羟基喜树碱体外抗癌药敏的实验研究

目的：研究羟基喜树碱抗癌药物在体外的敏感程度。方法：收集１９９７年￣１９９９年４２例手术切除的肿瘤标本，用ＭＴＴ法对羟基喜树碱进行体外药敏试验。同时用另外１２种药物作对照研究。结果：羟基喜树碱平均癌肿抑制庇为５３．２４％，在４２例中抑制率〉５０％有２５例（占５９．６％），耐药占１４．６％。结论：体外药敏试验证明羟基喜树碱对多种肿瘤抑制作用良好。将在未来的抗肿瘤治疗中具有广阔的前景。     

毛叶茶提取物对不同细胞生长的影响及体内协同抗瘤作用

应用噻唑兰（ＭＴＴ）法测得毛叶茶提取物（ＥＣＰＣ）对人肝癌细胞（ＢＥＬ－７４０２）的半数抑制浓度（ＩＣ５０）为３５１．１μｇ／ｍｌ，台盼蓝拒染法（ＴＢ）的ＩＣ５０为１１５．２μｇ／ｍｌ；用ＭＴＴ法测得ＥＣＰＣ对人红白血病细胞（Ｋ５６２）的ＩＣ５０〉１０００μｇ／ｍｌ，ＴＢＥ法测得其对Ｋ５６２细胞的ＩＣ５０为８１．４μｇ／ｍｌ。ＥＣＰＣ对人胎儿肺纤维细胞（ＨＦＬＦ）和人胎儿肾细胞（ＨＦＫ）显示促生长     

急性髓系白血病细胞四项耐药指标的评价

目的　为研究急性髓系白血病（ＡＭＬ）细胞耐药指标的敏感性和耐药方式。方法　ＭＴＴ药物敏感试验，白血病祖细胞（ＣＦＵＬ）体外生长类型，Ｂｃｌ２ 抗原表达和Ｂｃｌ２／Ｂａｘ 抗原比值，流式细胞仪测定细胞内柔红霉素（ＤＮＲ）荧光强度四项指标被综合评价。结果　６２ 例ＡＭＬ中，ＭＴＴ药敏试验阳性符合率为７３％ ，阴性符合率为７０％ ，三种临床常用药物中一种药物敏感预示缓解的符合率达７１％ 。５１例ＡＭＬ中，３１例完全缓解（ＣＲ）组中ＣＦＵＬ自主分泌生长２９例，不生长２例，２０ 例未缓解（ＮＲ）组中，自主分泌生长型１４ 例，不生长型６例，统计学有显著差异性（Ｐ＜ ０．０５）。３２ 例ＡＭＬ中，药敏组Ｂｃｌ２ 表达率为５９．５５％ ±１９．５６％ ，耐药组为７７．３６％ ±１１．９１％ （Ｐ＜ ０．０５），Ｂｃｌ２／Ｂａｘ 比值药敏组为７．５０±５．０４，耐药组为１４．３２±８．９９（Ｐ＜ ０．０５）。１５例临床耐药的ＡＭＬ，ＤＮＲ荧光直方图１２ 例呈现主峰左移，诊断为经典耐药，３ 例呈现主峰右移，诊断为再生耐药。结论　ＭＴＴ，ＣＦＵＬ检测可预示临床治疗是否耐药，Ｂｃｌ２，Ｂｃｌ２／Ｂａｘ 检测与患者预后有关，有ＤＮＲ直方     

几个植物生长控制剂的磺胺类衍生物的细胞毒作用研究

用噻唑蓝还原法（ＭＴＴ法）试验了６个植物生长控制剂的磺胺类衍生物在１００μｇ／ｍｌ浓度下对人宫颈癌Ｈｅｌａ细胞生长的影响，其中二氯苯氧乙酰（２，４－Ｄ）与磺胺甲基异唑（ＳＭＺ）结合的衍生物对Ｈｅｌａ细胞生长抑制率为８８．７％，半数抑制浓（ＩＣ５０）为２８μｇ／ｍｌ；２，３，５－三氯苯甲酰－ＳＭ２的生长抑制率为８５．４％，ＩＣ５０为５８μｇ／ｍ１；２，３，５－三氯苯甲酰ＳＭＺ的抑制率为５４％。上述三种化合物对Ｈｅｌａ细胞显示细胞毒作用，而其他３种植物生长控制剂与磺胺或水杨酸甲酯形成的衍生物的生长抑制率均在５Ｏ％以下，未见明显细胞毒作用。     

