PubMed Health⌕ Search

SEARCH · PubMed Health

Results for “Reduced Folate Carrier Protein”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 199 records · Page 11Linked to original sources
PubMed2004

A pleiotropic defect reducing drug accumulation in cisplatin-resistant cells.

The resistance of tumors to cisplatin remains a major cause of treatment failure in cancer patients. Multiple, simultaneous alterations are frequently encountered in cancer cells selected for cisplatin resistance. To determine whether the complex phenotype results from many different cellular alterations, single-step variants were isolated based on one-step selection in cisplatin. Reduced drug accumulation is a common feature of cisplatin-resistant (CP-r) cancer cells, which is probably caused by one or more dominant genes. Pulse-chase labeling and pulse-chase biotinylation of cell surface proteins suggest that membrane protein mislocalization occurs in CP-r cells, caused mainly by a defect in plasma membrane protein recycling, manifested also as a defect in acidification of lysosomes. This membrane protein mislocalization is presumed to reduce cell surface expression of a putative cisplatin carrier or carriers. In cells selected in several steps, decreased expression of folate-binding protein and arsenic-binding proteins, and reduced endocytosis were detected in CP-r cells, contributing to the reduced uptake of cisplatin, methotrexate and other related compounds. Multiple mechanisms in CP-r cells keep cytotoxic platinum compounds out of cells through defective expression of cell surface proteins such as transporters and carriers, and decreased expression of proteins involved in endocytosis.

Carboplatin↗
PubMed2004

Aberrant methylation in the promoter region of the reduced folate carrier gene is a potential mechanism of resistance to methotrexate in primary central nervous system lymphomas.

We investigated the prevalence and prognostic role of CpG island methylation of the reduced folate carrier (RFC) gene promoter region in primary central nervous system lymphoma (PCNSL) in immunocompetent patients. Genomic DNA from 40 PCNSL was used for methylation-specific polymerase chain reaction and bisulphite genomic sequencing of the RFC promoter region. Human immunodeficiency virus-negative systemic diffuse large B-cell lymphomas (DLBCL) were used as controls (n = 50). The impact on outcome of RFC promoter methylation was assessed in 37 PCNSL patients treated with high-dose methotrexate (HD-MTX)-based chemotherapy +/- radiotherapy. RFC promoter methylation occurred in 12 of 40 (30%) PCNSL and in four of 50 (8%) DLBCL (P = 0.01). Of 37 PCNSL treated with HD-MTX-based chemotherapy, methylation occurred in nine cases (24%, M-PCNSL), while 28 cases (76%, U-PCNSL) were negative. Three M-PCNSL (33%) and 15 U-PCNSL (54%) achieved complete remission (CR) after primary chemotherapy. Logistic regression confirmed the independent association between CR rate and International Extranodal Lymphoma Study Group score (P = 0.03), RFC promoter methylation (P = 0.07) and use of cytarabine (P = 0.08). The 3-year failure-free survival (FFS) and overall survival for M-PCNSL and U-PCNSL was 0% vs. 31 +/- 9% (P = 0.34) and 0% vs. 31 +/- 9% (P = 0.35) respectively. This is the first study to assess the methylation status of the RFC promoter in human tumour samples. RFC methylation is more common in PCNSL compared with systemic DLBCL, and is associated with a lower CR rate to HD-MTX-based chemotherapy. If confirmed in prospective trials on PCNSL treated with HD-MTX alone, these data may suggest the necessity for alternative strategies in M-PCNSL considering the increased risk of MTX resistance by tumour cells.

Adult↗
PubMed2003

Genomic imbalances associated with methotrexate resistance in human osteosarcoma cell lines detected by comparative genomic hybridization-based techniques.

