铂类络合物引起的DNA O6-AGT的耗竭及染色体损伤

本文测定了KB,CHL,HL-60和L1210细胞DNA鸟嘌呤O⁶-烷基转移酶(O⁶-AGT)活性。结果表明,KB细咆有较高的O⁶-AGT活性,系Mer⁺细胞。而CHL,HL-60和L1210细胞的O⁶-AGT活性低,属于Mer^-细胞。在此基础上我们观察了顺铂(DDP)、宁辛铂(樟脑胺氯乙酸铂,CCP)和碳铂(JM-8)对KB(Mer⁺)细胞O⁶-AGT的影响及Mer⁺和Mer^-细胞的杀伤及微核的诱发作用。结果表明,三种络合物在等毒性浓度下对O⁶-AGT耗竭程序是CCP>DDP>JM-8,但这种耗竭与杀细胞作用无明显相关性而与微核诱发作用相关。此结果提示铂类络合物对DNA鸟嘌呤O⁶的损伤可能是其致痛致突变的原因之一。     

异钩藤碱逆转人肺腺癌细胞A549/DDP对顺铂耐药性

目的 研究异钩藤碱(IR)在体外对耐药人肺腺癌细胞系A549/DDP对顺铂(DDP)的耐药性.方法 采用MTT法检测药物细胞毒性作用及耐药细胞逆转倍数,电感耦合等离子体质谱(ICP-MS)法测定细胞内顺铂浓度.结果 IR无细胞毒浓度组(3.0μg·mL-1)使A549/DDP细胞对DDP的IC50由16.81 mg·L-1降至3.36 mg·L-1;低细胞毒浓度(8.0μg·mL-1)组的IC50降至2.34 mg·L-1;2组均明显提高DDP在A549/DDP细胞内的浓度.结论 研究表明IR能部分逆转A549/DDP细胞的MDR,并可增加肿瘤细胞内DDP药物浓度.     

苦参碱对上皮性卵巢癌细胞株顺铂敏感性的影响及其机制研究

目的观察苦参碱对卵巢上皮性肿瘤患者肿瘤细胞OVCAR3体外顺铂药物敏感性的影响,并从凋亡抵抗角度探讨其潜在机制。方法体外培养上皮性卵巢癌细胞株OCCAR3,分为两组,不同浓度(10、40、160μmol/L)苦参碱处理组作为实验组,无药物处理组作为对照组。采用MTT法测定不同浓度的顺铂对两组细胞的抑制率,应用ELISA法测定两组细胞中的NF-κB核转录活性。采用Westernblot法检测Livin以及Survivin在两组细胞中的表达量。结果苦参碱处理细胞与对照组比较,对顺铂的药物敏感性不同,顺铂对3种浓度(10、40、160μmol/L)苦参碱处理后肿瘤细胞的IC50分别为(29.75±5.27)、(25.61±4.27)、(17.29±3.35)μmol/L,顺铂对对照组细胞的IC50为(31.65±2.23)μmol/L,差异有统计学意义。苦参碱(10、40、160μmol/L)处理的3组细胞的NF-κB蛋白表达分别为(0.507±0.027)、(0.375±0.019)和(0.311±0.017),对照组NF-κB蛋白表达为(0.705±0.032),差异有统计学意义。苦参碱作用于卵巢癌细胞后Livin、Survivin的表达呈浓度依赖性下调趋势,与对照组比较,差异有统计学意义。结论苦参碱处理后的上皮性卵巢癌细胞对顺铂药物的敏感性增加,其对NF-κB以及Livin、Survivin信号途径的影响是其机制之一。     

赛特铂抗肿瘤作用的临床前观察

目的观察赛特铂临床前抗肿瘤作用.方法体外观察赛特铂对人卵巢癌细胞A2780,肺腺癌细胞A549,结肠癌细胞HCT8,乳腺癌细胞MCF7和前列腺癌细胞DU145等10株人肿瘤细胞的细胞毒作用.体内观察赛特铂对人卵巢癌A2780,小鼠肉瘤S180和小鼠肝癌HepS的生长抑制作用.结果赛特铂的体外体内作用强度与对照药顺铂相当.对10种肿瘤细胞的半数抑制浓度IC50为0.51～3.29μmol·L-1,平均IC50为(1.44±0.32)μmo1·L-1;体内口服给药明显抑制裸鼠人卵巢癌A2780,小鼠肉瘤S180和小鼠肝癌HepS的生长,抑制率分别为61.3%,57.9%和50.6%.结论赛特铂的体外细胞毒和体内口服抗小鼠肿瘤生长抑制作用均与顺铂相仿.     

