Effect Of Thymidine Research Articles

α-Difluoromethylornithine-Induced Cytostasis is Reversed by Exogenous Polyamines, Not by Thymidine Supplementation.

Polyamine spermidine is essential for the proliferation of eukaryotic cells. Administration of polyamine biosynthesis inhibitor α-difluoromethylornithine (DFMO) induces cytostasis that occurs in two phases; the early phase which can be reversed by spermidine, spermine, and some of their analogs, and the late phase which is characterized by practically complete depletion of cellular spermidine pool. The growth of cells at the late phase can be reversed by spermidine and by very few of its analogs, including (S)-1-methylspermidine. It was reported previously (Witherspoon et al. Cancer Discovery 3(9); 1072–81, 2013) that DFMO treatment leads to depletion of cellular thymidine pools, and that exogenous thymidine supplementation partially prevents DFMO-induced cytostasis without affecting intracellular polyamine pools in HT-29, SW480, and LoVo colorectal cancer cells. Here we show that thymidine did not prevent DFMO-induced cytostasis in DU145, LNCaP, MCF7, CaCo2, BT4C, SV40MES13, HepG2, HEK293, NIH3T3, ARPE19 or HT-29 cell lines, whereas administration of functionally active mimetic of spermidine, (S)-1-methylspermidine, did. Thus, the effect of thymidine seems to be specific only for certain cell lines. We conclude that decreased polyamine levels and possibly also distorted pools of folate-dependent metabolites mediate the anti-proliferative actions of DFMO. However, polyamines are necessary and sufficient to overcome DFMO-induced cytostasis, while thymidine is generally not.

The effect of thymidine on the antibacterial and antiviral activity of zidovudine.

The effect of thymidine and deoxyadenosine on the antiviral and antibacterial effect of zidovudine was studied in human immunodeficiency virus type 1 (HIV-1) Escherichia coli and Salmonella typhimurium. In quantitative assays, 10 micrograms mL-1 thymidine was shown to increase the 50% inhibitory concentration (IC50) of zidovudine for HIV-1 by approximately 100-fold and to reduce zidovudine (1 microM)-induced protection of C8166 cells from 2.04 to 0.18 log syncytial-forming units. Thymidine also antagonized the antibacterial effect of zidovudine for two E. coli and three S. typhimurium species in a dose-dependent manner; 10 micrograms mL-1 of thymidine increased the minimum inhibitory concentration of zidovudine for E. coli strains by 10-40-fold and for S. typhimurium strains by three-fold. Deoxyadenosine reduced the minimum inhibitory concentration of zidovudine against all five bacterial strains but had no effect on the IC50 of zidovudine for HIV-1, nor did it significantly reverse the antagonism of the antibacterial and antiviral activity of thymidine. The induction of the SOS response in E. coli was reversed in a dose-dependent manner by thymidine while the presence of deoxyadenosine increased induction of the SOS response by zidovudine at suboptimal concentrations.

