 hours, 1.5 hours, 1.3 hours, about 1 hour, 0.5 hour or be no more than any other persistent period between about 4 hours to about 0.5 hour).

[0099] in exemplary embodiment, non-slow releasing injection comprises about 100 μ g to about 250 μ g (105 μ g for example, 110 μ g, 125 μ g, 150 μ g, 175 μ g, 190 μ g, 200 μ g, 210 μ g, 225 μ g or from about 100 μ g to any other amount about 250 μ g) former times the naphthoic acid salmaterol and about 500 μ g to about 1000 μ g (600 μ g for example, 650 μ g, 700 μ g, 730 μ g, 740 μ g, 800 μ g, 825 μ g, 875 μ g, 900 μ g, 930 μ g, 950 μ g or from about 500 μ g to any other amount about 1000 μ g) fluticasone propionate, be formulated in volume up to about 10ml (0.3ml for example, 0.5ml, 0.7ml, 1.1ml, 1.5ml, 2ml, 2.5ml, 3ml, 3.5ml, 4ml, 5ml, 6ml, 7ml, 8ml, 9ml or from about 0.3ml to any other volume about 10ml) with through the compatible excipient of socket of the eye administration.Excipient concentration can maintain below 1% (for example 0.05%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.8% or from about 0.05% any other concentration to below 1%.

[00100] in some embodiments, preparation described herein adopts any suitable method to send through eye, for example as the part with drop or by being placed on the drug storehouse storage under the eyelid.

[00101] in other embodiments, if preparation to be sent comprises long-acting beta-2 agonist (for example formoterol 2, salmaterol 1 or bambuterol 3) and glucocorticoid, then local the use is suitable.In some embodiments, but the slow releasing preparation of dermal delivery comprises biodegradable as mentioned above biocompatibility active component-polymer formulations or Liposomal formulation.

[00102] in some embodiments, provided herein is the method that is used for the treatment of thyroid eye diseas, and this method comprises one or more immunosuppressants (for example rapamycin (rapamycin), sirolimus (sirolimus) or everolimus (everolimus)) by local delivery preparation changes into adipose cell to suppress fibroblast proliferation and preceding adipose cell.In some embodiments, give one or more immunosuppressants with slow releasing preparation (for example depot formulation), described slow releasing preparation comprises immunosuppressant in the crystal fine grain suspensoid.In other embodiments, slow releasing preparation comprises the polymer particle (as described herein) that is loaded with one or more immunosuppressants.The administration of slow release immunosuppressant preparation can be socket of the eye, socket of the eye week or retrobulbar injection.

[00103] in some embodiments, in one of previous formulations, comprise thyroxin, thereby raise the beta receptor number on beta receptor or the increase cell surface, and the α acceptor number on downward modulation α receptor or the minimizing cell surface.Thyroxine (T4) 18 and trilute (triiodothyronine) (T3) stereoisomer of 19 both and these hormones can be used for this purpose.Levothyroxine is thyroxinic stereoisomer, has long half life.These thyroxins can share with beta-agonists, and through socket of the eye, socket of the eye week or retrobulbar injection administration.One or more thyroxins also can share with the beta-agonists in the above-mentioned slow releasing preparation (for example polymer or liposome), to reduce administration frequency.T3 and T4 can share with the long-acting β2Ji Dongji of selectivity of the polymerizing microballoons of the polylactide that is used for injection and slow releasing agent or polylactide/Acetic acid, hydroxy-, bimol. cyclic ester.

[00104] in some embodiments, above-mentioned preparation also comprises one or more class flavin (flavinoid) (for example Quercetin and fisetin) of flavone and flavin ketone (flavinone) group, they play the cAMP phosphodiesterase inhibitor, thereby improve the beta-adrenergic signal transduction.

Embodiment

[00105] to be interpreted as be exemplary to following specific embodiment, with the remainder of any alternate manner restriction present disclosure.We think and need not to further describe that those skilled in the art just can farthest utilize the present invention according to description herein.All publications that this paper quoted all are attached to herein by reference.If mention URL or other this class identifier or network address, should be understood that then this class identifier can change, customizing messages on the Internet changes constantly, but can find the information that is equal to during searching for Internet.The relevant reference material in the Internet shows the availability and the public dissemination of this category information.

