Effect of cytokines on the toxicity of cytostatic drugs to normal bone marrow cells in vitro

We evaluated in vitro how growth factors influenced the effect of cytostatic drugs on normal hematopoietic progenitor cells. Bone marrow was obtained from 15 donors for bone marrow transplantation. After separation the mononuclear fraction was incubated with granulocyte colony-stimulating factor (G-CSF) at 5 and 50 ng/ml, with granulocyte/macrophage colony-stimulating factor (GM-CSF) at 1 and 10 ng/ml, and with interleukin 3 (IL-3) at 0.5 and 5 ng/ml for 24 h prior to incubation with cytostatic drugs. These incubations were performed with 0.05 μM mitoxantrone and 0.2 μM daunorubicin for 1 h, and cells were thereafter cultured for colony-forming units – granulocyte/macrophage (CFU-GM) in soft agar for 10–12 days. Incubation with 0.05 μM cytosine arabinoside was performed continuously throughout the culture period. The proliferation of normal hematopoietic progenitor cells stimulated with GM-CSF at 10 ng/ml and with IL-3 at 5 ng/ml was significantly increased to 218% and 215% colonies, respectively, as compared with the control stimulated with conditioned medium only. Stimulation with G-CSF, on the other hand, did not induce any significantly enhanced proliferation relative to the control. Daunorubicin applied in combination with G-CSF at 5 ng/ml or with IL-3 at 0.5 ng/ml exerted a significantly higher degree of cytotoxicity on normal hematopoietic progenitor cells, resulting in 21% and 30% surviving colonies as compared with the 38% recorded for daunorubicin alone (P<0.05). Neither G-CSF nor IL-3 at a higher concentration nor GM-CSF exerted a significantly altered degree of toxicity relative to cells incubated with daunorubicin alone. The cytotoxic effect exerted on normal hematopoietic cells by mitoxantrone or ara-C was unchanged or significantly decreased after stimulation with growth factors as compared with the effect on cells incubated with cytostatic drugs alone. We conclude that G-CSF and IL-3 augment the effect of daunorubicin on normal hematopoietic progenitor cells. Received: 6 May 1997 / Accepted: 4 February 1998     

Lactacystin Exhibits Potent Anti-tumor Activity in an Animal Model of Malignant Glioma when Administered via Controlled-release Polymers

Summary Lactacystin, a proteasome-inhibitor, has been shown to induce apoptosis of experimental gliomas in vitro. However, its systemic toxicity prevents further clinical use. To circumvent this problem, lactacystin can be delivered intratumorally. We tested the efficacy of lactacystin incorporated into controlled-release polymers for treating experimental gliomas. 9L-gliosarcoma and F98-glioma cell lines were treated with lactacystin (10–100 μg/ml) for 72 h in vitro. Cell-viability was measured with MTT-assays. Toxicity of lactacystin/polycarboxyphenoxypropane-sebacic-acid (pCPP : SA) polymers was tested in vivo using Fischer-344 rats intracranially implanted with lactacystin polymers loaded from 0.1 to 2% lactacystin by weight. The efficacy of 1, 1.3, 1.5 and 1.7% lactacystin/pCPP : SA polymers was determined in Fischer-344 rats intracranially challenged with 9L and treated either simultaneously or 5 days after tumor implantation. Lactacystin was cytotoxic in 9L cells, causing a 16 ± 8% growth inhibition at 10-μg/ml that increased to 78 ± 4% at 100-μg/ml. Similarly, lactacystin inhibited growth of F98 by 18 ± 8% at 10-μg/ml and 74 ± 2% at 100-μg/ml in vitro. Polymers released lactacystin for 21 days and intracranial implantation in rats neither generate local nor systemic toxicity at doses lower than 2%. Treatment with lactacystin/pCPP : SA polymers with loading concentrations of 1.0, 1.3, and 1.5% prolonged survival of animals intracranially challenged with 9L when polymers where inserted in the day of tumor implantation. In conclusion, lactacystin exhibits potent cytotoxic-activity against 9L and F98 in vitro, it can be efficiently incorporated and delivered using controlled-release polymers, and at the proposed concentrations lactacystin polymers are safe for CNS delivery and prolong survival in the 9L model. These authors equally contributed in all aspects of this study.     

