关于“益今生”组分原人参二醇(PPD)的抗癌性研究

20(S)-原人参二醇药学与生物活性研究
王小峰;潘欣萍;徐建彬;陈伟;王玉芹
【期刊名称】《长春中医药大学学报》
【年(卷),期】2012(28)5
【摘要】@@%20(S)-原人参二醇是人参皂苷元中非常重要的一类,在治疗疾病及
保健方面,一直是研究热点.在抗肿瘤方面,20(S)-PPD可以通过增强机体免疫和直接的细胞毒作用来杀死肿瘤细胞,还可以抑制肿瘤间质血管生成,阻止肿瘤细胞的生长；在神经系统方面,20(S)-PPD可以抗癫痫、抗抑郁、增强学习能力等.20(S)-PPD是一个具有开发价值的化合物.
【总页数】3页(P911-913)
【作者】王小峰;潘欣萍;徐建彬;陈伟;王玉芹
【作者单位】上海中医药大学,上海201203;上海中药创新研究中心,上海201203;上海中药创新研究中心,上海201203;上海中药创新研究中心,上海201203;上海中医药大学,上海201203;上海中药创新研究中心,上海201203
【正文语种】中文
【中图分类】R284.2
【相关文献】
1.人参二醇的微生物转化及抗HSC-T6细胞活性研究 [J], 朱梦月;雷静;周垚;戴秋阳;朱腊梅;陈广通
2.人参二醇的微生物转化及抗HSC-T6细胞活性研究 [J], 朱梦月;雷静;周垚;戴秋
阳;朱腊梅;陈广通;
3.人参二醇的现代药学与生物活性研究进展 [J], 张贵明;赵余庆
4.原人参二醇及其衍生物的化学与抗癌活性研究进展 [J], 刘娜;朴虎日;李宁;赵余庆
5.人参二醇的现代药学与生物活性研究进展 [J], 刘林增
因版权原因，仅展示原文概要，查看原文内容请购买。

原人参二醇空心金纳米药物载体的制备及体外抗喉癌研究摘要原人参二醇〔PPD〕对多种肿瘤细胞具有抑制作用。

但由于水溶性差、利用度低，限制了其在临床上的应用。

基于此，本研究以空心金纳米颗粒为PPD的运输载体，合成PPD空心金纳米颗粒，采用高效液相色谱分析PPD空心金纳米药物载体的缓释效应性、MTT法检测对Hep-2细胞增殖的抑制作用、流式细胞术研究对Hep-2细胞凋亡的影响，以考察原人参二醇〔PPD〕空心金纳米药物载体的体外抗喉癌细胞Hep-2效应。

结果说明，PPD空心金纳米药物载体具有缓释效应性，与空白对照组、HAuNs组及PPD组相比，HAuNs-PEG-PPD组的Hep-2细胞生存率下降更显著，细胞凋亡率增加更显著〔p99%，吉林大学有机化学实验室合成〕；RPMI-1640培养基、胎牛血清〔美国Hyclone公司〕；二甲亚砜〔DMSO〕、青霉素和链霉素〔上海鼎国生物技术〕；MTT〔美國Sigma公司〕；TUNEL凋亡检测试剂盒〔瑞士罗氏公司〕。

实验用水为Milli-Q纯水仪〔美国Millipore公司〕制备的超纯水〔18.2MΩ·cm〕。

2.2空心金纳米颗粒〔HAuNs〕的合成参照文献[22]的方法合成空心金纳米颗粒〔HAuNs〕。

首先制备钴纳米颗粒，将氯化钴溶液〔0.1mL，0.4mol/L〕参加到含有新制备的硼氢化钠溶液〔0.1mL，1mol/L〕和柠檬酸钠溶液〔0.4mL，0.1mol/L〕中，混合溶液在氮气氛下反应60min。

将上述制备的无硼氢化钠的钴纳米颗粒溶液〔30mL〕注入脱氧氯金酸溶液〔10mL，1μmol/L〕中，在氮气氛下反应10min后，将溶液暴露于空气中至钴完全氧化。

以10000r/min离心，收集沉淀，用超纯水洗涤3次。

2.3HAuNs-PEG的合成通过配体交换法制备HAuNs-PEG[23]。

将合成的HAuNs分散在去离子水中〔1.0mg/mL〕，将150mgPEG-SH参加到40mL上述水溶液中，在室温下搅拌过夜。

20(S)-原人参二醇对肝癌生长的抑制作用及对癌细胞凋亡的影响秦俊杰;李永杰;付军;冷吉燕【期刊名称】《中国老年学杂志》【年(卷),期】2006(26)9【摘要】目的探讨原人参二醇(Ppd)抑制肿瘤生长和促进肿瘤细胞凋亡作用及其机制.方法建立肝癌动物模型,将实验动物分为五组;对照组、阳性药组、Ppd低剂量组(Ppd 25 mg/kg)、Ppd中剂量组(Ppd 50 mg/kg)、Ppd高剂量组(Ppd 100 mg/kg),2 w后处死动物测定肿瘤的重量和体积并做组织切片进行原位末端标记染色(TUNEL).结果 Ppd给药组肿瘤生长被抑制,其肿瘤体积、重量均较对照组减小(P ＜ 0.01),TUNEL 结果显示Ppd 100 mg/kg给药组细胞凋亡数量与对照组之间存在显著性差异(P＜ 0.01).结论 Ppd具有抑制肿瘤细胞的增殖的作用,其作用机制的可能是促进肿瘤细胞的凋亡.【总页数】2页(P1257-1258)【作者】秦俊杰;李永杰;付军;冷吉燕【作者单位】吉林大学第一医院,吉林,长春,130021;吉林大学第一医院,吉林,长春,130021;吉林大学第一医院,吉林,长春,130021;吉林大学第一医院,吉林,长春,130021【正文语种】中文【中图分类】R285.6【相关文献】1.20(s)-原人参二醇对肝癌血管内皮生长因子及碱性成纤维细胞生长因子蛋白表达的影响 [J], 冷吉燕;付军;王继萍2.20(S)-原人参二醇抑制肝癌血管内皮生长因子及其基因的表达 [J], 张国成;秦俊杰;冷吉燕3.20(s)-原人参二醇对前列腺癌RM-1细胞的生长抑制作用及其机制 [J], 郭亚雄;刘艳波;李德龙;韩向北;许多;程宏;赵丽晶;赵丽娟;董妍4.20(S)-原人参二醇对人胚肾293细胞生长的抑制作用 [J], 翟旭光;陈广通5.20(S)-原人参二醇对肺癌A549细胞增殖和荷瘤裸小鼠肿瘤生长的抑制作用 [J], 张锐;徐华丽;曲绍春;于小风;陈明侠;睢大赟因版权原因，仅展示原文概要，查看原文内容请购买。

