Purine analogue 6-methylmercaptopurine riboside inhibits early and late phases of the angiogenesis process. (1/28)

Angiogenesis has been identified as an important target for antineoplastic therapy. The use of purine analogue antimetabolites in combination chemotherapy of solid tumors has been proposed. To assess the possibility that selected purine analogues may affect tumor neovascularization, 6-methylmercaptopurine riboside (6-MMPR), 6-methylmercaptopurine, 2-aminopurine, and adenosine were evaluated for the capacity to inhibit angiogenesis in vitro and in vivo. 6-MMPR inhibited fibroblast growth factor-2 (FGF2)-induced proliferation and delayed the repair of mechanically wounded monolayer in endothelial GM 7373 cell cultures. 6-MMPR also inhibited the formation of solid sprouts within fibrin gel by FGF2-treated murine brain microvascular endothelial cells and the formation of capillary-like structures on Matrigel by murine aortic endothelial cells transfected with FGF2 cDNA. 6-MMPR affected FGF2-induced intracellular signaling in murine aortic endothelial cells by inhibiting the phosphorylation of extracellular signal-regulated kinase-2. The other molecules were ineffective in all of the assays. In vivo, 6-MMPR inhibited vascularization in the chick embryo chorioallantoic membrane and prevented blood vessel formation induced by human endometrial adenocarcinoma specimens grafted onto the chorioallantoic membrane. Also, topical administration of 6-MMPR caused the regression of newly formed blood vessels in the rabbit cornea. Thus, 6-MMPR specifically inhibits both the early and the late phases of the angiogenesis process in vitro and exerts a potent anti-angiogenic activity in vivo. These results provide a new rationale for the use of selected purine analogues in combination therapy of solid cancer.  (+info)

Stability of ribosilo-6-methylmercaptopurine in aqueous solutions. (2/28)

The kinetics of hydrolysis of ribosilo-6-methylmercaptopurine was studied in aqueous solution at 353 K over a pH range of 0.45-12.13. The decomposition was followed by HPLC method. The reaction of ribosilo-6-methylmercaptopurine hydrolysis includes: the reaction catalysed by hydrogen ions, that catalysed by hydroxide ions and spontaneous hydrolysis under the effect of water.  (+info)

Mechanisms of action of 6-thioguanine, 6-mercaptopurine, and 8-azaguanine. (3/28)

The effects of 6-thioguanine on purine biosynthesis and cell viability have been examined in H.Ep. 2 cells grown in culture. Toxicity is not reversed by aminoimidazolecarboxamide, suggesting that inhibition of purine biosynthesis de novo is not the sole mechanism of toxicity. Also, 6-(methylmercapto)purine ribonucleoside, a potent inhibitor of purine biosynthesis de novo, produces more marked reductions in cellular pools of purines than does 6-thioguanine without killing cells. There is no apparent inhibition by 6-thioguanosine 5'-monophosphate of other enzymes leading to the synthesis of guanosine 5'-triphosphate as determined in whole cells by measurements of radioactive hypoxanthine or guanine incorporation. Inhibition of DNA synthesis by 1 mM thymidine protects cells from 6-mercaptopurine or 6-thioguanine but fails to protect cells from 8-azaguanine toxicity. On the other hand, inhibition of RNA synthesis by 6-azauridine plus deoxycytidine protects cells against 8-azaguanine but does not protect against 6-thioguanine or 6-mercaptopurine toxicity. In agreement with the in vitro data, arabinosylcytosine (a potent inhibitor of DNA synthesis) fails to protect mice against 8-azaguanine but has previously been shown to protect mice from 6-mercaptopurine or 6-thioguanine toxicity. The results support the hypotheses of others that incorporation into DNA (as 6-thioguanine nucleotide) is a mechanism of toxicity for these thiopurines, whereas 8-azaguanine is toxic due to its incorporation into RNA.  (+info)

Thiopurine metabolism and identification of the thiopurine metabolites transported by MRP4 and MRP5 overexpressed in human embryonic kidney cells. (4/28)

