Enzyme-armed nanocleaner provides superior detoxification against organophosphorus compounds via a dual-action mechanism.

By inhibiting acetylcholinesterase (AChE) activity, organophosphate compounds (OPs) can quickly cause severe injury to the nervous system and death, making it extremely difficult to rescue victims after OP exposure. However, it is quite challenging to construct scavengers that neutralize and eliminate these harmful chemical agents promptly in the blood circulation system. Herein, we report an enzyme-armed biomimetic nanoparticle that enables a 'targeted binding and catalytic degradation' action mechanism designed for highly efficient in vivo detoxification (denoted as 'Nanocleaner'). Specifically, the resulting Nanocleaner is fabricated with polymeric cores camouflaged with a modified red blood cell membrane (RBC membrane) that is inserted with the organophosphorus hydrolase (OPH) enzyme. In such a subtle construct, Nanocleaner inherits abundant acetylcholinesterase (AChE) on the surface of the RBC membrane, which can specifically lure and neutralize OPs through biological binding. The OPH enzyme on the membrane surface breaks down toxicants catalytically. The in vitro protective effects of Nanocleaner against methyl paraoxon (MPO)-induced inhibition of AChE activity were validated using both preincubation and competitive regimens. Furthermore, we selected the PC12 neuroendocrine cell line as an experimental model and confirmed the cytoprotective effects of Nanocleaner against MPO. In mice challenged with a lethal dose of MPO, Nanocleaner significantly reduces clinical signs of intoxication, rescues AChE activity and promotes the survival rate of mice challenged with lethal MPO. Overall, these results suggest considerable promise of enzyme-armed Nanocleaner for the highly efficient removal of OPs for clinical treatment.

HI-6-Loaded Vehicle of Liposomes Mediated by an Amphiphilic Pillar[5]arene against Paraoxon Poisoning.

Organophosphate (OP) intoxication has become a severe common health matter all over the world. For the treatment of acute OP poisoning, the effective intracerebral delivery of acetylcholinesterase reactivators is crucial. Here, an amphiphilic hydrazide-pillar[5]arene (HP5A-6C), which could be readily integrated into liposomal bilayers' zwitterionic disaturated phosphatidylcholine (DSPC), was synthesized. A T7 peptide-containing guest (G) was attached on the surface via a noncovalent interaction to make mixed liposomes a particularly appealing candidate for brain-targeting delivery. Such coassembly could remain stable at room temperature for up to 6 weeks, and safety evaluations initially verified its fine biological compatibility. The hydrophilic interiors of T7/HP5A-6C@DSPC could further load HI-6 with 89.70% encapsulation efficiency. Support for brain-targeting potency came from imaging results. Notably, intravenous injection of HI-6-loaded vesicles exhibited a remarkable therapeutic effect on paraoxon (POX)-poisoned mice, effectively alleviating seizures and brain damage and significantly increasing the improving survival rate to 60% over the course of 7 days.

Unwashed grapes as a cause of organophosphate poisoning in a child in rural India.

This report describes the symptoms of pesticide poisoning in a previously healthy teenage girl. After consuming unwashed grapes for several days, the girl developed the following symptoms: drowsiness, vomiting, truncal weakness and fasciculations in the tongue and hands. Blood tests confirmed exposure to a small amount of the organophosphate (OP) compound, a type of chemical found in certain pesticides. The girl was treated with supportive care and cholinesterase reactivators, which minimised the damage caused by OP poisoning. Within 48 hours, the girl's symptoms improved and she made a full recovery. This case highlights that OP poisoning can present without classic cholinergic crisis symptoms (SLUDGING), including miosis. Fasciculations, as observed in this case, are a significant clue to the diagnosis.

Applications of Microbial Organophosphate-Degrading Enzymes to Detoxification of Organophosphorous Compounds for Medical Countermeasures against Poisoning and Environmental Remediation.

Mining of organophosphorous (OPs)-degrading bacterial enzymes in collections of known bacterial strains and in natural biotopes are important research fields that lead to the isolation of novel OP-degrading enzymes. Then, implementation of strategies and methods of protein engineering and nanobiotechnology allow large-scale production of enzymes, displaying improved catalytic properties for medical uses and protection of the environment. For medical applications, the enzyme formulations must be stable in the bloodstream and upon storage and not susceptible to induce iatrogenic effects. This, in particular, includes the nanoencapsulation of bioscavengers of bacterial origin. In the application field of bioremediation, these enzymes play a crucial role in environmental cleanup by initiating the degradation of OPs, such as pesticides, in contaminated environments. In microbial cell configuration, these enzymes can break down chemical bonds of OPs and usually convert them into less toxic metabolites through a biotransformation process or contribute to their complete mineralization. In their purified state, they exhibit higher pollutant degradation efficiencies and the ability to operate under different environmental conditions. Thus, this review provides a clear overview of the current knowledge about applications of OP-reacting enzymes. It presents research works focusing on the use of these enzymes in various bioremediation strategies to mitigate environmental pollution and in medicine as alternative therapeutic means against OP poisoning.

