Biomedical subjects

N Saijo

Publications and source records attributed to N Saijo.

At least 163 records · Page 9Linked to original sources

PubMed1996

A topoisomerase II inhibitor, NK109, induces DNA single- and double-strand breaks and apoptosis.

2,3-(Methylenedioxy)-5-methyl-7-hydroxy-8-methoxybenzo[c]phenanthr idinium hydrogensulfate dihydrate, called NK109, is a benzo[c]phenanthridine derivative, which inhibits DNA topoisomerase II activity by stabilizing the DNA-enzyme-drug complex, and shows strong growth-inhibitory effects on several human cancer cells. In the present study, NK109 treatment induced DNA fragmentation and a rise in the level of cytoplasmic nucleosomes, which are markers of apoptosis, in human small-cell lung carcinoma SBC-3 cells. These effects were inhibited by zinc ions and enhanced by cycloheximide or actinomycin D. Dose-dependent single- and double-strand DNA breaks were observed, using alkaline and neutral elution assays, in SBC-3 cells treated with more than 0.2 microM NK109 for 4 h. Treatment with NK109 caused more DNA single- and double-strand breaks than treatment with an equimolar amount of VP-16. These results suggest that NK109 induces DNA strand breaks and apoptosis. In addition, it appears that this process does not require protein or RNA synthesis, but involves a specific endonuclease which is inhibited by zinc ions.

DNA↗

PubMed1996

Roentgenographically occult small-cell lung cancer: case report and review of the literature.

This report describes a rare case of roentgenographically occult small-cell lung cancer in a 73-year-old man with hemoptysis. Fiberoptic bronchoscopy disclosed a 5-mm dome-shaped lesion; a biopsy established the diagnosis of small-cell lung cancer. The patient received a combination of chemotherapy and radiotherapy. More than 10 years later, he is still alive without recurrent disease. A review of the literature of roentgenographically occult small-cell lung cancer revealed the following: (1) a history of heavy smoking was common; (2) double primary bronchogenic carcinoma was noted; (3) hemoptysis or bloody sputum was an initial common symptom; (4) the sensitivity of sputum cytologic analysis was relatively low; (5) the tumor, which was shiny, smooth, and covered with bronchial epithelium, was often located at the bifurcation; and (6) lymph node metastatic involvement occurred.

Aged↗

PubMed1996

[Anticancer agents and apoptosis].

We reviewed recent investigations on apoptosis related to anticancer chemotherapy. The study of programmed cell death, apoptosis, has become one of the main stream in cellular biology, particularly in immunology, developmental biology and oncology. To determine whether the apoptotic cell death induced by anticancer agents could be inhibited by bcl-2 oncogene, we established a bcl-2 transfected human small cell lung cancer cell line, SBC-3/Bcl-2. SBC-3/Bcl-2 showed higher resistance to ADM, CPT-11 and MMC compared to the parental line SBC-3. Agarose gel electrophoresis showed typical DNA fragmentation of SBC-3 following treatment with CPT-11 or MMC. In contrast, the same concentration of the drugs did not induce DNA fragmentation in SBC-3/Bcl-2. However, there was no difference in sensitivity to CDDP, VP-16, ACNU, MTX and Taxol between SBC-3 and SBC-3/Bcl-2 (Ohmori T, et al: Biochem Biophys Res Commun 1993). These results suggest that bcl-2 can modulate the cytotoxicity of some anticancer agents by inhibiting the process of apoptosis. We speculate that some apoptotic pathways are bcl-2-dependent and others bcl-2-independent.

Animals↗

PubMed1996

[Early phase II study of BMS-181339 (paclitaxel) in patients with non-small cell lung cancer. BMS-181339 Non-Small Cell Lung Cancer Study Group].

We conducted a multi-institutional (11 facilities), early phase II study of BMS-181339 (paclitaxel), a novel anti-cancer drug, for non-small cell lung cancer (NSCLC). The 150 mg/m2 dose of paclitaxel was given by intravenous infusion over 24 hours every three weeks. When fifteen patients were accumulated, the interim review revealed that three of 15 eligible patients had a partial response for a response rate of 20%. The most common toxic effects were grade 3 or 4 leukopenia seen in 73.3% (11/15), and grade 4 neutropenia in 93.3% (14/15). One patient with neutropenia had suspected septic shock, which could be managed by G-CSF and antibiotics. No serious hypersensitivity reaction was seen with premedication of anti-allergic drugs, although mild allergic reactions such as skin rash and flush, were observed in 20.0% (3/15). Other adverse reactions, including alopecia, fever, arthralgia, myalgia and peripheral neuropathy, were mild in most cases. We conclude that it is relevant to proceed to a late phase II study for NSCLC.

