Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0699790 (colon cancer)
 28,837 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A series of 5-fluorouracil (5-FU)-resistant human colon H630 cancer cell lines were established by continuous exposure of cells to 5-FU. The concentration of 5-FU required to inhibit cell proliferation by 50% (IC50) in the parent colon line (H630) was 5.5 microM. The 5-FU IC50 values for the resistant H630-R1, H630-R10, and H630-R cell lines were 11-, 29-, and 27-fold higher than that for the parent H630 cell line. Using both the radioenzymatic 5-fluoro-2'-deoxyuridine-5'-monophosphate (FdUMP) binding and catalytic assays for measurement of thymidylate synthase (TS) enzyme activity, there was significantly increased TS activity in resistant H630-R1 (13- and 23-fold), H630-R10 (37- and 40-fold), and H630-R (24- and 34-fold) lines, for binding and catalytic assays, respectively, compared with the parent H630 line. The level of TS protein, as determined by western immunoblot analysis, was increased markedly in resistant H630-R1 (23-fold), H630-R10 (33-fold), and H630-R (26-fold) cells. Northern analysis revealed elevations in TS mRNA levels in H630-R1 (18-fold), H630-R10 (39-fold), and H630-R (36-fold) cells relative to parent H630 cells. Although no major rearrangements of the TS gene were noted by Southern analysis, there was significant amplification of the TS gene in 5-FU-resistant cells, which was confirmed by DNA slot blot analysis. These studies demonstrate that continuous exposure of human colon cancer cells to 5-FU leads to TS gene amplification and overexpression of TS protein with resultant development of fluoropyrimidine resistance.
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PMID:Thymidylate synthase gene amplification in human colon cancer cell lines resistant to 5-fluorouracil. 776 85

We examined the importance of dosing interval between leucovorin (LCV) and 5-fluorouracil (5-FU) on intracellular thymidylate synthase (TS) ternary complex, free TS and total TS protein levels in human MCF-7 breast and NCI H630 colon cancer cell lines. A 2- to 3-fold increase in total TS was noted when either cell line was exposed to 5-FU 10 microM plus LCV (0.01-10 microM) compared with a 1.4- to 1.6-fold increase in total TS due to 5-FU 10 microM alone. The amount of TS ternary complex formed was 2- to 3-fold higher in both cell lines treated with the combination of 5-FU and LCV compared with 5-FU alone. TS complex formation and total TS protein increased with LCV dose (0.1-10 microM). In MCF-7 cells, the maximal increase in total TS protein and TS ternary complex formation was observed when 5-FU was delayed for 4 h after the start of LCV exposure. In NCI H630 cells, maximal total TS protein and ternary complex formation occurred when 5-FU was delayed for 18 h after the start of LCV exposure. The amount of free TS did not change in either cell line whether 5-FU was given concurrently with LCV or delayed for up to 24 h. The accumulation rate of intracellular folates in the form of higher glutamates Glu3-Glu5 was rapid in MCF-7 cells (maximal formation after 4 h), whereas in H630 cells accumulation of higher polyglutamates continued to increase up to 18 h. The time of peak folate polyglutamate (Glu3-Glu5) formation coincided with the time of peak TS complex formation and total TS protein in each cell line. In these human carcinoma cell lines, the LCV dose and interval between 5-FU and LCV play a role in increased TS total protein and TS ternary complex; however, the amount of free TS is independent of the interval between 5-FU and LCV. The time-and dose-dependent increases in TS ternary complex and TS total protein are associated with differences in the accumulation of folate polyglutamates in these cell lines.
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PMID:The effect of dose and interval between 5-fluorouracil and leucovorin on the formation of thymidylate synthase ternary complex in human cancer cells. 777 3

