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It is generally accepted that selenium (Se) plays an important role in maintaining equilibrium of a healthy organism. It also participates in processes related to carcinogenesis such as inhibition of tumor formation and regression. Scientific data accumulated so far using experimental animal models and from clinical studies devoted to investigating the effects of Se confirm strong relationship or correlation between Se supplementation and tumor frequency of prostate, lungs, liver and colon. However, details of mechanisms of action of Se in modulation of carcinogenesis and cancer prevention are not yet fully elucidated. It is not clear yet whether Se deficiency itself is a cancer risk factor or whether it helps an already present cancer to progress. Additionally, the effects of other factors such as age, gender, life style, geographic location, comorbidities and use of drugs, are not clear. Despite the fact that some positive results were obtained with Se supplementation, it is necessary to verify these findings in more controlled experimental models including clinical studies. At the present time, data related to Se supplementation are not convincing enough as to allow general recommendation for using Se as an effective agent for chemoprevention of cancer. The goal of this minireview is to highlight present level of understanding of Se biological and prospects of its future clinical use. Information regarding Se, its effectiveness in various experimental models and in clinical tests, including combinations with other bioactive agents and anticancer drugs, is evaluated and summarized.

The extract of artichoke Cynara cardunculus L. (CCE) was investigated for its potential antigenotoxic and antioxidant effects using four experimental model systems. In the Saccharomyces cerevisiae mutagenicity/antimutagenicity assay, CCE significantly reduced the frequency of 4-nitroquinoline-N-oxide-induced revertants at the ilv1 locus and mitotic gene convertants at the trp5 locus in the diploid Saccharomyces cerevisiae tester strain D7. In the simultaneous toxicity and clastogenicity/anticlastogenicity assay, it exerted an anticlastogenic effect against N-nitroso-N′-methylurea-induced clastogenicity in the plant species Vicia sativa L. On the contrary, despite CCE not being mutagenic itself, in the preincubation Ames assay with metabolic activation, it significantly increased the mutagenic effect of 2-aminofluorene in the bacterial strain Salmonella typhimurium TA98. In the 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging assay, CCE exhibited considerable antioxidant activity. The SC50 value representing 0.0054% CCE corresponds to an antioxidant activity of 216.8 µm ascorbic acid which was used as a reference compound. Although the mechanism of CCE action still remains to be elucidated, different possible mechanisms are probably involved in the CCE antigenotoxic effects. It could be concluded that CCE is of particular interest as a suitable candidate for an effective chemopreventive agent. Copyright © 2007 John Wiley & Sons, Ltd.

Using four experimental model systems, it was demonstrated that glucomannan (GM) isolated from the cell wall of the industrial yeast Candida utilis revealed a broad range of protective activities. This effect depended on the nature and mode of action of the counteracting genotoxic compound as well as on the experimental model system used. In the Saccharomyces bioprotectivity assay, GM increased resistance towards ofloxacin-induced toxicity in the wild type and recombination repair-deficient yeast strains significantly enhancing survival of the cells. In the chromosomal aberration assay, GM exerted anticlastogenic effect against maleic hydrazide induced clastogenicity in Vicia faba L. In the DNA-topology assay, GM protected plasmid DNA from the breaks induced by Fe(2+) ions, but enhanced damage induced by bleomycin and hydrogen peroxide. In the cell-revitalization assay, it enhanced cytotoxic/cytostatic effect of teniposide applied to mouse leukemia cells. Thus, depending on the experimental model, GM acted as antimutagen, anticlastogen, DNA breaks inhibitor or inducer, and as cytotoxic/cytostatic effect enhancer. Several possible mechanisms of bioprotective action underlying the observed activities are suggested including iron chelation and free radical scavenging. The results imply that GM is a polysaccharide with marked biological activities and suggest its potential biomedical application, especially in combination with other bioactive compounds.

Nowadays naturally occurring compounds with the potential antimutagenic and anticarcinogenic effects are of great importance for their prospective use in cancer chemoprevention and treatment. The new water soluble derivative of microbial polysaccharide beta-D-glucan-carboxymethyl glucan (CMG) belongs to such a category of natural substances. CMG isolated from the cell wall of baker's yeast Saccharomyces cerevisiae is included into the class of biopolymers known as biological response modifiers (BRMs) with a broad range of activities, above all ones interfering with cancer therapy. It was demonstrated on four experimental model systems that biological and consequential medicinal importance of CMG is based on the combined application with another active compound. In the Saccharomyces cerevisiae antimutagenicity assay CMG significantly reduced ofloxacin-induced mutagenicity in the yeast strain D7. CMG exerted bioprotective (anti-toxic and antimutagenic) effect after its simultaneos application with methyl methanesulphonate on the repair-deficient strain uvs10 of the unicellular green alga Chlamydomonas reinhardtii. In the Vicia sativa simultaneous phytotoxicity and anticlastogenicity assay CMG exerted statistically significant anticlastogenic efect against maleic hydrazide-induced clastogenicity in Vicia sativa L. Only in the Salmonella/microsome assay CMG did not exert statistically significant antigenotoxic effect, despite of the fact that it reduced 9-aminoacridine-induced mutagenicity in S. typhimurium TA97, but his(+) revertants decreasing was statistically significant only at the highest CMG concentration used. The data presented unambiguously documented that even biopolysaccharides (e.g., derivatives of beta-glucan) belonging to the most abundant class of natural biopolymers may contribute to cancer prevention and therapy.

