Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: HUMANGGP:024500 (thymidylate synthase)
 2,970 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We evaluated antitumor effect histologically and assayed the tissue levels of 5-fluorouracil (5-FU) and thymidylate synthase (TS) activity using surgical specimens obtained from the patients with rectal cancer, who were given tegafur suppositories prior to surgery. The antitumor effect was evaluated histologically according to classification of the general rules for the gastric cancer study (Japanese Research Society for Gastric Cancer). In 39 patients, 16 tumor specimens revealed no effect (grade-0), 22 tumors grade-1 effect, and one was not evaluable because of the severe inflammatory changes. In 23 of these patients, resected specimens were available for the assay. 5-FU levels in cancer tissues were significantly higher than those in normal tissues, and TS inhibition rates (TSIR) were almost identical, averaging around 20%, in both cancer and normal tissues. Comparing the 5-FU levels and TS activity according to the histological effects (i.e.: 'grade-0' vs 'grade-1'), the 5-FU levels in the tumors achieved grade-1 were significantly higher than in the tumors showed 'grade 0' (p less than 0.01), and TSIR in the former were relatively greater than in the latter (p = 0.053). It is suggested that both tissue levels of 5-FU and TSIR may be useful parameters to predict the anti-tumor effect against rectal cancer after administration of 5-FU and its derivatives.
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PMID:[Neo-adjuvant chemotherapy with tegafur suppository for rectal cancer--evaluation of the antitumor effects, tissue levels of 5-FU and inhibition of thymidylate synthase. Tochigi Colorectal Cancer Study Group]. 151 26

Folinic acid (leucovorin, [LV]) can potentiate the growth inhibitory effects of fluorouracil (5-FU) in vitro and in vivo. LV is a precursor for 5,10-methylene-tetrahydrofolate (CH2-THF). Sufficient levels of CH2-THF enhance the inhibition of the enzyme thymidylate synthase by the 5-FU metabolite FdUMP. This study describes the effects of 5-FU and LV in two murine (C26-10, C38-1) and two human (WiDr, HT-29) colon carcinoma cell lines and in two murine tumors (Colon 26 and Colon 38). In vitro, only C38-1 was more sensitive for the combination of LV/5-FU compared with 5-FU alone. This effect was not dose or schedule dependent. l-LV, a purified stereo-isomere of LV, is thought to be the biological active form. Tests with this compound in vitro did not show a better effect than the mixture of d- and l-LV. In vivo, dl-LV could potentiate 5-FU antitumor effect in two murine colon tumors (Colon 26, Colon 38). This effect was clearly schedule dependent. dl-LV administered 1 hour before and together with 5-FU was much better than only simultaneous or posttreatment, but there was no dose dependency, while like in vitro l-LV effect was comparable to dl-LV. TdR was used to study the role of TS inhibition in the growth inhibitory effect of 5-FU with and without LV. TdR can reverse growth inhibition caused by TS inhibition due to 5-FU. In vitro, a partial reversal of growth inhibition of 5-FU and 5-FU/LV was observed, but in vivo there was no reversal. In vivo, TdR combinations led to high toxicity. Measurements of TS amounts in cells and tumors showed that those of human origin had much lower TS than the murine. C38-1 and Colon 38 with low TS were more sensitive to 5-FU than Colon 26 with higher TS amounts. TS inhibition was studied in the two murine colon tumors at several time points after weekly 5-FU or LV and 5-FU administration. LV did not increase the extent or retention of TS inhibition due to 5-FU during the first week. After three courses of treatment a fourfold increase of TS levels was seen in Colon 26 after 5-FU therapy. This resulted in a less effective TS inhibition after this treatment. Tumors treated with 5-FU and LV also showed an increase of TS, but to a lower extent, while the effect on TS inhibition remained the same.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Effect of folinic acid on fluorouracil activity and expression of thymidylate synthase. 153 71

