Sequence dependence of the hyperthermic potentiation of carboplatin-induced cytotoxicity and intracellular platinum accumulation in HeLa cells

We have examined the enhancement of cytotoxic effects of cis-diammine-1,1-cyclobutane dicarboxylate platinum(II) (carboplatin) by hyperthermia in HeLa cells using different regimes of timing and sequence. The results were compared with those obtained with cis-diamminedichloroplatinum(II) (cisplatin). We found that cisplatin simultaneously combined with heat was the most cytotoxic toward HeLa cells of the various timing and sequencing conditions studied. On the other hand, for carboplatin, drug treatment immediately following or during heat exposure showed the greatest effect. Intracellular platinum concentration in HeLa cells treated with heat before carboplatin showed a 2.75-fold increase over that in cells treated with the drug alone. The ratios for carboplatin given before, or during heating, were 0.67 and 1.42 respectively. Simultaneous exposure of cells to cisplatin and heat led to a 1.64-fold enhancement in cisplatin accumulation, compared to 0.92- and 1.24-fold increase for cells treated with cisplatin before and after heat respectively. Although each drug exposure prior to heat was less cytotoxic toward HeLa cells than any other heat/drug combination sequences, the platinum concentration was less than seen with each drug alone. Even though heat exposure prior to and during carboplatin showed a similar toxicity, platinum concentration in cells treated with heat prior to carboplatin was higher than that in cells treated with heat and carboplatin simultaneously. Thus, increased cytotoxicity cannot always be explained on the basis of intracellular platinum concentration. It is clear however that, differing from cisplatin, exposure of cells to heat prior to or during carboplatin administration results in the greatest cell kill.     

Tumor heterogeneity and in vitro chemosensitivity testing in ovarian cancer

OBJECTIVE: Our purpose was to study the heterogeneity of drug response in fresh human ovarian tumors to chemotherapeutic agents in an in vitro chemosensitivity assay. STUDY DESIGN: This assay evaluates total tumor cell kill by measuring the intracellular adenosine triphosphate levels of untreated controls and drug-exposed cells at various doses after culture for 6 days. The surviving fraction is calculated by dividing the treated values with the control values. One hundred tumors were tested against four single drugs (cisplatin, the active metabolite of cytoxan, 4-hydroxyperoxy-cyclophosphamide, Taxol, and carboplatin) and two drug combinations (cisplatin plus 4-hydroxyperoxy-cyclophosphamide; cisplatin plus Taxol). RESULTS: There is great variation in the degree of cell death for single drugs and drug combinations among the 100 tumors tested. CONCLUSION: More effective clinical response to chemotherapy may be achieved in patients with ovarian cancer by selecting the most active drugs for chemotherapy, on the basis of in vitro chemosensitivity test results for individual patients.     

A novel trans-platinum coordination complex possessing in vitro and in vivo antitumor activity

As part of a drug discovery program to discover more effective platinum-based anticancer drugs, a series of platinum complexes of trans coordination geometry centered on trans-ammine(cyclohexylaminedichlorodihydroxo)platinum(IV) (JM335) has been evaluated in vitro against a panel of cisplatin-sensitive and cisplatin-resistant human tumor cell lines (predominantly ovarian). In vitro, against 5 human ovarian carcinoma cell lines, JM335 was comparably cytotoxic to cisplatin itself and over 50-fold more potent than transplatin (mean 50% inhibitory concentrations: JM335, 3.1 microM; cisplatin, 4.1 microM; transplatin, 162 microM). With the use of seven pairs of human tumor cell lines (parent and subline with acquired resistance to cisplatin and encompassing all of the known major mechanisms of resistance to cisplatin) JM335 exhibited a different cross-resistance pattern to that of its cis isomer (JM149). JM335 showed non-cross-resistance in six of the seven resistant lines, cross-resistance in the A2780cisR line possibly being associated with high levels of glutathione. Preliminary intracellular DNA binding studies showed that in contrast to transplatin, JM335 was efficient at forming DNA-DNA interstrand cross-links. In vivo, JM335 produced growth delays in excess of 15 days against 4 of 6 human ovarian carcinoma xenografts and was unique among the complexes studied in retaining some efficacy against a cisplatin-resistant subline of the murine ADJ/PC6 plasmacytoma. JM335 is the first trans-platinum complex to demonstrate marked antitumor efficacy against both murine and human s.c. tumor models and represents a significant structural lead to complexes capable of circumventing cross-resistance to cisplatin.     

