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The multimodal effective compound (I) inhibits the anticholinesterase (AChE) activity in a submicromolar range and shows a significant reduction of AChE-induced anti-aggregating efficacy without a cytotoxicity in neuronal-like cells SH-SY5Y up to the highest concentration of 50 μM.

The reaction generates a C—C double bond in a very efficient manner with only AcOH as the side product.

Alzheimer’s and prion diseases etiological mechanisms have been linked to a conformational change of normally expressed proteins that leads to the aggregation and abnormal deposition of protease-resistant and insoluble isoforms, namely amyloid-beta (A-beta) and prion protein scrapie (PrPSc). As the fibrillar aggregates of both these proteins are toxic to neurons, it has long been hypothesized that fibrils cause the underlying neurodegeneration. Thus, the amyloid plaques have been historically considered the neuropathological hallmark of these diseases determined at autopsy and, more recently, the classical biomarker for diagnostic purposes. In a personalized medicine perspective, a molecular biomarker can be utilized in the diagnosis, but also in staging and monitoring of therapy. On these basis, we explored the possibility of devising imaging probes to image the amyloid deposition in vivo and potentially provide treatment strategies against both maladies. To this aim, we focused on styryl derivatives, because several styryl compounds have been employed to detect A-beta plaques and have been successfully tested against PrPSc. Since the quinoline fragment is contained in many anti-prion compounds, we proposed to generate a library of styrylquinoline derivatives, to detect, and potentially inhibit fibrillar aggregates. The synthesis of the designed derivatives was achieved via a vinylogous variation of the Povarov reaction. They exhibited a promising activity against prion replication in ScGT1 cells and, importantly, showed no appreciable cytotoxicity. We also studied their activity as inhibitors of A-beta and PrPSc aggregation in vitro. To corroborate the possibility of employing them in vivo, we tested their ability to cross the BBB and investigated their native fluorescence in a variety of polar and non-polar environments to model their interaction with proteins, and provide the information required for their possible use as theranostic agents.

In this work, we report a rational structure-based approach aimed at the discovery of new 2-aminoimidazoles as β-secretase inhibitors. Taking advantage of a microwave-assisted synthetic protocol, a small library of derivatives was obtained and biologically evaluated. Two compounds showed promising activities in both enzymatic and cellular assays. Moreover, one of them exhibited the capability to cross the blood-brain barrier as assessed by the parallel artificial membrane permeability assay. © 2011 Elsevier Ltd. All rights reserved.

A combinatorial library of quinone–polyamine conjugates designed to optimize the antitrypanosomatid profile of hit compounds 1 and 2 has been prepared by a solid-phase approach. The conjugates were evaluated against the three most important human trypanosomatid pathogens (Trypanosoma brucei rhodesiense, Trypanosoma cruzi, and Leishmania donovani), and several showed promising activity. A subset also inhibited trypanothione reductase in vitro and induced oxidase activity of the enzyme. A highly potent analogue (7) was identified with activity against T. brucei as low as 70 nM and a selectivity index of 72. Interestingly, the presence of a cadaverine tail confers to 7 the ability to target mitochondrial function in Leishmania. In fact, in L. donovani promastigotes, we verified for 7 a decrease of cytoplasmic ATP and mitochondrial potential. Therefore, the current results support the suitability of the conjugation approach for the development of novel polyamine conjugates with enhanced therapeutic potential.

The interaction between dopaminergic and cholinergic pathways in the induction of behavioral responses has been previously established. In the brain, M2 receptors are found predominantly in presynaptic cholinergic neurons as autoreceptors, and in dopaminergic neurons as heteroceptors, suggesting a control role of acetylcholine and dopamine release, respectively. Our aim was to investigate the role of M2 receptors on the yawning and genital grooming of rats induced by apomorphine, a dopaminergic receptor agonist, focusing on the interaction between cholinergic and dopaminergic pathways. Initially, the effect of atropine, a non-selective muscarinic antagonist, on yawning and genital grooming induced by apomorphine (100μg/kg s.c.) was analyzed. Atropine doses of 0.5, 1 and 2mg/kg i.p. were administered to Wistar rats 30min before induction of the behavioral responses by apomorphine. Number of yawns and time spent genital grooming were quantified over a 60min period. Apomorphine-induced yawning was increased by low dose (0.5mg/kg i.p.) but not by high doses (1 and 2mg/kg, i.p.) of atropine. Genital grooming was antagonized by 2mg/kg i.p. of atropine and showed no changes at the other doses tested. Tripitramine, a selective M2 cholinergic antagonist, was used as a tool for distinguishing between M2 and all other muscarinic receptor subtypes in yawning and genital grooming. Tripitramine doses of 0.01, 0.02 and 0.04μmol/kg i.p. were administered to Wistar rats 30min before apomorphine (100μg/kg s.c.). Number of yawns and time spent genital grooming were also quantified over a 60min period. Tripitramine 0.01μmol/kg increased all parameters. Higher doses, which possibly block all subtypes of muscarinic receptor, did not modify the response of apomorphine, suggesting a non-selective effect of tripitramine at these doses. Given that low doses of tripitramine increased the behavioral responses induced by apomorphine and that the main distribution of the M2 receptor is presynaptic, we raised the hypothesis that tripitramine may alter cholinergic and/or dopaminergic transmission in brain areas responsible for induction of yawning and genital grooming in rats, possibly by control of acetylcholine and/or dopamine release. In addition, the present study showed the involvement of M2 cholinergic receptors in the complex mechanisms of functional interactions between dopaminergic and cholinergic systems involved in the control of yawning and genital grooming.

