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1.
Int J Mol Sci ; 24(7)2023 Apr 06.
Article in English | MEDLINE | ID: mdl-37047792

ABSTRACT

Schistosomiasis is a neglected tropical disease with high morbidity. Recently, the Schistosoma mansoni phosphodiesterase SmPDE4A was suggested as a putative new drug target. To support SmPDE4A targeted drug discovery, we cloned, isolated, and biochemically characterized the full-length and catalytic domains of SmPDE4A. The enzymatically active catalytic domain was crystallized in the apo-form (PDB code: 6FG5) and in the cAMP- and AMP-bound states (PDB code: 6EZU). The SmPDE4A catalytic domain resembles human PDE4 more than parasite PDEs because it lacks the parasite PDE-specific P-pocket. Purified SmPDE4A proteins (full-length and catalytic domain) were used to profile an in-house library of PDE inhibitors (PDE4NPD toolbox). This screening identified tetrahydrophthalazinones and benzamides as potential hits. The PDE inhibitor NPD-0001 was the most active tetrahydrophthalazinone, whereas the approved human PDE4 inhibitors roflumilast and piclamilast were the most potent benzamides. As a follow-up, 83 benzamide analogs were prepared, but the inhibitory potency of the initial hits was not improved. Finally, NPD-0001 and roflumilast were evaluated in an in vitro anti-S. mansoni assay. Unfortunately, both SmPDE4A inhibitors were not effective in worm killing and only weakly affected the egg-laying at high micromolar concentrations. Consequently, the results with these SmPDE4A inhibitors strongly suggest that SmPDE4A is not a suitable target for anti-schistosomiasis therapy.


Subject(s)
Phosphodiesterase 4 Inhibitors , Schistosomiasis , Animals , Humans , Cyclic Nucleotide Phosphodiesterases, Type 4/genetics , Cyclic Nucleotide Phosphodiesterases, Type 4/metabolism , Schistosoma mansoni , Benzamides/pharmacology , Phosphodiesterase 4 Inhibitors/pharmacology , Schistosomiasis/drug therapy , Nucleotides, Cyclic
2.
J Med Chem ; 63(7): 3485-3507, 2020 04 09.
Article in English | MEDLINE | ID: mdl-32196340

ABSTRACT

Human African trypanosomiasis is causing thousands of deaths every year in the rural areas of Africa. In this manuscript we describe the optimization of a family of phtalazinone derivatives. Phosphodiesterases have emerged as attractive molecular targets for a novel treatment for a variety of neglected parasitic diseases. Compound 1 resulted in being a potent TbrPDEB1 inhibitor with interesting activity against T. brucei in a phenotypic screen. Derivative 1 was studied in an acute in vivo mouse disease model but unfortunately showed no efficacy due to low metabolic stability. We report structural modifications to achieve compounds with an improved metabolic stability while maintaining high potency against TbrPDEB1 and T. brucei. Compound 14 presented a good microsomal stability in mouse and human microsomes and provides a good starting point for future efforts.


Subject(s)
Phosphodiesterase Inhibitors/pharmacology , Phthalazines/pharmacology , Trypanocidal Agents/pharmacology , Animals , Crystallography, X-Ray , Drug Stability , Humans , Mice , Microsomes, Liver/metabolism , Molecular Structure , Parasitic Sensitivity Tests , Phosphodiesterase Inhibitors/chemical synthesis , Phosphodiesterase Inhibitors/metabolism , Phosphoric Diester Hydrolases/metabolism , Phthalazines/chemical synthesis , Phthalazines/metabolism , Protein Binding , Protozoan Proteins/metabolism , Structure-Activity Relationship , Trypanocidal Agents/chemical synthesis , Trypanocidal Agents/metabolism , Trypanosoma brucei brucei/drug effects
3.
J Antimicrob Chemother ; 75(4): 958-967, 2020 04 01.
Article in English | MEDLINE | ID: mdl-31860098