Thymidine phoshorylase as a target for antiangiogenesis treatment

Thymidine phosphorylase (TP) has emerged as a promising target for antiangiogenesis treatment of cancer. Angiogenesis, the formation of blood vessels, is essential for tumors to grow in order to be supplied with nutrients and oxygen. The association of TP with angiogenesis was demonstrated in several clinical studies in various tissue types. It has been postulated that the angiogenic effect of TP is related to its enzymatic activity, which catalyzes the breakdown of thymidine to thymine and deoxyribose-1-phosphate (dR-1-P). The latter, in its parent form or in its sugar form, deoxyribose, may play a role in angiogenesis. It may interfere in cellular energy metabolism or be substrate in a chemical reaction generating reactive oxygen species. L-deoxyribose and a specific TP inhibitor, TPI, can reverse these effects, supporting the role of the enzymatic reaction and that of the sugar. Although TP is usually high in the tumor, we also observed a high expression in tumor-associated stromal cells and macrophages. In order to elucidate the mechanism of TP induced angiogenesis we have investigated the association of TP with angiogenesis, the effect of thymidine and its metabolites on angiogenic parameters (e.g. invasion), the modulation by TPI, the formation and retention of the sugar metabolites of thymidine, and the potential signalling pathways involved in the angiogenic process. We used cell lines without/low TP expression (Colo320 and RT112) and TP transfected variants (Colo320TP1 and RT112/TP). Intrinsic TP expression in cancer cells did not stimulate these cells to invade more. On the other hand, Colo320 and Colo320TP1 cells could attract endothelial cells to a high extent, but Colo320TP1 did not attract them to a higher extent. RT112/TP cells attracted more endothelial cells than RT112 (2 fold). The difference between the RT112's and Colo320's may be related to different formation of sugars. Exposure of tumor cells to thymidine resulted in a rapid formation of dR-1-P, which was rapidly degraded to deoxyribose and further metabolized to other sugar derivatives. Of the possible sugars that can be produced by the conversion of TdR, dR-5-P seems to accumulate the most. dR accumulated 3 fold higher extent in RT112/TP than in Colo320/TP1 cells. dR could be converted to advanced glycation endproducts (AGE), however this was to a lower extent than ribose. Thymidine also induced several signalling pathways in the cells, involved in migration and invasion, such as the Focal adhesion kinase (FAK), which subsequently stimulated p70/S6 phosphorylation. The latter is a downstream kinase of rapamycin and its phosphorylation is inhibited by rapamycin, an mTOR inhibitor. The association between rapamycin and TP was shown by the protection by thymidine of rapamycin induced cytotoxicity, while TPI inhibited the effect of thymidine addition. These studies clearly show a mechanistic link between TP, signalling pathways, and cell migration.

Effects of thymidine and phorbol 12-myristate 13-acetate on erythroid differentiation of K562 cells and their sensitivity to nonspecific lysis by rat splenocytes

Induction of hemoglobin synthesis in K562 cells by thymidine (2 mM)in vitro did not significantly enhance major histocompatibility complex antigen-unrestricted lysis of splenocyte-exposed K562 cells. Inhibition of thymidine-induced hemoglobin synthesis by simultaneous incubation of cells with thymidine and phorbol 12-myristate 13-acetate (100 nM) decreased cytolytic activity of splenocytes against K562 cells. Preincubation of tumor cells with phorbol ester alone did not affect major histocompatibility complex antigen-unrestricted lysis induced by rat splenocytes but decreased the basal level of hemoglobin synthesis.

Thymidine inhibits the growth-arrest-specific degradation of thymidine kinase protein in transfected L fibroblasts

The expression of murine thymidine kinase (TK) is strictly dependent on the growth state of the cell. Expressing epitope-tagged TK in LTK − cells, we have previously shown that low TK enzyme levels in G 0 cells are in part due to a dramatic decrease in TK protein stability. Here we report that thymidine, one of the substrates of TK, is able to counteract the growth-arrest-specific decrease of TK expression. While TK mRNA levels and TK translation rate are almost unaffected by thymidine, the TK protein half-life rose more than sixfold after addition of the nucleoside to resting cells. The effect of thymidine is reversible and is independent of its presence during the protein synthesis of TK. Dideoxythymidine, a specific inhibitor of the TK enzyme activity, also has the capacity to increase TK protein levels in G 0 cells, indicating that the substrate itself exerts the stabilising effect on the TK protein.

Effects of thymidine and hydroxyurea on the metabolism and cytotoxicity of 1-B-D arabinofuranosylcytosine in highly resistant human leukemia cells

We have examined the effect of the ribonucleotide reductase inhibitors thymidine (dThd) and hydroxyurea (HU) on the metabolism and cytotoxicity of high concentrations (10 to 100 mumol/L) of cytosine arabinoside (Ara-C) in a deoxycytidine kinase-deficient, highly Ara-C- resistant human promyelocytic leukemic cell subline (HL-60/Ara-C). Administration of dThd or HU (0.5 to 3 mmol/L) in conjunction with high concentrations of Ara-C either sequentially or simultaneously lead to enhanced (and in some cases a supra-additive) inhibition of HL-60/Ara-C suspension culture growth and soft agar colony formation. In addition, exposure of cells to 0.5 mmol/L dThd or HU for 48 hours significantly increased the percentage of HL-60/Ara-C in S-phase. In contrast to previous studies involving low-dose Ara-C administration, we were unable to detect dThd- or HU-mediated increments in Ara-CTP formation in HL-60/Ara-C cells treated with high-dose Ara-C despite reductions in intracellular dCTP pools as great as 65%. However, dThd or HU did increase Ara-C incorporation into newly synthesized DNA. These studies show that administration of dThd or HU in conjunction with high concentrations of Ara-C results in an enhanced antiproliferative effect of Ara-C against Ara-C-resistant leukemic cells deficient in deoxycytidine kinase.