We have analysed cell cycle variations in thymidylate synthase (TS) protein in asynchronously growing NCl H630 and HT 29 colon cancer and MCF-7 breast cancer cell lines. Western immunoblot analysis using the TS 106 monoclonal antibody revealed a 14- to 24-fold variation in TS levels between the peak exponential and confluent growth phase in the three cell lines. Similar variations in TS levels and TS activity were detected using the 5-fluorodeoxyuridine monophosphate and deoxyuridine monophosphate biochemical assays. The percentage of cells in S-phase, which paralleled changes in TS levels, reached a maximum of 38-60% in asynchronous exponentially growing cells compared with 5-10% in confluent cells. In asynchronous exponential cells, analysis of TS levels in each cell cycle phase using two-parameter flow cytometric analysis revealed that TS protein levels were 1.3- to 1.5-fold higher in S than in G0/G1 phase cells, and 1.5- to 1.8-fold higher in G2/M than G0/G1 cells. Similar differences of 1.1- to 1.5-fold between G0/G1 and S-phase and 1.6- to 1.9-fold between G0/G1 and G2/M-phase were detected by Western immunoblot and biochemical assays. TS protein was not detectable by Western blot analysis, flow cytometry or biochemical analysis in the G0/G1 population of confluent cells. Twenty-six per cent of cells in this population were G0 cells compared with 2% in exponentially growing cells. In contrast to TS, a 4-fold difference in thymidine kinase (TK) was detected between G0/G1 and S-phase cells in exponentially growing MCF-7 cells. The level of TS enzyme is associated with cellular proliferation and the percentage of cells in S-phase; however, TS protein is not exclusively associated with S-phase in asynchronously growing cells. The variation in TS levels between exponentially growing and confluent cell population appears to be due to differences in TS levels between G0 and G1 cells.
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PMID:Increased thymidylate synthase protein levels are principally associated with proliferation but not cell cycle phase in asynchronous human cancer cells. 777 4

Recombinant human interleukin-1 alpha (rIL-1 alpha), at concentrations that were not growth-inhibitory when given alone (100-10,000 U/ml), enhanced the growth inhibition resulting from a 72-h fluorouracil (FUra) exposure in HCT116 colon cancer cells. Median-effect analysis of clonogenic assays indicated that rIL-1 alpha, given 24 h prior to and following a 24-h exposure to FUra, increased lethality in a more than additive fashion. rIL-1 alpha did not appear to significantly affect [3H]-FUra metabolism, total [3H]-FUra-RNA incorporation or RNA retention after drug removal, inhibition of thymidylate synthase, or thymidine triphosphate pool depletion. During continuous exposure to rIL-1 alpha, transient stimulation of RNA and DNA synthesis was observed at 72 h, with a return to normal by 96 h. A 24-h exposure to 10 microM FUra altered the elution profile of newly synthesized DNA as monitored by pH step alkaline elution. An accumulation of lower-MW single-stranded DNA species was noted with FUra compared to control, accompanied by a significantly decreased proportion of DNA retained on the polycarbonate filter: 10% retained vs. 32% for control (P = 0.01). A 48-h exposure to rIL-1 alpha alone did not affect the elution profile of nascent DNA species, nor did it enhance the effects of FUra. Although FUra did not appreciably affect pulse [3H]-uridine incorporation into RNA for the initial 8-24 h of FUra exposure, progressive inhibition of net RNA synthesis was observed thereafter. FUra prevented the stimulatory effect of rIL-1 alpha on RNA synthesis, and net RNA synthesis was significantly inhibited (by 64-79% after 72 and 96 h) with the combination compared to rIL-1 alpha alone. Continuous exposure to 10 microM thymidine did not rescue cells from the lethality of FUra alone or the combination of FUra plus rIL-1 alpha, suggesting that depletion of deoxythymidine triphosphate as a consequence of thymidylate synthase inhibition was not the most important component of FUra toxicity. In contrast, 1 mM uridine provided partial protection against the toxicity of FUra alone or with rIL-1 alpha. Although uridine did not affect FUra metabolism, it decreased FUra-RNA incorporation by 42-60%, presumably as a consequence of the 2-fold expansion of UTP pools. [125I]-rIL-1 alpha binding was nonspecific; with a 24-h exposure, however, internalized [125I]-rIL-1 alpha exceeded cell surface-bound material by 2-fold.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Enhanced cytotoxicity with interleukin-1 alpha and 5-fluorouracil in HCT116 colon cancer cells. 778 51