Binding studies in rat whole brain, frontoparietal cortex and brainstem membrane preparations revealed that benextramine displaced [3H]neuropeptide Y specific binding from a low and a high affinity site with IC50 values in the microM (36 +/- 2, 4.4 +/- 1.4 and 300 +/- 120 microM, respectively) and the pM (29.3 +/- 12.1, 0.35 +/- 0.11 and 0.42 +/- 0.03 pM, respectively) range, whereas in rat hippocampus benextramine displaced [3H]neuropeptide Y specific binding from one site only with an IC50 value of 22.8 +/- 5.7 microM. With the exception of frontoparietal cortex binding assay, benextramine was not able to completely inhibit [3H]neuropeptide Y specific binding revealing the presence of a benextramine nonsensitive third binding site. Benextramine pretreatment followed by membrane washing demonstrated that benextramine inhibited irreversibly both high and low affinity sites.

The April issue of ChemMedChem is dedicated to women in chemistry, medicinal chemistry in particular. Two of ChemMedChem's Board members, Professors Gloria Cristalli, University of Camerino, Italy, and Maria Laura Bolognesi, University of Bologna, Italy, introduce this issue with their Editorial, Remembering Marie Curie's Legacy, which highlights a few of the many examples in which female chemists have contributed enormously to their field.

Multitarget agents directed at selected molecular targets involved in the pathogenic cascade of Alzheimer's disease (AD) have been increasingly sought after in recent years, with the aim of achieving enhanced therapeutic efficiency with respect to single-target drugs and drug candidates. At the same time, much attention has been devoted to identifying high quality pharmacological tools to help explore the molecular mechanisms underlying AD without being exposed to physicochemical challenges. Herein, we discuss several examples of both types of compounds, taken from our own research and derived from the leads memoquin, lipocrine and bis(7)tacrine.

No abstract available

Eight monomeric congeners, related to the multitarget lead candidate memoquin, were prepared and evaluated at multiple targets to determine their profile against Alzheimer’s disease. 2–4 bind to AChE with similar low nanomolar affinities and function as effective inhibitors of amyloid aggregation. The most potent monovalent ligand 2 also inhibits BACE-1 in vitro and APP metabolism in primary chicken telencephalic neurons.

Focused microwave irradiation promoted the very effi- cient synthesis of 2-styrylquinolines by reaction between quinaldi- nes and benzaldehydes or cinnamaldehydes in the presence of acetic anhydride.

Prion diseases are prototypical conformational diseases. From a medicinal chemistry point of view, all conformational diseases are ‘black boxes’ because the three-dimensional structure and the mechanistic properties of the target are not fully available. Although reliable proof-of-principle was demonstrated in a variety of experimental models, and several small molecules have been identified as active against TSE [Trevitt, 2006], the molecular mechanism of action for most of these molecules remains largely unexplored. Thus, it emerges that the rational design of antiprion compounds is still a big challenge. In an effort to rationally discover antiprion lead compounds, we envisaged the planar 2,5-bis-diamino-benzoquinone and 3,6-dimethylenepiperazine-2,5-dione scaffolds as privileged motifs in modulating PPIs in prions. Building on these two fragments as appropriate spacers, we designed two series of symmetrical bifunctional ligands that effectively inhibited prion replication in ScGT1 cells [Tran, 2010; Bolognesi, 2010]. In particular, we reported that the Phe-benzoquinone conjugate 1 exhibited remarkable anti-prion activity in a cellular model (EC50 = 0.87 M) [Tran, 2010]. This allowed us to propose that the benzoquinone might be considered a promising central core in the search for novel multitarget anti-prion compounds. Starting from these results, a further library featuring a central BQ nucleus, with two spacers in position 2 and 5 connected to two terminal moieties, was designed. Some of the library entries showed significant inhibition against PrPSc aggregation in ScGT1 cell line. Notably, 6-chloro-1,2,3,4-tetrahydroacridine 2 displayed an EC50 of 0.17 μM, which was lower than that of the reference compound quinacrine. More importantly, 2 possessed the capability to contrast prion fibril formation and oxidative stress, together with a low cytotoxicity [Bongarzone, 2010]. These results validate our design rationale as a viable strategy for the identification of novel lead compounds with more than one activity against prion diseases.

