Effects of Modified Glucosamine on the Chondrogenic Potential of Circulating Stem Cells under Experimental Inflammation.

Glucosamine (GlcN) is a glycosaminoglycan (GAGs) constituent in connective tissues. It is naturally produced by our body or consumed from diets. In the last decade, in vitro and in vivo trials have demonstrated that the administration of GlcN or its derivates has a protective effect on cartilage when the balance between catabolic and anabolic processes is disrupted and cells are no longer able to fully compensate for the loss of collagen and proteoglycans. To date, these benefits are still controversial because the mechanism of action of GlcN is not yet well clarified. In this study, we have characterized the biological activities of an amino acid (AA) derivate of GlcN, called DCF001, in the growth and chondrogenic induction of circulating multipotent stem cells (CMCs) after priming with tumor necrosis factor-alpha (TNFα), a pleiotropic cytokine commonly expressed in chronic inflammatory joint diseases. In the present work, stem cells were isolated from the human peripheral blood of healthy donors. After priming with TNFα (10 ng/mL) for 3 h, cultures were treated for 24 h with DCF001 (1 µg/mL) dissolved in a proliferative (PM) or chondrogenic (CM) medium. Cell proliferation was analyzed using a Corning® Cell Counter and trypan blue exclusion technique. To evaluate the potentialities of DCF001 in counteracting the inflammatory response to TNFα, we measured the amount of extracellular ATP (eATP) and the expression of adenosine-generating enzymes CD39/CD73, TNFα receptors, and NF-κB inhibitor IκBα using flow cytometry. Finally, total RNA was extracted to perform a gene expression study of some chondrogenic differentiation markers (COL2A1, RUNX2, and MMP13). Our analysis has shed light on the ability of DCF001 to (a) regulate the expression of CD39, CD73, and TNF receptors; (b) modulate eATP under differentiative induction; (c) enhance the inhibitory activity of IκBα, reducing its phosphorylation after TNFα stimulation; and (d) preserve the chondrogenic potentialities of stem cells. Although preliminary, these results suggest that DCF001 could be a valuable supplement for ameliorating the outcome of cartilage repair interventions, enhancing the efficacy of endogenous stem cells under inflammatory stimuli.

Inhibition of Leishmania infantum Trypanothione Reductase by New Aminopropanone Derivatives Interacting with the NADPH Binding Site.

BACKGROUND: As a result of the paucity of treatment, Leishmaniasis continues to provoke about 60,000 deaths every year worldwide. New molecules are needed, and drug discovery research is oriented toward targeting proteins crucial for parasite survival. Among them, trypanothione reductase (TR) is of remarkable interest owing to its vital role in Leishmania species protozoan parasite life. Our previously identified compound 1 is a novel chemotype endowed with a unique mode of TR inhibition thanks to its binding to a formerly unknown but druggable site at the entrance of the NADPH binding cavity, absent in human glutathione reductase (hGR). METHODS: We designed and synthesized new 3-amino-1-arylpropan-1-one derivatives structurally related to compound 1 and evaluated their potential inhibition activity on TR from Leishmania infantum (LiTR). Cluster docking was performed to assess the binding poses of the compounds. RESULTS: The newly synthesized compounds were screened at a concentration of 100 µM in in vitro assays and all of them proved to be active with residual activity percentages lower than 75%. CONCLUSIONS: Compounds 2a and 2b were the most potent inhibitors found, suggesting that an additional aromatic ring might be promising for enzymatic inhibition. Further structure-activity relationships are needed to optimize our compounds activity.

New Inhibitors of the Human p300/CBP Acetyltransferase Are Selectively Active against the Arabidopsis HAC Proteins.

Histone acetyltransferases (HATs) are involved in the epigenetic positive control of gene expression in eukaryotes. CREB-binding proteins (CBP)/p300, a subfamily of highly conserved HATs, have been shown to function as acetylases on both histones and non-histone proteins. In the model plant Arabidopsis thaliana among the five CBP/p300 HATs, HAC1, HAC5 and HAC12 have been shown to be involved in the ethylene signaling pathway. In addition, HAC1 and HAC5 interact and cooperate with the Mediator complex, as in humans. Therefore, it is potentially difficult to discriminate the effect on plant development of the enzymatic activity with respect to their Mediator-related function. Taking advantage of the homology of the human HAC catalytic domain with that of the Arabidopsis, we set-up a phenotypic assay based on the hypocotyl length of Arabidopsis dark-grown seedlings to evaluate the effects of a compound previously described as human p300/CBP inhibitor, and to screen previously described cinnamoyl derivatives as well as newly synthesized analogues. We selected the most effective compounds, and we demonstrated their efficacy at phenotypic and molecular level. The in vitro inhibition of the enzymatic activity proved the specificity of the inhibitor on the catalytic domain of HAC1, thus substantiating this strategy as a useful tool in plant epigenetic studies.

