TSPO ligand residence time: a new parameter to predict compound neurosteroidogenic efficacy

The pharmacological activation of the cholesterol-binding Translocator Protein (TSPO) leads to an increase of endogenous steroids and neurosteroids determining benefic pleiotropic effects in several pathological conditions, including anxiety disorders. The relatively poor relationship between TSPO ligand binding affinities and steroidogenic efficacies prompted us to investigate the time (Residence Time, RT) that a number of compounds with phenylindolylglyoxylamide structure (PIGAs) spends in contact with the target. Here, given the poor availability of TSPO ligand kinetic parameters, a kinetic radioligand binding assay was set up and validated for RT determination using a theoretical mathematical model successfully applied to other ligand-target systems. TSPO ligand RT was quantified and the obtained results showed a positive correlation between the period for which a drug interacts with TSPO and the compound ability to stimulate steroidogenesis. Specifically, the TSPO ligand RT significantly fitted both with steroidogenic efficacy (Emax) and with area under the dose-response curve, a parameter combining drug potency and efficacy. A positive relation between RT and anxiolytic activity of three compounds was evidenced. In conclusion, RT could be a relevant parameter to predict the steroidogenic efficacy and the in vivo anxiolytic action of new TSPO ligands

Multiple Topoisomerase I (TopoI), Topoisomerase II (TopoII) and Tyrosyl-DNA Phosphodiesterase (TDP) inhibitors in the development of anticancer drugs

DNA Topoisomerases (Topos) are ubiquitous nuclear enzymes involved in regulating the topological state of DNA and, in eukaryotic organisms, Topos can be classified into two structurally and functionally different main classes: TopoI and TopoII. Both these enzymes proved to be excellent targets of clinically significant classes of anticancer drugs. Actually, TopoI or II inhibitors show considerable wide spectrum antitumor activities, an important feature to be included in many chemotherapeutic protocols. Despite their clinical efficacy, the use of inhibitors targeting only one of the two enzymes can increase the levels of the other one, favouring the onset of unwanted phenomena such as drug resistance. Therefore, targeting both TopoI and TopoII can reduce the probability of developing resistance, as well as side effects thanks to the use of lower doses, given the synergistic effect of the dual activity. Moreover, since drug resistance is also due to DNA repair systems such as tyrosyl-DNA phosphodiesterases I and II, inhibiting Topoisomerases concomitantly to Tyrosyl-DNA phosphodiesterase enzymes could allow more efficient and safe drugs. This review represents an update of previous works reporting about dual TopoI and TopoII inhibitors, but also an overview of the new strategy regarding the development of derivatives able to simultaneously inhibit Topo and TDP enzymes, with particular attention to structure-affinity relationship studies. The newly collected de-rivatives are described focusing attention on their chemical structures and their biological profiles. The final aim is to highlight the structural requirements necessary for the development of potent multiple modulators of these targets, thus providing new potential antitumor agents for the clinical usage

Allosterism vs. Orthosterism: Recent Findings and Future Perspectives on A2B AR Physio-Pathological Implications

The development of GPCR (G-coupled protein receptor) allosteric modulators has attracted increasing interest in the last decades. The use of allosteric modulators in therapy offers several advantages with respect to orthosteric ones, as they can fine-tune the tissue responses to the endogenous agonist. Since the discovery of the first A1 adenosine receptor (AR) allosteric modulator in 1990, several efforts have been made to develop more potent molecules as well as allosteric modulators for all adenosine receptor subtypes. There are four subtypes of AR: A1, A2A, A2B, and A3. Positive allosteric modulators of the A1 AR have been proposed for the cure of pain. A3 positive allosteric modulators are thought to be beneficial during inflammatory processes. More recently, A2A and A2B AR allosteric modulators have also been disclosed. The A2B AR displays the lowest affinity for its endogenous ligand adenosine and is mainly activated as a consequence of tissue damage. The A2B AR activation has been found to play a crucial role in chronic obstructive pulmonary disease, in the protection of the heart from ischemic injury, and in the process of bone formation. In this context, allosteric modulators of the A2B AR may represent pharmacological tools useful to develop new therapeutic agents. Herein, we provide an up-to-date highlight of the recent findings and future perspectives in the field of orthosteric and allosteric A2B AR ligands. Furthermore, we compare the use of orthosteric ligands with positive and negative allosteric modulators for the management of different pathological conditions

INDOLE AMIDE DERIVATIVES: SYNTHESIS, STUCTURE-ACTIVITY RELATIONSHIPS AND MOLECULAR MODELLING STUDIES OF A NEW SERIES OF HISTAMINE H1-RECEPTOR ANTAGONISTS.

