Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0023418 (leukemia)
 93,477 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

5,10-Dideazatetrahydrofolic acid (DDATHF) is a potent inhibitor of glycinamide ribonucleotide transformylase, one of the folate-dependent key enzymes in de novo purine biosynthesis. The present report demonstrates that multiple membrane-transport routes may be involved in the cellular uptake of DDATHF. These routes include the classic reduced folate carrier and a membrane-associated folate-binding protein (mFBP). The role of an mFBP in the uptake of DDATHF was suggested from observations that (a) the mFBP showed a very high binding affinity for DDATHF, (b) murine and human leukemia cells expressing an mFBP were highly sensitive to growth inhibition by DDATHF, and (c) protection against this growth inhibition could be achieved using folic acid rather than reduced folate compounds.
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PMID:Growth-inhibitory effects of 5,10-dideazatetrahydrofolic acid on variant murine L1210 and human CCRF-CEM leukemia cells with different membrane-transport characteristics for (anti)folate compounds. 206 81

N10-Propargyl-5,8-dideazafolic acid (CB3717) and 2-desamino-2-methyl-N10-propargyl-5,8-dideazafolic acid (ICI-198,583) are potent folate-based inhibitors of thymidylate synthase. We studied the membrane transport and the growth-inhibitory effects of the two thymidylate synthase inhibitors on human CCRF-CEM leukemia cells with different transport properties for folic acid, reduced folates, and methotrexate (MTX). Membrane transport of [3H]ICI-198,583 can proceed via the high affinity/low capacity reduced folate carrier as supported by findings that (a) uptake of [3H]ICI-198,583 was significantly impaired in CEM cells which have a transport defect for MTX, (b) variants of CEM cells which overproduce the reduced folate carrier system showed a concomitant increase in the uptake of [3H]ICI-198,583 as for [3H]MTX, (c) MTX inhibited transport of [3H]ICI-198,583, and (d) uptake of [3H]ICI-198,583 was inhibited after treatment of CEM cells with an N-hydroxysuccinimide ester of MTX, which is a potent inhibitor of MTX transport. However, a membrane-associated folate-binding protein (FBP) offers another route for entry of CB3717 and ICI-198,583. CEM-FBP cells that have an elevated amount of FBP and do not have a functional reduced folate carrier were 640- and 61-fold more sensitive to CB3717 and ICI-198,583, respectively, compared to control CEM cells expressing the reduced folate/MTX carrier. This high sensitivity was related to a high affinity of the FBP for CB3717 and ICI-198,583 (Kd 2-3 nM), which is only 3-fold lower than for folic acid (Kd 1 nM) but significantly higher than for MTX (Kd 100 nM). Furthermore, after incubation of CEM-FBP cells for 24 h at 10 nM [3H]ICI-198,583, the high affinity binding of the FBP for ICI-198,583 allowed a 600-fold concentrative uptake of [3H]ICI-198,583 and its conversion to polyglutamate forms. These results indicate that multiple folate transport systems may be involved in the uptake of folate-based thymidylate synthase inhibitors.
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PMID:Multiple membrane transport systems for the uptake of folate-based thymidylate synthase inhibitors. 225 2

Cl-920 is a structurally novel antitumor antibiotic which has activity against a wide spectrum of tumor cells in vitro and is curative in L1210 leukemia in vivo. Several lines of evidence indicate that this drug penetrates L1210 cells via the reduced folate carrier system. Reduced folates (100 microM) including leucovorin and 5-methyltetrahydrofolate completely protected L1210 cells from growth inhibition by Cl-920. Protective effects were not observed, however, with folic acid, a compound which is transported by a process distinct from that for reduced folates. Cl-920 was a potent inhibitor of methotrexate influx exhibiting a mixture of competitive and noncompetitive inhibition and having a Ki (slope) of 30.0 microM and a Ki (intercept) of 58.8 microM. The inhibition appeared to be irreversible since, after cells were preincubated with drug, the inhibitory effects persisted after cells were washed in drug-free media. The irreversibility could be eliminated, however, by dithiothreitol, suggesting that Cl-920 may interact with a thiol which is essential to this transport system. Cells made 71-fold resistant to Cl-920 by continuous exposure to increasing concentrations of this drug were 245-fold cross-resistant to methotrexate but were collaterally sensitive to the lipophilic antifolate trimetrexate and contained normal levels of dihydrofolate reductase. This mutant cell line had a severely impaired reduced folate carrier system exhibiting methotrexate influx rates of less than 1% of control cells. Finally, inhibition of methotrexate influx by a number of Cl-920 analogues showed that the intact lactone ring and the presence of the phosphate ester were required for maximum interaction with the carrier system and that the degree of inhibition correlated with relative antitumor potency. These observations are compatible with the concept that Cl-920 utilizes the folate carrier system and could be of fundamental importance for understanding the cytotoxicity and selectivity of Cl-920.
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PMID:Transport of the antitumor antibiotic Cl-920 into L1210 leukemia cells by the reduced folate carrier system. 654 36

