CN106580956B - Application of the Suo Lafen A in ischemic brain damage - Google Patents

Abstract

The present invention provides application of the Suo Lafen A in ischemic brain damage.Specifically, the present invention provides Suo Lafen A in the pharmaceutical composition for being used to prepare (i) prevention and/or treatment ischemic brain damage；And/or the purposes in the pharmaceutical composition of (ii) protection ischaemic neuronal.It is demonstrated experimentally that Suo Lafen A can effectively control the progress of acute apoplexy, reduce infarct size, slow down neuron and angioedema, and also plays positive effect in long-term nervous function protection.

Description

Application of the Suo Lafen A in ischemic brain damage

Technical field

The present invention relates to field of pharmaceutical biology.In particular it relates to which antineoplastic Suo Lafen A is to ischemic brain damage In new application.

Background technique

Nearest national coroner's inquest shows ischemic brain damage (cerebral ischemic injury) or ischemic Cerebral apoplexy (ischemic stroke) has leapt to disables and first of the cause of death for China human mortality, and morbidity has and increases year by year More trend.Acute ischemic cerebral apoplexy is the most common stroke types, accounts for the 60%~80% of whole cerebral apoplexies.But mesh It is preceding to there is the drug of definite therapeutic effect only there was only rt-PA acute apoplexy (recombinant tissue-plasminogen activator, rt-PA) is a kind of.Rt-PA be also current U.S.'s food with Unique drug of drug administration (US FDA) approval treatment acute apoplexy.But the therapeutic time window of rt-PA is extremely limited. In order to avoid the treatment-related Complications of Cerebral Hemorrhage of rt-PA occurs, it is presently recommended that it is after the onset of acute apoplexy in 4.5 hours Using.Thus only about 3% acute stroke patients below just have an opportunity to be treated using rt-PA.

Therefore, researching and developing new safely and effectively Treatment of Cerebral Stroke strategy seems more urgent.

Summary of the invention

The present invention provides the new applications of antineoplastic Suo Lafen-A a kind of, i.e., it is used to mitigate cerebral arterial thrombosis damage Therapeutic strategy.

First aspect present invention provides a kind of Suo Lafen-A (Soraphen-A) or the use of its pharmaceutically acceptable salt On the way, it is used to prepare the pharmaceutical composition of (i) prevention and/or treatment ischemic brain damage；And/or (ii) protects ischaemic neuronal Pharmaceutical composition, wherein the Suo Lafen-A have Formulas I shown in structure:

In another preferred example, the ischemic brain damage includes Acute ischemia rePerfusion or chronic ischemic brain damage Wound.

In another preferred example, the Acute ischemia rePerfusion include disease time be -3 days 2 hours between ischemic Property cerebral injury.

In another preferred example, the chronic ischemic cerebral injury includes the ischemic brain damage after disease time 3 days.

In another preferred example, the ischemic brain damage includes cerebral apoplexy, ischemia reperfusion injury, transience brain Ischemic episode, neonatal hypoxic ischemic encephalopathy.

In another preferred example, the prevention and/or treatment ischemic brain damage include one or more of to ischemic Property cerebral injury performance improvement:

(a) Brain stem injury is reduced, Infarction volume and/or area are such as reduced；

(b) improve limbs symmetry；

(c) improve limbs and use Preference；

(d) improve neural sensation and motor function damage；

(e) increase infarct location brain blood flow；

(f) remember after improving cerebral injury；

(g) reduce cerebral ischemia after neuron loss.

In another preferred example, the administration dosage of the Suo Lafen-A is 1-100mg/kg/ days.

In another preferred example, the administration dosage of the Suo Lafen-A is preferably, daily 1-50mg/kg, more preferably for 1-5mg/kg。

In another preferred example, the pharmaceutical composition contains Suo Lafen-A or its pharmaceutically acceptable salt as work Property ingredient and pharmaceutically acceptable carrier.

In another preferred example, the pharmaceutical composition also contains thrombolytics, rtPA and/or neuroprotective agent.

In another preferred example, cerebral apoplexy Advance of Intravascular Stent in Treating medicine box is also contained in the pharmaceutical composition.

Second aspect of the present invention provides a kind of method of external non-therapeutic protection ischaemic neuronal, comprising steps of Ischaemic neuronal is cultivated under conditions of containing Suo Lafen-A or its pharmaceutically acceptable salt, and/or at through ischemic Suo Lafen-A is added in the neuronal cultures of reason, to protect ischaemic neuronal.

