A-series agent A-234: initial in vitro and in vivo characterization.

A-series agent A-234 belongs to a new generation of nerve agents. The poisoning of a former Russian spy Sergei Skripal and his daughter in Salisbury, England, in March 2018 led to the inclusion of A-234 and other A-series agents into the Chemical Weapons Convention. Even though five years have already passed, there is still very little information on its chemical properties, biological activities, and treatment options with established antidotes. In this article, we first assessed A-234 stability in neutral pH for subsequent experiments. Then, we determined its inhibitory potential towards human recombinant acetylcholinesterase (HssAChE; EC 3.1.1.7) and butyrylcholinesterase (HssBChE; EC 3.1.1.8), the ability of HI-6, obidoxime, pralidoxime, methoxime, and trimedoxime to reactivate inhibited cholinesterases (ChEs), its toxicity in rats and therapeutic effects of different antidotal approaches. Finally, we utilized molecular dynamics to explain our findings. The results of spontaneous A-234 hydrolysis showed a slow process with a reaction rate displaying a triphasic course during the first 72 h (the residual concentration 86.2%). A-234 was found to be a potent inhibitor of both human ChEs (HssAChE IC50 = 0.101 ± 0.003 µM and HssBChE IC50 = 0.036 ± 0.002 µM), whereas the five marketed oximes have negligible reactivation ability toward A-234-inhibited HssAChE and HssBChE. The acute toxicity of A-234 is comparable to that of VX and in the context of therapy, atropine and diazepam effectively mitigate A-234 lethality. Even though oxime administration may induce minor improvements, selected oximes (HI-6 and methoxime) do not reactivate ChEs in vivo. Molecular dynamics implies that all marketed oximes are weak nucleophiles, which may explain the failure to reactivate the A-234 phosphorus-serine oxygen bond characterized by low partial charge, in particular, HI-6 and trimedoxime oxime oxygen may not be able to effectively approach the A-234 phosphorus, while pralidoxime displayed low interaction energy. This study is the first to provide essential experimental preclinical data on the A-234 compound.

Phenoxytacrine derivatives: Low-toxicity neuroprotectants exerting affinity to ifenprodil-binding site and cholinesterase inhibition.

Tacrine (THA), a long withdrawn drug, is still a popular scaffold used in medicinal chemistry, mainly for its good reactivity and multi-targeted effect. However, THA-associated hepatotoxicity is still an issue and must be considered in drug discovery based on the THA scaffold. Following our previously identified hit compound 7-phenoxytacrine (7-PhO-THA), we systematically explored the chemical space with 30 novel derivatives, with a focus on low hepatotoxicity, anticholinesterase action, and antagonism at the GluN1/GluN2B subtype of the NMDA receptor. Applying the down-selection process based on in vitro and in vivo pharmacokinetic data, two candidates, I-52 and II-52, selective GluN1/GluN2B inhibitors thanks to the interaction with the ifenprodil-binding site, have entered in vivo pharmacodynamic studies. Finally, compound I-52, showing only minor affinity to AChE, was identified as a lead candidate with favorable behavioral and neuroprotective effects using open-field and prepulse inhibition tests, along with scopolamine-based behavioral and NMDA-induced hippocampal lesion models. Our data show that compound I-52 exhibits low toxicity often associated with NMDA receptor ligands, and low hepatotoxicity, often related to THA-based compounds.

A-agents, misleadingly known as "Novichoks": a narrative review.

"Novichok" refers to a new group of nerve agents called the A-series agents. Their existence came to light in 2018 after incidents in the UK and again in 2020 in Russia. They are unique organophosphorus-based compounds developed during the Cold War in a program called Foliant in the USSR. This review is based on original chemical entities from Mirzayanov's memoirs published in 2008. Due to classified research, a considerable debate arose about their structures, and hence, various structural moieties were speculated. For this reason, the scientific literature is highly incomplete and, in some cases, contradictory. This review critically assesses the information published to date on this class of compounds. The scope of this work is to summarize all the available and relevant information, including the physicochemical properties, chemical synthesis, mechanism of action, toxicity, pharmacokinetics, and medical countermeasures used to date. The environmental stability of A-series agents, the lack of environmentally safe decontamination, their high toxicity, and the scarcity of information on post-contamination treatment pose a challenge for managing possible incidents.

