Casein kinase 1 inhibitor avoids TDP-43 pathology propagation in a patient-derived cellular model of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis is a fatal neurodegenerative disease without a cure to reverse its progression. Its main hallmark is the nuclear protein TDP-43, which undergoes different post-translational modifications leading to a loss of function in the nucleus and an increase in toxicity in the cytoplasm. Previous reports have indicated that pathogenic TDP-43 exhibits prion-like propagation in various contexts. With the aim of advancing therapeutics focused on preventing the propagation of TDP-43 pathology, we studied the potential role of pathogenic TDP-43 in lymphoblasts from sporadic ALS patients. We used lymphoblastoid cell lines from sporadic ALS patients as a source of pathogenic forms of TDP-43, and healthy human cells (lymphoblasts, myoblasts, neuroblastoma SH-SY5Y, or osteosarcoma U2OS) as recipient cells to investigate the seeding and spread of TDP-43 proteinopathy. Furthermore, we evaluated the potential of targeting TDP-43 phosphorylation with a CK-1 inhibitor to prevent the propagation of the pathology. The results presented herein indicate that pathogenic forms of TDP-43 are secreted into the extracellular medium of sporadic ALS lymphoblasts and could be transported by extracellular vesicles, spreading TDP-43 pathology to healthy cells. Moreover, tunneling nanotubes have also been discovered in pathological cells and may be involved in the transport of TDP-43. Interestingly, targeting TDP-43 phosphorylation with an in-house designed CK-1 inhibitor (IGS2.7) was sufficient to halt TDP-43 pathology transmission, in addition to its known effects on restoring the homeostasis of TDP-43 protein in patients-derived cells.

Druggable cavities and allosteric modulators of the cell division cycle 7 (CDC7) kinase.

Cell division cycle 7 kinase (CDC7) has been found overexpressed in many cancer cell lines being also one of the kinases involved in the nuclear protein TDP-43 phosphorylation in vivo. Thus, inhibitors of CDC7 are emerging drug candidates for the treatment of oncological and neurodegenerative unmet diseases. All the known CDC7 inhibitors are ATP-competitives, lacking of selectivity enough for success in clinical trials. As allosteric sites are less conserved among kinase proteins, discovery of allosteric modulators of CDC7 is a great challenge and opportunity in this field.Using different computational approaches, we have here identified new druggable cavities on the human CDC7 structure and subsequently selective CDC7 inhibitors with allosteric modulation mainly targeting the pockets where the interaction between this kinase and its activator DBF4 takes place.

Neuroprotective properties of exosomes and chitosan nanoparticles of Tomafran, a bioengineered tomato enriched in crocins.

Saffron has many pharmacological properties in addition to being a frequently used food seasoning. Crocin and picrocrocin which accumulate in saffron stigma, are responsible for these pharmacological properties. These natural products have health-promoting effects for the prevention and treatment of numerous diseases, including age-related cognitive and memory disfunction. Currently, crocin and picrocrocin are obtained from saffron, considered as the spice with the highest price in the market. To develop an efficient and low-cost approach to producing these compounds with high bioactivity, biosynthetic genes isolated from saffron can be exploited in the metabolic engineering of heterologous hosts and the production of crocins in productive crop plants. Recently, we engineered tomato fruit producing crocins (Tomafran). In this study, we demonstrated that crocin-rich extract, encapsulated in chitosan or in exosomes may function as a neuroprotective strategy. Crocins contained in the Tomafran extracts and much lower doses in chitosan nanoparticles or exosomes were enough to rescue the neuroblastoma cell line SH-SY5Y after damage caused by okadaic acid. Our results confirm the neuroprotective effect of Tomafran and its exosomes that may be useful for the delay or prevention of neurodegenerative disorders such as Alzheimer's disease.

Audiological evaluation (128-20,000Hz) in women with autoimmune thyroiditis: The role of antibodies vs. l-thyroxine deficiency.

