Additive and supra-additive cytotoxicity of cisplatin-taxane combinations in ovarian carcinoma cell lines

The purpose of this study was to compare the growth-inhibitory effect of cisplatin–paclitaxel with that obtained with a cisplatin–docetaxel combination and to assess the type of interaction. Concomitant use of taxanes and cisplatin was studied in seven human ovarian carcinoma cell lines, using the 96-well plate clonogenic assay. Chemosensitivity was expressed in terms of IC₅₀ values, the drug concentration causing 50% inhibition of clonogenic survival. The type of interaction was studied using the area under the survival curve ratios (AUC ratios) obtained by numerical integration. Comparison of the AUC ratio and the surviving fraction (SF) value after taxane alone was made using Student''s t-test. The influence of the drug concentration was tested by one-way analysis of variance (Anova). A supra-additive or additive effect was seen when seven ovarian carcinoma cell lines were exposed to paclitaxel or docetaxel concomitantly with cisplatin. A supra-additive effect was found in four cell lines (UT-OC-3, UT-OC-4, UT-OC-5 and SK-OV-3) after simultaneous use of cisplatin with all docetaxel concentrations tested, and in two cell lines (UT-OC-4 and SK-OV-3) when cisplatin was used concomitantly with paclitaxel. A more pronounced supra-additive effect was seen with the combination of cisplatin and docetaxel. The degree of supra-additivity was dose dependent, with increasing synergy after a higher taxane dose. The data obtained in this study suggest that a supra-additive or additive effect can be achieved in ovarian carcinoma with the concomitant use of cisplatin and a taxane. © 1999 Cancer Research Campaign     

Supra-additive effect with concurrent paclitaxel and cisplatin in vulvar squamous cell carcinoma in vitro

The effect of concurrent paclitaxel and cisplatin was tested in vitro in 5 vulvar squamous cell carcinoma (SCC) cell lines (UM-SCV-1A, -2, -4 and -7 and UT-SCV-3). Chemosensitivity was tested using the 96-well plate clonogenic assay. Paclitaxel concentrations used varied between 0.4 and 1.6 nM, and cisplatin concentrations varied between 0.1 and 0.9 microg/ml. These drug concentrations are clinically achievable. Survival data were fitted to the LQ model, and the area under the curve (AUC) value was obtained with numerical integration. The type of interaction was determined by comparing the AUC ratio of the 2 drugs with the survival fraction (SF) of paclitaxel alone. With all cell lines tested the growth-inhibitory effect of simultaneous paclitaxel and cisplatin was at least additive. The effect of the tested combination on the UM-SCV-1A and UT-SCV-3 cell lines was clearly supra-additive with all paclitaxel concentrations tested, and the UM-SCV-4 and UM-SCV-7 cell lines exhibited a supra-additive effect with increasing paclitaxel concentrations. The degree of supra-additivity was dose-dependent in the UM-SCV-7 cell line with increasing synergy at higher paclitaxel doses. In the current study the combination of paclitaxel and cisplatin had a clear additive or supra-additive cytotoxic effect on the vulvar SCC cell lines, and it has been successfully used in other gynecologic malignancies; therefore concurrent paclitaxel and cisplatin also deserves further testing in clinical settings in advanced-stage vulvar carcinoma, which has a poor prognosis.     

Effect of 41 degrees C and 43 degrees C on cisplatin radiosensitization in two human carcinoma cell lines with different sensitivities for cisplatin