青蒿酯钠的抗肿瘤作用

了解青蒿酯酯钠的抗肿瘤作用。方法：用蒿酯钠在体内、外进行抗肿瘤实验，通过伊红染色法检测青芳同酯钠对体外培养人宫颈癌细胞杀伤作用。用ＭＴＴ法检测青蒿酯钠对体外培养人体分化鳞状上皮鼻咽癌及ＳＵＮＥ－１）细胞杀伤作用。结果：体外实验表明青蒿酯钠对３种细胞均有杀作用，ＩＣ５０分别为４２．７μｇ／ｍｌ 、１０１．６μｇ／ｍｌ、１．２９μｇ／ｍｌ。体内实验表明在５０ｍｇ／ｋｇ．ｄ和１００ｍｇ／ｋｇ．ｄ剂量下灌     

琼脂培养MTT法测定混合培养HeLa细胞和人胚肺纤维母细胞药物敏感性

琼指培养ＭＴＴ法测定是结合人癌干细胞集落形成能力测定ＨＴＣＡ和ＭＴＴ比色法发展的一种新实验技术。应用这种方法测定了１：４比例混合培养ＨｅＬａ细胞和人胚肺纤维母细胞对顺氨氯铂、长春新碱、阿霉素、丝裂霉素和氟尿嘧啶的敏感性。这种方法与ＨＴＣＡ测得的ＩＣ５０值显著正相关，相关系数在０．８７４～０．９８５间（Ｐ＜０．０１）；与ＭＴＴ法测得的药敏结果有显著差异（Ｐ＜０．０１）。提示：该法可用于测定人实体瘤细胞的药物敏感性。     

若干植物生长调节剂衍生物对 DNA 多聚酶及肿瘤生长的抑制作用

目的：为了在体外培养的肿瘤细胞中证实一组有生物活性载体：磺胺（ＳＵ）、９－氨基吖啶（ＡＣＤ）、异烟肼（ＩＮＨ）的植物生长调节剂４－氯（Ｍ）、２，４－二氯（Ｄ）或２，４，５－三氯（Ｔ）的萘氧乙酸、２，３，５－三氯苯甲酸（ＴＢ）、α－萘乙酸（ＮＡＡ）的衍生物对小鼠艾氏腹水癌细胞ＤＮＡ多聚酶（ＤＰ）的抑制作用，并筛选出抑酶力最强的化合物。方法：参照Ｋ．Ｏｎｏ法分离、纯化并检测ＤＰ活性。ＭＴＴ法测定对体外培养肿瘤细胞的ＩＣ５０。结果：１１个植物生长调节剂中５个化合物在１００μｇ／ｍｌ浓度下对ＤＰ粗酶抑制作用大于５０％，抑制率分别是Ａ８４０３（２，４－Ｄ－ＡＣＤ）７８．５±５．５％，Ａ８８４５（２，４，５－Ｔ－ＳＭＺ）７５．８±１６．０％，Ａ８６０１（ＮＡＡ－ＳＮ）７４．５±６．０％，Ａ８８７４（２，４，５－Ｔ－ＩＮＨ）７３．４±２．９％，Ａ８８７３（４Ｍ－ＩＮＨ）７１．０±１２．１％。其中Ａ８４０３（９－２，４－二氯苯氧乙酸氨基吖啶，２，４－Ｄ－ＡＣＤ）抑制ＤＰ能力最强。对ＤＰα的ＩＣ５０为３．４１４２μｇ／ｍｌ，对ＤＰβ的ＩＣ５０为３．７５２３μｇ／ｍｌ。Ａ８４０３对体外培养的低分化鼻咽癌ＣＮＥ－２细胞及ＨｅＬａ     