[00106] Embodiment 1: the rat fat cell of beta-agonists and glucocorticoid is external The steatolysis experiment

[00107] decompose in the experiment at the body external fat, carry out chemical oxidation with hydrogen peroxide after, by spectrophotometry, detect the glycerol in the cell culture medium.In 3 hours, glycerol is measured.Only being exposed to beta-agonists, only being exposed to glucocorticoid or be exposed to one or more snippets preincubate after date of the two combination, the steatolysis level in people's adipose cell of cultivating to be measured, more details see below.

Isolate preceding adipose cell and be differentiated to form adipose cell:

[00108] end user's subcutaneous fat cells in the steatolysis experiment.Collect fatty tissue with suction lipectomy method or lipectomy, adipose cell before the following separation.Simply, with the fatty tissue chopping, be rich in the shaking channel of oxygen (shaking chamber) (5%CO ₂75 times/minute) in, in the Krebs-Ringer bicarbonate buffer that contains 1% bovine serum albumin and 0.1% collagenase, hatched 1 hour in 37 ℃.With suspension by behind the 400 μ m nylon net filters, under 100g centrifugal 1 minute.After the preceding adipose cell washed twice in the supernatant, be inoculated on 96 orifice plates by the density of described cells/well.Preceding adipose cell was cultivated in keeping culture medium 7 days, made it to be divided into adipose cell.

Reagent

Lavation buffer solution (serum-free Krebs Ringer buffer (KRB); [Sigma, K4002-10X1L])-be kept under 4 ℃

Measure the buffer (KRB that contains 1%FBS; [FBS derives from Gibco, 26140-079])-be kept under 4 ℃

Keeping culture medium is kept under 4 ℃

Behind glycerol reagent A (Zen-Bio, RGTL-15 or RGTL-40)-reprovision, keep in Dark Place under 4 ℃

Glycerol mother solution (1M) is diluted in glycerol [Sigma G2025-500ML] in the lavation buffer solution (serum-free) and prepares-to be kept under-20 ℃.

The steatolysis experiment:

[00109] tested preceding 21 hours at steatolysis, discard the former culture medium in each hole, what be replaced by that 75 μ l contain suitable drug or DMSO (solvent) concentration keeps culture medium (referring to following experimental design part).Each trial drug or control treatment are used for 8 hole/groups (12 processed group of per 96 orifice plates).Steatolysis was tested preceding 3 hours, each hole lavation buffer solution washed twice (200 μ l/ washing), fill it up with the testing liquid or the contrast solution (75 μ l/ hole) that in measuring buffer, prepare, hatched then 3 hours, promptly up to determining the glycerol content of measuring in the buffer.For some groups, only add medicine and reach 3 hour incubation period (referring to following experimental group and matched group).Tested preceding 1 hour, and prepared 7 glycerol standard solution that scope is 200 μ M-3.125 μ M by in measuring buffer, carrying out serial dilution.

[00110] hatches the glycerol content of measuring in the buffer in each hole, back and be used as the steatolysis index, if glycerol increases the expression steatolysis.Measure box (RandoxLaboratory, United Kingdom) with commercially available glycerol and measure the glycerol level, compare to determine content by glycerol serial dilution standard curve (3 μ M-200 μ M) by colorimetric.Make the glycerol concentration in each hole be normalized to cell density.

Experimental design:

[00111] in each group (n=8) of following matched group or experimental group, is equivalent to derive from the glycerol measured value in 8 holes of 96 porocyte culture plates.

[00112] as shown in Figure 2, after hatching in 3 hours, the dose dependent increase that long-acting beta-2 adrenergic receptor agonists formoterol induced lipolysis decomposes reaches more than 6 times, for 10 ^-6The above concentration of M, steatolysis is than viewed big with isoproterenol.Equally, after hatching in 3 hours, the long-acting beta-2 adrenergic receptor agonists salmaterol also dose dependent that decomposes of induced lipolysis increases, though effect (the steatolysis increase is slightly larger than 2 times) is not as observed strong with formoterol.The inductive steatolysis of salmaterol is equal to or less than with the viewed steatolysis of isoproterenol.

[00113] as shown in Figure 3, after 3 hours, glucocorticoid budesonide induced lipolysis decomposes increases (the highest about 1.5 times) slightly, and this is than observed low with isoproterenol (about 2.5 times).By contrast, only hatch and in fact made external steatolysis be subjected to slight inhibition in 18 hours with budesonide.

