A combined ligand- and structure-based approach for the identification of rilmenidine-derived compounds which synergize the antitumor effects of doxorubicin.

The clonidine-like central antihypertensive agent rilmenidine, which has high affinity for I1-type imidazoline receptors (I1-IR) was recently found to have cytotoxic effects on cultured cancer cell lines. However, due to its pharmacological effects resulting also from α2-adrenoceptor activation, rilmenidine cannot be considered a suitable anticancer drug candidate. Here, we report the identification of novel rilmenidine-derived compounds with anticancer potential and devoid of α2-adrenoceptor effects by means of ligand- and structure-based drug design approaches. Starting from a large virtual library, eleven compounds were selected, synthesized and submitted to biological evaluation. The most active compound 5 exhibited a cytotoxic profile similar to that of rilmenidine, but without appreciable affinity to α2-adrenoceptors. In addition, compound 5 significantly enhanced the apoptotic response to doxorubicin, and may thus represent an important tool for the development of better adjuvant chemotherapeutic strategies for doxorubicin-insensitive cancers.

Transfer of SAR information from hypotensive indazole to indole derivatives acting at α-adrenergic receptors: In vitro and in vivo studies.

In a search for novel antihypertensive drugs we applied scaffold hopping from the previously described α1-adrenergic receptor antagonists, 1-[(imidazolin-2-yl)methyl]indazoles. The aim was to investigate whether the α-adrenergic properties of the indazole core were transferable to the indole core. The newly obtained 1-[(imidazolin-2-yl)methyl]indole analogues were screened in vitro for their binding affinities for α1-and α2-adrenoceptors, which allowed the identification of the target-based SAR transfer (T_SAR transfer) as well as structure-based SAR transfer (S_SAR transfer) events. However, when screened in vivo with use of anaesthetized male Wistar rats, the new indole ligands showed a different hemodynamic profile than expected. Instead of the immediate hypotensive effect characteristic of peripheral vasodilatator α1 blockers, a biphasic effect was observed, reminiscent of clonidine-like centrally acting antihypertensive agents. This was supported by subsequent in vitro functional studies in [(35)S]GTPγS binding assay, where the indole analogues displayed partial agonist properties at α2-adrenergic receptors. Since no correlation was found between the in vitro binding to α-adrenoceptors and the in vivo hemodynamic effects of the two series of indazole and indole bioisosteric compounds, in a search for new imidazoline-containing adrenergic drugs, the structure-based SAR transfer information obtained from in vitro binding studies should be treated with caution.

Fluorinated analogues of marsanidine, a highly α2-AR/imidazoline I1 binding site-selective hypotensive agent. Synthesis and biological activities.

The aim of these studies was to establish the influence of fluorination of the indazole ring on the pharmacological properties of two selective α2-adrenoceptor (α2-AR) agonists: 1-[(imidazolidin-2-yl)imino]-1H-indazole (marsanidine, A) and its methylene analogue 1-[(4,5-dihydro-1H-imidazol-2-yl)methyl]-1H-indazole (B). Introduction of fluorine into the indazole ring of A and B reduced both binding affinity and α2-AR/I1 imidazoline binding site selectivity. The most α2-AR-selective ligands were 6-fluoro-1-[(imidazolidin-2-yl)imino]-1H-indazole (6c) and 7-fluoro-1-[(imidazolidin-2-yl)imino]-1H-indazole (6d). The in vivo cardiovascular properties of fluorinated derivatives of A and B revealed that in both cases the C-7 fluorination leads to compounds with the highest hypotensive and bradycardic activities. The α2-AR partial agonist 6c was prepared as a potential lead compound for development of a radiotracer for PET imaging of brain α2-ARs.

Synthesis and biological activities of 2-[(heteroaryl)methyl]imidazolines.