肿瘤相关抗原CA50监测泌尿男生殖系统恶性肿瘤的临床意义

作者用免疫放射测定法（ＩＲＭＡ）检测了８４例泌尿男生殖系统恶性肿瘤病人，４８例良性疾患病人及３５例正常人的血清ＣＡ５０值。结果发现，恶性肿瘤病人的血清ＣＡ５０值显著高于良性疾患病人及正常人（Ｐ＜０．０５～０．０１）。若以１７ｕ／ｍｌ为诊断界值，其阳性率分别为：肾细胞癌５５％（１１／２０），肾盂及输尿管癌８７％（７／８），膀胱癌６５％（３０／４６），男生殖系恶性肿瘤７０％（７／１０）。其敏感性为６６％（５５／８４），特异性为８８％（４２／４８）。ＣＡ５０水平的测定还能用于对疗效的评价及对肿瘤复发的预测。     

地昔帕明单用和合用替尼泊苷对大鼠脑胶质瘤C6细胞增殖的调控作用

目的：研究三环类抗抑郁药地昔帕明（ＤＭ）对脑胶质瘤Ｃ６细胞增殖的调控作用,并探讨与临床常用治疗脑瘤的化疗药物替尼泊苷（ＶＭ－２６）合用的协同效应。方法：采用ＭＴＴ比色法测定大鼠脑胶质瘤Ｃ６细胞增殖抑制作用和流式细胞术（ＦＣＭ）进行细胞周期分析。结果：ＤＭＩ（１０－８０μｍｏｌ／Ｌ）对Ｃ６细胞的增殖具有明显的抑制作用,呈浓度时间依赖关系,药物作用２４ｈ的ＩＣ５０（９５％置信区间）为２０．７（１７．３     

瑞香狼毒提取物尼地吗啉的抗癌活性

从中药瑞香狼毒（ＳｔｅｌｌｅｒａｃｈａｍａｅｊａｓｍｅＬ．）的甲醇提取物中分离到的二萜化合物尼地吗啉（ｇｎｉｄｉｍａｃｒｉｎ），以０．０２～０．０３ｍｇ／ｋｇ腹腔注射，可使小鼠白血病Ｐ－３８８和Ｌ－１２１０腹水型的生命延长７０％和８０％。以０．０１～０．０２ｍｇ／ｋｇ腹腔注射，可使小鼠实体瘤Ｌｅｗｉｓ肺癌、黑色素瘤Ｂ－１６和结肠癌２６的生命延长４０％、４９％和４１％。应用ＭＴＴ法和克隆形成法，观察了尼地吗啉对体外培养的人白血病Ｋ５６２和胃癌Ｋａｔｏ－Ⅲ、ＭＫＮ－２８、ＭＫＮ－４５及小鼠Ｌ－１２１０的细胞生长和克隆形成抑制作用，其ＩＣ＿５０在０．００７～０．０００１２μｇ／ｍｌ范围。这表明尼地吗啉具有较强的抗癌活性，是瑞香狼毒抗癌作用的主要成分。     

In vitro antitumor activity of teniposide against carcinoma of the lung in human tumor clonogenic assay

The antitumor activity of teniposide (VM26) on carcinoma of the lung was tested by comparing it with its congener, etoposide (VP16) in an in vitro soft agar clonogenic assay system (HTCA). Of 56 tumor biopsies placed in culture, 40 tumors were evaluable for drug sensitivity information (i.e., greater than or equal to 30 colonies in the control plate). The overall in vitro response rate (defined as less than or equal to 50% survival of TCFU) at the standard dose (10% peak plasma concentration) was 35%, which is higher than that of VP16 (28%). In cell lines derived from human carcinoma of the lung, VM26 showed 50% inhibition against colony formation of PC-7 cells at 0.45 microgram/ml which is less than that of VP16. A superior antitumor activity of VM26 on PC-9 cells was also observed. VM26 was observed to act faster than VP16 thus indicating its possible superior antitumor activity in vivo.     