The measured CSF/plasma donepezil concentration ratio but not individually measured CSF and plasma concentrations significantly increase over 24 h after donepezil treatment in patients with Alzheimer's disease.

BACKGROUND: The acetylcholinesterase inhibitor donepezil is administered as a treatment for Alzheimer's disease (AD). However, the appropriate donepezil dosage is still a matter of debate. METHODS: Forty AD patients receiving 10 mg/day of donepezil were randomly divided into four groups based on the time of plasma and cerebrospinal fluid (CSF) sampling: 6 h (n = 5), 12 h (n = 12), 18 h (n = 6) and 24 h (n = 17) after donepezil administration. High-performance liquid chromatography measured the donepezil concentration in plasma samples and CSF samples collected at 4-time points. RESULTS: Plasma and CSF levels among the groups were not significantly different. Conversely, the CSF/plasma donepezil concentration ratio considerably increased in the 24 h group compared to the 6 h (p < 0.005) and 12 h (p < 0.05) groups. CONCLUSION: The measurement of the CSF/plasma donepezil concentration ratio could be used to better evaluate the optimal dose of donepezil.

Nanofiber Wound Dressing Materials-A Comparative Study of Wound Healing on a Porcine Model.

BACKGROUND: Nanofiber wound dressings remain the domain of in vitro studies. The purpose of our study was to verify the benefits of chitosan (CTS) and polylactide (PLA)-based nanofiber wound dressings on a porcine model of a naturally contaminated standardized wound and compare them with the conventional dressings, i.e., gauze and Inadine. MATERIAL AND METHODS: The study group included 32 pigs randomized into four homogeneous groups according to the wound dressing type. Standardized wounds were created on their backs, and wound dressings were regularly changed. We evaluated difficulty of handling individual dressing materials and macroscopic appearance of the wounds. Wound swabs were taken for bacteriological examination. Blood samples were obtained to determine blood count values and serum levels of acute phase proteins (serum amyloid A, C-reactive protein, and haptoglobin). The crucial point of the study was histological analysis. Microscopic evaluation was focused on the defect depth and tissue reactions, including formation of the fibrin exudate with neutrophil granulocytes, the layer of granulation and cellular connective tissue, and the reepithelialization. Statistical analysis was performed by using SPSS software. The analysis was based on the Kruskal-Wallis H test and Mann-Whitney U test followed by Bonferroni correction. Significance was set at P < .05. RESULTS: Macroscopic examination did not show any difference in wound healing among the groups. However, evaluation of histological findings demonstrated that PLA-based nanofiber dressing accelerated the proliferative (P = .025) and reepithelialization (P < .001) healing phases, while chitosan-based nanofiber dressing potentiated and accelerated the inflammatory phase (P = .006). No statistically significant changes were observed in the blood count or acute inflammatory phase proteins during the trial. Different dynamics were noted in serum amyloid A values in the group treated with PLA-based nanofiber dressing (P = .006). CONCLUSION: Based on the microscopic examination, we have documented a positive effect of nanofiber wound dressings on acceleration of individual phases of the healing process. Nanofiber wound dressings have a potential to become in future part of the common wound care practice.

Neurotoxicity evoked by organophosphates and available countermeasures.