Methotrexate (MTX) is one of the most important drugs for osteosarcoma (OS) treatment. To identify genetic aberrations associated with the development of MTX resistance in OS cells, in addition to the previously reported expression changes of dihydrofolate reductase (DHFR) and reduced folate carrier (RFC) genes, comparative genomic hybridization (CGH)-based techniques were used. The direct comparison between MTX-resistant variants of U-2OS or Saos-2 human OS cell lines with their respective parental cell lines by CGH on chromosomes revealed that development of MTX resistance was associated with gain of the chromosomal regions 5q12-q15 and 11q14-qter in U-2OS variants, and with gain of 8q22-qter in Saos-2 variants. Further analyses by CGH on microarrays demonstrated a progressively increasing gain of mixed lineage leukemia (MLL) gene (11q23) in U-2OS MTX-resistant variants, which was also confirmed by fluorescence in situ hybridization (FISH), in addition to gain of FGR (1p36), amplification/overexpression of DHFR, and slight decrease of RFC expression. In Saos-2 MTX-resistant variants, gain of MYC (8q24.12-q24.13) was detected, together with a remarkable decrease of RFC expression. Further analyses of DHFR, MLL, MYC, and RFC gene status in four additional human OS cell lines revealed that only gain of DHFR and MLL were associated with an inherent lower sensitivity to MTX. These data demonstrate that genetic analyses with complementary techniques are helpful for the identification of new candidate genes, which might be considered for an early identification of MTX unresponsive tumors.

Antineoplastic Agents↗
PubMed1998

Reduced folate carrier gene expression in childhood acute lymphoblastic leukemia: relationship to immunophenotype and ploidy.

Reduced folate carrier (RFC) transcripts in human leukemias were measured by a competitive PCR assay. Total RNAs were reverse transcribed and amplified in the presence of competitive templates for RFC and beta-actin. RFC transcripts were normalized to transcripts for beta-actin. In a series of K562 sublines, a approximately 30-fold range of RFC transcripts measured by PCR assay closely agreed with results of Northern analysis and varied in proportion to RFC protein on Western blots and [3H]methotrexate transport. RFC transcripts varied over a 88-fold range in 49 specimens from 48 children with acute lymphoblastic leukemia (ALL). Median RFC transcripts were similar for 15 T-cell and 33 B-precursor ALL samples (RFC/beta-actin = 6.13 x 10(-3) and 7.92 x 10(-3), respectively) and for 41 diagnostic (7.20 x 10(-3)) and 8 relapse (5.58 x 10(-3)) samples. Whereas PCR measurements of RFC transcripts approximated changes in methotrexate transport in B-precursor ALL blasts (n = 10), for T-ALL blasts (n = 12) there was no apparent relationship between these parameters. For hyperdiploid B-precursor blasts (n = 11) with greater than 52 chromosomes and three to five copies of chromosome 21, the median RFC transcript level was approximately 3-fold higher than that for diploid B-precursor blasts. RFC transcripts were also elevated for two of three B-precursor specimens with acquired trisomy 21. Our results suggest that RFC gene expression is far more predictive of methotrexate uptake capacity in B-precursor than T-ALL and that increased copies of chromosome 21 in B-precursor ALL blasts are generally associated with increased RFC transcripts. Hence, the good prognosis for children with hyperdiploid B-precursor ALL treated with antimetabolite-based chemotherapy and the high levels of methotrexate and methotrexate polyglutamates accumulated may, in part, reflect elevated RFC gene expression and capacities for methotrexate transport.

Adolescent↗
PubMed2006

5-amino-4-imidazolecarboxamide riboside potentiates both transport of reduced folates and antifolates by the human reduced folate carrier and their subsequent metabolism.