Several wasp venoms contain philanthotoxins (PhTXs) that act as noncompetitive inhibitors (NCIs) on cation-selective ion channels including the nicotinic acetylcholine receptor (nAChR). In the search for a ligand with high affinity and specificity for the nAChR we tested a series of newly developed PhTX analogues. Modulation of the structural elements of PhTXs can significantly influence their binding affinities. This approach resulted in the development of the photolabile compound MR44. In photoaffinity labelling studies 125 I-MR44 was used to map the ligand-binding site at the Torpedo californica nAChR. Upon UV irradiation of the receptor–ligand complex, 125 I-MR44 was mainly incorporated into the receptor α-subunit. Proteolytic mapping and microsequencing identified the site of 125 I-MR44 cross-linking within the sequence αHis-186 to αLeu-199 that in its C-terminal region partially overlaps with the agonist-binding site. Since bound agonists had only minor influence on 125 I-MR44 photocross-linking, the site where the hydrophobic head group of 125 I-MR44 binds must be located outside the zone that is sterically influenced by agonists bound at the nAChR. A possible site of interaction of 125 I-MR44 would be the N-terminal region of the labelled sequence, in which aromatic amino-acid residues are accumulated. We suggest that the polyamine moiety of 125 I-MR44 interacts with the high affinity non-competitive inhibitor site deep in the ion channel, while the aromatic ring of this compound binds in the vestibule of the nAChR to a hydrophobic region on the α-subunit that is located close to the agonist binding site.

Memoquin (1) is a lead compound multitargeted against Alzheimer's disease (AD). It is an AChE inhibitor, free-radical scavenger, and inhibitor of amyloid-β (Aβ) aggregation. A new series of 1 derivatives was designed and synthesized by linking its 2,5-diamino-benzoquinone core with motifs that are present in the structure of known amyloid binding agents like curcumin, the benzofuran derivative SKF64346, or the benzothiazole bearing compounds KHG21834 and BTA-1. The weaker AChE inhibitory potencies and the concomitant nearly equipotent anti-amyloid activities of the new compounds with respect to 1 resulted in a more balanced biological profile against both targets. Selected compounds turned out to be effective Aβ aggregation inhibitors in a cell-based assay. By properly combining two or more distinct pharmacological properties in a molecule, we can achieve greater effectiveness compared to single-targeted drugs for investigating AD

Mitochondria-directed antioxidants 2-5 were designed by conjugating curcumin congeners with different polyamine motifs as vehicle tools. The conjugates emerged as efficient antioxidants in mitochondria and fibroblasts and also exerted a protecting role through heme oxygenase-1 activation. Notably, the insertion of a polyamine function into the curcumin-like moiety allowed an efficient intracellular uptake and mitochondria targeting. It also resulted in a significant decrease in the cytotoxicity effects. 2-5 are therefore promising molecules for neuroprotectant lead discovery.

A small library combining two different benzoquinone cores with seven ( l ) amino acid methyl esters (alanine, Nω-nitro-arginine, Ne-BOC-lysine, isoleucine, methionine, phenylalanine and tryptophan) was prepared and tested for prion replication inhibition in ScGT1 cells. The most potent hit, 6a, displayed an EC 50 value of 0.87 μM, which is very close to that of quinacrine (0.4 μM).

