Synthesis, activity, metabolic stability and cell permeability of new cytosolic phospholipase A2α inhibitors with 1-indolyl-3-phenoxypropan-2-one structure.

Cytosolic phospholipase A2α (cPLA2α), the key enzyme of the arachidonic acid cascade, is considered to be an interesting target for the development of new anti-inflammatory drugs. Potent inhibitors of the enzyme include indole-5-carboxylic acids with propan-2-one residues in position 1 of the indole. Previously, it was found that central pharmacophoric elements of these compounds are their ketone and carboxylic acid groups, which unfortunately are subject to pronounced metabolism by carbonyl reductases and glucuronosyltransferases, respectively. Here we show that the metabolic stability of these inhibitors can be improved by introducing alkyl substituents in the vicinity of the ketone group or by increasing their rigidity. Furthermore, permeability tests with Caco-2 cells revealed that the indole derivatives have only low permeability, which can be attributed to their affinity to efflux transporters. Among other things, the polar ketone group in the center of the molecules seems to be a decisive factor for their reverse transport. After its removal, the permeability increased significantly. The enhancement in metabolic stability and permeability achieved by the structural variations carried out was accompanied by a more or less pronounced decrease in the inhibitory potency of the compounds against cPLA2α.

Synthesis and pharmacokinetic properties of novel cPLA2α inhibitors with 1-(carboxyalkylpyrrolyl)-3-aryloxypropan-2-one structure.

Indole-5-carboxylic acids with 3-aryloxy-2­oxopropyl residues in position 1 have been shown to be potent inhibitors of cytosolic phospholipase A2α (cPLA2α), an enzyme involved in the formation of pro-inflammatory lipid mediators. Unfortunately, in animal experiments, only very low plasma concentrations could be achieved after peroral administration of this type of compound. Since insufficient metabolic stability was suspected as the cause, structural modifications were made to optimize this property. These included the conversion of the aromatic into an aliphatic carboxylic acid function as well as the rigidification of the lipophilic structural elements. A selected pyrrole-3-propionic acid was tested for its peroral in vivo bioavailability in mice. However, higher plasma concentrations could not be achieved also with this compound. Using the Caco2 cell permeation assay, substances investigated were found to be very good substrates for gastrointestinal efflux transporters, which explains their poor peroral absorption.

A FRET-Based Assay for the Identification of PCNA Inhibitors.

Proliferating cell nuclear antigen (PCNA) is the key regulator of human DNA metabolism. One important interaction partner is p15, involved in DNA replication and repair. Targeting the PCNA-p15 interaction is a promising therapeutic strategy against cancer. Here, a Förster resonance energy transfer (FRET)-based assay for the analysis of the PCNA-p15 interaction was developed. Next to the application as screening tool for the identification and characterization of PCNA-p15 interaction inhibitors, the assay is also suitable for the investigation of mutation-induced changes in their affinity. This is particularly useful for analyzing disease associated PCNA or p15 variants at the molecular level. Recently, the PCNA variant C148S has been associated with Ataxia-telangiectasia-like disorder type 2 (ATLD2). ATLD2 is a neurodegenerative disease based on defects in DNA repair due to an impaired PCNA. Incubation time dependent FRET measurements indicated no effect on PCNAC148S-p15 affinity, but on PCNA stability. The impaired stability and increased aggregation behavior of PCNAC148S was confirmed by intrinsic tryptophan fluorescence, differential scanning fluorimetry (DSF) and asymmetrical flow field-flow fractionation (AF4) measurements. The analysis of the disease associated PCNA variant demonstrated the versatility of the interaction assay as developed.

Improved Reversion of Calcifications in Porcine Aortic Heart Valves Using Elastin-Targeted Nanoparticles.

Calcified aortic valve disease in its final stage leads to aortic valve stenosis, limiting cardiac function. To date, surgical intervention is the only option for treating calcific aortic valve stenosis. This study combined controlled drug delivery by nanoparticles (NPs) and active targeting by antibody conjugation. The chelating agent diethylenetriaminepentaacetic acid (DTPA) was covalently bound to human serum albumin (HSA)-based NP, and the NP surface was modified using conjugating antibodies (anti-elastin or isotype IgG control). Calcification was induced ex vivo in porcine aortic valves by preincubation in an osteogenic medium containing 2.5 mM sodium phosphate for five days. Valve calcifications mainly consisted of basic calcium phosphate crystals. Calcifications were effectively resolved by adding 1-5 mg DTPA/mL medium. Incubation with pure DTPA, however, was associated with a loss of cellular viability. Reversal of calcifications was also achieved with DTPA-coupled anti-elastin-targeted NPs containing 1 mg DTPA equivalent. The addition of these NPs to the conditioned media resulted in significant regression of the valve calcifications compared to that in the IgG-NP control without affecting cellular viability. These results represent a step further toward the development of targeted nanoparticular formulations to dissolve aortic valve calcifications.