ABSTRACT

BACKGROUND: Chagas' disease, caused by the protozoan parasite Trypanosoma cruzi, needs urgent alternative therapeutic options as the treatments currently available display severe limitations, mainly related to efficacy and toxicity. OBJECTIVES: As phosphodiesterases (PDEs) have been claimed as novel targets against T. cruzi, our aim was to evaluate the biological aspects of 12 new phthalazinone PDE inhibitors against different T. cruzi strains and parasite forms relevant for human infection. METHODS: In vitro trypanocidal activity of the inhibitors was assessed alone and in combination with benznidazole. Their effects on parasite ultrastructural and cAMP levels were determined. PDE mRNA levels from the different T. cruzi forms were measured by quantitative reverse transcription PCR. RESULTS: Five TcrPDEs were found to be expressed in all parasite stages. Four compounds displayed strong effects against intracellular amastigotes. Against bloodstream trypomastigotes (BTs), three were at least as potent as benznidazole. In vitro combination therapy with one of the most active inhibitors on both parasite forms (NPD-040) plus benznidazole demonstrated a quite synergistic profile (xΣ FICI = 0.58) against intracellular amastigotes but no interaction (xΣ FICI = 1.27) when BTs were assayed. BTs treated with NPD-040 presented disrupted Golgi apparatus, a swollen flagellar pocket and signs of autophagy. cAMP measurements of untreated parasites showed that amastigotes have higher ability to efflux this second messenger than BTs. NPD-001 and NPD-040 increase the intracellular cAMP content in both BTs and amastigotes, which is also released into the extracellular milieu. CONCLUSIONS: The findings demonstrate the potential of PDE inhibitors as anti-T. cruzi drug candidates.


Subject(s)
Chagas Disease , Trypanocidal Agents , Trypanosoma cruzi , Chagas Disease/drug therapy , Humans , Phosphodiesterase Inhibitors/pharmacology , Phosphodiesterase Inhibitors/therapeutic use , Trypanocidal Agents/pharmacology , Trypanocidal Agents/therapeutic use
4.
Int J Mol Sci ; 20(9)2019 May 07.
Article in English | MEDLINE | ID: mdl-31067645

ABSTRACT

The caseinolytic protease proteolytic subunit (ClpP) is a serine protease playing an important role in proteostasis of eukaryotic organelles and prokaryotic cells. Alteration of ClpP function has been proved to affect the virulence and infectivity of a number of pathogens. Increased bacterial resistance to antibiotics has become a global problem and new classes of antibiotics with novel mechanisms of action are needed. In this regard, ClpP has emerged as an attractive and potentially viable option to tackle pathogen fitness without suffering cross-resistance to established antibiotic classes and, when not an essential target, without causing an evolutionary selection pressure. This opens a greater window of opportunity for the host immune system to clear the infection by itself or by co-administration with commonly prescribed antibiotics. A comprehensive overview of the function, regulation and structure of ClpP across the different organisms is given. Discussion about mechanism of action of this protease in bacterial pathogenesis and human diseases are outlined, focusing on the compounds developed in order to target the activation or inhibition of ClpP.


Subject(s)
Anti-Bacterial Agents/pharmacology , Bacterial Proteins/metabolism , Depsipeptides/pharmacology , Endopeptidase Clp/metabolism , Anti-Bacterial Agents/chemistry , Bacteria/drug effects , Bacteria/enzymology , Bacterial Proteins/agonists , Bacterial Proteins/chemistry , Depsipeptides/chemistry , Drug Design , Endopeptidase Clp/chemistry
5.
Article in English | MEDLINE | ID: mdl-30669086

ABSTRACT

We report the evaluation of 265 compounds from a PDE-focused library for their antischistosomal activity, assessed in vitro using Schistosoma mansoni. Of the tested compounds, 171 (64%) displayed selective in vitro activity, with 16 causing worm hypermotility/spastic contractions and 41 inducing various degrees of worm killing at 100 µM, with the surviving worms displaying sluggish movement, worm unpairing and complete absence of eggs. The compounds that did not affect worm viability (n = 72) induced a complete cessation of ovipositing. 82% of the compounds had an impact on male worms whereas female worms were barely affected. In vivo evaluation in S. mansoni-infected mice with the in vitro 'hit' NPD-0274 at 20 mg/kg/day orally for 5 days resulted in worm burden reductions of 29% and intestinal tissue egg load reduction of 35% at 10 days post-treatment. Combination of praziquantel (PZQ) at 10 mg/kg/day for 5 days with NPD-0274 or NPD-0298 resulted in significantly higher worm killing than PZQ alone, as well as a reduction in intestinal tissue egg load, disappearance of immature eggs and an increase in the number of dead eggs.