MTT法测定癌细胞对抗癌药物敏感性的实验研究

目的：研究ＭＴＴ法测定癌细胞对抗癌细胞对抗癌药物的敏感性。方法：采用ＭＴＴ比色法，测定４６例（胃癌１５例、肺癌１０例、大肠癌１２例、乳腺癌８例）体外原代癌细胞对８种常用化疗药物的敏感性。结果：不同肿瘤细胞对同一化疗药物的敏感性不同，不同肿瘤细胞有其各自的药敏谱。结论：研究表明ＭＴＴ法是一种简便、快捷的肿瘤细胞体外药敏试验方法，为临床实验个体化疗提供了实验依据。     

胶滴肿瘤药敏检测技术(CD-DST)的建立及初步临床应用研究

目的建立胶滴肿瘤体外药敏检测技术,探讨其在肿瘤个体化治疗中应用的可行性。方法用人类肿瘤细胞系建立胶滴肿瘤体外药敏检测技术,并用该技术对50例不同肿瘤类型的原代肿瘤标本进行体外药敏检测实验研究,每种肿瘤标本分别进行了4～5种抗肿瘤药物的敏感性测定。结果在体外成功建立了胶滴肿瘤药敏检测技术,标本整体评价率为82.0%(41/50),检测结果与临床经验有效率间有较好的符合性。结论CD-DST体外肿瘤药敏检测技术是一种较好的药敏检测方法,标本可评价率为82.0%(41/50),所需标本量小,可反映病人对不同抗肿瘤药物敏感性的差异,检测结果与临床经验有效率有较好的符合性,对抗肿瘤药物的体外筛选和对个体化治疗具有实际应用价值。     

Activity of SCH 66336, a tricyclic farnesyltransferase inhibitor, against human tumor colony-forming units

Background: The ras gene product regulates transduction of growth-proliferative signals from the membrane to the nucleus. Mutationally-activated Ras is the oncogene most frequently found in human tumors. In order to perform its function in cell signaling, Ras must be farnesylated on the CAAX motif present on the carboxyl terminus of the ras protein. This reaction is catalysed by farnesyl protein transferase. In the present study, SCH 66336, an orally bioavailable nonpeptide tricyclic farnesyltransferase inhibitor, was tested against a large variety of human tumors to define its preclinical activity profile, utilizing the human tumor cloning assay.Materials and methods: A soft agar cloning assay was used to determine the in vitro effects of SCH 66336 against primary human tumor specimens taken directly from patients. A total of 70 evaluable specimens were exposed to SCH 66336 for 14-day continuous exposure at concentrations ranging from 0.1 to 2.5 µM. In vitro responses were defined as an inhibition 50% of human tumor colony forming units at a given concentration.Results: There was a positive relationship between concentration and response to SCH 66336. With the highest concentration (2.5 µM), response was demonstrated in 50% (three of six) of breast tumors, 40% (6 of 15) of ovarian tumors, and 38% (5 of 13) of non-small-cell lung tumor colony forming units. Among the 69 specimens tested at the concentration of 2.5 µM, SCH 66336 had activity in 27% of tumor specimens that were resistant to doxorubicin, 38% of tumor specimens resistant to cisplatin, 33% of tumor specimens resistant to paclitaxel, and 27% of tumor specimens resistant to etoposide.Conclusions: The broad spectrum of soft agar growth inhibition by SCH 66336 in the human tumor cloning assay, and its efficacy at physiologically relevant concentrations in animal models, suggest that SCH 66336 may deserve future clinical trials in patients with ovarian, breast and non-small-cell lung cancer.     