Discrete spatio-temporal regulation of tyrosine phosphorylation directs influenza A virus M1 protein towards its function in virion assembly.

Small RNA viruses only have a very limited coding capacity, thus most viral proteins have evolved to fulfill multiple functions. The highly conserved matrix protein 1 (M1) of influenza A viruses is a prime example for such a multifunctional protein, as it acts as a master regulator of virus replication whose different functions have to be tightly regulated. The underlying mechanisms, however, are still incompletely understood. Increasing evidence points towards an involvement of posttranslational modifications in the spatio-temporal regulation of M1 functions. Here, we analyzed the role of M1 tyrosine phosphorylation in genuine infection by using recombinant viruses expressing M1 phosphomutants. Presence of M1 Y132A led to significantly decreased viral replication compared to wildtype and M1 Y10F. Characterization of phosphorylation dynamics by mass spectrometry revealed the presence of Y132 phosphorylation in M1 incorporated into virions that is most likely mediated by membrane-associated Janus kinases late upon infection. Molecular dynamics simulations unraveled a potential phosphorylation-induced exposure of the positively charged linker domain between helices 4 and 5, supposably acting as interaction platform during viral assembly. Consistently, M1 Y132A showed a defect in lipid raft localization due to reduced interaction with viral HA protein resulting in a diminished structural stability of viral progeny and the formation of filamentous particles. Importantly, reduced M1-RNA binding affinity resulted in an inefficient viral genome incorporation and the production of non-infectious virions that interferes with virus pathogenicity in mice. This study advances our understanding of the importance of dynamic phosphorylation as a so far underestimated level of regulation of multifunctional viral proteins and emphasizes the potential feasibility of targeting posttranslational modifications of M1 as a novel antiviral intervention.

Preparation of Sesquiterpene Lactone-Loaded PLA Nanoparticles and Evaluation of Their Antitrypanosomal Activity.

Human African trypanosomiasis (HAT), also commonly known as sleeping sickness, is a neglected tropical disease affecting millions of people in poorly developed regions in sub-Saharan Africa. There is no satisfactory treatment for this infection. The investment necessary to bring new drugs to the market is a big deterrent to drug development, considering that the affected communities form a non-lucrative sector. However, natural products and many sesquiterpene lactones (STLs) in particular are very strong trypanocides. Research and applications of nano-drug delivery systems such as nanoparticles (NPs) have undergone unprecedented growth in the recent past. This is mainly due to the advantages offered by these systems, such as targeted delivery of the drug to the place of action (cell, parasite, etc), sustained release of the drug, increased bioavailability, reduced drug dosage, and reduction of undesired side effects, among others. In this study, the STLs α-santonin, arglabin, schkuhrin II, vernolepin, and eucannabinolide, all trypanocides, were loaded into polylactic acid (PLA) NPs through an emulsification-diffusion method. The NPs were stable, homogenous, and spherical in shape with a rounded knotty depression like a navel orange. The average particle sizes were 202.3, 220.3, 219.5, 216.9, and 226.4 nm for α-santonin, arglabin, schkuhrin II, vernolepin, and eucannabinolide, respectively. The NPs had encapsulation efficiencies of 94.6, 78.1, 76.8, 60.7, and 78.9% for α-santonin, arglabin, schkuhrin II, vernolepin, and eucannabinolide, respectively. The NPs loaded with arglabin, vernolepin, and eucannabinolide exhibited considerable antitrypanosomal activity against Trypanosoma brucei rhodesiense (Tbr) with free drug equivalent IC50 values of 3.67, 1.11 and 3.32 µM, respectively. None of the NP formulations displayed cytotoxicity towards mammalian cells (rat skeletal myoblast cell line L6). These results provide new insights into the possibility of incorporating STLs into nanoparticles, which may provide new options for their formulation in order to develop new drugs against HAT.