Cell cycle perturbations in cisplatin-sensitive and resistant human ovarian carcinoma cells following treatment with cisplatin and low dose rate irradiation

Purpose: To investigate cell cycle pertubations in plateau-phase human ovarian carcinoma cells following treatment with cisplatin, low dose-rate irradiation (LDRI), or combined cisplatin and LDRI, in order to understand cell cycle mechanisms by which these two treatment modalities interact. Methods: Human ovarian carcinoma cells sensitive (A2780) and resistant (2780^CP) to cisplatin were grown to plateau phase and given protracted cisplatin treatments (A2780 0.7 and 2 μg/ml; 2780^CP 5 and 15 μg/ml) and/or LDRI (0.41 cGy/min). Cell cycle distribution following treatment was determined by two-parameter flow cytometry, based on bromodeoxyuridine (BrdU) uptake and DNA content using propidium iodide staining.Results: The cisplatin-sensitive A2780 cells exposed to cisplatin alone for up to 28 h showed depletion of the G1 fraction and accumulation in S-phase, although the percentage of S-phase cells actively incorporating BrdU dropped to almost zero. The cisplatin-resistant 2780^CP cells exposed to cisplatin alone showed a G1 arrest when exposed to 15 μg/ml, but not when exposed to 5 μg/ml. LDRI alone caused little cell cycle redistribution different from controls in either cell line. When LDRI was combined with cisplatin, no significant cell cycle redistribution was observed, apart from a decline in the actively incorporating S-phase fraction. Conclusions: Cisplatin caused A2780 cells to accumulate in nonincorporating S-phase, with no evidence of G1 arrest. Cisplatin-resistant 2780^CP cells showed a G1 block when exposed to a high enough cisplatin concentration. This could indicate a mechanism of cisplatin resistance in these cells. LDRI alone or in combination with cisplatin did not result in significant cell cycle redistribution. Received: 22 December 1995 / Accepted: 28 September 1996     

Anti-Tumor Effect of Curcumin on Human Cervical Carcinoma HeLa Cells In Vitro and In Vivo

Objective: To investigate the anti-tumor effect of curcumin on human cervical carcinoma HeLa cells in vitro and in vivo. Methods: (1) Human cervical carcinoma cell line HeLa was cultured in vitro. HeLa cells were treated with 5-50 μmol/L curcumin for 24. 48, 72 h and the growth inhibition rates of HeLa cells were measured by MTT method. Cell apoptosis was inspected by electron microscopy and flow cytometry (FCM). (2) A transplanted tumor model by injecting HeLa cells into subcutaneous tissue of BABL/C mice was established and its growth curve was measured. 30 BABL/C mice with tumors were divided into 2 groups at random and 0.2 ml saline or 0.2 ml 250 μmol/L curcumin was injected into abdominal cavity respectively once everyday and lasted for ten days. The changes of tumor volume were measured continuously and tumor inhibition rate was calculated. At last the expressions of caspase-3 and bax protein in transplanted tumors were detected by immunohistochemistry. Results: (1) Curcumin inhibited the proliferation of Lela cells on a dose-depending manner. Apoptosis of cells could be observed by FCM. Partial cells presented the characteristic morphological changes of apoptosis under electron microseope. (2) When 1×107 HeLa cells were inoculated for each mouse, 100% of the mice developed growing tumors after seven days. An inhibition effect was observed in treatment group, and the inhibition rate of curcumin was 74.33%. The expressions of caspase-3 and bax in the transplanted tumors were increased in curcumin group. Conclusion: Curcumin is effective as an anti-cancer drug not only in vitro but also in vivo.     