Genetic diagnosis of cardiovascular diseases and the therapeutic application

We have examined the effect of neutralizing TGF-beta antibodies on cisplatin-mediated cytotoxicity against MDA-231 human breast tumor cell spheroids. These tridimensional in vitro systems have been shown to recapitulate the drug sensitivity pattern of tumor cells in vivo. MDA-231 tumor cell spheroids exhibit higher protein levels of the cyclin-dependent kinase (Cdk) inhibitors p21 and p27 and >10-fold lower Cdk2 activity compared to adherent cell monolayers, as well as pRb hypophosphorylation, a predominant G1 population, and a cisplatin 1-h IC50 of approximately 100 microM. Treatment of MDA-231 cells in monolayer with cisplatin for 1 h, subsequently grown as spheroids, increased steady-state TGF-beta1 mRNA levels, secretion of active TGF-beta, cellular Cdk2 activity, pRb phosphorylation, and p21 protein levels, while downregulating p27. Accumulation of cells in G2M and progression into S were noted 48 h after treatment with 100 microM cisplatin. We tested whether drug-induced upregulation of TGF-beta1 and p21, perhaps by preventing cell cycle progression, were protective mechanisms against drug-mediated toxicity by using neutralizing anti-TGF-beta antibodies. Anti-TGF-beta antibodies diminished the induction of p21, enhanced the activation of Cdk2, and facilitated progression into S and G2M following cisplatin treatment. This resulted in a >twofold enhancement of drug-induced DNA fragmentation and a shift in the cisplatin 1-h IC50 from 100 to <10 microM. These data suggest that tumor cell TGF-beta1 may protect from DNA damage and that postchemotherapy administration of TGF-beta inhibitors may facilitate progression beyond G1/S, potentially increasing the efficacy of cytotoxic chemotherapy.     

Synergistic action of quercetin and genistein in human ovarian carcinoma cells

Ovarian carcinoma is the fourth most common cause of cancer death in women and there has been a steady increase in the age-adjusted cancer death rates in the past 25 years in the US. However, patients who become cisplatin resistant respond poorly to available cytotoxic agents; therefore, discovering novel targets for ovarian carcinoma is vital. Quercetin, an anticancer agent, arrests the cell cycle at G1 and S phase boundary. Genistein, a plant flavonoid, attacks the cell cycle at G2 and/or early M phases in most carcinoma cells. Quercetin and genistein block the phosphatidylinositol conversion to IP3 signal transduction pathway mainly by inhibiting 1-phosphatidylinositol 4-kinase (PI kinase, EC 2.7.1.67) and 1-phosphatidylinositol 4-phosphate 5-kinase (PIP kinase, EC 2.7.1.68), respectively. Because each drug attacks a different phase of the cell cycle and reduces IP3 concentration by attacking different signal transduction enzymes, we tested the hypothesis that the two drugs might be synergistic in human carcinoma cells. In human ovarian carcinoma OVCAR-5 cells in growth inhibition assay, the IC50S for quercetin and genistein were (mean +/- SE) 66 +/- 3.0 and 32 +/- 2.5 microM; in clonogenic assays they were 15 +/- 1.2 and 5 +/- 0.5 microM, respectively. When quercetin was added to the cultures of OVCAR-5 cells followed 8 h later by genistein, synergism was observed in growth inhibition and clonogenic assays. The synergistic action of quercetin and genistein may be of interest in clinical treatment of human ovarian carcinoma.     