In another preferred example, the protection includes preventive protection and/or therapeutic protection.

Third aspect present invention provides a kind of (i) prevention and/or treatment ischemic brain damage；And/or protection ischemic The method of neuron, including step, to required object application safe and effective amount Suo Lafen-A or its is pharmaceutically acceptable Salt, or contain the pharmaceutical composition of Suo Lafen-A or its pharmaceutically acceptable salt as active constituent.

In another preferred example, the required object includes the mammal with ischemic brain damage, such as small Mouse, rat or people.

It should be understood that above-mentioned each technical characteristic of the invention and having in below (eg embodiment) within the scope of the present invention It can be combined with each other between each technical characteristic of body description, to form a new or preferred technical solution.As space is limited, exist This no longer tires out one by one states.

Detailed description of the invention

Fig. 1 shows action site of the Suo Lafen-A in cellular process.

Fig. 2 is shown using the brain blood flow situation of change in laser speckle Doppler technology monitoring MCAO surgical procedure.Its In, after Fig. 2A-B shows the row sham-operation of Sham group mouse, Bilateral hemispheres brain blood flow is without significant changes；And in DMSO group and In SorA group mouse, the obstruction of arteria cerebri media causes the significant brain blood flow of side cerebral hemisphere and reduces, and brain blood flow Reduction degree is in this two groups of mouse and there was no significant difference；Fig. 2 C-D shows that the cerebral infarction volume through SorA treatment mouse is obvious Less than DMSO control mice.

Fig. 3 shows that the ratio that the cerebral ischemia mouse of Sor-A treatment turns to the left is lower than the ratio of DMSO group, represents The limbs symmetry of mouse is treated better than control mice.

Fig. 4 A-B shows that compared with DMSO mouse, cerebral ischemia mouse opposite side forelimb and hind leg mistake through Sor-A treatment walk The percentage of the total wrong step number of number Zhan significantly reduces, and approaches with sham group mouse.

Fig. 5 shows cerebral ischemia mouse after injury, and non-damaging side (right side) limbs preference use ratio significantly increases, The preference use ratio of Sor-A treatment mouse limb is substantially less than DMSO control group.

Fig. 6 A is limbs proprioception function score, and Fig. 6 B is climbing test scoring, and Fig. 6 C is forelimb walking experiment scoring, Fig. 6 D is that side turns experiment scoring, and Fig. 6 E is the comprehensive score of mouse, and Fig. 6 F is sticky paper experiment scoring；The above results show: Sor-A Treatment is remarkably improved nervous function comprehensive score after the chronic ischemic cerebral injury of mouse.

Fig. 7 A-B shows Morris water maze test, the results showed that, Sor-A significantly improves mouse in target quadrant Residence time, therefore can significantly improve learning and memory function；Fig. 7 C shows that the swimming rate of three groups of mouse is not significant Sex differernce.

Fig. 8 A-B shows that the neuron survived in the ischemic area that applied the mouse of Suo Lafen-A in advance is significantly more than DMSO control group mice.

Specific embodiment

The present inventor after extensive and in-depth study, for the first time it was unexpectedly observed that anti-tumor drug Suo Lafen-A for lack Brain after hemorrhagic cerebral injury has good neuroprotection.It is demonstrated experimentally that ischemic brain damage acute stage, chronic phase And ischemical reperfusion injury has preferable neuroprotection, occurs in advance using Suo Lafen-A or in ischemic brain damage Suo Lafen-A is used afterwards, can be effectively reduced ischemic focus area, be mitigated post-ischemic vascular oedema, and to long-lasting nerve function Repairing has very useful good effect.On this basis, the present invention is completed.

Suo Lafen A or its pharmaceutically acceptable salt

As used herein, term " inventive compound ", " Sora-A ", " Suo Lafen A " are used interchangeably, and refer to having There are the compound or its pharmaceutically acceptable salt of structure shown in Formulas I.