Transport is required before reduced folates and anticancer antifolates [e.g., methotrexate (MTX)] exert their physiologic functions or cytotoxic effects. The folate/antifolate transporter with the widest tissue distribution and greatest activity is the reduced folate carrier (RFC). There is little evidence that RFC-mediated influx is posttranscriptionally regulated. We show that [(3)H]MTX influx in CCRF-CEM human childhood T-leukemia cells is potentiated up to 6-fold by exogenous 5-amino-4-imidazolecarboxamide riboside (AICAr) in a AICAr and MTX concentration-dependent manner. Metabolism to more biologically active polyglutamate forms is also potentiated for MTX and other antifolates. That potentiation of influx by AICAr is mediated by effects on the RFC is supported by analyses +/-AICAr showing (a) similarity and magnitude of kinetic constants for [(3)H]MTX influx; (b) similarity of inhibitory potency of known RFC substrates; (c) lack of potentiation in a CCRF-CEM subline that does not express the RFC; and (d) similarity of time and temperature dependence. Potentiation occurs rapidly and does not require new protein synthesis. Effects of specific inhibitors of folate metabolism and the time and sequence of AICAr incubation with cells suggest that both dihydrofolate reductase inhibition and metabolism of AICAr are essential for potentiation. Acute folate deficiency or incubation of CCRF-CEM with AICAr-related metabolites (e.g., adenosine) does not initiate potentiation. AICAr increases growth inhibitory potency of MTX and aminopterin against CCRF-CEM cells when both AICAr and antifolate are present for the first 24 hours of a 120-hour growth period. AICAr is the first small molecule that regulates RFC activity.

Aminoimidazole Carboxamide↗
PubMed2001

Single nucleotide polymorphisms in the human reduced folate carrier: characterization of a high-frequency G/A variant at position 80 and transport properties of the His(27) and Arg(27) carriers.

The presence of sequence variants in the human reduced folate carrier (hRFC) was assessed in leukemia blasts from children with acute lymphoblastic leukemia (ALL) and in normal peripheral blood specimens. A CATG frame shift insertion at position 191 was detected in 10-60% of hRFC transcripts from 10 of 16 ALL specimens, by RFLP analysis and direct sequencing of hRFC cDNAs. In genomic DNAs prepared from 105 leukemia (n = 54) and non-leukemia (n = 51) specimens, PCR amplifications and direct sequencing of exon 3 identified a high-frequency G to A single nucleotide polymorphism at position 80 that resulted in a change of arginine-27 to histidine-27. The allelic frequencies of G/A80 were nearly identical for the non-leukemia (42.2% CGC and 57.8% CAC) and leukemia (40.7% CGC and 59.3% CAC) genomic DNAs. In cDNAs prepared from 10 of these ALL patients, identical allelic frequencies (40 and 60%, respectively) were recorded. In up to 62 genomic DNAs, hRFC-coding exons 4-7 were PCR-amplified and sequenced. A high-abundance C/T696 polymorphism was detected with nearly identical frequencies for both alleles, and a heterozygous C/A1242 sequence variant was identified in two ALL specimens. Both C/T696 and C/A1242 were phenotypically silent. In transport assays with [(3)H]methotrexate and [(3)H]5-formyl tetrahydrofolate, nearly identical uptake rates were measured for the arginine-27- and histidine-27-hRFC proteins expressed in transport-impaired K562 cells. Although there were no significant differences between the kinetic parameters for methotrexate transport for the hRFC forms, minor (approximately 2-fold) differences were measured in the K(i)s for other substrates including Tomudex, 5,10-dideazatetrahydrofolate, GW1843U89, and 10-ethyl-10-deazaaminopterin and for 5-formyl tetrahydrofolate.

Amino Acid Substitution↗
PubMed2001

Impact of the reduced folate carrier on the accumulation of active thiamin metabolites in murine leukemia cells.

The thiamin transporter encoded by SLC19A2 and the reduced folate carrier (RFC1) share 40% homology at the protein level, but the thiamin transporter does not mediate transport of folates. By using murine leukemia cell lines that express no, normal, or high levels of RFC1, we demonstrate that RFC1 does not mediate thiamin influx. However, high level RFC1 expression substantially reduced accumulation of the active thiamin coenzyme, thiamin pyrophosphate (TPP). This decreased level of TPP, synthesized intracellularly from imported thiamin, resulted from RFC1-mediated efflux of TPP. This conclusion was supported by the following observations. (i) Efflux of intracellular TPP was increased in cells with high expression of RFC1. (ii) Methotrexate inhibits TPP influx. (iii) TPP competitively inhibits methotrexate influx. (iv) Loading cells, which overexpress RFC1 to high levels of methotrexate to inhibit competitively RFC1-mediated TPP efflux, augment TPP accumulation. (v) There was an inverse correlation between thiamin accumulation and RFC1 activity in cells grown at a physiological concentration of thiamin. The modulation of thiamin accumulation by RFC1 in murine leukemia cells suggests that this carrier may play a role in thiamin homeostasis and could serve as a modifying factor in thiamin nutritional deficiency as well as when the high affinity thiamin transporter is mutated.