We studied the ability of five structurally related flavonoids (quercetin (QU), luteolin (LU), apigenin (AP), galangin (GA) and chrysin (ChR)) to modulate chemotherapeutic efficacy and affect chemotherapy-induced apoptosis and cell cycle arrest of murine leukemia L1210 cells. We found that QU and LU potentiated cytotoxicity of cisplatin, however the other flavonoids (AP, GA and ChR) decreased its cytotoxic effect. We proved that QU and LU positively modulated efficacy of cisplatin due to increasing of cisplatin-induced apoptotic cell death. In combination with doxorubicin all tested flavonoids decreased doxorubicin cytotoxicity and caused cells to shift from G2/M to S phase of the cell cycle. The findings that flavonoids can differentially modulate the efficacy of chemotherapeutics through interfering with chemotherapy-induced apoptosis or cell cycle arrest further improve our understanding of possible impacts of some dietary agents on chemotherapeutic intervention. © 2003

Water-soluble derivatives of the chitin–glucan (Ch–G) complex isolated from the fungal mycelium of the industrial strain of Aspergillus niger have been previously shown to possess potent antimutagenic protective activity in vivo. Their direct action on DNA has not been yet evaluated. Using carboxymethylation, sulfoethylation and subsequent ultrasonic treatment, lower molecular weight water-soluble derivatives were obtained from the crude fungal Ch–G. The biological effects of the prepared compounds were evaluated in direct interaction on plasmid DNA in vitro. Monitoring of electrophoretic mobility of different conformers of plasmid DNA implied that carboxymethyl chitin–glucan (CM-Ch–G) induced single- and double-strand breaks into supercoiled DNA in a concentration-dependent manner. On the other hand, sulfoethyl chitin–glucan (SE-Ch–G) alone did not induce any DNA breaks in plasmid DNA. However, process of DNA damaging induced by free-radical oxidation initiated with Fe 2+ was inhibited, while the process of DNA breakage induced by H 2 O 2 was increased in the presence of SE-Ch–G.

Two samples of the acid-mine water (AMW) containing heavy metals were collected in the year 1995 and 1998 in the former mining area of Banska Stiavnica-Sobov (Slovakia), and assayed for their genotoxic potential by the Ames assay and the DNA-topology assay. The content of toxic metal elements, determined by atomic absorption spectroscopy, was decreased in the sample of AMW collected in the year 1998 in comparison with that determined in the year 1995. The sample collected in 1995 was genotoxic after its application on bacterial strain Salmonella typhimurium TA97 (without metabolic activation), and TA102 (with metabolic activation). The sample collected in 1998 did not increase the frequency of his+ revertants in the same bacterial strains. Moreover, the DNA-topology assay (which facilitates electrophoretic monitoring and densitometric quantification of changes in a plasmid DNA structure) revealed the presence of 81.3% of the slowly migrating plasmid DNA, and 18.2% of the relaxed plasmid DNA after...

Benzamide riboside (BR), a recent synthetic nucleoside analogue, is a new compound demonstrating potent cytotoxic activity in malignant cell lines in vitro and in vivo in L1210 leukemia. It exhibits at least two different mechanisms of action. These are, first, the inhibition of inosine 5'-monophosphate dehydrogenase (IMPDH, EC 1.1.1.205), a rate-limiting enzyme for GTP and dGTP synthesis that plays a major role in DNA synthesis, cell proliferation and regulation; and second, the induction of apoptosis. Some aspects of BR activity in malignant cells in vitro and in vivo are reviewed as well as some of the mechanisms behind BR's anti-neoplastic effect.

Two experimental techniques, the DNA-topology assay and the Ames assay, were proved to be suitable for monitoring compounds with a genotoxic potential and/or with an antimutagenic effect. Both procedures were used in assaying the acid-mine water (AMW) containing toxic metals and sulfoethyl chitin-glucan (SE-Ch-G), a derivative of chitin-glucan, in which bioprotective activities were detected earlier. It was shown that after toxic metal concentrations were decreased due to AMW dilution to the limits that correspond with those set by the Slovak Technical Norm (STN) for drinking water, AMW was not genotoxic in the Ames assay. As it is possible to detect any single-strand DNA (ssDNA) break in the DNA-topology assay, the SE-Ch-G protective effect against the ssDNA breaks induced by Fe2+ in the DNA-topology assay was recorded. SE-Ch-G exhibited the antimutagenic potential after its application simultaneously with diagnostic mutagens in the Ames assay. These results demonstrate the complementarity of both experimental systems.

1. 1. Pharmacokinetic properties of a new derivative of the widely used and very potent antileukemic agent arabinosylcytosine (araC)—5′-chloro-5′-deoxy-arabinosylcytosine (5′-Cl-araC)—were investigated after intraperitoneal (i.p.) and oral routes of administration in rats and compared with the equimolar dose of araC administered orally. 2. 2. It was found that substitution of the hydroxyl group at position 5′ resulted in a change of pharmacokinetic parameters. 3. 3. There is a large difference in average serum concentrations of 5′-Cl-araC administered by the i.p. and oral routes; the average serum concentration obtained after i.p. injection being several times higher in comparison to those after oral administration. 4. 4. However, the latter are, at the same time, lower than the average serum concentrations of araC administered by the same route in an equimolar dose. 5. 5. On the other hand, the apparent volume of distribution is much larger, and the area under the curve of serum concentration of 5′-Cl-araC is smaller, after oral as compared to the i.p. route of administration indicating more extensive tissue distribution together with higher tissue binding of 5′-Cl-araC when compared to the parental drug araC.