Effects of Thymidine and Hydroxyurea on the Metabolism and Cytotoxicity of 1-B-D Arabinofuranosylcytosine in Highly Resistant Human Leukemia Cells

AbstractWe have examined the effect of the ribonucleotide reductase inhibitors thymidine (dThd) and hydroxyurea (HU) on the metabolism and cytotoxicity of high concentrations (10 to 100 µmol/L) of cytosine arabinoside (Ara-C) in a deoxycytidine kinase-deficient, highly Ara-C–resistant human promyelocytic leukemic cell subline (HL-60ZAra-C). Administration of dThd or HU (0.5 to 3 mmol/L) in conjunction with high concentrations of Ara-C either sequentially or simultaneously lead to enhanced (and in some cases a supra-additive) inhibition of HL-60Z Ara-C suspension culture growth and soft agar colony formation. In addition, exposure of cells to 0.5 mmol/L dThd or HU for 48 hours significantly increased the percentage of HL-60/Ara-C in S-phase. In contrast to previous studies involving low-dose Ara-C administration, we were unable to detect dThd- or HU-mediated increments in Ara-CTP formation in HL-60/Ara-C cells treated with high-dose Ara-C despite reductions in intracellular dCTP pools as great as 65%. However, dThd or HU did increase Ara-C incorporation into newly synthesized DNA. These studies show that administration of dThd or HU in conjunction with high concentrations of Ara-C results in an enhanced antiproliferative effect of Ara-C against Ara-C–resistant leukemic cells deficient in deoxycytidine kinase.

Cell Cycle Phase Dependence of Estrogen and Epidermal Growth Factor (EGF) Receptor Expression in MCF-7 Cells: Implications in Antiestrogen and EGF Cell Responsiveness*

In this report the effect of epidermal growth factor (EGF) and the antiestrogen hydroxytamoxifen (OH-TAM) on the cell cycle of the breast cancer cell line MCF-7 was investigated as a function of the presence of their respective receptors. For this study synchronized cells were obtained by cell incubation in the presence of 2 mM thymidine for 24 h at 37 C. The treatment led to a partial synchronization, since at the end of thymidine treatment, 80% of cells were accumulated in the G1 phase. The removal of thymidine allowed the cells to progress through the cell cycle, since between 6-9 h after the arrest of the treatment, about 50% of cells were found in the S phase. By 9-12 h, most of the cells entered the G2 phase, and by 24 h, the cells returned to the G1 phase. When MCF-7 cells were incubated in the presence of OH-TAM for various periods of time before thymidine exposure, the progression of the cells through the cell cycle was dramatically inhibited. Also, a short term antiestrogen treatment (2 h) before or immediately after the addition of thymidine led to an accumulation of MCF-7 cells in the G1 phase. However, when the cells were treated for 2 h with OH-TAM 22 h after thymidine addition or shortly after its removal from the cell culture, no effect of the antiestrogen on the cell cycle could be observed. In parallel, the effect of thymidine on the level of estrogen receptor was studied. Although low affinity estrogen-binding sites were maintained, high affinity ER were found to be dramatically reduced during the thymidine treatment. The comparison between the effect of OH-TAM on the cell cycle and the expression of ER revealed that the antiestrogen OH-TAM was effective only in the presence of ER. EGF was found to have no effect on the cell cycle of thymidine-synchronized cells, although it did partially reverse the G1 phase block induced by OH-TAM when added simultaneously to cell culture 24 h before thymidine exposure. The parallel analysis of EGF receptor level demonstrated that thymidine treatment also reduced EGF receptors that were found to reappear after the synchronization, during the S and G2 phases of the cell cycle.(ABSTRACT TRUNCATED AT 400 WORDS)