Adult↗

PubMed1996

[Selection of drugs for chemotherapy based on drug resistance marker].

There is no appropriate tumor marker for the selection of anti cancer drug. Some agents can be selected for the reversal of anti cancer drug resistance. For example, verapamil or cyclosporin A may be useful for p-glycoprotein related multidrug resistance, and amphotericin B, docosahexaenoic acid or 8-chloro cAMP can be used for the modification of cisplatin-resistance. Recently, bcl-2 or mutated p53 gene are demonstrated to be important markers for drug resistance. More studies are necessary to identify an appropriate markers for drug resistance and overcome it.

Biomarkers, Tumor↗

PubMed1996

Characterization of the protein kinase C signal transduction pathway in cisplatin-sensitive and -resistant human small cell lung carcinoma cells.

Protein kinase C (PKC) influences cellular sensitivity to cis-diamminedichloroplatinum(II) (cDDP). We have investigated whether the PKC signal transduction pathway is affected during the development of cellular resistance to cDDP. Activators of PKC, such as phorbol 12,13-dibutyrate (PDBu), enhanced the sensitivity of human small cell lung cancer H69 cells to cDDP by 2-fold but had no effect on the sensitivity of cDDP-resistant H69 cells (H69/CP) to cDDP. The maximum sensitization was achieved with 10 nM PDBu and blocked by down-regulation of PKC with higher concentrations of PDBu (1 microM) or bryostatin 1 (0.1 microM). PKC activity was decreased significantly in H69/CP cells compared to the drug-sensitive variant. A similar reduction in PKC activity was noted in ovarian carcinoma 2008 cells that were resistant to cDDP. A modest decrease in PKC activity was also observed in etoposide-resistant H69 (H69/VP-16) cells but not in Taxol-resistant H69 cells or bleomycin-resistant human head and neck carcinoma A-253 cells. H69 cells expressed conventional PKC alpha and-beta, novel PKC delta, atypical PKC zeta and-iota, and novel/atypical PKC mu. A decrease in cPKC alpha and-beta and an increase in nPKC delta were associated with the cDDP-resistant phenotype. The abundance of aPKC zeta or-iota was unaffected. H69/ VP-16 cells also displayed a reduction in cPKC beta and an increase in nPKC delta. Taxol-resistant H69 cells had no alteration in the expression of any of the PKC isozymes. Thus, a reduction in cPKCs and an increase in nPKC may be associated with cDDP resistance.

Antineoplastic Agents↗

PubMed1996

[Chemotherapy for non-small cell lung cancer].

The combination of cisplatin and either a vinca alkaloid or etoposide is one of the most effective chemotherapeutic regimens for non-small cell lung cancer. Meta-analyses of clinical trials in which cisplatin-based combination chemotherapy was compared with supportive care in patients with non-resectable non-small cell lung cancer showed that the chemotherapy reduced mortality. In the last decade, new agents, including vinorelbine, edatrexate, paclitaxel, docetaxel, irinotecan, topotecan and gemcitabine, have shown promise in the treatment of non-small cell lung cancer, and new agents combined with platinum compounds have reached the level of phase III testing. Randomized studies in which radiotherapy alone was compared with radiotherapy and cisplatin-based combination chemotherapy in patients with stage III disease showed that combined treatment can prolong survival and improve long-term outcome in patients with locally advanced non-small cell lung cancer. Recently, new agents combined with radiotherapy have been tested. Regimens in which platinum compounds are combined with new agents hold promise for more successful treatment of this disease.

Antineoplastic Agents, Phytogenic↗

PubMed1996

Serum levels of cytokines in patients with untreated primary lung cancer.