The effect of trimodality treatment consisting of hyperthermia, cisplatin and radiation was investigated in two cell lines with different sensitivities to cisplatin. Hyperthermia treatment was performed for 1 h at 41 degrees C and 43 degrees C in order to compare the effects of the two temperatures. Clonogenic assays were performed with cisplatin-sensitive SiHa human cervical carcinoma and cisplatin-resistant SW-1573 human lung carcinoma cell lines. Cells were treated with various combinations of hyperthermia, cisplatin and radiation. Radiation was performed after 1 h of simultaneous hyperthermia and cisplatin treatment. Cisplatin exposure was for 1 h or continuous without refreshment of the cisplatin-containing medium. SiHa cells were more sensitive to cisplatin than SW-1573 cells. Hyperthermia at 41 degrees C decreased survival in SW-1573 cells but was not cytotoxic in SiHa cells. Hyperthermia at 43 degrees C decreased survival dramatically in both cell lines with SiHa being the most sensitive. The addition of hyperthermia at 41 degrees C and 43 degrees C to cisplatin treatment led to enhanced cell kill in both cell lines compared with cisplatin alone. Radiosensitization was observed after continuous but not after 1 h of cisplatin treatment. Hyperthermia at 43 degrees C increased radiosensitivity whereas hyperthermia at 41 degrees C did not. A combination of 41 degrees C hyperthermia with continuous cisplatin treatment had an additive effect on SW-1573 cells but enhanced cisplatin radiosensitivity of SiHa cells. In SW-1573 cells trimodality treatment using 43 degrees C hyperthermia enhances cisplatin radiosensitivity. We conclude that hyperthermia at 43 degrees C enhances cisplatin-induced radiosensitization in both cisplatin-sensitive and -resistant cell lines. Hyperthermia at 41 degrees C was also able to increase cisplatin-induced radiosensitivity but only in the cisplatin-sensitive SiHa cell line.     

X-ray and cis-diamminedichloroplatinum(II) cross-resistance in human tumor cell lines

Clinical studies have suggested a close correlation between cis-diamminedichloroplatinum(II) (cisplatin) and radiation resistance. To determine whether this cross-resistance is due to an inherent cellular resistance to both agents, ten early passage human tumor cell lines were examined for their radiation and cisplatin sensitivity in vitro. Previous studies have suggested that these early passage tumor cell lines retain many of their in vivo characteristics and are therefore good models for tumor cells in vivo. Radioresistance was strongly associated with cisplatin resistance in these cell lines. Four of the cell lines examined were radioresistant, having Dos greater than 2.0 Gy. These four lines were also resistant to cisplatin, with the dose reducing survival to 10% greater than 1.29 microM. The remaining six cell lines had Dos ranging from 1.07 to 1.57 Gy of X-ray and doses reducing survival to 10% of less than 0.83 microM cisplatin. Because early passage human tumor cell lines were used, resistance or sensitivity to radiation and cisplatin most likely developed in vivo and was not due to selection in vitro. These results indicate that cross-resistance between cisplatin and radiation in vivo is probably due primarily to an inherent cellular resistance to these agents and not necessarily to the tumor microenvironment in situ.     

Radiosensitization by Cisplatin treatment in Cisplatin-resistant and sensitive human ovarian-carcinoma cell-lines

The responses of cisplatin resistant (A2780(cp)) and cisplatin sensitive (A2780) ovarian carcinoma cell lines to radiation, cisplatin and cisplatin plus radiation have been studied. The cisplatin resistant cell line showed cross resistance to radiation. When cells were exposed to 2 mu g/ml of cisplatin treatment for one hour radiosensitization was achieved. The degree of radiosensitization was treatment sequence dependent. Irradiation followed by cisplatin treatment resulted in synergistic interaction with dose modifying factors (DMFs) as high as 1.5 in the resistant line and 1.1 in the sensitive line. The reverse sequence resulted in antagonistic interaction with DMFs of 0.8 to 0.9 at the 10% survival level. At lower survival levels the interaction remained antagonistic for the resistant cell line. Increasing the cisplatin concentration to 4 mu g/ml did not increase the degree of cisplatin radiosensitization. Simultaneous treatment of irradiation during the middle of a one or two hour cisplatin treatment did not increase the degree of radiosensitization. The cisplatin resistant cell line had a much greater cisplatin radiosensitization effect than the sensitive parental cell line with maximum DMFs for the resistant cell line ranging from 0.9-1.5 and for the sensitive cell line from 0.8-1.1. These results indicate that cisplatin may be an effective radiosensitizer especially in cisplatin resistant cell lines. This effect may be related to the inhibition of repair of radiation damage which may be elevated as a mechanism of resistance in the resistant cell line.     