Antifolate inhibitors of thymidylate synthase (TS) have primarily been based on the structure of folic acid. This paper describes the identification and development of novel 6,7-imidazotetrahydroquinoline TS inhibitors by iterative ligand design, synthesis, and crystallographic analysis of protein-inhibitor complexes. Beginning with a high-resolution crystal structure of E. coli TS (TS, EC 2.1.1.45), an imidazotetrahydroquinoline inhibitor was designed de novo to occupy the folate binding pocket. Structural modifications of the initial compound 1h (Ki approximately 5 microM human/E. coli TS) were then made on the basis of feedback from additional cocrystal structures and activity data. An amino group in the 2-position of the imidazole was found to increase the potency of the series by 1-2 orders of magnitude. Other substitutions on the imidazole ring (1-CH3, 2-CH3, 2-NHCH3, 2-SCH3) generally led to weaker inhibition. Additional improvements in activity were obtained by modification of the substituents on the tetrahydroquinoline nitrogen, bringing the Ki of three of the compounds below 15 nM against the human TS enzyme. The compounds were tested for cytotoxicity and were shown to inhibit the growth of three tumor cell lines in vitro.
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PMID:Design and synthesis of novel 6,7-imidazotetrahydroquinoline inhibitors of thymidylate synthase using iterative protein crystal structure analysis. 154 76

Twenty-five mouse lung tumors induced by a single urethan treatment in female A/J, BALB/c, and (A/J x C3H/He)F1 (AC3) mice were analyzed for the presence of mutations at codon 61 of the Ki-ras gene and for the expression of the surfactant protein A (SP-A), retinoblastoma (Rb), growth arrest-specific-3 (gas-3), p53, c-myc, and thymidylate synthase (TS) genes. Ki-ras codon 61 mutations were detected in 22 of 25 tumor samples without differences among strains. In comparison with normal lungs, all the tumors showed increased SP-A mRNA levels, indicating their derivation from alveolar type II pneumocytes or Clara cells. Rb and gas-3 transcripts were instead found in all tumors at about tenfold and about 20-fold reduced levels, respectively. No apparent structural alterations or loss of heterozygosity at the Rb locus was detected in any tumors. The p53 mRNA was observed without variation in quantity or size in lung tumors and normal tissue. A threefold to fivefold c-myc overexpression was observed, without amplification of the gene. TS expression was only slightly increased, indicating no great differences in cell proliferation between lung tumors and normal tissue. Our data suggest that the pathogenesis of urethan-induced lung tumors in mice involves specific and recurrent molecular alterations (Ki-ras mutations, decrease of Rb and gas-3 expression, and increase of c-myc expression) that could represent different steps in lung carcinogenesis.
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PMID:Multiple molecular alterations in mouse lung tumors. 155 14

The inhibition of thymidylate synthase (TS) by the fluorouracil (5-FU) metabolite 5-fluoro-2'-deoxyuridine-5'-monophosphate (FdUMP) is considered to be one of the main mechanisms of action of 5-FU. The inhibition of TS is mediated by the formation of a ternary complex between TS, FdUMP, and the folate cofactor 5,10-methylene tetrahydrofolate. The activity of TS, its inhibition by FdUMP, and the binding of FdUMP to TS have been determined in biopsy specimens of colorectal tumors, liver metastases, normal colon mucosa, and liver obtained from patients who never had received chemotherapy, and patients treated with 5-FU or 5-FU with leucovorin (LV). In nontreated patients we observed a large variation in the activity of TS both at 1 microM and 10 microM dUMP (40- to 80-fold difference). In contrast, in normal colonic mucosa this variation was less than 10-fold. FdUMP binding in tumors also varied considerably but was not detectable in normal mucosa. The deviations from normal (ie, as found in mucosa) kinetic patterns of TS may represent a mutant TS form. Thirty-five patients with advanced colorectal cancer received 5-FU (500 mg/m2) at 1 to 48 hours prior to surgery. In biopsy specimens of tumor and normal tissues the residual catalytic activity of TS and the percentage of free-binding sites for FdUMP (TS-free) were determined. After dissociation of FdUMP, total catalytic activity of TS- and total FdUMP-binding sites (TS-tot) were determined. Total and residual catalytic TS activity in primary tumors and metastases showed a large variation. TS-tot and TS-free in tumors also varied considerably. At least eight patients with an undetectable TS-free showed response to subsequent intraarterial treatment with 5-FU. To several patients leucovorin (2-hour infusion of 500 mg/m2) was administered with a 5-FU bolus (500 mg/m2) in the middle of the infusion. Biopsy specimens were obtained about 48 hours after treatment. In these patients inhibition of TS was markedly enhanced compared with patients who did not receive LV. The large variation in TS may be related to the observed variation in clinical response to 5-FU treatment.
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PMID:Time course of inhibition of thymidylate synthase in patients treated with fluorouracil and leucovorin. 155 55