The cytotoxic and antineoplastic potential of two new duplex drugs, ECyd-5-FdU and ECyd- lipid- 5-FdU, were compared with the activity of the parent single-nucleoside analogues, 3-C-ethynylcytidine (ECyd) and 5-fluorodeoxyuridine (5-FdU), either applied as monotherapy or simultaneously in equimolar concentrations simulating their ratio in a duplex drug. Murine leukaemia L1210 cells were used for comparative in vitro tests of the duplex and the single drugs. The tested substances were evaluated for their cytotoxicity, combinatory potential and revitalisation properties. Additionally, an in vivo model of leukaemia L1210-bearing mice of the DBA/2J strain was used for testing of acute toxicity and antileukaemic activity using various chemotherapeutic regimes. Based on the results of this study, the suitability of ECyd and 5-FdU for forming a duplex drug was discussed from the perspective of their expected synergistic anticancer activities. We found an improvement of chemotherapy outcomes of the new duplex drugs tested by comparing their in vitro cytotoxicity and an increase of the time of survival of experimental leukaemia-bearing mice in a statistically significant manner.

Polysaccharides represent the major part of the yeast cell wall dry weight and build the skeletal carcass defining cell wall stability and cell morphology (beta-D-glucans) or constitute amorphous matrix and cell surface fibrous material (mannans and mannoproteins). It is known that yeast cell wall beta-D-glucans reveal immunomodulating properties, which allows for their application in anti-infective and antitumor therapy. Recent data also suggest that polysaccharides reveal antioxidant activity that can result in their protective function as antioxidants, antimutagens, and antigenotoxic agents. The paper provides a review of our continuing research involving water-soluble derivatives of beta-D-glucan isolated from the baker's yeast Saccharomyces serevisiae and of a glucomannan isolated from the industrial yeast Candida utilis. The results are confronted with the available literature data. The derivatives of beta-D-glucan demonstrated potent inhibitory effect on lipid peroxidation comparable to that of the known antioxidants and exerted DNA protection from oxidative damage. The free radical scavenging activity was confirmed by spin-trap electron paramagnetic resonance. Antimutagenic and antigenotoxic activity of the yeast polysaccharides was demonstrated using yeast, bacterial, and algal models. The derivatives of beta-D-glucan exerted potent enhancement of tumor necrosis factor alpha (TNF-alpha) released from murine macrophages and revealed synergistic effect with cyclophosphamide in the treatment of Lewis lung carcinoma and two types of lymphosarcoma in murine models. The results indicate significant protective antioxidant, antimutagenic, and antigenotoxic activities of the yeast polysaccharides and imply their potential application in anticancer prevention/therapy.

Cytotoxic/cytostatic activity of N-salicylidene-L-glutamato diaqua copper(II) complex (CuC) against mice leukemia cells L1210 has been estimated and their bioactivity was enhanced by addition of ascorbic acid. The Cu-complex with isoquinoline ligand (IQ-CuC) had stronger cytostatic effect (IC50 =15.6 microM) than parental complex (CuC) and its cytotoxicity several times increased in the presence of 0.1 mM ascorbic acid (IC50 =1.0 microM). The cytotoxicity has been caused by oxidative stress, enhanced creation of TBARS has been confirmed, and formation of 2',7'-dichlorofluorescein from 2',7'- dichlorodihydrofluorescein has been observed, also. Some hallmarks of apoptotic/necrotic death of L1210 cells have been observed by fluorescent microscopy after dyeing of cell with propidium iodide and Hoechst 33342. In addition, it was confirmed that both complexes in the presence of ascorbic acid cleavaged of pDNA. Although these copper complexes were initially prepared as substances with antioxidant properties we have showed that combined treatment of L1210 cells with IQCuC and ascorbic acid induced strong oxidative stress and death of cells. Our results confirmed that physiological concentration of ascorbic acid increases the cytostatic/cytotoxic efficiency of N-salicylidene-L-glutamato diaqua copper(II) complexes.

This review deals with alpha-lipoic acid (LA) from the point of its chemical and biological characteristics affecting enzymatic activities that are part of cellular biochemical processes in normal and cancer cells. This includes attributes of LA that are related to its ability to act as a free-radicals scavenger and also as a radical generator. LA is discussed in the light of its physico-chemical features, toxicity, biochemical bases of LA biological activities, and mechanisms of action. Additionally, it is discussed how these properties of LA are reflected by results of in vivo experiments with cancer cells and in experimental cancer chemotherapy. Finally, the results of LA use in human cancer chemotherapy and as chemopreventive agent are discussed in the light of LA future inclusion into chemotherapeutic protocols.

The extract of artichoke Cynara cardunculus L. (CCE) was investigated for its potential antigenotoxic and antioxidant effects using four experimental model systems. In the Saccharomyces cerevisiae mutagenicity/antimutagenicity assay, CCE significantly reduced the frequency of 4-nitroquinoline-N-oxide-induced revertants at the ilv1 locus and mitotic gene convertants at the trp5 locus in the diploid Saccharomyces cerevisiae tester strain D7. In the simultaneous toxicity and clastogenicity/anticlastogenicity assay, it exerted an anticlastogenic effect against N-nitroso-N'-methylurea-induced clastogenicity in the plant species Vicia sativa L. On the contrary, despite CCE not being mutagenic itself, in the preincubation Ames assay with metabolic activation, it significantly increased the mutagenic effect of 2-aminofluorene in the bacterial strain Salmonella typhimurium TA98. In the 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging assay, CCE exhibited considerable antioxidant activity. The SC50 value representing 0.0054% CCE corresponds to an antioxidant activity of 216.8 microm ascorbic acid which was used as a reference compound. Although the mechanism of CCE action still remains to be elucidated, different possible mechanisms are probably involved in the CCE antigenotoxic effects. It could be concluded that CCE is of particular interest as a suitable candidate for an effective chemopreventive agent.