Correlation of in vitro drug sensitivity testing of long-term small cell lung cancer cell lines with response and survival

In vitro drug sensitivity testing (DST) of long-term cultures from small cell lung cancer (SCLC) tumours was correlated with response and survival after four cycles of etoposide and cisplatin. 27 cell lines from 25 patients were tested by the semi-automated MTT assay after a median culture of 29 months. The logs of the IC₅₀ concentrations for etoposide and cisplatin were correlated with each other. For both drugs, median IC₅₀ values of patients with partial or complete responses (“responders”) were significantly lower (7–8 fold) than those of non-responders. When survival was plotted according to whether drug IC₅₀ values were in the upper or lower halves, curves for etoposide were significantly different, but those of cisplatin were not. DST of the long-term cell lines by MTT assay was significantly correlated with the Weisenthal dye exclusion assay of earlier passages of the same cell lines. DST of long-term SCLC cultures can predict clinical response and, for etoposide, survival. Disease-oriented panels of carefully selected, continuous, human tumour cell lines can be used to screen new drugs.     

Carboplatin- and cisplatin-induced potentiation of moderate-dose radiation cytotoxicity in human lung cancer cell lines.

The interaction between moderate-dose radiation and cisplatin or carboplatin was studied in a cisplatin-sensitive (GLC4) and -resistant (GLC4-CDDP) human small-cell lung cancer cell line. Cellular toxicity was analysed under oxic conditions with the microculture tetrazolium assay. For the platinum and radiation toxicity with the clinically relevant dose ranges applied, this assay was used to obtain information on cell survival after the treatments. Apart from effects on cell survival effects on DNA were also investigated. Configurational DNA changes could be induced by platinum drugs and thereby these drugs might change the frequency of DNA double-strand breaks (dsbs). DNA fragmentation assayed with the clamped homogeneous electric field (CHEF) technique was used as a measure for dsbs in DNA. The radiosensitising effect of the platinum drugs was expressed as enhancement ratio (ER) calculated directly from survival levels of the initial slope of the curve. The highest ER for cisplatin in GLC4 was 1.39 and in GLC4-CDDP 1.38. These were all at 75% cell survival. Carboplatin showed increased enhancement with prolonged incubation up to 1.21 in GLC4 and was equally effective as cisplatin in GLC4-CDDP. According to isobologram analysis, prolonged incubation with both platinum drugs showed at least additivity with radiation for both cell lines at clinically achievable doses. GLC4-CDDP showed cross-resistance to radiation. The radiosensitising capacity of both lung cancer cell lines was not dependent on their platinum sensitivity. The formation of dsbs in DNA directly after radiation was not influenced by pretreatment of either drug in the sensitive or in the resistant cell line. Drug treatment resulted in decreased DNA extractability in control as well as in irradiated cells. Modest enhancement ratio for radiosensitisation by platinum drugs cannot be explained on the level of dsb formation in DNA in both cell lines. Interaction of radiation with the clinically less toxic carboplatin can be improved by prolonged low-dose carboplatin exposure before irradiation and is as potent as cisplatin in the resistant lung cancer cell line. This suggests an advantage in combining radiation and carboplatin in lung cancer patients.     

Cyclopentenylcytosine does not enhance cisplatin-induced radiosensitization in human lung tumour cells

The search for agents that enhance the effect of ionizing radiation has been an object of study for decades. In this study, the sensitizing properties of cyclopentenylcytosine (CPEC) on radiation and cisplatin-induced radiosensitization in human squamous lung carcinoma cells were investigated. Human lung tumour SW-1573 cells (SWp, parental; SWg, gemcitabine-resistant) were incubated with CPEC and cisplatin and subsequently irradiated with different doses of γ-rays. Clonogenic survival was determined to measure the effectiveness of the treatments. CPEC (1 or 2 μM) treatment for 4 h decreased the plating efficiency to 75 and 50% in SWp and SWg cells, respectively. In the SWg cells, 0.1 and 1 μM CPEC for 4 h enhanced the cell killing effect of cisplatin. However, an increase was not noted in the SWp cells. Due to the moderate toxicity of 1 μM for 4 h, this CPEC dose was used in the radiosensitization experiments. However, CPEC neither radiosensitized the lung tumour cells nor enhanced the radiosensitizing effect of cisplatin. A 2-h incubation with 4 μM cisplatin also decreased the plating efficiency to 75-80% in the two cell lines. Using this cisplatin dose, radiosensitization was obtained in the two cell lines. Although cisplatin treatment clearly radiosensitized the lung tumour cells, CPEC treatment did not. Cisplatin-induced radiosensitization was also not enhanced by CPEC.     

