Toxicity of nutritionally available selenium compounds in primary and transformed hepatocytes

The essential trace element selenium is also toxic at low doses. Since supplementation of selenium is discussed as cancer prophylaxis, we investigated whether or not bioavailable selenium compounds are selectively toxic on malignant cells by comparing primary and transformed liver cells as to the extent and mode of cell death. Sodium selenite and selenate exclusively induced necrosis in a concentration-dependent manner in all cell types investigated. In primary murine hepatocytes, the EC50 was 20 microM for selenite, 270 microM for selenate, and 30 microM for Se-methionine. In the human carcinoma cell line HepG2, the EC50 for selenite was 40 microM, and for selenate 1.1 mM, whereas Se-methionine was essentially non-toxic up to 10 mM. Similar results were found in murine Hepa1-6 cells. Exposure of primary murine cells to selenate or selenite resulted in increased lipid peroxidation. Toxicity was inhibited by superoxide dismutase plus catalase, indicating an important role for reactive oxygen intermediates. In primary hepatocytes, metabolical depletion of intracellular ATP by the ketohexose tagatose, significantly decreased the cytotoxicity of Se-methionine, while the one of selenite was increased. These data do not provide any in vitro evidence that bioavailable selenium compounds induce preferentially apoptotic cell death or selectively kill transformed hepatocytes.     

Influence of O6-methylguanine on DNA damage and cytotoxicity of temozolomide in L1210 mouse leukemia sensitive and resistant to chloroethylnitrosoureas.

Temozolomide is a new anticancer agent which in the early clinical investigation has shown promising antitumor activity. It decomposes spontaneously to the active metabolite of DTIC (MTIC). Temozolomide is more cytotoxic against L1210 than against a subline L1210/BCNU, resistant to chloroethylnitrosoureas. Using [methyl-3H] temozolomide we found that after 1 h exposure the amount of O6-methylguanine (O6mGua) was twice as high in L1210 than in L1210/BCNU whereas the amount of N7 mGua was approximately the same in the two cell lines. O6-alkylguanine DNA alkyltransferase (AT) levels were higher in L1210/BCNU than in L1210, supporting the view that the resistance to methyltriazenes is probably related to the efficient repair of O6mGua in L1210/BCNU. Exposure of L1210/BCNU cells to 0.4 mM O6mGua for 24 h resulted in a depletion of AT and in a higher temozolomide-induced cytotoxicity. In the sensitive cell line L1210, temozolomide activity was not potentiated by O6mGua pretreatment. Moreover, in L1210/BCNU, O6mGua increased DNA single-strand breaks caused by temozolomide, suggesting that O6-guanine alkylation induces an excision repair mechanism in cells depleted in AT.     

Selenium compounds modulate the activity of recombinant rat AsIII-methyltransferase and the methylation of arsenite by rat and human hepatocytes

Formation of methylated metabolites is a critical step in the metabolism of inorganic arsenic or selenium. We have previously shown that under conditions of a concurrent exposure sodium selenite inhibits methylation of arsenite by cultured rat hepatocytes. Here, we compare the effects of sodium selenite and mono-, di-, and trimethylated selenium compounds on the methylation of arsenite by purified recombinant rat As(III)-methyltransferase (Cyt19) and by primary rat and human hepatocytes. Among these compounds, sodium selenite was the most potent inhibitor of the methylation of arsenite by the recombinant enzyme (K(i) = 1.4 microM) and by cultured cells. In both systems, methylseleninic acid was an order of magnitude less potent an inhibitor (K(i) = 19.4 microM) than was sodium selenite. Dimethylselenoxide and trimethylselenonium iodide were weak activators of recombinant As(III)-methyltransferase activity but were weak inhibitors of arsenite methylation in hepatocytes. These data suggest that selenite, rather than its methylated metabolites, is responsible for inhibition of arsenite methylation in cultured hepatocytes and that inhibition may involve direct interactions between selenite and As(III)-methyltransferase.     