Effects of thymidine and uridine on the phosphorylation of 3'-azido-3'-deoxythymidine (zidovudine) in human mononuclear cells

The effects of thymidine and uridine on the phosphorylation of 3'-azido-3'-deoxythymidine (AZT) were studied in various human mononuclear cell preparations. Thymidine suppressed [3H]AZT phosphorylation in the same concentration range (20 to 100 microM) in which it antagonizes the anti-human immunodeficiency virus activity of AZT. Uridine, in turn, had no influence on AZT phosphorylation, just as it has no effect on the anti-human immunodeficiency virus activity of AZT. These findings are consistent with a close relationship between the inhibition of AZT phosphorylation and the influence of physiological nucleosides on the antiviral activity of AZT.

Increased cAMP in proximal tubules is acute effect of nicotinamide analogues

The possible role of adenosine 3',5'-cyclic monophosphate (cAMP) in the mechanism of the acute inhibitory effects of nicotinamide and analogues on brush-border membrane (BBM) phosphate transport was investigated. Compared with basal values, cAMP content of rat renal proximal tubule suspensions was elevated two- to fivefold when incubated at 37 degrees C for 1 h with nicotinamide, 5-methylnicotinamide, or picolinamide at 1-3 mM and in the presence of a phosphodiesterase inhibitor. Thymidine had no effect on cAMP content. There was significant and specific inhibition of BBM transport of phosphate when proximal tubules were incubated with either nicotinamide or picolinamide at concentrations that increased tubule cAMP content. Thymidine had no effect on BBM transport of phosphate. These findings were independent of the dietary Pi intake of the rats. The absence of any effect of thymidine on phosphate transport strongly suggests that inhibition of poly(adenosine diphosphate ribose) polymerase does not play a role in nicotinamide action on phosphate transport. The change in phosphate transport induced by nicotinamide occurred with no change in NAD content. These findings indicate that an increase in cAMP, rather than NAD, is the important change that may mediate the acute inhibition of Na(+)-dependent phosphate transport by nicotinamide.

The effects of thymidine on deoxyribonucleotide pool levels, cytotoxicity and mutation induction in friend mouse erythroleukaemia cells

The ability of excess thymidine (10 −6−10 −3M) to enhance the frequency of 6-thioguanine (6-TG) resistant cell mutants and 2,6-diaminopurine (DAP) resistant cell mutants in Friend mouse erythroleukaemia cells, clone 707, was investigated. A significant increase in mutant frequency for both markers was observed at the higher (10 −4 and 10 −3 M) thymidine treatments. Measurements of deoxyribonucleoside triphosphate pool sizes in the cells revealed a dramatic elevation of the deoxythymidine triphosphate and deoxyguanosine triphosphate pools, an increase in the deoxyadenosine triphosphate pool and an almost complete disappearance of the deoxycytidine triphosphate pool at the higher thymidine treatments. This complemented the mutagenesis data. These results support the view that increases in mutant frequency may take place following perturbations in DNA precursor pools through a resultant decrease in the fidelity of DNA synthesis. Measurements of deoxyribonucleoside triphosphate pools were also carried out on clone 707 Friend cells and a thymidine kinase-deficient subclone, 707 BUF. The thymidine kinase-deficient subclone had significantly reduced deoxythymidine triphosphate and deoxyguanosine triphosphate pools relative to those observed inclone 707 cells. The previously observed mutagen hypersensitivity in thymidine kinase-deficient Friend cells may result through pool imbalance rendering DNA excision repair error prone.

AMP deaminase and thymidine kinase deficiencies in a mutant mouse S49 cell clone.