Examples of collateral sensitivity, even in experimental tumor systems, remain few. Preliminary data from this laboratory indicated that certain tumor cells expressed increased sensitivity to cisplatin after exposure in vitro to x-irradiation. To further clarify whether the type of fractionated radiation procedure used clinically can induce hypersensitivities to certain antitumor drugs we have pre-exposed the human ovarian carcinoma cell line JA-T/P derived from a tumor from an untreated patient to fractionated x-irradiation (total dose 50 Gy) in vitro. The resultant subline JA-T/DXR-10 expressed collateral sensitivity to cisplatin (CDDP), methotrexate (MTX) and fluorouracil (5-FU), but not to acute x-irradiation. Hypersensitivity to CDDP was associated with decreased activity of DNA polymerase beta (3.5-fold, P less than .01), but unaltered glutathione metabolism. Pre-incubation with cyclosporin A or with 3-aminobenzamide significantly enhanced (twofold, P less than .01) CDDP-induced cytotoxicity in JA-T/P cells, but not in the DXR-10 subline. Consistent with MTX hypersensitivity dihydrofolate reductase activity was significantly decreased (2.9-fold, P less than .01). Despite collateral sensitivity to 5-FU, however, thymidylate synthase activity was increased (twofold, P less than .05) suggesting alternative mechanisms for 5-FU-induced cytotoxicity in these JA-T/DXR-10 cells. These data demonstrate that DNA repair and associated reduced folate metabolism can be modified not only by drugs but also by fractionated x-irradiation.
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PMID:Expression of collateral sensitivity to cisplatin, methotrexate, and fluorouracil in a human ovarian carcinoma cell line following exposure to fractionated x-irradiation in vitro. 155 59

A sensitive method for determination of incorporation of non-radiolabeled 5-fluorouracil (FUra) into RNA (F-RNA) of tumor tissue samples was established and was shown to be applicable to clinical materials. RNA fractions containing incorporated FUra were extracted from perchloric acid precipitates of tissue sonicates by KOH hydrolysis. FUra in RNA fractions was released by HCl hydrolysis at 100 degrees C and its levels were determined by GC-MS. The sensitivity of this method was 100 fold higher than that of HPLC method. F-RNA levels were dose dependent and correlated well with antitumor efficacy of the drugs. F-RNA levels in FUra resistant murine tumors were significantly lower than that in parent tumors despite of the lack of difference in FUra concentrations in tumor tissue. Based on these results, it is suggested that measurement of F-RNA levels together with the determination of FUra concentration and the inhibition rate of thymidylate synthase should be considered as a good parameters of the evaluation of antitumor efficacy of FUra and its analogs both in experimental and clinical settings.
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PMID:[Significance of measuring 5-fluorouracil incorporated into RNA of tumor tissue as a parameter for the antitumor activity of 5-fluorouracil and its analogs]. 158 Jun 41

In a previous study, we showed that 5-fluorouracil (FU) is active against the dimethylhydrazine-induced colon tumor in rats; a 7-day infusion of FU at 30 mg/kg daily produced 85% tumor-free cures. The present study examined the effects of FU alone and in combination with leucovorin (LV) or D-glucarate (GT) using an ex vivo system that maintained the growth of the rat colon-tumor explants on collagen gels. The labeling index (LI) was determined by the incorporation of [3H]-thymidine and autoradiography. The mean LI of the untreated control was 64.8% +/- 19.8%. The IC50, IC90, and IC95 values following a 7-day exposure to FU were 0.36, 0.75, and 1.22 microM, respectively. In comparison, the steady-state FU concentration required to produce 67% tumor-free cures in rats following a 7-day infusion is 1.54 microM. LV alone did not produce any antiproliferative effect at concentrations as high as 10 microM. The addition of LV at concentrations of 0.001-10 microM did not significantly reduce the IC50 of FU. The lack of effect of LV may have been due to tissue saturation with folate provided in the culture medium. GT alone reduced the tumor LI by 20%-30% at concentrations of 0.1-10 microM. GT enhanced the effect of FU. As compared with FU alone, the addition of GT at concentrations of 0.1 and 1.0 microM reduced the IC50 of FU by 47% and 60% to 0.21 and 0.16 microM, respectively. Assessment of the potentiation of the inhibitory effect of FU by GT using two-way analysis of variance and the isobologram method indicated a significant synergistic interaction between FU and GT. This interaction occurred within the FU concentration range of 0.08 and 0.4 microM. In summary, these data indicate that (a) the IC values for FU are comparable in tumor explants and in rats, suggesting that the effects in cultured tumors reflect those in intact animals; (b) GT alone showed antitumor activity, albeit relatively minor as compared with FU; (c) FU and GT exhibited synergistic activity, which was most pronounced at FU concentrations that produced submaximal activity (less than 30% inhibition of tumor LI); and (d) GT and LV had different effects on the growth inhibition by FU, suggesting that GT acts by a mechanism different from the thymidylate synthase-directed effect of FU and LV.
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PMID:Effects of 5-fluorouracil, leucovorin, and glucarate in rat colon-tumor explants. 158 77