Mercaptopurines have been used as anticancer agents for more than 40 years, and most acute lymphoblastic leukemias are treated with 6-mercaptopurine (6MP) or 6-thioguanine (TG). Overexpression of the two related multidrug resistance proteins MRP4 and MRP5 has been shown to confer some resistance against mercaptopurines, which has been attributed to extrusion of mercaptopurine metabolites by these transporters. We have analyzed the mercaptopurine metabolites formed in human embryonic kidney cells and determined which metabolites are extruded by MRP4 and MRP5. Incubation with 6MP led to the formation of thioinosine and thioxanthosine metabolites and we found that thio-IMP was transported by both MRP4 and MRP5; MRP5 showed the highest transport rate. In contrast, only MRP5 transported thioxanthosine monophosphate (tXMP). During incubation with TG, the monophosphorylated form of thioguanosine was transported by both MRP4 and MRP5; the highest transport rate was for MRP4. Similarly, only 6-methyl-thio-IMP was formed during incubation with 6-methyl mercaptopurine riboside. This compound was a substrate for both MRP4 and MRP5; MRP4 showed the highest transport rate. Our results show that all major thiopurine monophosphates important in the efficacy of mercaptopurine treatment are transported by MRP4 and MRP5, although the substrate specificity of the two transporters differs in detail.  (+info)

The synthesis of oligoribonucleotides containing N6-alkyladenosines and 2-methylthio-N6-alkyladenosines via post-synthetic modification of precursor oligomers. (5/28)

The N6-alkyladenosines and 2-methylthio-N6-alkyladenosines are the most common modified adenosine nucleosides and transfer ribonucleic acids (tRNA) are particularly rich in these modified nucleosides. They are present at position 37 of the anticodon arm and the contribution of these hypermodified nucleosides to codon-anticodon interactions, as well as translation, are significant, although not fully understood. Herein we described a new chemical synthesis method of the oligoribonucleotides containing N6-alkyladenosines and 2-methylthio-N6-alkyladenosines via post-synthetic modifications of precursor oligoribonucleotides. To obtain oligoribonucleotides containing N6-alkyladenosines, the precursor oligoribonucleotide carrying 6-methylthiopurine riboside residue was used, whereas for the synthesis of oligoribonucleotides containing 2-methylthio-N6-alkyladenosines the precursor oligoribonucleotide carrying the 2-methylthio-6-chloropurine riboside was applied. Among the modified oligoribonucleotides of different length and secondary structures, there were several containing naturally occurring modified nucleosides such as: N6-isopentenyladenosine (i6A), N6-methyladenosine (m6A), 2-methylthio-N6-isopentenyladenosine (ms2i6A), and 2-methylthio-N6-methyladenosine (ms2m6A), as well as several unnaturally modified adenosine derivatives.  (+info)

Regulation of purine nucleotide synthesis in human B lymphoblasts with both hypoxanthine-guanine phosphoribosyltransferase deficiency and phosphoribosylpyrophosphate synthetase superactivity. (6/28)