Initial single-strand DNA damage and cellular pharmacokinetics of bleomycin A2

The cellular association and fate of high specific activity [3H]bleomycin A2 (BLM A2) were examined in three previously untreated cultured cell lines. Human head and neck A-253 carcinoma cells were 10-fold more sensitive to a 1-hr exposure to BLM A2 than either murine leukemic L1210 or human ovarian SK-OV cells. Both murine and human cells displayed rapid drug association with steady-state drug levels being reached within 15-30 min. At steady state, the T1/2 of drug dissociation was slow (between 65 and 155 min), unaltered by 100-fold excess of unlabeled BLM A2, and unrelated to cellular sensitivity to BLM. Approximately 15% of the total cellular drug was found in the nuclei at steady state. In intact cells, BLM hydrolase activity appeared latent; significant BLM hydrolase activity was detected using broken cell homogenates with all cell types, but no extensive drug metabolism was evident in intact cells. Murine L1210 cells differed from both human cell lines in that they had only 50% of the steady-state drug levels, had lower nuclear drug content, and had markedly less initial single-strand DNA damage. Human SK-OV cells had 2.4-fold greater initial single-strand DNA damage despite similar nuclear content and a much lower rate of DNA repair. Thus, cellular or nuclear factors, in addition to BLM A2 content, affect initial single-strand DNA damage. Collectively, our data support the proposition that lesions other than single-strand DNA breaks contribute to the cytotoxicity of BLM.     

Detection of DNA single-strand breaks induced by procarcinogens in Chinese hamster ovary cells cocultured with rat hepatocytes

DNA single-strand breaks induced by procarcinogens were detected in Chinese hamster ovary (CHO) cells cocultured with adult rat hepatocytes. Freshly isolated adult rat hepatocytes were added to the CHO cell culture prelabeled with [3H]thymidine. After allowing the hepatocytes to attach on or near the CHO cells, aflatoxin B1 or benzo[a]pyrene was added to the culture and incubated for the desired time. DNA single-strand breaks in CHO cells were measured by the alkaline elution technique. Aflatoxin B1 induced some DNA single-strand breaks in CHO cells cultured alone, but in coculture system with hepatocytes the number of DNA single-strand breaks increased greatly. The magnitude of the increase was related to the dose and the time of exposure to aflatoxin B1. Addition of proteinase-K to the cell lysates increased the elution of DNA compared to that of samples without proteinase-K. Benzo[a]pyrene did not induce any DNA single-strand breaks in CHO cells in the absence of liver cells, but a significant number of single-strand breaks were detected in the coculture system.     

Protein-associated deoxyribonucleic acid strand breaks produced in mouse leukemia L1210 cells by ellipticine and 2-methyl-9-hydroxyellipticinium

DNA intercalating agents, including ellipticine, had been found previously to produce protein-associated DNA single-strand breaks and double-strand breaks in mammalian cells. The relationship between these effects on DNA and cytotoxicity could not be determined reliably for ellipticine, because of the poor solubility characteristics of this compound. Studies were therefore carried out using the cationic derivative, 2-methyl-9-hydroxyellipticinium (2-Me-9-OH-E⁺), which has adequate water solubility and retains antitumor activity. DNA single-strand breaks (SSB) and DNA-protein crosslinks (DPC) were measured using the alkaline elution (pH 12) technique, and double-strand breaks (DSB) were measured by the neutral elution (pH 10) method. The effects of ellipticine and 2-Me-9-OH-E⁺ were compared in mouse leukemia L1210 cells. Like ellipticine, moderate concentrations of 2-Me-9-OH-E⁺ produced protein-associated SSB, indicated by the appearance of SSB and DPC at approximately equal frequencies and localized with respect to each other. Below 20 μM (1-hr treatments), the effects of the two drugs were comparable in magnitude. At higher concentrations, ellipticine produced extensive DNA breakage not associated with protein; this is perhaps secondary to an action on membranes or other non-DNA targets. However, 2-Me-9-OH-E⁺ produced only protein-associated strand breaks, even at 4-fold higher concentrations. The two compounds produced similar and relatively large extents of double-strand scission. The measured DSB/SSB ratio was higher than that produced by X-ray or certain other intercalators that have been similarly studied. The DNA effects of 2-Me-9-OH-E⁺, unlike those of ellipticine, were readily reversible and, therefore, permitted a meaningful correspondence between the magnitudes of the DNA effects and the inhibition of colony-forming ability. Comparison with two other types of intercalating agents indicated that neither the SSB nor the DSB predicts the magnitude of cell killing.     

