Synthesis, in vitro antiproliferative activity and DNA-interaction of benzimidazoquinazoline derivatives as potential anti-tumor agents.

The synthesis of benzimidazoquinazoline derivatives bearing different alkylamino side chains is reported. All new compounds tested by means of an in vitro assay exhibit antiproliferative activity toward human tumor cell lines. The cytotoxic effect depends on the type of side chain inserted in the planar nucleus and in some cases it is comparable to that of the well-known drug ellipticine. In order to understand the mechanism of action of these compounds, the interaction with DNA has been investigated. Linear flow dichroism measurements allowed us to verify the formation of a molecular complex with DNA and the corresponding geometry of interaction. Intrinsic binding constants have also been evaluated by performing fluorimetric titrations.

Thienocoumarin derivatives: interaction with nucleic acids and synthetic polydeoxyribonucleotides.

This paper reports the photobiological properties of two new thienocoumarins, 4,6,9-trimethyl-2H-thieno[3,2-g]-1-benzopyran-2-one (compound I) and the 6,9-dimethyl-4-methoxymethyl-2H-thieno[3,2-g]-1-benzopyran-2-one (compound II). Cell growth inhibition studies have revealed significant antiproliferative potency on human tumor cell lines. The photoaddition process of these tritium-labeled derivatives was investigated using various nucleic acid structures (calf thymus DNA, bacterial DNA, and synthetic polydeoxyribonucleotides). The results obtained show that both compounds photobind to DNA to a higher extent than 8-MOP, taken as the reference drug. The capacity to form interstrand crosslinks into DNA helix was also evaluated. Interestingly, notwithstanding the lack of cutaneous phototoxicity, II revealed a good ability to induce diadduct formation.

Pharmacological characterization of a new Ca(2+) sensitizer.

The benzimidazole molecule was modified to synthesize a Ca(2+) sensitizer devoid of additional effects associated with Ca(2+) overload. Newly synthesized compounds, termed 1, 2, 3, 4, and 5, were evaluated in spontaneously beating and electrically driven atria from reserpine-treated guinea pigs. Compound 3 resulted as the most effective positive inotropic agent, and experiments were performed to study its mechanism of action. In spontaneously beating atria, the inotropic effect of 3 was concentration-dependent (3.0 microM-0.3 mM). Compound 3 was more potent and more active than the structurally related Ca(2+) sensitizers sulmazole and caffeine, but unlike them it did not increase the heart rate. In electrically driven atria, the inotropic activity of 3 was well preserved and it was not inhibited by propranolol, prazosin, ranitidine, pyrilamine, carbachol, adenosine deaminase, or ruthenium red. At high concentrations (0.1-1.0 mM) 3 inhibited phosphodiesterase-III, whereas it did not affect Na(+)/K(+)-ATPase, sarcolemmal Ca(2+)-ATPase, Na(+)/Ca(2+) exchange carrier, or sarcoplasmic reticulum Ca(2+) pump activities of guinea pig heart. In skinned fibers obtained from guinea pig papillary muscle and skeletal soleus muscle, compound 3 (0.1 mM, 1 mM) shifted the pCa/tension relation curve to the left, with no effect on maximal tension and no signs of toxicity. Compound 3 did not influence the basal or raised tone of guinea pig isolated aorta rings, whose cells do not contain the contractile protein troponin. The present results indicate that the inotropic effect of compound 3 seems to be primarily sustained by sensitization of the contractile proteins to Ca(2+).

Synthesis and biological activity of linear and angular 4-methoxymethylthienocoumarins and 4-acetoxymethylthienocoumarins.

This paper reports the synthesis of 4-methoxymethyl and 4-acetoxymethyl-6,9-dimethyl-2H-thieno[3,2-g]-1-benzopyran-2-one as well as 4-methoxymethyl- and 4-acetoxymethyl-6,9-dimethyl-2H-thieno[2,3-h]-1- benzopyran-2-one. The synthesized derivatives were tested on human cells in UVA irradiation conditions. Skin phototoxicity and cross-link formation in DNA were also studied. Results indicate that the new thienocoumarins have good antiproliferative activity, greater than that of the well-known photochemotherapeutic drug 8-methoxypsoralen, but they are practically devoid of skin photosensitization effects.

Some new methyl-8-methoxypsoralens: synthesis, photobinding to DNA, photobiological properties and molecular modelling.

The tricyclic structure of known natural photochemotherapeutic drugs such as 8-methoxypsoralen and 5-methoxypsoralen is often taken as a model in the search of new photosensitizer agents with less phototoxic and mutagenic effects. This paper describes the synthesis, characterization, photobinding to DNA, photobiological properties and computational chemistry of some 8-methoxypsoralen derivatives bearing two or three methyl groups at the key positions of the two photoactive double bonds. Results showed that photoreactivity and photobiological behaviour depend on the pattern of methyl substitutions. Antiproliferative activity in cell lines shows good correlation with DNA interaction data.

Sequence specificity of psoralen photobinding to DNA: a quantitative approach.

