CN116211854A - 神经递质化合物ahn 1-055在制备抗肿瘤药物中的应用 - Google Patents
神经递质化合物ahn 1-055在制备抗肿瘤药物中的应用 Download PDFInfo
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Abstract
本发明提供神经递质化合物AHN 1‑055在制备抗肿瘤药物中的应用。属于医药技术领域。本发明结合体外细胞实验,如CCK8、EDU增殖染色以及Western Blot检测Caspases蛋白表达与切割情况等,发现AHN 1‑055(hydrochlride)对肝癌细胞有很强的抑制作用,能够抑制肝癌细胞增殖、促进凋亡,且对人肝正常细胞系没有明显的毒副作用。且经过数据库数据分析,发现药物的受体基因多巴胺转运蛋白在肿瘤样本中是高表达的,且随着肝癌的不同分期,多巴胺转运蛋白表达量逐步增加,符合其抑制剂的作用效果。因此,可将神经递质化合物AHN1‑055(hydrochlride)制备成治疗肝癌的药物。
Description
技术领域
本发明属于医药技术领域。
背景技术
原发性肝癌是世界范围内常见的恶性肿瘤之一,其发病率高,据估计,至2025年,每年将有超过100万人患肝癌。原发性肝癌以肝细胞癌(HCC)为主,约占总病例数的90%。乙型肝炎病毒(HBV)感染是HCC发展的最重要的风险因素,约占总病例数的50%。大部分患者在首诊时已处于中晚期阶段,预后差,五年生存率仅12.1%。肝癌晚期(尤其HCC)患者生存时间非常短,治疗手段极为有限。目前,以索拉非尼(sorafenib)为代表的多激酶抑制剂打破了癌症需全身治疗的僵局,获批应用于临床的靶向药物还有乐伐替尼(Lenvatinib)、瑞戈非尼(regorafenib)、卡博替尼(cabozantinib)和雷莫芦单抗(ramucirumab,VEGFR2单抗)等。靶向治疗对于晚期HCC治疗具有里程碑式的意义,但其疗效仍有很大提升空间。
近年来,新药研发的成功率持续走低,成本却与日俱增。为此,将高通量筛选和高内涵筛选(HCS,High-content screening)结合,希望能走出这种研发困境。高内涵以细胞为单位,可以进行多靶点分析,通过自动化细胞成像,综合生物信息学,对群体细胞表型进行定量分析,将细胞图像转化为数值数据。结合机器学习算法的高内涵筛选将广泛用于药物的研发。HCS在细胞活性、细胞周期、细胞迁移、毒性检测、受体蛋白转位、蛋白相互作用等许多方面都有很好的应用,是药物筛选、癌症研究、心血管疾病研究、干细胞研究、神经细胞研究等领域的重要研究工具。有研究表明,通过高内涵筛选并确定一种候选药物,该候选药物可在体外和体内挽救突变的GLIS3相关β细胞死亡;利用siRNA库,通过高内涵筛选出早衰综合征驱动基因;利用cDNA库,通过高内涵筛选出组蛋白H4K20me1的去甲基酶;筛选2800个生物活性和结构多样化的化合物(包括748种上市药物),鉴定出调节人胚胎干细胞自我更新和分化的小分子化合物。
神经递质与肿瘤关系密切,神经系统在调节器官干细胞中处于核心位置,神经调节肿瘤微环境的作用同样强大,神经元促进大脑、皮肤、前列腺癌、胰腺癌和胃癌的生长。神经系统调节干细胞和前体细胞的功能。癌症和神经系统活动相互调节,促进大脑中的神经元兴奋性增高,促进非神经系统肿瘤生长出新的神经分支。随着科学家对神经系统在肿瘤微环境中影响的认识,已经为大脑、前列腺、胰腺、胃和皮肤的特殊癌症开辟了新的潜在治疗途径。随着更多的研究,针对神经元-癌细胞的相互作用可能是一个强大的治疗策略。斯坦福大学医学院的Michelle Monje等研究发现成人和儿童脑胶质瘤细胞在和高度活跃的神经元相邻时生长速度快,这些研究表明,不仅是神经附近的癌细胞生长快,而且还对神经元分泌的化学信号作出反应。哥伦比亚大学的Timothy Wang等研究表明神经元进入到肿瘤微环境中是胃癌发展的必要条件。也有研究表明多巴胺受体拮抗剂硫利达嗪选择性地靶向白血病干细胞,同时避免正常的造血干细胞。同时肠5-羟色胺能神经元产生5-羟色胺(5-HT)作为结直肠癌干细胞(CSC)自我更新的调节剂。大量研究表明神经递质与肿瘤之间存在密切联系。
神经递质化合物AHN 1-055(hydrochlride)是一种多巴胺摄取抑制剂,与多巴胺转运蛋白(DAT)具有高亲和力,目前研究将AHN 1-055(hydrochlride)应用于可卡因滥用问题上,未有将其作为抗肿瘤药物方面的相关研究。
