CN113559143A - 一种用于抗肿瘤转移的中药组合物及其用途 - Google Patents
一种用于抗肿瘤转移的中药组合物及其用途 Download PDFInfo
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Abstract
本发明公开了提供一种用于抗肿瘤转移的中药组合物以及其制药用途，其由防风、黄芪、白术等制成，其能够有效预防或治疗恶性肿瘤的转移，尤其是在临床治疗过程中合并应用对乙酰氨基酚等非甾体类抗炎药时，对其诱导的肿瘤转移具有明显抑制作用。
Description
技术领域
本发明属于中药领域，具体地涉及一种用于抗肿瘤转移的中药组合物及其用途。
背景技术
西医学认为肿瘤转移是一个动态的极其复杂的过程，一部分肿瘤细胞在获得运动性之后从原发肿瘤脱落，内渗进入人体的脉管系统，随血液或者淋巴液向远处传播，并在特定情况下从血管或者淋巴管内皮外渗到达继发位置，增殖形成转移灶。且因恶性肿瘤易发生转移，而给临床用药、手术治疗等都带来极大困难，以致超过90％的恶性肿瘤患者死于肿瘤转移。中医学中虽未提到“肿瘤转移”这一名词，但对肿瘤转移的规律早有认识，如《灵枢·百病始生》。叶乃菁等通过研究发现三焦和经络系统一样，同为癌毒转移的通道。张晓梅等认为连续性的三焦管道是内生肿瘤从一个脏腑影响到多个脏腑的途径。因为三焦作为一个弥漫全身的多器官、组织及组织间隙构成的管道系统，不仅是五脏六腑之间联系沟通的共同通道，也是肿瘤细胞在原发肿瘤处脱落向远处转移的必经通道。郭子瑗等认为肿瘤炎性微环境不仅损伤人体的正气，而且会打破体内气血津液的健康稳定的动态平衡，逐渐形成一个有利于癌邪生存发展的内环境。而三焦膜性管道正常与否在改善该炎症微环境方面可以发挥重要的作用。如果三焦通路异常，一方面，充足的气血津液无法通过狭窄或拥堵的三焦管道到达发生炎性微环境的地带，不能对损伤组织及血管发挥及时的修复及濡养功能，新的组织无法生成。另一方面，不能及时代谢由于局部炎症导致的气血津液运行不畅而产生的瘀血、痰浊、郁热等病理产物及坏死组织废物，导致这些蓄积的病理产物在缺血缺氧的环境下诱发新的炎症，加重原来的炎症程度，扩大炎症范围，造成长时间的炎症刺激，使原有的微环境的稳定性被打破，促进肿瘤细胞转移侵袭的微环境形成。所以保证三焦通道的正常对于改善肿瘤微环境具有深远的意义。中医学普遍认为，从三焦的角度防治肿瘤的关键在于三焦气机的调护，杨亚萍认为平人防治肿瘤重在通过调节情志来保持三焦气化正常，达到体内和谐状态。赵书艺报道肿瘤转移与血瘀证互为因果，血瘀证可促进肿瘤转移，重视活血化瘀药的选用。因此对于治疗肿瘤患者肿瘤转移时，通常采用化瘀、化痰、利湿的主要治则。
在恶性肿瘤进展阶段，由于肿瘤细胞异常的代谢所产生内生性质热源，大约有一半以上的患者会伴有发热、癌痛等现象，治疗多采用物理降温、非甾体类抗炎药以及糖皮质激素为主，对乙酰氨基酚作为临床常用解热镇痛抗炎药。研究还发现70岁及以上的人服用阿司匹林后，无论是局部癌症还是晚期癌症的死亡率都较高，这表明阿司匹林可能对老年人的癌症演变产生不利影响。McNeil等人研究发现服用阿司匹林(ASPREE)的癌症患者能够增加转移性癌症的发生，同时阿司匹林也显著增加了癌症患者的死亡风险，其潜在的机制在于免疫功能低下，这是诱导老年人恶性细胞生长和转移的关键因素。