The effects of different DNA sequences on the photoreaction of various furocoumarin derivatives was investigated from a quantitative point of view using a number of self-complementary oligonucleotides. These contained 5'-TA and 5'-AT residues, having various flanking sequences. The furocoumarins included classical bifunctional derivatives, such as 8-methoxy- and 5-methoxypsoralen, as well as monofunctional compounds, such as angelicin and benzopsoralen. Taking into an account the thermodynamic constant for noncovalent binding of each psoralen to each DNA sequence, the rate constants for the photobinding process to each fragment were evaluated. The extent of photoreaction is greatly affected by the DNA sequence examined. While sequences of the type 5'-(GTAC)n are quite reactive towards all furocoumarins, 5'-TATA exhibited a reduced rate of photobinding using monofunctional psoralens. In addition terminal 5'-TA groups were the least reactive with 5- and 8-methoxypsoralen, but not with angelicin or benzopsoralen. Also 5'-AT-containing fragments exhibited remarkably variable responses toward monofunctional or bifunctional psoralen derivatives. As a general trend the photoreactivity rate of the former is less sequence-sensitive, the ratio between maximum and minimum being less than 2 for the examined fragments. The same ratio is about 3.4 for 8-methoxypsoralen and 6.2 for 5-methoxypsoralen. This approach, in combination with footprinting studies, appears to be quite useful for a quantitative investigation of the process of covalent binding of psoralens to specific sites in DNA.

Interaction between second generation anthracyclines and DNA in the nucleosomal structure.

The interaction between three anthracycline antibiotics of second generation (9-deoxydoxorubicin, 9-DAM, 4-demethoxydaunorubicin, 4-DDM, 4'-deoxydoxorubicin, 4'-DAM) and DNA in the nucleosomal structure was investigated using fluorescence and circular dichroism techniques. The thermodynamic parameters of the binding process were obtained at different ionic strength and temperature conditions, thus allowing the calculation of the electrostatic contribution to the free energy and the enthalpy of the process. The same measurements were performed on linear double stranded DNA for comparison. The parent compounds adriamycin and daunomycin were additionally considered. Although the examined drugs greatly vary in biological activity, their binding parameters are only slightly different. Like the parent compounds, 9-DAM, 4-DDM and 4'-DAM exhibit preference for isolated regions of the polynucleotide rather than for nucleosomes. This fact suggests a non-homogeneous distribution of the antibiotics in vivo. The enthalpy values are remarkably lower than the ones characterizing the interaction of adriamycin and daunomycin to DNA. According to CD spectra, all derivatives, but 4-DDM, intercalate into nucleosomal or free DNA in a manner similar to the first generation compounds, namely with the chromophore perpendicular to the hydrogen bonds between the bases. The demethoxy compound, on the other hand, seems to be able to insert its planar moiety in different orientations, which are related to the structure of the nucleic acid being examined. The lack of the methoxy group in the intercalating part of the molecule appears to be responsible for this behaviour. As far as biological activity is concerned, our findings indicate a qualitative correlation between cell cytotoxicity and ability of interaction with nucleosomes at physiological conditions, rather than with free DNA. The modified binding stereochemistry of 4-DDM could play an additive role in modulating the pharmacological effectiveness of the above compounds.

Photobiological activity of 3,4'-dimethyl-8-methoxypsoralen, a linear furocoumarin with unusual DNA-binding properties.

The furocoumarin derivative 3,4'-dimethyl-8-methoxypsoralen (DMe-8-MOP) exhibits remarkable antiproliferative activity, but is devoid of skin phototoxicity. To gain insight into this peculiar behaviour we investigated non-covalent and covalent binding of DMe-8-MOP to calf thymus DNA, along with DNA-synthesis inhibition and mutagenic activity. The non-covalent interaction of DMe-8-MOP with the nucleic acid is quite poor as shown by equilibrium dialysis, spectroscopic, chiroptical and hydrodynamic techniques. However, it exhibits relevant photobinding ability to DNA using both isolated nucleic acid samples and cellular systems. Unlike the large majority of congeners, DMe-8-MOP undergoes predominantly photochemical monoaddition to the double helical polynucleotide. Upon examination of the products obtained by enzymatic hydrolysis of DMe-8-MOP photomodified DNA, the formation of an unusual furan side adduct is proposed, which could account for the peculiar photochemical and photobiological properties of the 3,4'-dimethyl furocoumarin derivative.

New anthracenedione derivatives: interaction with DNA and biological effects.

Two anthracenedione derivatives [1 - (omega - diethylaminopropylamido) - 4 - hydroxy - 9,10 - anthracenedione hydrochloride (I) and 1 - (omega - diethylaminopropylamido) - 2 - methoxy - 4 - hydroxy - 9, 10 - anthracenedione hydrochloride (II)], having an electron-rich planar chromophore and an amino-substituted side chain, have been synthesized. Their binding ability to DNA was investigated by means of spectroscopic, equilibrium dialysis and fluorescence measurements. Their inhibition efficiency on nucleic acid synthesis was also evaluated both in mouse and human cells. Our results indicate that, in comparison with adriamycin, compound I shows a slightly weaker complexation ability to DNA, while compound II interacts with DNA at a substantially lower level. These data match quite well with the biological response on the inhibition of DNA and RNA synthesis exhibited by the above mentioned compounds; in fact compound I is slightly less efficient than adriamycin and about ten times more efficient than compound II. The close relationship between the results of physicochemical and biological studies is discussed.