Carboplatin is a better-tolerated alternative to cisplatin. JM216, the first p.o.-administrable platinum complex possesses toxicities comparable to carboplatin in Phase I studies. Together with the trans-platinum complex JM335, it provides new chemical guidelines for the development of compounds that may circumvent cisplatin resistance in tumors. Systematic structure-activity investigations have led to the discovery and development of ZD1694 (Tomudex), an inhibitor of thymidylate synthase that exploits both the reduced folate carrier and folylpolyglutamate synthetase as major determinants of its growth-inhibitory activity. Phase II studies have revealed encouraging activity against colon cancer, and Phase III studies are nearing completion. An associated structure-activity investigation has led to the development of ZD9331, a potent thymidylate synthase inhibitor which exploits the reduced folate carrier for cell entry, but which is independent of polyglutamation for its thymidylate synthase-inhibitory activity. This compound possesses antitumor activity in vivo and has been selected for full development.
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PMID:Initiatives with platinum- and quinazoline-based antitumor molecules--Fourteenth Bruce F. Cain Memorial Award Lecture. 779 1

Using an immunoprecipitation-reverse transcription-PCR technique, we characterized a thymidylate synthase (TS) ribonucleoprotein complex in cultured human colon cancer cells that consists of TS protein and the mRNA of the nuclear oncogene c-myc. TS protein is complexed in intact cells with the C-terminal coding region of c-myc mRNA that includes nucleotide positions 1625 to 1790. RNA electrophoretic gel mobility shift assays confirm a specific interaction between TS protein and c-myc mRNA and provide additional evidence that the C-terminal coding region represents an important cis-acting regulatory element. Further evidence demonstrates that the in vitro translational efficiency of c-myc mRNA is inhibited as a result of its direct interaction with TS protein. In addition, the presence of exogenous c-myc mRNA specifically relieves the inhibitory effects of TS protein on TS mRNA translation.
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PMID:Thymidylate synthase binds to c-myc RNA in human colon cancer cells and in vitro. 779 24

The cytotoxicity of idoxuridine (IdUrd), a thymidine analogue, and ICI D1694 (D1694), a folate-based thymidylate synthase (TS) inhibitor, were examined individually and in combination in two human tumor cell lines. MGH-U1 bladder cancer and HCT-8 colon cancer cells were grown as monolayer cultures with and without thymidine. The cytotoxicity of these agents alone and in combination were determined using normal human bone marrow colony-forming unit, granulocyte-macrophage (CFU-GM) as a surrogate for myelosuppression in vivo. Thymidylate synthase inhibition, IdUrd incorporation into DNA, and DNA single-strand breaks were measured in each cell line and related to cytotoxicity. The cytotoxicity of a 24-h exposure to IdUrd or D1694 increased with drug concentration in each cell line. The drug concentrations producing 50% and 10% clonogenic survival in MGH-U1 cells, respectively, were 0.006 and 0.009 microM for D1694 and 13.0 and 81.0 microM for IdUrd. Those for HCT-8 cells, respectively, were 0.009 and 0.018 microM for D1694 and 7.5 and 20.5 microM for IdUrd. The cytotoxicity of IdUrd combined with D1694 was synergistic in both MGH-U1 and HCT-8 cells as determined by median-effect analysis. The addition of thymidine at concentrations of 0.1, 0.3, and 1.0 microM to the culture medium did not decrease the cytotoxicity of D1694 in either tumor cell line. TS inhibition using the whole cell assay was observed with only D1694, producing 50% inhibition of TS activity at 0.002 microM for MGH-U1 and 0.007 microM for HCT-8 cells. IdUrd did not inhibit TS activity, nor did it enhance the TS inhibitory effects of D1694. The incorporation of IdUrd into DNA increased with increasing concentrations of D1694. This increased DNA incorporation correlated with the increase in DNA single-strand breaks. DNA single-strand breaks paralleled cytotoxicity. CFU-GM survival, exposed to the same drug concentrations as those used in the tumor cell lines, revealed that the therapeutic index was greater for the combination than for either agent alone. These findings suggest that IdUrd plus D1694 is a promising new drug combination, which may have a favorable therapeutic index in vivo.
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PMID:ICI D1694 and idoxuridine: a synergistic antitumor combination. 803 97