Subject(s)
Anthelmintics/pharmacology , Imidazoles/pharmacology , Schistosoma mansoni/drug effects , Small Molecule Libraries , 3',5'-Cyclic-AMP Phosphodiesterases/antagonists & inhibitors , Animals , Anthelmintics/chemistry , Drug Discovery , Fibroblasts/drug effects , High-Throughput Screening Assays , Humans , Imidazoles/chemistry , Male , Mice , Parasite Egg Count , Praziquantel/pharmacology
6.
J Med Chem ; 62(2): 774-797, 2019 01 24.
Article in English | MEDLINE | ID: mdl-30571121

ABSTRACT

Increased Gram-negative bacteria resistance to antibiotics is becoming a global problem, and new classes of antibiotics with novel mechanisms of action are required. The caseinolytic protease subunit P (ClpP) is a serine protease conserved among bacteria that is considered as an interesting drug target. ClpP function is involved in protein turnover and homeostasis, stress response, and virulence among other processes. The focus of this study was to identify new inhibitors of Escherichia coli ClpP and to understand their mode of action. A focused library of serine protease inhibitors based on diaryl phosphonate warheads was tested for ClpP inhibition, and a chemical exploration around the hit compounds was conducted. Altogether, 14 new potent inhibitors of E. coli ClpP were identified. Compounds 85 and 92 emerged as most interesting compounds from this study due to their potency and, respectively, to its moderate but consistent antibacterial properties as well as the favorable cytotoxicity profile.


Subject(s)
Endopeptidase Clp/antagonists & inhibitors , Escherichia coli Proteins/antagonists & inhibitors , Escherichia coli/enzymology , Organophosphonates/chemistry , Serine Proteinase Inhibitors/chemistry , Anti-Bacterial Agents/chemistry , Anti-Bacterial Agents/metabolism , Anti-Bacterial Agents/pharmacology , Binding Sites , Biphenyl Compounds/chemistry , Endopeptidase Clp/metabolism , Escherichia coli/drug effects , Escherichia coli Proteins/metabolism , Inhibitory Concentration 50 , Molecular Docking Simulation , Organophosphonates/metabolism , Organophosphonates/pharmacology , Protein Structure, Tertiary , Serine Proteinase Inhibitors/metabolism , Serine Proteinase Inhibitors/pharmacology , Structure-Activity Relationship
7.
Eur J Med Chem ; 151: 18-26, 2018 May 10.
Article in English | MEDLINE | ID: mdl-29604541

ABSTRACT

Human African trypanosomiasis is causing thousands of deaths every year in the rural areas of sub-saharan Africa. There is a high unmet medical need since the approved drugs are poorly efficacious, show considerable toxicity and are not easy to administer. This work describes the optimization of the pharmacokinetic properties of a previously published family of triazine lead compounds. One compound (35 (UAMC-03011)) with potent anti-trypanosomal activity and no cytotoxicity was selected for further study because of its good microsomal stability and high selectivity for Trypanosoma brucei over a panel including Trypanosoma cruzi, L.eishmania infantum, and Plasmodium falciparum. In vivo pharmacokinetic parameters were determined and the compound was studied in an acute in vivo mouse disease model. One of the important learnings of this study was that the rate of trypanocidal activity is an important parameter during the lead optimization process.


Subject(s)
Triazines/chemistry , Triazines/therapeutic use , Trypanocidal Agents/chemistry , Trypanocidal Agents/therapeutic use , Trypanosoma brucei brucei/drug effects , Trypanosomiasis, African/drug therapy , Animals , Disease Models, Animal , Humans , Mice , Structure-Activity Relationship , Triazines/pharmacokinetics , Triazines/pharmacology , Tropolone/analogs & derivatives , Tropolone/chemistry , Tropolone/pharmacokinetics , Tropolone/pharmacology , Tropolone/therapeutic use , Trypanocidal Agents/pharmacokinetics , Trypanocidal Agents/pharmacology
8.
Eur J Med Chem ; 143: 306-319, 2018 Jan 01.
Article in English | MEDLINE | ID: mdl-29197735

ABSTRACT

Human African trypanosomiasis (HAT), also known as sleeping sickness is a parasitic disease transmitted by the bite of the 'Glossina' insect, commonly known as the tsetse fly. This disease affects mostly poor populations living in remote rural areas of Africa. Untreated, it is usually fatal. Currently, safe and effective treatments against this disease are lacking. Phenotypic screening of triazine non-nucleoside HIV-1 reverse transcriptase inhibitors (monomers) resulted in potent and selective antitrypanosomal compounds. This serendipitous discovery and the presence of dimers in many compounds active against these neglected tropical diseases prompted us to investigate antitrypanosomal activity of triazine dimers. Optimization of the triazine dimers resulted in 3,3'-(((ethane-1,2-diylbis(azanediyl))bis(4-(mesityloxy)-1,3,5-triazine-6,2-diyl))bis(azanediyl))dibenzonitrile (compound 38), a compound with very potent in vitro and moderate in vivo antitrypanosomal activity.