Animals↗
PubMed2005

[Epidemiological study on reduced folate carrier gene(RFC1 A80G) polymorphism and other risk factors of neural tube defects].

OBJECTIVE: To study the reduced folate carrier gene (RFC1) A80G polymorphism and other factors influence on children with neural tube defects (NTDs) and provide the epidemiological evidence for finding genetic marker of NTDs. METHODS: RFC1(A80G) genotypes were detected using polymerase chain reaction-restriction fragment length polymorphism (RFLP-PCR) for blood DNA of 104 trios with NTDs-affected by child, and the 100 control families without child-affected by any birth defects. We performed the analysis of multifactors logistic regression for RFC1 genotypes and other factors in order to investigate the RFC1 genotype of the nuclear families and maternal periconceptional folic acid supplementation influence on NTDs independently. Transmission/disequilibrium test (TDT) for the RFC1 genotype of NTDs and control pedigree were carried out. RESULTS: The RFC1 G allele frequency of children with NTDs (64.42%) was higher than that of the control children (52.53%), and there was the significant difference between them (chi(2)=5.9198, P<0.05). We observed that the infants of the GG genotype were associated with a 2.56-fold increased risk of NTDs when compared with the AA genotype (95% CI=1.04-6.36), The risk of mothers who did not take folic acid for having an NTDs-affected infants was 7.69 (95% CI=2.86-21.75). There were significant differences between cases and controls in the other risk factors, such as paternal age (> or =30), maternal fever during the early pregnancy, the history of maternal spontaneous abortion. In the logistic regression analysis, of multifactors the three factors, for example, the offspring of the RFC1 GG genotype (OR=2.91, 95% CI=1.35-6.30), maternal periconceptional folic acid supplementation (OR=4.32, 95% CI=1.62-11.55), maternal fever during the early pregnancy, had the statistic significance for the risk of NTDs. There was the evidence of an association between G allele and the risk of the maternal having a child with NTDs (OR=1.56, 95% CI=1.07-2.28) in TDT analysis. CONCLUSION: Our findings indicate that the RFC1 genotype (GG) is a possible susceptible gene marker for an increased NTDs risk in this Chinese population, and there is a potential influence on the risk of NTDs in maternal periconceptional folic acid supplementation, and maternal fever during the early pregnancy.

Adult↗
PubMed2001

Mechanism of folate transport across the human colonic basolateral membrane.

Previous studies from our laboratory have demonstrated the existence of a folate transporter in the human colonic apical membranes. The current studies were undertaken to examine the possible presence and function of a folate carrier in the human colonic basolateral membrane vesicles (BLMV). BLMV were purified from mucosal scrapings of colons of organ donors by a Percoll-density gradient centrifugation technique, and uptake studies were performed using a rapid filtration technique. Our results on [(3)H]Pte-Glu uptake are summarized as follows: 1) uptake was sensitive to osmolarity of the incubation medium; 2) Na(+) removal from the incubation medium did not affect folate uptake into BLMV; 3) uptake was significantly increased with decreasing incubation buffer pH from 8 to 4; 4) uptake demonstrated saturation kinetics with an apparent Michaelis constant of 9.6 +/- 0.48 microM and a maximal velocity of 8.10 +/- 0.36 pmol x mg protein(-1) x 10 s(-1); 5) uptake was markedly inhibited by the structural analog methotrexate (inhibitory constant = 8.28 +/- 1.0 microM); 6) uptake into BLMV demonstrated a trans-stimulation phenomenon; 7) anion exchange inhibitors DIDS and SITS significantly inhibited folate uptake; and 8) uptake was potential-insensitive, as voltage clamping of vesicles or making them inside positive with K(+)/valinomycin failed to influence folate uptake. Western blot analysis using purified human colonic basolateral membrane preparations and specific polyclonal antibodies against the human reduced folate carrier (hRFC) has shown expression of the hRFC protein at this membrane domain. These data demonstrate the existence of a pH-dependent, DIDS-sensitive, electroneutral, carrier-mediated mechanism for folate transport across the human colonic basolateral membranes.