To evaluate the relationships between serum endogenous cytokine levels and their clinical implications in cancer patients, we measured the serum levels of endogenous granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), macrophage colony-stimulating factor (M-CSF), and interleukin 6 (IL-6) in patients with untreated primary lung cancer. The serum G-CSF level was measured using a chemiluminescent ELISA, and the other cytokine levels were measured using ELISA. Fifty healthy adults and 183 patients with primary lung cancer were studied. The mean M-CSF level in the lung cancer patients (1106.4 units/ml) was significantly higher than that in the healthy adults (836 units/ml, P = 0.0001). In patients with large cell carcinoma, endogenous G-CSF, M-CSF, and IL-6 levels were significantly higher than those in patients with carcinomas of other cell types (P < 0.05). Univariate analysis showed that survival of 159 non-small cell lung cancer patients with high (more than cutoff level) G-CSF, M-CSF, and IL-6 levels was significantly poorer than that of patients with low levels (Wilcoxon's test, P = 0.018, P < 0. 0001, and P < 0.0001, respectively). Survival of patients with high levels of two or more cytokines was poorer than that of those with high levels of one cytokine or normal cytokine levels (P < 0.0001). Multivariate analysis using Cox's proportional hazards model showed that high M-CSF and C-reactive protein levels correlated significantly with poor survival (P = 0.037 and 0.037, respectively). Our preliminary data suggest that high M-CSF levels in non-small cell lung cancer may be of poor prognostic value.

Adult↗

PubMed1996

Phase II study of 3-hour infusion of paclitaxel in previously untreated non-small cell lung cancer.

Paclitaxel has clinical activity in non-small cell lung cancer, with response rates of 21 and 24% in a 24-h infusion. Recent clinical studies have shown that a 3-h infusion of the drug with premedication did not result in hypersensitivity reactions, and that neutropenia was milder in the 3-h than in the 24-h schedule. In this Phase II study, we tried to evaluate the efficacy and toxicity of paclitaxel given over 3 h in patients with previously untreated, unresectable stage III or IV non-small cell lung cancer. In addition, we attempted to investigate the pharmacokinetics and pharmacodynamics of the drug. Paclitaxel was administered i.v. over 3 h at a dose of 210 mg/m2 every 3 weeks with premedication of dexamethasone, ranitidine, and diphenhydramine. Heparinized blood samples were obtained from 12 patients for pharmacokinetic studies. Twenty-three (38%) of 60 assessable patients achieved a partial response, with a median duration of 3.2 (range, 2.3-11.1) months. The median survival for all patients was 11.2 months, and the 1-year survival rate was 48%. Thirty (50%) patients developed grade 4 neutropenia. Nonhematological toxicities were mild, except for pulmonary toxicity in one (1.7%) patient who required mechanical ventilatory support for 4 days. The duration of the paclitaxel concentration above 0.1 microM correlated well with the percentage of decrease in the absolute neutrophil count. In conclusion, a 3-h infusion of paclitaxel was safe and probably not less effective than a 24-h infusion.

Adult↗

PubMed1995

Phorbol ester and okadaic acid-resistant cells: the crossroads of signal transduction and drug resistance.

Many factors are involved in the development of drug resistance for anticancer drugs. The drugs should pharmacokinetically attain the appropriate concentration. They should be metabolized to the active forms. Tumor cells should have sensitivity to them. Several molecular and biochemical mechanisms that may explain cellular drug resistance have been identified. The contribution of protein phosphorylation and dephosphorylation for drug resistance is demonstrated in phorbol ester and okadaic-acid-resistant cells. The modulation of drug resistance by substances that affect the signal transduction pathway is an important issue in the development of an effective method for overcoming drug resistance.

Antineoplastic Agents↗

PubMed1995

Enhanced interaction between tubulin and microtubule-associated protein 2 via inhibition of MAP kinase and CDC2 kinase by paclitaxel.