Human B lymphoblast lines severely deficient in hypoxanthine-guanine phosphoribosyltransferase (HGPRT) were selected for resistance to 6-thioguanine from cloned normal and phosphoribosylpyrophosphate (PP-Rib-P) synthetase-superactive cell lines and were compared with their respective parental cell lines with regard to growth and PP-Rib-P and purine nucleotide metabolism. During blockade of purine synthesis de novo with 6-methylthioinosine or aminopterin, inhibition of growth of all HGPRT-deficient cell lines was refractory to addition of Ade at concentrations which restored substantial growth to parental cell lines. Ade-resistant inhibition of growth of parental lines by 6-methylthioinosine, however, occurred during Ado deaminase inhibition. Insufficient generation of IMP (and ultimately guanylates) to support growth of lymphoblasts lacking HGPRT activity and blocked in purine synthesis de novo best explained these findings, implying that a major route of interconversion of AMP to IMP involves the reaction sequence: AMP----Ado----Ino----Hyp----IMP. PP-Rib-P generation and purine nucleoside triphosphate pools were unchanged by introduction of HGPRT deficiency into normal lymphoblast lines, in agreement with the view that accelerated purine synthesis de novo in this deficiency results from increased availability of PP-Rib-P for the pathway. Cell lines with dual enzyme defects did not differ from PP-Rib-P synthetase-superactive parental lines in rates of PP-Rib-P and purine synthesis despite 5-6-fold increases in PP-Rib-P concentrations, excretion of nearly 50% of newly synthesized purines, and diminished GTP concentrations. Fixed rates of purine synthesis de novo in PP-Rib-P synthetase-superactive cells appeared to reflect saturation of the rate-limiting amidophosphoribosyltransferase reaction for PP-Rib-P. In combination with accelerated purine excretion, increased channeling of newly formed purines into adenylates, and impaired conversion of AMP to IMP, fixed rates of purine synthesis de novo may condition cell lines with defects in HGPRT and PP-Rib-P synthetase to depletion of GTP with consequent growth retardation.  (+info)

Biochemical modulation of tumor cell energy: regression of advanced spontaneous murine breast tumors with a 5-fluorouracil-containing drug combination. (7/28)

This report describes a highly active chemotherapeutic drug combination, consisting of N-(phosphonacetyl)-L-aspartate plus 6-methylmercaptopurine riboside plus 6-aminonicotinamide plus 5-fluorouracil, in CD8F1 mice bearing spontaneous, autochthonous, breast tumors or first-passage advanced transplants of these spontaneous tumors. The combination and sequence of administration of these drugs were selected on the basis of known potentiating biochemical interactions. High performance liquid chromatography and nuclear magnetic resonance spectroscopy measurements of biochemical changes resulting from treatment with N-(phosphonacetyl)-L-aspartate plus 6-methylmercaptopurine riboside plus 6-aminonicotinamide indicated a severe depletion of cellular energy levels in the treated tumors. 6-Aminonicotinamide produced a severe block of the pentose shunt, and 5-fluorouracil severely inhibited both thymidylate synthase and thymidine kinase in the treated tumors. This quadruple drug combination, administered on a 10-11-day schedule, produced an impressive partial tumor regression rate of 67% of large, spontaneous, autochthonous, murine breast tumors and a tumor regression rate of 74% of first-passage transplants of the spontaneous breast tumors.  (+info)

Plasmodium falciparum purine nucleoside phosphorylase: crystal structures, immucillin inhibitors, and dual catalytic function. (8/28)

Purine nucleoside phosphorylase from Plasmodium falciparum (PfPNP) is an anti-malarial target based on the activity of Immucillins. The crystal structure of PfPNP.Immucillin-H (ImmH).SO(4) reveals a homohexamer with ImmH and SO(4) bound at each catalytic site. A solvent-filled cavity close to the 5'-hydroxyl group of ImmH suggested that PfPNP can accept additional functional groups at the 5'-carbon. Assays established 5'-methylthioinosine (MTI) as a substrate for PfPNP. MTI is not found in human metabolism. These properties of PfPNP suggest unusual purine pathways in P. falciparum and provide structural and mechanistic foundations for the design of malaria-specific transition state analogue inhibitors. 5'-Methylthio-Immucillin-H (MT-ImmH) was designed to resemble the transition state of PfPNP and binds to PfPNP and human-PNP with K(d) values of 2.7 and 303 nm, respectively, to give a discrimination factor of 112. MT-ImmH is the first inhibitor that favors PfPNP inhibition. The structure of PfPNP.MT-ImmH.SO(4) shows that the hydrophobic methylthio group inserts into a hydrophobic region adjacent to the more hydrophilic 5'-hydroxyl binding site of ImmH. The catalytic features of PfPNP indicate a dual cellular function in purine salvage and polyamine metabolism. Combined metabolic functions in a single enzyme strengthen the rationale for targeting PfPNP in anti-malarial action.  (+info)