Adult↗
PubMed1998

Analysis of the characteristics of folate binding proteins and its relationship with expression of multidrug resistance P-glycoprotein in myelodysplastic syndromes.

OBJECTIVE: To observe the characteristics of folate binding proteins (FBP) in myelodysplastic syndromes (MDS) and leukemia and to study the clinical significance of reduced folate carrier (RFC) present in MDS and its relationship with multidrug resistance (MDR). METHODS: The features of FBP on bone marrow cells were analyzed using radiolabeled 3H-folic acid (3H-FA) binding membrane proteins and SDS-polyacrylamide gel electrophoresis (SDS-PAGE). In the same time, P-glucoprotein and mRNA of MDR gene were detected using immunocytochemistry and reverse transcription polymerase chain reaction (RT-PCR) respectively in patients with MDS and leukemia. RESULTS: Two kinds of FBP, folate receptor (FR) and reduced folate carrier (RFC), were found on the leukemic cells. The same results were presented on mononuclear cells of bone marrow in 5 out of 14 MDS patients, and MDR positive was seen in 4 patiens of them. In normal control and other 9 cases of MDS FRs were only found on the mononuclear cells of bone marrow. CONCLUSION: Reduced folate carrier, which is present in the leukemic cell, is a product of neoplastic cell. It might reveal preleukmic state and have the same significance with MDR that RFC is found in MDS patients.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1991

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.

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.

Animals↗
PubMed2000

Adaptive regulation of intestinal folate uptake: effect of dietary folate deficiency.

Folate is an essential micronutrient that, in mammals, must be obtained from exogenous sources via intestinal absorption. Previous studies have characterized different aspects of the mechanism of the intestinal folate uptake process. Much less, however, is known about regulation of this process. In this study, we examined the effect of dietary folate deficiency on intestinal folate uptake using the rat as an animal model. The results showed that dietary folate deficiency leads to a significant (P < 0.01) and specific upregulation in the transepithelial transport of folic acid. The upregulation in transepithelial folate transport 1) was found to be due to an induction in carrier-mediated folate uptake across the brush-border membrane (BBM) and was mediated via a significant (P < 0.01) increase in the maximal velocity but not the apparent Michaelis constant of the uptake process, 2) was associated with a marked increase in the steady-state mRNA level of reduced folate carrier-1 and in the level of the expressed protein at the intestinal BBM, and 3) was associated with a marked (>10-fold) increase in the activity of the intestinal BBM form of folate hydrolase. Results of this study demonstrate, for the first time, that dietary folate deficiency leads to a marked upregulation in intestinal folate uptake and in the activity of folate hydrolase. Furthermore, the upregulation in folate uptake is associated with an increase in mRNA and protein levels of folate carrier, suggesting possible involvement of a transcriptional regulatory mechanism(s) in the upregulation.

Adaptation, Physiological↗
PubMed2003

Thymidylate synthase expression in advanced colorectal cancer predicts for response to raltitrexed.