Suo Lafen-A (Soraphen-A) is that the one kind found in oncology and immunological investigation in recent years can inhibit high The compound of metabolizing cells growing multiplication specific can inhibit (the Acetyl of acetyl-CoA carboxylase -1 in cellular process Coenzyme (CoA) carboxylase-1, ACC-1) activity,

(Acetyl coenzyme (CoA) carboxylase can be catalyzed acetyl coenzyme A to acetyl-CoA carboxylase (acetyl CoA) is converted into malonyl coenzyme A (malonyl CoA).Malonyl coenzyme A can regulate and control the life of long chain fatty acids Object synthesis and degradation.There are two types of the ACC, i.e. ACC1 and ACC2 of hypotype in vivo.ACC1 is predominantly located in cytoplasm, and ACC2 is main Positioned at mitochondria.In cytoplasm by the malonyl coenzyme A that ACC1 is catalyzed and synthesized be fatty acid synthetase catalysis long-chain fat The important carbon donor of acid synthesis, action site of the Suo Lafen-A in cellular process are shown in Fig. 1.And in mitochondrial surface The malonyl coenzyme A that catalyzes and synthesizes of ACC2 can then inhibit carnitine acyl transferase I, so that regulating and controlling long chain fatty acids enters line Plastochondria carries out beta oxidation.

In recent years, more and more researchs are all expected to the target for the treatment of metabolic disease and tumour using ACC1 as one Molecule.Knocking out ACC1 by liver specific genes can be with recombining for effective for fat loss acid and tiring out for triglycerides Product, and fat specific knockdown ACC1 then reduces the sluggishness of Skeletal Muscle Growth.And in metastatic tumo(u)r, ACC1 or fatty acid The expression of synzyme is all increased.

The present invention provides a kind of new applications of Suo Lafen-A, i.e., for the cerebral protection after ischemic brain damage.

Suo Lafen-A of the present invention can also be to be used by pharmacological or physiological acceptable salt form derived from acid or alkali. These salt include but is not limited to the salt formed with following acid: hydrochloric acid, hydrobromic acid, sulfuric acid, citric acid, tartaric acid, phosphoric acid, cream Acid, pyruvic acid, acetic acid, succinic acid, oxalic acid, fumaric acid, maleic acid, oxaloacetic acid, methanesulfonic acid, ethanesulfonic acid, benzene sulfonic acid or hydroxyl second Sulfonic acid.Other salt include: the salt formed with alkali or alkaline earth metal (such as sodium, potassium, calcium or magnesium), and with ester, carbamic acid The form of ester or other conventional " pro-drugs ".

Ischemic brain damage

As used herein, term " ischemic brain damage ", " ischemic cerebrovascular disease " are used interchangeably, and refer to local cerebral Tissue, denaturation, necrosis or the transient function occurred including nerve cell, spongiocyte and connection fiber due to blood supply disorder It loses.It is a kind of high lethal that clinic is common, crippling cranial vascular disease.

Usual ischemic brain damage can be due to causing ischaemia after local organization ischemic, postischemic reperfusion or bleeding Equal many reasons cause.

The ischemic brain damage that Suo Lafen-A of the present invention is applicable in is not particularly limited, and may include any local cerebral ischemia And with or without neurological dysfunction cerebral injury disease.Common ischemic brain damage includes: cerebral apoplexy, ischemic- Reperfusion injury, transient ischemic attack, neonatal hypoxic ischemic encephalopathy.

In ischemic brain damage, the destruction of the oedema, blood-brain barrier of most common pathological manifestations, that is, neuron, periphery is exempted from The brain tissue of epidemic disease cell infiltrates, the reconstruction of neural blood vessel unit.Therefore, the present invention also provides a kind of inhibition periphery immunocytes The method of maincenter infiltration.It is demonstrated experimentally that the periphery of the maincenter of acute phase of cerebral ischemia is exempted from after it applied Suo Lafen-A of the present invention Epidemic disease cellular infiltration significantly reduces, and blood-brain barrier disruption mitigates.From the point of view of the reparation of chronic phase, Suo Lafen-A can be by inhibiting nerve Inflammatory reaction plays the role of that neural blood vessel unit is promoted to rebuild.Periphery function of immune system can be caused after cerebral arterial thrombosis Disorder, show as immunocyte excessive activation in ischemic early stage, infiltration to brain tissue.There is immune system function in the phase after injury It can decline, cause infection-related complication.Suo Lafen-A can mitigate the immune of ischemic later period by the activation of regulation immunocyte Function inhibitio improves the prognosis of patients with cerebral apoplexy to reduce the infection-related complication of post-stroke.

