CN105974131A - Application of c-Kit serving as drug addiction treatment target - Google Patents
Application of c-Kit serving as drug addiction treatment target Download PDFInfo
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- CN105974131A CN105974131A CN201610435065.8A CN201610435065A CN105974131A CN 105974131 A CN105974131 A CN 105974131A CN 201610435065 A CN201610435065 A CN 201610435065A CN 105974131 A CN105974131 A CN 105974131A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
- G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/30—Psychoses; Psychiatry
- G01N2800/307—Drug dependency, e.g. alcoholism
Abstract
The invention discloses new application of c-Kit, and provides application of the c-Kit serving as a drug target in screening of drugs for treating drug addiction and application of the c-Kit in screening of drugs for treating substance dependence addiction psychological craving and excitability of addiction drugs. A classical rat sensitization and conditioned place preference animal model for evaluating addiction is adopted for observing influences of an inhibitor imatinib of the c-Kit on renewed behaviors after formation and withdrawal of rat sensitization behavior expression and conditioned place preferences respectively, the inhibiting effect of imatinib on morphine addiction and an anti-relapse effect after withdrawal of morphine addiction is achieved are evaluated, and application of an acting target c-Kit receptor serving as the drug treatment target of drug addiction is determined. The c-Kit is good in effect and is expected to fundamentally treat drug addiction.
Description
Technical field
The invention belongs to pharmaceutical technology field, be specifically related to c-Kit and become at screening medicine as drug targets
Application in addiction medicine.
Background technology
Drug dependence (drug abuse) or addiction (drug addiction) medical science that become international and sociology
Problem, drugs not only can serious harm human health, also bring a series of economy, society and political issue.
According to the United Nations's drugs and crime problem office 2013 world's drug Use Report, International Pharmaceutical abuse in 2011
Population accounts for the 0.3%～0.9% of whole world adult's sum, and the number of taking drugs that China registers on the books is 247.5 ten thousand.
In recent years, exogenous opioid (such as morphine, heroin) abuse has become the most serious social problem.
It the most seriously wrecks the physical and mental health of misuser, the most also gives family, society's bringing on a disaster property consequence.
Because opioid once abuses addiction, give up extremely difficult.And according to statistics, relapsing of China's drug dependence
Rate is up to more than 95%.
Drug dependence be a kind of with ability out of hand and compulsive drug use the chronic recurrent brain as principal character
Disease, its Forming Mechanism is still not clear (Nestler, 2004).At present, junkie is carried out drug addiction treatment main
Including natural drug withdrawal, alternative medicine (nicotine replacement therapy, NRT) and symptomatic treatment.Respectively
Plant therapy and all can obtain certain effect, but the best to the therapeutic effect of the drug craving of addiction.Addiction mechanism
Not clear, effective target for the treatment of narcotic addiction is limited, seeks the treatment for drug dependence drug craving of the treating narcotic addiction novel targets
Extremely urgent.
C-Kit gene is positioned at human chromosome 4q12-13, belongs to proto-oncogene, and its product is III type tyrosine kinase.
C-Kit as one of the important member of tyrosine kinase receptor protein family, itself and stem cell factor (SCF)
Be combined as important receptor-ligand complex by activate downstream signaling molecule complex, produce powerful company
Lock reactor, thus play very important effect in the regulation and control such as cell differentiation and proliferation.Up to now, two
Tens kinase mediated signals such as dimerization structure and phosphatidylinositol-3 kinase (PI3K), JAK-STAT
Path is confirmed closely related with the activation of c-kit successively, and for the targeted drug of suppression c-Kit activation design
Put into application the most successively, in the diagnosis of clinically relevant tumor and treatment etc. together with specific monoclonal antibody
Aspect serves critical effect.C-Kit expresses in the patient at 70% small cell lung cancer and GIST.Extremely
Till the present, whether whether c-Kit play an important role in addiction and can be as the target of anti-additive medicament effect
Mark has no report.
Summary of the invention
The problem existed for prior art, it is an object of the present invention to provide c-Kit and exists as drug targets
Application in the medicine of screening treatment of drug addiction.