20(s)-原人参二醇对体外培养宫颈癌Siha细胞的增殖抑制作用及其机制鲁育铭;赵丽晶;王贺彬;许多;赵丽娟;董妍【摘要】目的:观察20(s)-原人参二醇(PPD)对体外培养宫颈癌Siha细胞生长周期的作用及其对p53、p21及细胞周期素E(cyclin-E)基因转录和表达的影响,探讨PPD抑制Siha细胞增殖的机制.方法:体外培养人宫颈癌Siha细胞,将其分为阴性对照组(乙醇)和实验组(20 μg·L-1 PPD),分别用乙醇和PPD处理体外培养的宫颈癌Siha细胞,48 h后流式细胞术检测细胞周期,Real time PCR和Western blotting 法检测宫颈癌Siha细胞内p53、p21及cyclin-E mRNA及蛋白的表达情况.结果:与阴性对照组比较,20μg·L-1 PPD处理48 h后,G0/G1期Siha细胞比例增加(P＜0.01),G1期阻滞作用明显增强;Siha细胞中p53与p21 mRNA及蛋白表达水平上调(P＜0.01),cyclin-E mRNA及蛋白表达水平下调(P＜0.01).结论:PPD可抑制人宫颈癌Siha细胞增殖,其机制可能与上调p53、p21基因表达,下调cyclin-E基因表达有关.【期刊名称】《吉林大学学报（医学版）》【年(卷),期】2013(039)005【总页数】5页(P909-912,后插3)【关键词】20(s)-原人参二醇;Siha细胞;p53;p21;cyclin-E【作者】鲁育铭;赵丽晶;王贺彬;许多;赵丽娟;董妍【作者单位】吉林大学白求恩医学院病理生理学教研室,吉林长春130021;吉林大学白求恩医学院病理生理学教研室,吉林长春130021;吉林大学白求恩医学院病理生理学教研室,吉林长春130021;吉林大学白求恩医学院病理生理学教研室,吉林长春130021;吉林大学白求恩医学院病理生理学教研室,吉林长春130021;美国杜兰大学细胞生物学教研室,美国新奥尔良LA70062【正文语种】中文【中图分类】R737.33在发展中国家，子宫颈癌约占女性癌症的24%［1］。

人参二醇的现代药学与生物活性研究进展摘要：人参皂苷主要通过三种苷源组成，其中最典型的就是人参二醇，人参二醇具有相当高的生物活性，存在于多种植物当中，分析其生理活性和药理作用，这种物质具有广泛的药理作用，本研究主要分析人参二醇的现代药理和生物活性，希望所得内容能够为相关领域提供有价值的参考。

关键词：人参二醇；现代药学；生物活性最近这些年，因为对人参皂苷进行水解而得到的高活性的微量皂苷成分的研究实验开始不断进展，研究方向也主要围绕着酸碱水解和酶水解来进行。

人参二醇是一种三萜类化合物，所以人参皂苷当中，大多数原存形式都是人参二醇，在对其进行水检的时候，C-20位结构最容易受到酸的影响，这就会使它发生互变异构的情况，进而会生成以20（R）占优势的苷元混合物。

本研究据此主要分析人参二醇的现代药学和生物活性，结果取得较为满意的成效，现将主要研究情况作出如下综述。

1.人参二醇的现代药学研究进展人类对于人参的研究可以划分为两个阶段，在上个世纪六十年代以前，关于人参的研究进展较为缓慢，人们在研究的过程中，只能单凭感觉、直觉和直观的判断来进行，所以这阶段也称之为对人参进行研究的古代朴素的研究阶段。

现阶段，因为人类积累了大量的知识和临床经验，同时人们的各项技术也不断发展，所以人们对于人参的研究开始突飞猛进，浙江这一阶段的研究称之为现代科学研究阶段。

珍惜人参干燥类的化学成分，从上个世纪九十年代开始，一些学者在国外研究的基础上就对人参根和地面上其他的皂苷类成分进行了仔细的分离鉴定，单体皂苷的代谢化学和半合成、碱水点以及分析方法都进行了系统的研究[1]。

原人参二醇和人参三醇是人参皂苷当中的主要组成成分，本研究主要分析人参二醇，有研究对人参二醇进行分析，并以人参皂苷-Rb1和-Rb2为例，经过研究，可以得出人参皂苷-Rb1在胃中只有部分代谢的代谢产物和体外水解实验存在明显差异，它们在人类的胃肠当中会出现过氧化反应，过氧化反应所产生的过氧化物主要是Rb1的25-hydroperoxy-23-ene的衍生物[2]。

原人参二醇及Compound K抑制肝细胞肝癌的作用及机制张春蕾;宋海燕;许阳贤;季光【期刊名称】《中医学》【年(卷),期】2017(006)002【摘要】中晚期肝癌及术后复发转移性肝癌目前仍然缺乏有效治疗手段。

原人参二醇(protopanoxadiol, PPD)及人参皂苷Compound K是二醇型人参皂苷在体内的代谢产物。

近年来多项研究发现这两种天然产物可通过诱导细胞凋亡、抑制细胞增殖、抑制血管生成、抑制转移等机制对多种肿瘤发挥抑瘤效应。

其中对肝细胞肝癌的作用研究较多,抑癌效果尤为突出。

本文就近年来PPD及Compound K抗肝细胞肝癌的作用和相关机制研究进行系统综述,以期有助于开发出有效治疗肝癌的天然药物。

【总页数】6页(P104-109)【作者】张春蕾;宋海燕;许阳贤;季光【作者单位】[1]上海中医药大学附属龙华医院脾胃病研究所;;[1]上海中医药大学附属龙华医院脾胃病研究所;;[2]上海中医药大学附属龙华医院普外科;;[1]上海中医药大学附属龙华医院脾胃病研究所【正文语种】中文【中图分类】R73【相关文献】1.基因芯片技术分析20(S)-原人参二醇抑制人胃癌SGC-7901细胞作用机制 [J], 冷吉燕;李修英;张婧;秦俊杰2.20(s)-原人参二醇对体外培养宫颈癌Siha细胞的增殖抑制作用及其机制 [J], 鲁育铭;赵丽晶;王贺彬;许多;赵丽娟;董妍3.20(s)-原人参二醇对前列腺癌RM-1细胞的生长抑制作用及其机制 [J], 郭亚雄;刘艳波;李德龙;韩向北;许多;程宏;赵丽晶;赵丽娟;董妍4.丙泊酚抑制过氧化氢诱导细粒棘球蚴原头节凋亡作用机制的初步研究 [J], 马斌; 汤光耀; 王麟尧; 姜玉峰; 吕海龙5.丙泊酚抑制过氧化氢诱导细粒棘球蚴原头节凋亡作用机制的初步研究 [J], 马斌; 汤光耀; 王麟尧; 姜玉峰; 吕海龙因版权原因，仅展示原文概要，查看原文内容请购买。