Pharmaceutical composition and method of administration

The present invention also provides a kind of for treating the pharmaceutical composition of ischemic brain damage, the pharmaceutical composition In contain the Suo Lafen-A of safe and effective amount or its pharmaceutically acceptable salt and pharmaceutically acceptable carrier.In addition, this hair Drug, such as thrombolytics, neuroprotective agent etc. of the bright pharmaceutical composition also containing other treatment ischemic brain damage.

In general, these substances can be formulated in nontoxic, inert and pharmaceutically acceptable aqueous carrier medium, Middle pH is usually about 5~8, and preferably pH is about 6~8, although pH value can be with the property and disease to be treated for being formulated substance Disease and be varied.Prepared pharmaceutical composition can be administered by conventional route, including (but being not limited to): Oral, intramuscular, intravenous, subcutaneous, intradermal or local administration.

It, can be with one or more pharmaceutically acceptable carriers or figuration when Suo Lafen-A of the present invention is used for such use Agent mixing, such as solvent, diluent, preferably can be with sterile injectable solution or suspension form (containing about in isotonic medium 0.05-5% suspending agent) carry out parenteral routes.For example, these pharmaceutical preparations contain the about 0.01-99% mixed with carrier, The active constituent of more preferably about 0.1%-90% (weight).

The effective dose of active constituent used can be with the tight of compound used, the mode of administration and disease to be treated Weight degree and change, this usually can by those skilled in the art according to the actual situation and it is limited experiment be adjusted.So And usually when the compound of the present invention is given daily with the dosage of about 0.5-1000mg/kg the weight of animals, it can obtain making us full The effect of meaning is preferably given daily with 1-4 dosage.For most of large mammal, daily accumulated dose is about For 1-100mg/kg, it is preferably about 2-100mg/kg.This dosage is adjusted to provide optimal treatment response.For example, by An urgent demand of situation is treated, can daily be increased or decreased dosage pari passu.

The concentration or dosage of a kind of preferred active constituent are about daily 1-100mg/kg, preferably 1-50mg/kg, more It goodly is 1-5mg/kg.Preferably, the dosage can be according between mouse known in the art and human body weight and body surface area Formula convert.

The compounds of this invention or pharmaceutical composition preferably pass through the approach administration such as intravenous.Liquid carrier include: sterile water, Polyethylene glycol, propylene glycol, ethyl alcohol, dimethyl sulfoxide, nonionic surface active agent etc., if be suitble to active constituent characteristic and Required specific administration mode.Usually used adjuvant can also advantageously be included in preparation pharmaceutical composition, such as anti-corrosion Agent and antioxidant such as vitamin E, vitamin C, citric acid, BHT and BHA.

The medicament forms for being adapted to injection include: aseptic aqueous solution or dispersion liquid and aseptic powder (for extemporaneous preparation of sterile Inject solution or dispersion liquid).In all situations, these forms must be sterile and must be fluids to be easy to syringe row Fluid out.It must be stable under conditions of manufacture and storage, and must be able to prevent the pollution of microorganism (such as bacterium and fungi) It influences.Carrier can be solvent or decentralized medium, wherein containing as water, alcohol (such as glycerol, propylene glycol and liquid polyethylene glycol), They properly mix object and vegetable oil.

Clinical application

The present invention provides the new indications of Suo Lafen-A clinical application.Animal experiments show that in application Suo Lafen-in advance After A, the tolerance degree of cerebral ischemia is remarkably reinforced in mouse, is capable of the cerebral apoplexy severity of substantially reduced induction, and In the animal that cerebral apoplexy occurs, application Suo Lafen A can effectively control the progress of acute apoplexy, reduce infarct size, slow down Neuron and angioedema, and positive effect is also played in long-term nervous function protection.

Beneficial effect of the present invention

The present invention provides new application of the antineoplastic Suo Lafen-A in ischemic brain damage, can be used as preventative use Medicine can more be applied in acute attack stage and in chronic convalescence, extend the therapeutic time window of the ischemic brain damages such as cerebral apoplexy, It can substantially reduced cerebral apoplexy progress, improve the function of brain neuron, more favorable branch provided to patients with cerebral apoplexy prognosis It holds.