Spectrophotometric assay using streptavidin-alkaline phosphatase conjugate for studies on the proteolysis of polymer bead-bound peptides

Over the past 20 years ovarian cancer has provided a vivid illustration of the successes, failures and challenges for the medical oncologist. During that time the results of treatment have substantially improved; in the West of Scotland for example, for women aged under 55, 3-year survival rates have increased from 36% to 50%. One reason for this was probably the introduction of effective agents such as cisplatin in the mid-1970s and then carboplatin in the mid-1980s. The recent introduction of taxoids promises further improvement in the future. It is important to remember, however, that the best results will be obtained by an optimal organization for the delivery of treatment; national audit studies have shown that factors such as management in integrated clinics can have a major impact on outcome. Nevertheless, the majority of patients still die from the disease; when relapse occurs, clinical drug resistance eventually proves fatal despite further treatment. What are the fundamental mechanisms by which this resistance develops, and what means are available to attempt its circumvention? Factors involved could be described as pharmacological or cellular. Pharmacological resistance might best be addressed by increasing the doses of the drugs used, particularly, cis- or carboplatin. Three years ago we published the results of a randomized trial of 2 doses of cisplatin in 191 patients. At that stage a highly significant median survival advantage for the higher dose (100 mg/m2) of cisplatin was seen. However, a recent updated analysis with a median follow-up of 4 1/2 years shows a reduction in the survival benefit, with 4-year overall survival rates for high- and low-dose cisplatin of 32.4% and 26.6%, respectively. This suggests that a population of drug resistant ovarian cancer cells will eventually emerge despite the use of initial higher doses of cisplatin. A more dose-intensive approach is being pursued with carboplatin, and it seems clear that dose-increments over standard therapy of at least 4-fold will be necessary, to justify further randomized trials. Meanwhile, the alternative approach to delivering high drug concentrations, i.e. intraperitoneal (i.p.) chemotherapy, clearly merits further study, particularly in the light of a recently reported study in patients with minimal disease, which showed a significant survival benefit for i.p. cisplatin treatment. Cellular factors will probably prove to be crucial; studies using various cell lines suggest that multiple mechanisms are likely to be involved and these will need to be examined in relevant clinical material. After DNA damage induced by a range of cytotoxic agents has taken place in ovarian cancer cells, the key to sensitivity/resistance may well be the ability of these cells to engage the process of apoptosis. Several genes are involved in control of this process; these include the p53 gene, mutations of which have been linked to cisplatin resistance in our laboratory studies, as well as in clinical trials with carboplatin. We have also demonstrated an association in ovarian cancer cell lines between cisplatin resistance and microsatellite instability (indicative of defective mismatch repair) and the clinical relevance of this link is also being pursued. A thorough understanding of underlying mechanisms may lead to the rational development of therapeutic means for circumventing cisplatin-resistance in ovarian cancer; the emergence of new classes of drug such as taxoids as topoisomerase I inhibitors offers further promise of improvement in outcome in the next few years.     

Stability and cellular studies of [rac-1,2-bis(4-fluorophenyl)-ethylenediamine][cyclobutane-1,1- dicarboxylato]platinum(II), a novel, highly active carboplatin derivative

The synthesis of the diastereomeric [1,2-bis(4-fluorophenyl)ethylenediamine][cyclobutane-1, 1-dicarboxylato]platinum(II) complexes, rac- and meso-4F-Pt(CBDC), the evaluation of their structures, their tumor-inhibiting properties and their stability in physiological environment are described (reference complexes: the dichloro- and sulfatoplatinum(II) analogues, carboplatin and cisplatin). The most interesting diastereomer, rac-4F-Pt(CBDC), equals cisplatin and surpasses carboplatin in its effect on human breast cancer cell lines (MCF-7 and MDA-MB-231). Rac-4F-Pt(CBDC) is largely insensitive against attack of nucleophiles e.g. Cl-, a prerequisite for sufficient stability in vivo and for fewer side effects. In accordance with this, in vitro studies on the binding of rac-4F-Pt(CBDC) to albumin, the main plasma protein, show that the free, non-protein-bound fraction is relatively high, coming close to that of carboplatin. These properties are of importance for the transferability of the promising effects found in the cell culture experiments to in vivo conditions. The distinctly better anti-breast cancer activity of rac-4F-Pt(CBDC) than of carboplatin has been attributed to its ability to accumulate in the tumor cells. The human ovarian cancer cell line NIH-OVCAR-3 is also strongly inhibited by rac-4F-Pt(CBDC).     