CN102266434A公开了由三味药物五月茶、油桐叶、瑞香根组成，具有解毒散结，活血行气功效，针对肿瘤患者后期的“瘀、毒”病理状态，以解毒散结、活血祛瘀的治则配伍中药组合物，起到抗肿瘤转移作用。CN103877190A公开了由白术、白茯苓、猫爪草、藤梨根、石上柏、仙鹤草、绞股蓝、凌霄花和石见穿组成的中药具有解毒散结，活血益气健脾功效，针对肿瘤患者后期的“瘀、毒、虚”病理状态，以解毒散结、活血祛瘀、益气健脾的治则配伍，起到抗肿瘤转移作用。如何延长肿瘤患者的生命，提高病人的生活质量，寻找更加高效安全的药物治疗尤为重要。中医认为正气不足是恶性肿瘤发生、发展的内在原因和结果，临床常用扶正解表方剂(体虚兼外邪的表证)，故常在活血化瘀解毒的基础上配伍益气、滋阴、助阳、养血等来配伍方剂。可见目前的中药复方更多关注肿瘤本身的治疗作用，而对恶性肿瘤转移的进展关注不多，尤其是在肿瘤治疗过程中合并应用非甾体抗炎类药加速肿瘤转移的问题，缺乏有效的解决手段，过于强调活血化瘀解毒药物的配伍。
发明内容
本发明的目的是克服现有技术的不足，提供一种用于抗肿瘤转移的中药组合物以及其制药用途，其能够有效预防或治疗恶性肿瘤的转移，尤其是在临床治疗过程中合并应用对乙酰氨基酚等非甾体类抗炎药时，对其诱导的肿瘤转移具有明显抑制作用。
本发明的技术方案概述如下：
一种中药组合物在制备防治肿瘤转移药物中的用途，所述中药组合物按重量份数由下述原料制成：防风10-50份，炙黄芪30-120份、白术40-90份。
优选其重量份为：防风20-40份，炙黄芪60-120份、白术50-80份。
优选其重量份为：防风30份，炙黄芪60份、白术60份。
本发明中药还进一步包括大枣10-40份，优选20-30份。
所述中药组合物制备方法包括以下步骤：将各原料药粉碎后，加水或低级醇或低级醇和水的混合溶剂提取。
所述低级醇选自甲醇、乙醇、丙醇、丁醇中的一种或多种，优选乙醇，优选30-90％乙醇，优选40％、50％、60％、70％、80％乙醇。
所述中药组合物的剂型选自散剂、汤剂、合剂、丸剂、颗粒剂、胶囊、片剂、配方颗粒中的一种。
所述肿瘤转移为恶性肿瘤的肺转移，尤其是对于非甾体抗炎药物合并用药相关的肿瘤的肺转移，或非甾体抗炎药物合并用药诱导的肿瘤肺转移。
所述非甾体抗炎药物选自对乙酰氨基酚、布洛芬、阿司匹林中的一种或多种。
发明采用的中药经典方剂玉屏风散，其为托里固表，用于表虚自汗、风邪等疾病，《医方考》记载卫气一亏，则不足以固津液.而自渗泄矣，此自汗之由也。白术、黄芪所以益气，然甘者性缓，不能速达于表，故佐之以防风。东垣有言，黄芪得防风而功愈大，乃相畏相使者也。是自汗也，与伤风自汗不同，伤风自汗责之邪气实；杂证自汗责之正气虚，虚实不同，攻补亦异。《古今名医方论》柯韵伯：防风遍行周身，称治风之仙药，上清头面七窍，内除骨节疼痹、四肢挛急，为风药中之润剂，治风独取此味，任重功专矣。然卫气者，所以温分肉而充皮肤，肥腠理而司开阚。惟黄芪能补三焦而实卫，为玄府御风之关键，且无汗能发，有汗能止，功同桂枝，故又能治头目风热、大风癞疾、肠风下血、妇人子脏风，是补剂中之风药也。所以防风得黄芪，其功愈大耳。白术健脾胃，温分肉，培土即以宁风也。夫以防风之善驱风，得黄芪以固表，则外有所卫，得白术以固里，则内有所据，风邪去而不复来，当倚如屏，珍如玉也。《医方论》：此固表去风药，用以实表则可。若云加减即可代桂枝、麻黄等汤，则表实而邪无出路，断断不可。此等议论，误人不浅必，不可从。又《医方发挥》记载本方主治表虚卫阳不固的自汗证.当以益气固表止汗为主。故用黄芪甘温益气，大补脾肺为主；辅以白术健脾益气，固表止汗，且脾气旺则土能生金，肺气足则可固表实卫。故黄芪得白术则益气健脾，固表止汗之力更著。二药合用，能补中以资气血之源，使脾胃健旺，肌表充实，则邪不易侵，汗亦不能外泄；自汗虽属于表虚不固，风邪乘虚扰其卫阳亦为致病因素之一，故配伍防风走表而祛风邪，合黄芪、白术以益气散邪，且防风配黄芪一散表，一固表，二药合用，黄芪得防风则固表而不留邪，防风得黄芪则祛邪而不伤正，取相畏而相使的作用。三药合用，实系补中有疏，散中寓补之意，即可用于卫气不固的自汗，亦可用以实表而御风邪。可见，黄芪、白术与防风的配伍中相畏又相使，要发挥临床效果需在表虚的情况下应用，而对于表实邪者，反而会加重病情。虽然目前报道，本发明中药具有抗肿瘤作用，而对于肿瘤转移的作用未有发现报道。本发明在研究玉屏风散抗肿瘤过程中，还发现其具有显著的抗肿瘤转移作用。