Antitumor effects of bevacizumab in combination with fluoropyrimidine drugs on human oral squamous cell carcinoma

Vascular endothelial growth factor (VEGF) serves an important role in new blood vessel formation or angiogenesis, which is a critical event in tumor growth and metastasis. Bevacizumab is a humanized monoclonal antibody against VEGF-A, whereas S-1 is a fluoropyrimidine antineoplastic agent that induces apoptosis in various types of cancer cells. The present study evaluated the antitumor effects of bevacizumab in combination with 5-fluorouracil (5-FU) or S-1 against oral squamous cell carcinoma (OSCC) in vitro and in vivo. Two human OSCC cell lines were used, namely the high VEGF-A-expressing HSC-2 cells and the low VEGF-A-expressing SAS cells. MTT assay was used to evaluate the effect of bevacizumab and/or 5-FU against HSC-2 and SAS cell proliferation. Additionally, the antitumor effect of bevacizumab was evaluated alone and in combination with S-1 against HSC-2 tumors in nude mice. S-1 (6.9 mg/kg/day) was administered orally every day for 3 weeks, and bevacizumab (5 ml/kg/day) was injected intraperitoneally twice per week for 3 weeks. Apoptotic cells in mouse tumors were detected using the TUNEL method, and cell proliferation and microvessel density (MVD) were determined by immunohistochemical staining of Ki-67 and CD31, respectively. Bevacizumab alone did not inhibit OSCC cell proliferation in vitro, and did not exhibit any synergistic inhibitory effect in combination with 5-FU in vitro. However, combined bevacizumab and S-1 therapy exerted synergistic and significant antitumor effects in vivo on HSC-2 tumor xenografts, and induced apoptosis in tumor cells. Furthermore, this combination therapy led to decreased MVD and cell proliferative abilities, as well as increased apoptosis in residual tumors. The present findings suggested that the bevacizumab plus S-1 combination therapy may exert antitumor effects in high VEGF-A-expressing OSCC cells.     

Novel paclitaxel formulations for oral application: a phase I pharmacokinetic study in patients with solid tumours

Purpose To explore the pharmacokinetics (PKs) of paclitaxel and two major metabolites after three single oral administrations of a novel drinking solution and two capsule formulations in combination with cyclosporin A (CsA) in patients with advanced cancer. Moreover, the tolerability and safety of the formulations was studied. In addition, single nucleotide polymorphisms in the multidrug resistance (MDR1) gene were determined. Patients and methods Ten patients were enrolled and randomized to receive CsA 10 mg/kg followed by oral paclitaxel 180 mg given as (1) drinking solution (formulation 1), (2) capsule formulation 2B, and (3) capsule formulation 2C on day 1, 8, or 15. Results The median C _max of paclitaxel was 0.42 (0.23–0.96), 0.48 (0.08–0.59), and 0.39 (0.11–1.03) μg/ml and the area under the plasma concentration–time curve was 2.83 (1.69–5.12), 2.01 (1.57–3.04), and 2.67 (1.05–3.61) μg h/ml following administration of formulations 1, 2B, and 2C, respectively. The novel formulations were tolerated after single oral dose without causing relevant gastrointestinal or haematological toxicity. Conclusions The PK and metabolism of paclitaxel were comparable between the oral formulations co-administered with CsA.     

Effect of Poly (ADP-ribose) Polymerase-1 Inhibition on the Proliferation of Murine Colon Carcinoma CT26 Cells

To investigate effect of poly (ADP-ribose) polymerase inhibition on the proliferation of CT26 cells in vitro and the mechanism of this effect. CT26 cells were treated with a range of concentrations of 5-Aminoisoquinolin-1-one (PARP inhibitor) in vitro. MTT assays and flow cytometry were used to determine the proliferation and cell cycle distribution, respectively, of the cells. The expression of PARP-1 was investigated by Western blot. The binding of Nuclear Factor-κB to DNA was detected by electrophoretic mobility shift assay. The proliferation of CT26 cells was significantly inhibited by 5-AIQ induced PARP inhibition in a dose-dependent manner. Proliferation was inhibited by 17.5% at 100 μM 5-AIQ, by 27.6% at 500 μM 5-AIQ and by 39.9% at 1000 μM (P < 0.05). After treatment with 5-AIQ, the proportion of cells in G₀/G_l phases increased and the proportion of cells in S phase decreased. The expression of PARP-1 was attenuated in 5-AIQ-treated CT26 cells (P < 0.05) and the binding of NF-κB to DNA binding was similarly diminished (P < 0.05). These results suggest that PARP inhibition reduced the proliferation of CT26 cells in vitro and influences the cell cycle. This inhibition is mediated by inhibiting PARP-1, which then diminishes the activity of NF-κB.     