It is a further object to provide c-Kit to depend at screening treatment dependence producing drug material as drug targets
Application in the addiction drug craving relied and excitatoty medicine.
Imatinib (imatinib) is tyrosine kinase inhibitor, and this medicine is public by Novartis of Sweden (Novartis)
Department exploitation, inhibited to c-Kit, the present invention through gavage give experimental rat imatinib (0,
100mg/kg) or central nervous system's core group is after location gives, subcutaneous injection 10mg/kg morphine, warp is used
Allusion quotation evaluates rat sensitization and the Conditioned place preference animal model of addiction, observes imatinib respectively to rat
Sensitization Behavior Expression and Conditioned place preference form the impact giving up rear resume combustion behavior, and is it right to evaluate imatinib
The inhibitory action of coffee addiction and morphine addiction give up after preventing suction again effect；Determine that it is made by western-blot
By target spot c-Kit activity change after imatinib central nervous system's core group location gives, observe c-Kit
Probability as drug addiction treatment target spot.
Result is visible: imatinib can suppress the formation of morphine in rats sensitization, and locating injection imatinib is in prize
The activity of the core portion suppression c-Kit of reward related brain areas nucleus accumbens septi, can weaken the drug craving of morphine-addicted rats.
Result above shows: c-Kit plays an important role in drug dependence, is expected to give up poison for its design medicine
Addiction.
Dependence producing drug of the present invention refers to narcotics and psychotropic drugs etc..Wherein narcotics is divided into again
Opiates, cocaines, cannabis, opium that opiates comprises natural origin and the effective one-tenth therefrom extracted
Divide such as morphine, and effective ingredient is processed the product such as heroin obtained, the artificial conjunction of similar opium effect
Finished product such as methadone etc..Psychotropic drugs is divided into sedative hypnotic and antianxiety drugs, central stimulant, hallucinogen
Deng.Also include ethanol, Nicotiana tabacum L. and volatile organic solvent etc..
The present invention has the advantage that: (1) present invention is that treatment of drug addiction drug craving provides one effectively
The target spot for the treatment of；(2) present invention is all effective to the irritability of drug craving and drug-induced, and therapeutic effect is good,
Relapsing after being expected to fundamentally therapeutic substance addiction and preventing drug withdrawal.
Accompanying drawing explanation
Fig. 1 is spontaneous activity case apparatus figure；
Fig. 2 is the impact that morphine in rats sensitization is formed by imatinib；
A is for being administered experiment flow figure；B is the impact that morphine in rats sensitization is formed by imatinib
Fig. 3 is Conditioned place preference installation drawing；
Fig. 4 is that the location, core portion of nucleus accumbens septi gives the drug craving that imatinib suppresses morphine-addicted rats；
A is the time flow chart of nucleus accumbens septi site-specific delivery of drugs；B is that the location, core portion of nucleus accumbens septi gives imatinib to morphine
The inhibitory action of the drug craving of addictive rats；C is that the location, core portion of nucleus accumbens septi gives imatinib to c-Kit
The inhibitory action of activity
Detailed description of the invention
By combination accompanying drawing described further below it will be further appreciated that the features and advantages of the invention.Thered is provided
Embodiment be only the explanation to the inventive method, and limit never in any form the present invention disclose remaining in
Hold.Material used in following embodiment, reagent etc., if no special instructions, the most commercially obtain.
The impact that morphine in rats sensitization is expressed by [embodiment 1] imatinib
The present embodiment selection imatinib is as the medicine of the frequently-occurring activity of anti-morphine ab addiction, by setting up morphine
Sensitized rats model, inquires into c-Kit inhibitor imatinib to the frequently-occurring spontaneous activity caused by morphine in rats
Improvement result, it is intended to select determined curative effect, frequently-occurring spontaneous work caused by the anti-psychoactive drug substance that toxicity is little
Dynamic drug target mark.
Materials and methods
Medicine and reagent Morphine (Qinghai Pharmaceutic Plant)；Imatinib (Novartis PharmaStein
AG)；
Animal SPF level SD male rat, body weight 220-250g.Animal Experimental Study center, Hubei Province provides,
The animal quality certification number is NO.42000600012344, production licence number: SCXK (Hubei Province) 2015-2018.