The unnatural d diastereoisomer at carbon 6 of 5-methyltetrahydrofolate was only slightly less effective than the natural 1 diastereoisomer as a competitive inhibitor of the carrier-mediated membrane transport of [3H]methotrexate into L1210 murine leukemia cells. The apparent Ki for a mixture containing equal amounts of both natural and unnatural diastereoisomers was not significantly different from that found for the unnatural form. These results show that the reduced folate carrier system in these cells has a strong affinity for the unnatural stereoisomer, a finding in contrast to that obtained with the corresponding diastereoisomer of 5-formyltetrahydrofolate.
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PMID:Further studies stereospecificity at carbon 6 for membrane transport of tetrahydrofolates. Diastereoisomers of 5-methyltetrahydrofolates as competitive inhibitors of transport of methotrexate in L1210 cells. 709 44

This laboratory previously described an L1210 murine leukemia cell line with a functional defect in the reduced folate carrier and increased expression of folate receptor-beta (F2-MTXrA). This cell line was used to characterize methotrexate (MTX) influx mediated by folate receptor-beta and to compare this with influx mediated by the reduced folate carrier in L1210 parental cells. Influx of 0.2 microM MTX in F2-MTXrA cells was one-third that of L1210 cells and was abolished by very low concentrations of folic acid. Kinetic analysis revealed that MTX transport mediated by folate receptor-beta exhibited an influx kappa t one-third, and an influx Vmax one-fourth, that of the reduced folate carrier. Metabolic inhibitors markedly suppressed influx in F2-MTXrA cells but had no effect on MTX influx in L1210 cells. MTX influx in both cell lines was inhibited by the organic anions probenecid, sulfobromophthalein, and CI-920, but to a lesser extent in F2-MTXrA cells. The inhibitory effects of these anions on transport in F2-MTXrA cells could be attributed to their inhibition of MTX binding to the folate receptor. Although MTX influx in both cell lines was not sodium dependent, removal of extracellular chloride increased influx 2-fold in L1210 cells while markedly inhibiting influx in F2-MTXrA cells. Substitution of Cl- with isethionate or NO3- partially restored influx in the latter cells, whereas SO4(2-) was inhibitory. Anions enhanced MTX binding to folate receptor-beta with isethionate > SO4(2-) > Cl-. Decreasing the buffer pH to 6.2 produced a 69% reduction, and a 260% increase, in MTX influx in L1210 cells and F2-MTXrA cells, respectively. The data indicate that folate receptor-beta-mediated MTX influx has properties fundamentally different from transport mediated by the reduced folate carrier in terms of energy, ion, and pH dependence. There was no evidence indicating that these processes are functionally linked.
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PMID:Comparison of transport properties of the reduced folate carrier and folate receptor in murine L1210 leukemia cells. 748 46

This laboratory previously described an L1210 leukemia cell line (MTXrA) selected for resistance to methotrexate by virtue of impaired transport due to a functional defect in the translocation process. We now report on the sequence analysis of cDNAs encoding the reduced folate carrier from this line and identify a single mutation that results in the substitution of a proline for an alanine in a highly conserved transmembrane region of the protein. Transfection of the parental reduced folate carrier into MTXrA cells resulted in a cell line which exhibited a complete restoration of methotrexate uptake and an enhanced sensitivity to methotrexate. Northern analysis and specific [3H]MTX cell surface binding indicated that expression of the reduced folate carrier was elevated approximately 5-fold in the transfectant compared to parental and MTXrA cells. The MTX influx properties of the transfectant cell line were identical to those of the well characterized reduced folate carrier from parental L1210 cells in terms of: 1) patterns of sensitivity to competing folates, 2) sensitivity to the organic anion sulfobromophthalein, 3) lack of energy dependence, and 4) capacity for trans-stimulation. We also provide new data which suggests that the nucleotide sequence 5' of the predicted ATG initiation codon may encode additional protein information in the form of a leader sequence. Finally, we demonstrate that the MTXrA line has both the mutant and the parental reduced folate carrier alleles but that expression appears to be restricted to the mutant allele. Thus, the methotrexate transport phenotype and resultant drug resistance in this cell line result from genetic/regulatory events at both alleles.
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PMID:Characterization of a mutation in the reduced folate carrier in a transport defective L1210 murine leukemia cell line. 755 35