BACKGROUND AND AIM: Although sensorineural hearing loss may have different aetiologies, we focused on autoimmune hearing loss since it may be reversible with corticosteroid therapy; this entity is sometimes associated with systemic autoimmune diseases. Hashimoto's thyroiditis or chronic autoimmune thyroiditis shows antibodies and may be harmful to hearing thresholds regardless of hypothyroidism effect. To date this effect has not been sufficiently studied and never with extended high frequencies. The aim of this work is to study by age groups whether hearing thresholds in the human auditory range (128-20,000Hz) are affected in Hashimoto's disease. MATERIALS AND METHODS: Two groups of 128 patients affected by Hashimoto's thyroiditis were included. First group: patients with pathological antithyroid antibodies who do not need L-thyroxine treatment. Second group: patients controlled with L-thyroxine substitutive treatment. Audiometric threshold study comparing between the groups of patients and a group of 209 controls was performed. All patients underwent complete otorhinolaryngological examination, antithyroid antibodies, TSH, T3 and T4 blood levels, tympanometry, conventional pure-tone audiometry, and extended-high-frequency audiometry. RESULTS: All patients were women. Both groups showed worst audiometric thresholds than the control group; both study groups showed worse hearing than controls, this difference was statistically significant in all frequencies. In the 8-20kHz frequency range, this difference was more than 10dB, and in the 9-16kHz and 20kHz range this difference was more than 20dB. When separated by age groups, in younger subjects (20-29 years) these differences were found in all frequencies, except for conversational frequencies (500-4,000Hz); between 30 and 49 years the difference is statistically significant in all frequencies; and from 50 to 69 years differences are found, especially in the conversational frequencies. CONCLUSIONS: This first work studying the human auditory range in the chronic autoimmune thyroiditis or Hashimoto's thyroiditis confirms that hearing loss related to the autoimmune disorder predominates at extended-high-frequencies initially. But ends up involving all frequencies in pure-tone conventional audiometry, then it may be detected in routine clinical tests. These results support the role of extended-high-frequencies audiometry to diagnose subclinical hearing loss in patients affected by Hashimoto's thyroiditis.

Network pharmacology reveals multitarget mechanism of action of drugs to be repurposed for COVID-19.

The coronavirus disease 2019 pandemic accelerated drug/vaccine development processes, integrating scientists all over the globe to create therapeutic alternatives against this virus. In this work, we have collected information regarding proteins from SARS-CoV-2 and humans and how these proteins interact. We have also collected information from public databases on protein-drug interactions. We represent this data as networks that allow us to gain insights into protein-protein interactions between both organisms. With the collected data, we have obtained statistical metrics of the networks. This data analysis has allowed us to find relevant information on which proteins and drugs are the most relevant from the network pharmacology perspective. This method not only allows us to focus on viral proteins as the main targets for COVID-19 but also reveals that some human proteins could be also important in drug repurposing campaigns. As a result of the analysis of the SARS-CoV-2-human interactome, we have identified some old drugs, such as disulfiram, auranofin, gefitinib, suloctidil, and bromhexine as potential therapies for the treatment of COVID-19 deciphering their potential complex mechanism of action.

Phenotypic Characterization of Laryngospasm: The Utility of Laryngeal Neurophysiological Studies.

OBJECTIVE: To determine the existence of laryngeal neuropathy and the influence of gastroesophageal reflux in patients with episodic laryngospasm. STUDY DESIGN: Single -center, observational study with prospective clinical collection. STUDY SETTING: Prospective analysis of 18 Spanish patients with episodic laryngospasm in a Spanish Public Center collected by otolaryngologist. The recruitment dates were from January 2019 to December 2019. METHODS: Data collection of 18 patients with episodic laryngospasm. Clinical characteristics, laryngeal neurophysiological studies and reflux esophageal testing were analyzed. RESULTS: All patients have a trigger for the laryngospasms, being the most prevalent adopting the supine position (27.7%). EMG and ENG were pathological in 83.3 and 63.6 % respectively. Chronic bilateral denervation (increased amplitude and duration of potentials) with signs of reinnervation in the non-active chronic phase (large polyphasia), was the most prevalent finding. Evidence of gastroesophageal reflux either by pH meter, Gastroscopy or both was found in 38.8 % of patients. CONCLUSIONS: Neurophysiological studies have confirmed the existence of laryngeal neuropathy in the majority of patients with laryngospasm. A substantial percentage of patients (38.8%) with laryngospasm had objective GERD and improved with PPIs. Laryngeal EMG and ENG can establish a more accurate diagnostic for episodic laryngospams and may supports treatment with neuromodulators.

