a Abbreviations: CQ = chloroquine; FACS = Fluorescence-activated cell sorting; FPIX = ferriprotoporphyrin IX; HEPES = 4-(2-hydroxyethyl)-1-piperazinethanesulfonic acid; MEM = minimum essential medium; MOA = mechanism of action; µCi = microCurie; Pip = piperidine; SI = selectivity index.

In the present work the decomposition of the total Gibbs free energy of ligand-DNA binding onto various physical terms was accomplished for the group of nine DNA minor groove binders (MGB ligands) differing in both structure and charge state. The decomposition protocol includes the analysis of the most complete set of physical factors known to contribute to the complexation process, viz. the net change in the number of degrees of freedom (translational, rotational, vibrations of the chemical bonds and vibrations of the ligand as a whole within the binding site), the conformational entropy, van der Waals, electrostatic and hydrophobic interactions, the polyelectrolyte contribution and the net effect of changes in the number of hydrogen bonds. All of these processes are further decomposed into the interaction with the solvent and the interaction of the ligand with DNA. The principal outcome of the decomposition is the possibility of performing a comparative analysis of the energetic contribution of various physical terms and provide an answer to the question concerning what physical factors stabilize or destabilize the complexes of MGB ligands with DNA.

In this paper we report the synthesis of three families of new amidine-based aromatic derivatives as potential DNA minor groove binding agents for the treatment of cancer. The preparation of mono-guanidine, mono-2-aminoimidazoline and asymmetric diphenyl guanidine/2-aminoimidazoline derivatives (compounds 1a,b,c to 8a,b,c) is presented. The affinity of these substrates and of a family of mono-and bis-isoureas (previously prepared in Rozas' laboratory) for DNA was evaluated by means of DNA thermal denaturation measurements. In particular, compounds 2c, 5c, 6c, 7c, and 8c were found to bind strongly both to natural DNA and to Adenine-Thymine oligonucleotides, showing a preference for the Adenine-Thymine base pairs sequences.Abbreviations: HB, hydrogen bond; ΔT m , increment in DNA denaturation temperature; AT, Adenine-Thymine pairs; MES, 2-(N-morpholino)ethanesulfonic acid; P/D, ratio between base pairs and ligand (drug);

The efficient preparation and pharmacological characterization of different families of (bis)guanidine and (bis)2-aminoimidazoline derivatives (“twin” and “half” molecules) as potential α2-adrenoceptor antagonists for the treatment of depression is presented. The affinity toward the α2-adrenoceptor of all the compounds prepared was measured in vitro in human brain tissue. Additionally, the activity as agonist or antagonist of those compounds with a pK i larger than 7 was determined in functional [35S]GTPγS binding assays in human brain tissue. Finally, the activity of the most promising compounds was confirmed by means of in vivo microdialysis experiments in rats. Compounds 1, 2b, 3b, 12b, 13b, 17b, 18b, 22b, 25b, 26b, 28b, and 30 showed a good affinity toward the α2-ARs. In general, the 2-aminoimidazoline derivatives displayed higher affinities than their guanidine analogues. Finally and most importantly, compounds 18b and 26b showed antagonistic properties over α2-ARs not only in vitro [35S]GTPγS binding but also in vivo microdialysis experiments. Moreover, both compounds have shown to be able to cross the blood−brain barrier and, therefore, they can be considered as potential antidepressants.

The preparation of a number of (bis)guanidine and (bis)2-aminoimidazoline derivatives as potential alpha 2-adrenoceptor antagonists for the treatment of depression is presented. Human brain tissue was used to measure their affinity toward the alpha 2-adrenoceptors in vitro. Compounds 6b, 8b, 9b, 10b, 15b, 17b, 18b, 20b, and 21b displayed a good affinity (pKi > 7) and were evaluated in in vitro functional [(35)S]GTPgammaS binding assays in human prefrontal cortex to determine their agonistic or antagonistic activity. Among these compounds, 17b and 20b showed the expected behavior for an antagonist and were subject to in vivo microdialysis experiments in rats. Significantly, these experiments confirmed the antagonistic properties of 17b and 20b, and therefore both compounds can be considered as potential antidepressants.

Biophysical studies have been carried out on a family of asymmetric guanidinium-based diaromatic derivatives to assess their potential as DNA minor groove binding agents. To experimentally assess the binding of these compounds to DNA, solution phase biophysical studies have been performed. Thus, surface plasmon resonance, UV-visible spectroscopy and circular and linear dichroism have been utilized to evaluate binding constants, stoichiometry and mode of binding. In addition, the thermodynamics of the binding process have been determined by using isothermal titration calorimetry. These results show significant DNA binding affinity that correlates with the expected 1 : 1 binding ratio usually observed for minor groove binders. Moreover, a simple computational approach has been devised to assess the potential as DNA binders of this family of compounds.

In this paper we report the design and synthesis of a new family of asymmetric peptide linked diaromatic dications as potent DNA minor groove binders. These peptide-linked compounds, with a linear core, displayed a much larger affinity than other guanidinium-like derivatives from the same series with curved cores. As a first screening, the DNA affinity of these structures was evaluated by means of thermal denaturation experiments, finding that the nature of the cation (guanidinium vs 2-aminoimidazolinium) significantly influenced the binding strength. Their binding affinity was assessed by implementing further biophysical measurements such as surface plasmon resonance and circular dichroism. In particular, it was observed that compounds 6, 7, and 8 displayed both a strong binding affinity and significant selectivity for AT oligonucleotides. In addition, the thermodynamics of their binding was evaluated using isothermal titration calorimetry, indicating that the binding is derived from favorable enthalpic and entropic contributions.

In this paper, we report the synthesis of three new 2-aminoimidazoline (compounds 4b, 5b, and 6b) and three new guanidine derivatives (compounds 7b, 8b, and 9b) as potential alpha(2)-adrenoceptor antagonists for the treatment of depression. Their pharmacological profile was evaluated in vitro in human brain tissue and compared to the potential antidepressant 1 and the agonists 2 and 3. All new substrates were evaluated by in vitro functional [(35)S]GTPgammaS binding assays in human prefrontal cortex to determine their agonistic or antagonistic activity. Compound 8b was found to be an antagonist in vitro and was subjected to in vivo microdialysis experiments in rats. Moreover, a new synthesis of the precursor amines for compounds 4b-9b is presented.