CN101317844B - Application of N-methyl-D-asparagic acid acceptor inhibitor in preparing contraceptive medicament - Google Patents
Application of N-methyl-D-asparagic acid acceptor inhibitor in preparing contraceptive medicament Download PDFInfo
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- CN101317844B CN101317844B CN2007100417838A CN200710041783A CN101317844B CN 101317844 B CN101317844 B CN 101317844B CN 2007100417838 A CN2007100417838 A CN 2007100417838A CN 200710041783 A CN200710041783 A CN 200710041783A CN 101317844 B CN101317844 B CN 101317844B
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Abstract
The present invention discloses application of N-methyl-D-aspartate (NMDA) receptor depressor to preparation of contraceptive. In the present invention, the NMDA receptor depressor (+) 5-methyl-10, 11-dihydro-5H-dibenzanthracene[a, d]huangengxi-5, 10-imine ( MK-801) are tested on human spermatozoa. Results show that the NMDA receptor exists in the human spermatozoa and is expressed at tail of thehuman spermatozoa. The MK-801 can remarkably disturb the movement direction and velocity of the spermatozoa. Through test of effect that the NMDA receptor depressor MK-801 have on cervix mucus penetration of the human spermatozoa, results show that the MK - 801 can remarkably restrain the spermatozoa from passing through the cervix mucus. The NMDA receptor depressor of the present invention is non-steroid medicine; therefore, the NMDA receptor depressor has no side effect of steroid medicine. The NMDA receptor depressor is safe, effective, and simple in use and has great clinical application value.
Description
Technical field:
The present invention relates to the biological medicine technology field, be specifically related to chemical compound (+)-5-methyl isophthalic acid 0,11-dihydro-5H-dibenzo (a, d) cycloheptene-5, the application of 10-imines in the preparation contraceptive
Background technology:
World population has surpassed 6,000,000,000, and with every year on average newly-increased 7,700 ten thousand speed rapidly increase, thereby, control the population size, become the significant problem that the whole mankind faces, contraception is the great realistic problem that the couple at child-bearing age faces.Contraceptive device mainly contains at present: medical contraception and surgical contraception, wherein, medical contraception comprises oral medication and two kinds of methods of external applied contraceptive, the most frequently used oral contraceptive is a contraceptive steroid, because it is a hormone medicine, except each link of reproductive system is had the obvious effect, almost each system of human body all there is in various degree influence, there are some complication and contraindication simultaneously.External applied contraceptive is vagina medicinal for example, is one of human the most ancient method in order to the control fertility.Before oral contraceptive and intrauterine device appearance, vaginal contraception method and safe period, controlling desire, external row's essence, artificial abortion etc. are the common methods of human control fertility.Up to the later stage fifties in last century, oral contraceptive and intrauterine device develop in succession, have started the New Times of birth control.But the seventies in last century,, discovered their caused general untoward reaction and used contraindication from epiphytotics, and caused people's attention day by day along with the extensive use of oral contraceptive and intrauterine device.Therefore, people turn to research safety again, the external applied contraceptive that untoward reaction is little.At present, the external applied contraceptive that people use mostly is the preparation of making in alkane benzene polyalcohol ether ninth of the ten Heavenly Stems, menthylphenoxypolyethoxy ethanol, the emphasis of external contraceptive medicine effect link is in the research of spermicide, and the present invention is the unitransport albumen of the receptor that exists with human sperm's afterbody is target spot, pharmaceutical intervention makes the sperm can not proper motion, thereby reaches the purpose of contraception.
The inventor finds the existence of glutamate receptor on the human sperm after deliberation, a kind of N-methyl-D-aspartate (N-methyl-D-aspartate (NR2B), NMDA (NR2B)) receptor-specific is present in people's sperm tail, at making it lose the mobility that reaches fertilization behind the pharmaceutical intervention human sperm of this receptor, this research will provide new way for the mankind's contraception.
Glutamic acid is the topmost excitatory neurotransmitter of central nervous system (CNS), and its function is mediated by glutamate receptor, and glutamate receptor is divided into ion-type glutamate receptor and metabotropic glutamate receptor.The neurotransmission effect of glutamic acid is by multiple ligand-gated ion channel (ion-type glutamate receptor) mediation.According to sequence homology and ligand affinity, the ion-type glutamate receptor is divided into three kinds of hypotypes: N-methyl-D-aspartate (N-methyl-D-aspartate, NMDA) receptor, alpha-amido-3-hydroxy-5-methyl base-4-isoxazole propanoic acid (α-amino-3-hydroxy-5-methyl-4-isoxazole propionate, AMPA) receptor and kainic acid (kainate, KA) receptor, the AMPA/KA receptor is called as non-NMDA receptor usually.[HollmannM.Heinemann?S：loned?glutamate?receptors.Ann?Rev?Neurosci?17：31-108，1994；Nakanishi?S：Molecular?diversity?of?glutamate?receptors?and?implications?for?brain?function.Science?258：597-603，1992]。Wherein nmda receptor mainly contains two class subunit: NR1 and NR2; NR2 has four kinds, and NR2A.NR2B.NR2C and NR2D replace in different growth courses and express, and express the highest with NR2B in adults and people.All glutamate receptors all have in the central nervous system expresses and participates in many physiological and pathological factor [Collingridge GL, Lester RA:Excitatory amino acid receptors in the vertebrate centralnervous system.Pharmacol Rev 40:143-210,1989; Sims KD, Robinson MB:Expression patternsand regulation of glutamate transporters in the developing and adult nervous system.Crit RevNeurobio 13:169-197,1999].Yet in peripheral tissues, whether glutamate receptor exists, how not clear function is.Clear and definite is that some glutamate receptors exist on adrenal gland, hypophysis, pinus and islets of langerhans.The recent metabotropic glutamate receptor that studies show that has expression [Storto M on the rat and the mankind's testis, Sallese M, Salvatore L, Poulet R, Condorelli DF, Albani PD, Marcello, MF, Romeo R, Piombone P, Barone N, Nicolette F, Blasi AD:Expression of metabotropic glutamate receptors in the rat and human testis.J Endocrinol 170:71-78,2001], metabotropic glutamate receptor is a g protein coupled receptor.