20(S )-Protopanaxadiol (PPD)analogues chemosensitizemultidrug-resistant cancer cells to clinical anticancer drugsqJunhua Liu a , ,Xu Wang b , ,Peng Liu a ,Rongxin Deng b ,Min Lei a ,Wantao Chen b ,⇑,Lihong Hu a ,⇑a Shanghai Research Center for Modernization of Traditional Chinese Medicine,Shanghai Institute of Materia Medica,Chinese Academy of Sciences,Shanghai 201203,PR China bDepartment of Oral and Maxillofacial—Head and Neck Oncology,Ninth People’s Hospital,Shanghai Jiao Tong University,School of Medicine,Shanghai 200011,PR Chinaa r t i c l e i n f o Article history:Received 2April 2013Accepted 25April 2013Available online 3May 2013Keywords:20(S )-Protopanoxadiol Multidrug resistant P-glycoprotein ChemosensitizerStructure–activity relationshipa b s t r a c tNovel 20(S )-protopanoxadiol (PPD)analogues were designed,synthesized,and evaluated for the chemo-sensitizing activity against a multidrug resistant (MDR)cell line (KBvcr)overexpressing P-glycoprotein (P-gp).Structure–activity relationship analysis showed that aromatic substituted aliphatic amine at the 24-positions (groups V)effectively and significantly sensitized P-gp overexpressing multidrug resis-tant (MDR)cells to anticancer drugs,such as docetaxel (DOC),vincristine (VCR),and adriamycin (ADM).PPD derivatives 12and 18showed 1.3–2.6times more effective reversal ability than verapamil (VER)for DOC and VCR.Importantly,no cytotoxicity was observed by the active PPD analogues (5l M)against both non-MDR and MDR cells,suggesting that PPD analogues serve as novel lead compounds toward a potent and safe resistance modulator.Moreover,a preliminary mechanism study demonstrated that the chemo-sensitizing activity of PPD analogues results from inhibition of P-glycoprotein (P-gp)overexpressed in MDR cancer cells.Ó2013The Authors.Published by Elsevier Ltd.All rights reserved.1.IntroductionChemotherapeutic is an important tool in the treatment of can-cers.However,chemotherapy usually fails due to the development of tumor cell resistance to multiple drugs,a phenomenon known as multidrug resistance (MDR),1,2and essentially all cancer-related deaths are considered to be a result of chemotherapy failure.3MDR may originate from several biochemical mechanisms,but the major mechanism of drug resistance is the overexpression of active drug efflux transporters,such as ABCB1(also known as P-glycoprotein,P-gp)and ABCC1(multidrug resistance associate pro-tein 1,MRP 1).4,5In order to surmount MDR,a considerable re-search efforts have been put into developing clinically usable chemosensitizing agents over the past 2decades.Verapamil (VER)and cyclosporine A (CsA)are examples of first-generation chemosensitizers that inhibit the activity of P-gp and were evalu-ated clinically as adjuvants for chemotherapy.6,7Both compounds were precluded from clinical use because of significant toxicity at the dose required to attenuate P-gp funiction,8but are used inexperiments as positive controls.Since then,several potent and selective second-and third-generation chemosensitizers were developed and investigated.However,unsatisfactory toxicity and pharmacokinetic complications still impeded drug candidate development.Although several third-generation P-gp inhibitors,including tariquidar,are now in phase II cancer clinical trials,9their clinical efficiencies are not yet clear.10So far,no chemosensitizer has been approved for therapeutic use.Therefore,the discovery of safe and effective MDR modulators is still urgently needed to overcome the MDR of tumors.Ginsenosides are the active ingredients of Panax ginseng ,the root of which has been used in traditional herbal remedies/medi-cine in Eastern Asia for over 2000years.11It exhibits various phar-macological and physiological effects,including antioxidation,12immunostimulation,13antistress,14anticancer,15as well as stimu-lative effects on the central nervous system.16Within the ginseno-side family,ginsenoside Rg3(Fig.1)has been reported to reverse P-gp mediated multidrug resistance.1720(S )-Protopanoxadiol (PPD,Fig.1),an important metabolite of Rg3produced by intesti-nal bacteria,18has been paid more attention for its distinguished anti-cancer activity and significant inhibition of P-gp in tumor cells,19and extremely low toxicity.20–22Given extremely low toxicity of the compound,PPD is a potential candidate of chemo-sensitizer for treatment of multidrug resistant tumors.However,so far PPD analogues have not been prepared and reported,and their MDR reversal abilities and structure–activity relationship (SAR)studies have not been extensively performed.To explore more potent nontoxic MDR reversal analogues with lower effective0968-0896/$-see front matter Ó2013The Authors.Published by Elsevier Ltd.All rights reserved./10.1016/j.bmc.2013.04.067q This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial-No Derivative Works License,which per-mits non-commercial use,distribution,and reproduction in any medium,provided the original author and source are credited.⇑Corresponding authors.Tel.:+862163138341x5211;fax:+862163135412(W.C.);tel./fax:+862120231965(L.H.).E-mail addresses:chenwantao2003@ (W.Chen),lhhu@ (L.Hu).These authors contributed equally to this work.dosing and to study SAR,we designed and synthesized a series of PPD analogues.Herein,we report the chemosensitizing effects of newly synthesized PPD analogues.2.Design and synthesesOn the basis of previous literature,PPD had obvious effects on inhibition of P-gp and activation of MDR MCF-7/Adr.23–25Further-more,the predominant metabolic pathway of PPD observed was the oxidation of the 24,25-double bond.26,27Therefore,the 24,25-double bond was selected to modify in order to block the pathway of predominant metabolic.For the sake of more effective structural modifications at 24,25-double bond,we carefully analysed the structural properties of various known P-gp inhibitors,especially the first-,second-,and third-generation chemosensitizers (Fig.2),and found that most of P-gp inhibitors identified common features:(i)high hydrophobicity;(ii)two or more aromatic rings;(iii)a methoxy group on the aromatic ring (hydrogen bond acceptor);(iv)one or two protonatable nitrogens.28,29Building on this idea,we designed various amine-substituted PPD analogues 2–26(Scheme 1)in order to insert the nitrogen and the lipophilic moie-ties.The amine groups (R in Scheme 1)were selected by consider-ing size,hydrophobicity,and electron density.The diverse set included aliphatic acyclic amine (group I),aliphatic cyclic amine4280J.Liu et al./Bioorg.Med.Chem.21(2013)4279–4287(group II),heterocycle-aliphatic amine(group III),polar aliphatic amine(group IV),and aromatic substituted aliphatic amine(group V)groups,which could be transformed into water-soluble salts,if necessary.PPD werefirstly treated with O3,then the related amine (RNH2),and sodium triacetoxyborohydride(STAB)to produce2–26 (Scheme1).3.Results and discussion3.1.Evaluation of cytotoxicity and preliminary MDR reversal activity screeningBased on the preliminary experiment,PPD(5l M)did not exhi-bit cytotoxicity,but exhibit a weak MDR reversal effect in KBvcr cells.Thus,all synthesized compounds(5l M)were evaluated in a cytotoxic activity assay using two tumor cell lines,KB(epider-moid carcinoma of the nasopharynx),and a resistant subline,KBvcr (overexpression of P-gp selected using increasing concentrations of vincristine).Although compounds13,15,16,19,20and23were slightly cytotoxic,most of the PPD-derived compounds did not ex-hibit significant cytotoxicity,which implied low toxicity of these analogues.For evaluating chemosensitizing activity,KBvcr cells were co-treated with test compounds at5l M and the anticancer drug Docetaxel(DOC)(Fig.3).As shown in Figure3,PPD(1)did exhibit a weak MDR reversal effect at a concentration of5l M in KBvcr cells.PPD analogues with aliphatic acyclic amine(group I),ali-phatic cyclic amine(group II),heterocycle-aliphatic amine(group III),and polar aliphatic amine(group IV)did show an inactive activity(Fig.3).However,PPD analogues with aromatic–aliphatic amine substituents(group V)did show potent activity(Fig.3). The screening results suggested