TDP-43 Modulation by Tau-Tubulin Kinase 1 Inhibitors: A New Avenue for Future Amyotrophic Lateral Sclerosis Therapy.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease without any effective treatment. Protein TDP-43 is a pathological hallmark of ALS in both sporadic and familiar patients. Post-translational modifications of TDP-43 promote its aggregation in the cytoplasm. Tau-Tubulin kinase (TTBK1) phosphorylates TDP-43 in cellular and animal models; thus, TTBK1 inhibitors emerge as a promising therapeutic strategy for ALS. The design, synthesis, biological evaluation, kinase-ligand complex structure determination, and molecular modeling studies confirmed novel pyrrolopyrimidine derivatives as valuable inhibitors for further development. Moreover, compound 29 revealed good brain penetration in vivo and was able to reduce TDP-43 phosphorylation not only in cell cultures but also in the spinal cord of transgenic TDP-43 mice. A shift to M2 anti-inflammatory microglia was also demonstrated in vivo. Both these activities led to motor neuron preservation in mice, proposing pyrrolopyrimidine 29 as a valuable lead compound for future ALS therapy.

Improved Controlled Release and Brain Penetration of the Small Molecule S14 Using PLGA Nanoparticles.

Phosphodiesterase 7 (PDE7) is an enzyme responsible for the degradation of cyclic adenosine monophosphate (cAMP), an important cellular messenger. PDE7's role in neurotransmission, expression profile in the brain and the druggability of other phosphodiesterases have motivated the search for potent inhibitors to treat neurodegenerative and inflammatory diseases. Different heterocyclic compounds have been described over the years; among them, phenyl-2-thioxo-(1H)-quinazolin-4-one, called S14, has shown very promising results in different in vitro and in vivo studies. Recently, polymeric nanoparticles have been used as new formulations to target specific organs and produce controlled release of certain drugs. In this work, we describe poly(lactic-co-glycolic acid) (PLGA)-based polymeric nanoparticles loaded with S14. Their preparation, optimization, characterization and in vivo drug release profile are here presented as an effort to improve pharmacokinetic properties of this interesting PDE7 inhibitor.

Understanding the Molecular Mechanism of miR-877-3p Could Provide Potential Biomarkers and Therapeutic Targets in Squamous Cell Carcinoma of the Cervix.

No therapeutic targets and molecular biomarkers are available in cervical cancer (CC) management. In other cancer types, micro-RNA-877-3p (miR-877-3p) has been associated with events relevant for CC development. Thus, we aimed to determine miR-877-3p role in CC. miR-877-3p levels were examined by quantitative-PCR in 117 cervical lesions and tumors. Effects on CC cell proliferation, migration, and invasion were evaluated upon anti-miR-877-3p transfection. miR-877-3p dependent molecular mechanism was comprehensively explored by proteomics, dual-luciferase reporter assay, western blot, and immunohistochemistry. Cervical tumors expressed higher miR-877-3p levels than benign lesions. miR-877-3p promoted CC cell migration and invasion, at least partly by modulating cytoskeletal protein folding through the chaperonin-containing T-complex protein 1 complex. Notably, miR-877-3p silencing synergized with paclitaxel. Interestingly, miR-877-3p downregulated the levels of an in silico-predicted target, ZNF177, whose expression and subcellular location significantly distinguished high-grade squamous intraepithelial lesions (HSILs) and squamous cell carcinomas of the cervix (SCCCs). Cytoplasmic ZNF177 was significantly associated with worse progression-free survival in SCCC. Our results suggest that: (i) miR-877-3p is a potential therapeutic target whose inhibition improves paclitaxel effects; (ii) the expression and location of its target ZNF177 could be diagnostic biomarkers between HSIL and SCCC; and (iii) cytoplasmic ZNF177 is a poor-prognosis biomarker in SCCC.

Identification of potential inhibitors of protein-protein interaction useful to fight against Ebola and other highly pathogenic viruses.

Despite the efforts to develop new treatments against Ebola virus (EBOV) there is currently no antiviral drug licensed to treat patients with Ebola virus disease (EVD). Therefore, there is still an urgent need to find new drugs to fight against EBOV. In order to do this, a virtual screening was done on the druggable interaction between the EBOV glycoprotein (GP) and the host receptor NPC1 with a subsequent selection of compounds for further validation. This screening led to the identification of new small organic molecules with potent inhibitory action against EBOV infection using lentiviral EBOV-GP-pseudotype viruses. Moreover, some of these compounds have shown their ability to interfere with the intracellular cholesterol transport receptor NPC1 using an ELISA-based assay. These preliminary results pave the way to hit to lead optimization programs that lead to successful candidates.

Discovery of Amoebicidal Compounds by Combining Computational and Experimental Approaches.