Mus feedstuff, is purchased from Wuhan University's Experimental Animal Center.
Experimental technique
Animal packet is with process: rat is randomly divided into four groups, respectively normal saline+group of solvents, normal saline
+ imatinib administration group and morphine+group of solvents, morphine+imatinib administration group.All real Wuhan University animal
Testing in the SPF level environment of center and raise, temperature is 23 ± 2 DEG C, and humidity is 50 ± 5%, and light application time is
6:00-18:00, uses 12 hours light and shades alternately, and guarantee rat can freely obtain when raising food with
Drinking water, is provided which before experiment that week age environment adapts to.
Animal model: animal model: experiment the previous day (the 0th day), carries out activity base to experimental rat
Line measures, and is randomly divided into 4 groups (n=10) according to measurement result；Test the 1st～5 day, normal saline+solvent pair
According to group and normal saline-imatinib administration group equal subcutaneous injection normal saline (1ml/kg), morphine+solvent pair
According to group and morphine-imatinib administration group equal subcutaneous injection morphine (10mg/kg), after being administered, all of dynamic
Thing record spontaneous activity 60 minutes.So experiment repeats 5 days.Then give up 5 days, in experiment the 10th day,
Normal saline+solvent control group and morphine+solvent control group shift to an earlier date 45 minutes gavage normal saline (2ml/kg),
And normal saline-imatinib administration group and morphine+imatinib administration group shift to an earlier date 45 minutes gavage imatinibs
(100mg/kg), after administration, excite with dose morphine (5mg/kg) subcutaneous injection, detect its spontaneous work
Dynamic behavior 60 minutes, observes the change of spontaneous activity behavior, as shown in Figure 2 A.
Testing index:
Spontaneous activity detection case (such as Fig. 1) record spontaneous activity 1 hour is put into after each group rat injection morphine,
After sensitization is formed, spontaneous activity uses DigBehv spontaneous activity video analytic system (Chinese Academy of Medical Sciences's medicine
Thing Research Institute), it is by 4 spontaneous activity inspection boxes, video synthesizer, video pattern sampling card and to divide
The compositions such as analysis software.Native system can carry out video tracking to activities in rats, automatically record mice event trace,
The movable number of times of record rat.The index of spontaneous activity evaluation is: in rat certain period of time (such as 60min)
Movable total degree, i.e. total degree increase display spontaneous activity increase.
Experimental result
Result is visible, and rat, after 5 days morphine successive administrations, finds that packet gives imatinib and physiology
The spontaneous behavior of the rat of saline intervention has significant difference, i.e. system gives imatinib to normal dynamic
The inner directed behavior of thing has not significant impact, and can get rid of the medicine effect of imatinib self.Subcutaneous injection
The experimental group of morphine, system gives blank solvent, compares with saline control group, can observe SD rat
Spontaneous behavior showed increased, irritability increases, and sensibilization is obvious；After system gives imatinib, with
The morphine group of blank solvent compares discovery sensitization expression and receives substantially suppression.(* represents poor as shown in figure 2b
Different there is significance).
The location, core portion of [embodiment 2] nucleus accumbens septi gives imatinib suppression morphine in rats addiction and craves at heart
Being understood imatinib by the result of embodiment 1 can be as the medicine of anti-morphine ab addiction, and it is
The inhibitor of c-Kit, by setting up morphine Conditioned place preference (conditioned place preference, CPP)
Model, research imatinib location microinjection enters the core portion Syndrome of Morphine Dependent of nucleus accumbens septi and gives up rear drug craving
Impact, observes the c-Kit having great expression (Katafuchi, 2000) in addictive drug effect related brain areas simultaneously
Activity change.It is intended to determine that it becomes at the medicine screening a kind of determined curative effect, toxicity is little as drug targets
The effect played in addiction medicine.