Cytosine arabinoside is routinely used for treatment of leukemias and lymphomas. However, because of its extensive metabolic inactivation and limited activity in chemotherapy, new analogues of araC are being tested. The aim of this work was to synthetize two araC conjugates and evaluates their cytotoxic/antileukemic activity.Synthesis of araC-sulfonamide conjugates A and B was performed in anhydrous conditions using cyclostyling and 5'-chlorocyclocytidine as starting material. Elemental analysis and NMR, IR and UV spectrometry were used for structure confirmation. The synthesized araC conjugates were tested for their cytotoxicity in L1210 leukemia cells in vitro and for therapeutic activity and toxicity in vivo in leukemia L1210-bearing mice.The cytotoxic activities of araC and two synthesized conjugates A and B were expressed as IC(50) (micromol/l) and were compared respectively. The conjugate A is 303-times less active and the conjugate B is 757-times less active than araC. Consequently, the antileukemic activity and the acute toxicity of these compounds were examined in experiments involving leukemia L1210-bearing mice. Statistically significant therapeutic outcome was observed when the dosage of both araC conjugates was increased 10-times compared to araC. Next, the ration of cytotoxicity vs therapeutic activity for araC and both conjugates was performed. It was recorded that the ration between cytotoxicity and therapeutic activity for araC is 3333, for the conjugate A and B, the ration is significantly lower (110 and 44). This indicates that the inactivation of araC conjugate A is 30-times slower and the inactivation of conjugate B is 75-times slower as araC inactivation.The differences in cytotoxic and therapeutic activity registered in araC treatment and between two araC-analogues are most probably caused by slow liberation of araC from both conjugates. We are considered that prolonged araC liberation protected them from inactivation and extended the time period of therapeutic action both araC conjugates. The obtained results can serve as stimuli for further investigation of new araC-analogues.

Various amphiphilic heterodinucleoside phosphates containing 1-beta-D-arabinofuranosylcytosine (ara-C) and 5- fluorodeoxyuridine (5-FdUrd) have recently been synthesized in order to increase the efficacy of ara-C and 5-FdUrd. Employing growth inhibition and growth recovery assays, we evaluated the in vitro effects of four of these dimers (No. 2, 2A, 3, 10) in L1210 and P388D1 murine leukemia cells. Although ara-C and 5-FdUrd appeared equimolar in all dimers, their contribution to the cytotoxicity of these agents was different. Thus, the liberation of ara-C and 5-FdUrd from their dimeric origin and their subsequent metabolic activation had a different course. In another set of experiments, we examined the in vivo effects of these agents in mice. The dimer with the highest cytotoxicity in vitro exerted the lowest acute toxicity and yielded the lowest therapeutic effect in vivo. The obtained data indicate that dimers with slower liberation of ara-C and 5-FdUrd were less cytotoxic, but prolonged liberation of both antimetabolites protected them from inactivation and extended the time period of therapeutic action. Some of the dimers exceeded the synergistic effects yielded by simultaneous application of both ara-C and 5-FdUrd. The significantly higher therapeutic potential of these new antitumor agents indicates that further studies are warranted.

Naturally occurring polysaccharides isolated from the yeasts are the substances with versatile intriguing biomodulatory activities. One of the novel derivatives prepared from the (1 --3)-beta-D-glucan isolated from the cell walls of baker's yeast Saccharomyces cerevisiae is sulfoethyl glucan (SEG). Its DNA-protective, antimutagenic, anticlastogenic and cytotoxic/cytostatic enhancing effect was evaluated using five eukaryotic systems. SEG showed bioprotective effect in recombination- repair-deficient strain of alga Chlamydomonas reinhardtii against methyl methanesulfonate-induced genotoxicity, antimutagenic effect against ofloxacin-induced genetic changes in yeast Saccharomyces cerevisiae assay and anticlastogenic activity in plants Vicia sativa and Vicia faba assays against maleic hydrazide-induced clastogenicity. In the combined application with cytostatic drug vumon, SEG exerted enhancement of the drug's cytotoxic/cytostatic effect in the cell revitalization assay using mouse leukemia cells. The study sheds light on the possible mechanisms of actions and utilization of this microbial polysaccharide derivative in the cancer prevention and therapy.

Naturally occurring dietary compound resveratrol (RES), possessing chemopreventive and cytostatic properties, has been shown as potent sensitizer for apoptosis induced by a variety of anticancer drugs. Cell cycle analysis in sensitive promyelocytic leukemia HL60 cell line and its multidrug-resistant variant HL60/VCR (P-gp positive) treated with RES resulted in cell cycle arrest in S-phase in both cell variants. Flow cytometry measurements showed diverse activities of RES in combination with anticancer drugs doxorubicin (DOX), cycloheximide (CHX), busulfan (BUS), gemcitabine (GEM) and paclitaxel (PTX), in some cases resulting in apoptosis induction, preferentially at the expense of S-phase. Thus, RES could become a candidate to enhance the efficacy of combination anticancer therapy in a variety of human cancer cells inclusive leukemias.