Use of Light-Degradable Aliphatic Polycarbonate Nanoparticles As Drug Carrier for Photosensitizer.

Aliphatic poly(carbonate)s (APCs) with rapid and controlled degradation upon specific stimulation have great advantages for a variety of biomedical and pharmaceutical applications. In the present work, we reported a new poly(trimethylene carbonate) (PTMC)-based copolymer containing multiple 4,5-dimethoxy-2-nitrobenzyl photo cleavable groups as pendent chains. The six-membered light-responsive cyclic carbonate monomer (LrM) was first prepared from 2-(hydroxymethyl)-2-methylpropane-1,3-diol and 4,5-dimethoxy-2-nitrobenzyl alcohol and then copolymerized with trimethylene carbonate (TMC) by 1,8-diazabicyclo(5.4.0)undec-7-ene (DBU) catalyzed ring-opening polymerization (ROP) to afford the light-responsive polycarbonate (LrPC). The light-triggered decomposition of LrM and LrPC was studied by NMR, UV/vis spectroscopy, and size exclusion chromatography (SEC), as well as ESI-ToF mass spectrometry. Stable monodisperse nanoparticles with hydrodynamic diameter of 100 nm could be formulated from 25% LrPC and 75% poly(lactide- co-glycolide) (PLGA) and applied for the encapsulation of temoporfin. Upon irradiation with UV light these particles displayed a significant decrease of the particle countrate and increased the release rate of temoporfin in comparison to standard PLGA nanoparticles. This work demonstrated that a combination of encapsulation of photosensitizer and light degradation using light-responsive polymers is suitable to enhance photodynamic therapy (PDT).

Ligand-Modified Human Serum Albumin Nanoparticles for Enhanced Gene Delivery.

The development of nonviral gene delivery systems is a great challenge to enable safe gene therapy. In this study, ligand-modified nanoparticles based on human serum albumin (HSA) were developed and optimized for an efficient gene therapy. Different glutaraldehyde cross-linking degrees were investigated to optimize the HSA nanoparticles for gene delivery. The peptide sequence arginine-glycine-aspartate (RGD) and the HIV-1 transactivator of transduction sequence (Tat) are well-known as promising targeting ligands. Plasmid DNA loaded HSA nanoparticles were covalently modified on their surface with these different ligands. The transfection potential of the obtained plasmid DNA loaded RGD- and Tat-modified nanoparticles was investigated in vitro, and optimal incubation conditions for these preparations were studied. It turned out that Tat-modified HSA nanoparticles with the lowest cross-linking degree of 20% showed the highest transfection potential. Taken together, ligand-functionalized HSA nanoparticles represent promising tools for efficient and safe gene therapy.

Reaction of human macrophages on protein corona covered TiO2 nanoparticles.

The cytokine secretion of primary cells of human macrophages during the interaction of TiO2 nanoparticles (with an average primary size of 100-120 nm) is investigated down to concentration levels suggested to be relevant for in vivo conditions. We find that high TiO2 concentrations induce the cytokines Interleukin IL-1ß, IL-6, and IL-10 secretion, while at low concentrations only IL-6 secretion is observed. To obtain further evidence on in vivo conditions we investigated the development and structure of the protein corona of the nanoparticles. We demonstrated that the surface of TiO2 particles attract preferably secondary modified proteins which then induce cytokine secretion of macrophages. Our results indicate that concentration of corona covered TiO2 particles below 1 µg/ml induce IL-6 secretion which is reported to be responsible for the development of autoimmune diseases as well as for the secretion of acute phase proteins. FROM THE CLINICAL EDITOR: This study investigates the effects of protein corona covered titanium dioxide nanoparticles on human macrophages, concluding that concentration of such particles below 1 µg/ml induces IL-6 secretion, which may be responsible for the development of autoimmune diseases as well as for the secretion of acute phase proteins. This finding has important implications on future applications of such nanoparticles.

Covalent modification of human serum albumin by the natural sesquiterpene lactone parthenolide.

The reactivity of parthenolide (PRT), a natural sesquiterpene lactone from Tanacetum parthenium (Asteraceae), with human serum albumin (HSA) was studied by UHPLC/+ESI-QqTOF MS analysis after tryptic digestion of albumin samples after incubation with this compound. It was found that the single free cysteine residue, C34, of HSA (0.6 mM) reacted readily with PRT when incubated at approximately 13-fold excess of PRT (8 mM). Time-course studies with PRT and its 11ß,13-dihydro derivative at equimolar ratios of the reactants revealed that PRT under the chosen conditions reacts preferably with C34 and does so exclusively via its α-methylene-γ-lactone moiety, while the epoxide structure is not involved in the reaction.