Materials and methods
Medicine and reagent Morphine (Qinghai Pharmaceutic Plant)；Imatinib (Novartis PharmaStein AG)
Animal SPF level SD male rat, body weight 220-250g.Animal Experimental Study center, Hubei Province carries
Confession, the animal quality certification number is NO.42000600012016, production licence number: SCXK (Hubei Province) 2015-2018.
Mus feedstuff, is purchased from Wuhan University's Experimental Animal Center.
Experimental apparatus
Conditioned place preference instrument (institute of Materia Medica,Chinese Academy of Medical Sciences development): experiment uses computer automatically to control
System.Device is by the conditioned place preference case constituted for three casees: two side rooms and a medial compartment (such as Fig. 3).
Three Room by moveable dividing plate separately, inside and outside be black.Wherein A case and B case are positioned at the both sides of intermediate box,
Size is identical, and A case sidewall has the square that 9 Yellow light-emitting low temperature diodes are constituted, and base plate is rustless steel steel bar,
B box plate is Stainless steel mesh.Rat can be sent to calculate by data with going out indegree in each case time of staying
Machine, collects record behavioristics data automatically.
Experimental technique
The core portion row positioning operation of rat nucleus accumbens septi, carries out normal saline or morphine CPP training after the week,
As shown in Figure 4 A.
(1) foundation of morphine CPP model
Basic value is tested:
1st day, passage between open three casees, start CPP program on computer, rat is put into by medial compartment,
It is allowed in three casees the most movable 15 minutes, its residence time in each room of computer synchronous recording.
Conditioned place preference is trained:
2nd to 9 day, close passage between three casees.2nd, 4,6,8 days, experimental group subcutaneous injection morphine (10mg/kg)
And put into companion medicine side 45 minutes；Matched group subcutaneous injection normal saline (1ml/kg) also puts into 45 points, non-companion's medicine side
Clock.3rd, 5,7,9 days, experimental group and the equal injecting normal saline of control rats, experimental group put into non-companion
Medicine side, matched group is put into companion medicine side, is 45 minutes.The companion medicine side of every rat is fixing.Often group is big
Rearging cage it is put back into after Mus.
Morphine CPP tests:
Within 10th day, carry out CPP test, similar to basic value test phase.Passage between open three casees, not appoints
What injection, starts CPP program on computer, and rat is put into by medial compartment, and it is the most movable to be allowed in three casees
15 minutes, its residence time in each room of computer synchronous recording.Preference mark (CPP score) is defined
Difference by companion's coyote hole stayed time with non-companion's coyote hole stayed time.By rat measured value after CPP in companion's medicine-chest
Compare with front side value and determine whether rat forms CPP.And do not form CPP's according to measured value rejecting after CPP
Rat, carries out matched packet by animal.
(2) foundation of related environmental cues induction drug-seeking behavior model
At the 11st day of experiment, rat shifted to an earlier date 15 minutes micro-injection imatinibs (4 μ g/0.5 μ l) and physiology salt
Water (0.5 μ l), in the core portion of nucleus accumbens septi, is then exposed to, to medicine-chest, stop 10 minutes, and a part of rat returns again
To raising in cages in environment, with the ethological test of postcondition Place Preference in a week after 24 hours；Another portion
Divide rat to break end after 60 minutes and take brain, the method using western-blot, the core portion c-kit of detection nucleus accumbens septi
The relation that the change of activity changes with Conditioned place preference behavioristics.
(3) morphine CPP retests
Imatinib be administered after the 1st day and the 7th day, i.e. the 12nd and 18 day, test rat was to accompanying medicine-chest
Preference degree, 15 minutes, similar to basic value test phase.Middle 13rd day by the 17th day, to rat
It is left intact.
(4) the lighting of morphine CPP
Within 14th day, detect latter 24 hours, i.e. the 15th day, utilize dose morphine (3ml/kg) to light.
After injection of morphia 10 minutes, rat is put into the value of CPP test that intermediate box starts 15 minutes.
Testing index:
After rat training, using Conditioned place preference case detection morphine addiction situation, Conditioned place preference is marked
(CPP Score) i.e. reacts the formational situation of rat addictive behavior, and CPP Score increases and has significant difference,
Addictive behavior is formed.The core portion microinjection imatinib of nucleus accumbens septi expose again after 60min, detection c-kit work
Property, and the change of testing conditions Place Preference scoring behind 24 hours and 7 days.