Pathogenic and opportunistic free-living amoebae such as Acanthamoeba spp. can cause keratitis (Acanthamoeba keratitis [AK]), which may ultimately lead to permanent visual impairment or blindness. Acanthamoeba can also cause rare but usually fatal granulomatous amoebic encephalitis (GAE). Current therapeutic options for AK require a lengthy treatment with nonspecific drugs that are often associated with adverse effects. Recent developments in the field led us to target cAMP pathways, specifically phosphodiesterase. Guided by computational tools, we targeted the Acanthamoeba phosphodiesterase RegA. Computational studies led to the construction and validation of a homology model followed by a virtual screening protocol guided by induced-fit docking and chemical scaffold analysis using our medicinal and biological chemistry (MBC) chemical library. Subsequently, 18 virtual screening hits were prioritized for further testing in vitro against Acanthamoeba castellanii, identifying amoebicidal hits containing piperidine and urea imidazole cores. Promising activities were confirmed in the resistant cyst form of the amoeba and in additional clinical Acanthamoeba strains, increasing their therapeutic potential. Mechanism-of-action studies revealed that these compounds produce apoptosis through reactive oxygen species (ROS)-mediated mitochondrial damage. These chemical families show promise for further optimization to produce effective antiacanthamoebal drugs.

Targeting nuclear protein TDP-43 by cell division cycle kinase 7 inhibitors: A new therapeutic approach for amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with no known cure. Aggregates of the nuclear protein TDP-43 have been recognized as a hallmark of proteinopathy in both familial and sporadic cases of ALS. Post-translational modifications of this protein, include hyperphosphorylation, cause disruption of TDP-43 homeostasis and as a consequence, promotion of its neurotoxicity. Among the kinases involved in these changes, cell division cycle kinase 7 (CDC7) plays an important role by directly phosphorylating TDP-43. In the present manuscript the discovery, synthesis, and optimization of a new family of selective and ATP-competitive CDC7 inhibitors based on 6-mercaptopurine scaffold are described. Moreover, we demonstrate the ability of these inhibitors to reduce TDP-43 phosphorylation in both cell cultures and transgenic animal models such as C. elegans and Prp-hTDP43 (A315T) mice. Altogether, the compounds described here may be useful as versatile tools to explore the role of CDC7 in TDP-43 phosphorylation and also as new drug candidates for the future development of ALS therapies.

Antibiofilm activity of flavonoids on staphylococcal biofilms through targeting BAP amyloids.

The opportunistic pathogen Staphylococcus aureus is responsible for causing infections related to indwelling medical devices, where this pathogen is able to attach and form biofilms. The intrinsic properties given by the self-produced extracellular biofilm matrix confer high resistance to antibiotics, triggering infections difficult to treat. Therefore, novel antibiofilm strategies targeting matrix components are urgently needed. The Biofilm Associated Protein, Bap, expressed by staphylococcal species adopts functional amyloid-like structures as scaffolds of the biofilm matrix. In this work we have focused on identifying agents targeting Bap-related amyloid-like aggregates as a strategy to combat S. aureus biofilm-related infections. We identified that the flavonoids, quercetin, myricetin and scutellarein specifically inhibited Bap-mediated biofilm formation of S. aureus and other staphylococcal species. By using in vitro aggregation assays and the cell-based methodology for generation of amyloid aggregates based on the Curli-Dependent Amyloid Generator system (C-DAG), we demonstrated that these polyphenols prevented the assembly of Bap-related amyloid-like structures. Finally, using an in vivo catheter infection model, we showed that quercetin and myricetin significantly reduced catheter colonization by S. aureus. These results support the use of polyphenols as anti-amyloids molecules that can be used to treat biofilm-related infections.

Cognitive enhancement, TAU phosphorylation reduction, and neuronal protection by the treatment of an LRRK2 inhibitor in a tauopathy mouse model.

Leucine-rich repeat kinase 2 (LRRK2) is a protein kinase whose activity plays an important role in neurodegenerative diseases. Although mutations in LRRK2 gene are the most common cause of monogenic Parkinson's disease, it has been reported that LRRK2 may promote Tau phosphorylation, increasing its aggregation. Thus, the modulation of LRRK2 activity by small molecules able to inhibit this kinase activity could be an innovative therapeutic strategy for different tauopathies. We examined the therapeutic effects of a new benzothiazole-based LRRK2 inhibitor, known as JZ1.40, in a mouse model of tauopathy. Mice were injected in the right hippocampus with an adeno-associated vector expressing human-TAUP301L and treated daily with JZ1.40 (10 mg/kg, i.p) or vehicle for three weeks. JZ1.40 reaches the brain and modulates RAB10 and Tau phosphorylation at the epitopes modified by LRRK2. Moreover, JZ1.40 treatment ameliorates the cognitive impairment induced by TAUP301L overexpression, which correlates with prevention of granular cell layer degeneration by improving synaptic plasticity. These data show that JZ1.40 is neuroprotective in vivo, which is translated into cognition enhancement.