发明内容
本发明目的在于提供神经递质化合物AHN 1-055在制备抗肿瘤药物中的应用。
基于上述目的,本发明采取以下技术方案:
神经递质化合物AHN 1-055在制备抗肿瘤药物中的应用。
所述神经递质化合物AHN 1-055的结构式为
所述神经递质化合物AHN 1-055用于制备抑制肿瘤细胞活性的药物、治疗与多巴胺转运蛋白高表达量相关的肿瘤的药物、抑制肿瘤细胞增殖的药物或诱导肿瘤细胞凋亡的药物。
所述肿瘤为肝癌。
所述抗肿瘤药物以神经递质化合物AHN 1-055为活性成分,还包括药学上可接受的辅料,例如稀释剂、赋形剂、填充剂、溶剂或囊封材料等。
在本申请中,采用溶剂对神经递质化合物AHN 1-055进行溶解后使用,溶剂为水、DMSO或水、DMSO的组合。
优选的,溶剂为DMSO。
本发明的抗肿瘤药物的剂型和施用方式没有特别限制。
代表性的施用方式包括但并不限于:口服、(静脉内、肌肉内或皮下)注射、和局部给药。
口服给药包括固体剂型和液体制剂,固体剂型包括胶囊剂、片剂、丸剂、散剂和颗粒剂。
液体制剂包括神经递质化合物AHN 1-055,还包括水或其它溶剂。
与现有技术相比,本发明具有以下有益效果:
本发明提供了神经递质化合物AHN 1-055(hydrochlride)在制备抗肝癌药物中的应用,结合体外细胞实验,如CCK8、EDU增殖染色以及Western Blot检测Caspases蛋白表达与切割情况等,发现AHN 1-055(hydrochlride)对肝癌细胞有很强的抑制作用,能够抑制肝癌细胞增殖、促进凋亡,且对人肝正常细胞系没有明显的毒副作用。且经过数据库数据分析,发现药物的受体基因多巴胺转运蛋白在肿瘤样本中是高表达的,且随着肝癌的不同分期,多巴胺转运蛋白表达量逐步增加,符合其抑制剂的作用效果。因此,可将神经递质化合物AHN1-055(hydrochlride)制备成治疗肝癌的药物,为HCC治疗提供新方案。
附图说明
为了更清楚地说明本发明的技术方案,下面将对实施例中所需要使用的附图作简单地介绍,显而易见地,下面描述中的附图仅仅是本发明的一些实施例,对于本领域技术人员来讲,在不付出创造性劳动的前提下,还可以根据这些附图获得其他的附图。
图1为高内涵神经递质化合物在Hepa1-6细胞系中的初步筛选结果;
图2为神经递质化合物AHN 1-055对肝癌细胞系及人肝正常细胞系影响结果分析图;
图3为神经递质化合物AHN 1-055对WRL68细胞影响结果分析图;
图4,a为基于样本类型的SLC6A3在LIHC中的表达,b基于肿瘤不同分期的SLC6A3在LIHC中的表达,c为多巴胺转运蛋白在肝癌中表达量与预后关系图;
图5,a为Hepa1-6细胞系在不同时间节点不同神经递质化合物AHN 1-055药物浓度的吸光度值,b为CCK8结果分析图;
图6,a为Huh7细胞系在不同时间节点不同神经递质化合物AHN 1-055药物浓度的吸光度值,b为CCK8结果分析图;
图7,a为神经递质化合物AHN 1-055对Huh7细胞系增殖影响荧光图,b为EdU结果示意图;
图8为神经递质化合物AHN 1-055对Huh7细胞系凋亡影响,流式检测结果示意图,右图为Desloratadine(地氯雷他定)阳性对照;
图9为神经递质化合物AHN 1-055对Huh7细胞系凋亡影响,Western Blot检测结果示意图。
各图中涉及的统计学标识含义:*P<0.05,**P<0.01,***P<0.001。
具体实施方式
为使本发明的目的、技术方案和优点更加清楚,下面对本发明的技术方案进行详细描述,但下述实施例仅仅是本发明的一部分实施例,而不是全部的实施例。基于本发明中的实施例,本领域技术人员在没有做出创造性劳动的前提下所得到的所有其它实施方式,都属于本发明所保护的范围。
实施例1
神经递质化合物AHN 1-055在制备抗肝癌药物中的应用,抗肝癌药物为抑制肿瘤细胞活性的药物。神经递质化合物AHN 1-055作用于肝癌细胞时,能够抑制肿瘤细胞活性。
所述神经递质化合物AHN 1-055的结构式为
实施例2
神经递质化合物AHN 1-055用于制备治疗与多巴胺转运蛋白基因SLC6A3高表达量相关的肿瘤的药物。神经递质化合物AHN 1-055能够抑制巴胺转运蛋白。
实施例3
