Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.5.1.3 (dihydrofolate reductase)
5,819 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Methotrexate (MTX) linked to antitumor antibodies inhibits tumor growth better than free MTX, free antibody, or MTX linked to normal rabbit IgG (NRG), in spite of the less effective inhibition of the target enzyme dihydrofolate reductase (DHFR) by conjugated MTX. In addition to the demonstrated higher uptake of MTX linked to antitumor antibodies (compared with the uptake of free MTX or nonspecific IgG conjugates), a contributory factor to the superior tumor inhibitory action of MTX-IgG conjugates may be the prolonged release of active drug from the internalized conjugate. Therefore, we have investigated whether an MTX-IgG conjugate could be hydrolyzed to release free MTX or fully active MTX-containing fragments after incubation with liver homogenates and have characterized the catabolites according to the presence of free MTX and their capacity to inhibit DHFR. Catabolism was optimal at pH 4.6, activated by dithiothreitol, and inhibited by antipain and N-alpha-p-tosyl-L-lysine chloromethyl ketone, thus implicating lysosomal enzymes. Liver homogenates produced an MTX-containing, low-molecular-weight fraction that was isolated by gel filtration. Further purification of this fraction by DEAE-cellulose chromatography gave two MTX-containing peaks, neither of which migrated as free MTX on thin-layer chromatography or inhibited DHFR more effectively than the parent conjugate. However, the presence of amino acid residues in these catabolites could contribute to their observed prolonged intracellular retention and superior antitumor action.
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PMID:Hydrolysis of methotrexate-immunoglobulin conjugates by liver homogenates and characterization of catabolites. 340 49

A new analogue of methotrexate was synthesized from 4-amino-4-deoxy-N10-methylpteroic acid and D,L-homocysteic acid. The product (mAPA-HCysA) was bound tightly to L1210 mouse leukemia dihydrofolate reductase (IC50 = 1 nM), inhibited L1210 cell proliferation in culture (IC50 = 0.3 microM), and prolonged the survival of L1210 leukemic mice (98% increase in lifespan at 120 mg/kg, qdx9). Studies on the interaction of mAPA-HCysA with partially purified mouse liver folyl polyglutamate synthetase revealed that mAPA-HCysA was not a substrate. Hence, the increased dose of mAPA-HCysA required to inhibit tumor growth in vitro and in vivo relative to methotrexate may reflect, in part, the inability of this compound to form non-effluxing polyglutamates. Folyl polyglutamate synthetase was competitively inhibited by mAPA-HCysA (K1 = 190 +/- 70 microM) when folate was the variable substrate. Thus, mAPA-HCysA is the first known compound to inhibit both mammalian dihydrofolate reductase and mammalian folyl polyglutamate synthetase.
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PMID:Methotrexate analogues--XVII. Antitumor activity of 4-amino-4-deoxy-N10-methylpteroyl-D,L-homocysteic acid and its dual inhibition of dihydrofolate reductase and folyl polyglutamate synthetase. 654 90

Analogues of classical antifolates with the 4-aminobenzoyl group replaced by 4-amino-1-naphthoyl were synthesized for study after molecular modeling indicated ample spatial accommodation for the naphthalene ring and even larger groups in models based on reported X-ray crystallographic data describing the binding of methotrexate to human dihydrofolate reductase (DHFR). The side-chain precursors, N-(4-amino- and 4-(methylamino)-1-naphthoyl)-L-glutamic acid diethyl esters, were synthesized, and the 2,4-diamino-substituted heterocyclic groups were attached using several methods. Target compounds included naphthoyl analogues of aminopterin (AMT), methotrexate (MTX), 5-deazaAMT, 5-deazaMTX, 5-methyl-5-deazaAMT, 5-methyl-5-deazaMTX, and 5,8-dideazaAMT. A 5,6,7,8-tetrahydronaphthoyl analogue of 5-deazaAMT was also prepared. None of the naphthoyl analogues showed loss in binding to DHFR compared with the corresponding antifolate bearing the benzoyl group, thus confirming the anticipated bulk tolerance. Only the 5,6,7,8-tetrahydronaphthoyl analogue displayed reduced antifolate effects. Substrate activity toward folylpolyglutamate synthetase was, however, severely compromised. The naphthoyl compounds were transported into L1210 cells 3-6 times more readily than MTX, and despite apparently low levels of intracellular polyglutamylation, each compound was found to be significantly more potent than MTX in inhibiting tumor cell growth in vitro in three lines (L1210, HL60, and S180). The MTX, 5-methyl-5-deazaAMT, and 5-methyl-5-deazaMTX analogues were evaluated in vivo alongside MTX against E0771 mammary adenocarcinoma in mice. All three proved more effective than MTX in retarding the tumor growth. The naphthoyl analogue of 5-deazaAMT strongly inhibited DHFR from Pneumocystis carinii, Toxoplasma gondii, and rat liver giving IC50 (pM) values of 0.53, 2.1, and 1.6 respectively, but this compound did not inhibit in vitro growth of T. gondii, thus indicating lack of transport.
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PMID:Studies on analogues of classical antifolates bearing the naphthoyl group in place of benzoyl in the side chain. 827 97