Ultrastructural Damages to H1N1 Influenza Virus Caused by Vapor Essential Oils.

Influenza viruses are transmitted from human to human via airborne droplets and can be transferred through contaminated environmental surfaces. Some works have demonstrated the efficacy of essential oils (EOs) as antimicrobial and antiviral agents, but most of them examined the liquid phases, which are generally toxic for oral applications. In our study, we describe the antiviral activity of Citrus bergamia, Melaleuca alternifolia, Illicium verum and Eucalyptus globulus vapor EOs against influenza virus type A. In the vapor phase, C. bergamia and M. alternifolia strongly reduced viral cytopathic effect without exerting any cytotoxicity. The E. globulus vapor EO reduced viral infection by 78% with no cytotoxicity, while I. verum was not effective. Furthermore, we characterized the EOs and their vapor phase by the head-space gas chromatography-mass spectrometry technique, observing that the major component found in each liquid EO is the same one of the corresponding vapor phases, with the exception of M. alternifolia. To deepen the mechanism of action, the morphological integrity of virus particles was checked by negative staining transmission electron microscopy, showing that they interfere with the lipid bilayer of the viral envelope, leading to the decomposition of membranes. We speculated that the most abundant components of the vapor EOs might directly interfere with influenza virus envelope structures or mask viral structures important for early steps of viral infection.

Recent Advances in Recovery of Lycopene from Tomato Waste: A Potent Antioxidant with Endless Benefits.

Growing attention to environmental protection leads food industries to adopt a model of "circular economy" applying safe and sustainable technologies to recover, recycle and valorize by-products. Therefore, by-products become raw material for other industries. Tomato processing industry produces significant amounts of by-products, consisting of skins and seeds. Tomato skin is very rich in lycopene, and from its seeds, high nutritional oil can be extracted. Alternative use of the two fractions not only could cut disposal costs but also allow one to extract bioactive compounds and an oil with a high nutritional value. This review focused on the recent advance in extraction of lycopene, whose beneficial effects on health are widely recognized.

Design, Synthesis and Biological Evaluation of New Pyrimidine Derivatives as Anticancer Agents.

BACKGROUND: Anticancer drug resistance is a challenging phenomenon of growing concern which arises from alteration in drug targets. Despite the fast speed of new chemotherapeutic agent design, the increasing prevalence of this phenomenon requires further research and treatment development. Recently, we reported a new aminopyrimidine compound-namely RDS 344-as a potential innovative anticancer agent. METHODS: Herein, we report the design, synthesis, and anti-proliferative activity of new aminopyrimidine derivatives structurally related to RDS 3442 obtained by carrying out substitutions at position 6 of the pyrimidine core and/or on the 2-aniline ring of our hit. The ability to inhibit cell proliferation was evaluated on different types of tumors, glioblastoma, triple-negative breast cancer, oral squamous cell carcinomas and colon cancer plus on human dermal fibroblasts chosen as control of normal cells. RESULTS: The most interesting compound was the N-benzyl counterpart of RDS 3442, namely 2a, that induced a significant decrease in cell viability in all the tested tumor cell lines, with EC50s ranging from 4 and 8 µM, 4-13 times more active of hit. CONCLUSIONS: These data suggest a potential role for this class of molecules as promising tool for new approaches in treating cancers of different histotype.

Discovery of a pyrimidine compound endowed with antitumor activity.