Endocrine therapy testing of human breast cancers in the soft agar clonogenic assay

Summary The human tumor soft agar cloning assay has been used to assess the biological effects of cytotoxic drugs and other agents on human cancers. In this study we have examined the effects of two hormonal agents, tamoxifen (Tam) and medroxyprogesterone acetate (MPA), on colony growth of the MCF-7 human breast cancer cell line as well as fresh human breast cancer specimens. Using standard criteria for a colony (>50 cells or >60 microns in diameter) Tam (1.0µM) reduced MCF-7 colony formation by only 30% to 50%, and MPA (1.0µM) had no effect. However, both agents dramatically reduced the formation of larger colonies; less than 10% of colonies larger than 124 microns survived Tam exposure, and less than 25% survived with MPA.In vitro sensitivity (< 30% colony survival) of fresh human breast cancer specimens was observed infrequently with either Tam (1/39 evaluable assays) or MPA (3/36 evaluable assays). Colony growth of human breast cancer was unaltered when cells were plated in charcoal-stripped serum to reduce the endogenous estrogen concentration.In vitro sensitivity to Tam or MPA was not increased under these conditions. No correlation was found between estrogen receptor (ER) concentration and inhibition of colony survival with Tam or MPA. None of 16 assays from ER-positive specimens treated with Tam and 2 of 18 ER-positive specimens treated with MPA were sensitivein vitro. In contrast, 2 of 12 ER-negative specimens tested with Tam and 3 of 7 ER-negative specimens tested with MPA were sensitivein vitro. Stimulation of colony growth was observed in about 20% of Tam or MPA-treated specimens. Of the assays with ER data available, 10 of 11 with enhanced colony growth were ER-positive. Human breast cancer specimens did not grow well enough to assess the effect of these agents on large-sized colonies.These data suggest that the standard human tumor cloning assay will need modification before it can be used to predict hormonal sensitivity of fresh human breast cancer specimens.Abbreviations ER estrogen receptor - PgR progesterone receptor - Tam tamoxifen - MPA medroxyprogesterone acetate     

体外双层琼脂法与~3H—TdR参入法测定抗癌药敏感性比较

本文采用修改Hamburger—salmon体外双层琼脂法与~3H—TdR参入法同时对人类宫颈癌细胞系细胞和卵巢癌细胞进行药物敏测。观察了13例卵巢癌患者用两法测得顺铂体外敏感性与临床疗效的关系。结果表明:HeLa细胞和卵巢癌细胞接种数与克隆产率、CPM值在一定范围内均呈线性相关。用剂量——存活曲线测得阿霉素、顺氯氨铂和长春新碱对HeLa细胞系细胞的IC_(50)值无明显差异。两法测定顺铂对卵巢癌的体外敏感性与临床疗效相关。体内外阳性相关率为88％(8/9);阴性相关率为100％(4/4)。两种方法测得药敏结果无明显差异。我们的实验数据表明~3H～TdR参入法有可能取代双层琼脂法为指导临床化疗选药提供一定依据。     

In vitro chemotherapy sensitivity testing of primary human colorectal carcinomas

The cells obtained from 138 tumor samples taken from 135 patients with colorectal malignancies were cultured in vitro in a soft agar colony formation assay similar to that of Salmon and colleagues [1]. Significant colony formation occurred for 63 (51%) of evaluable tumor cultures, 53 of which were also tested against chemotherapeutic agents in vitro. The median number of drugs tested per tumor was 15. Using 70% inhibition of colony formation as the criterion for significant drug-induced cytotoxicity, only 3/53 (6%) of the tumors were noted to be sensitive to any drug. When colony counts generated by initially plated small tumor cell aggregates were taken into account, 8/40 (20%) of the tumors were noted to be sensitive in vitro to one or more agents. Because of the low rate of drug sensitivity found with this in vitro assay, its current role in the prospective assignment of chemotherapeutic treatment for patients with colorectal carcinoma is somewhat limited.     

Stimulation of colony formation of various human carcinoma cell lines by rhGM-CSF and rhIL-3

We studied the effects human recombinant granulocyte-macrophage colony-stimulating factor and human recombinant interleukin-3 on the colony formation of three human solid tumor cell lines. Using a modified doublelayer soft agar clonogenic assay rhGM-CSF enhanced colony formation of all cell lines tested in a dose dependent manner (up to twofold for the breastcancer cell line BT-20, up to 163% of the control for the hypernephroma cell line C 94 and up to 147% for the non-small cell lung cancer cell line CCL 185 at a concentration of 100 ng/ml). RhIL-3 stimulated colony formation of the cell lines C 94 and BT-20, whereas on the cell line CCL 185 rhIL-3 had no effect even at the highest dose level tested (100 ng/ml). Combinations of growth factors showed subadditive stimulation on two cell lines tested (BT-20, C 94). These data indicate that haematopoietic growth factors exert a growth promoting activity on certain solid tumor cells in vitro at physiological concentrations. Therefore our results suggest that the application of these factors in immuno- and myelosuppressed cancer patients after high dose chemotherapy should be seen in light of a possible co-stimulation of the malignant cells.     