In vitro exposure of the TR170 ovarian carcinoma cell line to six intermittent 24-h treatments with a 90% inhibitory concentration of cisplatin (CDDP) (0.15 micrograms/ml; 0.5 microM) resulted in a 2-fold stably resistant subline designated TR170/CP+ (B.T. Hill et al., Int. J. Cancer, 39: 219-225, 1987). Resistance to CDDP in these CP+ cells has now been associated with reduced uptake of 195mCDDP (2-fold; P less than 0.01) and decreased removal of specific Pt-DNA adducts, quantitated immunochemically, indicative of an apparent increased tolerance of CDDP-induced DNA damage. Specifically these resistant cells appeared deficient in removal of the major cis-Pt-(NH3)2d(pGpG) adduct and the difunctional cis-Pt(NH3)2d(GMP)2 lesion, showed less efficiency in removing cis-Pt(NH3)2d(pApG) adducts, but proved as proficient as the parental cell line in removing DNA-DNA interstrand cross-links. Activities of DNA polymerase-alpha and -beta were comparable in both lines, and no significant alterations in glutathione metabolism were identified. Response to acute X-irradiation was not modified in these TR170/CP+ cells, but they showed marked (10-fold) cross-resistance to 5-fluorouracil and, unusually, proved collaterally sensitive (12-fold) to methotrexate. Resistance to 5-fluorouracil was associated with significantly increased thymidylate synthase activity (P less than 0.01), but this was not reflected in altered gene expression, while increased sensitivity to methotrexate was accompanied by increased drug uptake but by unaltered activity and expression of dihydrofolate reductase. These results indicate that exposure to CDDP can result in numerous alterations, both intracellularly and at the cellular membrane, reflected in significant changes in the tumor cells' responses to the cytotoxic effects of a range of antitumor drugs. The clinical relevance of these observations remains to be established.
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PMID:Characterization of a cisplatin-resistant human ovarian carcinoma cell line expressing cross-resistance to 5-fluorouracil but collateral sensitivity to methotrexate. 159 24

Rapid and quantitative polymerase chain reaction (PCR) assays based upon the competitive template technique have been developed for human dihydrofolate reductase (DHFR; E.C.1.5.1.3) and thymidylate synthase (TS; E.C.2.1.1.45) mRNAs. In various tumor cell lines and clinical tumor biopsies, TS mRNA levels correlated with TS levels as determined by [3H]-fluorodeoxyuridylate binding. Levels of DHFR and TS mRNAs, determined by PCR, correlated with mRNA quantitation by conventional dot blot methodology. The ratio of TS/DHFR mRNAs in a number of human carcinoma cell lines varies from 0.4 to 9.9 but ranges from 1 to greater than 1.5 x 10(3) in a number of tumor samples. Differences in the TS/DHFR mRNA ratio in tumors as compared with cultured cells reflects low levels of DHFR mRNA in some tumors. In patients treated with a combination of 5-fluorouracil and leucovorin, mRNA levels for TS increased approximately an order of magnitude in tumor samples 4 and 24 hr after drug treatment, whereas TS levels decreased. These results have significance for the biochemical pharmacology of antifolates and fluorinated pyrimidines in vivo and the relevance of cell culture models for antifolate chemotherapy and drug resistance.
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PMID:Quantitation of dihydrofolate reductase and thymidylate synthase mRNAs in vivo and in vitro by polymerase chain reaction. 159 83


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