顺铂聚乳酸微球的药物释放特性及肝动脉栓塞研究

对顺铂聚乳酸微球进行了体外药物释放和家犬肝动脉栓塞研究。该微球粒径范围为50～200μm,平均粒径为115.76±35.94μm,顺铂含量为37.16%(W/W);体外药物释放机制符合Higuchi方程;肝动脉栓塞后8h,肝组织顺铂浓度高达21.55±12.18μg/g,明显高于肝动脉灌注顺铂组:3.16±0.09μg/g(P<0.05);肝动脉栓塞组的顺铂血浓峰值、各取血点浓度及曲线下面积AUC皆低于肝动脉灌注顺铂组。可望达到提高栓塞部位的药物疗效,降低全身毒副反应的作用。     

大鼠静注双环铂、卡铂与顺铂后铂的肾排泄与肾分布比较研究

目的比较顺铂、卡铂和双环铂3种铂制剂的肾排泄速度和肾组织中浓度,探讨铂类抗癌药的肾毒性机制,为临床用药提供参考。方法大鼠按铂元素10mg·kg-1剂量静脉注射3种铂制剂后,采用原子吸收法测定大鼠尿液中3种铂的排泄量,计算4h尿药累积排泄百分率。结果顺铂组4h的累积排泄量平均为(33.7±5.7)%;卡铂组为(89.1±8.5)%;双环铂组为(70.1±9.8)%,由此结果可见顺铂排泄最少,双环铂和卡铂则较多;静注顺铂、卡铂和双环铂后4h测定肾组织中铂的浓度分别为(70.6±31.6),(217.7±97.6)和(278.8±112.0)μg·g-1。结论与顺铂相比,双环铂与卡铂肾排泄较快,肾组织中的铂浓度相对较高,肾组织中铂浓度与肾毒性间可能无直接关系。     

大鼠静脉注射3种铂制剂后铂的肾排泄与肾分布比较研究

目的:探讨铂类抗癌药的肾毒性机制。方法:12只大鼠均分为3组,分别按铂元素10mg·kg-1剂量静脉注射顺铂、卡铂和双环铂,采用原子吸收法测定并计算4h尿药累积排泄率及肾组织中浓度。结果:4h累积排泄率顺铂、卡铂、双环铂分别平均为(33.7±5.7)%、(89.1±8.5)%、(70.1±9.8)%,肾组织中铂的浓度分别为(70.6±31.6)、(217.7±97.6)、(278.8±112.0)μg.g-1。结论:与顺铂相比,双环铂与卡铂肾排泄较快,肾组织中的铂浓度相对较高;肾组织中铂浓度高低与肾毒性大小之间可能无直接关系。     