1-Benzylindoles as inhibitors of cytosolic phospholipase A2α: synthesis, biological activity, aqueous solubility, and cell permeability.

Cytosolic phospholipase A2α (cPLA2α) is considered an interesting target for the development of new anti-inflammatory drugs, as it is significantly involved in the formation of pro-inflammatory lipid mediators. Recently, in a ligand-based virtual screening approach, 2,4-dichlorobenzyl-substituted 4-[2-(indol-3-ylmethylene)hydrazineyl]benzoic acid 7 was found to be an inhibitor of cPLA2α with micromolar activity. This compound has now been systematically varied to increase inhibitory potency. The studies performed led to 5-(1-benzylindol-3-ylmethyl)-2H-tetrazol-2-yl)pentanoic acid derivatives that exhibited submicromolar activity against the enzyme. The most potent compounds were also tested for their water solubility and for permeability in a Caco-2 model. Among other things, it was found that in Caco-2 cells, the pentanoic acid chain of the molecules can be metabolised to a considerable extent to propionic acid by ß-oxidation.

Development of a screening platform for the formulation of poorly water-soluble drugs as albumin-stabilized nanosuspensions using nab™ technology.

The nanoparticle albumin bound™ (nab™) technology generally offers great potential for the formulation of poorly water-soluble drugs as albumin-stabilized nanosuspensions for intravenous use while avoiding solubilizers and cross-linking agents. The nab™ technology is a three-step process consisting of emulsification, high-pressure homogenization and solvent evaporation. Within this work, a screening approach was developed to predict whether active pharmaceutical ingredients are suitable for nab™ formulations. A design of experiments approach was used to investigate the effects of ultrasonic homogenization on an albumin-stabilized itraconazole nanosuspension. Based on this, a screening platform was developed, and subsequently evaluated and applied to a selection of poorly water-soluble drugs. The screening process to produce albumin-stabilized nanosuspensions consists of two process steps: Ultrasonic treatment, which combined emulsification and homogenization, followed by solvent evaporation. The results of the screening process were fully transferable to the standard three-step process of nab™ technology. In addition, based on drug screening, drug properties were highlighted that are important for the development of nab™ formulations. All in all, the nab™ technology is a promising but not universal formulation platform for poorly water-soluble drugs. Nevertheless, for some poorly soluble drugs it offers a valuable approach for the formulation of nanosuspensions for intravenous use.

Challenges of nanoparticle albumin bound (nab™) technology: Comparative study of Abraxane® with a newly developed albumin-stabilized itraconazole nanosuspension.

Nanoparticle albumin bound™ (nab™) technology is an established delivery platform for development of albumin stabilized nanoparticles as drug delivery systems for poorly water-soluble drugs. By using albumin for particle stabilization, nab™ technology does not require solubilizers or emulsifiers for the formulation of poorly water-soluble drugs for intravenous use. Despite the great potential, however, to date only two products based on nab™ technology have been approved by the Food and Drug Administration: Abraxane® (nab™ paclitaxel) and Fyarro® (nab™ rapamycin). In this study, the commercially available product Abraxane® was characterized in comparison to an albumin stabilized nanosuspension for the poorly water-soluble drug itraconazole. The aim of this study was to identify critical product parameters of the nanosuspensions depending on the manufacturing process in order to assess the transferability of nab™ technology to other drugs. The colloidal properties, stabilizing protein composition and particle disintegration behavior were analyzed. In addition, studies were carried out on the impact of the key process step, the high-pressure homogenization, using a design of experiments (DoE) approach. A nanosuspension comprising spherical, stable drug nanoparticles stabilized by a large fraction of dissolved albumin around the nanoparticles were identified. During the manufacturing process, the drug core was coated with a layer of albumin, which was cross-linked to a certain level. The Abraxane® and itraconazole suspensions differed in the analyzed protein fraction, with stronger cross-linking at the particle surface for Abraxane®. Both active pharmaceutical ingredients were present in the amorphous state as nanoparticles. In vitro disintegration studies performed to mimic a strong dilution during intravenous application showed the disintegration of the nanoparticles. All in all, the analysis underlined the transferability of the nab™ technology to selected other poorly water-soluble drugs with the great advantage of eliminating solubilizers and emulsifiers for intravenous applications.