Sodium thiosulfate administered six hours after cisplatin does not compromise antineuroblastoma activity.

PURPOSE: We determined if the potentially otoprotective agent sodium thiosulfate (STS) could be given 6 h after cisplatin without diminishing the antineuroblastoma activity of cisplatin in human neuroblastoma cell lines in vitro (including cisplatin-resistant cell lines) and in neuroblastoma xenografts in vivo. EXPERIMENTAL DESIGN: We determined the antineuroblastoma activity of cisplatin with or without the addition of STS at 0 or 6 h after cisplatin in six neuroblastoma cell lines, both in standard cell culture conditions (20% O(2)) and in physiologic hypoxia (2% O(2)). Drug cytotoxicity was measured using the DIMSCAN fluorescence/digital imaging microscopy assay. In vivo studies of cisplatin combined with STS used a human neuroblastoma subcutaneous xenograft model (SMS-SAN) in athymic nu/nu mice. RESULTS: A significant protection against cisplatin cytotoxicity was seen when the neuroblastoma cells were exposed to cisplatin directly combined with STS. However, when cisplatin was given first and STS exposure occurred 6 h later, no effect on cisplatin cytotoxicity was observed. In a subcutaneous neuroblastoma xenograft model in nu/nu mice, mice receiving cisplatin alone or cisplatin + STS at 6 h had significantly better progression-free survival rates (P < 0.03) compared with controls or mice treated with cisplatin + STS concurrently. There was no statistically significant difference in outcomes between mice treated with cisplatin alone and the group treated with cisplatin followed by STS 6 h later (P = 0.9). CONCLUSION: These preclinical data suggest that the use of STS 6 h after cisplatin for otoprotection is unlikely to compromise the antineuroblastoma activity of cisplatin.     

Anti-Proliferative Effects of Compound A and Its Effect in Combination with Cisplatin in Cholangiocarcinoma Cells

Background: Cholangiocarcinoma (CCA) is a fatal cancer with high resistance to anticancer drugs.  The development of new drugs or compounds to be used alone or in combination with currently available chemotherapeutic agents to improve the treatment of CCA is needed. Compound A (CpdA), which is a small plant-derived glucocorticoid receptor modulator, strongly inhibited the growth and survival of several cancers.  However, the effect of CpdA on cholangiocarcinoma has not been elucidated. The aim of this study was to investigate the effect of CpdA on CCA. Methods: Cytotoxicity of CpdA was tested in primary cells including peripheral blood mononuclear cells (PBMCs), fibroblasts, and human umbilical vein endothelial cells (HUVECs), as well as on CCA cell lines (KKU-100, KKU-055, and KKU-213) was examined. Cell cycle distribution and IL-6 expression was assessed by flow cytometry and real-time polymerase chain reaction, respectively.  The effect of combination CpdA and cisplatin was evaluated by cell viability assay. Results: CpdA significantly inhibited cell cycle at G1 phase in CCA cell lines, and reduced IL-6 mRNA expression.  However, combination CpdA and cisplatin did not enhance the inhibitory effect. TGFβR-II expression was increased in CCA cells after the combination treatment. Conclusions: These results indicate the potential of CpdA for CCA treatment. However, combination treatment with CpdA and cisplatin increased CCA cell survival. The molecular mechanism is likely attributable to promotes cell survival via the TGFβR-II signaling pathway. The combination of CpdA with other anticancer drugs for CCA treatment should be further examined.     

Enhancement of osteosarcoma cell sensitivity to cisplatin using paclitaxel in the presence of hyperthermia

Abstract

Purpose: This paper aimed to evaluate the effects of a combination of paclitaxel and cisplatin on osteosarcoma (OS) cell lines in the presence of hyperthermia and to investigate the related mechanism.