Twenty-four out of twenty-nine quinoxalines were selected at the National Cancer Institute, Bethesda, Md, USA, for in vitro anticancer screening. Among these, 10 derivatives exhibited high values of percent tumor growth inhibition at a concentration of 10(-4) M in all cancer cell lines. Four of these compounds maintained these values at 10(-5) M, whereas a certain number exhibited significant values of percent inhibition at the most diluted concentrations (10(-8)-10(-6) M). Inhibitory activity against dihydrofolate reductase (DHFR) (bovine and rat liver) was determined for the most active compounds. This test showed that this type of quinoxaline exhibited an appreciable activity in comparison with the previously described aza analogues. In the other test (Lactobacillus casei, thymidylate synthase (TS), human HTS) no or poor activity was detected in both series of compounds.
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PMID:Quinoxaline chemistry. Part 11. 3-Phenyl-2[phenoxy- and phenoxymethyl]-6(7) or 6,8-substituted quinoxalines and N-[4-(6(7)-substituted or 6,8-disubstituted-3-phenylquinoxalin-2-yl)hydroxy or hydroxymethyl] benzoylglutamates. Synthesis and evaluation of in vitro anticancer activity and enzymatic inhibitory activity against dihydrofolate reductase and thymidylate synthase. 983 61

The centromeric region of rat chromosome 2 (2q1) harbors unidentified quantitative trait loci of genes that control tumor growth or development. To improve the mapping of this chromosome region, we microdissected it and generated 10 new microsatellite markers, which we included in the linkage map and/or radiation hybrid map of 2q1, together with other known markers, including four genes: Pcsk1 (protein convertase 1), Dhfr (dihydrofolate reductase), Ndub13 (NADH ubiquinone oxidoreductase subunit b13), and Ccnb1 (cyclin B1). To generate anchor points between the different maps, the gene Ndub13 and the microsatellite markers D2Ulb25 and D2Mit1 were also localized cytogenetically. The radiation map generated in region 2q1 extends its centromeric end of about 150 cR.
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PMID:Localization of new, microdissection- generated, anonymous markers and of the genes Pcsk1, Dhfr, Ndub13, and Ccnb1 to rat chromosome region 2q1. 1077 84

Although the VEGF-Flk-1-pathway has been known as the major driving force of angiogenesis, new evidence has shown that VEGFR-1/Flt-1 plays important roles during the neovascularization under pathological conditions including tumor, atherosclerosis and arthritis. In search of Flt-1 receptor antagonizing peptides, we screened a phage display 12-mer-peptide library with recombinant Flt-1 protein. Seven candidate peptides were identified that specifically bound to VEGF receptor Flt-1, of which peptide F56 (WHSDMEWWYLLG) almost abolished VEGF binding to receptor Flt-1 in vitro. In vivo, F56 fused with DHFR (DHFR-F56) inhibited angiogenesis in a CAM assay. Moreover, DHFR-F56 significantly inhibited the growth of nodules of human gastric cancer cell line MGC-803 in BALB/c nude mice. Histological analyses showed that necrosis of the implanted tumor was markedly enhanced following treatment with DHFR-F56. In the severe combined immunodeficiency disease (SCID) mouse model for studying metastasis of the human breast cancer cell line BICR-H1, synthetic peptide F56 significantly inhibited tumor growth and lung metastases. Taken together, our results have demonstrated that peptide F56, as a Flt-1 receptor antagonist, fulfilled the antiangiogenic and antimetastatic effects by specifically interfering with the interaction between VEGF and receptor Flt-1. Thus, short peptide F56 may have clinical potential in tumor therapy.
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PMID:Suppression of tumor growth and metastasis by a VEGFR-1 antagonizing peptide identified from a phage display library. 1519 67

The p14(ARF) protein, the product of an alternate reading frame of the INK4A/ARF locus on human chromosome 9p21, disrupts the ability of MDM2 to target p53 for proteosomal degradation and causes an increase in steady-state p53 levels, leading to a G(1) and G(2) arrest of cells in the cell cycle. Although much is known about the function of p14(ARF) in the p53 pathway, not as much is known about its function in human tumor growth and chemosensitivity independently of up-regulation of p53 protein levels. To learn more about its effect on cellular proliferation and chemoresistance independent of p53 up-regulation, human HT-1080 fibrosarcoma cells null for p14(ARF) and harboring a defective p53 pathway were stably transfected with p14(ARF) cDNA under the tight control of a doxycycline-inducible promoter. Induction of p14(ARF) caused a decrease in cell proliferation rate and colony formation and a marked decrease in the level of dihydrofolate reductase (DHFR) protein. The effect of p14(ARF) on DHFR protein levels was specific, because thymidylate kinase and thymidylate synthase protein levels were not decreased nor were p53 or p21WAF1 protein levels increased. The decrease in DHFR protein was abolished when the cells were treated with the proteasome inhibitor MG132, demonstrating that p14(ARF) augments proteasomal degradation of the protein. Surprisingly, induction of p14(ARF) increased resistance to the folate antagonists methotrexate, trimetrexate, and raltitrexed. Depletion of thymidine in the medium reversed this resistance, indicating that p14(ARF) induction increases the reliance of these cells on thymidine salvage.
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PMID:p14ARF expression increases dihydrofolate reductase degradation and paradoxically results in resistance to folate antagonists in cells with nonfunctional p53. 1520 49