[00114] adipose cell and salmaterol (10 ^-6M) hatch 18 hours reduction steatolysiss (Fig. 4).Similarly, caused steatolysis to reduce in 18 hours with the isoproterenol processing.Yet when salmaterol being mixed with budesonide when reaching 18 hour incubation period, observing steatolysis increases (Fig. 4).

[00115] according to these data, we conclude that in the adipose cell of cultivating formoterol and salmaterol be induced lipolysis decomposition effectively in 3 hours.Yet after using 18 hours, in fact salmaterol reduces steatolysis, and this is likely by due to receptor desensitization or the receptor down-regulated.In addition, in the presence of the glucocorticoid budesonide, even salmaterol also can decompose by induced lipolysis after 18 hours.Therefore, budesonide can be used to keep or recovers the beta-2 adrenergic agonist ability that induced lipolysis decomposes in over a long time, and this may be to have prevented due to the beta-2 adrenergic receptor downward modulation.

[00116] Embodiment 2: the lipogenesis of beta-agonists and glucocorticoid suppresses

[00117] non-limiting example of this class lipogenesis inhibition is as follows:

Cell culture:

(ATCC, Manassas is VA) by 4 * 10 with the 3T3-L1 pre-adipose cell lines ⁵Cell/T75ml culture bottle is inoculated in the improved Eagle culture medium of Dulbecco (DMEM), and culture medium contains 10% normal Ox blood serum and 1% penicillin/streptomycin antibiotic.With cell at 37 ℃, 5%CO ₂Under hatch.After 3 days, cell is peeled off, behind the counting, be suspended in again and contain 6 * 10 with trypsin ⁵In the 2ml culture medium of 24 orifice plates of cells/well.After 1-2 days, cell is prepared the beginning lipogenesis near converging.

The lipogenesis material:

Lipogenesis starts culture medium: DMEM/10% hyclone/0.5mM IBMX/1 μ M dexamethasone

Lipogenesis carries out culture medium: DMEM/10% hyclone/10 μ g/ml insulins

Lipogenesis is kept culture medium: the DMEM/10% hyclone

The negative control culture medium: the normal Ox blood serum of DMEM/10%

The lipogenesis scheme:

After from each hole, discarding the 1.8ml culture medium, in every hole, add the 2ml lipogenesis and start culture medium.With plate at 37 ℃, 5%CO ₂Under hatched 48 hours.After discarding the 2ml culture medium, in every hole, add the 2ml lipogenesis and carry out culture medium.With plate at 37 ℃, 5%CO ₂Under hatched 48 hours.After discarding the 2ml culture medium, in every hole, add the 2ml lipogenesis and keep culture medium.With plate at 37 ℃, 5%CO ₂Under hatched at least 48 hours.Born of the same parents' inner lipid drops in to be accumulated in the cell 5 days at least.

Experimental design:

Before lipogenesis, with adipose cell before the 3T3-L1 of different phase β2Ji Dongji and/or glucocorticoid pretreatment.Adding lipogenesis startup culture medium preceding 24 hours, following processing cell:

The 1st group: be untreated

The 2nd group: 10 ^-10The M salmaterol

The 3rd group: 10 ^-8The M salmaterol

The 4th group: 10 ^-6The M salmaterol

The 5th group: 10 ^-4The M salmaterol

The 6th group: 10 ^-10M salmaterol+10 ^-6The M budesonide

The 7th group: 10 ^-8M salmaterol+10 ^-6The M budesonide

The 8th group: 10 ^-6M salmaterol+10 ^-6The M budesonide

The 9th group: 10 ^-4M salmaterol+10 ^-6The M budesonide

The 10th group: 10 ^-6The M budesonide

The 11st group: 10 ^-10The M budesonide

The 12nd group: 10 ^-6M capsaicin (capsaicin), this is a kind of known lipogenesis inhibitor

Another set of cell was handled 24 hours by above-mentioned each group before the adding lipogenesis carries out culture medium, and another jacket cell was handled before lipogenesis is kept culture medium 24 hours.In a cover contrast, cell was handled 24 hours by above-mentioned each group before adding the negative control culture medium.Each 12 groups in other two cover, 1 cover replaces to long-acting beta 2 agonist formoterol with salmaterol, another set ofly replaces to fugitive β2Ji Dongji albuterol.