Use of mammalian DNA repair-deficient mutants to assess the effects of toxic metal compounds on DNA

Wild-type and repair-deficient cell lines ( EM9 ) of Chinese Hamster Ovary cells were utilized to assess cytotoxic responses towards metals that produce lesions in DNA. Alkaline elution studies indicated that both CaCrO4 and HgCl2 induced single-strand breaks in the DNA. CaCrO4 and HgCl2 treatments of intact Chinese hamster ovary cells also caused the induction of DNA cross links. The mutant cells, which are thought to have a defect in the repair polymerase enzyme and therefore exhibit greater sensitivity towards a variety of agents that produce lesions in the DNA such as X-rays and ultraviolet-light, also displayed a greater sensitivity, compared to wild-type cells, towards the cytotoxic response of HgCl2 and CaCrO4 . For example, the IC50 (concentration producing a 50% growth inhibition) following exposure for 6-hr to CaCrO4 or 1 hr to HgCl2 was 3.4-fold or 1.8- to 3.9-fold greater in wild-type cells compared to repair-deficient cells respectively. Mutant cells compared to wild-type cells were not more sensitive to growth inhibition by agents whose primary site of action was not at the DNA level (i.e. amphotericin B, trifluoroperazine and cycloheximide). The DNA crosslinks induced by exposure to 10 microM CaCrO4 for 6 hr were almost completely repaired in wild-type cells within 24 hr, whereas in similarly exposed mutant cells this lesion was initially more pronounced and was only partially repaired following a 24-hr recovery period in the absence of CaCrO4 . The repair of single-strand breaks induced by CaCrO4 was more rapid and similar in both wild-type and mutant cells. Since Hg(II) inhibits repair of single-strand breaks, we could not study repair of this lesion induced by this agent; however, at very low concentrations (1 microM) binding of 203Hg(II) to DNA was greater in the mutant cells compared to the wild-type cells. Following removal of 203Hg(II) from the media, mutant cells generally retained more 203Hg bound to DNA relative to the total 203Hg(II) present in the cell. These results demonstrate that an important toxic action of CaCrO4 and HgCl2 involves injury to DNA since the concentrations of these metals causing measurable DNA damage were consistent with their respective cytotoxic concentrations and DNA repair-deficient mutants displayed both enhanced cytotoxicity and decreased repair of metal-induced lesions.     

The role of thioredoxin reductase activity in selenium-induced cytotoxicity

The selenoprotein thioredoxin reductase is a key enzyme in selenium metabolism, reducing selenium compounds and thereby providing selenide to synthesis of all selenoproteins. We evaluated the importance of active TrxR1 in selenium-induced cytotoxicity using transfected TrxR1 over-expressing stable Human Embryo Kidney (HEK-293) cells and modulation of activity by pretreatment with low concentration of selenite. Treatment with sodium selenite induced cytotoxity in a dose-dependent manner in both TrxR1 over-expressing and control cells. However, TrxR1 over-expressing cells, which were preincubated for 72h with 0.1 microM selenite, were significantly more resistant to selenite cytotoxicity than control cells. To demonstrate the early effects of selenite on behaviour of HEK-293 cells, we also investigated the influence of this compound on cell motility. We observed inhibition of cell motility by 50 microM selenite immediately after administration. Moreover, TrxR1 over-expressing cells preincubated with a low concentration of selenite were more resistant to the inhibitory effect of 50 microM selenite than those not preincubated. It was also observed that the TrxR over-expressing cells showed higher TrxR1 activity than control cells and the preincubation of over-expressing cells with 0.1 microM selenite induced further significant increase in the activity of TrxR1. On the other hand, we demonstrated that TrxR1 over-expressing cells showed decreased glutathione peroxidase activity compared to control cells. These data strongly suggest that TrxR1 may be a crucial enzyme responsible for cell resistance against selenium cytotoxicity.     