Organophosphorus compounds (OP) are a constant problem, both in the military and in the civilian field, not only in the form of acute poisoning but also for their long-lasting consequences. No antidote has been found that satisfactorily protects against the toxic effects of organophosphates. Likewise, there is no universal cure to avert damage after poisoning. The key mechanism of organophosphate toxicity is the inhibition of acetylcholinesterase. The overstimulation of nicotinic or muscarinic receptors by accumulated acetylcholine on a synaptic cleft leads to activation of the glutamatergic system and the development of seizures. Further consequences include generation of reactive oxygen species (ROS), neuroinflammation, and the formation of various other neuropathologists. In this review, we present neuroprotection strategies which can slow down the secondary nerve cell damage and alleviate neurological and neuropsychiatric disturbance. In our opinion, there is no unequivocal approach to ensure neuroprotection, however, sooner the neurotoxicity pathway is targeted, the better the results which can be expected. It seems crucial to target the key propagation pathways, i.e., to block cholinergic and, foremostly, glutamatergic cascades. Currently, the privileged approach oriented to stimulating GABAAR by benzodiazepines is of limited efficacy, so that antagonizing the hyperactivity of the glutamatergic system could provide an even more efficacious approach for terminating OP-induced seizures and protecting the brain from permanent damage. Encouraging results have been reported for tezampanel, an antagonist of GluK1 kainate and AMPA receptors, especially in combination with caramiphen, an anticholinergic and anti-glutamatergic agent. On the other hand, targeting ROS by antioxidants cannot or already developed neuroinflammation does not seem to be very productive as other processes are also involved.

MITOCHONDRIA IN BIODOSIMETRY: FLOW CYTOMETRY ASSESMENT IN VITRO.

The JC-1 dye is widely used in apoptosis studies to monitor mitochondrial health. The probe was tested in vitro on two established cell lines and peripheral porcine blood lymphocytes after gamma irradiation (IR) to assess its potential in biodosimetric evaluation. In brief, we stained irradiated and non-irradiated cells with the JC-1 dye to determine the existing changes in mitochondrial membrane potential and monitor cell health through flow cytometry. The stage of injury in these cells was evaluated through an irradiated versus non-irradiated ratio (IVNIR), comparing the relative proportion of polarised cells containing red JC-1 aggregates. We observed a decreasing IVNIR as the radiation dose increased (i.e. 0.5; 1; 2; 4; 6; 8 and 10 Gy), performing the analysis at 4, 8 and 24 h after IR in all the tested cells. The results from the JC1-dye test showed that CD4 T lymphocytes were more sensitive to irradiation than other subpopulations.

Non-covalent acetylcholinesterase inhibitors: In vitro screening and molecular modeling for novel selective insecticides.

Insecticides represent the most crucial element in the integrated management approach to malaria and other vector-borne diseases. The evolution of insect resistance to long-used substances and the toxicity of organophosphates (OPs) and carbamates are the main factors contributing to the development of new, environmentally safe pesticides. In our work, fourteen compounds of 7-methoxytacrine-tacrine heterodimers were tested for their insecticidal effect. Compounds were evaluated in vitro on insect acetylcholinesterase from Anopheles gambiae (AgAChE) and Musca domestica (MdAChE). The evaluation was executed in parallel with testing on human erythrocyte acetylcholinesterase (HssAChE) and human butyrylcholinesterase (HssBChE) using a modified Ellman's method. Compound efficacy was determined as IC50 values for the respective enzymes and selectivity indexes were expressed to compare the interspecies selectivity. Docking studies were performed to predict the binding modes of selected compounds. K1328 and K1329 provided high HssAChE/AgAChE selectivity outperforming standard pesticides (carbofuran and bendiocarb), and thus can be considered as suitable lead structure for novel anticholinesterase insecticides.

Mitigation of Ionizing Radiation-Induced Gastrointestinal Damage by Insulin-Like Growth Factor-1 in Mice.