a rough SAR,including(a)the aro-J.Liu et al./Bioorg.Med.Chem.21(2013)4279–42874281matic substituted aliphatic amine (group V)tended to enhance the reversal ability of PPD analogues,but aliphatic acyclic amine (group I),aliphatic cyclic amine (group II),heterocycle-aliphatic amine (group III),and polar aliphatic amine (group IV)led to re-duced activity;(b)one-or two-carbon linear chain in the aromatic substituted aliphatic amine was effective in reversing -pounds (12,14,17,18,21,22,and 24–26)which contained one-or two-carbon linear chain displayed the most significant activity.But compound with 4-methoxy-benzenamine 11was inactive,and compound 23containing three-carbon linear chain showed low toxicity;(c)The electronic effect from the aromatic ring of the aro-matic substituted aliphatic amine had a significant impact on activity.For example,compounds with aromatic ring carrying elec-tron-withdrawing (15,16,19,and 20)showed low toxicity,but compounds with aromatic ring carrying electron-donating substit-uents (14,17,21,and 22)displayed the most significant activity;(d)Compounds with hetero-aromatic ring (24–26)also showed potent activity.From these results,the most active compounds (12,18,21,and 25)were selected for further investigation.3.2.Chemoreversal ability of PPD analogues with docetaxel (DOC),vincristine (VCR),and adriamycin (ADM)A quantitative evaluation of the reversal ability of PPD ana-logues 12,18,21,and 25was performed using MDR KBvcr cells with various concentrations of DOC,VCR,and ADM,which are clin-ically used and known as significant P-gp substrates,partly accounting for their resistance.The IC 50value of anticancer drugs in the presence of test compounds at 5l M concentration was cal-culated,and fold reversal was determined by dividing the IC 50of anticancer drug alone by the IC 50of anticancer drugs plus PPD ana-logue (Table 1).Most of the tested analogues,including VER,showed great reversal against DOC,VCR,and ADM resistance.Especially,compounds 12and 18were 1.3–2.6times more potent than the positive control VER for chemoreversal ability against DOC,VCR,and ADM resistance.The following SAR correlations were proposed based on the chemosensitizing effects against DOC,VCR,and pounds with aromatic ring carrying no substituents (12and 18)were more potent activity than com-pounds with hetero-aromatic ring (25)and aromatic ring carrying electron-donating substituents (21).On the basis of the above results,compounds 12and 18were selected for further evaluation of chemosensitizing efficacy.The dose–response proliferation inhibitory effects of DOC,VCR,and ADM at 5l M were analyzed against KB and KBvcr cells (Fig.4).In the absence of PPD compound or VER,KBvcr cells were resistant to all three anticancer drugs,resulting in IC 50values over 1000nM.When 5l M of compound 12or 18or VER was added,the sensitiv-ity of KBvcr cells to each anticancer drug was dramatically increased.The chemosensitizing efficacy of 12or 18was either similar or better than that of VER.These results demonstrated that 5l M of 12and 18effectively chemosensitized MDR cells.3.3.Dose–response effect of compounds 12and 18on sensitization of KBvcr to DOCTo evaluate the reversal activity of 12and 18in a dose–response manner,KBvcr cells were cultured with nontoxic concen-tration of DOC (100nM)in the presence of variousconcentrationsreversal abilities against KBvcr.Note:(a)Concentration of compounds:5l M;(b)survival rate (%)was measured by MTT method absence (À)of docetaxel (DOC).Compounds with cell viability below 20%were considered very potent and moved to further experiments;(II)aliphatic cyclic group,(III)heterocycle-aliphatic group,(IV)polar group,(V)aromatic–aliphatic group.Table 1Effect of compounds on the cytotoxicity of DOC,VCR and ADM in the P-gp overexpressing KBvcr cancer cells Compd a IC 50of DOC b (nM)Fold c IC 50of VCR (nM)Fold IC 50of ADM (nM)Fold 5628(DOC alone)—5091(VCR alone)—13120(ADM alone)—VER 75.974.285.266.1301.143.6PPD 525.510.7645.18.7935.614.01250.9110.764.986.7233.256.31828.7196.148.0117.3211.861.921142.639.5201.827.9452.129.025117.747.8188.329.9278.547.1a Concentration of compound:5l M.b SD is shown in Supplementary data .cThe reversal fold values were calculated as the following:reversal fold =IC 50(anticancer drug alone)/IC 50(anticancer drug +test compound).of compounds(Fig.5).As we expected,compounds12and18 exhibited reversal activity in a dose-dependent manner.The effec-tive concentration(EC50)values of12(1.367l M)and18 (1.310l M)were similar to that of VER(1.253l M).These results demonstrate that the effect of12and18could be similar to that of VER in chemosensitizing the MDR cells to DOC.3.4.The effect of PPD analogues on P-gp function in KBvcr cellsTo confirm our hypothesis that PPD analogues inhibit efflux activity of P-gp resulting in elevated concentration of anticancerin MDR cells,the effect of compounds12and18on pounds.Although18was slightly less potent than VER,12was more potent than VER,especially at concentrations around the EC50value(1.367l M)of12.Therefore,these results clearly indi-cated that PPD analogues,especially12,are effective P-gp inhibitors.To demonstrate the effective efflux inhibition of anticancer drugs,direct measurement of cellular accumulation of ADM in KBvcr cells was studied as the intensity of intrinsicfluorescence of ADM(Fig.7).KBvcr cells were pretreated with compounds fol-lowed by addition of ADM.Intracellular accumulation of ADM was measured as thefluorescence intensity and standardized as fold ratio.All PPD analogues induced ADM accumulation in KBvcrFigure4.Reversal of chemosensitivity of KBvcr by12or18.Chemoresistant KBvcr cells were incubated with various concentrations of anticancer drugs DOC(A),VCR (B),or ADM(C)in the presence of test compounds,as indicated,for72h to evaluate the effect on chemosensitization.Chemosensitization of KBvcr cells was observed when the cells were cotreated with5l M of12,18,or VER.5.Dose–response effect of compounds12and18on sensitizationMultidrug-resistant KBvcr cells were treated with various concentrations compounds12or18in the presence of100nM DOC,an absolutely concentration for KBvcr.Data are expressed as mean±SD of three independent experiments.Calculated median effective concentration(EC50)of compounds VER was also listed.Cellular accumulation of Rho123012345101512VER18Compound(µM)CellularaccumulationofRho123(14RFU)6.Effect of compounds on P-gp function in KBvcr cells.KBvcr cells pretreated with compounds followed by addition of Rhodamine123.The accumulation of Rhodamine123is represented by the relativefluorescent RFU).Cellular accumulation of Rhodamine123demonstrates inhibitionactivity of P-gp.Data are shown with mean±SD of three independent experiments.cells at1.3-to2.1-fold.The cellular accumulation of ADM by PPD analogues was consistent with sensitization of KBvcr cells to ADM(Table1).Thus,these data further support that PPD-derived chemosensitizers function as P-gp inhibitors resulting in cellular accumulation of anticancer drugs.3.5.Hydrophobicity evaluation of active PDD analoguesP-gp is a large polytopic membrane protein which contains a large drug-binding pocket including predominantly hydrophobic and aromatic residues.30,31Because of similarity,hydrophobic and aromatic substrates would bind to the hydrophobic and aromatic residues.Thus,it is possible that hydrophobicity of PPD analogues could influence their MDR reversal activity.The c log P values of synthesized compounds are showed in Table2.Although a few exceptions were present,the chemosensitizing effects of compounds were moderately correlated with their c log P values. Active compounds had c log P values of4–7.The c log P values of 12,18,and25,which were significantly active as described above,were6.10,6.17and4.6,which is close to that of VER.This fact im-plied that hydrophobicity is an important parameter in P-gp inhibition.4.ConclusionIn summary,we selected PPD as a lead,and twenty-five new PPD analogues were newly designed and synthesized.All synthe-sized analogues were evaluated for MDR chemosensitizing effects on clinically used anticancer drugs,such as DOC,VCR,and ADM. We