Triglyceride-filled albumin-based nanocapsules: A promising new system to avoid discarding poorly water-soluble drug candidates.

Even though current drug discovery provides a variety of potential drug candidates, many of those substances are difficult to formulate due to their poor water-solubility. To overcome this obstacle a technological formulation is crucial. Albumin-based nanocarriers are a possible intravenous delivery system which is already approved and commercially available. However, no universal carrier for poorly water-soluble substances is found yet. In the present study, new preparation processes for nanocapsules consisting of a medium-chain triglyceride (MCT) core and a human serum albumin (HSA) shell were developed. The nanocarrier system exhibits desirable physicochemical properties with a hydrodynamic diameter of 150 nm and a polydispersity index of 0.1. Furthermore, the nanocapsules were stable towards the addition of electrolytes and also in basic to neutral pH range. The nanocapsules were storage stable for at least 7 months at 4 °C and could also be lyophilized to reach an even longer shelf life of at least 21 months. In addition, the nanocapsule system showed no cytotoxicity in cell culture. The developed system represents a suitable carrier for a variety of different poorly water-soluble drug substances (e.g., fenofibrate, naproxen, indomethacin) showing a high potential for a universal formulation platform for further lipophilic active pharmaceutical ingredients (APIs).

Effects of Generic Exchange of Levodopa Medication in Patients With Parkinson Disease.

OBJECTIVES: Generic exchange is common practice in most healthcare systems. This study investigated how patients with Parkinson disease (PD) perceived a switch of their levodopa medication and the resulting effects on their PD symptoms. METHODS: A questionnaire was developed, piloted, and finally distributed to 13,857 members of the national PD patient support group. It was designed to be completed by patients and their pharmacies. χ 2 tests for independence statistics with or without Monte Carlo simulation were performed. Cramér φ and Cramér V were calculated. McNemar test was used to investigate whether a generic switch of a levodopa-containing medication had an impact on PD symptoms. RESULTS: Analyses were done with 410 finalized respondents of 13,857 distributed questionnaires. More than half of the responders were 75 years or older and rated themselves Hoehn and Yahr stages 3 to 5. Most patients were confused by a change of their medication. A total of 54.7% of the switchers (n = 148) reported swallowing difficulties with medication, which was significantly more frequent than with nonswitchers (37.3% of 204, P = 0.001). Adverse effects related to the switch were reported by 26.6% of all switchers (switchback rate, 20.5%). The patients at higher Hoehn and Yahr stages were affected to a greater extend. CONCLUSIONS: Patients, who experienced any brand switch of their levodopa medication, frequently expressed distrust and confusion. Swallowing difficulties and negative effects on their symptoms were problems, which were more pronounced in advanced disease stages. It remains unclear whether the detrimental impact on therapy was caused by the brand switch or by a nocebo effect.

Different dissolution conditions affect stability and dissolution profiles of bioequivalent levodopa-containing oral dosage forms.

Parkinson's disease is frequently treated with combinations of levodopa/carbidopa and, at the occurrence of motor fluctuations, levodopa/carbidopa/entacapone. For these (and other) medications, using generic versions can reduce costs. To show that generic drugs are equivalent to the originator drug, regulations usually refer to the bioavailability of active ingredients, which is influenced by the selected dosage form and the chosen excipients. However, while registration trials administer drugs under standardized conditions, these conditions often do not reflect the conditions of patients' daily intake. Thus, this study aimed to characterize levodopa combinations from different manufacturers in biorelevant media. Dissolution profiles of bioequivalent levodopa/carbidopa combinations and levodopa/carbidopa/entacapone combinations were tested in different media, such as tap water, gastric fluid without pepsin and whole milk. Results showed distinct discrepancies in the drugs' dissolution profiles between manufactures. Using whole milk as a dissolution medium led to the most differing dissolution profiles. Furthermore, carbidopa was unstable in tap water and milk, and it rapidly degraded. This effect was less pronounced if entacapone was present. In contrast to reports in the literature, stability testing did not show that vitamin C helps to protect against carbidopa degradation. Entacapone hardly dissolved in an acidic environment. This study found that dissolution of bioequivalent levodopa formulations varied with changing media. Further, the stability of carbidopa was found to be critical. As an implication, an acidic environment must be ensured when these drugs are applied, and generic exchange of levodopa combinations should be considered only with great caution.