Purpose: Insulin-like growth factor-1 (IGF-1) stimulates epithelial regeneration but may also induce life-threatening hypoglycemia. In our study, we first assessed its safety. Subsequently, we examined the effect of IGF-1 administered in different dose regimens on gastrointestinal damage induced by high doses of gamma radiation. Material and methods: First, fasting C57BL/6 mice were injected subcutaneously with IGF-1 at a single dose of 0, 0.2, 1, and 2 mg/kg to determine the maximum tolerated dose (MTD). The glycemic effect of MTD (1 mg/kg) was additionally tested in non-fasting animals. Subsequently, a survival experiment was performed. Animals were irradiated (60Co; 14, 14.5, or 15 Gy; shielded head), and IGF-1 was administered subcutaneously at 1 mg/kg 1, 24, and 48 h after irradiation. Simultaneously, mice were irradiated (60Co; 12, 14, or 15 Gy; shielded head), and IGF-1 was administered subcutaneously under the same regimen. Jejunum and lung damage were assessed 84 h after irradiation. Finally, we evaluated the effect of six different IGF-1 dosage regimens administered subcutaneously on gastrointestinal damage and peripheral blood changes in mice 6 days after irradiation (60Co; 12 and 14 Gy; shielded head). The regimens differed in the number of doses (one to five doses) and the onset of administration (starting at 1 [five regimens] or 24 h [one regimen] after irradiation). Results: MTD was established at 1 mg/kg. MTD mitigated lethality induced by 14 Gy and reduced jejunum and lung damage caused by 12 and 14 Gy. However, different dosing regimens showed different efficacy, with three and four doses (administered 1, 24, and 48 h and 1, 24, 48, and 72 h after irradiation, respectively) being the most effective. The three-dose regimens supported intestinal regeneration even if the administration started at 24 h after irradiation, but its potency decreased. Conclusion: IGF-1 seems promising in the mitigation of high-dose irradiation damage. However, the selected dosage regimen affects its efficacy.

Toxicity, pharmacokinetics, and effectiveness of the ortho-chlorinated bispyridinium oxime, K870.

Oxime reactivators are causal antidotes for organophosphate intoxication. Herein, the toxicity, pharmacokinetics, and reactivation effectiveness of o-chlorinated bispyridinium oxime K870 are reported. Oxime K870 was found to have a safe profile at a dose of 30 mg/kg in rats. It exhibited rapid absorption and renal clearance similar to those of other charged oximes after intramuscular administration. Its isoxazole-pyridinium degradation product was identified in vivo. Although it showed some improvement in brain targeting, it was nevertheless rapidly effluxed from the central nervous system. Its reactivation effectiveness was evaluated in rats and mice intoxicated with sarin, tabun, VX, and paraoxon and compared with pralidoxime and asoxime. K870 was found to be less effective in reversing tabun poisoning compared to its parent unchlorinated oxime K203. However, K870 efficiently reactivated blood acetylcholinesterase for all tested organophosphates in rats. In addition, K870 significantly protected against intoxication by all tested organophosphates in mice. For these reasons, oxime K870 seems to have a broader reactivation spectrum against multiple organophosphates. It seems important to properly modulate the oximate forming properties (pKa) to obtain more versatile oxime reactivators.

Novel D2/5-HT receptor modulators related to cariprazine with potential implication to schizophrenia treatment.

Schizophrenia is a serious mental disorder without a fully understood pathomechanism, but which involves dysregulation of neurotransmitters and their receptors. The best option for the management of schizophrenia comprises so-called multi-target ligands, similar to the third generation of neuroleptics. Dopamine type 2 receptors (D2Rs) are the main target in the treatment of schizophrenia, in particular for mitigation of the positive symptoms. Due to the high expression of 5-hydroxytryptamine type 3 receptors (5-HT3Rs) in human brain areas responsible for emotional behavior, motivation, and cognitive function, 5-HT3Rs represent a potential target for modulating the cognitive and negative symptoms of schizophrenia. Here we present the design, synthesis, and both in vitro and in vivo biological evaluation of 1,4-disubstituted aromatic piperazines. Screening of in vitro properties revealed the two most promising drug candidates (21 and 24) which were found to be potent D2Rs and moderate 5-HT3R antagonists, and which were forwarded to in vivo studies in Wistar rats. Considering toxicity, administration of the maximal feasible dose of 21 (2 mg/kg) did not produce any side effects. By contrast, the higher solubility of 24 led to revelation of mild and temporary side effects at the dose of 20 mg/kg. Importantly, both 21 and 24 showed facile crossing of the blood-brain barrier, even exerting higher levels in the brain in comparison to plasma. In a behavioral study using the acute amphetamine model of psychosis, we showed that compound 24 ameliorated both positive and negative effects of amphetamine including hyperlocomotion, social impairments, and disruption of prepulse inhibition. The effect of the highest dose (10 mg/kg) was comparable to the effect of the reference dose of aripiprazole (1 mg/kg).