Acquisition of chemoresistance in a human ovarian carcinoma cell is linked to a defect in cell cycle control

Chemoresistance is a major concern in cancer erradication; it involves various mechanisms, including defects in the apoptosis program induced by anticancer drugs. In order to further explore the mechanisms underlying the development of chemoresistance in ovarian carcinoma after cisplatin treatment, we established an in vitro model, mimicking a clinical protocol of administration of cisplatin. Therefore, IGROV1 ovarian carcinoma cells were exposed for 2 hr to the drug and allowed to recover for several weeks; this way of exposure was reiterated with escalating doses. We followed changes in cytotoxicity of the drug, cell cycle kinetics and long-term survival of cells after cisplatin treatment, and found that resistance to cisplatin was not associated with altered apoptosis pathway, since both cisplatin sensitive and resistant cells underwent apoptosis in a similar way. Acquisition of resistance to cisplatin was associated with the ability of the treated cells to progress through the cell cycle beyond the G1/S checkpoint; although most cells died by apoptosis, a few surviving cells proliferated and recolonized the cultures. Compared to sensitive cells, the chemoresistant variants were able to override the G1/S checkpoint whatever the dose, and the recurrent cells recolonized the cultures much faster. Analysis of alterations in gene expression suggests that the defect in cell cycle regulation could take place at the level of the cdk inhibitor p21(CIP1/WAF1).     

Chemotherapy in advanced ovarian cancer: four systematic meta-analyses of individual patient data from 37 randomized trials. Advanced Ovarian Cancer Trialists' Group

The purpose of this systematic study was to provide an up to date and reliable quantitative summary of the relative benefits of various types of chemotherapy (non-platinum vs platinum, single-agent vs combination and carboplatin vs cisplatin) in the treatment of advanced ovarian cancer. Also, to investigate whether well-defined patient subgroups benefit more or less from cisplatin- or carboplatin-based therapy. Meta-analyses were based on updated individual patient data from all available randomized controlled trials (published and unpublished), including 37 trials, 5667 patients and 4664 deaths. The results suggest that platinum-based chemotherapy is better than non-platinum therapy, show a trend in favour of platinum combinations over single-agent platinum, and suggest that cisplatin and carboplatin are equally effective. There is no good evidence that cisplatin is more or less effective than carboplatin in any particular subgroup of patients.     

Decreased drug accumulation and increased tolerance to DNA damage in tumor cells with a low level of cisplatin resistance

In an attempt to examine the cellular changes associated with cisplatin resistance, we selected a cisplatin-resistant (A43 1/Pt) human cervix squamous cell carcinoma cell line following continuous in vitro drug exposure. The resistant subline was characterized by a 2.5-fold degree of resistance. In particular, we investigated the expression of cellular defence systems and other cellular factors probably involved in dealing with cisplatin-induced DNA damage. Resistant cells exhibited decreased platinum accumulation and reduced levels of DNA-bound platinum and interstrand cross-link frequency after short-term drug exposure. Analysis of the effect of cisplatin on cell cycle progression revealed a cisplatin-induced G2M arrest in sensitive and resistant cells. Interestingly, a slowdown in S-phase transit was found in A431/Pt cells. A comparison of the ability of sensitive and resistant cells to repair drug-induced DNA damage suggested that resistant cells were able to tolerate higher levels of cisplatin-induced DNA damage than their parental counterparts. Analysis of the expression of proteins involved in DNA mismatch repair showed a decreased level of MSH2 in resistant cells. Since MSH2 seems to be involved in recognition of drug-induced DNA damage, this change may account for the increased tolerance to DNA damage observed in the resistant subline. In conclusion, the involvement of accumulation defects and the increased tolerance to cisplatin-induced DNA damage in these cisplatin-resistant cells support the notion that multiple changes contribute to confer a low level of cisplatin resistance.     