Born of the same parents' inner lipid manifests:

5 days harvestings after the adding lipogenesis is kept culture medium.After discarding cell culture medium, each plate phosphate buffered saline(PBS) (PBS) washed twice.0.5ml oil red O solution (0.36% oil red O/60% isopropyl alcohol) is added in every hole, plate was at room temperature hatched 15 minutes.After discarding staining solution, each hole is with 60% washed with isopropyl alcohol 3 times.Then dyed plate is taken pictures and/or scan to carry out visual analyzing.Lipid is dyed redness.

The lipid quantitative assay:

The 0.25ml dyestuff is extracted solution (CHEMICON International) to be added in the hole of respectively dyeing.Plate was placed on rail mounted agitator (orbital shaker) or the shaking machine 15-30 minute.To contain the solution that extracts dyestuff and transfer in the cuvette, under 520nm, read absorbance with spectrophotometer.

[00118] Embodiment 3: β-2 agonist and glucocorticoid combination alleviate epididymal adipose tissues The pad quality

[00119] we attempt to determine whether glucocorticoid can reduce fat in vivo by the mode with external steatolysis data consistent described in the embodiment 1.For this reason, we only measured with the treatment of long-acting beta-2 2-adrenergic agonist components formoterol and with the rat epididymis fat pad quality of budesonide therapeutic alliance.

[00120] use Matrx 3000 carburator in 4% isoflurane (isoflurane) with the (～500g) anesthesia of male Sprague Dawley rat.Then, give as the injection 0.4ml solvent (2%PEG) in 5mm place before the fat pad rear end of listed animal in the following table 2; Formoterol in the solvent (3.48 μ g/ml; Dosage=1.39 μ g); Or the formoterol in the solvent (3.48 μ g/ml) adds budesonide (10 μ g/ml; Dosage=1.39 μ g formoterols, 4 μ g budesonides).Every animal is subjected to Drug therapy at a side joint, is subjected to solvent (2%PEG) treatment at a relative side joint; With regard to medicine and solvent, make and respectively organize left and right sides equilibrium (referring to table 2).

[00121] duplicate injection after 24 hours and 48 hours, totally 3 injections.Injected the last time back 24 hours, and made animal euthanasia, from every animal, win left and right sides epididymal adipose tissues pad, weigh (the results are shown in Table 3 and Fig. 6) through the excessive pentobarbital of peritoneal injection (150mg/kg).Use paired t-test and standard t check carrying out statistical analysis.

[00122] only formoterol shown in the his-and-hers watches 3 and formoterol+budesonide treatment data have been carried out this figure of pairing and are stepped on t check (paired Student t-test) and analyze, and the results are shown in Table 4.

[00123] as shown in table 4, the 1st treated animal (only treating) with formoterol show difference and variation with treat naturally contrast not consistent.Only the mean clinical value of formoterol is+0.028g ± 0.140g.The p value that statistical analysis obtains is 0.63, and is consistent with the trend of not having the trend therapeutic effect.On the other hand, the 2nd treated animal (formoterol+budesonide treatment) shows therapeutic effect.Mean clinical value is-0.353 ± 0.270, p value=0.079.

[00124] we have also carried out statistical analysis (this figure steps on the t check) to the significant difference between single medicine treatment and the combination medicine therapeutic effect, also the therapeutic alliance and the significant difference of not treating between control animal are analyzed.These statistic analysis result see Table 5.

[00125] as shown in table 5 and Fig. 6, significant difference (p=0.05) is arranged between the 2nd group and the 1st group.The 2nd group alleviates and significant difference is arranged, p=0.013 not treating in the animal average fat pad quality.

[00126] in tracking test, we add the ability that budesonide alleviates the fat pad quality to formoterol in the above-mentioned experiment and study.As shown in Figure 7, unite and give dosage 1.4 μ g/ days formoterols+4.0 μ g/ days budesonides, in the treatment group treatment is not organized and caused significant difference (p=0.01) aspect the average epididymis mass discrepancy.Equally, even making up under (0.7 μ g/ days formoterol+2.0 μ g/ days budesonides, administration every other day) than low dosage, therapeutic alliance animal and the difference of not treating between the control animal have significance (p=0.04).

[00127] according to these data and analysis, we infer that long-acting beta-2 agonist (for example formoterol) and glucocorticoid (for example budesonide) combination are effective to induced lipolysis decomposition and fatty minimizing the in the body probably.