说明书附图：
图1本发明中药对肺组织病理形态
图2本发明中药的肺转移的HE染色图(50X)
图3本发明中药的肺组织免疫组化染色结果
有益效果：本发明从中医的整体理论出发，发现本发明中药能够有效预防或治疗恶性肿瘤的转移，尤其是在临床治疗过程中合并应用对乙酰氨基酚等非甾体类抗炎药时，对其诱导的肿瘤转移具有明显抑制作用。
具体实施方式
下面结合具体实施例对本发明作进一步的说明。
实施例1
取防风30g、蜜炙黄芪、白术各60g，加12倍量水常温浸泡30min，加热回流煎煮1.5h，滤过，将药渣第二次煎煮，加入10倍量水煎煮1h，滤过，将药渣第三次煎煮，加入8倍量水煎煮1h，滤过，将滤液合并，加热浓缩成浸膏后，干燥、粉碎过筛，加入糊精湿法制粒，干燥即得颗粒剂。
实施例2
取防风30g、蜜炙黄芪90g、白术各60g，加15倍量水常温浸泡30min，加热回流煎煮2h，滤过，将药渣第二次煎煮，加入12倍量水煎煮1h，滤过，将药渣第三次煎煮，加入8倍量水煎煮1h，滤过，将滤液合并，加热浓缩成浸膏后，干燥、粉碎过筛，制成散剂。
实施例3
取防风30g、蜜炙黄芪、白术各60g，加入10倍量的80％乙醇回流提取1.5h，滤过，将药渣第二次煎煮，加入10倍量的70％乙醇回流提取1h，滤过，将滤液合并，加热浓缩成浸膏后，干燥、粉碎过筛，加入微晶纤维素和蔗糖湿法制粒，再加入硬脂酸镁压制成片剂。
实施例4
取防风40g、蜜炙黄芪60g、白术各90g，其中防风加12倍量的80％乙醇回流提取两次，过滤取滤液，防风药渣和剩余的黄芪、白术加15倍量水加热回流煎煮2h，滤过，将药渣第二次煎煮，加入10倍量水煎煮1h，滤过，将滤液全部合并，加热浓缩后，加入甜味剂制成合剂。
实施例5
取防风30g、蜜炙黄芪、白术各60g，加入10倍量的80％乙醇回流提取1.5h，滤过，将药渣第二次煎煮，加入10倍量的70％乙醇回流提取1h，滤过，将滤液合并，加热浓缩成浸膏后，干燥、粉碎过筛，加入淀粉湿法制粒，干燥，填充至胶囊，制得胶囊剂。
实施例6
取防风30g、蜜炙黄芪、白术各60g、大枣30g，加12倍量水常温浸泡45min，加热回流煎煮2h，滤过，将药渣第二次煎煮，加入10倍量水煎煮1h，滤过，将药渣第三次煎煮，加入8倍量水煎煮1h，滤过，将滤液合并，加热浓缩成浸膏后，干燥、粉碎过筛，加入糊精湿法制粒，干燥即得颗粒剂。
实施例7
取防风40g、蜜炙黄芪60g、白术各90g、大枣40g，其中防风加12倍量的80％乙醇回流提取两次，过滤取滤液，防风药渣和剩余的黄芪、白术加10倍量水加热回流煎煮2h，滤过，将药渣第二次煎煮，加入10倍量水煎煮1h，滤过，将滤液全部合并，加热浓缩后，加入甜味剂制成合剂。
实验例1
荷瘤模型小鼠的制备：在无菌条件下，用含10％胎牛血清的DMEM培养Lewis肺癌细胞，传代3次后，将培养后的癌细胞，用无菌的PBS调整细胞密度至1×107个/ml，接种至5只小鼠的右腋处0.2ml(即右前肢腋窝靠背部0.3-0.5cm处，常规使用碘伏棉球消毒)。注射完毕后，停留数10秒左右，用无菌棉球轻轻按压进针点。然后放入新垫料的笼具中，次日观察小鼠的精神状态、活动以及进食情况，并观察注射部位有无红肿。