From a mutagenized population of wild type S49 cells, a clone was isolated in a single step that possessed functional and biochemical deficiencies in both AMP deaminase and thymidine kinase activities. This mutant cell line, DTB6, was selected in semi-solid medium containing 1mM thymidine and 1mM dibutyryl cyclic AMP. In comparative growth rate experiments, DTB6 cells were considerably less sensitive than parental cells to the growth inhibitory effects of thymidine. In contrast, DTB6 cells were much more sensitive to the cytotoxic effects of adenine and adenosine. The supersensitivity of DTB6 cells toward adenine could be ameliorated by the addition of hypoxanthine to the culture medium. The growth phenotype of the mutant cells could be attributed to deficiencies in two enzyme activities. First, DTB6 cells possessed a 60-70% deficiency in AMP deaminase activity, although the residual activity appeared kinetically similar to the wild type enzyme. Second, DTB6 cells possessed a virtual complete deficiency in thymidine kinase activity. Both enzyme deficiencies behaved in a recessive fashion in intraspecies hybrids. Revertants of DTB6 cells possessed wild type levels of AMP deaminase activity but remained deficient in thymidine kinase activity, while another revertant of DTB6 cells expressed 11% of the wild type thymidine kinase level but did not perceptibly change its AMP deaminase activity. The ability to isolate single step mutants with two seemingly independent biochemical abnormalities raises the speculation that there may be some link between cellular functions responsible for purine nucleotide and thymidine metabolism.

Study of human chromosome V. The effects of thymidine concentration and timing on the expression of uridine-induced constitutive fragile sites.

The induction of fragile sites on human chromosomes has been demonstrated under various conditions that cause thymidylate stress, including exposure to uridine. In this study, we examined common fragile site expression by initially exposing peripheral lymphocytes to uridine, followed by repair of the fragile sites with media containing various concentrations of thymidine. Lymphocytes were cultured in medium 199 with 2 mM uridine. At 0.5, 1, 2, 3, 8, 10, 12, and 18 h before harvest, the uridine medium was removed and replaced by medium containing thymidine at various concentrations. Our results demonstrate that the effect of uridine on chromosome fragility can be reversed by low concentrations of thymidine (2 microM up to 200 microM) and the "rescuing" effect of thymidine can be achieved if the cells were treated prior to 2-3 h before harvest. No repair was found if thymidine was added to culture within 2 h prior to harvesting, suggesting that packing of chromosomes is also an important factor in the expression and repair of fragile sites.

The effects of exogenous thymidine on endogenous deoxynucleotides and mutagenesis in mammalian cells.

The intracellular deoxyribonucleoside triphosphate pools in mammalian cells affect diverse biological functions including the spontaneous or induced mutability. We have isolated from murine T-lymphosarcoma S49 cells, a mutant that is unable to convert dCMP to dUMP, contains deranged intracellular dNTP pools, and exhibits a mutator phenotype. The enzymatic defect in araC-6-1 cells is a deficiency of deoxycytidylate deaminase, which accounts for the high dCTP and low TTP intracellular pools. The addition of increasing concentrations of exogenous thymidine to araC-6-1 cells alters these dNTP pools in a predictable manner: increasing the TTP and diminishing the dCTP. Concomitant with this reversal of the dCTP:TTP ratio is a marked decrease in the mutation rate followed by an increase in the mutation rates at higher exogenous thymidine concentrations. This response of the mutation rate is in contrast to that seen in the control cell line containing normal deoxycytidylate deaminase. In the latter case, increasing thymidine concentration induces an enhanced mutation rate that parallels the later phase of the thymidine-induced mutation rate in araC-6-1 cells. The deficiency of deoxycytidylate deaminase, the endogeneous dNTP pool alterations, and the mutator phenotype of araC-6-1 cells are all recessive traits in cell-cell hybrids. These observations allow one to predict whether exogenous thymidine will be mutagenic, antimutagenic, or both for a given cell line and provide a basis for understanding conflicting reports in the literature concerning the effects of the thymidine on genomic stability.