Surgical technique modifies the postoperative atrioventricular block rate in sutureless prostheses.

BACKGROUND: The main objective was to analyse the impact of the modification of the Perceval S implantation technique on the prevalence of postoperative atrioventricular block, which requires a permanent pacemaker, in our aortic valve replacement series. In addition, we attempted to identify those risk factors that are related to the appearance of this complication. METHODS: Five hundred and seventy-two valve replacements were carried out with PERCEVAL S in our centre up to July 2018. Use of modified technique (n=302). Minimally invasive approach (n=340). Associated coronary surgery (n=95). Patients with pacemakers prior to surgery (n=27) and associated mitral or tricuspid valve surgery (n=26) were excluded. We analysed variables of interest that could influence the increase in postoperative atrioventricular block. Technique performed, disorders of intraventricular conduction and pre/intraoperative characteristics. The influence of the modified technique was analysed. RESULTS: Five hundred and nineteen aortic valve replacements with PERCEVAL S. Age (years) (median 77, interquartile range 8). Height (cm) (159, 13.5). Euroscore II (%) (2.25, 2.27). Postoperative atrioventricular block standard technique (n=23, 10.14%). Modified technique (n=14, 4.30%) (P=0.009). Multivariate regression analysis. Final model AUC =0.740, maximum model AUC =0.774 (P>0.05). Includes: Technique used (P=0.024), height (P=0.043) and disorders of interventricular conduction, right bundle branch block (P=0.005), trifascicular block (P=0.008). CONCLUSIONS: In our experience, the modified technique significantly decreases the incidence of postoperative atrioventricular block that requires a permanent pacemaker in the aortic valve replacement with PERCEVAL S. The prior electrocardiographic presence of right bundle branch block, trifascicular block and the height of the patient are associated with an increased risk of blocking.

Analysis of the PD-1/PD-L1 axis in human autoimmune thyroid disease: Insights into pathogenesis and clues to immunotherapy associated thyroid autoimmunity.

Autoimmune thyroid diseases (AITDs), i.e., Graves' disease (GD) and Hashimoto thyroiditis (HT), are the most prevalent organ-specific autoimmune diseases, but their pathogenesis is still incompletely understood. The PD-1/PD-L1 pathway is an important mechanism of peripheral tolerance that has not been investigated in AITDs. Here, we report the analysis of the expression of PD-1, PD-L1 and PD-L2 in PBMCs, infiltrating thyroid lymphocytes (ITLs) and in thyroid follicular cells (TFCs) in GD, HT and multinodular goiter (MNG) patients and healthy controls PBMCs (HC). By combining flow cytometry, tissue immunofluorescence and induction experiments on primary and thyroid cell line cultures, we show that: 1) while PD-1+ T cells are moderately expanded in PBMCs from GD vs HC, approximately half of T cells in the infiltrate are PD-1+ including some PD-1hi; 2) PD-L1, but not PD-L2, is expressed by 81% of GD glands and in 25% of non-autoimmune glands; 3) PD-L1, was expressed by TFCs in areas that also contain abundant PD-1 positive T cells but; 4) co-localization in TFCs indicated only partial overlap between the smaller areas of the PD-L1+ and the larger areas of HLA class II+ expression; 5) IFNγ is capable of inducing PD-L1 in >90% of TFCs in primary cultures and cell lines. Collectively these results indicate that the PD-1/PD-L1 axis is operative in AITD glands and may restrain the autoimmune response. Yet the discrepancy between easy induction in vitro and the limited expression in vivo (compared to HLA) suggests that PD-L1 expression in vivo is partially inhibited in GD and HT glands. In conclusions 1) the PD-1/PD-L1 pathway is activated in AITD glands but probably not to the extent to inhibit disease progression and 2) Thyroid autoimmunity arising after PD-1/PD-L1 blocking therapies in cancer patients may result from interfering PD-1/PD-L1 tolerance mechanism in thyroid with minimal (focal) thyroiditis. Finally acting on the PD-1/PD-L1 pathway could be a new approach to treat AITD and other organ-specific autoimmunity in the future.

Screening of a PDE-focused library identifies imidazoles with in vitro and in vivo antischistosomal activity.

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.

l-Thyroxine does not prevent immunemediated sensorineural hearing loss in autoimmune thyroid diseases.