Materials and methods: Two types of OS cell lines (OS732 and MG63) were treated with paclitaxel and cisplatin in the presence of hyperthermia. The survival rate was measured by MTT assay, and the clonogenic rate was measured by a clonogenic assay. The cellular changes were observed with an inverted phase contrast microscope and a fluorescence microscope. The apoptotic effect was analysed with flow cytometry (FCM). Fas expression by the OS cell lines was measured by western blot. Fas expression in OS tissue was measured by immunohistochemistry.

Results: Our study indicated that 1?h after the application of a combination of 10?μg/mL paclitaxel and 5?μg/mL cisplatin to OS cells at 43?°C, the survival rate of the OS cells was 11.96%, which was significantly lower than when either 10?μg/mL paclitaxel (45.02%) or 5?μg/mL cisplatin (48.69%) was applied alone (p?<?0.01). Additionally, the clonogenic assay demonstrated that the clonogenic survival rate in the OS cells of the combination group was lower than that in the individual groups. Moreover, the cellular changes and apoptosis rates indicated that apoptosis in the combined application group was much greater than when either drug was applied individually. Fas expression by OS cell lines was increased by the combination of paclitaxel and cisplatin under hyperthermic conditions. More importantly, our study revealed low Fas expression in OS, which better explained the up-regulation of Fas achieved by the combination of paclitaxel and cisplatin in the presence of hyperthermia.

Conclusions: The combination of paclitaxel and cisplatin increases the effects of thermochemotherapy on OS cell lines, primarily through the induction of apoptosis by the up-regulation of Fas expression.     

Carboplatin-paclitaxel- and carboplatin-docetaxel-induced cytotoxic effect in epithelial ovarian carcinoma in vitro

BACKGROUND: The combination of paclitaxel and cisplatin is standard for patients with newly diagnosed epithelial ovarian carcinoma. The role of another taxane, docetaxel, currently is being studied. Due to its milder nonhematologic toxicity carboplatin increasingly is being substituted for cisplatin in taxane-based combinations. The purpose of this study was to compare the combination of carboplatin-paclitaxel with carboplatin-docetaxel in ovarian carcinoma in vitro, and to assess the type of interaction, if any. METHODS: Sensitivity to carboplatin and the concomitant use of a taxane and carboplatin was studied in 4 ovarian carcinoma cell lines using the 96-well plate clonogenic assay. Chemosensitivity was expressed as the IC50 value (i.e., the drug concentration causing 50% inhibition of clonogenic survival). IC50 values were obtained from dose-response curves after fitting the data to the linear quadratic equation. Synergism was studied by the area under the survival curve ratios (AUC ratios), obtained by numeric integration. The AUC ratio and the surviving fraction (SF) value after the administration of taxane alone were compared using the Student t test for paired data. RESULTS: The IC50 values for carboplatin were between 0.5-1.6 microgram/mL; there was only a 3.2-fold difference between individual cell lines. Carboplatin administered concomitantly with a taxane had either an additive or supra-additive growth inhibitory effect on all four ovarian carcinoma cell lines. A supra-additive effect occurred after simultaneous exposure of the cells to carboplatin at all tested paclitaxel concentrations in three of four cell lines (UT-OC-3, UT-OC-5, and SK-OV-3). The carboplatin-docetaxel combination had a supra-additive effect at the two highest docetaxel concentrations in two cell lines (UT-OC-4 and UT-OC-5) and at the highest docetaxel concentration in the other two cell lines (UT-OC-3 and SK-OV-3). CONCLUSIONS: Carboplatin has a synergistic effect when used with paclitaxel or docetaxel. A supra-additive effect is achieved with a wider range of paclitaxel concentrations than docetaxel concentrations.     