Plasma concentrations of polysorbate 80 measured in patients following administration of docetaxel or etoposide

 Docetaxel (Taxotere, Rhone-Poulenc Rorer) and etoposide are water-insoluble drugs formulated with polysorbate 80 for intravenous administration. We have previously reported that surfactants, including polysorbate 80 and Cremophor EL, can reverse the multidrug resistance (MDR) phenotype in an experimental system and that plasma Cremophor EL concentrations measured following a 3-h infusion of paclitaxel were ≥1 μl/ml, sufficient to modulate MDR in vitro. The purpose of this study was to measure polysorbate 80 plasma concentrations in patients following intravenous administration of etoposide or docetaxel using a bioassay in which MDR-expressing cells are incubated with daunorubicin (DNR) plus 50/50 growth medium/plasma and equilibrium intracellular DNR fluorescence is measured by flow cytometry. In vitro experiments show maximal reversal of MDR at concentrations of 1.0–2.0 μl/ml and 50% reversal at 0.2–0.3 μl/ml. Patients received docetaxel at 75 mg/m² (five patients) or 100 mg/m² (four patients) (total dose 125–178 mg, containing 3.12–4.45 ml polysorbate 80) over 60 min. The median end-infusion polysorbate 80 concentration was 0.1 μl/ml (range 0.07–0.41 μl/ml). Only one patient had a level of >0.2 μl/ml. Five patients received intravenous etoposide at 120 mg/m² over 45–120 min (total dose 180–250 mg, containing 0.67–0.93 ml polysorbate 80). In the end-infusion plasma sample, polysorbate 80 was not detectable (<0.06 μl/ml) in any patient. Plasma polysorbate 80 levels following an intravenous infusion of 120 mg/m² etoposide or of docetaxel at doses used in Phase II trials, are insufficient to show modulation of MDR in vitro. Received: 21 July 1996 / Accepted: 4 November 1996     

Antiproliferative activity of contragestazol (DL111-IT) in murine and human tumor models in vitro and in vivo

Purposes: To evaluate the antiproliferative activity of contragestazol (DL111-IT) in vitro and in vivo and to elucidate potential molecular mechanisms. Methods: Cell killing ability of DL111-IT was measured by MTT/Trypan blue exclusion method and murine and human tumor models; cell cycle was analyzed by flow cytometry; pRb, CDK4 and Cyclin D1 expressions were detected by western blotting. Results: DL111-IT exhibited high efficiency on cell growth inhibition of 12 cancer cell lines, the IC₅₀ values were 4.1–19.7 μg/ml. In Sarcoma-180 (S180) and Hepatoma-22 (H22) tumor bearing mice models, the inhibition rates were 55.9 and 55.6%, respectively, at the doses of DL111-IT 12.5–50.0 mg/kg for 9 days consecutive administration. Human ovarian carcinoma (HO-8910) xenograft study showed that, nine administrations (within 15 days) of DL111-IT (12.5–50.0 mg/kg) significantly inhibited tumor growth with the inhibition rates ranging from 17.0 to 64.3%. DL111-IT induced G1 arrest and overexpression of pRb, CDK4 and Cyclin D1 were observed in HO-8910 cell line, suggesting that cell cycle regulation might contribute to the anticancer property of DL111-IT. Conclusions: DL111-IT could inhibit the proliferation of cancer cells both in vitro and in vivo via a cell cycle regulation pathway.     

Cisplatin’s tumoricidal effect on human breast carcinoma MCF-7 cells was not attenuated by American ginseng