SB203580, a specific inhibitor of p38-MAPK pathway, is a new reversal agent of P-glycoprotein-mediated multidrug resistance.

P-glycoprotein (P-gp) is the plasma membrane transport pump responsible for efflux of chemotherapeutic agents from cells and is one of the systems that secures multidrug resistance (MDR) of neoplastic cells. In the present study, drug sensitive L1210 and multidrug resistant L1210/VCR (characterized by overexpression of P-gp) mouse leukemic cell lines were used as an experimental model. We have found that SB203580, a specific inhibitor of p38-MAPK pathway, significantly reduced the degree of the vincristine resistance in L1210/VCR cells. This phenomenon was accompanied by a decrease in the LC(50) value of vincristine from 3.203+/-0.521 to 0.557+/-0.082 microM. The LC(50) value of sensitive cells for vincristine was about 0.011 microM. The effect of SB203580 on L1210/VCR cells was associated with significantly increased intracellular accumulation of [3H]-vincristine in the concentration dependent manner. Prolonged exposure of resistant cells to 30 microM SB203580 did neither significantly influence the gene expression of P-gp, nor change the protein levels of p38-MAPK. Western blot analysis revealed that the MDR phenotype in L1210/VCR cells was associated with increased level and activity of cytosolic p38-MAPK. In resistant cells, the enhanced phosphorylation of both, p38-MAPK and ATF-2 (endogenous substrate for p38-MAPK) was found as well. In conclusion we could remark that SB203580, an inhibitor of p38 kinase pathway, reversed the MDR resistance of L1210/VCR cells. MDR phenotype of these cells is connected with increased levels and activities of p38-MAPK. These findings point to the possible involvement of the p38-MAPK pathway in the modulation of P-gp mediated multidrug resistance in the L1210/VCR mouse leukemic cell line. However, the mechanisms of SB203580 action should be further investigated.     

In vivo effects of mercury (II) on deoxyuridine triphosphate nucleotidohydrolase, DNA polymerase (alpha, beta), and uracil-DNA glycosylase activities in cultured human cells: relationship to DNA damage, DNA repair, and cytotoxicity

The effect of mercuric acetate on the activities of deoxyuridine triphosphate nucleotidohydrolase (dUTPase), DNA polymerase (alpha, beta), and uracil-DNA glycosylase has been studied in cultured human KB cells. There was a dose- and time-dependent inactivation of both dUTPase and DNA polymerase alpha activities by mercuric acetate. In cells exposed to low concentrations (10 microM) of mercuric acetate, dUTPase was most sensitive to inhibition with 30% of the activity being inhibited after a 1-hr exposure. At higher concentrations or for longer exposure times, DNA polymerase alpha was most sensitive to inhibition with greater than 60% of the activity being inhibited by 25 microM mercuric acetate after a 15-min exposure. There was no inhibition of DNA polymerase beta or uracil-DNA glycosylase activities in cells exposed to 50 microM mercuric acetate for 90 min. In fact, there was a time- and dose-dependent activation of uracil-DNA glycosylase activity with maximum activation occurring in cells exposed to 50 microM mercuric acetate. The inhibition of dUTPase and DNA polymerase alpha activities and the activation of uracil-DNA glycosylase activity correlated with the induction of single-strand breaks in DNA by mercuric acetate and with the decrease in cell viability.     

Concurrent unilateral chromatid damage and DNA strand breakage in response to 6-thioguanine treatment