Recently, some synthetic nitrogen-based heterocyclic molecules, such as PJ34, have shown pronounced antitumor activity. Therefore, we designed and synthesized new derivatives characterized by a nitrogen-containing scaffold and evaluated their antiproliferative properties in tumor cells. We herein report the effects of three newly synthesized compounds on cell lines from three different human cancers: triple-negative breast cancer, colon carcinoma and glioblastoma. We found that two of these compounds did not affect proliferation, while the third significantly inhibited replication of the three cell lines. Moreover, this third molecule at 20 µM led to the upregulation of p21 and p27 and blockage of the cell cycle at G0/G1; in addition, it induced apoptosis in all three cell lines when used at higher concentrations (30-50 µM). The results demonstrate that this compound is a potent inhibitor of replication, an inducer of apoptosis and a negative regulator of cell cycle progression for cancer cells of different histotypes. Our data suggest a potential role for this new molecule as an interesting and powerful tool for new approaches in treating various cancers.

Structure-guided approach to identify a novel class of anti-leishmaniasis diaryl sulfide compounds targeting the trypanothione metabolism.

Leishmania protozoans are the causative agent of leishmaniasis, a neglected tropical disease consisting of three major clinical forms: visceral leishmaniasis (VL), cutaneous leishmaniasis, and mucocutaneous leishmaniasis. VL is caused by Leishmania donovani in East Africa and the Indian subcontinent and by Leishmania infantum in Europe, North Africa, and Latin America, and causes an estimated 60,000 deaths per year. Trypanothione reductase (TR) is considered to be one of the best targets to find new drugs against leishmaniasis. This enzyme is fundamental for parasite survival in the human host since it reduces trypanothione, a molecule used by the tryparedoxin/tryparedoxin peroxidase system of Leishmania to neutralize the hydrogen peroxide produced by host macrophages during infection. Recently, we solved the X-ray structure of TR in complex with the diaryl sulfide compound RDS 777 (6-(sec-butoxy)-2-((3-chlorophenyl)thio)pyrimidin-4-amine), which impairs the parasite defense against the reactive oxygen species by inhibiting TR with high efficiency. The compound binds to the catalytic site and engages in hydrogen bonds the residues more involved in the catalysis, namely Glu466', Cys57 and Cys52, thereby inhibiting the trypanothione binding. On the basis of the RDS 777-TR complex, we synthesized structurally related diaryl sulfide analogs as TR inhibitors able to compete for trypanothione binding to the enzyme and to kill the promastigote in the micromolar range. One of the most active among these compounds (RDS 562) was able to reduce the trypanothione concentration in cell of about 33% via TR inhibition. RDS 562 inhibits selectively Leishmania TR, while it does not inhibit the human homolog glutathione reductase.

Inhibition of Polycomb Repressive Complex 2 activity reduces trimethylation of H3K27 and affects development in Arabidopsis seedlings.

BACKGROUND: Polycomb repressive complex 2 (PRC2) is an epigenetic transcriptional repression system, whose catalytic subunit (ENHANCER OF ZESTE HOMOLOG 2, EZH2 in animals) is responsible for trimethylating histone H3 at lysine 27 (H3K27me3). In mammals, gain-of-function mutations as well as overexpression of EZH2 have been associated with several tumors, therefore making this subunit a suitable target for the development of selective inhibitors. Indeed, highly specific small-molecule inhibitors of EZH2 have been reported. In plants, mutations in some PRC2 components lead to embryonic lethality, but no trial with any inhibitor has ever been reported. RESULTS: We show here that the 1,5-bis (3-bromo-4-methoxyphenyl)penta-1,4-dien-3-one compound (RDS 3434), previously reported as an EZH2 inhibitor in human leukemia cells, is active on the Arabidopsis catalytic subunit of PRC2, since treatment with the drug reduces the total amount of H3K27me3 in a dose-dependent fashion. Consistently, we show that the expression level of two PRC2 targets is significantly increased following treatment with the RDS 3434 compound. Finally, we show that impairment of H3K27 trimethylation in Arabidopsis seeds and seedlings affects both seed germination and root growth. CONCLUSIONS: Our results provide a useful tool for the plant community in investigating how PRC2 affects transcriptional control in plant development.

Inhibition of Leishmania infantum trypanothione reductase by diaryl sulfide derivatives.