Paclitaxel, an anti-mitotic anti-cancer agent, is active against solid tumors. The inhibition of depolymerization and promotion of microtubular assembly are essential for the anti-tumor activity of paclitaxel. Microtubule-associated proteins (MAPs) co-polymerize with tubulin and play some roles in microtubular dynamics. We examined the effect of paclitaxel on the interaction between tubulin and MAPs. Human lung-cancer cells, PC-14, were synchronized to G1/S border by the thymidine-double-block technique. After release from exposure to thymidine, the cells were treated briefly with 2 nM paclitaxel and the levels of alpha and beta tubulins and MAPs were examined after various times. Immunoblot analysis of paclitaxel-treated cells showed no changes in the overall expression of alpha and beta tubulins, microtubule-associated protein 2 (MAP2) or MAPs in comparison with controls. The samples were immunoprecipitated with anti-alpha- and anti-beta-tubulin antibodies and reblotted with an anti-MAP2 antibody, which showed that the amount of co-immuno-precipitated MAP2 in the synchronized cells, were increased by the brief paclitaxel treatment. These results suggest that paclitaxel treatment enhances the interaction between alpha and beta tubulins and MAP2. Since the phosphorylation state of MAP2 regulates the affinity of MAP2 for tubulins, and mitogen-activated protein (MAP) kinase is considered to be one of the kinases responsible for MAP2 phosphorylation, the effect of paclitaxel treatment on the MAP-kinase activity of synchronized PC-14 cells was examined. Two bands with molecular masses of 42 and 44 kDa were detected by an "intra-gel" MAP-kinase assay using myelin basic protein as the substrate. Paclitaxel treatment inhibited the MAP-kinase activity of PC-14 cells and inhibition was maximal at the G2/M phase of the cell cycle. Similar, concentration-dependent inhibition by paclitaxel of cellular MAP kinase of human synchronized small-cell lung carcinoma, H69, was observed. No inhibition of the MAP kinase of the paclitaxel-resistant sub-line H69/Txl by paclitaxel was observed, suggesting that some change of the MAP-kinase cascade had occurred in these cells. No direct inhibition of MAP-kinase activity by paclitaxel was observed in the cell-free assay (in vitro), suggesting that paclitaxel did not inhibit MAP kinase directly. Since it has been speculated that p34cdc2 kinase is also a kinase that phosphorylates MAP2, the effect of paclitaxel treatment on the p34cdc2-kinase activity of synchronized PC-14 and PC-9 cells was examined. Paclitaxel inhibited p34cdc2-kinase activation at the G2/M phase. These results suggest that paclitaxel inhibited MAP kinase and p34cdc2 kinase in vivo indirectly. These actions of paclitaxel may be responsible for the increased affinity between MAP2 and tubulins that it induces.

Amino Acid Sequence↗

PubMed1995

Gamma-glutamylcysteine synthetase gene overexpression results in increased activity of the ATP-dependent glutathione S-conjugate export pump and cisplatin resistance.

The ATP-dependent glutathione S-conjugate export pump (GS-X pump) has been suggested to play a role in the mechanism of cisplatin resistance. The purpose of this study was to determine the relationship between intracellular glutathione (GSH) levels and GS-X pump activity and whether GS-X pump overexpression results in cisplatin resistance. We transfected the human gamma-glutamylcysteine synthetase (gamma-GCS) gene into a human small-cell lung cancer cell line, SBC-3, producing SBC-3/GCS. The intracellular GSH content of SBC-3/GCS was twice that of the parental line, its GS-X pump activity was significantly enhanced and cellular cisplatin accumulation decreased. SBC-3/GCS showed higher resistance (relative resistance value of 7.4) to cisplatin than the parental line SBC-3. These data indicate that gamma-GCS gene overexpression induces cellular cisplatin resistance associated with increases in both the GSH content and GS-X pump activity, resulting in reduced cisplatin accumulation. In conclusion, GS-X pump expression is related to cellular GSH metabolism and involved in cisplatin resistance.

Adenosine Triphosphate↗

PubMed1995

Long-term human hematopoiesis in SCID-hu mice bearing transplanted fragments of adult bone and bone marrow cells.

An attempt was made to establish an SCID-hu murine model of long-term human hematopoiesis by coimplantation of a bone fragment and bone marrow (BM) cells from an adult human. The SCID-hu mice were treated with a cytokine mixture (recombinant human stem cell factor, interleukin-3, granulocyte/macrophage colony-stimulating factor, and granulocyte colony-stimulating factor) for 4 months and were then maintained for further 8 months under cytokine-free conditions. In the peripheral blood, spleen, and implanted bone fragments in the SCID-hu mice that had received both a bone fragment and BM cells, human CD59+ cells were detected 1 year after transplantation; however, they were not detectable in SCID-hu mice that had received either a bone fragment or BM cells only. Thus, implantation of both a bone fragment and BM cells appears to provide a model of long-term adult human hematopoiesis in SCID mice.

Aged↗

PubMed1995

Antitumor activities of a new indolocarbazole substance, NB-506, and establishment of NB-506-resistant cell lines, SBC-3/NB.