Purpose We previously observed that American ginseng berry and ginsenoside Re attenuated cisplatin-induced emesis in a rat model, suggesting that the herb may have a value in treating chemotherapy-induced nausea/vomiting. However, it is not clear whether consuming ginseng concurrently with chemotherapy affects the efficacy of chemotherapeutic agents. In this study, we explored if the ginseng extract and its constituents, ginsenosides Rb₁, Rb₃, and Re, alter tumoricidal activity of cisplatin in human cancer cells. Methods Tumoricidal effects of cisplatin, and/or American ginseng berry extract (AGBE) and ginsenosides Rb₁, Rb₃, and Re, on human breast carcinoma MCF-7 cells were measured as cell proliferation in vitro. Cell counts were performed in MCF-7 cells pretreated with test agents for 72 h. Results Cisplatin decreased MCF-7 cell proliferation significantly in a concentration-dependent manner. Compared to control group, cisplatin reduced the cell proliferations to 56.5±3.3% at 1 μg/ml, to 36.6±2.4% at 5 μg/ml, and to 26.9±2.4% at 15 μg/ml (P<0.01). AGBE also inhibited the cell proliferation significantly, although in a less extended manner. When the berry extract at 0.5 mg/ml was used with cisplatin at 1 μg/ml, a significant enhancement of cisplatin’s activity was observed (35.8±2.5%; P<0.05). We also observed that Rb₁ did not change cisplatin’s activity; Rb₃, at a higher concentration, increased cisplatin’s anti-proliferation activity (48.0±1.2%; P<0.05); Re increased cisplatin’s activity (Re 0.1 mg/ml, 48.0±2.8%; Re 0.3 mg/ml, 31.9±2.2%, P<0.01). Conclusion Our data suggest that AGBE and the tested ginsenosides do not attenuate cisplatin’s tumoricidal activity in MCF-7 cells, but in fact may actually enhance it. Additionally, the ginseng extract and ginsenoside Re by themselves exerted anti-proliferative activity against MCF-7 cells. The herb might potentially serve a complementary role with the chemotherapeutic agents in treating cancer, in addition to decreasing chemotherapy-induced nausea/vomiting.     

Anti-tumor Activities of Novel Estrogen Compound 17a α-D-Homo-Ethynylestradiol-3-Acetate

Objective： To study the anti-tumor activities of novel estrogen compound 17a α-D-homo-ethynylestradiol-3-acetate in vitro and in vivo. Methods： In vitro anti-tumor activity was assayed in adenoma cells A549 and human liver cancer cells Bel-7402 using MTT method, and half-inhibitory concentration （IC50） were observed. In vivo the pulmonary adenoma LA795 cells was selected and the conventional assay method of anti-tumor activity was employed. 5, 7.5, 10 mg/kg of 17a α-D-homo-ethynylestradiol-3-acetate was administered by i.p., and tumor-inhibitory rate, thymus and spleen indexes, bone marrow cells （BMC） were observed. Results： IC50 of 17a α-D-homo-ethynylestradiol-3-acetate in vitro for A549 and Bel-7402 cells were 12.28 μg/ml and 17.79 μg/ml, respectively. In vivo the highest tumor-inhibitory rates for LA795 was 60.0% （P〈0.01）. The drug had hardly any side-effect in spleen indexes, thymus indexes, and BMC compared with control mice. Nevertheless, compared with the positive control drug cyclophosphamide （CY）, thymus and spleen indexes, BMC showed obvious differences （P〈0.01）. Conclusion： 17a α-D-homo-ethynylestradiol-3-acetate has obvious anti-tumor activities in vitro and in vivo with low side-effect, thus worth further investigation.     

Antitumor activity and low intestinal toxicity of S-1, a new formulation of oral tegafur,in experimental tumor models in rats