Prion diseases are fatal neurodegenerative and infectious disorders for which effective pharmacological tools are not yet available. This unmet challenge and the recently proposed interplay between prion diseases and Alzheimer's have led to a more urgent demand for new antiprion agents. Herein, we report the identification of a novel bifunctional diketopiperazine (DKP) derivative 1 d, which exhibits activity in the low micromolar range against prion replication in ScGT1 cells, while showing low cytotoxicity. Supported by properly addressed molecular modeling studies, we hypothesized that a planar conformation is the major determinant for activity in this class of compounds. Moreover, studies aimed at assessing the mechanism-of-action at the molecular level showed that 1 d might interact directly with recombinant prion protein (recPrP) to prevent its conversion to the pathogenic misfolded prion protein (PrP Sc )-like form. This investigation suggests that DKP based antiprion compounds can serve as a promising lead scaffold in developing new drugs to combat prion diseases.

Trypanosomiases and Leishmaniases are neglected tropical diseases that affect the less developed countries. For this reason, they did not and still do not have high visibility in Western societies. The name neglected diseases also refers to the fact that they often received little interest at the level of public investment, research and development. The drug discovery scenario, however, is changing dramatically. After a period in which different socioeconomic factors have prevented massive research efforts in this field, such efforts have increased considerably in the very recent years, with significant scientific advancements. In this context, we have embarked on a new drug discovery project devoted to identification of new small molecules for the treatment of trypanosomal and leishmanial diseases. Two complementary approaches have been pursued and are reported here. The first deals with a structure-based drug design, and a privileged structure-guided synthesis of quinazoline compounds able to modulate trypanothione reductase activity was accomplished. In the second, a combinatorial library, built on a natural product-based strategy, was synthesized. Using whole parasite assays, different quinones have been identified as promising lead compounds. A combination of both approaches to hopefully overcome some of the challenges of anti-trypanosomatid drug discovery has eventually been proposed.

A library of 11 entries, featuring a 2,5-diamino-1,4-benzoquinones nucleus as spacer connecting two aromatic prion recognition motifs, was designed and evaluated against prion infection. Notably, 6-chloro-1,2,3,4-tetrahydroacridine 10 showed an EC(50) of 0.17 μM, which was lower than that displayed by reference compound BiCappa. More importantly, 10 possessed the capability to contrast prion fibril formation and oxidative stress, together with a low cytotoxicity. This study further corroborates the bivalent strategy as a viable approach to the rational design of anti-prion chemical probes.

A small library combining two different benzoquinone cores with seven (L) amino acid methyl esters (alanine, Nomega-nitro-arginine, Nepsilon-BOC-lysine, isoleucine, methionine, phenylalanine and tryptophan) was prepared and tested for prion replication inhibition in ScGT1 cells. The most potent hit, 6a, displayed an EC(50) value of 0.87 microM, which is very close to that of quinacrine (0.4 microM).

Neglected tropical diseases (NTD) are parasitic infections that affect the poorest people in less developed countries. For this reason, they did not and still do not have high visibility in Western societies. The name neglected diseases reflects also the failure of pharmaceutical companies to invest in research and development of new medicines. In fact, most of the drugs still used to treat these diseases were developed in colonial times. These are often expensive, have variable efficacy, toxic side effects and require long courses of administration. Furthermore, several of them are also becoming ineffective because of increasing parasite resistance. This gloomy outlook has changed during the past decade as a result of important advances in Kinetoplastida (comprising the genera Trypanosoma and Leishmania) basic research. In 2005, the genome sequences of the ‘Tritryp’ (T. brucei, T. cruzi and L. major) and in 2007 those of two other species of Leishmania (L. infantum and L. braziliensis) were completed, eliminating many of the hurdles to performing state-of-the-art drug discovery research. The vast amount of available information has made a more comprehensive and accurate identification and validation of novel possible drug targets. However, new licensed therapies have not yet resulted from genome-derived investigations, and trypanosomiases and leishmaniases, even in the post-genomic era, may remain incurable and often fatal. We have proposed the multi-target drug design strategy as a means to overcome the challenges of current drug discovery in NTD.[1, 2] This approach is based on the design of single chemical entities able to simultaneously modulate multiple targets. Although these concepts emerged less than 10 years ago, multi-target drugs are already a reality in the therapy of complex diseases. In NTD, multi-target agents have an added value with respect to the single-target ones: the onset of drug resistance might be significantly delayed