Based on differential levels of membrane-associated folate binding protein (mFBP) expression, murine L1210 leukemia, human KB epidermoid carcinoma, and human IGROV1 ovarian carcinoma cells maintained under low (physiological) folate conditions (2 nM folinic acid) were used as model systems to investigate the potential role of mFBP in antifolate transport. In addition, L1210 parental cells were compared to a subline, L1210A, expressing high levels of mFBP and defective reduced folate carrier. Antifolates for which KB-derived mFBP has high affinity (5, 10-dideazatetrahydrofolic acid [DDATHF] and homo-DDATHF [0.24 and 0.78 respectively relative to folic acid]) and low affinity (methotrexate [0.002]) were chosen for this study. Protection against DDATHF/homo-DDATHF induced cytotoxicity was achieved preferentially by folic acid compared to folinic acid in IGROV1 and L1210A cells. In IGROV1 cells, cytotoxicity IC50s were increased 18- and 5.5-fold for DDATHF and homo-DDATHF respectively by 20 nM folic acid. Moreover, greater protection was observed in L1210A cells, where IC50s were increased 354- and 80-fold for these same compounds by 20 nM folic acid. Similar protection was not observed in KB cells, suggesting that KB mFBP was not functional in DDATHF transport. Although mFBP expression may be an important determinant in the cytotoxicity of antifolates for certain tumor cells, our data demonstrate a lack of correlation between levels of mFBP and function of mFBP for DDATHF transport in the models studied.
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PMID:Role of membrane-associated folate binding protein in the cytotoxicity of antifolates in KB, IGROV1, and L1210A cells. 757 32

L1210 leukemia cells transport reduced folates and methotrexate via a well defined reduced folate carrier system and, in the absence of low folate selective pressure, do not express an alternate endocytotic route mediated by cell surface folate receptors. This laboratory previously described an L1210 leukemia cell line, MTXrA, with acquired resistance to methotrexate (MTX) due to the loss of mobility of the reduced folate carrier. We now report on the transfection of MTXrA with a cDNA encoding the murine homolog of the human folate receptor isoform of KB cells to produce MTXrA-TF1, which constitutively expresses high levels of FR-alpha. MTXrA-TF1 and L1210 cells were utilized to compare transport of methotrexate mediated by FR-alpha and the reduced folate carrier, respectively. Methotrexate influx in the two lines was similar when the extracellular level was 0.1 microM, but as the methotrexate concentration increased, influx via the reduced folate carrier increased in comparison to influx mediated by FR-alpha. Transport kinetics indicated both a approximately 20-fold lower influx Kb and Vmax for MTXrA-TF1 as compared to L1210 cells. The two cell lines exhibited distinct influx properties. Methotrexate influx in MTXrA-TF1 was markedly inhibited by 50 nM folic acid and metabolic poisons. In L1210 cells, 1.0 microM folic acid did not affect MTX influx, and metabolic poisons either had no effect on or increased methotrexate influx. Removal of extracellular chloride markedly inhibited transport in MTXrA-TF1 but stimulated influx in L1210 cells. When the pH was decreased to 6.2, methotrexate influx was not altered in MTXrA-TF1 but was reduced in L1210 cells. Probenecid and sulfobromophthalein inhibit methotrexate influx in both L1210 and MTXrA-TF1 cell lines; however, inhibition in MTXrA-TF1 could be accounted for on the basis of inhibition of methotrexate binding to FR-alpha. The data indicate that the reduced folate carrier and FR-alpha function independently and exhibit distinct properties. FR-alpha expressed at sufficient levels can mediate influx of MTX and folates into cells at rates comparable to the reduced folate carrier and hence has pharmacologic and physiologic importance.
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PMID:Distinguishing between folate receptor-alpha-mediated transport and reduced folate carrier-mediated transport in L1210 leukemia cells. 771 75