在第10天左右，瘤体积长至约1000mm3时，脱颈后处死，无菌条件下取生长良好的肺癌组织(粉红色的瘤组织)，并将周围非肿瘤组织剪除干净，然后将之剪碎(剪得越小越好)，按瘤重(g)：生理盐水(ml)＝1：3制成匀浆，200目细胞筛过滤，得瘤细胞混悬液，1000r/min，4℃离心5min，弃除上层液体，留下细胞沉淀。再用生理盐水稀释细胞，吹打均匀，经台盼蓝染液染色后并细胞计数，当活细胞比例大于90％，调整细胞浓度为1×107/mL，制得肺癌细胞悬液，最后接种于40只小鼠的右腋皮下处。其中对乙酰氨基酚诱导的高转移模型，需预先连续3天，给予对乙酰氨基酚(0.26g/kg)，每天间隔4h，分2次造模，再将瘤细胞接种至小鼠的右腋处，建立乙酰氨基酚诱导的荷瘤高转移模型。
观察到有肿瘤形成时，采用随机数字表法进行随机分组，每组10只，共分为5组，分别为正常组(NC)、荷瘤组(MOD)、荷瘤治疗组(MOD_YPF)；对乙酰氨基酚诱导的高转移荷瘤模型，则分为对乙酰氨基酚荷瘤组(MOD_AP)和对乙酰氨基酚荷瘤治疗组(MOD_AP_YPF)。其中正常组(NC)和荷瘤组(MOD)给予蒸馏水，对乙酰氨基酚(AP)诱导的高转移模型，在接种瘤细胞后，为了维持AP对上皮屏障功能的干扰，此后需每隔3天，再连续3天给予对AP干预，一天2次，干预12天；荷瘤治疗组(MOD_YPF)和对乙酰氨基酚荷瘤治疗组(MOD_AP_YPF)给予40g/kg的实施例1药物，连续给药实施例1药物15天，每只0.1mL/10g。
表1 分组与给药
对乙酰氨基酚的配制：将对乙酰氨基酚片置于研钵中，研碎后加0.5％羧甲基纤维素钠(CMC-Na)制备含有260mg/kg(26mg/mL)的对乙酰氨基酚混悬液，每日需要现配现用。
实施例1药物的配制：精密称取实施例1的颗粒，蒸馏水溶解，制成40g/kg(4g/mL)的药物，每日需要现配现用。
每天观察小鼠的状态：毛发色泽、饮食、排便、精神状态、活动、聚集等现象。
结果表明，与NC相比，MOD组和MOD_AP组中小鼠精神状态不佳，毛发暗淡无光泽，活动次数减少；与MOD组相比，MOD_AP组中小鼠精神状态较差，毛发色泽暗淡，活动次数明显减少。但本发明中药给药干预后，MOD_YPF组和MOD_AP_YPF组中小鼠精神状态良好，毛发稍有光泽。
肺组织表面转移灶比较：取出肺脏，经生理盐水漂洗后在显微镜下观察肺表面的肿瘤转移灶，并进行计数。转移灶计算方法：转移灶为类圆形小突起呈结节状、半透明点状，部分转移灶可能融合到一起。根据肺结节直径将其分为四级：I级＜0.5mm，0.5mm≤Ⅱ级≤1mm，1mm≤Ⅲ级≤2mm，IV≥2mm，肺表面转移结节总数＝I×1+Ⅱ×2+Ⅲ×3+Ⅳ×4。
肺转移抑制率(％)＝(荷瘤模型组平均肺表面转移结节数-治疗组平均肺表面转移结节数)/荷瘤模型组平均肺转移结节数×100％。
表2 对Lewis肺癌荷瘤小鼠肺转移的影响(
n＝10)
(注:与MOD相比,*P＜0.05；与MOD_AP相比,#P＜0.05)
由图1和表2可知：与NC组相比，MOD组和MOD_AP组中肺组织均含有肿瘤转移灶(箭头所示)；与MOD组相比，MOD_AP组中肺组织表面含有明显的肿瘤转移灶(箭头所示)，表明对乙酰氨基酚存在加速肺癌转移的风险；经本发明中药给药治疗后，MOD_YPF组和MOD_AP_YPF组中肺组织均未发现肿瘤转移灶。因此，本发明中药能够显著抑制肿瘤的肺转移，而且对于对乙酰氨基酚诱导加速的肿瘤肺转移有显著的抑制作用。
实验例2
肺组织HE染色
