本文選題：抗抑郁藥 + YL-0919�。� 參考：《中國人民解放軍軍事醫(yī)學科學院》2014年博士論文

【摘要】：抑郁癥是致病率和致殘率最高的精神疾病之一,因其具有高發(fā)病、高自殺、高復發(fā)、高致殘率,和低識別、低就診、低治療率成為嚴重的全球性公共衛(wèi)生問題和突出社會問題。WHO預測,到2020年抑郁癥將成為危害人類健康的第二大疾患。目前臨床最為廣泛的一線絕大多數抗抑郁藥物,包括5-HT重攝取抑制劑(SSRIs,如氟西汀)、5-HT/NE雙重重攝取抑制劑(SNRIs,如度洛西汀)等均存在起效延遲、有效率不高、導致性功能障礙和自殺傾向等較嚴重缺陷。近年來基于“優(yōu)化的單胺策略”,5-HT1A受體部分激動和選擇性的5-HT再攝取抑制雙靶標抗抑郁藥(serotonin partial agonist and reuptake inhibitors, SPARIs),因其具有強效、快速起效、潛在低自殺傾向等特點,以及不良反應(包括性功能障礙和體重增加等)發(fā)生率低的優(yōu)勢,己成為新型藥物的代表方向。鹽酸羥哌吡酮(YL-0919)是軍事醫(yī)學科學院毒物藥物研究所正在研發(fā)的兼有5-HT1A部分激動和5-HT重攝取抑制雙重靶標的1.1類抗抑郁專利新藥,具有較強的新藥研發(fā)前景,本研究旨在系統(tǒng)探討其行為活性及靶標機制。 目的：主要研究了兼有5-HT1A受體(5-HT1AR)激動和5-HT重攝取抑制雙重活性抗抑郁新藥YL-0919的活性特點及其靶標調節(jié)機制。 方法：采用放射性配體競爭結合實驗分別研究YL-0919與大鼠突觸體蛋白中5-HT轉運蛋白(SERT)和5-HT1A受體的親合性以及受體選擇性,并進一步制備含有SERT的大鼠突觸體蛋白以及采用穩(wěn)定表達人源SERT (hSERT)的HEK-293細胞系進行YL-0919對5-HT的重攝取抑制作用研究。分別采用小鼠懸尾和小鼠強迫游泳兩個經典行為絕望模型評價新藥YL-0919單次給藥的抗抑郁活性,并采用相關5-HT受體阻斷劑,明確YL-0919抗抑郁作用的相關受體。在小鼠獲得性無助模型上評價YL-0919亞慢性給藥的抗抑郁活性,在zero-迷宮、高架十字迷宮、Vogel電擊沖突飲水、新奇抑制攝食行為模型中探討單次或慢性給予大鼠YL-0919的抗焦慮活性。采用小鼠新奇抑制攝食、懸尾實驗和強迫游泳實驗對YL-0919和同靶標抗抑郁藥維拉佐酮的抗焦慮、抗抑郁效果進行比較。在受體后機制研究中,首先在離體水平用競爭免疫熒光法檢測YL-0919對正常大鼠前額皮層突觸膜腺苷酸環(huán)化酶(AC)活性的影響。在整體水平,檢測YL-0919長期給藥后小鼠海馬、前額皮層Foskolin激活的AC活性的變化。應用酶聯免疫吸附法(ELISA)檢測長期給藥后小鼠海馬PKA活性的變化,并在小鼠懸尾和小鼠強迫游泳模型上檢測腦室注射選擇性蛋白激酶A(PKA)抑制劑H-89對YL-0919抗抑郁活性的阻斷作用。采用蛋白免疫印跡分析法(Western Blot)考察慢性給藥YL-0919對小鼠海馬BDNF、p-CREB、p-ERK表達的調控作用。 結果：研究表明,YL-0919對SERT以及5-HT1A受體均具有高親和力結合,Ki分別為0.72nM和0.19nM; YL-0919與NET(NE轉運蛋白)和DAT (DA轉運蛋白)的幾乎沒有親和力。在大鼠前額皮層粗制突觸體中,YL-0919強效抑制[3H]-5-HT的重攝取,ICso為1.78nM；在穩(wěn)定表達人源SERT的HEK-293細胞系中,YL-0919同樣對[3H]-5-HT的重攝取具有強效的抑制作用,IC50為1.93nM。并且YL-0919的5-HT重攝取抑制作用強于臨床一線藥物氟西汀和度洛西汀。YL-0919在濃度為10μM時對D3、D2、D4、D1、D5、5-HT1B、5-HT1D、5-HT2A、5-HT2C、5HT5A、5-HT6、β1A、α1B、α2A、M1、M2、M3、M4、M5等19種受體無親和力。這些結果提示YL-0919對5-HT1A和SERT具有高親和力和高選擇性結合,并且是強效的5-HT重攝取抑制劑。 單次給予YL-0919(0.625-2.5mg/kg, i.g)能夠劑量依賴的顯著縮短小鼠懸尾和強迫游泳不動時間。使用5-HT1A受體阻斷劑WAY-100635預處理能夠完全阻斷YL-0919(2.5mg/kg,i.g)的抗抑郁活性。而5-HT2受體拮抗劑米安色林、5-HT3受體拮抗劑曲匹西隆、α2腎上腺素受體拮抗劑育亨賓對YL-0919的抗抑郁活性無阻斷作用。提示YL-0919急性給藥具有抗抑郁作用,并且與激動5-HT1A受體有關,這與本實驗室的其他研究一致。YL-0919亞慢性給藥(1.25-5mg/kg, i.g)在第2-5天顯著縮短獲得性無助小鼠逃避潛伏期及減少逃避失敗次數,表現出了顯著的抗抑郁活性。單次或慢性給予大鼠YL-0919(0.625-2.5mg/kg,i.g)顯著縮短新奇抑制攝食潛伏期、增加大鼠在zero-迷宮開臂的停留時間和進入開臂的次數、增加大鼠在高架十字迷宮進入開臂次數百分比和在開臂停留時間百分比、增加電擊飲水次數,提示YL-0919單次或慢性給藥具有抗焦慮作用。慢性給予YL-0919(0.625-2.5mg/kg, i.g.)和維拉佐酮(1-4mg/kg, i.g.)均可顯著縮短新奇抑制攝食潛伏期,并都顯著縮短小鼠懸尾、游泳不動時間,提示YL-0919和維拉佐酮抗抑郁、抗焦慮活性相當,但YL-0919的起效劑量較維拉佐酮小1-3倍。 進一步的受體后機制研究發(fā)現,在離體水平YL-0919(10-9-10-5mol/L)與正常大鼠前額皮層突觸膜孵育,在1-10μM濃度時可顯著激活AC活性,表現出與陽性藥氟西汀和丁螺環(huán)酮一致的作用。5-HT1A受體阻斷劑WAY-100635能顯著拮抗YL-0919對AC的激活作用。