Many lines of evidence suggest that oxidative stress and mitochondria impairment have a central role in age-related neurodegenerative diseases such as Alzheimer’s disease (AD). In addition, the recent understanding that mitochondria are at the intersection of the life and death of a cell, particularly through the involvement of mitochondrial oxidative stress, has made mitochondria a promising target for drug discovery and therapeutic interventions. Thus, considering the oxidative stress as a crucial event on the degenerative cascade, we designed a series of compounds presenting a constrained curcumin-like moiety. Curcumin and its constrained analogs have currently received remarkable interest as they have a unique conjugated structure which shows a pleiotropic biological profile. Actually, beside the well-known direct antioxidant activity, curcumin displays a wide range of properties including anti-inflammatory and anti-amyloidogenic activities. Moreover, dietary antioxidants, like curcumin, have recently been demonstred in vitro to be neuroprotective through the activation of heme oxigenase 1 pathway, showing an additional indirect antioxidant behaviour. On the basis of the consideration that mitochondria are a major source of ROS and are particularly vulnerable to oxidative stress, it is convincible that antioxidants that alleviate mitochondrial dysfunction could be beneficial in AD. This prompted us to design mitochondria-targeted antioxidants by connecting curcumin analogs to different polyamine chains. Polyamines might deliver the corresponding conjugates into the cell with active polyamine transporters and, acting as lipophilic cations, drive the antioxidant moiety selectively into mitochondria in a membrane potential-dependent manner across the inner membrane. In order to validate if such compounds were preferentially taken up by mitochondria, a fluorescent probe was designed by combining the phenyl benzothiazole group with the polyamine residue.

The binding of polyamines to a variety of receptors and other defined recognition sites has been widely reported. It is well-known that polyamines interact with aspartate, glutamate, and aromatic residues of a given receptor and/or enzyme mainly through the formation of ion bonds, since at physiological pH, protonation of amino groups is nearly complete. From this, the hypothesis arises that a polyamine may be a universal template able to recognize different receptor systems. This hypothesis suggests that both affinity and selectivity may be fine-tuned by inserting appropriate substituents onto the amine functions and by varying the methylene chain lengths between them on the polyamine backbone. In this paper, we detail several application of this design strategy aimed at discovering potent and selective polyamines able to bind neurotransmitter receptors and enzymes, such as muscarinic receptor subtypes, muscle-type nicotinic receptors and acethylcholinesterase.

The present article expands on the study of structure–activity relationships of the novel class of quinone-bearing polyamines, as multi-target-directed ligands against Alzheimer’s disease. Namely, the effect of inserting a methyl substituent at the α position of the terminal benzyl amine moieties of lead candidate 1 (memoquin) was evaluated at the multiple targets involved in the multifunctional mechanism of action. The RR stereoisomer 2 resulted more effective than 1 in reverting two important effects mediated by acetylcholinesterase (AChE), that is, acetylcholine hydrolysis and AChE-induced amyloid-β aggregation.

In this work, we review and comment upon the challenges and the 'quo vadis' in Alzheimer's disease drug discovery at the beginning of the new millennium. We emphasize recent approaches that, moving on from a target-centric approach, have produced innovative molecular probes or drug candidates. In particular, the discovery of endosome-targeted BACE1 inhibitors and mitochondria-targeted antioxidants represents a significant advance in Alzheimer's research and therapy. The case study of the development of rasagiline provides an excellent example to support the validity of the multitarget-designed ligand approach to the search for effective medicines for combating Alzheimer's disease.

Novel multitargeted antioxidants 3-6 were designed by combining the antioxidant features, namely, a benzoquinone fragment and a lipoyl function, of two multifunctional lead candidates. They were then evaluated to determine their profile against Alzheimer's disease. They showed antioxidant activity, improved following enzymatic reduction, in mitochondria and T67 cell line. They also displayed a balanced inhibitory profile against amyloid-beta aggregation and acetylcholinesterase, emerging as promising molecules for neuroprotectant lead discovery.

Alzheimer’s disease (AD) is a major neurodegenerative disease affecting a substantial proportion of elderly population and imposing a significant burden on public health and quality of life. Despite of this, for AD no cure still exists. The finding that it is a complex multifactorial disorder arising from a complex array of neurochemical factors may explain, at least partially, why the currently available drugs, developed according to the classic drug discovery paradigm of "one-molecule-one-target," have turned out to be palliative, rather than curative. In light of this, drug combinations that can act at different levels of the neurotoxic cascade offer new hopes toward effectively treating AD. This finding was also the logical starting point for achieving disease modifying therapeutics by the identification of ligands possessing a multi-modal mechanism of action. Multi-target-directed ligands (MTDLs), which are small molecule rationally designed with the deliberate aim of simultaneously modulating multiple targets in the disease network, should be especially suited to address the extreme complexity of AD etiology. In recent years, the MTDL concept has been exploited by us and others to design different ligands hitting different biological targets. Here, we report on memoquin, a novel MTDL designed by inserting the benzoquinone nucleus of coenzyme Q into a polyamine scaffold. Memoquin furnished us with the ‘proof of concept’ of the MTDL drug discovery approach. In fact, a detailed in vitro and in vivo investigation allowed us to demonstrate that memoquin combats AD on different crucial fronts: it inhibits acetylcholinesterase, amyloid production and aggregation, and also oxidative stress. Memoquin was also the lead molecule for an optimization program aimed at developing novel MTDLs with more balanced affinity profiles and physicochemical properties consistent with good oral absorption. The results obtained further support the potential for the development of efficacious multi-targeted drug candidates against AD.