succeeded to improve chemoreversal action by introducing an aromatic substituted aliphatic amine group to PPD,and SAR studies are summarized.The aromatic substituted aliphatic amine (group V)tended to enhance the reversal ability of PPD analogues, but aliphatic acyclic amine(group I),aliphatic cyclic amine(group II),heterocycle-aliphatic amine(group III),and polar aliphatic amine(group IV)led to reduce activity.Among all tested com-pounds,PPD derivatives12and18were1.3–2.6times more effec-tive than VER for DOC and VCR reversal ability.Importantly,active PPD analogues(5l M)displayed no cytotoxicity against tumor cells,suggesting that our novel PPD analogues are significant lead compounds for further clinical development to overcome the MDR phenotype.Intracellular accumulation studies using calcein-AM and ADM in KBvcr cells clearly demonstrated that PPD analogues interfere with the P-gp drug efflux pump.To conclude,newly syn-thesized PPD analogues were identified as P-gp inhibitors possess-ing a new scaffold for nontoxic chemosensitizer drug development.5.Experimental5.1.GeneralThe reagents(chemicals)were purchased from Lancaster,Alfa Aesar,and Shanghai Chemical Reagent Co.and used without fur-ther purification.Nuclear magnetic resonance(NMR)spectroscopy was performed on a Bruker AMX-300NMR(IS as TMS).Chemical shifts were reported in parts per million(ppm,d)downfield from tetramethylsilane.Proton coupling patterns were described as sin-glet(s),doublet(d),triplet(t),quartet(q),multiplet(m),and broad (br).HRESIMS were determined on a Micromass Q-Tif Global mass spectrometer and ESIMS were run on a Bruker Esquire3000Plus Spectrometer.All reactions were monitored by thin-layer chroma-tography(TLC)on HSGF254silica gel plates(150–200l m thick-ness;Yantai Huiyou Co.,China).Ozone was produced with a BGF-YQ ozone generator(2.0L/min O2,100V;Beijing ozone Co., China).Allfinal compounds are>95%pure based on HPLC. Anhydrous solvents were purchased from commercial suppliers.5.2.General synthetic procedure for compounds2–26Into a solution of PPD(1,200mg,0.435mmol)in CH2Cl2 (15mL)and cooled to–78°C,ozone was bubbled(at aflow rate of2.0L/min of oxygen containing5%of ozone)with stirring.The mixture was maintained at–78°C for5min.The reaction was monitored by thin-layer chromatography(TLC).The excess of ozone was eliminated by bubbling nitrogen into the solution. Amine(RNH2)(0.1mL),NaBH(OAc)3(368.8mg, 1.7mmol),and CH3OH(8mL)were successively added,and the mixture was al-lowed to reach0°C.After completion of the reaction(TLC monitor-ing,CH2Cl2/CH3OH,30:1,v/v,on silica gel plate),water was added, and the product was isolated by extraction with dichloromethane. The organic phase was washed with water,brine,dried over anhy-drous sodium sulfate,filtered,and concentrated under vacuum to obtain the desired crude products.The appropriate compounds (2–26)were obtained following purification by silica gel columnFigure7.Recovered ADM accumulation in PPD analogue-treated drug resistantcells.KBvcr cells were incubated in ADM medium(final concentrationM)for3h in the presence of5l M PDD analogues,and then cellularaccumulation of ADM was measured as the intrinsicfluorescence intensityThefluorescence intensity of ADM was expressed as the ratio of effectcompound to negative control(ADM).Intracellular accumulation of ADMclearly observed in the presence of PPD-derived compounds.Data are representedmean±SD,n=3.P-gp inhibitor VER(5l M)was used as a positive control.Table2c log P values of synthesized PPD-derived compoundsCompd c log P a Compd c log P1 6.7814 6.022 4.0715 6.243 4.6016 6.814 5.1317 6.125 4.9118 6.176 5.3119 6.897 4.6520 5.918 4.3821 6.279 6.2222 6.1310 3.9023 6.5511 6.3324 4.612 6.1025 4.613 6.4126 4.6VER 4.58a c log P was calculated by ChemDraw Ultra Version12.0.Chem.21(2013)4279–4287chromatography with dichloromethane/methanol/triethyl-amine=60:1:0.5(v/v/v).5.2.1.(3S,8R,10R,12R,14R,17S)-17-((S)-2-Hydroxy-5-(methyl am ino)pentan-2-yl)-4,4,8,10,14-pentamethylhexadecahydro-1H-cy clopenta[a]phenanthrene-3,12-diol(2)Yield85%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d4.12(m,1H),3.52(td,J=13.0,6.2Hz,1H), 3.21(dd,J=10.9,5.0Hz,1H),2.94(m,1H),2.62(m,1H),2.47(s, 3H),1.13(s,3H),0.99(s,3H),0.98(s,3H),0.88(s,6H),0.78(s, 3H),0.73(d,J=11.0Hz,1H);HRMS(ESI)m/z calcd for C28H52NO3 (M+H)+450.3942,found450.3945.5.2.2.(3S,8R,10R,12R,14R,17S)-17-((S)-5-(Ethylamino)-2-hydrox ypentan-2-yl)-4,4,8,10,14-pentamethylhexadecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(3)Yield86%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d 3.55(td,J=12.9, 6.3Hz,1H), 3.19(dd, J=10.9, 4.9Hz,1H),2.93(m,1H),2.72(m,1H), 2.58(m,1H), 2.48(m,1H),2.09(m,1H),1.15(s,3H),1.11(t,J=7.2Hz,3H), 0.99(s,3H),0.98(s,3H),0.89(s,3H),0.88(s,3H),0.78(s,3H), 0.73(d,J=10.7Hz,1H);HRMS(ESI)m/z calcd for C29H54NO3 (M+H)+464.4098,found464.4094.5.2.3.(3S,8R,10R,12R,14R,17S)-17-((S)-2-Hydroxy-5-(propy lam i no)pentan-2-yl)-4,4,8,10,14-pentamethylhexadecahydro-1H-cy clopenta[a]phenanthrene-3,12-diol(4)Yield87%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d 3.55(td,J=12.7, 6.2Hz,1H), 3.20(dd, J=10.8, 5.2Hz,1H),2.94(m,1H),2.67(m,1H), 2.52(m,2H), 2.11(m,1H), 1.14(s,3H),0.99(s,3H),0.97(s,3H),0.93(t, J=7.3Hz,3H),0.89(s,3H),0.88(s,3H),0.78(s,3H),0.73(d, J=10.9Hz,1H);HRMS(ESI)m/z calcd for C30H56NO3(M+H)+ 478.4255,found478.4257.5.2.4.(3S,8R,10R,12R,14R,17S)-17-((S)-2-Hydroxy-5-(isopropyl amino)pentan-2-yl)-4,4,8,10,14-pentamethylhexadecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(5)Yield88%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d 3.57(td,J=12.5, 6.0Hz,1H), 3.19(dd, J=10.8,5.2Hz,1H),2.95(d,J=12.6Hz,1H),2.85(m,1H),2.50 (m,1H),2.10(m,1H),1.14(s,3H),1.13(d,J=7.2Hz,6H),0.97(s, 3H),0.95(s,3H),0.87(s,3H),0.86(s,3H),0.76(s,3H),0.73(d, J=10.9Hz,1H);HRMS(ESI)m/z calcd for C30H56NO3(M+H)+ 478.4255,found478.4258.5.2.5.(3S,8R,10R,12R,14R,17S)-17-((S)-5-(tert-Butylamino)-2-hy droxypentan-2-yl)-4,4,8,10,14-pentamethylhexadecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(6)Yield88%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d5.30(s,1H),3.57(td,J=12.3,6.1Hz,1H), 3.19(dd,J=10.7,5.3Hz,1H),2.93(d,J=10.8Hz,1H),2.44(t, J=11.5Hz,1H),2.09(m,1H),1.13(s,12H),0.97(s,6H),0.88(s, 6H),0.77(s,3H),0.72(d,J=10.8Hz,1H);HRMS(ESI)m/z calcd for C31H58NO3(M+H)+492.7966,found492.7969.5.2.6.(3S,8R,10R,12R,14R,17S)-17-((S)-5-(Cyclopropylamino)-2-hydroxypentan-2-yl)-4,4,8,10,14-pentamethylhexadecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(7)Yield80%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d 3.54(td,J=12.6, 6.2Hz,1H), 3.20(dd, J=10.7,5.3Hz,1H),3.05(m,1H),2.95(dd,J=11.0,7.2Hz,1H), 2.56(t,J=11.0Hz,1H),2.11(m,1H),2.07(m,1H),1.11(s,3H), 0.97(s,3H),0.96(s,3H),0.87(s,6H),0.76(s,3H),0.72(d, J=10.9Hz,1H);HRMS(ESI)m/z calcd for C30H54NO3(M+H)+ 476.4098,found476.4095.5.2.7.(3S,8R,10R,12R,14R,17S)-17-((S)-5-((3-(1H-Imidazol-1-yl) propyl)amino)-2-hydroxypentan-2-yl)-4,4,8,10,14-pentame thylhexadecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol (8)Yield90%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d7.49(s,1H),7.05(s,1H),6.92(s,1H),4.00 (m,2H),3.55(td,J=12.7,6.0Hz,1H),3.19(dd,J=11.0,5.2Hz, 1H),2.88(m,1H),2.68(m,1H),2.52(m,2H),1.13(s,3H),0.98 (s,3H),0.97(s,3H),0.89(s,6H),0.78(s,3H),0.72(d,J=10.9Hz, 1H);HRMS(ESI)m/z calcd for C33H58N3O3(M+H)+544.4473,found 544.4470.5.2.8.(3S,8R,10R,12R,14R,17S)-17-((S)-2-Hydroxy-5-((2-(pipera zin-1-yl)ethyl)amino)pentan-2-yl)-4,4,8,10,14-pentamethyl hexadecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(9) Yield80%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d 5.31(d,J=1.4Hz,1H), 3.57(td,J=13.0, 5.6Hz,1H),3.20(dd,J=10.9,5.3Hz,1H),2.80(t,J=6.0Hz,2H), 2.47(m,12H),1.14(s,3H),0.98(s,6H),0.89(s,6H),0.78(s,3H), 0.73(d,J=10.9Hz,1H);HRMS(ESI)m/z calcd for C33H62N3O3 (M+H)+548.4786,found548.4784.5.2.9.(3S,8R,10R,12R,14R,17S)-17-((S)-2-Hydroxy-5-((3-hydrox ypropyl)amino)pentan-2-yl)-4,4,8,10,14-pentamethylhexadeca hydro-1H-cyclopenta[a]phenanthrene-3,12-diol(10) Yield80%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d 3.74(t,J=5.7Hz,2H), 3.54(td,J=12.6, 5.8Hz,1H),3.20(dd,J=10.8,5.4Hz,1H),2.89(m,2H),2.77(m, 1H),2.60(m,1H),1.15(s,3H),0.98(s,3H),0.97(s,3H),0.88(s, 3H),0.86(s,3H),0.78(s,3H),0.72(d,J=10.7Hz,1H);HRMS (ESI)m/z calcd for C30H56NO4(M+H)+494.4204,found494.4207.5.2.10.