Induction of sister-chromatid exchanges (SCEs) by 5-fluorodeoxyuridine The role of 5-bromodeoxyuridine incorporated into parental DNA

The effect of thymidine (dT) and 5-fluorodeoxyuridine (FdU) on the frequency of sister-chromatid exchanges (SCEs) was examined in root-tip cells of Allium cepa. dT was unable to induce SCEs at all the concentrations tested. In contrast, FdU was efficient in the induction of SCEs. A nearly 3-fold increase in the frequency of SCEs was obtained with 5 · 10 −7 M FdU when it was present only for the first 5-bromodeoxyuridine (BrdU) for 2 consecutive cell cycles. Also, when FdU was present only for the first round of replication, the rate of SCEs in M2 chromosomes was notably higher than when it was given only during the second replication round. By analyzing M3 chromosomes treated during the second replication round with FdU the results also supported the hypothesis that the majority of the FdU-induced SCEs arise as a consequence of a higher rate of BrdU incorporation, and that most of them are formed when BrdU-containing strands are used as template for DNA synthesis.

The effect of thymidine on the induction of micronuclei by alkylating agents in V79 Chinese hamster cells

Incubation in thymidine-containing medium resulted in increased lethality and micronucleus frequency in V79 cells treated with ethyl nitrosourea (ENU), methyl nitrosourea (MNU) and ethyl methanesulphonate (EMS) but not with methyl methanesulfonate (MMS). Thymidine had no effect in ENU treated HeLa cells. In V79 cells, the presence of thymidine during post-treatment DNA replication was necessary for the effect. It is suggested that the increase in chromosome damage was the result of an increased O 6-alkylguanine-thymine mispairing in cells which are defective in the repair of O 6-alkylguanine. Treatment of V79 cells with O 6-ethylguanine resulted in increased production of both micronuclei and polyploid cells. These effects might be explained by spindle dysfunction caused by the alkylated guanine.

Deoxyribonucleoside triphosphate metabolism and the mammalian cell cycle: Effects of thymidine on wild-type and dCMP deaminase-deficient mouse S49 T-lymphoma cells

The size of the dCTP pool has been implicated as a possible regulator of DNA synthesis. In this investigation we correlate large intracellular variations in deoxyribonucleoside triphosphate levels to the growth rates and cell-cycle kinetics of mouse S49 T-lymphoma cells. Wild-type and a mutant line AzidoC-100-5, lacking dCMP-deaminase activity resulting in a 10-fold expanded dCTP pool were studied and compared using flow cytometry, centrifugal elutriation and nucleoside triphosphate determinations. An increase in the dCTP pool was closely correlated to the passage of cells from G1 to S phase in both cell types. Addition of thymidine to wild-type and mutant cells resulted in an accumulation of cells in early S phase, concomitant with a decreased dCTP level. Mutant cells excreted large amounts of deoxycytidine into the medium which partially protected the cells from thymidine inhibition. The doubling times for the mutant and wild-type cells were very similar but the mutant had a somewhat prolonged S phase and shortened G1 phase compared with the wild-type cells. Large changes in the DNA precursor levels were produced by addition of thymidine to mutant cultures. This gave no change in the growth rate but a somewhat shortened S phase and prolonged G1. The biochemical background for these effects is discussed.

Modulation of anti-metabolite effects: Effects of thymidine on the efficacy of the quinazoline-based thymidylate synthetase inhibitor, cb3717

CB3717 ( N-(4-(N-((2- amino-4- hydroxy-6- quinazolinyl) methyl) prop-2- ynylamino) benzoyl)- l- glutamic acid ) is an antitumour agent that inhibits thymidylate synthetase (TS). A dose-dependent fall in plasma thymidine (dThd) (1.43 μM to 0.47 μM) occurred in non-tumour-bearing mice following the administration of CB3717. Further, in mice carrying the L1210/CBRI tumour, the drug's antitumour properties were ablated by co-administration of dThd, an effect consistent with TS being the cytotoxic locus. In vitro studies of protection by dThd against CB3717 cytotoxicity were carried out in an attempt to quantify this reversal. The metabolism of [ 14C]-dThd was measured in cultures of L1210 cells (10 4/ ml) exposed to a completely cytotoxic dose of CB3717 (50 μM). The cytotoxicity of the drug was only expressed when the dThd concentration (0.5–2 μm) had fallen to <0.1 μm in the media. This reduction was due to: (1) dThd incorporation into DNA, (2) catabolism of dThd to thymine. By reducing the initial cell concentration to 10 3/ml the depletion of dThd was substantially reduced and consequently cells continued to grow for a longer period. The critical concentration of dThd, below which growth in the presence of CB3717 could not be supported was estimated to be between 0.026 and 0.1 μM. Thus even the minimum level of dThd achieved in vivo was still in excess of that required for protection from CB3717 toxicity in vitro. There was a small accumulation of deoxyuridine (dUrd) (~2-fold) in mouse plasma 24 hr after completion of a 5-day course of CB3717 (200 mg/kg) but in vitro studies demonstrated that this was unlikely to modulate CB3717 toxicity in the presence of dThd. We caution against the use of rodent tumour models (or human tumour xenografts) for antitumour or toxicity testing of compounds designed to inhibit the de novo synthesis of thymidylate; they may be misleading because the high dThd levels found in these animals compared with man may mask the cytotoxic effects of these drugs.