A series of 2-[(heteroaryl)methyl]imidazolines was synthesized and tested for their activities at α(1)- and α(2)-adrenoceptors and imidazoline I(1) and I(2) receptors. The most active 2-[(indazol-1-yl)methyl]imidazolines showed high or moderate affinities for α(1)- and α(2)-adrenoceptors. However, their intrinsic activities at α(2A)-adrenoceptors proved to be negligible. A selected 7-chloro derivative behaved as a potent α(1)-adrenoceptor antagonist and exhibited peripherally mediated hypotensive effects in rats.

3-[(Imidazolidin-2-yl)imino]indazole ligands with selectivity for the α(2)-adrenoceptor compared to the imidazoline I(1) receptor.

A series of 3-[(4,5-dihydroimidazolidin-2-yl)imino]indazoles has been synthesized as positional analogues of marsanidine, a highly selective α(2)-adrenoceptor ligand. Parent compound 4a and its 4-chloro (4c) and 4-methyl (4d) derivatives display α(2)-adrenoceptor affinity at nanomolar concentrations (K(i)=39.4, 15.9 and 22.6nM, respectively) and relatively high α(2)/I(1) selectivity ratios of 82, 115 and 690, respectively. Evidence was obtained that these compounds act as partial agonists at α(2A)-adrenoceptors. Compound 4d with intrinsic activity comparable with that of marsanidine, but lower than that of clonidine, elicited pronounced cardiovascular effects in anesthetized rats at doses as low as 0.01mg/kg iv.

Octopamine receptors from the barnacle balanus improvisus are activated by the alpha2-adrenoceptor agonist medetomidine.

G protein-coupled octopamine receptors of insects and other invertebrates represent counterparts of adrenoceptors in vertebrate animals. The alpha(2)-adrenoceptor agonist medetomidine, which is in clinical use as a veterinary sedative agent, was discovered to inhibit the settling process of barnacles, an important step in the ontogeny of this crustacean species. Settling of barnacles onto ship hulls leads to biofouling that has many harmful practical consequences, and medetomidine is currently under development as a novel type of antifouling agent. We now report that medetomidine induces hyperactivity in the barnacle larvae to disrupt the settling process. To identify the molecular targets of medetomidine, we cloned five octopamine receptors from the barnacle Balanus improvisus. We show by phylogenetic analyses that one receptor (BiOctalpha) belongs to the alpha-adrenoceptor-like subfamily, and the other four (BiOctbeta-R1, BiOctbeta-R2, BiOctbeta-R3, and BiOctbeta-R4) belong to the beta-adrenoceptor-like octopamine receptor subfamily. Phylogenetic analyses also indicated that B. improvisus has a different repertoire of beta-adrenoceptor-like octopamine receptors than insects. When expressed in CHO cells, the cloned receptors were activated by both octopamine and medetomidine, resulting in increased intracellular cAMP or calcium levels. Tyramine activated the receptors but with much lesser potency than octopamine. A hypothesis for receptor discrimination between tyramine and octopamine was generated from a homology three-dimensional model. The characterization of B. improvisus octopamine receptors is important for a better functional understanding of these receptors in crustaceans as well as for practical applications in development of environmentally sustainable antifouling agents.

Alpha2-adrenoreceptors profile modulation. 4. From antagonist to agonist behavior.

The goal of the present study was to modulate the receptor interaction properties of known alpha 2-adrenoreceptor (AR) antagonists to obtain novel alpha 2-AR agonists with desirable subtype selectivity. Therefore, a phenyl group or one of its bioisosteres or aliphatic moieties with similar steric hindrance were introduced into the aromatic ring of the antagonist lead basic structure. The functional properties of the novel compounds allowed our previous observations to be confirmed. The high efficacy of 7, 12, and 13 as alpha 2-AR agonists and the significant alpha 2C-AR subtype selective activation displayed by 11 and 15 demonstrated that favorable interactions to induce alpha 2-AR activation were formed between the pendant groups of the ligands and the aromatic cluster present in transmembrane domain 6 of the binding site cavity of the receptors.