Permeability of dopamine D2 receptor agonist hordenine across the intestinal and blood-brain barrier in vitro.

Hordenine, a bioactive food compound, has several pharmacological properties and has recently been identified as a dopamine D2 receptor (D2R) agonist. Since the pharmacokinetic profile of hordenine has been described to a limited extent, the present study focused on the transfer and transport of hordenine across the intestinal epithelium and the blood-brain barrier (BBB) in vitro. Hordenine was quickly transferred through the Caco-2 monolayer in only a few hours, indicating a rapid oral uptake. However, the high bioavailability may be reduced by the observed efflux transport of hordenine from the bloodstream back into the intestinal lumen and by first pass metabolism in intestinal epithelial cells. To determine the biotransformation rate of hordenine, the metabolite hordenine sulfate was synthesized as reference standard for analytical purposes. In addition, transfer studies using primary porcine brain capillary endothelial cells (PBCEC) showed that hordenine is able to rapidly penetrate the BBB and potentially accumulate in the brain. Thus, a D2R interaction of hordenine and activation of dopaminergic signaling is conceivable, assuming that the intestinal barrier can be circumvented by a route of administration alternative to oral uptake.

Spacer length and serum protein adsorption affect active targeting of trastuzumab-modified nanoparticles.

Receptor-mediated active targeting of nanocarriers is a widely investigated approach to specifically address cancerous cells and tissues in the human body. The idea is to use these formulations as drug carriers with enhanced specificity and therefore reduced systemic side effects. Until today a big obstacle to reach this goal remains the adsorption of serum proteins to the nanocarrier's surface after contact with biological fluids. In this context different nanoparticle characteristics could be beneficial for effective active targeting after formation of a protein corona which need to be identified. In this study trastuzumab was used as an active targeting ligand which was covalently attached to human serum albumin nanoparticles. For coupling reaction different molecular weight spacers were used and resulting physicochemical nanoparticle characteristics were evaluated. The in vitro cell association of the different nanoparticle formulations was tested in cell culture experiments with or without fetal bovine serum. For specific receptor-mediated cell interaction SK-BR-3 breast cancer cells with human epidermal growth factor receptor 2 (HER2) overexpression were used. MCF-7 breast cancer cells with normal HER2 expression served as control. Furthermore, serum protein adsorption on respective nanoparticles was characterized. The qualitative and quantitative composition of the protein corona was analyzed by SDS-PAGE and LC-MS/MS and the influence of protein adsorption on active targeting capability was determined.

Erratum to 'Spacer length and serum protein adsorption affect active targeting of trastuzumab-modified nanoparticles' [Biomaterials and Biosystems 5 (2022) 100032].

[This corrects the article DOI: 10.1016/j.bbiosy.2021.100032.].

Hexafluoroisopropyl Carbamates as Selective MAGL and Dual MAGL/FAAH Inhibitors: Biochemical and Physicochemical Properties.

A series of hexafluoroisopropyl carbamates with indolylalkyl- and azaindolylalkyl-substituents at the carbamate nitrogen was synthesized and evaluated for inhibition of the endocannabinoid degrading enzymes fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL). The synthesized derivatives with butyl to heptyl spacers between the heteroaryl and the carbamate moiety were inhibitors of both enzymes. For investigated compounds in which the alkyl chain was partially incorporated into a piperidine ring, different results were obtained. Compounds with a methylene spacer between the piperidine ring and the heteroaromatic system were found to be selective MAGL inhibitors, while an extension of the alkyl spacer to two to four atoms resulted in dual inhibition of FAAH/MAGL. The only small change in enzyme inhibitory activity with variation of the heteroaromatic system indicates that the reactive hexafluoroisopropyl carbamate group is mainly responsible for the strength of the inhibitory effect of the compounds. Selected derivatives were also tested for hydrolytic stability in aqueous solution, liver homogenate and blood plasma as well as for aqueous solubility and for permeability in a Caco-2 cell model. Some compounds showed a slightly higher MAGL inhibitory effect than the known selective MAGL inhibitor ABX-1431 and also partly surpassed this substance with regard to certain physicochemical and biochemical properties such as water solubility and cell permeability.