PURPOSE: The purpose of this research was to evaluate the predictive value of expression of thymidylate synthase (TS) and other genes for response to raltitrexed (RTX). EXPERIMENTAL DESIGN: Twenty-five patients with metastatic colorectal cancer received RTX 3 mg/m(2) 3-weekly. Pretreatment tumor biopsies were analyzed for TS, dihydropyrimidine dehydrogenase (DPD), thymidine phosphorylase (TP), folylpolyglutamate synthetase, and reduced folate carrier mRNA expression by real-time reverse transcription-PCR. TS protein expression was evaluated by immunohistochemistry using a polyclonal TS antibody. RESULTS: Twenty patients were evaluable for response and gene expression. Six of 20 (30%) achieved a partial response. Median TS/beta-actin was 5.7 x 10(3) (range, 2.2-42 x 10(3)). Median TS/beta-actin was 3.7 x 10(3) in responding patients and 6.1 x 10(3) in nonresponders (P = 0.048). Five of 6 patients with TS/beta-actin </=4.1 x 10(3) and 1 of 14 with higher values responded (P = 0.002). Overall survival was 21.7 months in patients with TS/beta-actin </=4.1 x 10(3) and 5.7 months in patients with higher values (P = 0.013). No correlations were seen between expression of TP, DPD, reduced folate carrier, or folylpolyglutamate synthetase mRNA and response or survival. Weak TS staining was seen in 10 of 21 tumors evaluable for immunohistochemistry, including 5 responders All 4 of the patients with both weak staining and TS/beta-actin </=4.1 x 10(3) responded. CONCLUSIONS: High TS mRNA expression predicts nonresponse to RTX. By contrast with 5-fluorouracil, high levels of TP and DPD mRNA are not associated with RTX resistance. Limited genomic fingerprinting could optimize single-agent therapy, allowing combinations to be reserved for high TS-expressing patients or for treatment failures, with potential reductions in toxicity and cost.

Antimetabolites, Antineoplastic↗
PubMed2000

Thiamine intestinal transport and related issues: recent aspects.

In the intestinal lumen thiamine is in free form and very low concentrations. Absorption takes place primarily in the proximal part of the small intestine by means of a dual mechanism, which is saturable at low (physiological) concentrations and diffusive at higher. Thiamine undergoes intracellular phosphorylation mainly to thiamine pyrophosphate, while at the serosal side only free thiamine is present. Thiamine uptake is enhanced by thiamine deficiency, and reduced by thyroid hormone and diabetes. The entry of thiamine into the enterocyte, as evaluated in brush border membrane vesicles of rat small intestine in the absence of H+ gradient, is Na+- and biotransformation-independent, completely inhibited by thiamine analogs and reduced by ethanol administration and aging. The transport involves a saturable mechanism at low concentrations of vitamin and simple diffusion at higher. Outwardly oriented H+ gradients enhance thiamine transport, whose saturable component is a Na+-independent electroneutral uphill process utilizing energy supplied by the H+ gradient, and involving a thiamine/ H+ 1:1 stoichiometric exchange. The exit of thiamine from the enterocyte, as evaluated in basolateral membrane vesicles, is Na+-dependent, directly coupled to ATP hydrolysis by Na+-K+-ATPase, and inhibited by thiamine analogs. Transport of thiamine by renal brush border membrane vesicles is similar to the intestinal as far as both H+ gradient influence and specificity are concerned. In the erythrocyte thiamine transport is a Na+-independent, electroneutral process yet with two components: saturable, prevailing at low thiamine concentrations, and diffusive at higher. The saturable (specific) component is missing in patients of the rare disease known as thiamine-responsive megaloblastic anaemia (TRMA), producing a general disturbance of thiamine transport up to thiamine deficiency. The TRMA gene is located in chromosome 1q23.3. Recently, the thiamine transporter has been cloned: it is a protein of 497 amino acid residues with high homology with the reduced-folate transporter.

Absorption↗
PubMed2004

Reduced expression of small GTPases and hypermethylation of the folate binding protein gene in cisplatin-resistant cells.