封闭ERCC1基因表达对卵巢癌细胞耐药的影响

目的:探讨通过封闭ERCC1基因表达干扰DNA损伤修复路径及其对卵巢癌细胞对铂类药物耐药性的影响.方法:构建ERCC1基因反义表达载体,转染人卵巢癌细胞A2780及A2750/CP70,采用Northern杂交及Western Blot分析人卵巢癌细胞内ERCC1基因RNA及蛋白表达水平变化;利用荧光素酶报告基因系统观察宿主细胞对DNA损伤的修复作用;采用MTT实验研究细胞对铂类药物耐药性的影响.结果:卵巢癌细胞转染反义ERCC1基因后,ERCC1基因转录水平、蛋白表达水平明显下降.荧光素酶报告基因系统结果证明,宿主细胞的DNA修复活性下降.MTT实验表明,伴随ERCC1基因表达水平下降,细胞对顺铂的耐药性降低.结论:转染ERCC1反义基因显著影响宿主细胞的核苷酸切除修复,影响细胞对顺铂的耐药性.     

PARP抑制剂对顺铂抑制肺癌细胞增殖影响的研究

目的：以顺铂（DDP）敏感和耐药肺癌细胞系为实验对象,探讨聚腺苷二磷酸核糖聚合酶[Poly（ADP—ribose）polymerase,PARP3抑制剂对DDP抑制肺癌细胞增殖作用的影响。方法：MTT法检测DDP、PARP抑制剂单药及其两者联合对DDP敏感和耐药肺癌细胞系增殖的抑制作用。应用蛋白免疫印迹方法,检测肺癌细胞系中PARP一1和PARP一2的表达,以及DDP激活PARP生成聚腺苷二磷酸核糖（Poly（ADP—ribose）,PAR]与PARP抑制剂对PARP的抑制作用。结果：蛋白质印迹法结果显示PARP：1在PC9、PCI4、SBC3亲代DDP敏感肺癌细胞系及其子代DDP耐药细胞系之间的表达差异有统计学意义,P值分别为0．028、0．013和0．008。DDP作用于肺癌细胞系激活PARP生成PAR在40min达到高峰,但在60min即迅速降低,且DPQ可抑制PAR的生成。而MTT结果显示,DPQ联合DDP抑制肺癌敏感和耐药细胞系的抗增殖作用与DDP单药作用比较差异均无统计学意义（P〉O．05,￡〈O）,表明DPQ没有增效DDP的作用。结论：在敏感和耐药肺癌细胞系中,PARP抑制剂与DDP联合无明显的协同增效作用,提示PARP-1表达差异与肺癌细胞对DDP敏感或者耐药无关,说明尚需进一步探讨PARP抑制剂与DNA损伤药物联合使用对肺癌的影响。     

In vitro oxygen-dependent survival of 2 human cell lines after radiation combined with tirapazamine (SR-4233) and cisplatin]

Recent data have shown that the in vitro and in vivo cytotoxicity of bioreductive drugs could be significantly increased by combination with ionising radiation or chemotherapy. Various parameters such as oxygen tension and timing of administration of the drugs could play a crucial role in the efficacy of combined treatment modalities. The aim of this study was to define the oxygen dependency of cell survival after in vitro irradiation and incubation with tirapazamine, a bioreductive drug, and cisplatin given alone or simultaneously. Two human cell lines were studied: one cell line sensitive to tirapazamine, Na11+, a pigmented melanoma with a high percentage of hypoxic cells, and a less sensitive cell line to tirapazamine, HRT18, a rectal adenocarcinoma. Gas changes were made to study cell survival at four different oxygen concentrations (pO2): air (20.9% O2), 10.2 and 0.2% O2. Cells were incubated with tirapazamine and cisplatin alone or combined for one hour at 37 degrees C, then irradiated and cultured. For Na11+, cell survival after irradiation was comparable in air and at 10% oxygen with the two drugs given alone or combined. At 2 and 0.2% oxygen, cell killing was largely increased by tirapazamine and was not modified by the addition of cisplatin. For HRT18, cell survival was not modified when cisplatin was added to radiation, whatever the oxygen partial pressure. At low pO2 (2 and 0.2%) the cytotoxic effect of tirapazamine was not significantly decreased by the addition of cisplatin. When cytotoxic and bioreductive drugs are combined to radiation, the magnitude of the observed effect is highly dependent on the partial oxygen pressure and on the sensitivity of the cell line to the individual drugs. This has very important implications for clinical strategies based on combined chemo-radiotherapy.     