Summary of the invention
The technical problem to be solved in the present invention is how to use the nmda receptor inhibitor is reduced human sperm's mobility and penetrating of cervical mucus, designs new contraceptive.
The invention provides chemical compound (+)-5-methyl isophthalic acid 0,11-dihydro-5H-dibenzo (a, d) cycloheptene-5, the application of 10-imines in the preparation contraceptive.
The inventor has disclosed the existence of nmda receptor on the human sperm, and immunofluorescence experiment shows that nmda receptor exists on the human sperm, and NMDA (NR2B) expression of receptor is in human sperm's afterbody.Inhibitor (+) 5-methyl isophthalic acid 0 with nmda receptor, 11-dihydro-5H-dibenzo (a, d) cycloheptene-5, the function of nmda receptor on 10-imines (MK-801) the blocking-up sperm, the motion of finding sperm has been subjected to appreciable impact, and especially the rectilinear motion rate of sperm significantly descends; Find that simultaneously the ability that sperm penetrates cervical mucus has significantly weakened, illustrate with the nmda receptor to be that the inhibitor of target spot can be used for contraception clinically.
The inhibitor of nmda receptor of the present invention is MK-801:
Chemical name: (+)-5-methyl isophthalic acid 0,11-dihydro-5H-dibenzo (a, d) cycloheptene-5, the 10-imines has following structural:
At present, the nmda receptor inhibitor is the medicine that is used to suppress the central nervous system, the invention provides the new purposes that this type of medicine is used to practise contraception.
Nmda receptor inhibitor of the present invention can be made external-application cream, unguentum, liniment, varnish or gel according to a conventional method when being used to practise contraception.During use, the effective dose of preparation of the present invention can be imposed in male genital organ appearance skin or the vagina, can reach the effect of contraception.
Nmda receptor inhibitor of the present invention is non-hormone medicine, does not therefore have the side effect of hormone medicine.Safe and effective, easy to use, bigger clinical value arranged.
The specific embodiment:
One, embodiment 1
There is test in nmda receptor on the human sperm
1. method is tested the source of required medicine: the antiglutamic acid primary antibody is available from Sigma company, and anti-NB2B antibody is available from SantaCruz company.Two of FITC labelling resists available from Sigma company.Human sperm's sample from 21-43 year man controlling desire after 3 days masturbation obtain.Sperm detected eupyrene sperm in 30 minutes in 37 ℃ of liquefaction, fixed 30 minutes in the phosphate buffer that contains 4% paraformaldehyde, was applied on the microscope slide of gelatin bag quilt, and is dry, fixing.
Human sperm's smear seals to reduce nonspecific immunity dyeing with the PBS buffer that contains 10% goat or rabbit anteserum and 0.1%Triton X-100 then with 0.01M PBS washing.Spend the night for 4 ℃ with primary antibody or as the normal NIS of negative control, after the hatching, smear is with PBS rinsing 3 times, and with corresponding two anti-(being connected FITC) room temperatures under hatched 2 hours, use the PBS rinsing then, last smear detects under the Leica fluorescence microscope and takes pictures.
2, fructufy is issued after examination and approval present sperm tail and can clearly be observed the specific fluorescence of NR2B but sperm head immunologic incompetence (Fig. 1), illustrates that ion-type glutamate receptor NMDA is positioned human sperm's afterbody.
Two, embodiment 2
Nmda receptor inhibitor MK-801 is to the influence test of human sperm's mobility
1, method: the source of testing required medicine: MK-801 is available from Sigma company, human sperm's masturbation method is taken from ascetic 5-7 days of 25-40 year, the healthy man that obsterical history is arranged, after the sperm liquefaction, to suspend 30 minutes in 37 ℃ of water-baths of BWW liquid, adjusting sperm concentration is 6 * 107, specimen adds respectively in 37 ℃ of water-baths of variable concentrations MK-801 fed 30 minutes, detect with U.S. Hamilton-IVOS sperm analyser, repeat 3 times, experimental result is analyzed with the origin statistical software.