(3S,8R,10R,12R,14R,17S)-17-((S)-2-Hydroxy-5-((4-metho xyphenyl)amino)pentan-2-yl)-4,4,8,10,14-pentamethylhexa decahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(11) Yield90%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d6.76(d,J=9.0Hz,2H), 6.68(d,J=9.0Hz, 2H),3.71(s,3H),3.54(td,J=12.8,6.3Hz,1H),3.14(dd,J=11.0, 5.2Hz,1H),3.05(t,J=6.4Hz,2H),2.04(m,1H),1.14(s,3H),0.99 (s,3H),0.96(s,3H),0.91(s,6H),0.78(s,3H),0.75(d,J=10.6Hz, 1H);HRMS(ESI)m/z calcd for C34H56NO4(M+H)+542.4204,found 542.4208.5.2.11.(3S,8R,10R,12R,14R,17S)-17-((S)-5-(Benzylamino)-2-hydroxypentan-2-yl)-4,4,8,10,14-pentamethylhexadecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(12)Yield90%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d7.32(m,5H),3.74(s,2H),3.57(td,J=12.9, 6.0Hz,1H), 3.20(dd,J=10.8, 5.2Hz,1H), 2.97(d,J=12.3Hz, 1H),2.78(dd,J=12.3,7.2Hz,1H),2.52(t,J=11.0Hz,1H),2.12 (m,1H),1.15(s,3H),0.99(s,3H),0.97(s,3H),0.90(s,3H),0.89 (s,3H),0.78(s,3H),0.73(d,J=10.7Hz,1H);HRMS(ESI)m/z calcd for C34H56NO3(M+H)+526.4255,found526.4253.5.2.12.(3S,8R,10R,12R,14R,17S)-17-((S)-2-Hydroxy-5-(((S)-1-phenylethyl)amino)pentan-2-yl)-4,4,8,10,14-pentamethylhexadecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(13)Yield86%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d7.29(m,5H),3.82(q,J=6.7Hz,1H),3.56 (td,J=12.8,5.9Hz,1H),3.19(dd,J=10.8,5.3Hz,1H),2.69(m, 1H),2.40(m,1H),2.11(m,1H),1.40(d,J=6.7Hz,3H),1.18(s, 3H),0.97(s,3H),0.96(s,3H),0.89(s,3H),0.87(s,3H),0.77(s, 3H),0.77(d,J=10.8Hz,1H);HRMS(ESI)m/z calcd for C35H58NO3 (M+H)+540.4411,found540.4414.J.Liu et al./Bioorg.Med.Chem.21(2013)4279–428742855.2.13.(3S,8R,10R,12R,14R,17S)-17-((S)-2-Hydroxy-5-((4-metho xybenzyl)amino)pentan-2-yl)-4,4,8,10,14-pentamethylhexad ecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(14) Yield91%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d7.21(dd,J=8.6,4.2Hz,2H),6.87(dd,J=8.6, 4.2Hz,2H), 3.80(s,3H), 3.79(s,2H), 3.69(s,1H), 3.57(td, J=12.8, 6.1Hz,1H), 3.20(dd,J=10.8, 5.3Hz,1H), 2.96(d, J=12.0Hz,1H), 2.52(t,J=11.0Hz,1H), 2.11(m,1H), 1.14(s, 3H),0.99(s,3H),0.98(s,3H),0.90(s,3H),0.89(s,3H),0.78(s, 3H),0.73(d,J=10.9Hz,1H);HRMS(ESI)m/z calcd for C35H58NO4 (M+H)+566.4360,found566.4362.5.2.14.(3S,8R,10R,12R,14R,17S)-17-((S)-5-((4-Fluorobenzyl) amino)-2-hydroxypentan-2-yl)-4,4,8,10,14-pentamethylhexa decahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(15) Yield88%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d7.32(dd,J=8.6,5.4Hz,2H),7.03(dd,J=8.6, 5.4Hz,2H),5.30(s,1H),3.84(s,2H),3.53(td,J=12.9,6.0Hz, 1H),3.20(dd,J=11.0,5.2Hz,1H),2.98(m,1H),2.65(m,1H), 2.09(m,1H),1.12(s,3H),0.98(s,6H),0.88(s,6H),0.78(s,3H), 0.73(d,J=10.8Hz,1H);HRMS(ESI)m/z calcd for C34H55NO3 (M+H)+544.4161,found544.4165.5.2.15.(3S,8R,10R,12R,14R,17S)-17-((S)-5-((4-Chlorobenzyl) amino)-2-hydroxypentan-2-yl)-4,4,8,10,14-pentamethylhexad ecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(16) Yield87%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d7.29(d,J=6.1Hz,2H),7.20(d,J=6.1Hz, 2H),3.84(s,1H),3.71(s,2H),3.56(td,J=12.9,5.9Hz,1H),3.19 (dd,J=10.9,5.3Hz,1H),2.94(m,1H),2.52(t,J=10.1Hz,1H), 2.10(m,1H),1.13(s,3H),0.98(s,3H),0.97(s,3H),0.89(s,3H), 0.87(s,3H),0.77(s,3H),0.74(d,J=10.7Hz,1H);HRMS(ESI)m/z calcd for C34H55ClNO3(M+H)+560.3865,found560.3863.5.2.16.(3S,8R,10R,12R,14R,17S)-17-((S)-5-((3,4-Dimethoxy benzyl)amino)-2-hydroxypentan-2-yl)-4,4,8,10,14-pentamethy lhexadecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(17) Yield92%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d6.93(s,1H),6.86(d,J=8.4Hz,1H),6.84(d, J=8.4Hz,1H),5.30(s,1H),3.89(s,3H),3.87(s,3H),3.82(s,2H), 3.53(td,J=12.8,6.0Hz,1H),3.20(dd,J=10.8,5.3Hz,1H),2.96 (m,1H),2.69(m,1H),2.08(m,1H),1.12(s,3H),0.98(s,6H), 0.88(s,6H),0.78(s,3H),0.71(d,J=10.8Hz,1H),HRMS(ESI)m/z calcd for C36H60NO5(M+H)+586.4466,found586.4468.5.2.17.(3S,8R,10R,12R,14R,17S)-17-((S)-2-Hydroxy-5-(phenethylamino)pentan-2-yl)-4,4,8,10,14-pentamethylhexadecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(18)Yield90%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d7.23(m,5H),3.56(td,J=13.1,6.1Hz,1H), 3.20(dd,J=10.9,4.9Hz,1H),2.83(m,5H),2.54(m,1H),2.07(m, 1H),1.13(s,3H),0.98(s,3H),0.97(s,3H),0.89(s,3H),0.88(s, 3H),0.77(s,3H),0.71(d,J=10.9Hz,1H);HRMS(ESI)m/z calcd for C35H58NO3(M+H)+540.4411,found540.4414.5.2.18.(3S,8R,10R,12R,14R,17S)-17-((S)-5-((4-Chlorophenethyl) amino)-2-hydroxypentan-2-yl)-4,4,8,10,14-pentamethyl hexadecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(19) Yield84%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d7.29(d,J=6.2Hz,2H),7.17(d,J=6.2Hz, 2H),3.58(td,J=12.7,5.9Hz,1H),3.19(dd,J=10.8,5.2Hz,1H), 2.73(m,4H),2.38(m,1H),2.11(m,1H),1.16(s,3H),0.98(s, 3H),0.97(s,3H),0.88(s,3H),0.87(s,3H),0.77(s,3H),0.72(d, J=10.9Hz,1H);HRMS(ESI)m/z calcd for C35H57ClNO3(M+H)+ 574.4022,found574.4025.5.2.19.(3S,8R,10R,12R,14R,17S)-17-((S)-2-Hydroxy-5-((4-nitroph enethyl)amino)pentan-2-yl)-4,4,8,10,14-pentamethylhexadeca hydro-1H-cyclopenta[a]phenanthrene-3,12-diol(20) Yield83%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d8.16(d,J=8.8Hz,2H),7.40(d,J=8.8Hz, 2H),3.47(m,2H),3.18(m,2H),3.08(m,4H),2.80(m,1H),1.10 (s,3H),1.00(s,3H),0.97(s,3H),0.88(s,6H),0.77(s,3H),0.72 (d,J=11.0Hz,1H);HRMS(ESI)m/z calcd for C35H57N2O5(M+H)+ 585.4262,found585.4260.5.2.20.(3S,8R,10R,12R,14R,17S)-17-((S)-5-((3,4-Dimethoxyph enethyl)amino)-2-hydroxypentan-2-yl)-4,4,8,10,14-pentamethy lhexadecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(21) Yield93%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d6.80(d,J=8.2Hz,1H), 6.73(d,J=8.2Hz, 1H),6.71(s,1H),5.30(s,1H),3.86(s,3H),3.85(s,3H),3.56(td, J=12.8,6.0Hz,1H),3.20(dd,J=10.9,5.3Hz,1H),2.80(m,5H), 2.52(t,J=9Hz,1H),1.13(s,3H),0.98(s,3H),0.97(s,3H),0.90 (s,3H),0.89(s,3H),0.77(s,3H),0.73(d,J=11.0Hz,1H);HRMS (ESI)m/z calcd for C37H62NO5(M+H)+600.4623,found600.4620.5.2.21.(3S,8R,10R,12R,14R,17S)-17-((S)-5-((2-(Benzo[d][1,3] dioxol-5-yl)ethyl)amino)-2-hydroxypentan-2-yl)-4,4,8,10,14-pentamethylhexadecahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(22)Yield92%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d 6.72(d,J=7.8Hz,1H), 6.65(d,J=1.6Hz, 1H),6.61(dd,J=7.8,1.6Hz,1H),5.91(s,2H),3.56(td,J=12.8, 5.8Hz,1H),3.19(dd,J=10.9,5.3Hz,1H),2.78(m,6H),2.47(m, 1H),2.09(m,1H),1.13(s,3H),0.98(s,3H),0.97(s,3H),0.89(s, 3H),0.87(s,3H),0.77(s,3H),0.73(d,J=10.7Hz,1H);HRMS (ESI)m/z calcd for C36H58NO5(M+H)+584.4310,found584.4312.5.2.22.(3S,8R,10R,12R,14R,17S)-17-((S)-2-Hydroxy-5-((3-phenyl propyl)amino)pentan-2-yl)-4,4,8,10,14-pentamethylhexadeca hydro-1H-cyclopenta[a]phenanthrene-3,12-diol(23) Yield90%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d7.22(m,5H),3.53(td,J=13.1,6.5Hz,1H), 3.19(dd,J=10.8,5.3Hz,1H),2.90(m,1H),2.66(m,6H),2.07(m, 1H),1.10(s,3H),0.98(s,3H),0.97(s,3H),0.88(s,3H),0.87(s, 3H),0.72(s,3H),0.72(d,J=10.7Hz,1H);HRMS(ESI)m/z calcd for C36H60NO3(M+H)+554.4568,found554.4565.5.2.23.(3S,8R,10R,12R,14R,17S)-17-((S)-2-Hydroxy-5-((pyridin-2-ylmethyl)amino)pentan-2-yl)-4,4,8,10,14-pentamethylhexa decahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(24) Yield82%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d8.53(d,J=4.9Hz,1H),7.64(t,J=4.9Hz, 1H),7.20(d,J=4.9Hz,1H),7.16(t,J=4.9Hz,1H),3.85(s,1H), 3.83(s,1H), 3.55(d,J=12.8, 5.9Hz,1H), 3.19(dd,J=11.0, 5.1Hz,1H),2.92(m,1H),2.47(m,1H),2.06(m,1H),1.15(s,3H), 0.98(s,3H),0.97(s,3H),0.88(s,6H),0.77(s,3H),0.73(d, J=10.9Hz,1H);HRMS(ESI)m/z calcd for C33H55N2O3(M+H)+ 527.4207,found527.4204.5.2.24.(3S,8R,10R,12R,14R,17S)-17-((S)-2-Hydroxy-5-((pyridin-3-ylmethyl)amino)pentan-2-yl)-4,4,8,10,14-pentamethylhexa decahydro-1H-cyclopenta[a]phenanthrene-3,12-diol(25) Yield81%(starting from200mg of1);white power;1H NMR (300MHz,CDCl3):d8.52(s,1H),8.43(d,J=4.7Hz,1H),7.85(d, J=7.4Hz,1H),7.41(dd,J=7.4,4.7Hz,1H),3.79(s,2H),3.53(d, J=13.1,5.7Hz,1H),3.14(dd,J=10.8,5.2Hz,1H),2.63(m,2H), 2.02(m,1H),1.14(s,3H),1.00(s,3H),0.96(s,3H),0.91(s,6H), 0.77(s,3H),0.74(d,J=10.7Hz,1H);HRMS(ESI)m/z calcd for C33H55N2O3(M+H)+527.4207,found527.4205.4286J.Liu et al./Bioorg.Med.Chem.21(2013)4279–4287。