在整體水平,長期反復給予YL-0919(1.25-2.5mg/kg,i.g.)和維拉佐酮(4mg/kg, i.g.)均能增強小鼠海馬、前額皮層forskolin激活的AC活性,同時增強小鼠海馬PKA活性,并顯著上調小鼠海馬BDNF、p-CREB、 p-ERK的表達。進一步研究證實,腦室注射選擇性PKA阻斷劑H-89能完全阻斷單次給予YL-0919(2.5mg/kg, i.g.)在小鼠懸尾和小鼠強迫游泳模型上的抗抑郁活性。這些結果提示激活5-HT1A受體,并增強cAMP-CREB信號通路的功能是YL-0919抗抑郁作用的重要機制之 結論：YL-0919與SERT蛋白和5-HT1A受體蛋白具有高親合活性并且這種結合具高選擇性；YL-0919具有強效的5-HT重攝取抑制功能,并且強于一線藥物氟西汀和度洛西�。籝L-0919單次、亞慢性或慢性給藥在多種動物模型上,表現出顯著的抗抑郁、抗焦慮作用；YL-0919具有與維拉佐酮相當的抗抑郁、抗焦慮活性,但起效劑量更低；激活5-HT1A受體功能,并增強前額皮層、海馬cAMP-CREB信號通路功能,上調海馬BDNF水平從而促進神經營養(yǎng)是YL-0919抗抑郁、抗焦慮活性的重要機制。
[Abstract]:Depression is one of the mental diseases with the highest morbidity and disability rate. Because of its high incidence, high suicide, high relapse, high disability rate, low recognition, low treatment and low rate of treatment, it has become a serious global public health problem and a prominent social problem.WHO forecast. By 2020, depression will become the second major disease endangering human health. Most of the most widely used antidepressants in the bed, including 5-HT reuptake inhibitors (SSRIs, such as Fu Xiting), 5-HT/NE double reuptake inhibitors (SNRIs, such as duloxetine) have delayed onset and low efficiency, resulting in severe defects such as sexual dysfunction and suicidal tendencies. In recent years, the "optimized monoamine strategy", 5-HT1 A receptor partial excitation and selective 5-HT reuptake inhibits the dual target antidepressant (serotonin partial agonist and reuptake inhibitors, SPARIs), and has become a new drug because of its strong effect, rapid onset, potential low suicide, as well as the low incidence of adverse reactions (including sexual dysfunction and weight gain). YL-0919 is the 1.1 patent new antidepressant drug, which is being developed by the Toxicological Research Institute of Military Medical Science Academy of the PLA, which has both 5-HT1A partial excitation and 5-HT reuptake inhibition target. It has a strong prospect for new drug research and development. The aim of this study is to systematically explore its activity activity and target mechanism.
Objective: To study the activity characteristics and target regulation mechanism of the double active antidepressant antidepressant YL-0919, which has both 5-HT1A receptor (5-HT1AR) and 5-HT reuptake inhibition.