The in vitro effects of lonidamine combined with cisplatin in human small cell lung cancer cell lines.

Two human small cell lung cancer (SCLC) cell lines were used to evaluate the in vitro response to lonidamine and cisplatin exposure. The two cell lines both showed growth inhibition when exposed to lonidamine alone at concentrations greater than 50 micrograms/ml; however, one cell line (H69) was more sensitive. When cisplatin was combined with lonidamine a synergistic interaction was observed when cells were exposed to 10 microM cisplatin for 1 hour combined with lonidamine at concentrations of 50 micrograms/ml or greater. At a concentration of 25 micrograms/ml lonidamine combined exposure with cisplatin had no effect on cell growth or viability.     

Herpes oncolytic therapy of salivary gland carcinomas

Bcl-xL functions as a dominant regulator of apoptotic cell death and is implicated in chemotherapeutic resistance of malignant pleural mesothelioma (MPM). Mesothelioma cell lines demonstrate increasing levels of Bcl-xL as resistant clones are selected in vitro. Moreover, upon introduction of antisense oligonucleotides specific to Bcl-xL mRNA, MPM cells are sensitized to chemotherapeutic agents. Here we describe the therapeutic effects of a novel combination therapy, Bcl-xL antisense oligonucleotide (ASO 15999) and cisplatin, on mesothelioma cell lines in vitro and in vivo; in addition, efficacy of ASO 15999 in decreasing tumor load as well as its effect on survival in an animal model. Finally, we initiated preliminary toxicity studies involved with intraperitoneal (IP) injections of ASO 15999 into mice. This novel combination, with doses of cisplatin four times below established IC(50) levels, significantly decreased viability of MPM cell lines after 48 hr. The growth of established mouse flank human tumor xenografts was reduced with intra-tumor administration of ASO 15999. Local spread and development of IP xenografts was reduced with treatments of ASO alone, and survival of mice afflicted with these xenografts was prolonged after administration of ASO alone and ASO 15999 + cisplatin combination therapy. These findings suggest that ASO 15999 sensitizes MPM cell lines to the toxic effects of cisplatin. ASO 15999 induced reduction of Bcl-xL is effective in slowing the progression of human mesothelioma cell lines both in vitro and in vivo. More notably, the combination of Bcl-xL ASO and cisplatin extends survival in an orthotopic tumor xenograft model.     

白杨素增强顺铂抗肿瘤作用的研究

目的：探讨白杨素增强顺铂抗肿瘤作用的最佳条件并筛选敏感细胞。方法：设立阴性对照组、空白调零组及40 μmol/L白杨素、5.0 μg/mL顺铂、40 μmol/L白杨素+5.0 μg/mL顺铂3个处理组,选取人结直肠癌细胞（HCT-116）、人鼻咽癌细胞（CNE1）、人肝癌细胞（HepG2）、人胃癌细胞（BGC-823）分别作用24 h。采用四甲基噻唑蓝（MTT）法测定肿瘤细胞的增殖抑制率,采用Hoechst 33342荧光染色法测定诱导肿瘤细胞的凋亡率,通过抑制率和凋亡率筛选出敏感细胞。采用L₉（3³）正交设计方法,设立3个白杨素水平（10、20、40 μmol/L）,3个顺铂水平（1.3、2.5、5.0 μg/mL）,以及3种作用时间（12、24、36 h）处理敏感细胞,确定白杨素增强顺铂抗肿瘤作用的最佳作用条件。结果：与白杨素或顺铂单独处理组相比,MTT试验结果表明,白杨素联合顺铂作用组对上述4种肿瘤细胞增殖的抑制率均明显增加,差异均具有统计学意义（P均 < 0.01）;Hoechst 33342染色实验发现,白杨素联合顺铂作用组诱导各肿瘤细胞的凋亡率亦均明显增加（P均 < 0.01）。白杨素联合顺铂作用组对人胃癌细胞BGC-823的增殖抑制率为（78.0±2.0）%、诱导细胞的凋亡率为（72.3±6.5）%。在4种细胞中,人胃癌细胞株BGC-823对白杨素联合顺铂抗肿瘤作用最敏感。正交实验的体外模型结果表明,最佳作用条件为白杨素40 μmol/L联合顺铂5.0 μg/mL,作用时间24 h。结论：白杨素能增强顺铂抑制人肿瘤细胞增殖和诱导人肿瘤细胞凋亡的作用。人胃癌细胞BGC-823是白杨素和顺铂联合作用的敏感细胞株。     