20(S)-原人参二醇对SMMC-7721细胞体内外作用的研究张锐;徐华丽;于小风;曲绍春;陈明侠;睢大筼【期刊名称】《中国药理学通报》【年(卷),期】2008(24)11【摘要】目的观察不同剂量的20(S)-原人参二醇(Protopanaxadiol,PPD)在体内外对人肝癌细胞株SMMC-7721抗肿瘤作用.方法建立人肝癌裸鼠皮下移植瘤模型,观察20(S)-原人参二醇的肿瘤抑制作用.MTT比色法检测20(S)-原人参二醇对SMMC-7721细胞的增殖抑制作用,Hoechst33342核染色观察细胞凋亡形态学改变,采用FITC-AnnexinⅤ/PI双染流式细胞术分析凋亡情况,同时检测Caspase-3活性.结果在体内,PPD可抑制SMMC-7721细胞裸鼠异种移植瘤生长;在体外,PPD 对SMMC-7721细胞的增殖具有明显的抑制及诱导其凋亡作用,呈时间和剂量依赖性,Hoechst33342核染色可见凋亡小体,同时伴有Caspase-3活性的增加.结论20(S)-原人参二醇在体内外均可抑制SMMC-7721细胞增殖,并诱导其凋亡,其机制可能通过活化Caspase-3诱导细胞凋亡而发挥抗肿瘤作用.【总页数】5页(P1504-1508)【作者】张锐;徐华丽;于小风;曲绍春;陈明侠;睢大筼【作者单位】吉林大学药学院药理学教研室,吉林,长春,130021;吉林大学药学院药理学教研室,吉林,长春,130021;吉林大学药学院药理学教研室,吉林,长春,130021;吉林大学药学院药理学教研室,吉林,长春,130021;吉林大学药学院药理学教研室,吉林,长春,130021;吉林大学药学院药理学教研室,吉林,长春,130021【正文语种】中文【中图分类】R-332;R284.1;R329.24;R329.25;R73-352;R735.702.2【相关文献】1.三氧化二砷对人肝癌SMMC-7721细胞的体内外作用及可能的机制 [J], 朱正;沈宇;李德春2.20(s)-原人参二醇对体外培养宫颈癌Siha细胞的增殖抑制作用及其机制 [J], 鲁育铭;赵丽晶;王贺彬;许多;赵丽娟;董妍3.茅苍术麸炒品对人肝癌SMMC-7721细胞的体内外抑制作用 [J], 罗吉辉;杨熙华;肖方涛;周美华4.小檗胺在体内外对SMMC-7721细胞的增殖抑制作用及机制研究 [J], 刘丽丽;严佳栋;许晓东;徐志英;曹莺5.人参二醇型皂苷抑制肝癌细胞转移体内外实验研究 [J], 郑文秀;何梦娇;王娜;顾正位;韩春超因版权原因，仅展示原文概要，查看原文内容请购买。