Reduced accumulation of cisplatin is the most consistent feature seen in cisplatin-resistant (CP-r) cells that are cross-resistant to other cytotoxic compounds, such as methotrexate. In this report, defective uptake of a broad range of compounds, including [(14)C]-carboplatin, [(3)H]MTX, [(3)H]folic acid (FA), [(125)I]epidermal growth factor, (59)Fe, [(3)H]glucose, and [(3)H]proline, as well as (73)As(5+) and (73)As(3+), was detected in CP-r human hepatoma and epidermal carcinoma cells that we have previously shown are defective in fluid-phase endocytosis. Downregulation of several small GTPases, such as rab5, rac1, and rhoA, which regulate endocytosis, was found in CP-r cells. However, expression of an early endosomal protein and clathrin heavy chain was not changed, suggesting that the defective endocytic pathway is clathrin independent. Reduced expression of the cell surface protein, folate-binding protein (FBP), which is a carrier for the uptake of MTX, was also observed in the CP-r cells by confocal immunofluorescence microscopy and Real-Time PCR. Reactivation of the silenced FBP gene in the CP-r cells by a DNA demethylation agent, 2-deoxy-5-aza-cytidine (DAC) demonstrates that hypermethylation occurred in the CP-r cells. The uptake of [(14)C]carboplatin, [(3)H]FA, and [(3)H]MTX increased in an early stage CP-r cell line (KB-CP1) after treatment with DAC. Both a defective endocytic pathway and DNA hypermethylation resulting in the downregulation of small regulatory GTPases and cell surface receptors contribute to the reduced accumulation of a broad range of compounds in CP-r cells.

Antineoplastic Agents↗
PubMed2000

Identification and characterization of the human and mouse SLC19A3 gene: a novel member of the reduced folate family of micronutrient transporter genes.

We report here the isolation, characterization, and chromosomal localization of the genes encoding the human and corresponding murine orthologue of solute carrier family 19A member 3 (SLC19A3). Human SLC19A3 encodes a 496-amino-acid residue protein with a predicted molecular weight of 56 kDa that shares sequence similarity to both SLC19A1 (reduced folate transporter (RFC-1)) and SLC19A2 (high affinity thiamine transporter (THTR-1)). Like the SLC19A1 and SLC19A2 proteins, SLC19A3 contains 12 putative transmembrane domains. The human SLC19A3 gene is widely expressed, with the most abundant expression observed in placenta, kidney, and liver, and has been mapped to chromosome 2q37. The murine SLC19A3 gene maps to central chromosome 1 in the region defined as a seizure susceptibility locus in the DBA/2J mouse strain. This article describes the identification of SLC19A3, a gene encoding a novel solute transporter, and establishes murine SLC19A3 as a candidate gene for seizures in the DBA/2J mouse.

Amino Acid Sequence↗
PubMed1995

Suramin sodium: pronounced effects on methotrexate transport and anti-folate activity in cultured tumor cells.

Suramin is an experimental anti-neoplastic agent which has shown promising activity against prostatic carcinoma and lymphoma in clinical trials. To elucidate its mechanism of action, suramin was examined for an effect on the transport of folate compounds by tumor cells. Influx of the anti-folate methotrexate via the reduced-folate carrier system of CCRF-CEM cells was found to be highly sensitive to inhibition by suramin but not to various other arylsulfonic acids. Inhibition by suramin was competitive, and the inhibition constant Ki was 1.3 microM, a value 3-fold lower than the Kt for half-maximal influx of methotrexate. Folate binding to the membrane-associated folate-binding protein of KB cells was not affected by suramin. Growth studies revealed that the response of human CCRF-CEM, KB, PC-3 and MCF-7 cells to methotrexate was antagonized from 6- to 17-fold by pharmacological levels (10-200 microM) of suramin. Conversely, growth inhibition was additive or synergistic when suramin was combined with metoprine, a lipophilic anti-folate which enters cells by diffusion. Synergism was observed between metoprine and suramin in CCRF-CEM cells, which take up folate exclusively through the reduced-folate carrier (inhibitable by suramin), whereas additivity was observed for KB cells, which rely largely on the folate-binding protein (unaffected by suramin) for folate import. Our results indicate that inhibition of cellular transport of folate compounds may explain part of the anti-neoplastic effects of suramin on tumor cells.

Antineoplastic Agents↗