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Several general and gene- and cell-selective transcription factors are required for specific transcription to occur. Many of them exert their functions through specific contacts either in the promoter region or at distant sequences regulating the initiation. These contacts may be altered by anticancer drugs which form non-covalent complexes with DNA. Covalent modifications of DNA by alkylating agents may prevent transcription factors from recognizing their specific sequences or may constitute multiple "unnatural" binding sites in DNA which attract the factors thus decreasing their availability in the cell. The anticancer drug-transcription factor interplay which is based on specific interactions with DNA may contribute to pharmacological properties of the former and provide a basis for the search for new drugs.

9-Aminoacridine carboxamide derivatives studied here form with DNA intercalative complexes which differ in the kinetics of dissociation. Inhibition of total RNA synthesis catalyzed by phage T7 and Escherichia coli DNA-dependent RNA polymerases correlates with the formation of slowly dissociating acridine-DNA complex of time constant of 0.4-2.3 s. Their effect on RNA synthesis is compared with other ligands which form with DNA stable complexes of different steric properties. T7 RNA polymerase is more sensitive to distamycin A and netropsin than the E. coli enzyme while less sensitive to actinomycin D. Actinomycin induces terminations in the transcript synthesized by T7 RNA polymerase. Despite low dissociation rates of DNA complexes with acridines and pyrrole antibiotics no drug dependent terminations are observed with these ligands.

The mode of action of many anticancer drugs involves DNA interactions. We here examine the ability of actinomycin D to alter the specific binding of transcription factors Spl and NFkappaB to their DNA sequences. Employing an electrophoretic mobility shift assay, it is shown that actinomycin D inhibits complex formation between nuclear proteins present in the extracts from stimulated human umbilical vein endothelial cells and the Sp1-binding site. Actinomycin D is also able to induce disruption of preformed DNA-protein complexes, pointing to the importance of an equilibrium of three components: actinomycin D, protein and DNA for drug action. The effect of actinomycin D is sequence-specific, since no inhibition is observed for interaction of nuclear proteins with the NFkappaB binding site. The results support the view that DNA-binding drugs displaying high sequence-selectivity can exhibit distinct effects on the interaction between DNA and different DNA-binding proteins.

The uptake of six 9-aminoacridinecarboxamide derivatives by L1210 cells in relation to their lipophilicity and cytotoxic activity was studied. The amount of acridines taken up by cells was estimated by fluorimetric measurements. It was found that the uptake efficiency of this class of compounds by cells depends on the size of carboxamide residue as well as on position of the substituent. The increase of size of carboxamide chain resulted in the loss of capability of acridines to penetrate cell membrane. Cytotoxic effects of acridines were well correlated with the level of drugs accumulated by cells, whereas no clear correlation between uptake and lipophilicity was observed. It is concluded that uptake of 9-aminoacridinecarboxamides is the most important factor determining their antiproliferative activity.

The cytotoxic activity, susceptibility to thiol activation and ability of eight 1-nitroacridine derivatives to stabilize the topoisomerase I-DNA cleavable complex, were compared. Among the acridines tested three compounds exhibited high ability to stabilize the cleavable complex. This ability was correlated with susceptibility to thiol activation as well as with cytotoxic activity. Our results suggest that 1-nitroacridine-DNA adducts interfering with topoisomerase I action may contribute to the lethal effects of some 1-nitroacridine derivatives.

Methylthioadenosine (MTA) phosphorylase purified 615-fold from human liver cleaved phosphorolytically nucleoside analogues at the decreasing specific activity: 5'-deoxyadenosine > 5'-iodo-5'-deoxyadenosine > MTA > adenosine > 2-chloroadenosine > 2-chloro-5'-O-methyl-2'-deoxyadenosine > 2-chloro-2'-deoxyadenosine > > 2'-deoxyadenosine. Adenosine and analogues of 5'-deoxyadenosine were strong competitive inhibitors of MTA phosphorolysis catalysed by the human liver enzyme.

Increased catalase activity was observed in the liver microsomal fraction of ethanol-treated rats (10% v/v aqueous ethanol solution per os for 5 weeks). In contrast, cytochrome P-450 concentration and specific activity of NADPH-cytochrome c reductase remained at the same level as in the liver of control rats (drinking water). The ratio of microsomal H2O2-generation to catalase activity was lower in the "ethanol" group than in the control one. This phenomenon seems to be related to the increased contribution of the "peroxidatic" reaction (increased rate of ethanol oxidation). Administration of mesitylene (1,3,5-trimethylbenzene) by gastric tube for 3 days (5 mmoles per kg daily) increased cytochrome P-450 concentration, specific activity of NADPH-cytochrome c reductase and ethanol metabolism.

Adenosine deaminase (ADA) was partially purified 486- and 994-fold from rat liver mitochondria and cytosol, respectively. Relative molecular mass of the enzymes from both fractions was 34,000. Km for adenosine and 2'-deoxy-adenosine were 3.08 x 10(-5) M and 3.03 x 10(-5) M for mitochondrial ADA and 3.12 x 10(-5) M and 2.87 x 10(-5) M for cytosolic ADA. The enzyme from both subcellular fractions had the maximum activity at pH 7.5-8.0, and pI 5.2 and 4.2 for mitochondrial and cytosolic enzyme, respectively. The enzyme was inhibited by erythro-9-(2-hydroxy-3-nonyl)adenine and 2'-deoxycoformycin with Ki 4.4 x 10(-7) M and 3.2 x 10(-7) M for mitochondrial ADA and 4.9 x 10(-7) M 2.8 x 10(-7) M for cytosolic ADA. Among the natural nucleoside and deoxynucleotide derivatives tested, deoxy-GTP and UTP inhibited only cytosolic adenosine deaminase by 60% and 40%, respectively.

New pyrimidine acyclonucleosides, prepared by us, were phosphorylated by nucleoside phosphotransferase from wheat shoots as well as from Enterobacter agglomerans. Conditions and parameters of enzymic phosphorylation were optimized and the results obtained with the two phosphotransferases were compared.

The activity of glycerol ester hydrolase (GEH) from aorta wall, at optimum pH and triacylglycerol substrate concentration (optimal for each substrate) decreased in the following order: C18:1 greater than or equal to C18:2 greater than or equal to C18:0 greater than or equal to C18:3 greater than or equal to C16:0. At optimum pH and the same substrate concentration (1 mM), the activity of GEH from aorta wall decreased in a slightly different order: C18:1 greater than or equal to C18:2 greater than or equal to C18:3 greater than C18:0 greater than C16:0 and that of the enzyme from serum in the order: C18:1 = C18:3 greater than C18:2 greater than or equal to C16:0 greater than or equal to C18:0. These differences in substrate affinity of GEH may influence the metabolism and accumulation of acylalcohols and alcohols in arterial wall and serum.

A series of 9-aminoacridine carboxamide derivatives of systematically varied structure was assayed in an RNA synthesis in vitro system. Escherichia coli DNA-dependent RNA polymerase and DNA derived from phage T7 or calf thymus were used to measure the effect of the drugs on overall RNA and the initiating dinucleotide (pppApU) syntheses. By means of multiple linear regression analysis it was shown that the inhibition of these reactions depends both on the drug equilibrium binding constant and kinetic parameters of dissociation of drug-DNA complexes.