[00128] Embodiment 4: with the combination treatment that comprises beta-agonists and glucocorticoid The clinical experiment of graves' ophthalmopathy

The non-limiting example of this class clinical trial that is used for the treatment of graves' ophthalmopathy is as follows:

The patient selects:

The patient does not have allergy to institute to medicine over more than 18 years old or 18 years old.Be diagnosed as the exophthalmos symptom patient relevant by ultrasonic examination, ophthalmostatometry, MRI or computer body-layer scanning with graves' ophthalmopathy.Specifically, select to suffer from outstanding or outstanding 0.5mm of double eyeball of simple eye ball or 3mm or more than, the patient of other symptom that occurs together or do not occur together.It is low that the patient also has limited and movable obviously limited, the corneal ulcer of diplopia, extreme position eye activity, pain, appearance deformity (cosmetic deformity) and quality of life.The patient may carry out the thyroidectomy of treatment hyperthyroidism.Other Steroid treatment should not be used for treating hyperthyroidism.All researchs are all carried out after acquisition Ethics Committee of mechanism (institutional ethics committee) approval and patient's agreement.

Research design:

Test 1: this test is the polycentric dose escalation study of salmaterol (a kind of long-acting β2Ji Dongji) and budesonide (a kind of glucocorticoid) conjoint therapy.The patient accepts the parenteral pharmaceutical composition that injection gives every day.After the treatment of 1 week, partially or completely reaction perhaps appears in the not improved patient of exophthalmos but the patient of stable disease will accept the therapy in extra 1 week, and dosage is higher than initial selected dosage.The composition of medicine that patient's acceptable dose of 3-6 name cohort increases progressively is up to definite maximum tolerated dose (MTD).The definition of MTD is to have among 3 patients among 2 or 6 patients 2 dosage before the experience dose limitation toxicity are arranged.

Test 2: this test is a multicenter study at random.Research lasts 60 days.The patient is assigned randomly to one of 18 treatment groups.For the 1st group, only give patient's salmaterol or formoterol every day by MTD.For the 2nd group, only give patient's budesonide every day by MTD.For the 3rd group, give patient's salmaterol and budesonide simultaneously by MTD.For the 4th group, give patient's salmaterol or formoterol every day, and every other day give budesonide.For the 5th group, give patient's budesonide every day, and every other day give salmaterol or formoterol.For the 6th group, whenever the odd number sky gives the patient budesonide, whenever the even number sky gives salmaterol.The dosage of 7-12 group is identical with the 1-6 group, and just dosage is 1/4MTD.Same identical with 1-6 of the dosage of 13-18 group, be dosage be 1/10MTD.Except that the treatment group, matched group is untreated.

Terminal point is estimated:

When research finished, the minimizing that patient's exophthalmos is alleviated and extraocular muscles long-pending with corpus adiposum orbitae was estimated.Also estimated the improvement of catacleisis and eye movement.Suppose and in 60 days, reduce by 20% positive result.

[00129] Embodiment 5: pharmaceutical composition

The non-limiting example of this class pharmaceutical composition is as follows:

The parenteral compositions

Embodiment 5AIn order to prepare the parenteral pharmaceutical composition that is applicable to drug administration by injection, water soluble salt and about 3mg ketotifen of about 100 μ g formoterols is dissolved in DMSO, mix with 10ml 0.9% Sterile Saline then.Mixture packed into be suitable in the unit dosage forms of drug administration by injection.

Embodiment 5BIn order to prepare the parenteral pharmaceutical composition that is applicable to drug administration by injection, water soluble salt and about 100 μ g fluticasone propionates of about 50 μ g salmaterol are dissolved in DMSO, mix with 0.9% Sterile Saline that 10ml contains 20% (volume/volume) PEG-400 that has an appointment then.With hyaluronidase be added in the mixture to final concentration be 8IU/ml.The gained mixture packed into be suitable in the unit dosage forms of drug administration by injection.

Embodiment 5CIn order to prepare the parenteral pharmaceutical composition that is applicable to drug administration by injection, the water soluble salt of about 50-100 μ g salmaterol is dissolved in DMSO, mix with 0.9% Sterile Saline that 10ml contains 20% (volume/volume) PEG-400 that has an appointment then.Mixture packed into be suitable in the unit dosage forms of drug administration by injection.