A systematic evaluation of the cucurbit[7]uril pharmacokinetics and toxicity after a single dose and short-term repeated administration in mice.

Cucurbit[n]urils are macrocyclic compounds capable of forming host-guest complexes with different molecules. In this study, we focused on cucurbit[7]uril (CB[7]) safety and pharmacokinetics. We investigated CB[7] cytotocixity in human renal cells ACHN using the xCELLigence system. We also determined maximum tolerated doses (MTD) and no observed adverse effect levels (NOAEL) after intramuscular (i.m.), intraperitoneal (i.p.), and intragastric (i.g.) administration in mice using clinical observation, blood biochemistry, and histopathology. At NOAELs, we studied its pharmacokinetics in plasma and kidneys. Finally, we performed a 7 day repeated-dose toxicity study at 50% of NOAEL after i.p. administration, assaying CB[7] concentration in plasma, brain, kidney, and liver; we also assessed the liver and kidney histopathology. In vitro, CB[7] did not show toxicity up to 0.94 mg/mL. MTDs in vivo were set at 300, 350, and 600 mg/kg, and NOAEL were established at 150, 100, and 300 mg/kg after i.m., i.p., and i.g. administration, respectively. Parenteral administration produced tissue damage mainly to the kidney, while i.g. administration caused only minor liver damage. Parenteral CB[7] administration led to fast elimination from blood, accompanied with kidney accumulation; absorption from the gastrointestinal tract was minimal. Short repeated i.p. administration was well tolerated. After initial CB[7] accumulation in blood and kidney, the concentrations stabilised and decreased during the experiment. Approximately 3.6% of animals showed signs of nephrotoxicity. Although CB[7] appears to be a promising molecule, nephrotoxicity may be the most critical drawback of its parenteral use, because the kidney represents the main organ of its elimination.

A Comparison of the Neuroprotective and Reactivating Efficacy of a Novel Bispyridinium Oxime K870 with Commonly Used Pralidoxime and the Oxime HI-6 in Tabun-Poisoned Rats.

AIM: The comparison of neuroprotective and central reactivating effects of the oxime K870 in combination with atropine with the efficacy of standard antidotal treatment in tabun-poisoned rats. METHODS: The neuroprotective effects of antidotal treatment were determined in rats poisoned with tabun at a sublethal dose using a functional observational battery 2 h and 24 h after tabun administration, the tabun-induced brain damage was investigated by the histopathological evaluation and central reactivating effects of oximes was evaluated by the determination of acetylcholinesterase activity in the brain using a standard spectrophotometric method. RESULTS: The central reactivating efficacy of a newly developed oxime K870 roughly corresponds to the central reactivating efficacy of pralidoxime while the ability of the oxime HI-6 to reactivate tabun-inhibited acetylcholinesterase in the brain was negligible. The ability of the oxime K870 to decrease tabun-induced acute neurotoxicity was slightly higher than that of pralidoxime and similar to the oxime HI-6. These results roughly correspond to the histopathological evaluation of tabun-induced brain damage. CONCLUSION: The newly synthesized oxime K870 is not a suitable replacement for commonly used oximes in the antidotal treatment of acute tabun poisonings because its neuroprotective efficacy is only slightly higher or similar compared to studied currently used oximes.

Strong Antimicrobial and Healing Effects of Beta-Acids from Hops in Methicillin-Resistant Staphylococcus aureus-Infected External Wounds In Vivo.