This laboratory previously described an L1210 leukemia cell line (MTXrA) selected for resistance to methotrexate by virtue of impaired transport. In this line, the reduced folate carrier had unchanged affinity for methotrexate, was present at the cell surface in usual quantity, but did not deliver drug into the cell, indicative of a functional defect in the translocation process. In this study, we further characterize this cell line along with a subline (F2-MTXrA) selected for growth in low levels of folic acid. This subline demonstrates continued high resistance to methotrexate and very low influx of [3H]methotrexate and 5-[3H]formyltetrahydrofolate, indicating the persistence of the defect in the reduced folate carrier. Both MTXrA and F2-MTXrA are shown to overexpress FBP2, the murine homolog of a folate-binding protein initially isolated from human placenta. Compared with parent L1210 cells, Northern analysis revealed FBP2 expression to be elevated 40-fold in the MTXrA line and 500-fold in F2-MTXrA. The large increase in FBP2 expression in the F2-MTXrA line correlates with a 10-fold increase in [3H]folic acid membrane surface binding and a 1000-fold decrease in the folic acid growth requirement compared with parental L1210 cells. Also, there are 20- and 500-fold decreases in the 5-formyltetrahydrofolate growth requirement compared with parent L1210 and MTXrA cells, respectively. Finally, the genomic organization of the FBP2 locus is presented. The results of Northern analyses using probes specific to FBP2 5'-untranslated sequences or to a splice junction within this region suggest that the up-regulated FBP2-specific message in F2-MTXrA utilizes 5'-noncoding sequences distinct from those used in the message encoded in L1210 cell lines with low level FBP2 expression. The MTXrA cells provide an example of a line selected for primary resistance to methotrexate that also exhibits concomitant increased expression of a folate-binding protein. Further overexpression of this folate-binding protein (which has homology to that initially identified in placenta) provides cells with the ability to meet cellular folate needs in a folate-deprived environment.
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PMID:Increased expression and genomic organization of a folate-binding protein homologous to the human placental isoform in L1210 murine leukemia cell lines with a defective reduced folate carrier. 830 91

The biological activity and cellular metabolism of ZD1694, a novel folate-based thymidylate synthase (TS) inhibitor, were analyzed in a human leukemia cell line, MOLT-3, and its antifolate-resistant sublines with different mechanisms of resistance to methotrexate (MTX), trimetrexate (TMQ) and N10-propargyl-5,8-dideazafolic acid (CB3717). MOLT-3/CB3717(40), which was selected for CB3717 resistance, demonstrated impaired membrane drug transport via reduced folate carrier (RFC) and lower accumulation of [3H]ZD1694-polyglutamates in the cells with a shift in the polyglutamate distribution profile to shorter chain length polyglutamates, indicating an alteration in polyglutamation capacity in this subline. Impaired RFC and reduced rate of polyglutamation could explain the cross-resistance (12-fold) of this subline to ZD1694. On the other hand, there was little or no cross-resistance to this drug in a subline (MOLT-3/TMQ800) reportedly resistant to TMQ through impaired membrane transport for TMQ and an increase in dihydrofolate reductase (DHFR) activity. Total amount of ZD1694 polyglutamated to a level higher than diglutamate was approximately 1.7-fold higher in the TMQ-resistant cells than that in the parent cells, but a low degree of increase in TS activity in the cells counteracted the supposed increase in sensitivity to ZD1694. MOLT-3/TMQ800-MTX10000 cells, which were established by sequential exposure of the TMQ-resistant cells to MTX and were previously shown to amplify mutated DHFR with low affinity for MTX, showed a decreased accumulation of polyglutamated ZD1694 as compared with the parent line and this was consistent with cross-resistance to ZD1694 in this subline. Overproduction of variant DHFR scarcely influenced the sensitivity to this drug. These results indicate that ZD1694 could overcome antifolate resistance through a mechanism such as amplified DHFR activity, and the biological activity of this drug against the cells paralleled the amount of polyglutamated drug inside the cells. Determination of polyglutamation capacity in tumor cells may allow prediction of sensitivity to this drug.
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PMID:Biological activity and intracellular metabolism of ZD1694 in human leukemia cell lines with different resistance mechanisms to antifolate drugs. 869 29


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