Embodiment 5DIn order to prepare the parenteral pharmaceutical composition that is applicable to drug administration by injection, about 50 μ g fluticasone propionate water soluble salts are dissolved in DMSO, mix with 10ml 0.9% Sterile Saline then.It is 10IU/ml that hyaluronidase is added to mixture to final concentration.The gained mixture packed into be suitable in the unit dosage forms of drug administration by injection.

The topical gel compositions

Implement 5EIn order to prepare the topical gel pharmaceutical composition, about 100mg salmaterol is mixed with 1.75g hydroxypropyl cellulose, 10ml propylene glycol, 10ml isopropyl myristate and 100ml straight alcohol USP with about 100mg prednisolone.Then the gained gel mixture is packed into and be suitable for the container of topical, for example pipe.

Embodiment 5FIn order to prepare the topical gel pharmaceutical composition, about 100mg formoterol is mixed with about 10mg hyaluronidase, 1.75g hydroxypropyl cellulose, 10ml propylene glycol, 10ml isopropyl myristate and 100ml straight alcohol USP with about 100mg budesonide.Then the gained gel mixture is packed into and be suitable for the container of topical, for example pipe.

Embodiment 5GIn order to prepare the topical gel pharmaceutical composition, about 100mg salmaterol is mixed with about 10ml PEG-400,1.75g hydroxypropyl cellulose, 10ml isopropyl myristate and 100ml straight alcohol USP.Then the gained gel mixture is packed into and be suitable for the container of topical, for example pipe.

Embodiment 5HIn order to prepare the topical gel pharmaceutical composition, about 100mg prednisolone is mixed with about 10ml PEG-400,1.75g hydroxypropyl cellulose, 10ml isopropyl myristate and 100ml straight alcohol USP.Then the gained gel mixture is packed into and be suitable for the container of topical, for example pipe.

The ophthalmic solution compositions

Embodiment 5IIn order to prepare the medicament for the eyes liquid composite, with the mixture of about 100mg salmaterol and about 100mg budesonide with after 0.9g NaCl/100ml pure water mixes, with 0.2 micron filter filtration.Then the gained isosmotic solution is installed to the medicament for the eyes unit of sending that is suitable for administration through eye, for example eye drop container.

Embodiment 5JIn order to prepare the medicament for the eyes liquid composite,, filter with 0.2 micron filter with the mixture of about 100mg formoterol and about 100mg budesonide and the pure aqueous solution of 0.9g NaCl/100ml about 10% (volume/volume) PEG-400.Then the gained isosmotic solution is installed to the medicament for the eyes unit of sending that is suitable for administration through eye, for example eye drop container.

Embodiment 5KIn order to prepare the medicament for the eyes liquid composite, will about 100mg formoterol and hyaluronidase (to final concentration be 10IU/ml) mixture and the pure aqueous solution of 0.9g NaCl/100ml about 10% (volume/volume) PEG-400, filter with 0.2 micron filter.Then the gained isosmotic solution is installed to the medicament for the eyes unit of sending that is suitable for administration through eye, for example eye drop container.

Embodiment 5LIn order to prepare the medicament for the eyes liquid composite, will about 100mg ketotifen chemical compound and the pure aqueous solution of 0.9g NaCl/100ml about 20% (volume/volume) PEG-400, filter with 0.2 micron filter.Then the gained isosmotic solution is installed to the medicament for the eyes unit of sending that is suitable for administration through eye, for example eye drop container.

Orally administered composition

Embodiment 5MIn order to prepare combination of oral medication, about 100mg prednisolone chemical compound mixes with 750mg starch.Mixture packed into be suitable for the oral dosage units of oral administration, for example hard gelatin capsule.

Embodiment 5NIn order to prepare combination of oral medication, about 50mg budesonide chemical compound is mixed with the 375mg gelatin.Mixture packed into be suitable for the oral dosage units of oral administration, for example hard gelatin capsule.

Embodiment 5OIn order to prepare combination of oral medication, about 200mg ketotifen chemical compound mixes with the 1500mg hydroxypropyl emthylcellulose.Mixture packed into be suitable for the oral dosage units of oral administration, for example hard gelatin capsule.

Embodiment 5PIn order to prepare combination of oral medication, about 50mg fluticasone propionate chemical compound is mixed with 600mg starch.Mixture packed into be suitable for the oral dosage units of oral administration, for example hard gelatin capsule.