Chemotherapy testing for human ovarian cancer using in vitro colony assay

The in vitro evaluation of anticancer drug efficacy was performed using the human tumor clonogenic assay developed by Hamburger and Salmon, and correlation between the in vitro and clinical efficacy was analyzed retrospectively. The in vitro colony assay method used in this study was a minor modification of the above method. Thirty-two out of forty-eight samples from patients with ovarian cancer formed more than five colonies per plate on in vitro colony assay. The median plating efficiency was 0.06% (range 0.02-1.3%) and the median colony count per plate was 279 (range: 8-4,000). With regard to colony formation of ovarian cancer according to the source of the specimen, the colony-forming rate of solid tumor was high (72%) as compared with 63% for ascites and 43% for pleural effusion. In vitro chemosensitivity was defined as more than a 50% decrease in colony formation and the rates for standard drugs on ovarian cancer were as follows: adriamycin (0.04 micrograms/ml) 29%, bleomycin (0.1 micrograms/ml) 24% cisplatin (0.2 micrograms/ml) 31%, 5-FU (1.0 micrograms/ml) 22%, hexamethylmelamine (1.0 micrograms/ml) 19%, L-PAM (0.4 micrograms/ml) 44%, mitomycin C (0.1 micrograms/ml) 38% and THP-adriamycin (0.5 micrograms/ml) 36%. A group of patients who had not been exposed to any anticancer drug showed higher sensitivity in vitro as compared with a group of patients who had received prior chemotherapy (35% vs 22%, p less than 0.05). Correlation between in vitro drug sensitivity and clinical responses in patients treated with the same drugs were analysed retrospectively. In all twenty cases, two were true positive cases (29%), while in ten cases, the results were true negative (77%), The overall predictive accuracy was 60%. In conclusion, ovarian cancer cells can form colonies well when the soft agar method is used and this assay method is suitable for the evaluation of various anticancer drugs in vitro.     

Morphometric and colorimetric analyses of human tumor cell line growth and drug sensitivity in soft agar culture.

Previous studies have demonstrated the suitability of image analysis of tetrazolium-stained colonies to assess growth and drug sensitivity of human tumor cells cultivated in soft agar culture. In the present study, the potential utility of colorimetric analysis to expedite experimental drug evaluations using human tumor cell lines was investigated. The same culture dishes were assessed by image analysis and by formazan colorimetry for purposes of comparing multiple methods of measuring growth as well as growth inhibition. Replicate cultures treated with 2-(p-iodonitrophenyl)-3-p-nitrophenyl-5-phenyltetrazolium chloride or 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide exhibited nearly identical colony count and volume indices as well as excellent correlation in colorimetric end points. Colony-forming unit volume analysis versus colorimetric assessment of the same cultures following dimethyl sulfoxide extraction of protamine sulfate-rinsed, dried soft agar cultures exhibited excellent linear correlation for both growth (Pearson r ranging from 0.95 to 1.00) and drug sensitivity (Pearson r ranging from 0.90 to 0.99, and Spearman r ranging from 0.82 to 0.97) and similar drug sensitivity profiles. Results of the current investigation indicate that end points of soft agar culture remain stable for a period of at least 2 weeks following assay termination. In addition, a colorimetric detection range of 1.3-2.2 log units permits determinations of survival levels ranging from 100 to 5% of respective control levels. Colorimetric analysis is anticipated to expedite soft agar colony formation assay evaluations (a) by reducing the need to use the more rigorous and time-consuming image analysis procedures to measure activity in preliminary drug sensitivity assays and (b) by permitting the determination of effective concentration ranges of new experimental agents for subsequent, more detailed investigations.