Endometrial cancer cell lines are sensitive to paclitaxel

We have previously reported a 24 fold difference in the cisplatin sensitivity and a 12 fold difference in carboplatin sensitivity of endometrial carcinoma cell lines. In this study as evaluate paclitaxel sensitivity of the same cell lines. We tested nine endometrial cancer cell lines with the 96-well plate clonogenic assay using limiting dilution. The chemosensitivity was expressed as IC50 value, 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. The IC50 values for paclitaxel were 0.49 - 2.3 nM showing only a 4.7 fold difference between various cell lines. No correlation could be demonstrated between in vitro paclitaxel and platinum analog sensitivities of endometrial adenocarcinoma cell lines. The variance in paclitaxel sensitivity of different cell lines was little. Our results suggest that endometrial adenocarcinoma cell lines tested with the same methods. The clinical efficacy of paclitaxel in the treatment of endometrial cancer should further be evaluated in clinical trials.     

cis-Amminedichloro(2-methylpyridine) platinum(II) (AMD473), a novel sterically hindered platinum complex: in vivo activity, toxicology, and pharmacokinetics in mice

A novel sterically hindered platinum complex, AMD473 [cis-amminedichloro(2-methylpyridine) platinum(II)], designed primarily to be less susceptible to inactivation by thiols, has shown in vitro activity against several ovarian carcinoma cell lines. Notably, AMD473 has shown activity in vitro in human carcinoma cells that have acquired cisplatin resistance due to reduced drug transport (41M/41McisR) or enhanced DNA repair/increased tolerance of platinum-DNA adducts (CH1/CH1cisR). In this study, we show that AMD473, at its maximum tolerated dose of 35-40 mg/kg i.p. administration, produced marked in vivo antitumor activity against a variety of murine (ADJ/PC6 plasmacytoma, L1210 leukemia) and human ovarian carcinoma xenograft models, including several possessing acquired resistance to cisplatin [ADJ/PC6cisR, L1210cisR, CH1cisR, and HX110 (carboplatin-resistant)]. In the ADJ/PC6 model, an increased therapeutic index was noted following oral as opposed to i. p. administration. In a head-to-head comparison using CH1cisR xenografts and equitoxic doses (q7dx4 schedule), comparative growth delays were as follows: AMD473, 34 days; cisplatin, 10.4 days; carboplatin, 6.4 days; and JM216 (p.o. administration), 3.5 days (in a previous experiment, the trans-platinum complex JM335 induced a growth delay of 5.4 days against this model). In this model, oral activity was also noted with a growth delay of 34 days at 400 mg/kg every 7 days (total of four doses). In addition, AMD473 showed promising activity against CH1 xenografts that had regrown following initial treatment with cisplatin (additional growth delay of 30 days over that observed for retreatment with cisplatin). Across the whole panel of cisplatin-sensitive to cisplatin-resistant human ovarian carcinoma xenografts, AMD473 showed improved or at least comparable activity to that observed for an equitoxic dose (4 mg/kg) and schedule of cisplatin. Platinum pharmacokinetics showed that following i.v. administration of 20 mg/kg AMD473 in saline to Balb/c- mice bearing murine plasmacytoma (ADJ/PC6), a biexponential decay was observed in the plasma with a rapid distribution t1/2alpha of 24 min followed by a slow elimination t1/2beta of 44 h. Platinum accumulated in various organs with platinum tissue to plasma area under the curve ratios of 8.6 for liver and kidney, 5.7 for spleen, 3.7 for heart, 5.2 for lung, and 5 for tumor. The plasma and tissue concentration time curve following i.p. administration was similar to that observed following i.v. administration, with a bioavailability of 89%. When AMD473 was given p.o., the platinum absorption was rapid (K01 of 30 min) and the bioavailability was 40%. A less than proportional increase in area under the curve and Cmax was noted in tissue, plasma, and plasma ultrafiltrate following increasing oral doses of AMD473. In vitro, with AMD473, the rate of binding to different plasma proteins was approximately half of that of cisplatin. Following administration of 45 mg/kg i.p. in oil, 33% of the administered platinum was eliminated in the urine after 24 h, and 40% was eliminated after 72 h. Fecal recovery represented 13% of the administered dose after 3 days. Similar results were observed following oral and i.v. administration of 20 mg/kg, but significantly more was excreted in the feces (over 50% of the administered dose) following oral administration of 400 mg/kg, showing that absorption might be a limiting factor by this route of administration. The dose-limiting toxicity for AMD473 in mice was myelosuppression, and no renal toxicity was observed. The promising antitumor activity of AMD473, together with its lack of nephrotoxicity and favorable pharmacokinetic profile, suggests that AMD473 is a good candidate for clinical development. AMD473 is entering Phase I clinical trials under the auspices of the United Kingdom Cancer Research Campaign in 1997.     