Staphylococcus (S.) aureus is an important causative agent of wound infections with increasing incidence in the past decades. Specifically, the emergence of methicillin-resistant S. aureus (MRSA) causes serious problems, especially in nosocomial infections. Therefore, there is an urgent need to develop of alternative or supportive antimicrobial therapeutic modalities to meet these challenges. Purified compounds from hops have previously shown promising antimicrobial effects against MRSA isolates in vitro. In this study, purified beta-acids from hops were tested for their potential antimicrobial and healing properties using a porcine model of wounds infected by MRSA. The results show highly significant antimicrobial effects of the active substance in both the powder and Ambiderman-based application forms compared to both no-treatment control and treatment with Framycoin. Moreover, the macroscopic evaluation of the wounds during the treatment using the standardized Wound Healing Continuum indicated positive effects of the beta-acids on the overall wound healing. This is further supported by the microscopic data, which showed a clear improvement of the inflammatory parameters in the wounds treated by beta-acids. Thus, using the porcine model, we demonstrate significant therapeutic effects of hops compounds in the management of wounds infected by MRSA. Beta-acids from hops, therefore, represent a suitable candidate for the treatment of non-responsive nosocomial tissue infections by MRSA.

The Effect of Lactobacillus casei on Experimental Porcine Inflammatory Bowel Disease Induced by Dextran Sodium Sulphate.

BACKGROUND: Gastrointestinal injury caused by dextran sodium sulphate (DSS) is a reliable porcine experimental model of inflammatory bowel disease (IBD). The purpose of this study was to evaluate the effect of probiotic Lactobacillus casei DN 114001 (LC) on DSS-induced experimental IBD. RESULTS: Eighteen female pigs (Sus scrofa f. domestica, weight 33-36 kg, age 4-5 months) were divided into 3 groups (6 animals per group): controls with no treatment, DSS, and DSS + LC. LC was administered to overnight fasting animals in a dietary bolus in the morning on days 1-7 (4.5 × 1010 live bacteria/day). DSS was applied simultaneously on days 3-7 (0.25 g/kg/day). On day 8, the pigs were sacrificed. Histopathological score and length of crypts/glands (stomach, jejunum, ileum, transverse colon), length and width of villi (jejunum, ileum), and mitotic and apoptotic indices (jejunum, ileum, transverse colon) were assessed. DSS increased the length of glands in the stomach, length of crypts and villi in the jejunum and ileum, and the histopathological score of gastrointestinal damage, length of crypts and mitotic activity in the transverse colon. Other changes did not achieve any statistical significance. Administration of LC reduced the length of villi in the jejunum and ileum to control levels and decreased the length of crypts in the jejunum. CONCLUSIONS: Treatment with a probiotic strain of LC significantly accelerated regeneration of the small intestine in a DSS-induced experimental porcine model of IBD.

Strong Antimicrobial Effects of Xanthohumol and Beta-Acids from Hops against Clostridioides difficile Infection In Vivo.

Clostridioides (C.) difficile is an important causative pathogen of nosocomial gastrointestinal infections in humans with an increasing incidence, morbidity, and mortality. The available treatment options against this pathogen are limited. The standard antibiotics are expensive, can promote emerging resistance, and the recurrence rate of the infection is high. Therefore, there is an urgent need for new approaches to meet these challenges. One of the possible treatment alternatives is to use compounds available in commonly used plants. In this study, purified extracts isolated from hops-alpha and beta acids and xanthohumol-were tested in vivo for their inhibitory effect against C. difficile. A rat model of the peroral intestinal infection by C. difficile has been developed. The results show that both xanthohumol and beta acids from hops exert a notable antimicrobial effect in the C. difficile infection. The xanthohumol application showed the most pronounced antimicrobial effect together with an improvement of local inflammatory signs in the large intestine. Thus, the hops compounds represent promising antimicrobial agents for the treatment of intestinal infections caused by C. difficile.

Silver distribution in chronic wounds and the healing dynamics of chronic wounds treated with dressings containing silver and octenidine.