Trypanosomiases and Leishmaniases are neglected tropical diseases that affect the less developed countries. For this reason, they did not and still do not have high visibility in Western societies. The name neglected diseases also refers to the fact that they often received little interest at the level of public investment, research and development. The drug discovery scenario, however, is changing dramatically. After a period in which different socioeconomic factors have prevented massive research efforts in this field, such efforts have increased considerably in the very recent years, with significant scientific advancements. In this context, we have embarked on a new drug discovery project devoted to identification of new small molecules for the treatment of trypanosomal and leishmanial diseases. Two complementary approaches have been pursued and are reported here. The first deals with a structure-based drug design, and a privileged structure-guided synthesis of quinazoline compounds able to modulate trypanothione reductase activity was accomplished. In the second, a combinatorial library, built on a natural product-based strategy, was synthesized. Using whole parasite assays, different quinones have been identified as promising lead compounds. A combination of both approaches to hopefully overcome some of the challenges of anti-trypanosomatid drug discovery has eventually been proposed.

Alzheimer's disease is currently thought to be a complex, multifactorial syndrome, unlikely to arise from a single causal factor; instead, a number of related biological alterations are thought to contribute to its pathogenesis. This may explain why the currently available drugs, developed according to the classic drug discovery paradigm of "one-molecule-one-target," have turned out to be palliative. In light of this, drug combinations that can act at different levels of the neurotoxic cascade offer new avenues toward curing Alzheimer's and other neurodegenerative diseases. In parallel, a new strategy is emerging-that of developing a single chemical entity able to modulate multiple targets simultaneously. This has led to a new paradigm in medicinal chemistry, the "multi-target-directed ligand" design strategy, which has already been successfully exploited at both academic and industrial levels. As a case study, we report here on memoquin, a new molecule developed following this strategy. The in vitro and in vivo biological profile of memoquin demonstrates the suitability of the new strategy for obtaining innovative drug candidates for the treatment of neurodegenerative diseases.

Novel quinazoline-type compounds were designed as inhibitors of the parasite specific enzyme trypanothione reductase (TR), and their biological activities were evaluated. Some of our compounds inhibited TR, showed selectivity for TR over human glutathione reductase, and inhibited parasite growth in vitro. We propose that the quinazoline framework is a privileged structure that can be purposely modified to design novel TR inhibitors. Furthermore, the use of privileged motifs might emerge as an innovative approach to antiparasitic lead candidates.

The photostability properties of memoquin, a multifunctional compound in preclinical development for the treatment of Alzheimer's disease (AD) were investigated in solutions exposed to radiations, using a xenon arc lamp to simulate the natural sunlight. Reversed phase liquid chromatography coupled with diode array detection and electrospray ionization mass spectrometry (LC–UV/DAD–ESI-MS/MS) was applied to follow the photodegradation and disappearance of memoquin after irradiation. Under optimized chromatographic conditions, memoquin was separated with high resolution from the photoproducts formed in the photoexposed solutions. The results showed that memoquin is more stable at physiological and acid pHs, while it has a slow degradation pattern at more drastic conditions such as basic pH (t1/2 = 389 min) and in methanolic solutions (t1/2 = 465 min). In the irradiated solutions the appearance of photoproducts with lower retention times and molecular weight than memoquin was observed, thus indicating that some fragments were lost from its structure. The photodegradation products were characterized by LC–ESI-MS/MS and LC–UV/DAD analysis. The photoreactive centers were found on the amino groups of the side chains while the 1,4-benzoquinone functionality was maintained. Conversely, memoquin was found to be stable in the dark. These results suggest that, with appropriate handling and storage, memoquin's activity is not impaired.