 S-1, a new oral antitumor agent, is composed of 1-(2-tetrahydrofuryl)-5-fluorouracil (Tegafur, FT), 5-chloro-2,4-dihydroxypyridine (CDHP) and potassium oxonate (Oxo) in a molar ratio of 1 : 0.4 : 1. FT which is a masked compound of 5-fluorouracil (5-FU) acts as an effector, while both CDHP and Oxo which do not have antitumor activity themselves act as modulators. In this study, the antitumor activity and intestinal toxicity of S-1 were investigated using experimental tumor models in rats, and compared with those of other oral fluoropyrimidines, namely 5-FU, FT, FCD (1 M FT/0.4 M CDHP) and UFT (combination of FT and uracil). In rats bearing subcutaneous Yoshida sarcoma, S-1 inhibited tumor growth at the lowest dose (ED₅₀ value: S-1 5, UFT 22, FT 82, FCD 5, and 5-FU 19 mg/kg per day), and induced the least host body weight suppression, leading to the highest therapeutic index (TI) (S-1 4.5, UFT 1.4, FT 1.8, FCD 2.0, and 5-FU 1.4). S-1 also showed a higher therapeutic effect than UFT against AH-130 and Sato lung carcinoma. After administration of S-1 and UFT at equitoxic doses, S-1 showed a higher and more prolonged concentration of 5-FU than UFT both in plasma (AUC_0-∞: S-1 28 nmol h/ml, UFT 15 nmol⋅h/ml) and in tumor tissue (AUC_0-∞: S-1 95 nmol h/g tissue, UFT 52 nmol h/g tissue), leading to a higher 5-FU level incorporated into the RNA fraction (F-RNA level) in tumor tissue (AUC_0-24: S-1 7.0 nmol h/mg RNA, UFT 4.3 nmol h/mg RNA) and 5–8% higher thymidylate synthase (TS) inhibition in tumor tissue at every time-point through 24 h. Compared with other oral fluoropyrimidines after administration of the maximal tolerable dose (MTD), S-1 caused the lowest rates of intestinal toxicities, such as diarrhea and occult blood in feces. S-1 also showed a higher antitumor effect on Yoshida sarcoma implanted intracolonically than UFT at an equitoxic dose (tumor weight: S-1 64±30 mg, UFT 133±52 mg; P<0.05). These results suggest that CDHP, which is a potent inhibitor of 5-FU degradation, increases the antitumor activity of FT, and that Oxo, which is an inhibitor of 5-FU phosphorylation, locally protects the gastrointestinal tract from 5-FU-induced toxicity without decreasing the antitumor activity. Received: 13 October 1996/Accepted: 18 May 1996     

Radiosensitizing potential of the selective cyclooygenase-2 (COX-2) inhibitor meloxicam on human glioma cells

The COX-2 protein is frequently overexpressed in human malignant gliomas. This expression has been associated with their aggressive growth characteristics and poor prognosis for patients. Targeting the COX-2 pathway might improve glioma therapy. In this study, the effects of the selective COX-2 inhibitor meloxicam alone and in combination with irradiation were investigated on human glioma cells in vitro. A panel of three glioma cell lines (D384, U87 and U251) was used in the experiments from which U87 cells expressed constitutive COX-2. The response to meloxicam and irradiation (dose-range of 0–6 Gy) was determined by the clonogenic assay, cell proliferation was evaluated by growth analysis and cell cycle distribution by FACS. 24–72 h exposure to 250–750 μM meloxicam resulted in a time and dose dependent growth inhibition with an almost complete inhibition after 24 h for all cell lines. Exposure to 750 μM meloxicam for 24 h increased the fraction of cells in the radiosensitive G₂/M cell cycle phase in D384 (18–27%) and U251 (17–41%) cells. 750 μM meloxicam resulted in radiosensitization of D384 (DMF:2.19) and U87 (DMF:1.25) cells, but not U251 cells (DMF:1.08). The selective COX-2 inhibitor meloxicam exerted COX-2 independent growth inhibition and radiosensitization of human glioma cells.     

The Inhibitory Effect of Oridonin on the Growth of Fifteen Human Cancer Cell Lines

OBJECTIVE To study the inhibitory effect of oridonin on the growth of cancer cells. METHODS Fifteen human cancer cell lines were subjected to various concentrations of oridonin in culture medium. The inhibitory rate of cell growth was measured by the MTT assay, and compared with a negative control and 5-Fu-positive control. RESULTS The 50% inhibiting concentration （IC50） and maximal inhibition （Imax） of oridonin shown by studying the growth of the cancer cell lines were as follows： leukemias （HL60 cells： 3.9 μg/ml and 73.8%, K562 cells： 4.3 μg/ml and 76.2%）; esophageal cancers（SHEEC cells： 15.4 μg/ml and 99.2%, Eca109 cells： 15.1 μg/ml and 84.6%, TEl cells： 4.0 μg/ml and 70.2%）; gastric cancers （BGC823 cells： 7.6 μg/ml and 98.7%, SGC7901 cells： 12.3 μg/ml and 85.7%）; colon cancers （HT29 cells： 13.6 μg/ml and 97.2%, HCT cells： 14.5 μg/ml and 96.5%）; liver cancers （Bel7402 cells： 15.2 μg/ml and 89.2%, HepG2 cells： 7.1 μg/ml and 88.3%）; pancreatic cancer （PC3 cells： 11.3 μg/ml and 68.4%）; lung cancer （A549 cells： 18.6 μg/ml and 98.0% ）; breast cancer （MCF7 cells： 18.4 μg/ml and 84.7%）; uterine cervix cancer （Hela cells： 13.7 μg/ml and 98.5%）. CONCLUSION Oridonin had a relatively wide anti-tumor spectrum, and a relatively strong inhibitory effect on the growth of the 15 human cancer cells. Inhibitory effects were concentration dependent.     