Translation of thymidylate synthase (TS) mRNA is controlled by its own protein product, TS, in an autoregulatory manner. Direct binding of TS protein to two different cis-acting elements on the TS mRNA is associated with this translational regulation. In this study, an immunoprecipitation-reverse transcription-PCR technique was used to identify a TS ribonucleoprotein (RNP) complex in cultured human colon cancer cells. Using antibodies specific for TS protein, we show that TS is complexed in vivo with its own TS RNA. Furthermore, evidence demonstrating a direct interaction between the mRNA of the nuclear oncogene c-myc and TS protein is presented.
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PMID:Identification of a thymidylate synthase ribonucleoprotein complex in human colon cancer cells. 826 88

The cytotoxicity of 3'-azido-3'-deoxythymidine (AZT), a thymidine analogue, and ICI D1694, a folate-based thymidylate synthase (TS) inhibitor, was examined individually and in combination in two human tumor cell lines, MGH-U1 bladder cancer and HCT-8 colon cancer cells, grown as a monolayer culture with and without thymidine (TdR). In addition, TS inhibition, [3H]AZT incorporation into DNA, [3H]AZT-MP (monophosphate) production, and DNA double-strand breaks were measured. Twenty-four hour exposure of AZT at 0.5, 5, 50 and 500 microM was not cytotoxic to MGH-U1 or HCT-8 cells in a colony-forming assay. ICI D1694 cytotoxicity increased with drug concentration, and the IC50 and IC90, respectively, were 0.0064 and 0.01 microM in MGH-U1 cells and 0.009 and 0.018 microM in HCT-8 cells. TdR in concentrations of 0.1 to 1.0 microM did not affect ICI D1694 cytotoxicity in either cell line. AZT at 5, 50 or 500 microM increased ICI D1694 cell kill. The IC50 and IC90 for MGH-U1 were 0.0037 and 0.0075 microM for 50 microM AZT combined with ICI D1694. The IC50 and IC90 for HCT-8 were 0.0075 and 0.015 microM for 50 microM AZT plus ICI D1694. The incorporation of [3H]AZT into DNA increased with increasing concentrations of ICI D1694. Concentrations producing an IC50 and IC90 of ICI D1694, respectively, increased incorporation of [3H]AZT into DNA by 319 and 569% in MHG-U1, and 243 and 400% in HCT-8 cells. The formation of [3H]AZT-MP paralleled the increase in [3H]AZT incorporated into DNA. AZT, ICI D1694 and the combination of AZT and ICI D1694 caused DNA double-strand breaks, with the combination of these agents being additive. CFU-GM survival, exposed to drug concentrations, as those used in the tumor cell lines, revealed that the therapeutic index was greater for AZT plus ICI D1694 than for ICI D1694 alone. These findings suggest that AZT plus ICI D1694 may increase antitumor effect with minimal myelosuppression. We conclude that AZT increases the cytotoxicity of ICI D1694 with increasing AZT incorporation into DNA.
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PMID:Combination studies with 3'-azido-3'-deoxythymidine (AZT) plus ICI D1694. Cytotoxic and biochemical effects. 826 49

The thymidylate synthase (TS) inhibition rate was measured after tegafur (FT) administration (1.5 g/day, at least 10 days) in 7 sigmoid colon cancer patients. The TS inhibition rate decreased as the interval between the time of the last administration and the time of the tumor resection increased longer. This study provides basic data for considering methods of drug administration and assessment of modification, for example, by leucovorin.
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PMID:Dependence of the thymidylate synthase inhibition rate on the interval after the last administration of tegafur in sigmoid colon cancer patients. 846 74


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