A library of 24 derivatives designed by combining two natural products-derived fragments was prepared and tested to determine their anticancer potential in HT29 colon cancer cells. All library members inhibit cell proliferation as measured by MTT mitochondrial functional assay, with IC50 values in the 1-100 microM range. Entry 1b caused apoptotic EGFR-mediated intracellular signaling. Thus, polyamino-quinones emerged as readily accessible and easily diversified scaffolds for anticancer lead discovery.

Our understanding of the pathogenesis of diseases has advanced enormously in recent decades. As a consequence, drug discovery has gradually shifted from an entirely humanphenotype-based endeavor to today’s reductionist approach centered on single molecular targets. The focus has shifted from the early animal models to isolated proteins via cellular models. This change has led to a decrease in complexity but also to a decrease in relevance to the human condition. In this context, drug research has become (and still is) aimed mainly at the discovery of small molecules able to modulate the biological function of a single protein target thought to be fully responsible for a certain disease. Much effort has been devoted to achieving selectivity for that given target, and indeed, nowadays, many ligands endowed with outstanding in vitro selectivity are available. This one-molecule, one-target paradigm has led to the discovery of many successful drugs, and it will probably remain a milestone for years to come. However, it should be noted that a highly selective ligand for a given target does not always result in a clinically efficacious drug. This may be because (a) the ligand does not recognize the target in vivo, (b) the ligand does not reach the site of action, or (c) the interaction with the respective target does not have enough impact on the diseased system to restore it effectively. Reasons for the latter might lie in both the multifactorial nature of many diseases and the fact that cells can often find ways to compensate for a protein whose activity is affected by a drug, by taking advantage of the redundancy of the system, i.e., of the existence of parallel pathways. Medicinal chemists are often faced with these frustrating aspects of drug research. Drawbacks a and b can be addressed through the well-established rational ligand modification approaches. But issue c is more problematic and needs to be carefully discussed. This is one of the aims of the present article.

Novel multi-target-directed ligands were designed by replacing the inner dipiperidino function of 3 with less flexible or completely rigid moieties to obtain compounds endowed with multiple biological properties that might be relevant to Alzheimer's disease. 15 was the most interesting, inhibiting AChE in the nanomolar range and inhibiting AChE-induced and self-promoted beta-amyloid aggregation in the micromolar range.

Aimed at discovering innovative drug candidates for the treatment of trypanosomatid infections, we decided to follow two strategies that are nowadays well known in the med chem community: structure-based drug design; library design and synthesis. In the first approach, the 3D structure of validated molecular targets are used as starting point for docking simulations and de novo design. In the second approach, a compound library of 15-to-20 molecules was generated and tested by means of whole-parasite assays. In this case, the molecular target is initially unknown, and a posteriori by means of forward-chemical-genetics approaches, like the chemical proteomics, the target is tentatively fished out from parasitic cell extracts.

Prion diseases, also known as transmissible spongiform encephalopathies (TSE), are fatal neurodegenerative disorders of the central nervous system. The main molecular mechanism underlining TSE is based on the aberrant misfolding of the cellular form of the prion protein (PrPC) into its pathological counterpart denominated PrPSc. To date there are no identified therapies. One therapeutic strategy against this disease is focused on the stabilization of the PrPC in order to prevent its conversion to PrPSc . Designing ligands targeting PrPC with a high binding affinity might augment its stability and prevent its misfolding. Here, we have set-up a computational protocol aiming at addressing this issue.

Pirenzepine ( 2 ) is one of the most selective muscarinic M 1 versus M 2 receptor antagonists known. A series of 2 analogs, in which the piperazyl moiety was replaced by a cis - and trans -cyclohexane-1,2-diamine ( 3 – 6 ) or a trans - and cis -perhydroquinoxaline rings ( 7 and 8 ) were prepared, with the aim to investigate the role of the piperazine ring of 2 in the interaction with the muscarinic receptors. The structural change leading to compounds 3 – 6 abolished in binding assays the muscarinic M 1 /M 2 selectivity of 2 , due to an increased M 2 affinity. Rather, compounds 3 – 6 displayed a reversed selectivity showing more affinity at the muscarinic M 2 receptor than at all the other subtypes tested.

Taking advantage of the structural features of natural products showing anti-trypanosomatid activity, we designed and synthesized a small library of 2-phenoxy-1,4-naphthoquinone and 2-phenoxy-1,4-anthraquinone derivatives. The library was obtained following a parallel approach and using readily available synthons. All the derivatives showed inhibitory activity toward either Trypanosoma or Leishmania species, with 8, 10, and 16 being the most active compounds against Trypanosoma brucei rhodesiense, Leishmania donovani, and Trypanosoma cruzi cells (IC(50)=50nM, IC(50)=0.28microM, and IC(50)=1.26microM, respectively).