The study presented here aimed at identifying a new class of compounds acting against Leishmania parasites, the causative agent of Leishmaniasis. For this purpose, the thioether derivatives of our in-house library have been evaluated in whole-cell screening assays in order to determine their in vitro activity against Leishmania protozoan. Among them, promising results have been achieved with compound RDS 777 (6-(sec-butoxy)-2-((3-chlorophenyl)thio)pyrimidin-4-amine) (IC50 = 29.43 µM), which is able to impair the mechanism of the parasite defence against the reactive oxygen species by inhibiting the trypanothione reductase (TR) with high efficiency (Ki 0.25 ± 0.18 µM). The X-ray structure of L. infantum TR in complex with RDS 777 disclosed the mechanism of action of this compound that binds to the catalytic site and engages in hydrogen bonds the residues more involved in the catalysis, namely Glu466', Cys57 and Cys52, thereby inhibiting the trypanothione binding and avoiding its reduction.

New Thiazolidine-4-One Derivatives as SARS-CoV-2 Main Protease Inhibitors.

It has been more than four years since the first report of SARS-CoV-2, and humankind has experienced a pandemic with an unprecedented impact. Moreover, the new variants have made the situation even worse. Among viral enzymes, the SARS-CoV-2 main protease (Mpro) has been deemed a promising drug target vs. COVID-19. Indeed, Mpro is a pivotal enzyme for viral replication, and it is highly conserved within coronaviruses. It showed a high extent of conservation of the protease residues essential to the enzymatic activity, emphasizing its potential as a drug target to develop wide-spectrum antiviral agents effective not only vs. SARS-CoV-2 variants but also against other coronaviruses. Even though the FDA-approved drug nirmatrelvir, a Mpro inhibitor, has boosted the antiviral therapy for the treatment of COVID-19, the drug shows several drawbacks that hinder its clinical application. Herein, we report the synthesis of new thiazolidine-4-one derivatives endowed with inhibitory potencies in the micromolar range against SARS-CoV-2 Mpro. In silico studies shed light on the key structural requirements responsible for binding to highly conserved enzymatic residues, showing that the thiazolidinone core acts as a mimetic of the Gln amino acid of the natural substrate and the central role of the nitro-substituted aromatic portion in establishing π-π stacking interactions with the catalytic His-41 residue.

Synergistic Effects of Caffeine in Combination with Conventional Drugs: Perspectives of a Drug That Never Ages.

Plants have been known since ancient times for their healing properties, being used as preparations against human diseases of different etiologies. More recently, natural products have been studied and characterized, isolating the phytochemicals responsible for their bioactivity. Most certainly, there are currently numerous active compounds extracted from plants and used as drugs, dietary supplements, or sources of bioactive molecules that are useful in modern drug discovery. Furthermore, phytotherapeutics can modulate the clinical effects of co-administered conventional drugs. In the last few decades, the interest has increased even more in studying the positive synergistic effects between plant-derived bioactives and conventional drugs. Indeed, synergism is a process where multiple compounds act together to exert a merged effect that is greater than that of each of them summed together. The synergistic effects between phytotherapeutics and conventional drugs have been described in different therapeutic areas, and many drugs are based on synergistic interactions with plant derivatives. Among them, caffeine has shown positive synergistic effects with different conventional drugs. Indeed, in addition to their multiple pharmacological activities, a growing body of evidence highlights the synergistic effects of caffeine with different conventional drugs in various therapeutic fields. This review aims to provide an overview of the synergistic therapeutic effects of caffeine and conventional drugs, summarizing the progress reported to date.

Role of a Novel Heparanase Inhibitor on the Balance between Apoptosis and Autophagy in U87 Human Glioblastoma Cells.

BACKGROUND: Heparanase (HPSE) is an endo-ß-glucuronidase that cleaves heparan sulfate side chains, leading to the disassembly of the extracellular matrix, facilitating cell invasion and metastasis dissemination. In this research, we investigated the role of a new HPSE inhibitor, RDS 3337, in the regulation of the autophagic process and the balance between apoptosis and autophagy in U87 glioblastoma cells. METHODS: After treatment with RDS 3337, cell lysates were analyzed for autophagy and apoptosis-related proteins by Western blot. RESULTS: We observed, firstly, that LC3II expression increased in U87 cells incubated with RDS 3337, together with a significant increase of p62/SQSTM1 levels, indicating that RDS 3337 could act through the inhibition of autophagic-lysosomal flux of LC3-II, thereby leading to accumulation of lipidated LC3-II form. Conversely, the suppression of autophagic flux could activate apoptosis mechanisms, as revealed by the activation of caspase 3, the increased level of cleaved Parp1, and DNA fragmentation. CONCLUSIONS: These findings support the notion that HPSE promotes autophagy, providing evidence that RDS 3337 blocks autophagic flux. It indicates a role for HPSE inhibitors in the balance between apoptosis and autophagy in U87 human glioblastoma cells, suggesting a potential role for this new class of compounds in the control of tumor growth progression.