Triterpenoids are natural, biologically active compounds extracted from many plants. They possess antiinflammatory, anticancer, and antioxidant properties. In the report presented, antiproliferative effects and leukemia cell growth and apoptosis modulating activities of ursolic acid (UA) and oleanolic acid (OA) were investigated. Both triterpenoids are inhibitors of leukemia cell growth and inductors of apoptosis. However, when applied in combination with anthracycline antitumor antibiotic doxorubicin (Dox), UA and OA diversely modulate therapeutic efficacy of Dox, due to different antioxidant activities. Compare to OA showing synergism/additive effect with Dox, UA (stronger antioxidant) acts antagonistically and reduces leukemia cell growth inhibiting and differentiation effects induced by Dox. In conclusion, these findings suggest that although triterpenoids UA and OA can induce apoptosis, their antioxidant activities can interfere with the therapeutic effect of antitumor antibiotic Dox which mechanism of action is attributed to the production of reactive oxygen species.

We describe the use of the new ribonucleotide reductase inhibitor, trimidox (TDX), in combination chemotherapy under in vitro and in vivo conditions with cisplatin and cyclophosphamide. In vitro, the combination of TDX and cisplatin was tested in L1210 cells. The combination caused concentration dependent antagonistic or additive effects. However, the combination of TDX-cisplatin-cyclophosphamide in vivo is highly synergistic in both, the L1210 and P388D1 leukemia mouse models. Both combinations, TDX with cisplatin or TDX with cyclophosphamide were also synergistic in the L1210 and P388D1 leukemia animal models.

Antimutagenic, anticlastogenic, and bioprotective effect of polysaccharide glucomannan (GM) isolated from Candida utilis was evaluated in four model test systems. The antimutagenic effect of GM against 9-aminoacridine (9-AA)- and sodium azide (NaN3)-induced mutagenicity was revealed in the Salmonella typhimurium strains TA97 and TA100, respectively. GM showed anticlastogenic effect against N-nitroso-N'-methylurea (NMU) induced chromosome aberrations in the Vicia sativa assay. The bioprotective effect of GM co-treated with methyl-methane-sulphonate (MMS) was also established in Chlamydomonas reinhardtii repair deficient strains uvs10 and uvs14. The statistically significant antimutagenic potential of GM was not proved against 4-nitro-quinoline-1-oxide (4-NQO)-induced mutagenicity in Saccharomyces cerevisiae D7 assay. It may be due to bioprotectivity of alpha-mannan and beta-glucan, which are integral part of S. cerevisiae cell walls. Due to the good water solubility, low molecular weight (30 kDa), antimutagenic/anticlastogenic, and bioprotective activity against chemical compounds differing in mode of action, GM appears to be a promising natural protective (antimutagenic) agent.

In this study we have examined the antitumor effect of combined administrations of indomethacin (IND) with doxorubicin (DOX) on growth of K-562 leukemia cells. Although, as single drug treatment, only high concentrations of IND reduced growth (200 microM) and induced apoptosis (800 microM) of the K-562 cells, a synergistic effects on DOX-induced cell growth inhibition, apoptosis and differentiation were observed during the co-administration of DOX with 10 microM IND. Cells treated with this combination had elevated GSHt level compare to DOX-treated cells. Modulation of GSHt level of DOX-treated cells with Cd2+ ions or BSO confirmed its important role in processes of DOX-induced differentiation. Results of this study showed that IND has a positive effect on therapeutic efficacy of DOX, and could be a perspective modulator in cancer chemotherapy.

Effects of three flavonoids, quercetin (QU), galangin (GA), and chrysin (ChR) on cisplatin (cis-Pt)-induced apoptosis of human promyelocytic leukemia HL-60 cells and murine leukemia L1210 cells were investigated. The quantitative analysis of apoptotic DNA fragmentation was used to show that preincubation of cells with flavonoids can influence cis-Pt-induced apoptosis in different way. ChR had no effect, QU enhanced, and GA reduced apoptotic DNA fragmentation. It is also shown that combined treatment with QU and cis-Pt showed synergistic effect, however, GA combined with cis-Pt exhibited antagonism on cytotoxicity in L1210 murine leukemia cells. We assume that tested flavonoids affect the important biological activities connected with cancer chemotherapy and chemoprevention as they differently modulated the sensitivity of cells to cis-Pt treatment. QU is presented as pro-apoptotic agent and GA as agent with anti-apoptotic potential.

Several metal complex agents have already been introduced into clinical tumor therapy and others are subject of antitumor studies. In this study we focused on the tetraaza macrocyclic copper complex (Cu(TAAB)Cl-2). We studied the influence of the substance on cell growth, cell cycle, membrane integrity, necrosis, apotosis and glutathione level on the leukemic cell line L1210 in 1-day (22 h) and 3-day (72 h) experiments. The metal complex shows a dose-dependent antiproliferative effect, without affecting cell cycle phases. The present results confirm that copper complex can damage plasmatic membranes and trigger apoptosis, and that after treatment of leukemic cells with the copper complex, glutathione levels were increased. (C) 2002 Elsevier Science Ltd. All rights reserved.