神经递质化合物AHN 1-055用于制备抑制肝癌细胞增殖的药物。神经递质化合物AHN 1-055作用于肿瘤细胞时,能够抑制肿瘤细胞增殖。
实施例4
神经递质化合物AHN 1-055用于制备诱导肝癌细胞凋亡的药物,是指神经递质化合物AHN 1-055作用于肿瘤细胞时,能够诱导肿瘤细胞凋亡。
实施例5
如无特别说明,本申请中所用试剂、生物材料、培养基和溶液均为本领域常用、公众可以得到或市售,或者通过常规制备能够得到的物品。
在本申请中,神经递质化合物AHN 1-055先用DMSO溶解,实验过程中用再用DMEM培养基稀释至相应的药物浓度。如,5mg神经递质化合物AHN 1-055加入1.3162mL DMSO溶解,此时,药物储备液的浓度为10mM(mM为浓度单位,mmol/L),实验过程中使用DMEM培养基进行稀释至相应浓度即可。
5.1前期神经递质化合物筛选及其他肝癌细胞系以及人肝正常细胞系的验证
试验方法:1、鼠源Hepa1-6细胞系以2000个细胞/孔铺到96孔板中,每孔中药物浓度为10μM,放置细胞培养箱共孵育48h;
2、PBS洗涤细胞2-3次,hochest33342活细胞核染料以1:5000稀释后每孔加入100μL,避光染色30min;
3、调整高内涵参数,拍照分析,对每孔中活细胞计数。
数据分析,加药组的细胞个数/对照组细胞数×100%,计算细胞存活率(即细胞活力,用于确定细胞的总体健康、优化培养或实验条件,以及测量经化合物处理后的细胞存活);在其他细胞系上的验证方式和上述方法一致。
本发明首先在鼠源Hepa1-6肝癌细胞系上,从665种神经递质化合物中初步筛选出27种抑制细胞生长及影响生存的神经递质化合物,再进一步的验证。将化合物对肿瘤抑制程度排序,如图1所示,图中,编号7是神经递质化合物AHN1-055(由于筛选出的27种神经递质化合物,还有待进一步做相关科研研究,因此,出于技术保密需要,在本申请中不方便披露图中其余编号相对应的神经递质化合物)。
由图得知,神经递质化合物AHN 1-055细胞活力为0.35(35%),综合考虑上述神经递质化合物在后续试验过程中对几种不同肝细胞癌细胞系的作用,以及抑制肝癌细胞系生长的效果,确定神经递质化合物AHN 1-055为研究对象。
将神经递质化合物AHN 1-055(hydrochlride)在人源肝癌细胞系Hepa1-6、HepG2、Hep3B、PLC/PRF/5以及Huh7上进行验证,结果如图2所示。
人源和鼠源肝癌细胞系购自ATCC;WRL68为人肝正常细胞系,来自郑州大学基础医学院。培养、复苏/传代细胞均采取现有技术。
由图2可知,神经递质化合物AHN 1-055(hydrochlride)对人源肝癌细胞系具有显著的抑制效果。
将神经递质化合物AHN 1-055(hydrochlride)在人肝正常细胞系WRL68上进行验证,以同样的浓度处理,结果如图3所示。
由图3得知,本发明以人肝正常细胞系WRL68为研究对象,发现神经递质化合物AHN1-055(hydrochlride)对正常细胞系的作用不明显,也即无明显副作用。5.2多巴胺转运蛋白基因SLC6A3在肝癌细胞系、肿瘤和正常样本中的表达情况以及预后相关情况
1、登录Depmap数据库,输入SLC6A3基因进行检索,检索完成后,点击肿瘤类型选项选择肝癌,页面选择DATA plot进行数据下载,选择相应的细胞系进行分析;
2、登录UALCAN数据库,点击癌症基因,TCGA数据库,输入SLC6A3基因,选择肝癌,分析得到SLC6A3基因在正常和肿瘤样本中的表达量。结果如图4所示。
由图4a、b、c得知,本发明进行查询Depmap数据库,发现多巴胺转运蛋白基因SLC6A3在多种肝癌细胞系上都有表达,并且进行qPCR验证。另外又进行TCGA数据库进行查询基因在肿瘤和正常样本中表达量差异,发现相较于正常组织基因,SLC6A3在肿瘤样本中是高表达的,符合筛选到的药物是多巴胺转运蛋白抑制剂,且随肿瘤分期表达量逐渐增加,且与预后相关。
5.3神经递质化合物AHN 1-055(hydrochlride)对肿瘤细胞系生长的影响及其IC50测定
1、首先设置不同的药物浓度(神经递质化合物AHN 1-055药物浓度)分别为0(空白组),0.01μM(10nM)、0.1μM(100nM)、1μM、2μM、4μM、6μM、8μM、10μM这9个浓度梯度,每个浓度梯度6个重复;
2、96孔板2000个细胞/孔铺板,空白组为DMEM培养基(含血清)无细胞,对照组为1%DMSO;