Diketo acid inhibitors of nsp13 of SARS-CoV-2 block viral replication.

For RNA viruses, RNA helicases have long been recognized to play critical roles during virus replication cycles, facilitating proper folding and replication of viral RNAs, therefore representing an ideal target for drug discovery. SARS-CoV-2 helicase, the non-structural protein 13 (nsp13) is a highly conserved protein among all known coronaviruses, and, at the moment, is one of the most explored viral targets to identify new possible antiviral agents. In the present study, we present six diketo acids (DKAs) as nsp13 inhibitors able to block both SARS-CoV-2 nsp13 enzymatic functions. Among them four compounds were able to inhibit viral replication in the low micromolar range, being active also on other human coronaviruses such as HCoV229E and MERS CoV. The experimental investigation of the binding mode revealed ATP-non-competitive kinetics of inhibition, not affected by substrate-displacement effect, suggesting an allosteric binding mode that was further supported by molecular modelling calculations predicting the binding into an allosteric conserved site located in the RecA2 domain.

Pyrrolyl and Indolyl α-γ-Diketo Acid Derivatives Acting as Selective Inhibitors of Human Carbonic Anhydrases IX and XII.

Solid tumors are active tissues containing hypoxic regions and producing metabolic acids. By decreasing pH, cancer cells create a hostile environment for surrounding host cells and foster tumor growth and progression. By governing acid/base regulation, carbonic anhydrases (CAs) are involved in several physiological/pathological processes, including tumors. Indeed, CAs are clinically relevant in cancer therapy as among the fifteen human isoforms, two of them, namely CA IX (overexpressed in solid tumors and associated with increased metastasis and poor prognosis) and CA XII (overexpressed in some tumors) are involved in tumorigenesis. Targeting these two isoforms is considered as a pertinent approach to develop new cancer therapeutics. Several CA inhibitors (CAIs) have been described, even though they are unselective inhibitors of different isoforms. Thus, efforts are needed to find new selective CAIs. In this work, we described new diketo acid derivatives as CAIs, with the best acting compounds 1c and 5 as nanomolar inhibitors of CA IX and XII, being also two orders of magnitude selective over CAs I and II. Molecular modeling studies showed the different binding poses of the best acting CAIs within CA II and IX, highlighting the key structural features that could confer the ability to establish specific interactions within the enzymes. In different tumor cell lines overexpressing CA IX and XII, the tested compounds showed antiproliferative activity already at 24 h treatment, with no effects on somatic not transformed cells.

Miconazole-like Scaffold is a Promising Lead for Naegleria fowleri-Specific CYP51 Inhibitors.

Developing drugs for brain infection by Naegleria fowleri is an unmet medical need. We used a combination of cheminformatics, target-, and phenotypic-based drug discovery methods to identify inhibitors that target an essential N. fowleri enzyme, sterol 14-demethylase (NfCYP51). A total of 124 compounds preselected in silico were tested against N. fowleri. Nine primary hits with EC50 ≤ 10 µM were phenotypically identified. Cocrystallization with NfCYP51 focused attention on one primary hit, miconazole-like compound 2a. The S-enantiomer of 2a produced a 1.74 Å cocrystal structure. A set of analogues was then synthesized and evaluated to confirm the superiority of the S-configuration over the R-configuration and the advantage of an ether linkage over an ester linkage. The two compounds, S-8b and S-9b, had an improved EC50 and KD compared to 2a. Importantly, both were readily taken up into the brain. The brain-to-plasma distribution coefficient of S-9b was 1.02 ± 0.12, suggesting further evaluation as a lead for primary amoebic meningoencephalitis.

Oxidative stress and visual system: a review.