The stability of the new antileukemic kojic acid derivative, 5-benzyloxy-2-thiocyanatomethyl-4-pyranone (BTMP) was investigated. The degradation of BTMP was studied using specific and reproducible HPLC and LC-MS methods. Accelerated stability studies of BTMP were conducted in 0.1 M hydrochloric acid solution, physiological phosphate buffer solution (pH 7.5) and basic phosphate buffer solution (pH 9.0) at 30, 40 and 60 degrees C, respectively. The degradation of BTMP was found to follow pseudo-first order kinetics. In basic solution (pH 9.0) BTMP underwent rapid hydrolysis at a degradation rate constant (0.183-0.638 h-1) and degradation half-life (3.67-1.06 h) depending on the temperature setting. On the other hand, BTMP was significantly stable in 0.1 M hydrochloric acid solution (kdeg: 0.0017-0.0052 h-1; degradation half-life t1/2: 408.6-135.7 h), whereas in physiological phosphate buffer solution (pH 7.5), BTMP was only moderately stable (kdeg: 0.006-0.231 h-1; degradation half-life: 117.7-3.0 h). Arrhenius plots were constructed to predict the degradation kinetic parameters of BTMP at 25 degrees C and 4 degrees C. LC-MS analyses confirmed the degradation of BTMP in basic solutions and indicated at least two degradation products; namely 5-benzyloxypyran-2-ol-4-one (m/z 217.8) and 2-thiocyanatomethylpyran-5-ol-4-one (m/z 181.6).

Fourteen substituted 4-anilinoquinazolines have been tested for cytotoxic effect and structure activity relationships. The most active derivatives were substituted by chlorine or bromine group in the aromatic ring, in the pyrimidine ring by morpholine group and in the aniline skeleton by nitro group in position 4 or 2. Derivatives 6-bromo-2-(morpholin-1-yl)-4-(4'-nitroanilino)quinazoline, 6-bromo-2-morpholin-1-yl)-4-anilinoquinazoline, 2-(morpholin-1-yl)-4-(4'-bromoanilino)-quinazoline and 6-chloro-2-(morpholin-1-yl)-4-(4'-nitroanilino)quinazoline inhibited growth of tumor cell lines HeLa, B16 and L1210. Mutagenic data provided by Ames test showed, that the compounds 6-bromo-2-morpholin-1-yl)-4-anilinoquinazoline and 2-(morpholin-1-yl)- 4-(4'-bromoanilino)quinazoline did not exhibit the mutagenic effect, whereas the compounds 6-bromo-2-(morpholin-1-yl)-4-(4'-nitroanilino)quinazoline and 6-chloro-2-(morpholin-1-yl)-4-(4'-nitroanilino) quinazoline increased slightly the number of revertants of the strain TA 98 without metabolic activation. Concentration 26 micromol/L of 6-bromo-2-(morpholin-1-yl)-4-anilinoquinazoline induced necrosis of tumor cells B16. Concentration 5.2 micromol/l induced a significant increase of filamentous actin in the transformed HepG2 cells. Derivatives 6-bromo-2-(morpholin-1-yl)-4-(4'-nitroanilino)quinazoline, 6-bromo-2-morpholin-1-yl)-4-anilinoquinazoline, 2-(morpholin-1-yl)-4-(4'-bromoanilino)quinazoline and 6-chloro-2-(morpholin-1-yl)-4-(4'-nitroanilino)quinazoline exhibited antiprotease effect on plasmine. This results could be relevant for the anticancer properties of these compounds.

Benzamide riboside (BR), a new synthetic nucleoside analogue, has demonstrated a potent cytotoxic activity in murine leukemia in vitro. The purpose of the present investigation was to examine the antitumor activity of BR in mice bearing leukemia L1210. The results revealed that BR possesses a potent antitumor activity in vivo. It increases life-span of mice with leukemia. Synergistic cytotoxicity of BR with select DNA damaging agents, cisplatin (cis-Pt) and staurosporine (STP) was examined in MTT chemosensitivity assay, FACS analyses and apoptotic DNA fragmentation on L1210 cells in culture. A simultaneous treatment of leukemia L1210 cells with the combination of BR and STP resulted in synergistic cytotoxicity that correlated with increased apoptotic activity in those cells. On the other hand, treatment of L1210 cells with combination of BR and cis-Pt resulted in antagonistic cytotoxic effect. Finally, to elucidate the synergistic effect of BR and STP in inducing apoptosis, the attention was directed to the activation of cell death processes through various cell cycle signals. This is the first report describing in vivo antitumor activity of BR and its utilization in combination chemotherapy.

Tamoxifen belongs among relatively new drugs. As it has already been shown, it undoubtedly brings a benefit to oncology patients. However, there are still questions regarding its broader use in therapy or cancer prevention. This review puts together some data available at present time with the aim of elucidating the most important aspects of its use in medical oncology.

5'-Chloro-5'-deoxy arabinosylcytosine (Cl-araC) is a more lipophilic analog of the clinically used drug--arabinosylcytosine (araC). The resistance toward the enzyme cytidine-deaminase action was described as an characteristic feature of this synthetic nucleoside. The kinetics of the Cl-araC transformation in acid and alkaline solutions was studied at various temperatures. When compared with parent compound araC, the chlorine atom at the 5' position of the nucleoside sugar moiety increases the Cl-araC stability. The chlorine atom stabilizing effect is higher in acidic conditions. Cl-araC increased stability, antileukemic activity accompanied by higher lipophilicity confirm the fact that Cl-araC belong among interesting compounds from the point of view of cancer chemotherapy.

Kojic acid as a molecule of natural origin may serve as template for the synthesis of new biologically active compounds. The synthetic KA (pyranone) derivatives possess various kinds of biological activities which are related by their similarity to flavonoids. The most important property is the antifungal and antineoplastic activity and capability of chelating metals. It is shown that the antineoplastic activity of kojic acid derivatives is based on various mechanisms of action on different levels of cellular metabolism and functions what makes this compound interesting for future investigation as cytotoxic agent.