OBJECTIVE: This is the first report dealing with immune-mediated inner ear disease (IMIED) hearing loss in a group of patients affected with autoimmune thyroid disease (AITD), whose treatment required corticosteroids, despite being treated with levothyroxine. Immunopathology linking the inner ear and the thyroid gland is also presented. PATIENTS: A total of 220 patients were selected with sensorineural hearing loss (SNHL) of causes other than presbycusis. Audiometry was performed and pure tone average was calculated before and after treatment with corticosteroids. RESULTS: Eighty-four (84) patients had SNHL of autoimmune origin, and 15 patients were diagnosed with AITD (Hashimoto's disease). Bilateral hearing loss was observed in 10 patients (66.5%). Sudden sensorineural hearing loss was the most frequent clinical form of presentation. Nine patients showed a hearing recovery greater than 10dB after corticosteroid treatment. CONCLUSIONS: Acquired hypothyroidism is thought to affect hearing due to different mechanisms. Although specific hormonal therapy may improve peripheral or central auditory disorders associated with hypothyroidism, the presence of IMIED in AITD patients requires another approach. Altered immune regulatory mechanisms involving Treg cells and CD4+CD45RO cells have been suggested in patients with AITD and IMIED. In the present study, although all the patients with hypothyroidism and subclinical hypothyroidism were being treated with levothyroxine, immune-mediated hearing loss was observed. Therapy with corticosteroids could achieve hearing recovery. Since inner ear and thyroid gland share possible antigen targets, we highlight the existence of IMIED in AITD patients and the importance of implementing appropriate therapy with corticosteroids.

Cyclic Nucleotide-Specific Phosphodiesterases as Potential Drug Targets for Anti-Leishmania Therapy.

The available treatments for leishmaniasis are less than optimal due to inadequate efficacy, toxic side effects, and the emergence of resistant strains, clearly endorsing the urgent need for discovery and development of novel drug candidates. Ideally, these should act via an alternative mechanism of action to avoid cross-resistance with the current drugs. As cyclic nucleotide-specific phosphodiesterases (PDEs) of Leishmania major have been postulated as putative drug targets, a series of potential inhibitors of Leishmania PDEs were explored. Several displayed potent and selective in vitro activity against L. infantum intracellular amastigotes. One imidazole derivative, compound 35, was shown to reduce the parasite loads in vivo and to increase the cellular cyclic AMP (cAMP) level at in a dose-dependent manner at just 2× and 5× the 50% inhibitory concentration (IC50), indicating a correlation between antileishmanial activity and increased cellular cAMP levels. Docking studies and molecular dynamics simulations pointed to imidazole 35 exerting its activity through PDE inhibition. This study establishes for the first time that inhibition of cAMP PDEs can potentially be exploited for new antileishmanial chemotherapy.

Amyloid ß-induced impairments on mitochondrial dynamics, hippocampal neurogenesis, and memory are restored by phosphodiesterase 7 inhibition.

BACKGROUND: The phosphodiesterase (PDE) 7 inhibitor S14 is a cell-permeable small heterocyclic molecule that is able to cross the blood-brain barrier. We previously found that intraperitoneal treatment with S14 exerted neuroprotection in an Alzheimer's disease (AD) model (in APP/PS1 mice). The objective of this study was to investigate the neurogenic and cellular effects of oral administration of S14 on amyloid ß (Aß) overload. METHODS: We orally administered the PDE7 inhibitor S14 (15 mg/kg/day) or vehicle in 6-month-old APP/PS1 mice. After 5 weeks of S14 treatment, we evaluated cognitive functions and brain tissues. We also assessed the effects of S14 on the Aß-treated human neuroblastome SH-SY5Y cell line. RESULTS: Targeting the cyclic adenosine monophosphate (cAMP)/cAMP-response element binding protein (CREB) pathway, S14 rescued cognitive decline by improving hippocampal neurogenesis in APP/PS1 transgenic mice. Additionally, S14 treatment reverted the Aß-induced reduction in mitochondrial mass in APP/PS1 mice and in the human neuroblastoma SH-SY5Y cells co-exposed to Aß. The restoration of the mitochondrial mass was found to be a dual effect of S14: a rescue of the mitochondrial biogenesis formerly slowed down by Aß overload, and a reduction in the Aß-increased mitochondrial clearance mechanism of mitophagy. CONCLUSIONS: Here, we show new therapeutic effects of the PDE7 inhibitor, confirming S14 as a potential therapeutic drug for AD.