将上述实验例1中小鼠的左肺放入4％多聚甲醛固定，常规石蜡包埋、切片，苏木-伊红染色，具体步骤如下：(1)石蜡切片脱蜡至水，二甲苯I：20min 二甲苯Ⅱ：20min 无水乙醇I：5min 无水乙醇Ⅱ：5min 75％乙醇：5min 自来冲洗：30s。(2)苏木素染色：苏木素染色：5min 自来水洗：30s 1％盐酸-酒精分化：15s 自来水洗：30s 返蓝液染色：1min流水冲洗：30s。(3)伊红染色：将切片依次放入85％乙醇：5min 95％乙醇：5min，再入伊红染液中染色：5min流水冲洗：30s。(4)脱水封片：切片依次放入无水乙醇I：5min无水乙醇II：5min 无水乙醇Ⅲ：5min 二甲苯I：5min 二甲苯Ⅱ：5min，透明后，采用中性树胶封片。在50倍镜下观察肺组织的病理及微小肿瘤转移灶。
肺组织免疫组化染色
常规石蜡切片脱蜡至水，二甲苯I：15min，二甲苯Ⅱ：15min，二甲苯III：15min，无水乙醇I:5min，无水乙醇Ⅱ:5min，85％乙醇：5min，75％乙醇：5min，蒸馏水洗30s；柠檬酸缓冲液进行抗原修复，采用微波炉中火8min，停火8min，再低火7min；3％H2O2溶液室温避光孵育25min，洗涤3次，每次5min；3％BSA室温封闭30min；轻轻甩掉封闭液，滴加一抗：癌胚抗原(1：200)，切片平放于湿盒内，4℃孵育过夜；第二天，洗涤3次，每次5min；滴加HRP标记的山羊抗兔(1:50)，室温孵育50min；洗涤3次，每次5min；DAB显色；苏木素染液复染细胞核，脱水封片；显微镜下镜检左肺组织中癌胚抗原的表达量。
实验结果表明，图2所示，与NC组相比，MOD组和MOD_AP组中左肺组织含有微小肿瘤转移灶；与MOD组相比，MOD_AP组中左肺组织一侧可见肿瘤转移灶(箭头)，与邻近组织有明显的分界；本发明中药给药治疗后，MOD_YPF组和MOD_AP_YPF组中的肺组织大体形态有所恢复，具有显著的治疗效果。
肺组织免疫组化染色结果如图3所示，与NC组相比，MOD组和MOD_AP组中肺组织均含有明显的癌胚抗原表达；与MOD组相比，MOD_AP组中肺组织的免疫组化染色相对偏深，以棕黄色为主；本发明中药给药治疗后，MOD_YPF组和MOD_AP_YPF组中肺组织含有的癌胚抗原表达量均有所减少，说明本发明中药可以缓解肿瘤的肺部转移。
Claims (10)
1.一种中药组合物在制备防治肿瘤转移药物中的用途，其特征在于：所述中药组合物按重量份数由下述原料制成：防风10-50份，炙黄芪30-120份、白术40-90份。
2.如权利要求1所述的用途，其特征在于：其重量份为：防风20-40份，炙黄芪60-120份、白术50-80份。
3.如权利要求1所述的用途，其特征在于：其重量份为：防风30份，炙黄芪60份、白术60份。
4.一种中药组合物在制备防治肿瘤转移药物中的用途，其特征在于：所述中药组合物按重量份数由下述原料制成：防风10-50份，炙黄芪30-120份、白术40-90份、大枣10-40份。
5.如权利要求1-4任意一项所述的用途，其特征在于：所述中药组合物制备方法包括以下步骤：将各原料药粉碎后，加水或低级醇或低级醇和水的混合溶剂提取。
6.如权利要求5所述的用途，其特征在于：所述低级醇选自甲醇、乙醇、丙醇、丁醇中的一种或多种，优选乙醇。
7.如权利要求1-5任意一项所述的用途，其特征在于：所述中药组合物的剂型选自散剂、汤剂、合剂、丸剂、颗粒剂、胶囊、片剂、配方颗粒中的一种。
8.根据权利要求1-5任意一项所述的用途，其特征在于：所述肿瘤转移为恶性肿瘤的肺转移。
9.根据权利要求1-5任意一项所述的用途，其特征在于：所述肿瘤转移为非甾体抗炎药物合并用药相关的肿瘤的肺转移。
10.根据权利要求8任意一项所述的用途，其特征在于：所述非甾体抗炎药物选自对乙酰氨基酚、布洛芬、阿司匹林中的一种或多种。
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