The novel anticancer glucosyl derivative of indolo-carbazole (NB-506), an inhibitor of DNA topoisomerase I, exhibited strong in vitro cytotoxicity against various human cancer cell lines. In order to elucidate its cytotoxic mechanisms, we established nine NB-506-resistant sublines with different resistance ratios from human small cell lung cancer cells (SBC-3/P) by stepwise and brief exposure (24 h) to NB-506. Among them, SBC-3/NB#9 was 454 times more resistant to NB-506 than the parent cell line. The SBC-3/NB#9 cells showed cross-resistance only to topoisomerase I inhibitors, such as 11,7-ethyl-10-[4-(1-piperidino)-1-piperidino] carbonyloxycamptothecia and 7-ethyl-10-hydroxy-camptothecin, and not to other anticancer drugs, such as vincristine, vinblastine, Adriamycin, etoposide, and teniposide. These results indicate that the difference on the effect of topoisomerase I was considered to be related to a resistance mechanism. The topoisomerase I activities of nuclear extracts eluted from SBC-3/NB#9 cells was only one-tenth of the parent cell activity. A Western blotting study indicated that this lower activity was due to a lower amount of DNA topoisomerase I. Furthermore, we found correlations between topoisomerase I activity and sensitivity to NB-506 in sublines with different degrees of resistance. Accumulation of 3H-labeled NB-506 by SBC-3/NB#9 cells was only one-fifth of that by the parent cells, whereas intracellular accumulation of 3H-labeled camptothecin by both cell lines did not differ. The reduction of accumulation was specific to NB-506, and this result may explain why the resistance ratio for NB-506 was higher than those for 11,7-ethyl-10-[4-(1-piperidino)-1-piperidino] carbonyloxycamptothecin and 7-ethyl-10-hydroxy-camptothecin.

Antineoplastic Agents↗

PubMed1995

Correlation of therapeutic outcome in non-small cell lung cancer and DNA damage assayed by polymerase chain reaction in leukocytes damaged in vitro.

A pilot study was conducted in patients with advanced non-small cell lung cancer to examine whether the gene-specific damage in mononuclear cells (MNCs) incubated with cisplatin in vitro correlates with chemotherapeutic outcome in cisplatin-based chemotherapy. Twenty-one patients received cisplatin-based chemotherapy, consisting of cisplatin (80 mg/m2 i.v. on day 1), vindesine (3 mg/m2 i.v. on days 1 and 8), with or without mitomycin (8 mg/m2 i.v. on day 1). MNCs from peripheral blood were obtained from each patient before chemotherapy. The cells were incubated with cisplatin for 3 h in vitro and the 2.7-kb fragment of the hypoxanthine phosphoribosyltransferase gene was amplified by PCR for quantitation of DNA damage. There was a 4-fold interpatient variation in DNA damage in MNCs. Seven of 21 patients had a partial response to chemotherapy. When the dose of cisplatin required to reduce amplification of the hypoxanthine phosphoribosyltransferase sequence by 63% (D63 value) of MNCs was compared in each patient (defined by a Poisson distribution as the dose that produced an average of one lesion per single strand of the 2.7-kb fragment), the mean D63 value in patients showing a partial response (n = 7; 52 +/- 11 micrograms/ml) was significantly lower than that in patients showing no change (n = 10; 81 +/- 20 micrograms/ml; P = 0.0045) and in patients with disease progression (n = 4; 115 +/- 34 micrograms/ml; P = 0.0012). The mean D63 in patients with no change was also significantly lower than that in the patients with disease progression (P = 0.0386). Seven (70%) of 10 patients with a D63 value < 70 micrograms/ml were responders. No relationship was observed between the D63 values and hematological and nonhematological toxicities. It is suggested that DNA damage in MNCs incubated by cisplatin treatment in vitro in responders was greater than that in nonresponders. Gene-specific damage in MNCs from peripheral blood incubated with cisplatin in vitro assayed by PCR may predict the chemotherapeutic response in cisplatin-based chemotherapy for non-small cell lung cancer.

Adult↗

PubMed1995

In vitro cytotoxicity of a novel antitumor antibiotic, spicamycin derivative, in human lung cancer cell lines.