Camptothecin (CAM) and cisplatin (cis-diamminedichloroplatinum(II), cis-Pt) were used as inducers of apoptosis in the mouse leukemic L1210 cells. Relatively high concentrations of 50 micromol cis-Pt and 50 micromol CAM, respectively, were used to induce the apoptotic DNA ladder. The simultaneous treatment of L1210 cells by the drug and pentoxifylline (PTX) resulted in a decrease of drug concentrations necessary for the induction of apoptosis. This study revealed that a cell cycle G2 checkpoint inhibitor PTX reduces time intervals necessary for the onset of drug-induced apoptosis in these cells. This fact might be important as the earlier onset of programmed cell death may decrease a risk of tumor cells to become resistant to drug therapy.

Ribonucleotide reductase is the rate limiting enzyme of de novo DNA synthesis; its activity is significantly increased in tumor cells related to the proliferation rate of the tumor cell. Therefore the enzyme is considered to be an excellent target for cancer chemotherapy. In the present study we tested the in vitro and in vivo antitumor effects of a drug combination using trimidox (3,4,5-trihydroxybenzohydroxamidoxime), a novel inhibitor of rib onucleotide reductase with adriamycin, a widely used anticancer drug. This combination was selected because adriamycin generates free radicals, which are responsible for cardiotoxic side effects of adriamycin treatment, and because trimidox has been shown to be a good free radical scavenger. The in vitro cytotoxic effect of the drug combination was examined in L 1210 mouse leukemia cells employing an MTT chemosensitivity assay. Simultaneous in vitro incubation of these cells yielded antagonistic cytotoxic effects compared to either drug alone. These effects were not caused by the involvement of p-glycoprotein mediated drug efflux. However, when the effect of trimidox and adriamycin in combination was examined in L 1210 leukemia bearing mice, antitumor effects of adriamycin could be enhanced by the presence of trimidox. Animals were treated on day two after tumor cell injection with 5 mg/kg adriamycin and received 250 mg/kg trimidox on days 2,3 and 4. Mice treated with adriamycin or trimidox alone yielded a 41 and 38% increase in life span, respectively. However, animals, which were treated with both drugs, showed a 89% increase of their life span. Our data indicate, that in vitro results of drug combinations should be interpreted with extreme caution and suggest that the in vivo combination of adriamycin together with trimidox might be beneficial for the treatment of malignancies.

The DNA-protective activity of hydroxyurea (HU) and novel ribonucleotide reductase (RR) inhibitors amidox (AX), didox (DX) and trimidox (TX) was examined using hydrogen peroxide as the DNA-damaging agent. The exposure of superspiralized plasmid DNA molecules (pBR 322) to H2O2 under precisely defined in vitro conditions initiates a change in DNA topology (DNA from I relaxes to DNA form II). This electrophoretically monitored change in the plasmid DNA topology is related to the induction of ss-DNA breaks and corresponds with DNA exposition to free radicals. The inhibition of DNA relaxation (the prevention of DNA damage induced by hydrogen peroxide) depended on the free radical scavenging capacity of the drugs investigated. HU exerted DNA protective activity at a concentration of 4 mM, AX at concentration of 1 microM, TX at a concentration of 5 microM and DX at a concentration of 25 microM (the free radical scavenging activity increases from HU to AX in following manner: HUDXTXAX). It can be concluded that the new synthetic RR-inhibitor AX which is being investigated at the preclinical level as a potential anti-cancer drug possess the highest capacity for scavenging of free radicals.

The human pre-B acute lymphoblastic leukemia cell line REH6 was utilized for characterization of CD45 glycoprotein by monoclonal antibodies (mAb) recognizing four distinct CD45 antigen specificities, i.e. nonrestricted CD45, restricted CD45RA, CD45RB and CD45R0. Immunoprecipitation revealed two antigen specificities on REH6 cells of m.w. 220 kDa and 190 kDa, both presenting wide range of isoelectric point pI approximately 6.0-7.5. Nonrestricted CD45 epitopes were not affected by the sialyl acid cleavage with sodium metaperiodate or neuraminidase, but were sensitive to both, tunicamycin, the N-glycosylation inhibitor and monensin, an inhibitor of protein transport through the Golgi compartment. O-sialoglycoprotein endopeptidase from Pasteurella haemolytica A1 partially cleaved CD45RA and CD45RB epitopes, while nonrestricted CD45 determinants were not affected by this enzyme. Limited proteolysis of this antigen resulted in the appearance of 160-180 kDa peptide domains which retained CD45 epitopes. Further, the treatment of cells with phorbol myristate acetate (PMA) induced marked down-regulation of 220 and 190 kDa isoforms and the appearance of new 210, 180 and 170 kDa variant glycoprotein forms which were not found on parental cells. This PMA effect was not accompanied by the programmed cell death and was markedly blocked by a nonselective protein kinase (PK) inhibitor isoquinoline sulfonamide H7. Modulation of CD45 by phorbol esters might serve as an in vitro model for an additional insight into the function of CD45 in hematopoietic cells.