In vitro determination of cytotoxic drug response in ovarian carcinoma using the fluorometric microculture cytotoxicity assay (FMCA)

The fluorometric microculture cytotoxicity assay (FMCA), a short-term in vitro assay based on the concept of total tumor cell kill, was used for testing the cytotoxic drug sensitivity of tumor cells from patients with ovarian carcinoma. A total of 125 fresh specimens was obtained, 98 (78%) of which were analyzed successfully. Data from 45 patients were available for clinical correlations. The FMCA appeared to yield clinically relevant cytotoxic drug sensitivity data for ovarian carcinoma as indicated by a comparison with tumor samples obtained from patients with non-Hodgkin's lymphoma or kidney carcinoma. Considering the most active single agent in vitro actually given in vivo, and using the median drug activity among all ovarian carcinoma samples as a cut-off, the sensitivity of the assay and its specificity were 75 and 52%, respectively. Cross-resistance in vitro was frequently observed between standard drugs but not between standard drugs and Taxol. Ten percent of the specimens showed an extreme resistance for at least 4 of 6 of the drugs investigated.     

Cell biological markers of drug resistance in ovarian carcinoma

The aim of the study is to review the mechanisms of resistance to four classes of drugs that are widely used in ovarian carcinoma: platinum (cisplatin/carboplatin) compounds, classical alkylating agents (cyclophosphamide/melphalan), natural drugs (doxorubicin), and "new drugs" (taxol and taxotere). Both platinum and classical alkylating agents mediate their cytotoxicity by the formation of drug-DNA adducts, resulting in DNA damage. Therefore, drug resistance mechanisms are (in part) comparable. In ovarian carcinoma cell lines increased repair of DNA damage and increased detoxification by binding of drugs to glutathione, possibly catalyzed by glutathione S-transferases, have been identified as the most prominent resistance mechanisms to these drugs. Studies on the role of DNA repair mechanisms and glutathione in human ovarian carcinoma are hampered by the complexity of enzyme systems involved in DNA repair and intratumor heterogeneity for glutathione. Resistance to doxorubicin appears to be mediated by enhanced efflux from the cell by increased expression of membrane glycoproteins acting as a drug efflux pump, such as P-glycoprotein. Resistance to doxorubicin can also be due to quantitative and/or qualitative changes in the nuclear target of doxorubicin, topisomerase (Topo) II. Finally, resistance to taxol may be mediated by enhanced expression of P-glycoprotein, while presumed other mechanisms such as alterations in tubulin structure, the cellular "target" of taxol, and changes in polymerization of tubulin are still largely unresolved. Several ways to modulate the reviewed resistance mechanisms are also described. In conclusion, this review shows that many cell biological factors may be involved in drug resistance. The relevance of the identification of most of these factors in ovarian carcinoma patients however remains to be established.     