双氢青蒿素通过PI3K/Akt通路逆转肺癌顺铂耐药A549/DDP细胞的分子机制

摘要：目的:研究双氢青蒿素通过磷脂酰肌醇-3-激酶/丝苏氨酸蛋白激酶（PI3K/Akt）通路逆转肺癌顺铂耐药A549/DDP细胞的分子机制。方法:体外培养人肺癌耐顺铂株（A549/DDP）,给予不同浓度双氢青蒿素(20,40,80,160,200,250,300μmol·L-1)处理48 h后,通过MTT法检测双氢青蒿素对A549/DDP细胞增殖影响,选取双氢青蒿素最佳实验浓度及在不同浓度DDP(0,20,40,60,80,100μmol·L-1)作用下观察并计算顺铂对A549/DDP细胞IC50和双氢青蒿素对A549/DDP逆转倍数;通过流式细胞术检测各组细胞凋亡率,Western blot法检测各组细胞中凋亡相关因子B淋巴细胞瘤-2基因（Bcl-2）和半胱氨酸蛋白酶3（Caspase3）及PI3K/Akt通路相关蛋白PI3K、AKT、磷酸化蛋白激酶B（p-Akt）的表达。结果:随双氢青蒿素浓度升高,细胞增殖抑制率依次升高（P<0.05）,且具有浓度依赖性,IC10（32.07±1.04）μmol·L-1是双氢青蒿素为最佳逆转耐药浓度;随DDP浓度的升高,双氢青蒿素A549/DDP细胞增殖抑制率随之升高（P<0.05）,DDP+双氢青蒿素组顺铂对A549/DDP的IC50为（26.42±1.23）μmol·L-1,顺铂组为（58.16±1.32）μmol·L-1,双氢青蒿素的逆转耐药倍数为2.21;与对照组相比,DDP组、双氢青蒿素组、DDP+双氢青蒿素组细胞凋亡率、caspase-3表达显著升高（P<0.05）,Bcl-2、PI3K、p-Akt/Akt表达显著降低（P<0.05）;与DDP组、双氢青蒿素组相比,DDP+双氢青蒿素组细胞凋亡率、caspase-3表达显著升高（P<0.05）,Bcl-2、PI3K、p-Akt/Akt表达显著降低（P<0.05）。结论:双氢青蒿素通过抑制PI3K/Akt通路促进A549/DDP细胞凋亡,在一定程度上可逆转肺癌A549/DDP细胞株对顺铂的耐药性。     

干扰核黄素代谢对细胞卵巢癌HO8910细胞顺铂敏感性的影响（英文）北大核心CSCD

目的研究干预核黄素(RIB)代谢途径对卵巢癌HO8910细胞顺铂(DDP)敏感性的影响。方法采用慢病毒包装sh RNA载体干扰RIB转运体2(RFT2)和RIB竞争性抑制剂氯丙嗪(CHL)干预RIB代谢途径。HO8910卵巢癌细胞分为正常细胞对照组、sh RNA对照组、RFT2 sh RNA组、CHL 50μmol·L^(-1)组、DDP 20μmol·L^(-1)组、RFT2 shR NA+DDP组、CHL+DDP组和DDP+RIB 20μmol·L^(-1)组。各组细胞处理48 h后CCK-8方法检测细胞存活;结晶紫染色方法检测克隆形成能力;流式细胞仪检测凋亡、线粒体膜电位、活性氧(ROS)含量和CD44^+CD133^+细胞比例。结果与单独DDP处理组对比,RFT2 sh RNA或CHL联合DDP能明显降低HO8910细胞活性(P<0.01),减少细胞集落形成能力(P<0.01),降低细胞线粒体膜电位(P<0.01),增加细胞ROS含量和细胞凋亡比例(P<0.01),同时减少CD44^+CD133^+细胞比例(P<0.01),RIB可减弱DDP对HO8910细胞的作用(P<0.01)。结论干扰RIB代谢途径能增强DDP对卵巢癌HO8910细胞的杀伤作用,该作用与增强DDP氧化损伤和降低干细胞样肿瘤细胞比例有关;RIB可减弱DDP对HO8910细胞的作用。     

多聚二磷酸腺苷核糖聚合酶抑制剂对顺铂在肺癌治疗中增敏的作用研究

目的探讨多聚二磷酸腺苷核糖聚合酶(PARP-1)抑制剂4-氨基-1,8-萘二胺(4-AN)对顺铂在肺腺癌治疗中的增敏作用及相关机制。方法应用MTT法和克隆形成试验检测4-AN与顺铂联合作用对A549细胞的细胞毒性作用;应用单细胞凝胶电泳和微核试验检测4-AN与顺铂联合作用对A549细胞的遗传毒性作用。结果 4-AN可以增加顺铂对A549细胞的杀伤作用,且杀伤作用随4-AN浓度增加而增强;4-AN可以增加顺铂导致的DNA单双链断裂和染色体损伤,而且随药物浓度的增加,损伤作用增强。结论抑制PARP-1可以有效的增加A549对顺铂的敏感性,其作用机制可能是通过抑制A549细胞DNA单双链损伤修复,继而引起染色体损伤,导致细胞的生长和克隆受到抑制。     

Chromosomal aberrations in V79 cells induced by superoxide radical generated by the hypoxanthine-xanthine oxidase system

The effect of the superoxide radical, generated by the hypoxanthine-xanthine oxidase system, on chromosomal mutation was examined in Chinese hamster V79 cells. When cells were treated with this system for 1 h in Hanks' solution, the incidence of metaphases with chromosomal aberrations was increased with hypoxanthine at concentrations of 2.5 to 10 micrograms/ml. On the other hand, in Eagle's minimum essential medium (MEM) or MEM supplemented with 10% fetal bovine serum, only hypoxanthine at 5 micrograms/ml plus xanthine oxidase induced chromosomal aberrations and higher concentrations of hypoxanthine were cytotoxic to V79 cells. The increased frequency of chromosomal aberrations and the cytotoxicity of hypoxanthine plus xanthine oxidase were not affected by superoxide dismutase, but were strongly inhibited by catalase.     