Experimental result
Result is shown in that Fig. 4 B, administration group difference compared with matched group have significance.The core portion microinjection of nucleus accumbens septi
After imatinib, Conditioned place preference weakens, and is not ignited after 2 weeks；The rat of non-administration, condition position
Preference still exists.Western-blot method is used to detect that the core portion of nucleus accumbens septi gives c-kit after imatinib
Phosphorylation activity substantially reduce (Fig. 4 C).Illustrate microinjection imatinib in the core portion of nucleus accumbens septi, by pressing down
C-kit processed activation blocks and is administered the drug craving that environment causes, and can improve morphine addiction symptom.
Experiment conclusion
In a word, imatinib is administered systemically the formation of the sensitization activity that morphine can be suppressed to cause, microinjection c-Kit
Inhibitor is the formation of drug craving after the core portion of nucleus accumbens septi can suppress morphine addiction, and c-Kit signaling molecule is described
In the addictive behavior caused by morphine, play the effect of key, can reach for its design its activity of Drug inhibition
The effect for the treatment of narcotic addiction, is the useful effect target for anti-additive medicament research and development.
Claims (3)
1.c-Kit as drug targets screening treatment dependence producing drug substance depilatory medicine in application.
2.c-Kit is as drug targets application in the screening treatment addiction drug craving of dependence producing drug substance depilatory and excitatoty medicine.
Application the most according to claim 1 and 2, it is characterized in that, described dependence producing drug refers to (1) narcotics, wherein narcotics is divided into again opiates, cocaines, cannabis, opium that opiates comprises natural origin and the effective ingredient morphine therefrom extracted, and effective ingredient is processed the product heroin obtained, the Synthetic artifact of similar opium effect；(2) psychotropic drugs, psychotropic drugs is divided into sedative hypnotic and antianxiety drugs barbiturates, central stimulant amphetamine, hallucinogen Lysergide；(3) ethanol, Nicotiana tabacum L. and volatile organic solvent.
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112569355A (en) * | 2019-09-30 | 2021-03-30 | 武汉大学 | Application of imatinib and derivative thereof and addiction substance combined medicine or compound preparation in prevention and treatment of addiction and prevention and treatment of relapse |
| CN112569238A (en) * | 2019-09-30 | 2021-03-30 | 武汉大学 | Application of imatinib and derivative thereof and analgesic combined medicine or compound preparation in treating pain |
| CN112569352A (en) * | 2019-09-30 | 2021-03-30 | 武汉大学 | Application of c-Kit as target for behavior addiction treatment |
| CN112569237A (en) * | 2019-09-30 | 2021-03-30 | 武汉大学 | Application of combination or compound of imatinib and derivatives thereof and nicotine or analogues thereof in preventing and treating nicotine addiction and relapse |
| CN112575073A (en) * | 2019-09-30 | 2021-03-30 | 武汉大学 | Application of c-Kit as addiction diagnosis and monitoring marker |
| WO2021063388A1 (en) * | 2019-09-30 | 2021-04-08 | 武汉大学 | Application of c-kit as addiction diagnosis and monitoring marker and addiction treatment target |
| CN116497031A (en) * | 2023-06-02 | 2023-07-28 | 徐州医科大学 | SiRNA for relieving mouse morphine addiction and application thereof |
| CN117461602A (en) * | 2023-10-25 | 2024-01-30 | 六合熙诚(北京)信息科技有限公司 | Construction method of morphine addiction animal model |
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| CN116497031A (en) * | 2023-06-02 | 2023-07-28 | 徐州医科大学 | SiRNA for relieving mouse morphine addiction and application thereof |
| CN117461602A (en) * | 2023-10-25 | 2024-01-30 | 六合熙诚(北京)信息科技有限公司 | Construction method of morphine addiction animal model |
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| Publication number | Publication date |
|---|---|
| CN105974131B (en) | 2017-12-26 |
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