2, result: the point-to-point speed (VSL) that adds sperm behind 100 μ M and the 500 μ MMK-801 and average path velocity (VAP) obviously reduce to be compared difference with the blank group significance is arranged.Show that MK801 can significantly disturb the direction of motion and the speed of sperm.See Fig. 2 and Fig. 3.
Three, embodiment 3
Nmda receptor inhibitor MK-801 penetrates the influence test of cervical mucus to the human sperm
1, method: the source of testing required medicine: MK-801 is available from Sigma company, human sperm's masturbation method is taken from ascetic 5-7 days of 25-40 year, the healthy man that obsterical history is arranged, after the sperm liquefaction, to suspend 30 minutes in 37 ℃ of water-baths of BWW liquid, adjusting sperm concentration is 6 * 107, specimen adds respectively to feed in 37 ℃ of water-baths of variable concentrations MK-801 and was used in 30 minutes testing, cervical mucus, adopt the absorption method, obtained in 1-2 days before ovulation, inclusion criteria is: there is the healthy women of obsterical history in 25-40 year, presses the Moghissi marking system, score value＞10 minute, the application cervical mucus interacts and tests (slide method), and experiment repeats 3 times, and experimental result uses the origin statistical software to analyze.
2, result: the blank group with add number that 10 μ M, 30 μ M, 100 μ M and 300 μ M, MK-801 group sperm the penetrate seminal fluid-cervical mucus contact interface difference of comparing significance arranged.Show that MK-801 can significantly suppress sperm and pass through cervical mucus.See Fig. 4
Above embodiment medicament sources, (+) 5-methyl isophthalic acid 0,11-dihydro-5H-dibenzo (a, d) cycloheptene-5,10-imines (MK-801) is available from Sigma company.
Description of drawings
Fig. 1 immunofluorescence experiment shows that NMDA (NR2B) receptor-specific is expressed in human sperm's afterbody.
Fig. 2 nmda receptor inhibitor MK-801 influences * p＜0.05 to human sperm's point-to-point speed
Transverse axis is represented: the intervention of variable concentrations MK-801
The longitudinal axis is represented: the numerical value of human sperm's point-to-point speed
The influence of MK-801 pair of human sperm's average path speed of Fig. 3 nmda receptor inhibitor
*P＜0.05
Transverse axis is represented: the intervention of variable concentrations MK-801
The longitudinal axis is represented: the numerical value of human sperm's average path speed
100 μ M MK-801 intervene the result
Fig. 4 nmda receptor inhibitor MK-801 couple human sperm penetrates the influence of cervical mucus
*P＜0.05,
*P＜0.01,
* *P＜0.001
Transverse axis is represented: the intervention of variable concentrations MK-801
The longitudinal axis is represented: sperm-cervical mucus contact test (sperm cervical mucus contact test, SCMC, slide method) is by the sperm count at interface
Claims (2)
1. chemical compound (+)-5-methyl isophthalic acid 0,11-dihydro-5H-dibenzo (a, d) cycloheptene-5, the application of 10-imines in the preparation contraceptive, its effect valid density scope is between 30 μ M to the 300 μ M.
2. chemical compound according to claim 1 (+)-5-methyl isophthalic acid 0,11-dihydro-5H-dibenzo (a, d) cycloheptene-5, the application of 10-imines in the preparation contraceptive is characterized in that described medicine is external-application cream, unguentum, liniment, varnish or gel.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1309562A (en) * | 1998-07-16 | 2001-08-22 | 纪念斯隆-凯特林癌症中心 | Topical compositions comprising opioid analgesic and NMDA antagonist |
| CN1479617A (en) * | 2000-12-13 | 2004-03-03 | �Ʒ� | N-methyl-D-aspartate receptor agonist pharmaceutical compositions |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1309562A (en) * | 1998-07-16 | 2001-08-22 | 纪念斯隆-凯特林癌症中心 | Topical compositions comprising opioid analgesic and NMDA antagonist |
| CN1479617A (en) * | 2000-12-13 | 2004-03-03 | �Ʒ� | N-methyl-D-aspartate receptor agonist pharmaceutical compositions |
Non-Patent Citations (4)
| Title |
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| 徐岩等.地佐环平对左旋多巴引起帕金森病大鼠行为和基底节Fos表达的影响.中华神经科杂志第36 卷 第6 期.2003,第36 卷(第6 期),参见第439页. |
| 徐岩等.地佐环平对左旋多巴引起帕金森病大鼠行为和基底节Fos表达的影响.中华神经科杂志第36 卷 第6 期.2003,第36 卷(第6 期),参见第439页. * |
| 蒋青松等.地佐环平对缺血突触体氨基酸释放和游离钙的影响.中国药理学与毒理学杂志第16卷 第3期.2002,16(3),参见第161-164页. |
| 蒋青松等.地佐环平对缺血突触体氨基酸释放和游离钙的影响.中国药理学与毒理学杂志第16卷 第3期.2002,16(3),参见第161-164页. * |
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