Present invention will be further explained below with reference to specific examples.It should be understood that these embodiments are merely to illustrate the present invention Rather than it limits the scope of the invention.In the following examples, the experimental methods for specific conditions are not specified, usually according to conventional strip Part, such as Sambrook et al., molecular cloning: laboratory manual (New York:Cold Spring Harbor Laboratory Press, 1989) condition described in, or according to the normal condition proposed by manufacturer.Unless otherwise stated, no Then percentage and number are weight percent and parts by weight.

1 Suo Lafen-A of embodiment has the work of neuroprotection to Acute ischemia rePerfusion caused by middle cerebral artery occlusion With

Experimental method:

1) 8-12 weeks male C57/BL6 wild-type mice (being purchased from Shrek experimental animal company) is selected, is randomly divided into 5 Group, Sham group (n=8), DMSO group (n=8), SorA-1 group (n=8), SorA-5 group (n=8), Sor-A-25 group (n=8).

2) each group mouse is given respectively and establishes animal model, Sham group mouse row sham-operation, and separating blood vessel is not placed in line bolt. The SorA group mouse row middle cerebral artery occlusion of DMSO and three dosage group performs the operation (MCAO).MCAO surgical procedure is as follows: will After 1.5% isoflurane anesthesia of mouse, fixation of lying on the back.Exposure neck, median incision about 3cm, blunt separation, exposure tracheae are blunt Property separation two sides thyroid gland, exposure right side nutator and sternohyoideus between trigonum.Internal carotid is separated to drum Bubble, carefully separates and ligatures arteria pterygopalatina.On the right side of coagulation after external carotid artery distal end, is temporarily pressed from both sides with micro- artery clamp and close neck and always move Arteries and veins leads external carotid artery stump to outer lower side, is allowed to cut an osculum in external carotid artery stump in straight line with internal carotid, No. 8 line bolts that diameter is 0.23mm are inserted into internal carotid from external carotid artery, are softly slowly pushed ahead to the power that is hampered, by line bolt It is ligatured jointly with external carotid artery with fixing line bolt, unclamps artery clamp, sew up the incision.Arteria cerebri media opening, resistance are enclosed at this time Broken the blood flow of arteria cerebri media, and MCAO model is made.It blocks and exposes cervical incision after sixty minutes, slowly pull out Outlet bolt and realize again Perfusion.

The MCAO model of mouse can lead to the infarct of volume parietal cortex and caudate nucleus, and the death rate is low (< 10%), be suitable for growing The assessment of phase behavioral function.

In order to reduce physiologic variables to ischemic brain damage more after potential impact, during the experiment by rectal temperature and life Manage norm controlling in the normal range.Local cerebral blood flow during monitoring ischemic using Speckle laser-Doppler sxemiquantitative (rCBF).If cortex rCBF decline < 20%, will be removed outside during mouse MCAO.

3) bloodstream blocking after sixty minutes, pulls out line bolt, realizes that brain blood flow fills again.After filling 1 hour again, abdomen is given according to grouping Chamber administration: DMSO group mouse gives 200ulDMSO；SorA-1 group gives 10% that Suo Lafen-A1mg/kg is dissolved in 200ul In DMSO；SorA-5 group is given Suo Lafen-A 5mg/kg and is dissolved in 10% DMSO of 200ul；SorA-25 group gives Suo La Sweet smell-A 25mg/kg is dissolved in 10% DMSO of 200ul.

4) 1 day, 2 days and 3 days after performing the operation, the scoring of mouse Nerve functional defect is measured respectively, and standards of grading are as follows: being divided into 0- 5 points: 0 points are impassivity dysfunction；1 point is unable to full extension for right side fore paw；2 points are to turn-take to the right；3 points are to roll to the right ?；4 points are that cannot walk；5 points are death.

5) mouse is put to death in third day after performing the operation, and Application mouse brain section mold is from after caudate nucleus front to hippocampus after taking brain Tissue does continuous coronal slice between portion (with a thickness of 1mm).Brain piece is infiltrated on 2%TTC (2,3,5-chlorinated triphenyl base, four nitrogen Azoles).Since TTC is pyridine in respiratory chain-nucleotide structure enzyme system proton acceptor, with the dehydrogenase reaction in normal tissue And take on a red color, and dehydrogenase activity declines in ischemic tissue, cannot react, therefore variation will not be generated in pale.Therefore TTC dyeing The ischemic infraction of detection mammalian tissues can be used to dye.Paraformaldehyde after dyeing using 4% is fixed overnight, and second day It can be taken off shooting TTC stained photographs.According to the method for Swanson, it is based on ischemic side and non-ischemic Ce Wei infarction tissue, analysis is every The infarcted region of brain on piece is opened, can avoid infarcted region angioedema obscures effect.Infarction volume=infarct size summation × All brain piece thickness.