20（s）-原人参二醇、人参皂苷Rh2及人参皂苷Rg3抗肿瘤作用的对比张虹;付雯雯;徐华丽;于晓风;曲绍春;睢大筼【期刊名称】《中国老年学杂志》【年(卷),期】2014(000)017【摘要】目的：对比20（s）-原人参二醇（Ppd）与人参皂苷Rh2（Rh2）及人参皂苷 Rg3（Rg3）的抗肿瘤作用。

方法建立小鼠肝癌 H22、Lewis肺癌及黑色素瘤B16三种移植瘤动物模型，将小鼠随机分成11组，对照组给予0.5％羧甲基纤维素钠，阳性药组给予环磷酰胺（CTX）30 mg/kg，Ppd、Rh2及Rg3三个剂量组均给予相应受试药25、50、100 mg/kg。

阳性药组隔日腹腔注射给药1次，其余各组均每日灌胃给药1次，给药容积为20 ml/kg，连续10 d。

每天记录小鼠体重，末次药后称取小鼠体重及肿瘤重量并计算肿瘤抑瘤率。

结果 Ppd、Rh2及Rg3在25、50、100 mg/kg剂量下，对肝癌H22、Lewis肺癌及黑色素瘤B16均有一定的抑瘤作用，仅Ppd在50、100 mg/kg剂量下对肝癌H22、Lewis肺癌及黑色素瘤B16抑制效果明显，其抑瘤率超过40％，Rh2与Rg3的抑瘤率无明显差别。

结论 Ppd、Rh2及 Rg3对小鼠三种移植瘤均具有抗肿瘤活性，以 Ppd 的抗肿瘤作用最明显， Rh2与Rg3之间无明显差别。

【总页数】3页(P4911-4913)【作者】张虹;付雯雯;徐华丽;于晓风;曲绍春;睢大筼【作者单位】吉林大学药学院药理教研室，吉林长春 130021;吉林大学药学院药理教研室，吉林长春 130021;吉林大学药学院药理教研室，吉林长春 130021;吉林大学药学院药理教研室，吉林长春 130021;吉林大学药学院药理教研室，吉林长春 130021;吉林大学药学院药理教研室，吉林长春 130021【正文语种】中文【中图分类】R285.5;R979.1【相关文献】1.人参皂苷Rh2抗肿瘤作用机制的研究进展 [J], 张丽媛;吴铁2.人参皂苷Rh2和Rg3抗肿瘤作用研究进展 [J], 黄月云;夏婷;赵成国;陈素云;张裔辉3.人参皂苷Rh2对荷Lewis肺癌小鼠抗肿瘤作用 [J], 崔艳茹;屈飞4.人参皂苷Rh2抗肿瘤作用机制研究进展 [J], 吴颉;5.HPLC法分离与检测20(S)-人参皂苷Rg3和20(R)-人参皂苷Rg3 [J], 汤明辉;姜志宏;赵中振;张钧寿因版权原因，仅展示原文概要，查看原文内容请购买。

关于“益今生”组分原人参二醇（PPD）的抗癌性研究关于“益今生”组分原人参二醇（PPD）的抗癌性研究人参被认为是千百年来一味名贵的中药，人参具有温补、滋养的功效，被认为是滋补的名药。

但是，事实上，人参的功效病不仅在于滋补，还有它的药性。

陶弘景编的《名医别录》则认为人参还对胃肠机能衰退而引起诸症有良好的治疗作用。

近代研究发现人参还具有抗肿瘤作用，越来越引起国内外的重视。

肿瘤已经成为近年来人们广泛关注的话题，肿瘤从一至多，虽然治疗手段在不断完善，但是仍然是全世界难以攻克的难题，癌症，至今仍然是绝症。

越来越多的科研人员将人参、红豆杉等天然植物作为抗肿瘤药物的重点研究对象。

对人参的七十多年研究中发现，人参中的物质除了在主体中发现的皂苷类之外，还有人参根中提取分离出糖类、挥发油类、脂肪油、街醇类及氨基酸类、维生素类等物质。

而从人参主体中提取的皂苷类物质经过十几年的研究，在药理和临床试验中发现了人参皂苷不仅有与人参相似的药理作用，且发现它可能具有抗肿瘤的作用。

其中人参中原人参二醇型(20（S）一protopanaxadiol type)简称（PPD）和原人参三醇型(20（S）一protopanaxatriol tyPe)简称（PPT）皂苷及齐墩果酸类皂苷是药理活性的主体。

原人参二醇类皂苷（PPD）与原人参三醇类皂苷（PPT）是人参皂苷中的两类主要成分，均有相同的类固醇骨架结构，在既往的研究中发现，原人参二醇类皂苷（PPD）包含Ra1、Rb1、Rb2、Rb3、Rc、Rd、Rg3、Rh2。

在“益今生”新药成分20（S）—原人参二醇（PPD）对肝癌作用的研究显示，PPD从人参西洋参中提纯，通过体内试验增效减毒试验肿瘤免疫学试验及分子生物学试验证实，PPD具有良好的抗癌活性，对肝癌H22、Lew is肺癌及黑色素瘤B16均具有明显的生长抑制作用，并可增强机体免疫力，具有提高自然杀伤细胞( NK) 白细胞介素-2( IL-2) 活性的作用。