Enhancement of cisplatin-induced cytotoxicity by 7-hydroxystaurosporine (UCN-01), a new G2-checkpoint inhibitor

DNA-damaging agents arrest cell cycle progression at either G1 or G2. A variety of agents such as caffeine have been shown to abrogate the DNA damage-dependent G2 checkpoint and enhance cytotoxicity. Unfortunately, this strategy has not enhanced therapeutic activity because adequate concentrations of these modulators are not tolerated in vivo. Here, using Chinese hamster ovary cell lines, we show that the potent protein kinase inhibitor 7-hydroxy-staurosporine (UCN-01) abrogates the G2 arrest induced by the DNA-damaging agent cisplatin. UCN-01 not only was effective at inducing mitosis when added to G2-arrested cells but also prevented cells from arresting in G2 when added to S-phase cells. Furthermore, UCN-01 did not cause premature mitosis of S-phase cells; rather, the cells progressed to G2 before undergoing mitosis. These effects were observed at noncytotoxic concentrations of UCN-01 that alone had no effect on cell cycle passage. Furthermore, the same concentrations of UCN-01 that resulted in abrogation of the cisplatin-induced G2 arrest also enhanced cisplatin-induced cytotoxicity, as determined by a colony formation assay. UCN-01 enhanced cisplatin cytotoxicity up to 60-fold and reduced by 3-fold the concentration of cisplatin required to kill 90% of the cells. The concentrations of UCN-01 required for this enhancement have been shown to be well tolerated in animal models, suggesting that this combination may represent an effective strategy for enhancing cisplatin-based chemotherapeutic regimens.     

Immediate eradication of Helicobacter pylori in patients with previously documented peptic ulcer disease: clinical and economic effects

The introduction of adenovirus 5 E1A into the SKOV3ip1 ovarian cancer cell line was shown previously to suppress HER2/neu expression and reduce the malignant potential of these cells (Yu et al., Cancer Res., 53: 891-898, 1993). In this report, we show that reduction of p185 in cells stably expressing E1A protein was coincident with increased sensitivity to cytotoxic agents. The LD50 of cisplatin was reduced 6-fold, and the LD50 of paclitaxel and doxorubicin was reduced 10-fold in E1A-expressing cells compared with control cells. The growth of SKOV3ip1 and control cells was unchanged in the presence of 150 ng/ml of tumor necrosis factor-alpha, whereas the growth of E1A-expressing cells was reduced by 30 to 40%. When we used a physiologically obtainable concentration of paclitaxel (0.5 microM), DNA laddering consistent with apoptotic cell death was seen after a 24-h exposure in the E1A-expressing cells, whereas laddering and DNA fragmentation were only detected in DNA from control cells after longer exposure (48 h) at a 20-fold higher concentration of paclitaxel. The SKOV3ip1 cells do not express p53 protein; hence, the induction of apoptosis by paclitaxel is through a p53-independent pathway. Despite their diverse mechanisms of action, the cytotoxic effects of cisplatin, doxorubicin, paclitaxel, and tumor necrosis factor-alpha were enhanced by the expression of E1A proteins in the SKOV3ip1 ovarian cancer cells. This suggests that these agents share a common final pathway of cell killing, which may represent a potential therapeutic target in resistant ovarian cancers.     