[00130] Embodiment 6: beta-agonists and glucocorticoid dosage regimen

The non-limiting example of this class dosage regimen is as follows:

Embodiment 6AFirst day and subsequently whenever treatment odd number sky, suffers from the compositions 5C of the patient treatment effective dose of graves' ophthalmopathy.In the even number sky, give the compositions 5D of patient treatment effective dose.

Embodiment 6BFirst day and and subsequently whenever treatment odd number sky, suffer from the compositions 5D of the patient treatment effective dose of graves' ophthalmopathy.In the even number sky, give the compositions 5C of patient treatment effective dose.

Embodiment 6CThe compositions 5C that suffers from the patient treatment effective dose of graves' ophthalmopathy every day.In the even number sky, give the compositions 5D of patient treatment effective dose.

Embodiment 6DThe compositions 5D that suffers from the patient treatment effective dose of graves' ophthalmopathy every day.In the even number sky, give the compositions 5C of patient treatment effective dose.

Embodiment 6EThe compositions 5C that suffered from the patient treatment effective dose of graves' ophthalmopathy in first day then had a rest two days.Second day off, give the compositions 5D of patient treatment effective dose.Repeat this dose regimen then.

Embodiment 6FThe compositions 5D that suffered from the patient treatment effective dose of graves' ophthalmopathy in first day then had a rest two days.Second day off, give the compositions 5C of patient treatment effective dose.Repeat this dose regimen then.

Embodiment 6GFirst day and subsequently whenever treatment odd number sky, suffers from the compositions 5M of the patient treatment effective dose of graves' ophthalmopathy.In the even number sky, give the compositions 5C of patient treatment effective dose.

Embodiment 6HThe compositions 5C that suffers from the patient treatment effective dose of graves' ophthalmopathy every day.In the even number sky, give the compositions 5M of patient treatment effective dose.

[00131] should be understood that embodiment described herein and embodiment only are used for illustration purpose, various modifications on its basis or change all fall in the application's the spirit and scope, and fall in the scope of appended claims.The publication that all this paper quoted, patent and patent application all are attached to by reference and are used for all purposes herein.

Claims

1. lipopectic method of socket of the eye that is used to reduce the patient that needs are arranged, this method comprise and give described patient:

(a) at least a Beta-3 adrenergic agonist of treatment effective dose; With

(b) at least a chemical compound that is used to reduce the Beta-3 adrenergic receptor desensitization of treatment effective dose.

2. the process of claim 1 wherein that described patient suffers from thyroid eye diseas.

3. the process of claim 1 wherein described in parenteral, oral, ophthalmic, socket of the eye, behind eye, socket of the eye week, eyeball, in the cone, part, intramuscular, percutaneous, Sublingual, intranasal or respiratory tract carry out.

4. the process of claim 1 wherein and before giving at least a Beta-3 adrenergic agonist, give described at least a chemical compound.

5. the method for claim 4 wherein gave described at least a chemical compound in about 3 days to about 7 days before giving at least a Beta-3 adrenergic agonist.

6. the method for claim 5, wherein said at least a chemical compound route of administration in ophthalmic, socket of the eye, behind eye, socket of the eye week, eyeball or in the cone gives.

7. the method for claim 6, wherein said at least a chemical compound gives with crystal fine grain suspensoid form.

8. the method for claim 4, wherein said at least a chemical compound oral administration gives, and described at least a Beta-3 adrenergic agonist gives through eye.

9. the method for claim 8, wherein said at least a Beta-3 adrenergic agonist gives with the crystal fine grain suspensoid.

10. the process of claim 1 wherein that described at least a Beta-3 adrenergic agonist comprises the long-acting beta 2-adrenergic agonist components.

11. the process of claim 1 wherein that described at least a Beta-3 adrenergic agonist comprises beta-2 adrenergic receptor Beta-3 adrenergic agonist selectively.

12. the process of claim 1 wherein that described at least a Beta-3 adrenergic agonist comprises salmaterol, formoterol or its any combination.

13. the method for claim 12, wherein said at least a Beta-3 adrenergic agonist comprises salmaterol, and the salmaterol of treatment effective dose is about 0.01 μ g/ days to about 100 μ g/ days.

14. the method for claim 13, wherein said at least a Beta-3 adrenergic agonist comprises formoterol, and the formoterol of treatment effective dose is about 0.001 μ g/ days to about 50 μ g/ days.