Experimental result:

It is small such as Fig. 2A-B using the brain blood flow situation of change in laser speckle Doppler technology monitoring MCAO surgical procedure After mouse row sham-operation, Bilateral hemispheres brain blood flow is without significant changes；In DMSO group and SorA group mouse, arteria cerebri media Obstruction cause the significant brain blood flow of side cerebral hemisphere and reduce, and the reduction degree of brain blood flow in this two groups of mouse simultaneously There was no significant difference (Fig. 2 B).By TTC dye method, calculate the volume of ischemic tissue of brain, and it was found that through SorA (1, 5,25) cerebral infarction volume of the mouse for the treatment of be significantly less than DMSO control mice (Fig. 2 C-D).Therefore, it is demonstrate,proved by this experiment Bright, SorA has protective effect to Acute ischemia rePerfusion.

2 Suo Lafen-A of embodiment can improve long-term nervous function after ischemic brain damage

This experiment uses a series of effective, sensitive, the reproducible behaviouristics detection method of rodents to assess Neurological dysfunction after ischemic brain damage.All appraisal procedures MCAO operation after or sham-operation after 1 week beginning weekly into Row test.

1) Corner is tested: the sensorimotor function that Corner tests after capable of detecting MCAO to 90 days damages.It will experiment Animal is placed in the clamping plate entrance of 30 ゜, it is made voluntarily to advance to corner.It, can be leftward or rightward when mouse advances to angle head end It turns to, then mouse is taken out and is replaced in clamping plate entrance, repeat above step.Each mouse gives 10 advance turning machines Meeting records the evaluation index that the number turned to the left is tested as corner.

2) wrong step test: for evaluating forelimb and hind leg function.Mouse is placed in 30 (length) × 35 (width) × 31 (height) cm Grid on (grid 2.5cm²).Mouse steps on steel wire and moves on grid.Claw, which falls or makes a misstep, is just denoted as primary wrong step. Each animal testing 1min, totally 3 times, every minor tick 1min.Calculate the percentage of the total wrong step number of contralateral limbs mistake step number Zhan.

3) cylinder test (Cylinder Test): cylinder test can assess long-term feeling and motor impairment and Limbs symmetry was detected at mouse postoperative 3 days, 5 days, 7 days, 10 days, 14 days, 21 days and 28 days in this research.Specifically It is as follows: experimental animal being put into diameter 8cm, in the transparent plastic drum of high 15cm, 2-4 animal is placed every time, guarantees every A mouse is placed in cylinder.It records a video 5 minutes after marking.After completing all video recording work, by one to experimental group not Informed onlooker person contacts the number progress technology of cylindrical wall to upper limb in video recording mouse 5 minutes upwards.Final evaluation index Be expressed as non-damaging side (right side) limbs preference use ratio, i.e., (right side-left side)/(right side+left side+bilateral) frequency of exposure × 100。

4) neural sensation and motor function are assessed:

A) limbs feel function score: contacting every side limbs of mouse with a blunt stick, and observe it to the anti-of stimulation It answers.Standards of grading are as follows: 3, mouse is equally sensitive to the stimulation of two sides, and has the reaction of rotary head；2, bilateral habituation or right Side reaction weakens；1, mouse is blunt to the stimulate the reaction in left side；With 0, mouse is reactionless to the stimulation in left side.

B) climbing experiment: mouse is placed on wire mesh cage.Under normal conditions, mouse climbs up wire mesh cage with four all legs Metope.After mouse removes from wire mesh cage, record mouse catches the muscular strength of wire mesh cage.Standards of grading are as follows: 3, mouse is light Ground is climbed up, and wire netting is held on to；2, left side is impaired, and left side muscular strength is weaker than right side；1, mouse can not climb or pirouette Circle.