Electropermeabilization in bladder cancer chemotherapy

 Purpose: Electropermeabilization has been used for the introduction of genes into cells. Using this technique, we introduced the cytotoxic drug bleomycin (BLM) into cells and examined whether the technique might be useful for the treatment of bladder cancer. Materials and methods: For electropermeabilization in vitro, we used YTS-1 cells, a human transitional cell carcinoma line. Aliquots of cell suspension were mixed with a solution of BLM and immediately exposed to electric pulses. A high-power pulse generator was used to supply square-shaped pulses of 1250 V/cm (100 μs, eight pulses). After a 2-h post-shock incubation, cells were washed and incubated for one further hour. Then the concentration of BLM in the cells was measured using a bioassay. For electropermeabilization of tissue, we used normal male Wistar rats. The bladder was exposed and 10 mg/kg BLM was injected into the caudal vein. A series of eight pulses with a time constant of 100 μs at an electric field intensity of 1000 V/cm was applied. The bladder, liver and lungs were extracted 1 h later and prepared for quantification of the BLM concentration using the bioassay. Results: Electrotreated cells contained significantly higher concentrations of BLM than nonelectrotreated cells. The concentration of BLM 1 h after electrotreatment in bladder tissue was 2.7 times higher than that in nonelectrotreated bladder tissue. Conclusion: The electropermeabilization technique has the potential to serve as a new and effective modality for the treatment of bladder cancer. Received: 8 August 1995/Accepted: 18 March 1996     

The anticancer effect and anti-DNA topoisomerase II effect of extracts of camellia ptilophylla chang and camellia sinesis

The cytotoxic effect of extract of camellia ptilophylla chang (ECPC) and extract of camellia sinensis (ECS) on HeLa cell line, poorly differentiated nasopharyngeal carcinoma cell line (CNE₂) and gastric cancer cell line (MGC-803)in vitro was studied using MTT assay method. The results showed that ECPC and ECS possessed significant cytotoxic effect on above three cell lines. The anticancer test in mice showed that ECPC had marked inhibitory effect against Ehrlich solid carcinoma (ESC) with inhibition rates of 17.8–48.3% and with inhibition rates of 28.3–54.5% against reticular cell sarcoma (L₂) and that ECS had inhibition rates of 31.5–49.4% against ESC and 35.8–50% against L₂. These two extracts had only marginal inhibitory effect against sarcoma- 180. The unknotting activity of DNA topoisomerase II was inhibited completely by ECPC and ECS at the concentration of 50 μg/ml suggesting that DNA topoisomerase II might be a target enzyme of these two extracts.     

The significance of the sequence of administration of topotecan and etoposide

 Purpose: Often the best method of integrating chemotherapeutic agents is unknown. Recently there has been interest in the use of combinations of the topoisomerase II inhibitors and the topoisomerase I inhibitors as these agents have shown individual activity in malignancies such as non-small-cell lung cancer. This study examined the interaction of the topoisomerase II inhibitor etoposide with the topoisomerase I inhibitor topotecan (Tpt) in V79 cells (hamster lung fibroblast cells) to determine the optimal method of delivering these agents. Methods and results: Cell survival was assessed by colony formation. Synergistic interactions were assessed by the median effect principle in which a combination index (CI) of less than one suggests a synergistic interaction. The V79 cells were exposed to sequential 24-h incubations with the two chemotherapeutic agents. Initially, equitoxic doses of the two agents were delivered (i.e. 0.0275 μg/ml of topotecan alone or 0.089 μg/ml of etoposide alone resulting in a surviving fraction of 70%; Tpt : etoposide ratio 1 : 3.2). It was determined that a sequence-dependent synergistic interaction (CI<1) resulted at a lower level of cytotoxicity if the etoposide exposure followed the Tpt exposure compared to the opposite sequence. This same effect was seen after treatment of cells with various concentration (μg/ml) ratios of Tpt : etoposide (1 : 4.0, 1 : 1, 2.5 : 1). Conclusions: These results suggest that maximum synergy occurs for the delivery of etoposide following Tpt exposure (compared to the opposite sequence) and these findings may have important clinical implications. Received: 29 September 1995/Accepted: 25 March 1996     