Methods: the affinity and acceptor selectivity of 5-HT transporter (SERT) and 5-HT1A receptor in YL-0919 and rat synaptosomes were studied by radioactivity ligand competition assay, and the synaptosomes in rats with SERT and HEK-293 cell lines with stable expression of human SERT (hSERT) were further prepared and YL-0919 pairs of 5-HT were carried out. The antidepressant activity of the new drug YL-0919 single dose was evaluated by two classical behavioral despair models of mouse hanging tail and forced swimming in mice. The related 5-HT receptor blockers were used to identify the related receptors of the antidepressant effect of YL-0919. The subchronic YL-0919 administration was evaluated in the mouse acquired helplessness model. Antidepressant activity in the zero- labyrinth, the elevated cross maze, the Vogel electric shock drinking water, the novel inhibition feeding behavior model to explore the single or chronic anti anxiety activity of YL-0919 in rats. The antidepressant and antidepressant of YL-0919 and the same target antidepressant, vistadone, were used in the novel inhibition feeding, tail suspension experiment and forced swimming test. The effects were compared. In the post receptor mechanism study, the effects of YL-0919 on the activity of synaptic adenylate cyclase (AC) in the prefrontal cortex of normal rats were first detected by competitive immunofluorescence. At the overall level, the changes in the AC activity of the hippocampus in mice and the Foskolin activated in the frontal cortex after the long-term administration of YL-0919 were detected. The changes of PKA activity in the hippocampus of mice after long-term administration were detected by ELISA. The blocking effect of the selective protein kinase A (PKA) inhibitor H-89 on the antidepressant activity of YL-0919 was detected on the mouse suspension tail and the mice forced swimming model. The chronic administration of YL-0919 to mice was examined by the protein immunoblotting analysis (Western Blot). The regulation of the expression of BDNF, p-CREB and p-ERK in the hippocampus.
Results: the study showed that YL-0919 had high affinity for both SERT and 5-HT1A receptors, and Ki was 0.72nM and 0.19nM, respectively, and there was almost no affinity between YL-0919 and NET (NE transporter) and DAT (DA transporter). In the HEK-293 cell line of human SERT, YL-0919 also has a strong inhibitory effect on [3H]-5-HT, IC50 is 1.93nM. and the 5-HT reuptake inhibition of YL-0919 is stronger than the clinical frontline drug fluoxetine and duloxetine.YL-0919 at the concentration of 10 mu M. 19 receptors, such as 2A, M1, M2, M3, M4, M5, have no affinity. These results suggest that YL-0919 has a high affinity and high selectivity for 5-HT1A and SERT, and is a potent inhibitor for 5-HT reuptake.
