Author Report for: Galdeano C

Soluble epoxide hydrolase (sEH) is a drug target with the potential for therapeutic utility in the areas of inflammation, neurodegenerative disease, chronic pain, and diabetes, among others. Proteolysis-targeting chimeras (PROTACs) molecules offer new opportunities for targeting sEH, due to its capacity to induce its degradation. Here, we describe that the new ALT-PG2, a PROTAC that degrades sEH protein in the human hepatic Huh-7 cell line, in isolated mouse primary hepatocytes, and in the liver of mice. Remarkably, sEH degradation caused by ALT-PG2 was accompanied by an increase in the phosphorylated levels of AMP-activated protein kinase (AMPK), while phosphorylated extracellular-signal-regulated kinase 1/2 (ERK1/2) was reduced. Consistent with the key role of these kinases on endoplasmic reticulum (ER) stress, ALT-PG2 attenuated the levels of ER stress and inflammatory markers. Overall, the findings of this study indicate that targeting sEH with degraders is a promising pharmacological strategy to promote AMPK activation and to reduce ER stress and inflammation.

Multitarget anti-Alzheimer agents are the focus of very intensive research. Through a comprehensive bibliometric analysis of the publications in the period 1990-2020, we have identified trends and potential gaps that might guide future directions. We found that: (i) the number of publications boomed by 2011 and continued ascending in 2020; (ii) the linked-pharmacophore strategy was preferred over design approaches based on fusing or merging pharmacophores or privileged structures; (iii) a significant number of in vivo studies, mainly using the scopolamine-induced amnesia mouse model, have been performed, especially since 2017; (iv) China, Italy and Spain are the countries with the largest total number of publications on this topic, whereas Portugal, Spain and Italy are the countries in whose scientific communities this topic has generated greatest interest; (v) acetylcholinesterase, beta-amyloid aggregation, oxidative stress, butyrylcholinesterase, and biometal chelation and the binary combinations thereof have been the most commonly pursued, while combinations based on other key targets, such as tau aggregation, glycogen synthase kinase-3beta, NMDA receptors, and more than 70 other targets have been only marginally considered. These results might allow us to spot new design opportunities based on innovative target combinations to expand and diversify the repertoire of multitarget drug candidates and increase the likelihood of finding effective therapies for this devastating disease.

The inhibition of the enzyme soluble epoxide hydrolase (sEH) has demonstrated clinical therapeutic effects in several peripheral inflammatory-related diseases, with 3 compounds in clinical trials. However, the role of this enzyme in the neuroinflammation process has been largely neglected. Herein, we disclose the pharmacological validation of sEH as a novel target for the treatment of Alzheimer's disease (AD). Evaluation of cognitive impairment and pathological hallmarks were used in 2 models of age-related cognitive decline and AD using 3 structurally different and potent sEH inhibitors as chemical probes. sEH is upregulated in brains from AD patients. Our findings supported the beneficial effects of central sEH inhibition, regarding reducing cognitive impairment, neuroinflammation, tau hyperphosphorylation pathology, and the number of amyloid plaques. This study suggests that inhibition of inflammation in the brain by targeting sEH is a relevant therapeutic strategy for AD.

Neuroinflammation is a crucial process associated with the pathogenesis of neurodegenerative diseases, including Parkinson's disease (PD). Several pieces of evidence suggest an active role of lipid mediators, especially epoxy-fatty acids (EpFAs), in the genesis and control of neuroinflammation; 14,15-epoxyeicosatrienoic acid (14,15-EET) is one of the most commonly studied EpFAs, with anti-inflammatory properties. Soluble epoxide hydrolase (sEH) is implicated in the hydrolysis of 14,15-EET to its corresponding diol, which lacks anti-inflammatory properties. Preventing EET degradation thus increases its concentration in the brain through sEH inhibition, which represents a novel pharmacological approach to foster the reduction of neuroinflammation and by end neurodegeneration. Recently, it has been shown that sEH levels increase in brains of PD patients. Moreover, the pharmacological inhibition of the hydrolase domain of the enzyme or the use of sEH knockout mice reduced the deleterious effect of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) administration. This paper overviews the knowledge of sEH and EETs in PD and the importance of blocking its hydrolytic activity, degrading EETs in PD physiopathology. We focus on imperative neuroinflammation participation in the neurodegenerative process in PD and the putative therapeutic role for sEH inhibitors. In this review, we also describe highlights in the general knowledge of the role of sEH in the central nervous system (CNS) and its participation in neurodegeneration. We conclude that sEH is one of the most promising therapeutic strategies for PD and other neurodegenerative diseases with chronic inflammation process, providing new insights into the crucial role of sEH in PD pathophysiology as well as a singular opportunity for drug development.