Subject(s)
Antiprotozoal Agents/pharmacology , Reverse Transcriptase Inhibitors/pharmacology , Triazines/pharmacology , Trypanosoma brucei brucei/drug effects , Trypanosoma brucei rhodesiense/drug effects , Trypanosoma cruzi/drug effects , Animals , Antiprotozoal Agents/chemical synthesis , Antiprotozoal Agents/chemistry , Cell Proliferation/drug effects , Cell Survival/drug effects , Dimerization , Dose-Response Relationship, Drug , Drug Discovery , Fibroblasts/drug effects , Humans , Leishmania infantum/drug effects , Male , Mice , Molecular Structure , Reverse Transcriptase Inhibitors/chemical synthesis , Reverse Transcriptase Inhibitors/chemistry , Structure-Activity Relationship , Triazines/chemical synthesis , Triazines/chemistry
9.
Eur J Med Chem ; 138: 328-342, 2017 Sep 29.
Article in English | MEDLINE | ID: mdl-28688273

ABSTRACT

Leucine-rich repeat kinase 2 (LRRK2) is one of the most pursued targets for Parkinson's disease (PD) therapy. Moreover, it has recently described its role in regulating Wnt signaling and thus, it may be involved in adult neurogenesis. This new hypothesis could give rise to double disease-modifying agents firstly by the benefits of inhibiting LRRK2 and secondly by promoting adult neurogenesis. Herein we report, the design, synthesis, biological evaluation, SAR and potential binding mode of indoline-like LRRK2 inhibitors and their preliminary neurogenic effect in neural precursor cells isolated from adult mice ventricular-subventricular zone. These results open new therapeutic horizons for the use of LRRK2 inhibitors as neuroregenerative agents. Moreover, the indolinone derivatives here prepared, inhibitors of the kinase activity of LRRK2, may be considered as pharmacological probes to study the potential neuroregeneration of the damaged brain.


Subject(s)
Indoles/pharmacology , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/antagonists & inhibitors , Neural Stem Cells/drug effects , Neuroprotective Agents/pharmacology , Protein Kinase Inhibitors/pharmacology , Animals , Cell Differentiation/drug effects , Cell Proliferation/drug effects , Dose-Response Relationship, Drug , Humans , Indoles/chemical synthesis , Indoles/chemistry , Leucine-Rich Repeat Serine-Threonine Protein Kinase-2/metabolism , Mice , Mice, Inbred C57BL , Models, Molecular , Molecular Structure , Neuroprotective Agents/chemical synthesis , Neuroprotective Agents/chemistry , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/chemistry , Structure-Activity Relationship
10.
Future Med Chem ; 9(8): 731-748, 2017 05.
Article in English | MEDLINE | ID: mdl-28485668

ABSTRACT

AIM: Since neuroinflammation is partially mediated by cAMP levels and PDE10A enzyme is able to regulate these levels being highly expressed in striatum, its inhibitors emerged as useful drugs to mitigate this inflammatory process and hence the neuronal death associated with Parkinson's disease (PD). Methodology & results: To study the utility of PDE10A as a pharmacological target for PD, in this work we propose the search and development of new PDE10A inhibitors that could be useful as pharmacological tools in models of the disease and presumably as potential drug candidates. By using different medicinal chemistry approaches we have discovered imidazole-like PDE10A inhibitors and showed their neuroprotective actions. CONCLUSION: Here, we demonstrate the neuroprotective effect of PDE10A inhibitors in cellular models of PD. [Formula: see text].


Subject(s)
Imidazoles/pharmacology , Parkinson Disease/drug therapy , Phosphodiesterase Inhibitors/pharmacology , Phosphoric Diester Hydrolases/metabolism , Cell Line, Tumor , Cell Survival/drug effects , Humans , Imidazoles/chemical synthesis , Imidazoles/chemistry , Models, Molecular , Molecular Structure , Parkinson Disease/metabolism , Phosphodiesterase Inhibitors/chemical synthesis , Phosphodiesterase Inhibitors/chemistry
11.
ACS Omega ; 2(8): 5215-5220, 2017 Aug 31.
Article in English | MEDLINE | ID: mdl-30023743

ABSTRACT

Parkinson's disease (PD), an age-related neurodegenerative disorder that results from a progressive loss of dopaminergic neurons has an enormous economical and human cost. Unfortunately, only symptomatic treatment such as dopamine replacement therapy is available. Therefore, drugs with new mechanisms of action able to protect against neuronal cell death are an urgent need. We here report the in vivo efficacy on dopaminergic neuronal protection in a PD mouse model and the lack of toxicity in zebrafish and Ames test of benzothiazole-based casein kinase-1δ (CK-1δ) nanomolar inhibitors. On the basis of these results, we propose protein kinase CK-1δ inhibitors as the possible disease-modifying drugs for PD, benzothiazole 4 being a promising drug candidate for further development as a new therapy of this neurodegenerative disease.