A single dose of YL-0919 (0.625-2.5mg/kg, i.g) could significantly shorten the dose dependent mice suspension and forced swimming time. The 5-HT1A receptor blocker WAY-100635 pretreatment could completely block the antidepressant activity of YL-0919 (2.5mg/kg, i.g). The 5-HT2 receptor antagonist, michalin, the 5-HT3 receptor antagonist, traepimacon, and the alpha 2 adrenaline The anti depressant activity of YL-0919 was not blocked by a prime receptor antagonist, suggesting that YL-0919 acute administration has antidepressant effect and is associated with the activation of 5-HT1A receptor. This is consistent with other studies in the laboratory, which is consistent with the other studies in the laboratory (1.25-5mg/kg, i.g), which significantly shortened the escape latency and decrease of the Acquired Helplessness mice on the 2-5 day. The single or chronic YL-0919 (0.625-2.5mg/kg, i.g) significantly shortened the incubation period of new inhibition of feeding, increased the duration of the opening of the arm in the zero- maze and the number of times to enter the arm, and increased the percentage of the opening arm of the rat in the high cross maze and the open arm in the open arm. The percentage of retention time increased the frequency of electric shock and drinking water, suggesting that the single or chronic administration of YL-0919 had anti anxiety effect. Chronic YL-0919 (0.625-2.5mg/kg, i.g.) and Vilazzo ketone (1-4mg/kg, i.g.) could significantly shorten the incubation period of new inhibition of feeding, and shorten the tail of mice and swim time, suggesting YL-0919 and Vera R. Antidepressant and anti anxiety activity were equivalent, but the dose of YL-0919 was 1-3 times smaller than that of group.
Further postreceptor mechanism study found that the activity of AC was significantly activated at the concentration of YL-0919 (10-9-10-5mol/L) in the prefrontal cortex of the normal rat and at the concentration of 1-10 micron M, showing that the.5-HT1A receptor blocker WAY-100635, in accordance with the positive drug fluoxetine and buspirone, could significantly antagonize the activation of YL-0919 to AC. At the overall level, YL-0919 (1.25-2.5mg/kg, i.g.) and Vilazzo ketone (4mg/kg, i.g.) can enhance the activity of forskolin activated in the hippocampus and the prefrontal cortex, and enhance the activity of PKA in the hippocampus of mice, and increase the expression of BDNF, p-CREB, p-ERK in the hippocampus of mice. Further studies have proved that selective PKA blockers are injected into the ventricle. -89 can completely block the antidepressant activity of a single dose of YL-0919 (2.5mg/kg, i.g.) on mouse tail and mouse forced swimming. These results suggest that activating the 5-HT1A receptor and enhancing the function of the cAMP-CREB signaling pathway is an important mechanism for the antidepressant effect of YL-0919.
Conclusion: YL-0919 has high affinity to SERT protein and 5-HT1A receptor protein and this binding has high selectivity; YL-0919 has strong inhibitory function of 5-HT reuptake, and is stronger than fluoxetine and duloxetine; YL-0919 single, subchronic or chronic administration in multiple animal models, showing significant antidepressant. Anti anxiety effect; YL-0919 has antidepressant, anti anxiety activity with Vera R, but lower starting dose; activating 5-HT1A receptor function, enhancing the function of the prefrontal cortex, hippocampal cAMP-CREB signal pathway, up regulating the BDNF level of the hippocampus and promoting neuronutrition is an important mechanism of YL-0919 antidepressant and anti anxiety activity.
【學位授予單位】：中國人民解放軍軍事醫(yī)學科學院
【學位級別】：博士
【學位授予年份】：2014
【分類號】：R965

【共引文獻】

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本文編號：2077550