C) forelimb walking experiment: lift the tail of mouse, so that its hind leg is left ground, only walked with forelimb.3, trippingly It goes ahead and symmetrical；2, it offsets to one side；1, it turn-takes；0, forelimb movements cannot.

D) side turns experiment: mouse, which is raised, turns ability from 10 centimetres of desktop or more in the sky to observe the side of mouse, scoring Standard is as follows: 3. turn to bilateral equality side, and 45 degree of side gyration >；2. turning to bilateral equality side, but side gyration < 45 degree； 1, unequal side turns；0, turn without side.

E) overall neurological function score: it will score above and be added and obtain.

F) sticky paper is tested: being sticked sticky paper in the centre of the palm of the forelimb of mouse, and is measured mouse and feel after the presence of sticky paper it Any location contacts of limbs remove the required time to the time of sticky paper and by sticky paper.If mouse can not still remove in 120s Sticky paper, then recording and removing the maximum duration of sticky paper is 120s.

5) Morris water maze test assesses the obstacle of long-term cognitive function: the impaired learning and memory of detection MCAO mouse Function: mouse learns to find hiding platform in open Morris water maze to flee from the swimming task being forced.It is postoperative Mouse was placed in 13 days in the water maze of no platform and it is allowed to adapt to, altogether twice.Hereafter, the 2nd, 4,6 and 8 week after surgery daily row water Labyrinth test, 4 times a day.Record the required time (incubation period) that animal reaches submerged platform.When finally training twice, remove flat Platform allows mouse arbitrarily to swim 30 seconds, and records mouse and mutually limit (the mutually limit that platform was once placed) interior residence time and swimming in target Speed.

As a result

1) Corner test: in 10 advance turning machine meetings of mouse, through Sor-A treatment cerebral ischemia mouse to The ratio of left steering is lower than the ratio of DMSO group, and the limbs symmetry for representing treatment mouse is better than control mice.See Fig. 3.

2) wrong step test: compared with DMSO mouse, cerebral ischemia mouse opposite side forelimb and hind leg mistake step number through Sor-A treatment The percentage of the total wrong step number of Zhan significantly reduces, and approaches with sham group mouse.See Fig. 4.

3) cylinder test: after injury, non-damaging side (right side) limbs preference use ratio significantly rises cerebral ischemia mouse The preference use ratio of height, Sor-A treatment mouse limb is substantially less than DMSO control group.See Fig. 5.

4) neural sensation and motor function are assessed:

A) limbs feel function score: the cerebral ischemia mouse damage opposite side of Sor-A treatment is significantly higher than the reaction of stimulation DMSO group, see from.

B) climbing experiment: after cerebral ischemia, the climbing ability of mouse is significantly reduced, and treats through Sor-A, climbs ability Higher than DMSO group, Fig. 6 B is seen.

C) forelimb walking experiment: the forelimb walking ability of mouse is remarkably decreased after cerebral ischemia, through the small of Sor-A treatment Mouse fall reduces, and sees Fig. 6 C.

D) side turns experiment: mouse is raised in the sky from 10 centimetres of desktop or more to observe discovery when the side of mouse turns ability Compared with DMSO control group mice, side turns ability and is significantly increased mouse through Sor-A treatment, sees Fig. 6 D.

E) after above-mentioned scoring adding up, the comprehensive score of mouse is shown, Sor-A treats the neural function for being remarkably improved mouse Energy comprehensive score, is shown in Fig. 6 E.

F) sticky paper is tested: mouse is after cerebral ischemia, and the time needed for the sticky paper in the centre of the palm is removed dramatically increases, Sor-A Treating the time needed for mouse removes sticky paper significantly reduces, and sees Fig. 6 F.

5) Morris water maze test: mouse is significant after cerebral ischemia in the residence time of target quadrant in water maze It reduces, Sor-A treatment significantly increases it in the residence time of target quadrant.And the swimming rate of three groups of mouse does not have conspicuousness Difference.See Fig. 7.

In conclusion behaviouristics testing result is shown, Suo Lafen A can improve long-term feeling after ischemic brain damage, fortune It is dynamic, limbs symmetry and learning and memory function.

The nervous function damage of (or ischaemic neuronal) after 3 Suo Lafen-A of embodiment can prevent ischemic brain damage

8-12 weeks male C57/BL6 wild-type mice (being purchased from Shrek experimental animal company) of selection, is randomly divided into 3 groups, Sham group (n=8), DMSO group (n=8), SorA group (n=8).SorA group gives 1mg/kg/d, and intraperitoneal injection pre-processes 3 days It is followed by being performed the operation by Sham or MCAO.