An SFG-based retroviral bicistronic vector containing a double-mutant dihydrofolate reductase-cytidine deaminase fusion cDNA (F/S DHFR-CD) with IRES-eGFP confers resistance to both methotrexate (MTX) and cytarabine (ara-C). Two weeks after transplantation with marrow transduced with either a fusion or a control gene (eGFP-IRES-NeoR), human lymphoma (SKI-DLCL-1) cells were injected sc into the flanks of nonobese diabetic/severe combined immune deficiency mice. In mock-transplanted mice, maximal tolerated dose (MTD) of posttransplant MTX/ara-C (15/10 mg/kg/day, x3) was unable to control tumor growth. Transfer of the fusion gene allowed doses of MTX/ara-C (25/15 mg/kg/day, x4) twofold higher than the MTD to be tolerated. The tumor burden defined the efficiency of posttransplant chemotherapy; early treatment, 48 h after tumor inoculation, provided tumor-free survival, while starting treatment after having palpable tumor growth (7 days) delayed tumor growth a median time of 28 days. In addition, the early treated group had higher gene expression in peripheral blood and marrow cells than the late treated group (P < 0.05), suggesting that early treatment allowed for enrichment of transduced marrow progenitors. These results encourage clinical studies using this retroviral fusion gene construct.
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PMID:Methotrexate and cytarabine inhibit progression of human lymphoma in NOD/SCID mice carrying a mutant dihydrofolate reductase and cytidine deaminase fusion gene. 1533 57

Methotrexate (MTX), a stoichiometric inhibitor of dihydrofolate reductase, is a chemotherapeutic agent for treating a variety of neoplasms. Impairment of drug import into cells and increase in drug export from cells may render cells resistant to MTX. MTX, when locally administered in a soluble form, is rapidly absorbed through capillaries into the circulatory system, which may also account for therapeutic failure in patients. To retain MTX within tumor cells for longer duration and alter its pharmacokinetic behavior, we proposed a new formulation of MTX bound to the gold nanoparticle (AuNP) that serves as drug carriers. In this study, we developed the MTX-AuNP conjugate and examined its cytotoxic effect in vitro and antitumor effect in vivo. Spectroscopic examinations revealed that MTX can be directly bound onto AuNP via the carboxyl group (-COOH) to form the MTX-AuNP complex and kinetically released from the nanoparticles. The accumulation of MTX is faster and higher in tumor cells treated with MTX-AuNP than that treated with free MTX. Notably, MTX-AuNP shows higher cytotoxic effects on several tumor cell lines compared with an equal dose of free MTX. This can be attributed to the "concentrated effect" of MTX-AuNP. Administration of MTX-AuNP suppresses tumor growth in a mouse ascites model of Lewis lung carcinoma (LL2), whereas an equal dose of free MTX had no antitumor effect. In conclusion, these results suggest that by combining nanomaterials with anticancer drugs MTX-AuNP may be more effective than free MTX for cancer treatment.
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PMID:Methotrexate conjugated to gold nanoparticles inhibits tumor growth in a syngeneic lung tumor model. 1770 53

Human melanoma is a significant clinical problem because it is resistant to treatment by most chemotherapeutic agents, including antifolates. It is therefore a desirable goal to develop a second generation of low-toxicity antifolate drugs to overcome acquired resistance to the prevention and treatment of this skin pathology. In our efforts to improve the stability and bioavailability of green tea polyphenols for cancer therapy, we synthesized a trimethoxy derivative of epicatechin-3-gallate, which showed high antiproliferative and proapoptotic activity against melanoma. This derivative, 3-O-(3,4,5-trimethoxybenzoyl)-(-)-epicatechin (TMECG), is a prodrug that is selectively activated by the specific melanocyte enzyme tyrosinase. Upon activation, TMECG generates a stable quinone methide product that strongly inhibits dihydrofolate reductase in an irreversible manner. The treatment of melanoma cells with TMECG also affected cellular folate transport and the gene expression of DHFR, which supported the antifolate nature of this compound. In addition, its pharmacological efficacy has been confirmed in a mouse melanoma model, in which tumor growth and metastasis were inhibited, significantly enhancing the mean survival of the treated groups. TMECG, therefore, shows a potential for clinical use in melanoma therapy.
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PMID:Melanoma activation of 3-o-(3,4,5-trimethoxybenzoyl)-(-)-epicatechin to a potent irreversible inhibitor of dihydrofolate reductase. 1935 68


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