Symptomatic treatment of myasthenia gravis is based on the use of peripherally-acting acetylcholinesterase (AChE) inhibitors that, in some cases, must be discontinued due to the occurrence of a number of side-effects. Thus, new AChE inhibitors are being developed and investigated for their potential use against this disease. Here, we have explored two alternative approaches to get access to peripherally-acting AChE inhibitors as new agents against myasthenia gravis, by structural modification of the brain permeable anti-Alzheimer AChE inhibitors tacrine, 6-chlorotacrine, and huprine Y. Both quaternization upon methylation of the quinoline nitrogen atom, and tethering of a triazole ring, with, in some cases, the additional incorporation of a polyphenol-like moiety, result in more polar compounds with higher inhibitory activity against human AChE (up to 190-fold) and butyrylcholinesterase (up to 40-fold) than pyridostigmine, the standard drug for symptomatic treatment of myasthenia gravis. The novel compounds are furthermore devoid of brain permeability, thereby emerging as interesting leads against myasthenia gravis.

Dual submicromolar trypanocidal-antiplasmodial compounds have been identified by screening and chemical synthesis of 4-aminoquinoline-based heterodimeric compounds of three different structural classes. In Trypanosoma brucei, inhibition of the enzyme trypanothione reductase seems to be involved in the potent trypanocidal activity of these heterodimers, although it is probably not the main biological target. Regarding antiplasmodial activity, the heterodimers seem to share the mode of action of the antimalarial drug chloroquine, which involves inhibition of the haem detoxification process. Interestingly, all of these heterodimers display good brain permeabilities, thereby being potentially useful for late stage human African trypanosomiasis. Future optimization of these compounds should focus mainly on decreasing cytotoxicity and acetylcholinesterase inhibitory activity.

Multitarget compounds are increasingly being pursued for the effective treatment of complex diseases. Herein, we describe the design and synthesis of a novel class of shogaol-huprine hybrids, purported to hit several key targets involved in Alzheimer's disease. The hybrids have been tested in vitro for their inhibitory activity against human acetylcholinesterase and butyrylcholinesterase and antioxidant activity (ABTS(+), DPPH and Folin-Ciocalteu assays), and in intact Escherichia coli cells for their Abeta42 and tau anti-aggregating activity. Also, their brain penetration has been assessed (PAMPA-BBB assay). Even though the hybrids are not as potent AChE inhibitors or antioxidant agents as the parent huprine Y and [4]-shogaol, respectively, they still exhibit very potent anticholinesterase and antioxidant activities and are much more potent Abeta42 and tau anti-aggregating agents than the parent compounds. Overall, the shogaol-huprine hybrids emerge as interesting brain permeable multitarget anti-Alzheimer leads.

A family of huprine-tacrine heterodimers has been developed to simultaneously block the active and peripheral sites of acetylcholinesterase (AChE). Their dual site binding for AChE, supported by kinetic and molecular modeling studies, results in a highly potent inhibition of the catalytic activity of human AChE and, more importantly, in the in vitro neutralization of the pathological chaperoning effect of AChE toward the aggregation of both the beta-amyloid peptide (Abeta) and a prion peptide with a key role in the aggregation of the prion protein. Huprine-tacrine heterodimers take on added value in that they display a potent in vitro inhibitory activity toward human butyrylcholinesterase, self-induced Abeta aggregation, and beta-secretase. Finally, they are able to cross the blood-brain barrier, as predicted in an artificial membrane model assay and demonstrated in ex vivo experiments with OF1 mice, reaching their multiple biological targets in the central nervous system. Overall, these compounds are promising lead compounds for the treatment of Alzheimer's and prion diseases.

BACKGROUND: Multifactorial diseases such as Alzheimer's disease (AD) should be more efficiently tackled by drugs which hit multiple biological targets involved in their pathogenesis. We have recently developed a new family of huprine-tacrine heterodimers, rationally designed to hit multiple targets involved upstream and downstream in the neurotoxic cascade of AD, namely beta-amyloid aggregation and formation as well as acetylcholinesterase catalytic activity. OBJECTIVE: In this study, the aim was to expand the pharmacological profiling of huprine-tacrine heterodimers investigating their effect on muscarinic M(1) receptors as well as their neuroprotective effects against an oxidative insult. METHODS: Sprague-Dawley rat hippocampus homogenates were used to assess the specific binding of two selected compounds in competition with 1 nM [(3)H]pirenzepine (for M(1) receptors) or 0.8 nM [(3)H]quinuclidinyl benzilate (for M(2) receptors). For neuroprotection studies, SHSY5Y cell cultures were subjected to 250 muM hydrogen peroxide insult with or without preincubation with some huprine-tacrine heterodimers. RESULTS: A low nanomolar affinity and M(1)/M(2) selectivity has been found for the selected compounds. Huprine-tacrine heterodimers are not neurotoxic to SHSY5Y cells at a range of concentrations from 1 to 0.001 muM, and some of them can protect cells from the oxidative damage produced by hydrogen peroxide at concentrations as low as 0.001 muM. CONCLUSION: Even though it remains to be determined if these compounds act as agonists at M(1) receptors, as it is the case of the parent huprine Y, their low nanomolar M(1) affinity and neuroprotective effects expand their multitarget profile and increase their interest as disease-modifying anti-Alzheimer agents.