Influence of genetic predisposition to thrombosis on natural history of acute promyelocytic leukaemia. MRC Adult Leukaemia Working Party

Clinical evidence and the majority of experimental data suggest a cross-resistance between cisplatin and radiation in ovarian cancer. The authors are not aware of any report of a human ovarian cancer cell line for which the dose-response relationships to both radiation and chemotherapy including paclitaxel and cisplatin have been evaluated with the same methodology. The present study investigated the radiosensitivity profiles of four established human ovarian cancer cell lines in vitro using the ATP assay, which measures total cell kill. The CAOV-3 cell line showed the highest degree of radiosensitivity of the four cell lines. SKOV-3 cells were the most resistant. The BG-1 cell line, previously shown to be highly resistant to cisplatin but sensitive to paclitaxel, was distinctly sensitive to radiation. This was particularly true for the lower dosages (2-6 Gy). The four cell lines tested are a good representation of cell lines with different radiosensitivities. The different response patterns to cytotoxic agents and radiation, make the BG-1 cell line in particular an interesting candidate for future studies on mechanisms of resistance and combination effects between radiation and chemotherapy.     

Direct delivery of platinum-based antineoplastics to the central nervous system: a toxicity and ultrastructural study

Platinum drugs are playing an increasingly major role in cancer treatment, but systemic administration of these agents has resulted in significant toxicity. To examine the effects of cisplatin and two newer agents, iproplatin and carboplatin, we injected the agents directly into the cerebrospinal fluid of rats and found that neurotoxic reactions resulted from doses of cisplatin (10 nmol) much lower than those of iproplatin (40 nmol) or carboplatin (80 nmol). Moreover, central nervous system tissue appeared to be less adversely affected by direct exposure to carboplatin since chronic toxicity was not observed in any of the animals receiving carboplatin until a lethal dose was reached. Furthermore, only the animals receiving cisplatin showed histologic damage in their spinal cords, and ultrastructural studies confirmed that while significant abnormalities were observed in the spinal cords of rats receiving 40 nmol cisplatin, no architectural changes were detected in the spinal cords of animals receiving 240 nmol carboplatin. We conclude that platinum drugs can be delivered intrathecally to achieve a much greater concentration of active drug than can be achieved by intravenous administration and that carboplatin appears to be the most suitable platinum-based drug for use in systems delivering drugs directly to the brain and spinal cord.     

Knowledge and attitudes among California dental care providers regarding child abuse and neglect

Combination chemotherapy with paclitaxel plus a platinum compound (carboplatin or cisplatin) is the current regimen of choice for the treatment of advanced epithelial ovarian cancer. The two most widely used combinations are paclitaxel (135 mg/m2, 24-hour infusion) plus cisplatin (75 mg/m2) or paclitaxel (175 mg/m2, 3-hour infusion) plus carboplating dosed to an area under curve of 7.5. Randomized trials are in progress comparing these two regimens. Numerous other clinical issues remain regarding how to maximize the effectiveness of this therapy, including dose and schedule, duration of treatment, route of administration, and incorporation of other agents with novel mechanisms of cytotoxicity. New agents currently undergoing evolution as part of novel induction regimens have been shown to have significant activity in recurrent ovarian cancer and include topotecan, gemcitabine, oral etoposide, and encapsulated doxorubicin.     

A sensitive postcolumn derivatization/UV detection system for HPLC determination of antitumor divalent and quadrivalent platinum complexes

A sensitive postcolumn derivatization/UV detection system has been developed for HPLC analysis of antitumor divalent and quadrivalent platinum complexes. It is based on the derivatization of platinum complexes by reaction with sodium bisulfite to corresponding product(s) which has enhanced absorptivity at 280-300 nm. Platinum complexes examined in this study were cisplatin, carboplatin and oxaliplatin (divalent platinum complexes) and oxoplatin and tetraplatin (quadrivalent ones). The proposed detection system was sensitive to all these complexes. Under the detection conditions optimized for individual complexes, the HPLC gave linear relationships between the complex concentration and the peak height. Detection limits at 290 nm with 100 microliters injection were 20 nM for cisplatin, 40 nM for oxoplatin, 60 nM for carboplatin and tetraplatin and 100 nM for oxaliplatin (S/N = 3 at 0.005 AUFS). The proposed system was successfully applied for the determination of cisplatin and oxoplatin in plasma and urine. Pharmacokinetic behavior of oxoplatin and its reduced product cisplatin following a single intravenous injection of oxoplatin in rabbits has been discussed.