A number of indole amide derivatives bearing a basic side chain, in which the indole ring replaces the isoster benzimidazole nucleus typical of some well-known antihistamines, were prepared and tested for their H1-antihistaminic activity. The 1-benzyl-3-indolecarboxamides 32–42 showed antihistaminic (H1) activity (pA2 6–8); the 3-indolylglyoxylylamides 7–16 and the 2-indolecarboxamides 48–56 showed little or no activity. Insertion of the basic side chain of the active 3-indolecarboxamide derivatives into a piperazine ring (compounds 57–59) led to a dramatic loss of activity. All the active compounds proved to be competitive antagonists, since the values of the regression slope were not statistically different from 1. The most active compounds, 32, 33, 38–41, were also tested both in vitro for their anticholinergic activity and in vivo for their ability to antagonize histamine-induced cutaneous vascular permeability in rats. The biological results and the structure–activity relationships of the novel compounds are discussed in the light of molecular modelling studies, taking the molecule of astemizole as a model, and referring to proposed H1-receptor pharmacophore model

A2A receptors and methamphetamine toxicity: a role of adenosine as an endogenous neurotoxin

Adenosine A2A are a class of purinergic receptors largely expressed in dopamine (DA)-rich areas of the central nervous system. In particular, they are abundant within basal ganglia, where they modulate the activity of various neurotransmitters, including DA. Despite the lack of knowledge on their fine physiological mechanisms, it is worth to mention that A2A antagonists prevent neuronal death and dyskinesia in Parkinsonism. Moreover the neuroprotective effects observed after blockade of adenosine A2A receptors in several models of neurotoxicity suggests a toxic effect for endogenous adenosine In the light of these evidences, in the present study, by using in vitro models of DA neurons, we investigated: (i) whether A2A antagonists protect DA containing neurons against methamphetamine (METH); (ii) whether activation of A2A receptors produce neurodegeneration. This was done either using A2A agonist receptor NECA or the endogenous compound adenosine; (iii) whether specific cell mechanisms are involved in these phenomena. We found that A2A antagonists protect DA cells against METH neurotoxicity. Moreover, we found that NECA and adenosine both produced a toxic effects. In the light of the key role of autophagy in modulating the survival of DA neurons we found that A2A antagonists increase, while A2A agonists decrease, autophagy. These results suggest that neuroprotection induced by A2A antagonists may be mediated by enhancement of autophagy. As expected we found that pre-treatment with a non-adenosine related inducer of autophagy produced the same protective effects obtained with A2A antagonists. Our data indicate for the first time, that A2A antagonists are protective in DA neurons against METH. Such an effect appear to be mediated by the enhancement of autophagy. On the other hand we found that activation of A2A receptor produces neurotoxicity. Interestingly these effects was reproduced by administering endogenous adenosine. This suggests that adenosine may produce neurodegeneration by inhibiting the autophagy pathway

Long lasting MDM2/Translocator protein modulator: a new strategy for irreversible apoptosis of human glioblastoma cells

The development of multi-target drugs and irreversible modulators of deregulated signalling proteins is the major challenge for improving glioblastoma multiforme (GBM) treatment. Reversible single-target drugs are not sufficient to sustain a therapeutic effect over time and may favour the activation of alternative signalling pathways and the onset of resistance phenomena. Thus, a multi-target compound that has a long-lasting mechanism of action might have a greater and longer life span of anti-proliferative activity. Recently, a dual-target indol-3ylglyoxyldipeptide derivative, designed to bind to the Translocator Protein (TSPO) and reactivate p53 function via dissociation from its physiological inhibitor, murine double minute 2 (MDM2), has been developed as a potent GBM pro-apoptotic agent. In this study, this derivative was chemically modified to irreversibly bind MDM2 and TSPO. The new compound elicited a TSPO-mediated mitochondrial membrane dissipation and restored p53 activity, triggering a long-lasting apoptosis of GBM cells. These effects were sustained over time, involved a stable activation of extracellular signal regulated kinases and were specifically observed in cancer cells, in which these protein kinases are deregulated. Dual-targeting and irreversible binding properties combined in the same molecule may represent a useful strategy to overcome the time-limited effects elicited by classical chemotherapies