Different types of tissues respond differently to the action of oxidative stress. The visual system is very sensitive to oxidative action due to continuous exposure to light. In consideration of the growing interest of scientific studies towards various compounds endowed with antioxidant and anti-inflammatory properties, we performed a review of the literature focusing on the use of some antioxidant molecules for the treatment of conditions affecting the visual system. In this study, we focused on the ability of two antioxidant agents, the small molecule α-lipoic acid (ALA) and the enzyme superoxide dismutase (SOD), to influence the neurodegenerative physiological processes related to aging and oxidative stress affecting the ocular segment. The literature data report that ALA and SOD can protect against neurodegenerative effects both the optic nerve and retina and, if administered together, they are able to lower the levels of oxidative stress, thus preventing neurodegeneration and reducing the apoptotic process.

Design, Synthesis, and In Vitro, In Silico and In Cellulo Evaluation of New Pyrimidine and Pyridine Amide and Carbamate Derivatives as Multi-Functional Cholinesterase Inhibitors.

Alzheimer disease is an age-linked neurodegenerative disorder representing one of the greatest medical care challenges of our century. Several drugs are useful in ameliorating the symptoms, even if none could stop or reverse disease progression. The standard approach is represented by the cholinesterase inhibitors (ChEIs) that restore the levels of acetylcholine (ACh) by inhibiting the acetylcholinesterase (AChE). Still, their limited efficacy has prompted researchers to develop new ChEIs that could also reduce the oxidative stress by exhibiting antioxidant properties and by chelating the main metals involved in the disease. Recently, we developed some derivatives constituted by a 2-amino-pyrimidine or a 2-amino-pyridine moiety connected to various aromatic groups by a flexible amino-alkyl linker as new dual inhibitors of AChE and butyrylcholinesterase (BChE). Following our previous studies, in this work we explored the role of the flexible linker by replacing the amino group with an amide or a carbamic group. The most potent compounds showed higher selectivity against BChE in respect to AChE, proving also to possess a weak anti-aggregating activity toward Aß42 and tau and to be able to chelate Cu2+ and Fe3+ ions. Molecular docking and molecular dynamic studies proposed possible binding modes with the enzymes. It is noteworthy that these compounds were predicted as BBB-permeable and showed low cytotoxicity on the human brain cell line.

Acetylcholinesterase inhibitors for the treatment of Alzheimer's disease - a patent review (2016-present).

Introduction - AD, the most common form of dementia, has a multifactorial etiology, and the current therapy (AChEIs and memantine) is unable to interrupt its progress and fatal outcome. This is reflected in the research programs that are oriented toward the development of new therapeutics able to operate on multiple targets involved in the disease progression.Areas covered - The patents from 2016 to present regarding the use of AChEIs in AD, concerns the development of new AChEIs, multitarget or multifunctional ligands, or the associations of currently used AChEIs with other compounds acting on different targets involved in the AD.Expert opinion - The development of new multitarget AChEIs promises to identify compounds with great therapeutic potential but requires more time and effort in order to obtain drugs with the optimal pharmacodynamic profile. Otherwise, the research on new combinations of existing drugs, with known pharmacodynamic and ADME profile, could shorten the time and reduce the costs to develop a new therapeutic treatment for AD. From the analyzed data, it seems more likely that a response to the urgent need to develop effective treatments for AD therapy could come more quickly from studies on drug combinations than from the development of new AChEIs.

New Pyrimidine and Pyridine Derivatives as Multitarget Cholinesterase Inhibitors: Design, Synthesis, and In Vitro and In Cellulo Evaluation.

A new series of pyrimidine and pyridine diamines was designed as dual binding site inhibitors of cholinesterases (ChEs), characterized by two small aromatic moieties separated by a diaminoalkyl flexible linker. Many compounds are mixed or uncompetitive acetylcholinesterase (AChE) and/or butyrylcholinesterase (BChE) nanomolar inhibitors, with compound 9 being the most active on Electrophorus electricus AChE (EeAChE) (Ki = 0.312 µM) and compound 22 on equine BChE (eqBChE) (Ki = 0.099 µM). Molecular docking and molecular dynamic studies confirmed the interaction mode of our compounds with the enzymatic active site. UV-vis spectroscopic studies showed that these compounds can form complexes with Cu2+ and Fe3+ and that compounds 18, 20, and 30 have antioxidant properties. Interestingly, some compounds were also able to reduce Aß42 and tau aggregation, with compound 28 being the most potent (22.3 and 17.0% inhibition at 100 µM on Aß42 and tau, respectively). Moreover, the most active compounds showed low cytotoxicity on a human brain cell line and they were predicted as BBB-permeable.