The delayed cytotoxicity of 6-thioguanine (TG) may relate to the arrest of cells in G2 upon completion of one cell cycle after drug exposure. In Chinese hamster ovary (CHO) cells, both the unilateral chromatid damage in G2 chromosomes, determined by induction of premature condensed chromosome condensation [Maybaum and Mandel, Cancer Res. 43, 3852 (1983)], and incorporation of TG into DNA resulting in DNA strand breakage [Christie et al., Cancer Res. 44, 3665 (1984)] were correlated with cytotoxicity. We have studied the correlation between strand breakage and unilateral chromatid damage in L1210 cells. DNA breaks were detected only when cells were treated with TG (0.25 microM) for one cell cycle time (12 hr) followed by 12 hr in drug-free medium containing [3H]thymidine (TdR) to label the DNA. After simultaneous incubation of cells with drug and label during the first or second 12-hr period, strand breaks were not found. Strand breaks increased with dose, which correlated with greater cytotoxicity (0.01 to 0.25 microM). Treatment of cells with 0.25 microM TG for 12 hr, and transfer to drug-free medium for 12 hr prior to making prematurely condensed chromosomes (PCC), resulted in unilateral chromatid damage. Prominent curving of G2 chromosomes with gapping and diffuse staining of one of the sister chromatids occurred. The 4-fold increase in the percentage of cells in G2 compared with control cells suggested G2 arrest. When cells were treated with TG for 12 hr and PCC made immediately, neither the arrest of cells in G2 nor unilateral chromatid damage was observed. These data suggest that strand breaks and unilateral chromatid damage occur in the second cell cycle after TG exposure and that this damage may be important in TG-delayed cytotoxicity.     

Induction of DNA single- and double-strand breaks by diethylstilbestrol in murine L5178Y lymphoblasts in vitro

The mechanism of action of the synthetic estrogen diethylstilbestrol (DES) was investigated in murine L5178Y lymphoblasts. The dose-survival curve of cells treated with DES in serum-free medium for 1 hr was characterized by a prominent shoulder followed by a simple exponential decline; the Do, the dose of DES reducing cell survival to 1/e, was 1.52 nmoles/ml. DNA single-strand breaks, as measured by the alkaline elution method, were observed in DES-treated cells, and these followed a dose-response relationship after an apparent threshold of 10 microM DES was exceeded. Protein-associated strand breaks, which represent the increment in single-strand breaks that occurs by exposing drug-treated cells to proteinase K, were also noted. DNA double-strand breaks as measured by filter elution technology at pH 9.6 were observed and increased markedly to reach a level of approximately 9000 rad equivalents at a DES concentration of 20 microM. The measured ratio (mean +/- S.E.) of single- to double-strand breaks induced by DES in L5178Y limphoblasts was 0.09 +/- 0.035. A comparison of the ratio of single- to double-strand breaks induced by DES to that observed following radiation suggested that all of the single-strand breaks produced by DES could be attributed to double-strand breaks. The close correspondence of the dose-response curve for cytocidal activity of DES with that obtained for induction of DNA double-strand breaks suggested that such breaks may play an important role in the mechanism of cell kill by DES.     

Selenium modifies the metabolism and toxicity of arsenic in primary rat hepatocytes

Arsenic and selenium are metalloids with similar chemical properties and metabolic fates. Inorganic arsenic (iAs) has been shown to modify metabolism and toxicity of inorganic and organic selenium compounds. However, little is known about effects of selenium on metabolism and toxicity of iAs. The present work examines the effects of selenite (Se(IV)) on the cellular retention, methylation, and cytotoxicity of trivalent iAs, arsenite (iAs(III)), in primary cultures of rat hepatocytes. The concurrent exposure to Se(IV) (0.1 to 6 microM) inhibited methylation and/or significantly increased cellular retention of iAs(III) in cultured cells. The ratio of the methylated metabolites produced from iAs(III), dimethylarsenic (DMAs) to methylarsenic (MAs), decreased considerably in cells treated with Se(IV), suggesting that synthesis of DMAs from MAs may be more susceptible to inhibition by Se(IV) than is the production of MAs from iAs(III). The 24-h preexposure to 2 microM Se(IV) had a similar but less pronounced inhibitory effect on the methylation of iAs(III) in cultured cells. The exposure to 2 microM Se(IV) alone for up to 24 h had no effect on the viability of cultured hepatocytes. However, concurrent exposure to 2 microM Se(IV) increased the cytotoxicity of iAs(III) and its mono- and dimethylated metabolites that contain trivalent arsenic, MAs(III) and DMAs(III). These data suggest that pre- or coexposure to inorganic selenium may enhance the toxic effects of iAs, increasing its retention in tissues and suppressing its methylation, which may be a pathway for detoxification of iAs.     