Novel positive allosteric modulators of A2B adenosine receptor acting as bone mineralisation promoters

Small-molecules acting as positive allosteric modulators (PAMs) of the A2B adenosine receptor (A2B AR) could potentially represent a novel therapeutic strategy for pathological conditions characterised by altered bone homeostasis, including osteoporosis. We investigated a library of compounds (4-13) exhibiting different degrees of chemical similarity with three indole derivatives (1-3), which have been recently identified by us as PAMs of the A2B AR able to promote mesenchymal stem cell differentiation and bone formation. Evaluation of mineralisation activity of 4-13 in the presence and in the absence of the agonist BAY60-6583 allowed the identification of lead compounds with therapeutic potential as anti-osteoporosis agents. Further biological characterisation of one of the most performing compounds, the benzofurane derivative 9, confirmed that such a molecule behaves as PAM of the A2B AR

The mesenchymal stem cell differentiation to osteoblasts is potentiate by the allosteric modulation of A2B adenosine receptors.

The A2B adenosine receptor (A2BAR) has been recently emerged as the major adenosine receptor involved in the mesenchymal stem cell differentiation to osteoblast and bone formation, highlighting this receptor as a new target in bone diseases. In the present study, we characterized a new 3-keto-indole-derivative (KI-7) as the first positive allosteric modulator (PAM) of the human A2B AR in mesenchymal stem cells (MSCs), and we investigated the potential activity of this compound as osteogenic agent. KI-7 was able to increase the effects of A2B AR of both endogenous and orthosteric agonists on the expression of osteogenic markers and on osteoblast mineralization. In the early phase of differentiation program, KI-7 significantly potentiated physiological and A2B agonist-mediated down-regulation of IL-6 release. Conversely, during the late stage of differentiation, when most of the cells have an osteoblast phenotype, KI-7 caused a sustained raise in IL-6 levels and an improvement in osteoblast viability. These data suggest that positive allosteric modulation of A2B AR not only favors MSC commitment to osteoblasts, but also ensures a greater survival of mature osteoblasts. Our study paves the way for a therapeutic use of selective positive allosteric modulators of A2B AR in the control of osteoblast differentiation, bone formation and fracture repair

Sulfonamides incorporating heteropolycyclic scaffolds show potent inhibitory action against carbonic anhydrase isoforms I, II, IX and XII

Three series of polycyclic compounds possessing either primary sulfonamide or carboxylic acid moieties as zinc-binding groups were investigated as inhibitors of four physiologically relevant CA isoforms, the cytosolic hCA I and II, as well as the transmembrane hCA IX and XII. Most of the new sulfonamides reported here showed excellent inhibitory effects against isoforms hCA II, IX and XII, but no highly isoform-selective inhibition profiles. On the other hand, the carboxylates selectively inhibited hCA IX (KIs ranging between 40.8 and 92.7 nM) without inhibiting significantly the other isoforms. Sulfonamides/carboxylates incorporating polycyclic ring systems such as benzothiopyranopyrimidine, pyridothiopyranopyrimidine or dihydrobenzothiopyrano[4,3-c]pyrazole may be considered as interesting candidates for exploring the design of isoform-selective CAIs with various pharmacologic applications

Acid Derivatives of Pyrazolo[1,5-a]pyrimidine as Aldose Reductase Differential Inhibitors

Aldose reductase (AKR1B1), the key enzyme of the polyol pathway, plays a crucial role in the development of long-term complications affecting diabetic patients. Nevertheless, the expedience of inhibiting this enzyme to treat diabetic complications has failed, due to the emergence of side effects from compounds under development. Actually AKR1B1 is a Janus-faced enzyme which, besides ruling the polyol pathway, takes part in the antioxidant defense mechanismof the body. In this workwe report the evidence that a class of compounds, characterized by a pyrazolo[1,5-a]pyrimidine core and an ionizable fragment, modulates differently the catalytic activity of the enzyme, depending on the presence of specific substrates such as sugar, toxic aldehydes, and glutathione conjugates of toxic aldehydes. The study stands out as a systematic attempt to generate aldose reductase differential inhibitors (ARDIs) intended to target long-term diabetic complications while leaving unaltered the detoxifying role of the enzyme