3、实验设置6h、24h、48h、72h、96h 5个时间点,每孔加入10μL的cck8溶液在培养箱中孵育20min,用酶标仪(波长450nm)测定吸光度值;
4、数据分析。
肿瘤细胞系的细胞存活率(细胞活力)=(加药组吸光度值-空白组吸光度值)/(对照组吸光度值-空白组吸光度值)×100%,IC50值在作图软件GraphPad Prime8中分析计算。
以Hepa1-6、Huh7细胞系为研究对象,结果分别如图5和图6所示。其中,图5a为Hepa1-6细胞系在不同时间节点不同药物浓度的吸光度值(基于不同浓度折线图更好区分考虑,试验数据原图中0.01μM浓度与空白组折线图接近,因此,在该图中未显示0.01μM对应的折线图,图6a同理),5b为CCK8结果分析图;图6以Huh7细胞系为研究对象,6a为Huh7细胞系在不同时间节点不同药物浓度的吸光度值,6b为CCK8结果分析图。
由上图得知,本申请以Hepa1-6、Huh7细胞系为研究对象,进行CCK8实验,计算AHN1-055(hydrochlride)的IC50值,96h的IC50为5.9μM,由此说明,神经递质化合物AHN 1-055能在较低浓度下发挥杀伤肝癌细胞作用。
5.4神经递质化合物AHN 1-055(hydrochlride)对细胞增殖的影响
选择以Huh7细胞系为研究对象。
试验方法:
1、96孔板,2000个细胞/孔,实验组每孔的药物浓度6μM;细胞与AHN1-055(hydrochlride)培养箱共孵育48h;
2、加入37℃EdU试剂,室温孵育2h;
3、EdU标记完成后,去除原培养基,加入4%多聚甲醛,室温固定10min;
4、去除固定液,加PBS洗涤3次,每次5min;
5、去除洗涤液,加入0.3% Triton X-100的PBS,室温孵育10-15min;透化完成后洗涤2-3次;
6、按照96孔板的大小加入50μL的Click反应液,室温孵育30min;
7、去除Click反应液,PBS洗涤2次每次5min;
8、再加入含1/20的DAPI核染料,孵育20min;高内涵选择488nm荧光波长及450nm进行拍照,并分析DAPI蓝光(细胞核的数量)和FITC绿光(增殖细胞数量)所代表的细胞个数,荧光图如图7a所示(为便于分析,申请人将说明书附图7的彩色荧光图以其他证明文件的形式提交),增殖的细胞比例=绿色荧光细胞数量/蓝色荧光细胞数量×100%,如图7b所示。
由图7a和7b得知,本发明进行EDU增殖实验验证,AHN 1-055(hydrochlride)对Huh7肿瘤细胞系增殖的影响,AHN 1-055(hydrochlride)药物能够显著抑制细胞系的增殖。
5.5AHN 1-055(hydrochlride)影响的肝癌细胞系死亡方式
1、细胞凋亡流式检测
(1)以20万个/孔的细胞数铺到6孔板中,加药组浓度为6μM,药物与细胞共孵育48h;
(2)收集细胞,用PBS洗涤一次,加入100μL 1×Binding Buffer重悬;
(3)加入5μL Annexin V-Alexa Fluor 647(用于检测细胞凋亡的膜连蛋白-V偶联物)和10μL PI(碘化丙啶),轻轻混匀;室温下,避光反应15min;加入400μL1×BindingBuffer混匀,放置于冰上;
(4)Alexa Fluor647最大激发波长为651nm,最大发射波长为667nm;PI-DNA复合物的最大激发波长为535nm,最大发射波长为615nm,调节流式细胞仪的波长后,进行收集10000个细胞分析细胞凋亡情况。结果如图8所示(为便于分析,申请人将说明书附图8的彩色图以其他证明文件的形式提交)。
2、细胞凋亡Western Blot检测
(1)照流式检测细胞凋亡同样的步骤,药物与细胞孵育48h;
(2)收集实验和对照组细胞加入1×loading Buffer蛋白电泳上样缓冲液裂解,沸水浴15min;(3)按照对照组:实验组为5μL:25μL的上样量进行SDS-PAGE,条件为浓缩胶80V,30min,分离胶120V,1h;
(4)半干转膜仪进行18V,1h转膜;
(5)spases3蛋白及其切割条带的位置分别是35kDa、19kDa及17kDa,内参β-actin对应的45kDa,5%的脱脂牛奶封闭1h;
(6)洗涤3次每次5min;
(7)加一抗孵育2h;
(8)洗涤3次每次5min;
(9)加二抗孵育1h;