A series of 19 huprines has been evaluated for their activity against cultured bloodstream forms of Trypanosoma brucei and Plasmodium falciparum. Moreover, cytotoxicity against rat myoblast L6 cells was assessed for selected huprines. All the tested huprines are moderately potent and selective trypanocidal agents, exhibiting IC(50) values against T. brucei in the submicromolar to low micromolar range and selectivity indices for T. brucei over L6 cells of approximately 15, thus constituting interesting trypanocidal lead compounds. Two of these huprines were also found to be active against a chloroquine-resistant strain of P. falciparum, thus emerging as interesting trypanocidal-antiplasmodial dual acting compounds, but they exhibited little selectivity for P. falciparum over L6 cells.

Two novel families of dual binding site acetylcholinesterase (AChE) inhibitors have been developed, consisting of a tacrine or 6-chlorotacrine unit as the active site interacting moiety, either the 5,6-dimethoxy-2-[(4-piperidinyl)methyl]-1-indanone fragment of donepezil (or the indane derivative thereof) or a 5-phenylpyrano[3,2-c]quinoline system, reminiscent to the tryciclic core of propidium, as the peripheral site interacting unit, and a linker of suitable length as to allow the simultaneous binding at both sites. These hybrid compounds are all potent and selective inhibitors of human AChE, and more interestingly, are able to interfere in vitro both formation and aggregation of the beta-amyloid peptide, the latter effects endowing these compounds with the potential to modify Alzheimer's disease progression.

Dual binding site acetylcholinesterase inhibitors have recently emerged as a new class of anti-Alzheimer agents with potential to positively modify the course of the disease. These compounds exhibit a multifunctional pharmacological profile arising from interaction with several biological targets involved upstream and downstream in the neurodegenerative cascade of Alzheimer's disease (AD). The primary target of these compounds is the enzyme acetylcholinesterase (AChE). Interaction of dual binding site AChE inhibitors with AChE results in a potent inhibitory activity of AChE and AChE-induced beta-amyloid peptide (Abeta) aggregation. Some dual binding site AChE inhibitors take on added value a significant ability to additionally inhibit the enzymes butyrylcholinesterase and BACE-1, involved in the co-regulation of the hydrolysis of the neurotransmitter acetylcholine and in Abeta formation, respectively. The structural determinants which mediate the interaction of dual binding site AChE inhibitors with these three important enzymes for AD treatment are herein reviewed.

A new family of dual binding site acetylcholinesterase (AChE) inhibitors has been designed, synthesized, and tested for their ability to inhibit AChE, butyrylcholinesterase (BChE), AChE-induced and self-induced beta-amyloid (Abeta) aggregation and beta-secretase (BACE-1), and to cross the blood-brain barrier. The new heterodimers consist of a unit of racemic or enantiopure huprine Y or X and a donepezil-related 5,6-dimethoxy-2-[(4-piperidinyl)methyl]indane moiety as the active site and peripheral site to mid-gorge-interacting moieties, respectively, connected through a short oligomethylene linker. Molecular dynamics simulations and kinetics studies support the dual site binding to AChE. The new heterodimers are potent inhibitors of human AChE and moderately potent inhibitors of human BChE, AChE-induced and self-induced Abeta aggregation, and BACE-1, and are predicted to be able to enter the central nervous system (CNS), thus constituting promising multitarget anti-Alzheimer drug candidates with the potential to modify the natural course of this disease.

Two isomeric series of dual binding site acetylcholinesterase (AChE) inhibitors have been designed, synthesized, and tested for their ability to inhibit AChE, butyrylcholinesterase, AChE-induced and self-induced beta-amyloid (Abeta) aggregation, and beta-secretase (BACE-1) and to cross blood-brain barrier. The new hybrids consist of a unit of 6-chlorotacrine and a multicomponent reaction-derived pyrano[3,2-c]quinoline scaffold as the active-site and peripheral-site interacting moieties, respectively, connected through an oligomethylene linker containing an amido group at variable position. Indeed, molecular modeling and kinetic studies have confirmed the dual site binding of these compounds. The new hybrids, and particularly 27, retain the potent and selective human AChE inhibitory activity of the parent 6-chlorotacrine while exhibiting a significant in vitro inhibitory activity toward the AChE-induced and self-induced Abeta aggregation and toward BACE-1, as well as ability to enter the central nervous system, which makes them promising anti-Alzheimer lead compounds.

A novel series of donepezil-tacrine hybrids designed to simultaneously interact with the active, peripheral and midgorge binding sites of acetylcholinesterase (AChE) have been synthesized and tested for their ability to inhibit AChE, butyrylcholinesterase (BChE), and AChE-induced A beta aggregation. These compounds consist of a unit of tacrine or 6-chlorotacrine, which occupies the same position as tacrine at the AChE active site, and the 5,6-dimethoxy-2-[(4-piperidinyl)methyl]-1-indanone moiety of donepezil (or the indane derivative thereof), whose position along the enzyme gorge and the peripheral site can be modulated by a suitable tether that connects tacrine and donepezil fragments. All of the new compounds are highly potent inhibitors of bovine and human AChE and BChE, exhibiting IC50 values in the subnanomolar or low nanomolar range in most cases. Moreover, six out of the eight hybrids of the series, particularly those bearing an indane moiety, exhibit a significant A beta antiaggregating activity, which makes them promising anti-Alzheimer drug candidates.