Interaction of DNA intercalator 3-nitrobenzothiazolo (3,2-a)quinolinium with DNA topoisomerases: a possible-mechanism for its biological activity

There is multiple evidence linking the inhibition of DNA topoisomerases I and II with the cytotoxic effects of antitumor drugs such as camptothecin and the DNA intercalators, 4-(9-acridinylamino)methanesulfon-m-anisidine) (mAMSA) and Adriamycin. We have assessed the effect of the DNA intercalator 3-nitrobenzothiazolo(3,2-a)quinolinium (NBQ) on the DNA topoisomerase I and II activities. NBQ has no effect on the activity of purified topoisomerase I, whereas it induced purified topoisomerase II binding to DNA without inducing DNA scission. Above 30 microM, NBQ stimulated formation of double- and single-strand breaks mediated by topoisomerase II in plasmid DNA. Using the alkaline elution technique we determined that NBQ induced single-strand and DNA-protein-associated breaks in the human promyelocytic leukemia cell line HL-60. At sublethal concentrations (less than or equal to 1 microM), NBQ induce HL-60 cells to differentiate. Topoisomerase II-mediated DNA cleavage induced by mAMSA was substantially reduced in NBQ-differentiated cells. Our data suggest that topoisomerase II could play a major role in the biological activity of NBQ in vivo.     

Cytotoxicity and deoxyribonucleic acid damage associated with bromoacetate

Bromoacetate, one of the hydrolysis products of bromoacetylcholine, has been shown previously to inhibit the growth of neuroblastoma cells in culture. Its mechanism of action is unknown. In this work we have further characterized the cytotoxic effect of bromoacetate in C-1300 neuroblastoma cells in culture and extended it to a cell line not of neural origin, the mouse leukemia L-1210 line. Doses required to inhibit the growth of L-1210 cells in culture were similar to those that inhibited the C-1300 line. Cytotoxicity depended on concentration and duration of exposure. L-1210 colony formation in soft agar was also inhibited by bromoacetate. Bromoacetate was shown to produce DNA single strand breaks in L-1210 cells by the alkaline elution assay. These breaks continued to form after the drug was removed from the medium. Taken together, the toxicity demonstrated toward L-1210 cells and the evidence of DNA damage following drug exposure indicate that the anti-tumor action of bromoacetate may be based on monofunctional alkylation of DNA and not related to inhibition of the cholinergic receptors.     

Toxicity of selenite in the unicellular green alga Chlamydomonas reinhardtii: comparison between effects at the population and sub-cellular level

The toxicity of selenium in aquatic ecosystems is mainly linked to its uptake and biotransformation by micro-organisms, and its subsequent transfer upwards into the food chain. Thus, organisms at low trophic level, such as algae, play a crucial role. The aim of our study was to investigate the biological effects of selenite on Chlamydomonas reinhardtii, both at the sub-cellular level (effect on ultrastructure) and at the population level (effect on growth). The cells were grown under batch culture conditions in well-defined media and exposed to waterborne selenite at concentrations up to 500 microM; i.e. up to lethal conditions. Based on the relationship between Se concentration and cell density achieved after a 96 h exposure period, an EC(50) of 80 microM with a 95% confidence interval ranging between 64 and 98 microM was derived. No adaptation mechanisms were observed: the same toxicity was quantified for algae pre-contaminated with Se. The inhibition of growth was linked to impairments observed at the sub-cellular level. The intensity of the ultrastructural damages caused by selenite exposure depended on the level and duration of exposure. Observations by TEM suggested chloroplasts as the first target of selenite cytotoxicity, with effects on the stroma, thylakoids and pyrenoids. At higher concentrations, we could observe an increase in the number and volume of starch grains. For cells collected at 96 h, electron-dense granules were observed. Energy-dispersive X-ray microanalysis revealed that these granules contained selenium and were also rich in calcium and phosphorus. This study confirms that the direct toxicity of selenite on the phytoplankton biomass is not likely to take place at concentrations found in the environment. At higher concentrations, the link between effects at the sub-cellular and population levels, the over-accumulation of starch, and the formation of dense granules containing selenium are reported for the first time in the literature for a phytoplankton species after exposure to selenite.     