(10)洗涤3次每次5min;
(11)在膜上滴加适量的发光液进行显影。结果如图9所示。
由图8和9得知,本发明通过流式检测细胞凋亡,Western blot检测Caspases蛋白表达与切割情况,表面AHN 1-055(hydrochlride)能够诱导肝癌细胞凋亡。
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Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US5792775A (en) * | 1995-06-21 | 1998-08-11 | The United States Of America As Represented By The Department Of Health And Human Services | 4' and 4', 4"-substituted-3-α-(diphenylmethoxy) tropane analogs as cocaine therapeutics |
US20080226559A1 (en) * | 2005-08-24 | 2008-09-18 | Government Of The Usa,Represented By The Secretary Department Of Health And Human Services | Benztropine Compounds and Uses Thereof |
CN105853420A (zh) * | 2016-04-05 | 2016-08-17 | 中山大学 | 苯扎托品及其药学上可接受的盐的新应用 |
-
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- 2023-01-19 CN CN202310060898.0A patent/CN116211854B/zh active Active
Patent Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US5792775A (en) * | 1995-06-21 | 1998-08-11 | The United States Of America As Represented By The Department Of Health And Human Services | 4' and 4', 4"-substituted-3-α-(diphenylmethoxy) tropane analogs as cocaine therapeutics |
US20080226559A1 (en) * | 2005-08-24 | 2008-09-18 | Government Of The Usa,Represented By The Secretary Department Of Health And Human Services | Benztropine Compounds and Uses Thereof |
CN105853420A (zh) * | 2016-04-05 | 2016-08-17 | 中山大学 | 苯扎托品及其药学上可接受的盐的新应用 |
Non-Patent Citations (2)
Title |
---|
CHIHARU SOGAWA等: "Antiparkinson Drug Benztropine Suppresses Tumor Growth, Circulating Tumor Cells, and Metastasis by Acting on SLC6A3/DAT and Reducing STAT3", CANCERS, vol. 12, pages 523 * |
XIELING YIN等: "Screening and Validation of a Carvacrol-Targeting Viability-Regulating Protein, SLC6A3, in Liver Hepatocellular Carcinoma", DISEASE MARKERS, vol. 2022, pages 3736104 * |
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