A novel candidate compound with urethane structure for anticancer drug development

Diethyl-4,4'-methylenebis(N-phenylcarbamate) (MDU) is a urethane compound that we originally synthesized, along with three other compounds, to investigate how polyurethane is hydrolysed. We tested the four compounds for cytotoxicity in two Chinese hamster cell lines (CHL and V79) and a human cancer cell line (HeLa S3). MDU showed the strongest cytotoxicity in all the cell lines with an IC50 of around 0.1 microg/ml. We further investigated MDU for its ability to induce chromosome aberrations (CAs) and micronuclei (MN) in CHL cells. MDU induced around 100% polyploid cells at 0.5 microg/ml after 24- and 48-h treatment in the CA test and a significantly increased frequency of micronuclei, polynuclear cells, and mitotic cells in the MN test, suggesting that it may induce numerical CAs. MDU's ability to cause mitotic arrest in CHL cells was greater than that of taxol and colchicine. Based on a COMPARE analysis using JFCR39, a panel of cancer cell lines, we predicted MDU to be a tubulin inhibitor. We confirmed this possibility in nerve growth factor-stimulated PC12 cells as well as in HT1080 cells, in which MDU exhibited the activity to inhibit tubulin polymerization. MDU is simpler in structure than existing anticancer drugs taxol and vincristine and can be synthesized relatively easily. Here we offer MDU as a potential new type of anticancer drug, stable even at room temperature, and inexpensive.     

Contribution of the major copper influx transporter CTR1 to the cellular accumulation of cisplatin, carboplatin, and oxaliplatin

The goal of this study was to determine the ability of the major copper influx transporter CTR1 to mediate the cellular accumulation of cisplatin (DDP), carboplatin (CBDCA), and oxaliplatin (L-OHP). Wild-type murine embryonic fibroblasts (CTR1+/+) and a subline in which both alleles of CTR1 were deleted (CTR1-/-) were tested for their ability to accumulate platinum when exposed to increasing concentrations of DDP, CBDCA, or L-OHP for 1 h. They were also tested for their sensitivity to the growth-inhibitory effect of each drug. Platinum content was measured by ion-coupled plasmon mass spectroscopy. The experimental model was validated by measuring copper accumulation and cytotoxicity. CTR1-/- cells accumulated only 5.7% as much copper as CTR1+/+ cells during a 1-h exposure to 2 microM copper. When exposed to DDP, CBDCA, or L-OHP at 2 microM, accumulation in the CTR1-/- cells was only 35 to 36% of that in the CTR1+/+ cells. When tested at a 5-fold higher concentration, this deficit remained for DDP and CBDCA, but accumulation of L-OHP was no longer CTR1-dependent. There was an association between the effect of loss of CTR1 function on uptake of the platinum drugs and their cytotoxicity. The CTR1-/- cells were 3.2-fold resistant to DDP, 2.0-fold resistant to CBDCA, but only 1.7-fold resistant to L-OHP. Thus, whereas CTR1 controls the cellular accumulation of all three drugs at low concentrations, accumulation of L-OHP is not dependent on CTR1 at higher concentrations. We conclude that L-OHP is a substrate for some other cellular entry mechanism, a feature consistent with its different clinical spectrum of activity.     