Effect of thymidine on the toxicity and antitumor activity of cis-diamminedichloroplatinum (II).

The effect of thymidine (TdR) on the preclinical toxicity of cis-diamminedichloroplatinum (II) (DDP) was investigated in the BDF1 mouse and the Sprague-Dawley rat. The effect of TdR on the antitumor activity of DDP was investigated using the ascites P388 murine leukemia model. TdR at 500 mg/kg consistently decreased the recovery of body weight after DDP treatment IP, but did not affect the lethal toxicity of DDP to non-tumor-bearing mice or those with the P388 murine leukemia. This effect was greatest when TdR was injected 30 min prior to DDP and at higher doses of DDP. A 500-mg/kg dose of TdR did not affect the antitumor activity of DDP 5 mg/kg administered on days 1, 5, and 9. Treatment of rats with TdR 500 mg/kg according to various schedules of timing relative to a 5-mg/kg dose of DDP did not consistently affect the DDP-related loss in body weight or nephrotoxicity at day 3. Pretreatment of mice with TdR 1,500 mg/kg 30 min prior to DDP 5 mg/kg (every 4 days X 3) resulted in a slower recovery of body weight, which became more pronounced with increasing doses of DDP. Pretreatment of ascites P388-bearing mice with TdR 1,500 mg/kg increased the number of early deaths when mice were treated with DDP 5 mg/kg (days 1, 5, and 9). These data suggest that the cytotoxicity of DDP is increased by TdR only at higher doses of either drug, but that the antitumor activity against P388 murine leukemia is not affected.

Modulation of 5-fluorouracil metabolism by thymidine: In vivo and in vitro studies on RNA-directed effects in rat liver and hepatoma

The effects of thymidine (TdR) co-administration on the cytotoxicity and incorporation of 5-fluorouracil (5-FU) into RNA of various tissues was studied in rats bearing an ascites hepatoma (AH 130). The role of pyrimidine degradation in determining the modulating effects of TdR on the formation of FU-RNA was studied in hepatocytes and AH 130 cells in vitro. TdR (500 mg/kg) potentiated the antitumour effect of 5-FU (150 mg/kg) and also increased host toxicity as judged by changes in body weight. TdR given alone did not significantly affect tumour growth and body weight gain. Examination of the effect of TdR on the incorporation of 5-FU into RNA revealed a differential modulation of RNA-directed toxicity in different tissues. Incorporation of 5-FU into RNA in tumour and bone marrow was increased 2- and 4-fold, respectively. In spleen and kidney the incorporation increased by approximately 50%, but the values did not reach statistical significance. In contrast, the incorporation into RNA of liver and intestinal mucosa was decreased to ca 35% of the control. TdR at concentrations of 40 μM–40 mM progressively inhibited the degradation of 5-FU and decreased the incorporation of 5-FU into RNA of hepatocytes in vitro. In AH 130 cells in vitro TdR did not significantly influence the metabolism of 5-FU and the incorporation into RNA. These results demonstrate that the enhanced incorporation of 5-FU into tumour RNA in vivo after pretreatment with TdR is related not to local effects on the tumour cells but rather to an increased bioavailability of the drug. Although co-administration of TdR did not selectively enhance the antitumour effect of 5-FU, a differential toxicity in host tissues was indicated by the modulated incorporation of 5-FU into RNA