Spicamycin (SPM), produced by Streptomyces alanosinicus, induces potent differentiation in a human leukemia cell line, HL60. One of the derivatives of SPM (SPM-D), KRN5500, has a wide range of antitumor activity against human cancer cell lines. We examined the cytotoxicity of SPM-D in small and non-small cell lung cancer cell lines using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and colony assays. SPM-D was active against a wide range of lung cancer cell lines. All three cisplatin (CDDP)-resistant cell lines established in our laboratory (PC-9/CDDP, PC-14/CDDP, and H69/CDDP) showed collateral sensitivity to SPM-D with relative resistance values of 0.43, 0.34, and 0.32, respectively. Intracellular SPM-D in PC-14/CDDP was 35% higher than that for PC-14 suggesting that intracellular accumulation can explain the collateral sensitivity to SPM-D at least in PC-14/CDDP. On the other hand, in PC-9/CDDP cells, no increase of intracellular SPM-D accumulation was observed, but the conversion ratio of a metabolite (the amino nucleoside moiety of spicamycin binding with glycine, SAN-G) from SPM-D evaluated by TLC was higher as compared with that of parental PC-9 cells (45.5% versus 37%; PC-9/CDDP versus PC-9). The increased intracellular metabolism of SPM-D could explain the mechanism of collateral sensitivity in PC-9/CDDP cisplatin-resistant cell lines. To elucidate the determinant of the SPM-D-induced cytotoxicity, we established SPM-D-resistant cell lines, PC-9/SPM-D, PC-14/SPM-D, and H69/SPM-D, by exposing cells to stepwise increases in SPM-D concentration. The relative resistances of these sublines were more than 5000, 46.6, and 37.8 times those of the parental cell lines, respectively. The intracellular concentration of the active metabolite, SAN-G, was found to be decreased in the SPM-D-resistant sublines. This result indicates that the intracellular metabolism of SPM-D to SAN-G is one of the determinants of cellular sensitivity to SPM-D in these SPM-D-resistant cell lines. In conclusion, both drug accumulation and metabolism may contribute to the sensitivity/resistance to SPM-D and both may merit investigation.

Adenocarcinoma↗

PubMed1995

In vivo screening models of cisplatin-resistant human lung cancer cell lines using SCID mice.

In vivo screening models of a cisplatin (CDDP)-resistant human small-cell lung cancer cell (SCLC) line, H69/CDDP, and a non-small-cell lung cancer cell (NSCLC) line, PC-14/CDDP, were evaluated. The transplantability of the tumor xenografts to SCID mice was more than 90%. Tumor xenografts of H69/CDDP and PC-14/CDDP showed CDDP resistance during in vivo treatment. The novel anticancer agent 254-S showed only a partial effect on the growth of H69/CDDP and PC-14/CDDP while ormaplatin showed no cross resistance to CDDP. The in vivo results correlated well with the results of the in vitro MTT assay. In this in vivo sensitivity test, H69/CDDP and PC-14/CDDP were more sensitive to ormaplatin than its parental cell lines. In vivo sensitivity testing using SCID mice bearing transplanted CDDP-resistant tumors was shown to be useful for evaluating the effects of new anti-cancer drugs, especially those that might overcome CDDP resistance.

Animals↗

PubMed1995

Induction of apoptosis in cultured retinoblastoma cells by the protein phosphatase inhibitor, okadaic acid.

The induction of apoptosis in cultured retinoblastoma cells by diverse drugs was examined by analyzing DNA fragmentation, a hallmark of apoptosis. First, the ability of six retinoblastoma cell lines to undergo apoptosis was surveyed using etoposide (30 micrograms/ml, 20 h exposure). The NCC-RbC-60 cell line, established in this laboratory showed DNA fragmentation clearly, whereas the other cell lines tested, including the representative retinoblastoma cell line, Y-79, did not show distinct DNA fragmentation. Biochemical modulators, such as A23187, forskolin, retinoic acid, phorbol 12-myristate 13-acetate and okadaic acid, were examined to ascertain whether they could induce apoptosis in NCC-RbC-60 and Y-79 cells after exposure for 20 h. Only okadaic acid induced DNA fragmentation in all the retinoblastoma cell lines tested and it induced DNA fragmentation in Y-79 cells in a time- and concentration-dependent manner. Flow-cytometric analysis and microscopic examination revealed that Y-79 cells treated with okadaic acid for 24-48 h accumulated at the G2/M, especially M, phases, before undergoing DNA fragmentation. Other mitotic poisons, nocodazole, colcemid and taxol, also induced apoptosis in Y-79 cells. In the K1034 cell line, established from non-malignant retinal pigmented epithelium, okadaic acid failed to induce both G2/M arrest and DNA fragmentation. These findings suggest that okadaic-acid-induced apoptosis occurs as a result of metaphase arrest.

Apoptosis↗