Antiproliferative action of metformin in human lung cancer cell lines

The oral antidiabetic agent metformin has anticancer properties, probably via adenosine monophosphate-activated protein kinase activation. In the present study, growth inhibition was assessed by a clonogenic and by a cell survival assay, apoptosis induction was assessed by Hoechst staining and caspase activities and cell cycle alteration after exposure to metformin, and the interaction of metformin with cisplatin in vitro were elucidated in four human lung cancer cell lines representing squamous, adeno-, large cell and small cell carcinoma. Clonogenicity and cell proliferation were inhibited by metformin in all the cell lines examined. This inhibitory effect was not specific to cancer cells because it was also observed in a non-transformed human mesothelial cell line and in mouse fibroblast cell lines. Inhibition of clonogenicity was observed only when the cells were exposed to metformin for a long period, (10 days) and the surviving fraction, obtained after inhibiting proliferation by increasing the dose, reached a plateau at approximately 0.1-0.3, indicating the cytostatic characteristics of metformin. Metformin induced significant apoptosis only in the small cell carcinoma cell line. A tendency of cell cycle accumulation at the G0/G1 phase was observed in all four cell lines. Cisplatin, in a dose-dependent manner, severely antagonized the growth inhibitory effect of metformin, and even reversed the effect in three cell lines but not in the adenocarcinoma cell line. The present data obtained using various histological types of human lung cancer cell lines in vitro illustrate the cytostatic nature of metformin and its cytoprotective properties against cisplatin.     

Combination photoimmunotherapy and cisplatin: effects on human ovarian cancer ex vivo.

BACKGROUND: Patients with ovarian cancer that is clinically resistant to cisplatin-based chemotherapy have little hope of a cure of their disease. Photoimmunotherapy, which involves the antibody-targeted delivery of a nontoxic photosensitizer that is activated to a cytotoxic state with visible light, may offer a new treatment option. Photoimmunotherapy may be applied intraperitoneally to target disseminated tumor. We tested the hypothesis that this treatment in combination with cisplatin potentiates cytotoxicity in ovarian cancer cell lines and primary cultures of human tumors. METHODS: Five human cancer cell lines (ovarian and breast) and 19 primary cultures were studied. The primary cultures were from solid and ascites tumor samples obtained from 14 patients with ovarian cancer who were undergoing primary surgery. The photosensitizer chlorin e(6) was conjugated to the F(ab')(2) fragment of the murine monoclonal antibody OC-125, which is directed against the antigen CA 125. Cytotoxicity was measured by the microculture tetrazolium assay. Treatments consisted of cisplatin alone, photoimmunotherapy alone, and photoimmunotherapy followed by cisplatin. The fractional product method was used to assess synergy in treatment effects. Ex vivo cultured human cells exhibiting 80% or greater survival at cisplatin concentrations of 10 microM for 24 hours were defined as cisplatin resistant for this study. RESULTS: When all cell types (cisplatin sensitive and cisplatin resistant) were considered together, combination treatment yielded cytotoxicity that was, on average, 6.9 times (95% confidence interval = 1.86-11.94) greater than that of cisplatin alone (two-sided P =.023). Cisplatin-resistant cells showed a synergistic effect of the two treatments (two-sided P =.044), while cisplatin-sensitive cells showed an additive effect. CONCLUSION: These ex vivo data suggest that platinum resistance in human ovarian cancer cells may be reversible by pretreatment with OC-125-targeted photoimmunotherapy. Further studies are required to confirm the efficacy of this approach in vivo.