Activity of platinum(II) intercalating agents against murine leukemia L1210

Four series of intercalating, square-planar Pt(II) complexes derived from the ligands 2,2'-bipyridine, 2,2':6',2"-terpyridine, 1,10-phenanthroline, and 3,4,7,8-tetramethyl-1,10-phenanthroline were synthesized and aspects of their activity against murine leukemia L1210 cells investigated. The 2,2':6',2"-terpyridine-thiolato complexes are growth inhibitory in culture, with IC50 values in the range 6-32 microM, and cause cell lysis at high concentrations. Of the remaining three series, the 2,2'-bipyridine complexes are the least potent in their effects. There is a general enhancement in activity on moving from the 1,10-phenanthroline complexes to the 3,4,7,8-tetramethyl-1,10-phenanthroline analogues. Flow cytometric analysis on representative complexes shows that they are not cell cycle specific. Alkaline elution experiments indicate no damage to DNA of cells exposed to (thiophenolato)(2,2':6',2"-terpyridine)platinum(II) chloride monohydrate and (ethylenediamine)(1,10-phenanthroline)platinum(II) dichloride dihydrate although (ethylenediamine)(3,4,7,8-tetramethyl-1,10-phenanthroline)platinum(II) dichloride dihydrate causes both single-strand breaks and DNA cross-links. Compounds 2a, 5a, and 6a showed no antitumor activity against L1210 in mice.     

Synthetic polyamine analogues as antineoplastics

In this paper, we report on the synthesis and biological activity of a number of N-alkylated spermine compounds. The dialkylspermines N1,N12-dimethylspermine (DMSPM-2), N1,N12-diethylspermine (DESPM-3), and N1,N12-dipropylspermine (DPSPM-4) are all shown to inhibit the growth of L1210 cells in culture with IC50 values of less than 1 microM at 96 h. Furthermore, DESPM-3 is shown to be similarly active against Daudi and HL-60 cells in culture. A structure-activity relationship is shown to exist between the position at which spermine is alkylated and its antiproliferative properties. The activity of 10 microM DESPM-3 against L1210 cells was shown to be cytostatic, with greater than 90% cell viability by trypan blue exclusion, even after a 144-h exposure. When L1210 cells were treated with 10 microM DESPM-3 over a 144-h period, their size and mitochondrial DNA content were gradually but substantially diminished. However, flow cytometric measurements of the nuclear DNA content of these treated cells at 96 h indicated only slightly reduced S and G2 populations and significant changes only after 144 h. A cloning assay performed on the cells after 96 h of exposure to this drug (10 microM) indicated that the cells were not growing. Finally, when male DBA/2 mice, inoculated with L1210 leukemia cells, were treated with DESPM-3, their life span was increased in excess of 200% relative to untreated controls. Moreover, many long-term survivors were apparently tumor free at the end of the experiment (60 days).     

The influence of selinium on methyl mercury toxicity in rat hepatoma cells, human embryonic fibroblasts and human lymphocytes in culture

The effect of methyl mercury and two selenium compounds have been studied in cell cultures. Methyl mercury in concentrations above 1 microM had a pronounced inhibiting effect on the growth of rat Morris hepatoma cells. Glucose and lactate uptake in relation to cell protein was appreciably stimulated by the organic mercury compound. Selenite in low concentration (0.5 microM) and seleno-di-N-acetyl glycine in thousandfold higher concentrations offered considerable protection against these effects of methyl mercury. The same selenite concentration (0.5 microM), which did not affect cell growth, caused an appreciable protection against methyl mercury (6 microM), even if it was added 3 days after methyl mercury. The methyl mercury inhibited the growth of human embryonic fibroblasts and the DNA-synthesis in the human lymphocytes. However, no protective effect of selenite were observed in these cell types. These results suggest that selenium compounds exert their protective effect through cell specific processes rather than by a direct chemical reaction between selenite and methyl mercury.