Temozolomide treatment does not affect topiramate and oxcarbazepine plasma concentrations in chronically treated patients with brain tumor-related epilepsy

Objective Medical management of brain tumor-related epilepsy is complicated by interactions between antiepileptic and chemotherapeutic drugs. We studied the effect of temozolomide therapy on the disposition of the new antiepileptic drugs topiramate (TPM) or oxcarbazepine (OXC). Methods Fifteen patients chronically treated with TPM or OXC in monotherapy starting a chemotherapeutic treatment with temozolomide were enrolled in the study, of which ten were available for the final analyses. Blood samples were collected before temozolomide treatment (T₀), at its end (T₇) and after further 1–3 weeks (T₁₄–T₂₈). For each patient, more than one treatment cycle was studied. Topiramate and OXC mono-10-hydroxy derivative (MHD) plasma concentrations were determined by hplc coupled with ion spray mass spectrometer (TPM) or ultraviolet (MHD) detection. Results Mean TPM concentrations were 5.4 ± 2.4 μg/ml at T₀ vs. 5.5 ± 2.4 μg/ml at T₇ (n = 14), and 5.4 ± 2.4 μg/ml at T₀ vs. 5.6 ± 2.8 μg/ml at T₁₄–T₂₈ (n = 14). Mean MHD concentrations were 16.4 ± 7.6 μg at T₀ vs. 18.5 ± 9.0 μg/ml at T₇ (n = 5), and 16.8 ± 7.0 μg/ml at T₀ vs. 18.0 ± 8.7 μg/ml at T₁₄–T₂₈ (n = 8) (all comparisons not statistically significant; Student’s t-test for paired samples). Conclusion Temozolomide treatment did not affect TPM plasma concentrations in chronically treated patients. Data for MHD in OXC-treated patients were similar, but, due to the small sample size, results should be interpreted cautiously.These findings confirm that TPM (and possibly OXC) are a reasonable choice of antiepileptic drug in patients with brain tumor-related epilepsy.     

THE ANTITUMOR ACTIVITIES OF GNIDIMACRIN ISOLATED FROM STELLERA CHAMAEJASMEL．

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Asterosaponin 1, a cytostatic compound from the starfish Culcita novaeguineae, functions by inducing apoptosis in human glioblastoma U87MG cells

Summary Malignant glioblastoma is one of the most common malignant tumors in the neurological system. Asterosaponin 1, a new cytostatic agent from the starfish Culcita novaeguineae appear to exhibit various biological activities, including antitumor effect, but the function and mechanism of this new agent on glioblastoma cells has not previously been determined. In the present study, we investigated the proliferation change of human glioblastoma U87MG cells exposed to different concentrations (2.5–20.0 μg/ml) of asterosaponin 1 for a certain time. The results showed that asterosaponin 1 significantly suppressed U87MG cell proliferation in a time- and dose-dependent manner (IC₅₀ =4.3 μg/ml). Flow cytometric analysis of DNA in U87MG cells showed that asterosaponin 1 induces the prominent appearance of a sub-G1 peak in the cell cycle suggestive of apoptosis identical with the result of annexin V/PI assay. Furthermore, U87MG cells treatment with asterosaponin 1 resulted in nuclear condensation with apoptotic bodies observed by both fluorescence and electron microscopy. Agarose gel electrophoresis of DNA from asterosaponin 1-treated cells revealed a typical “ladder” consistent with apoptotic DNA fragmentation. Western-blot staining showed asterosaponin 1 decreased the expression of Bcl-2 protein and increased the expression of Bax protein. The novel findings suggest that the cytostatic actions of asterosaponin 1 toward U87MG cells result from the induction of cell apoptosis. Overall, our data demonstrate that asterosaponin 1 is fully equipped for an efficient apoptotic killing of glioblastoma cells and suggest that this mechanism may play a critical role in anti-tumor chemotherapy.Guang Cheng, Xiang Zhang and Hai-Feng Tang contributed equally to this work