The initial transition of amyloid beta (1-42) (Abeta42) soluble monomers/small oligomers from unordered/alpha-helix to a beta-sheet-rich conformation represents a suitable target to design new potent inhibitors and to obtain effective therapeutics for Alzheimer's disease. Under optimized conditions, this reliable and reproducible CD kinetic study showed a three-step sigmoid profile that was characterized by a lag phase (prevailing unordered/alpha-helix conformation), an exponential growth phase (increasing beta-sheet secondary structure) and a plateau phase (prevailing beta-sheet secondary structure). This kinetic analysis brought insight into the inhibitors' mechanism of action. In fact, an increase in the duration of the lag phase can be related to the formation of an inhibitor-Abeta complex, in which the non-amyloidogenic conformation is stabilized. When the exponential rate is affected exclusively, such as in the case of Congo red and tetracycline, then the inhibitor affinity might be higher for the pleated beta-sheet structure. Finally, by adding the inhibitor at the end of the exponential phase, the soluble protofibrils can be disrupted and the Abeta amyloidogenic structure can revert into monomers/small oligomers. Congo red and tetracycline preferentially bind to amyloid in the beta-sheet conformation because both decreased the slope of the exponential growth, even if to a different extent, whereas no effect was observed for tacrine and galantamine. Some very preliminary indications can be derived about the structural requirements for binding to nonamyloidogenic or beta-sheet amyloid secondary structure for the development of potent antiaggregating agents. On these premises, memoquin, a multifunctional molecule that was designed to become a drug candidate for the treatment of Alzheimer's disease, was investigated under the reported circular dichroism assay and its anti-amyloidogenic mechanism of action was elucidated.

One of the characteristics of Alzheimer's disease (AD) that hinders the discovery of effective disease-modifying therapies is the multifactorial nature of its etiopathology. To circumvent this drawback, the use of multi-target-directed ligands (MTDLs) has recently been proposed as a means of simultaneously hitting several targets involved in the development of the AD syndrome. In this paper, a new class of MTDLs based on a polyamine-quinone skeleton, whose lead (memoquin, 2) showed promising properties in preclinical investigations (Cavalli et al. Angew. Chem., Int. Ed. 2007, 46, 3689-3692), is described. 3-29 were tested in vitro against a number of isolated AD-related targets, namely, AChE and BChE, and Abeta aggregation (both AChE-mediated and self-induced). Furthermore, the ability of the compounds to counteract the oxidative stress in a human neuronal-like cellular system (SH-SY5Y cells) was assayed, in both the presence and absence of NQO1, an enzyme able to generate and maintain the reduced form of quinone.

Memoquin, a recently discovered new drug candidate for the treatment of Alzheimer's disease (AD), is a compound able to interfere with different key targets of AD neurodegeneration. In the present work, the electroactivity of Memoquin, due to the presence of a quinone ring in the molecule, was exploited for the development of a sensitive and selective HPLC chromatographic method with coulometric detection system. For this purpose, a monolithic silica-based stationary phase (Chromolith C18) was coupled with ESA detector and under high flow rate condition (2 mL/min) gave an efficient coulometric detection without a significant backpressure. The monolithic stationary phase and the electrochemical detection were combined to obtain a very fast (less than 7 min) and sensitive analysis of the compound of interest. For this reason, the method was found suitable to determine Memoquin with very high sensitivity (LOD, 0.005 microg/mL; LOQ, 0.016 microg/mL), better than with UV and fluorescence detections, and selectivity resulting potentially suitable for its analysis in biological samples. Moreover, the HPLC method was employed for the separation of Memoquin from other quinone analogs (endowed with different substituent on the quinone ring and length of the lateral chain) and allowed the comparison of the electrochemical properties of the compounds of the series. In the optimized chromatographic conditions it was possible to separate each quinone and to evaluate the influence of the substituent of the quinone ring on the electrochemical properties of the molecule.

A design strategy to convert a dual-binding site AChE inhibitor into triple functional compounds with promising in vitro profile against multifactorial syndromes, such as Alzheimer's disease, is proposed. The lead compound bis(7)-tacrine (2) was properly modified to confer to the new molecules the ability of chelating metals, involved in the neurodegenerative process. The multifunctional compounds show activity against human AChE, are able to inhibit the AChE-induced amyloid-beta aggregation, and chelate metals, such as iron and copper.