蛇床子素逆转人膀胱肿瘤T24／ADM细胞耐药作用及其机制

目的研究蛇床子素（osthole,OST）对T24／ADM细胞多药耐药的逆转及对P-gP表达、功能的影响。方法采用MTT法筛选OST对T24／ADM细胞无毒或低毒浓度,以筛选出的浓度作为耐药逆转的实验浓度。分别利用MTF法分别检测阿霉素（adriamycin,ADM）、氟尿嘧啶（fluorouracil,Fu）及顺铂（cisplatin,DDP）对细胞株T24和T24／ADM以及OST处理后的T24／ADM细胞的半数抑制浓度IC50。用流式细胞仪测定OST对T24／ADM细胞上P-gP表达的影响。结果浓度17Ixmol／LOST对T24／ADM无明显细胞毒性,T24／ADM对ADM、FU和DDP均有不同程度的。耐药,耐药倍数分别为18．86、5．09、3．28,使用17μmol／LOST处理后T24／ADM对上述ADM、Fu和DDP的敏感性均有不同程度的提高,ADM对耐药细胞株的IC蜘由（0．962±0．017）μxg／mL降至（0．364±0．031）μg／mL,逆转指数为2．64;FU对耐药细胞株的IC。由（0．723±0．003）μg／mL降至（0．463±0．021）μxg／mL,逆转指数为1．56;DDP对耐药细胞株的IC50由（0．664±0．007）μg／mL降至（0．587±0．016）μg／mL,逆转指数为1．13,使T24／ADM细胞内P-gP蛋白表达明显下降,下降了16．32％（P〈0．05）。结论蛇床子素在体外能部分逆转T24／ADM的耐药性,逆转机制可能是通过抑制P—gP蛋白表达实现的。     

IN VITRO CHROMOSOME ABERRATION ASSAY USING HUMAN BRONCHIAL EPITHELIAL CELLS

The determination of chromosome aberrations (CA) in Chinese hamster lung fibroblast (CHL) cells was compared with that in normal human bronchial epithelial (BEAS-2B) cells, which upon adenovirus infection were reported to possess carcinogen metabolizing capacities similar to polycyclic aromatic hydrocarbons. CHL and BEAS-2B cells were treated with increasing concentrations of benzo\[a]pyrene ( BaP) or N -nitrosodiethylamine (NDEA) . In BEAS-2B cells, BaP, at a concentration of 50 mug/ml, produced a significant increase in the CA frequency, while NDEA did not markedly alter the number of aberrations in the absence of S9 mixture. The CHL cells exposed to BaP and NDEA in the presence of S9 mixture responded as anticipated with a 30% and 14% frequency of CA observed in the BaP (50 mug/ ml) and NDEA (1000 mug/ ml) treated cells, respectively. The results of this study show that the CA assay using human cell line with intrinsic metabolic activation system, such as BEAS-2B cells, may be a useful model for predicting human clastogens and carcinogens.     

Genetic toxicity of taxol

研究了抗肿瘤新药紫杉醇的致突变性及致癌潜能，结果表明，紫杉醇在１～５０００μｇ／皿范围内，无论有无Ｓ９活化系统对沙门氏菌株四株（ＴＡ９７，ＴＡ９８，ＴＡ１００，ＴＡ１０２）标准测试菌株均无致突变性．但在小鼠骨髓细胞微核试验和ＣＨＬ细胞染色体畸变分析试验中，均显示了阳性反应．紫杉醇在２０，４０，８０ｍｇ·ｋｇ－１诱导的小鼠骨髓多染红细胞微核率分别为１９．３‰，２９．３‰，４７．１‰，均明显高于阴性对照的２．２‰（Ｐ＜０．０１）。染色体畸变分析表明，非活化条件下在２０～１６０μｇ·Ｌ－１范围内，活化条件下在８０～６４０μｇ·Ｌ－１范围内，紫杉醇可使ＣＨＬ细胞染色体畸变增加．畸变类型主要为染色体数目改变，表明其作用点是纺缍体而不是ＤＮＡ．本研究结果显示了紫杉醇的致癌潜能，也为其临床应用的风险效益评价提供了有价值的资料．关键词紫杉醇；沙门氏菌；微核；染色体畸变     

Mutagenicity studies on catena-(S)-[mu-[N alpha-(3-aminopropionyl)histidinato(2-)-N1,N2,O:N tau]-zinc]