Potential gentoxicity of five new local anesthetics, derivatives of phenylcarbamic acid differing in the length of the alkyl chain of the alkoxy substituent, was studied on five test systems. There was a direct relationship with increased toxic effect in bacteria and yeast as a function of the elongation of the alkyl chain of the alkoxy substituents of the phenylcarbamic acid esters. On the other hand, no structure-toxicity relationship was found after application of 3-(2-alkoxyphenylcarbamoyloxy)-quinuclidium chlorides on plants and Drosophila. All anesthetics were nonmutagenic to Salmonella typhimurium strains TA97, TA98, TA100, and TA102 in the absence and in the presence of S9 mix. Pentyloxy and heptyloxy derivatives increased rates of genetic changes in Saccharomyces cerevisiae, mainly revertants at the isoleucine locus. Pentyloxy and hexyloxy derivatives increased the frequency of chromosome aberrations in Vicia faba root-tip meristems. No chlorophyll mutations were detected after treatment of Hordeum vulgare with pentyloxy, hexyloxy and heptyloxy derivatives. No sex-linked recessive lethals were scored in Drosphila melanogaster males. The rates of aneuploids induced in their germ cells were significantly increased after treatment with butoxy and octyloxy derivatives. However, the local toxic and genotoxic effects of test anesthetics on the microorganisms of the anesthetized tissues may be of some importance. In particular, the genotoxic effect exhibited in fungi by the heptyloxy derivative, a potent local anesthetic, was remarkable.

Hydrochloride of 5'-chloro-5'-deoxy-cyclocytidine (Cl-cC) is an analogue of cyclocytine hydrochloride (cC), a prodrug of the compound with the strong antileukemic activity arabinosylcytosine (araC). This paper is devoted to the study of its cytotoxic activity in vitro and to the effect of acid and alkaline conditions and temperature on its stability. Cl-cC inhibits not only the growth of L1210 leukemia cells in vitro and the DNA synthesis (IC50 = 0.09 mumol/l) but, at the same time, it has a weak effect on RNA synthesis (IC50250 mumol/l) and no effect on proteosynthesis. In alkaline conditions Cl-cC is transformed to 5'-chloro-araC and 2',5'-anhydro-araC but is more stable in acid solutions.

1. Occurrence of lesions induced in plasmid DNA by cis-DDP and by HA was quantified both as a transforming activity and as conformation integrity of supercoiled pBR322 DNA. Fifty per cent decrease of the biological activity of plasmid DNA, not accompanied by measurable change of DNA conformation, was observed after a single exposure of DNA to cis-DDP (1 hr/37 degrees C). 2. HA induced conversion of supercoiled DNA to other topological forms in a dose-dependent manner. 3. One- and two-strand DNA breaks were determined electrophoretically with high sensitivity. Cis-DDP exposed DNA relaxed at 30 times lower HA concentration compared to intact DNA. 4. This effect may be connected with a local distortion of DNA structure at the cis-DDP--DNA bond, which makes possible high effectivity of HA-DNA interaction. 5. On other hand, biological activity stayed at the 50% level despite breaks induced in DNA. 6. This finding supports the idea that DNA breaks occur at the locations which were modified during the exposure of DNA to cis-DDP. 7. The importance of the DNA structure during interaction with HA may be seen during HA-DNA interaction at heat-denaturation of supercoiled DNA. At this condition, the DNA breaks were induced at 100 times lower concentration of HA. 8. We conclude, on the basis of these results published earlier, that local distortion of supercoiled DNA structure, which is caused by the cis-DDP bond, and the local DNA uncoiling caused by heat-denaturation are related to high HA-DNA reactivity.

Pyridoxal and cis-diamminedichloroplatinum (II) (cis-DDP) have a different mode of interaction with DNA. Cis-DDP caused extensive transforming inactivation of pBR322 DNA but did not form strand breaks in DNA molecules. On the other hand, pyridoxal formed frank strand breaks in the DNA chains and decreased DNA transforming activity. A higher level of DNA transforming inactivation was obtained during simultaneous application of both compounds than would be an additive effect of these compounds applied to DNA independently. This synergistic effect can be ascribed to the introduction of thermolabile sites by the combined action of cis-DDP and pyridoxal. These lesions were converted by heat-treatment to DNA strand breaks and detected electrophoretically. Using two different methods, cis-DDP and pyridoxal, during simultaneous application, interacted with DNA with higher efficiency.

The aim of this study was to determine the selenium content in various tissues of the mouse employing the galvanostatic stripping chronopotentiometry (SCP) technique and to investigate the distribution profile of selenium as well as its pharmacokinetics in a mouse model.The animals received 0.25 μg/g Se orally for 5 days. Samples of whole blood and various tissues comprising kidney, liver and brain were harvested from mice and then analysed for Se content employing the SCP technique.The SCP method was validated over Se concentration range of 10 – 100 ng/mL and showed good linearity (r²0.999). The precision (over 5 days) of the assay in various mouse tissues (liver, kidney, and brain) ranged from 0.03 to 2.9% with accuracy results that varied from -6.69 to 0.28%. The mean (n = 5) recoveries of Se from the mouse tissues ranged from 93.31 to 100.28%. The lower limit of Se detection in the mouse tissues was 0.2 ng/mL. The present method was successfully applied in evaluating the distribution of Se in various tissues as well as its pharmacokinetics in the mouse model. The Se tissue concentrations in the mouse model showed that the maximum Se levels in most tissues were attained within 3-4 days following its administration. Furthermore, the pharmacokinetic profile of Se in the mouse model indicates that the element is slowly absorbed from the gastrointestinal tract (GIT) reaching a plateau in 4 days and then it is slowly eliminated from the body with a half-life of about 4.5 days.The present SCP method was employed to analyse Se in various mouse tissues. The method was characterized by excellent performance parameters necessary for the determination of Se in biological matrices. Se distributes in whole blood as well as into various tissues of the mouse with high concentrations in the kidney and liver and low levels in the blood and brain tissues. The absorption of Se from the GIT was very slow and the data suggest that the elimination of Se seems to be through the kidney at a very slow rate as well. The data of the present study thus suggest that Se remains in the mouse body for a long period of time.