12.
Mol Neurodegener ; 11(1): 36, 2016 04 30.
Article in English | MEDLINE | ID: mdl-27138926

ABSTRACT

BACKGROUND: Mutations in the progranulin gene (GRN) are the most common cause of frontotemporal lobar degeneration with TDP-43 inclusions (FTLD-TDP). TDP-43 pathology is characterized by the hyperphosphorylation of the protein at Serine 409/410 residues. Casein kinase-1δ (CK-1δ) was reported to phosphorylate TDP-43 directly. Previous works from our laboratory described the presence of CDK6/pRb-dependent cell cycle alterations, and cytosolic accumulation of TDP-43 protein in lymphoblast from FTLD-TDP patients carriers of a loss-of function mutation in GRN gene (c.709-1G > A). In this work, we have investigated the effects of two brain penetrant CK-1δ inhibitors (IGS-2.7 and IGS-3.27) designed and synthetized in our laboratory on cell proliferation, TDP-43 phosphorylation and subcellular localization, as well as their effects on the known nuclear TDP-43 function repressing the expression of CDK6. RESULTS: We report here that both CK-1δ inhibitors (IGS-2.7 and IGS-3.27) normalized the proliferative activity of PGRN-deficient lymphoblasts by preventing the phosphorylation of TDP-43 fragments, its nucleo-cytosol translocation and the overactivation of the CDK6/pRb cascade. Moreover, ours results show neuroprotective effects of CK-1δ inhibitors in a neuronal cell model of induced TDP-43 phosphorylation. CONCLUSIONS: Our results suggest that modulating CK-1δ activity could be considered a novel therapeutic approach for the treatment of FTLD-TDP and other TDP-43 proteinopathies.


Subject(s)
Casein Kinase Idelta/antagonists & inhibitors , Cell Proliferation/physiology , Frontotemporal Dementia/metabolism , Protein Kinase Inhibitors/pharmacology , TDP-43 Proteinopathies/metabolism , Casein Kinase Idelta/metabolism , Cells, Cultured , DNA-Binding Proteins/genetics , Frontotemporal Dementia/drug therapy , Frontotemporal Dementia/genetics , Humans , Intercellular Signaling Peptides and Proteins/genetics , Lymphocytes/metabolism , Mutation/genetics , Phosphorylation
13.
J Med Chem ; 57(6): 2755-72, 2014 Mar 27.
Article in English | MEDLINE | ID: mdl-24592867

ABSTRACT

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease where motor neurons in cortex, brain stem, and spinal cord die progressively, resulting in muscle wasting, paralysis, and death. Currently, effective therapies for ALS are lacking; however, identification of pathological TAR DNA-binding protein 43 (TDP-43) as the hallmark lesion in sporadic ALS suggests new therapeutic targets for pharmacological intervention. Pathological TDP-43 phosphorylation appears to drive the onset and progression of ALS and may result from upregulation of the protein kinase CK-1 in affected neurons, resulting in postranslational TDP-43 modification. Consequently, brain penetrant specific CK-1 inhibitors may provide a new therapeutic strategy for treating ALS and other TDP-43 proteinopathies. Using a chemical genetic approach, we report the discovery and further optimization of a number of potent CK-1δ inhibitors. Moreover, these small heterocyclic molecules are able to prevent TDP-43 phosphorylation in cell cultures, to increase Drosophila lifespan by reduction of TDP-43 neurotoxicity, and are predicted to cross the blood-brain barrier. Thus, N-(benzothiazolyl)-2-phenyl-acetamides are valuable drug candidates for further studies and may be a new therapeutic approach for ALS and others pathologies in which TDP-43 is involved.


Subject(s)
Amyotrophic Lateral Sclerosis/drug therapy , Casein Kinase I/antagonists & inhibitors , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/therapeutic use , Animals , Animals, Genetically Modified , Benzothiazoles/chemical synthesis , Benzothiazoles/pharmacology , Blood-Brain Barrier/drug effects , Blood-Brain Barrier/metabolism , Cell Membrane Permeability/drug effects , Cells, Cultured , DNA-Binding Proteins/metabolism , Drosophila , Drug Design , Drug Discovery , HEK293 Cells , Heterocyclic Compounds/chemical synthesis , Heterocyclic Compounds/pharmacology , High-Throughput Screening Assays , Humans , Magnetic Resonance Spectroscopy , Models, Molecular , Molecular Conformation , Neurons/drug effects , Neurons/enzymology , Neurotoxicity Syndromes/drug therapy , Phosphorylation , Substrate Specificity
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