Z-103 (catena-(S)-[mu-[N alpha-(3-aminopropionyl)histidinato (2-)-N1,N2,O:N tau]-zinc], CAS 107667-60-7) was examined in the bacterial mutation test, a chromosomal aberration test with mammalian cells in culture and the micronucleus test using male mice. 1. Z-103 did not increase the number of revertants in Escherichia coli WP2 uvrA when tested at up to 5000 micrograms/plate in the presence or absence of metabolic activation. And Z-103 did not increase mutants in Salmonella typhimurium SD 100 (streptomycin dependent strain) or in Salmonella typhimurium TM677 (8-azaguanine sensitive strain) when tested at up to 5000 micrograms/ml in the presence or absence of metabolic activation. 2. The chromosomal aberration test was carried out with cultured Chinese hamster lung cells (CHL). For the direct assay procedure, the cells were treated with 3.3 x 10(-4)-3.3 x 10(-6) mol/l Z-103 for 24 or 48 h, after which time chromosome preparations were made. For the metabolic activation assay procedure, the cells were treated with 1.0 x 10(-3)-3.3 x 10(-6) mol/l of Z-103 for 6 h in the presence or absence of metabolic activation, and the chromosome preparations were made after a further 18-h incubation in the absence of Z-103 and metabolic activation. Z-103 did not cause chromosome aberrations either in the presence or in the absence of metabolic activation. 3. The micronucleus test was performed in ddY male mice. Z-103 was administered orally to mice at a dose of 400, 200 or 100 mg/kg.(ABSTRACT TRUNCATED AT 250 WORDS)     

XRCC1 protects against particulate chromate-induced chromosome damage and cytotoxicity in Chinese hamster ovary cells.

Water-insoluble hexavalent chromium compounds are well-established human lung carcinogens. Lead chromate, a model insoluble Cr(VI) compound, induces DNA damage, chromosome aberrations, and dose-dependent cell death in human and Chinese hamster ovary (CHO) cells. The relationship between lead chromate-induced DNA damage and chromosome aberrations is unknown. Our study focus was on examining the role of XRCC1 in lead chromate-induced cytotoxicity and structural chromosomal aberrations in CHO cells. Three different cell lines were used: AA8 (parental), EM9 (XRCC1 mutant), and H9T3 (EM9 complemented with human XRCC1 gene). Cytotoxicity was significantly higher in EM9 cells when compared to AA8 and H9T3 cells, indicating that XRCC1 is important for protecting cells from lead chromate particles-induced cell death. The frequency of damaged metaphase cells was not affected by XRCC1 deficiency. However, the total amount of Cr(VI)-induced chromosome damage was exacerbated by XRCC1 deficiency, and the spectrum of damage changed dramatically. Chromatid and isochromatid lesions were the most prominent aberrations induced in all cell lines. XRCC1 was essential to reduce the formation of chromatid lesions but not for isochromatid lesions. In addition, XRCC1 deficiency resulted in a dramatic increase in the number of chromatid exchanges, indicating that XRCC1 is involved in protection from lead chromate-induced chromosome instability.     

XRCC1 protects against particulate chromate-induced chromosome damage and cytotoxicity in Chinese hamster ovary cells.

Water-insoluble hexavalent chromium compounds are well-established human lung carcinogens. Lead chromate, a model insoluble Cr(VI) compound, induces DNA damage, chromosome aberrations, and dose-dependent cell death in human and Chinese hamster ovary (CHO) cells. The relationship between lead chromate-induced DNA damage and chromosome aberrations is unknown. Our study focus was on examining the role of XRCC1 in lead chromate-induced cytotoxicity and structural chromosomal aberrations in CHO cells. Three different cell lines were used: AA8 (parental), EM9 (XRCC1 mutant), and H9T3 (EM9 complemented with human XRCC1 gene). Cytotoxicity was significantly higher in EM9 cells when compared to AA8 and H9T3 cells, indicating that XRCC1 is important for protecting cells from lead chromate particles-induced cell death. The frequency of damaged metaphase cells was not affected by XRCC1 deficiency. However, the total amount of Cr(VI)-induced chromosome damage was exacerbated by XRCC1 deficiency, and the spectrum of damage changed dramatically. Chromatid and isochromatid lesions were the most prominent aberrations induced in all cell lines. XRCC1 was essential to reduce the formation of chromatid lesions, but not for isochromatid lesions. In addition, XRCC1 deficiency resulted in a dramatic increase in the number of chromatid exchanges, indicating that XRCC1 is involved in protection from lead chromate-induced chromosome instability.