Although silver is an efficient antimicrobial and is a widely used antiseptic in wound healing, previous studies have reported the cytotoxic in vitro effects of silver dressings. Moreover, few studies have addressed the distribution of silver in chronic wounds. The study compares the healing of chronic wounds treated with a standard-of-care silver dressing (Ag-CMC) and a dressing containing antiseptic octenidine (OCT-HA). Biopsies were taken from two wound areas before the commencement of treatment (baseline), after 2 weeks and after 6 weeks (the end of the study). We analyzed the histopathologic wound-healing score, silver distribution, and expression of selected genes. The wound-healing score improved significantly in the wounded area treated with OCT-HA after 2 weeks compared to the baseline and the Ag-CMC. The Ag-CMC wound areas improved after 6 weeks compared to the baseline. Moreover, collagen maturation and decreases in the granulocyte and macrophage counts were faster in the OCT-HA parts. Treatment with OCT-HA resulted in less wound slough. The silver, visualized via autometallography, penetrated approximately 2 mm into the wound tissue and associated around capillaries and ECM fibers, and was detected in phagocytes. The metallothionein gene expression was elevated in the Ag-CMC wound parts. This exploratory study determined the penetration of silver into human chronic wounds and changes in the distribution thereof during treatment. We observed that silver directly affects the cells in the wound and elevates the metallothionein gene expression. Octenidine and hyaluronan dressings provide a suitable alternative to silver and carboxymethyl cellulose dressings without supplying silver to the wound.

Phenothiazine-Tacrine Heterodimers: Pursuing Multitarget Directed Approach in Alzheimer's Disease.

Since 2002, no clinical candidate against Alzheimer's disease has reached the market; hence, an effective therapy is urgently needed. We followed the so-called "multitarget directed ligand" approach and designed 36 novel tacrine-phenothiazine heterodimers which were in vitro evaluated for their anticholinesterase properties. The assessment of the structure-activity relationships of such derivatives highlighted compound 1dC as a potent and selective acetylcholinesterase inhibitor with IC50 = 8 nM and 1aA as a potent butyrylcholinesterase inhibitor with IC50 = 15 nM. Selected hybrids, namely, 1aC, 1bC, 1cC, 1dC, and 2dC, showed a significant inhibitory activity toward τ(306-336) peptide aggregation with percent inhibition ranging from 50.5 to 62.1%. Likewise, 1dC and 2dC exerted a remarkable ability to inhibit self-induced Aß1-42 aggregation. Notwithstanding, in vitro studies displayed cytotoxicity toward HepG2 cells and cerebellar granule neurons; no pathophysiological abnormality was observed when 1dC was administered to mice at 14 mg/kg (i.p.). 1dC was also able to permeate to the CNS as shown by in vitro and in vivo models. The maximum brain concentration was close to the IC50 value for acetylcholinesterase inhibition with a relatively slow elimination half-time. 1dC showed an acceptable safety and good pharmacokinetic properties and a multifunctional biological profile.

Determination of K869, a Novel Oxime Reactivator of Acetylcholinesterase, in Rat Body Fluids and Tissues by Liquid-Chromatography Methods: Pharmacokinetic Study.

Oxime reactivators of acetylcholinesterase (AChE) represent an integral part of standard antidote treatment of organophosphate poisoning. Oxime K869 is a novel bisquaternary non-symmetric pyridinium aldoxime with two pyridinium rings connected by a tetramethylene bridge where two chlorines modify the pyridinium ring bearing the oxime moiety. Based on in vitro assays, K869 is a potent AChE and butyrylcholinesterase (BChE) reactivator. For the investigation of the basic pharmacokinetic properties of K869 after its intramuscular application, new HPLC-UV and LC-MS/MS methods were developed and validated for its determination in rat body fluids and tissues. In this study, the SPE procedure for sample pretreatment was optimized as an alternative to routine protein precipitation widely used in oxime pharmacokinetics studies. K869 oxime is quickly absorbed into the central compartment reaching its maximum in plasma (39 ± 4 µg/mL) between 15 and 20 min. The majority of K869 was eliminated by kidneys via urine when compared with biliary excretion. However, only a limited amount of K869 (65 ± 4 ng/g of brain tissue) was found in the brain 30 min after oxime administration. Regarding the brain/plasma ratio calculated (less than 1%), the penetration of K869 into the brain did not exceed conventionally used oximes.