Postoperative 3 days, 5 days, 7 days, 14 days, row Animal Behavior Science detection in 28 days.Method is same as above.3 days after surgery execution mouse, Perfusion carries out NeuN immunofluorescence dyeing after taking brain, measures the existing state of neuron.

It is significantly more than the experimental results showed that applied the neuron survived in the ischemic area of the mouse of Suo Lafen-A in advance DMSO control group mice, is shown in Fig. 8.

The dose screening of 4 Suo Lafen-A of embodiment is tested

1 same mouse of this experiment Example, and mouse is divided into 6 groups；N=8, the Suo Lafen A dosage point of every group of use Not are as follows: G0 placebo is as control；G1 Suo Lafen-A 0.1/kg mg；G2 Suo Lafen-A 1/kg mg；G3 Suo Lafen-A 5/kg mg；G4 Suo Lafen-A 25/kg mg；G5 Suo Lafen-A 50/kg mg.The experiment content of embodiment 1-3 is repeated in G0-G5.

As a result (not shown) shows when the mouse dose of Suo Lafen A is 0.1 or 1/kg mg, Suo Lafen A to acute or Chronic ischemic cerebral injury has certain effect, but no significant difference compared with compareing G0 group；

When the mouse dose of Suo Lafen A is 5,25,50/kg mg, Suo Lafen A is to dosage form or chronic ischemic cerebral injury It is significantly improved effect (kinesitherapy nerve, sensory nerve have significant recovery), but the result between G3, G4, G5 is without aobvious Write sex differernce.

Therefore, Suo Lafen A has the effect of the nerve recovery after ischemic brain damage in low dose faint, but works as When dosage rises to 5mg/kg, then start to generate obvious action, and effect rises there is no obvious after escalated dose.

All experiment mices are without generating obvious or serious adverse reaction.

All references mentioned in the present invention is incorporated herein by reference, independent just as each document It is incorporated as with reference to such.In addition, it should also be understood that, after reading the above teachings of the present invention, those skilled in the art can To make various changes or modifications to the present invention, such equivalent forms equally fall within model defined by the application the appended claims It encloses.

Claims (10)

1. the purposes of Suo Lafen-A (Soraphen-A) or its pharmaceutically acceptable salt, which is characterized in that it is pre- to be used to prepare (i) Anti- and/or treatment ischemic brain damage pharmaceutical composition；And/or the pharmaceutical composition of (ii) protection ischaemic neuronal, In, the Suo Lafen-A has structure shown in Formulas I:

2. purposes as described in claim 1, which is characterized in that the ischemic brain damage includes Acute ischemia rePerfusion Or chronic ischemic cerebral injury.

3. purposes as claimed in claim 2, which is characterized in that the Acute ischemia rePerfusion includes that disease time is 2 small When -3 days between ischemic brain damage.

4. purposes as claimed in claim 2, which is characterized in that the chronic ischemic cerebral injury include disease time 3 days it Ischemic brain damage afterwards.

5. purposes as described in claim 1, which is characterized in that the ischemic brain damage includes cerebral apoplexy, transient ischemia/reperfusion Infuse damage, transient ischemic attack, neonatal hypoxic ischemic encephalopathy.

6. purposes as described in claim 1, which is characterized in that the administration dosage of the Suo Lafen-A is 1-100mg/kg/ days.

7. purposes as described in claim 1, which is characterized in that the pharmaceutical composition contain Suo Lafen-A or its pharmaceutically Acceptable salt is as active constituent and pharmaceutically acceptable carrier.

8. purposes as claimed in claim 7, which is characterized in that the pharmaceutical composition also contains thrombolytics and/or nerve Protective agent.

9. purposes as claimed in claim 8, which is characterized in that the thrombolytics is rtPA.

10. a kind of method of external non-therapeutic protection ischaemic neuronal, which is characterized in that comprising steps of being drawn containing rope Cultivate ischaemic neuronal under conditions of sweet smell-A or its pharmaceutically acceptable salt, and/or to the neuron handled through ischemic Suo Lafen-A is added in culture, to protect ischaemic neuronal.