15. the process of claim 1 wherein that described at least a chemical compound comprises glucocorticoid, antihistaminic or its any combination.

16. the process of claim 1 wherein that described at least a chemical compound comprises dexamethasone, prednisolone, methylprednisolone, fluticasone propionate, budesonide, ketotifen or its any combination.

17. the method for claim 6, wherein said at least a chemical compound is a glucocorticoid, and described at least a long-acting beta 2-adrenergic agonist components is salmaterol, formoterol or its combination.

18. the method for claim 1, this method also are included in the immunosuppressant that the approach in ophthalmic, socket of the eye, behind eye, socket of the eye week, eyeball or in the cone of giving before the described at least a chemical compound is treated effective dose.

19. the method for claim 18, the immunosuppressant of wherein said treatment effective dose gives with the form of crystal fine grain suspensoid.

20. one kind is used for the treatment of pop-eyed method, this method comprises that the patient that needs are arranged comprises the compositions of at least a Beta-3 adrenergic agonist for the treatment of effective dose.

21. the method for claim 20, wherein said at least a Beta-3 adrenergic agonist comprises the long-acting beta 2-adrenergic agonist components.

22. the method for claim 20, wherein said at least a Beta-3 adrenergic agonist comprise beta-2 adrenergic receptor Beta-3 adrenergic agonist selectively.

23. the method for claim 20, wherein said at least a Beta-3 adrenergic agonist comprises salmaterol, formoterol, bambuterol, eformoterol, isoproterenol, albuterol or fenoterol.

24. the method for claim 20, wherein said compositions comprise the mixture of at least a long-acting beta 2-adrenergic agonist components and at least a fugitive Beta-3 adrenergic agonist.

25. the method for claim 23, wherein said compositions comprises salmaterol, and the salmaterol of treatment effective dose is about 0.01 μ g/ days to about 100 μ g/ days.

26. the method for claim 23, wherein said compositions comprises formoterol, and the formoterol of treatment effective dose is about 0.001 μ g/ days to about 50 μ g/ days.

27. the method for claim 23, wherein said in parenteral, oral, ophthalmic, socket of the eye, in the cone, behind eye, eyeball, part, intramuscular, percutaneous, Sublingual, intranasal or respiratory tract carry out.

28. the method for claim 20, wherein said compositions also comprises the hyaluronidase for the treatment of effective dose.

29. comprising, the lipopectic method of socket of the eye that is used to reduce the patient that needs are arranged, this method give described patient:

(a) one or more adrenoreceptor pathway stimulation chemical compounds of treatment effective dose; With

30. the method for claim 29, wherein said one or more adrenoreceptor pathway stimulation chemical compounds comprise catecholamine, alpha-1 adrenergic antagonists, forskolin, aminophylline or its analog.

31. a medical composite for eye, described compositions comprise the methylprednisolone acetate or the fluticasone propionate of the crystal fine grain suspensoid form of acceptable excipient of ophthalmology and treatment effective dose.

32. the medical composite for eye of claim 31, described compositions also comprise solubilising methylprednisolone acetate or solubilising fluticasone propionate.

33. the medical composite for eye of claim 33, described compositions also comprise at least a long-acting beta-2 agonist that is crystal fine grain suspensoid form for the treatment of effective dose.

34. a medical composite for eye, described compositions comprise acceptable excipient of ophthalmology and at least a long-acting beta-2 agonist that is crystal fine grain suspensoid form of treatment effective dose.

35. the medical composite for eye of claim 34, wherein said at least a long-acting beta-2 agonist comprises salmaterol or formoterol.

36. the medical composite for eye of claim 34, described compositions also comprise at least a solubilising long-acting beta-2 agonist for the treatment of effective dose.

37. the medical composite for eye of claim 34, described compositions also comprise at least a chemical compound that is used to reduce the crystal fine grain suspensoid form of Beta-3 adrenergic receptor desensitization of treatment effective dose.

38. at least a Beta-3 adrenergic agonist and at least aly be used for reducing target tissue is used for the treatment of the medicine of the disease that comprises the socket of the eye lipopexia in preparation to the chemical compound of Beta-3 adrenergic agonist desensitization purposes.

39. at least a Beta-3 adrenergic agonist and at least aly be used for reducing target tissue to the chemical compound of Beta-3 adrenergic agonist desensitization purposes in the method that is used for the treatment of the disease that comprises the socket of the eye lipopexia.