N-acetylcysteine as a potentially safe adjuvant in the treatment of neurotoxicity due to pirimiphos-methyl poisoning.

Exogenous, well-established antioxidant N-acetylcysteine can reduce or prevent the deleterious effects of pesticides. In this study, utilizing a mouse model of daily single dose of N-acetylcysteine administration, we investigated the impact of this adjuvant on the treatment with atropine and/or obidoxime as well as oxidative stress response in pyrimiphos-methyl-induced toxicity. We found that N-acetylcysteine significantly reduces the oxidative stress generated by pyrimiphos-methyl. The therapy consisting of atropine and/or obidoxime routinely used in organophosphorous insecticide poisonings, including pyrimiphos-methyl, had no effect on the antioxidant properties of N-acetylcysteine. Adjunctive treatment offered by N-acetylcysteine fills therapeutic gap and may provide the full potential against pyrimiphos-methyl-induced toxicity.

Serum Organophosphorus compound levels as a surrogate marker for severity assessment and management in Acute Organophosphorus Poisoning - A novel approach.

Organophosphorus (OP) compounds are the most extensively used pesticides' class worldwide; cause most self­poisoning deaths especially in India. Thus, it is utmost important for early identification and aggressive management of OP poisoning from the clinical perspective to prevent serious complications by using sophisticated LC-MS/MS approach. This was a prospective study involving 103 patients of OP cases admitted to Karnataka Institute of Medical Sciences from June 2022 to May 2023, based on the inclusion and exclusion criteria patients were subjected to study. On admission, venous blood was collected from patient with Malathion and Profenofos OP poisoning history and subjected to serum biomarker and to LC-MS/MS analysis. Out of the 103 patients, 68 patients consumed Profenofos (66%) and 35 patients consumed Malathion (34%). Pseudocholinesterase levels among the of OP cases revealed that the 33 patients had mild toxicity, 40 patients had moderate toxicity and 30 patients had severe toxicity of OP poisoning. Subsequently LC-MS/MS analysis showed that the results obtained are not in correlation with indirect serum marker pseudocholinesterase levels. On the other side, LC-MS/MS results are in correlation with the clinical outcome of the patients with respect to morbidity and mortality. Thus, LC-MS/MS approach to assess the OP levels in patients could be used as potential diagnostic and prognostic marker for the absolute quantification of OP compounds compared to indirect OP levels estimation.

The use of bispyridinium non-oxime analogues for the restoration of nerve agent impaired neuromuscular transmission in rat hemidiaphragms - Structure optimization.

Organophosphate pesticide poisoning challenges health care systems worldwide. Furthermore, nerve agents remain a continuous threat. The treatment options for organophosphate poisoning have virtually been unchanged for decades, relying on symptomatic treatment and the use of oximes to indirectly restore neuromuscular function. Hence, compounds targeting directly nicotinic acetylcholine receptors (nAChRs) might substantially improve treatment options. The current study investigated a series of bispyridinium analogues with a trimethylene or 2,2'-diethyloxy linker in a rat hemidiaphragm model, using indirect field stimulation. Methyl- and ethyl-substituted bispyridinium analogues restored neuromuscular function up to 37 ± 17% (MB419, a 3-methyl analogue) at a stimulation frequency of 20 Hz. The bispyridinium analogues with a 2- or 3-methyl group, or a 2- or 3-ethyl group, tended towards a higher restoration of neuromuscular function than those with a 4-methyl or 4-ethyl group, respectively. The current data can be used for future studies to optimize structure-based molecular modeling of compounds targeting the nAChR.

Synthesis and biological evaluation of novel MB327 analogs as resensitizers for desensitized nicotinic acetylcholine receptors after intoxication with nerve agents.

Poisoning with organophosphorus compounds, which can lead to a cholinergic crisis due to the inhibition of acetylcholinesterase and the subsequent accumulation of acetylcholine (ACh) in the synaptic cleft, is a serious problem for which treatment options are currently insufficient. Our approach to broadening the therapeutic spectrum is to use agents that interact directly with desensitized nicotinic acetylcholine receptors (nAChRs) in order to induce functional recovery after ACh overstimulation. Although MB327, one of the most prominent compounds investigated in this context, has already shown positive properties in terms of muscle force recovery, this compound is not suitable for use as a therapeutic agent due to its insufficient potency. By means of in silico studies based on our recently presented allosteric binding pocket at the nAChR, i.e. the MB327-PAM-1 binding site, three promising MB327 analogs with a 4-aminopyridinium ion partial structure (PTM0056, PTM0062, and PTM0063) were identified. In this study, we present the synthesis and biological evaluation of a series of new analogs of the aforementioned compounds with a 4-aminopyridinium ion partial structure (PTM0064-PTM0072), as well as hydroxy-substituted analogs of MB327 (PTMD90-0012 and PTMD90-0015) designed to substitute entropically unfavorable water clusters identified during molecular dynamics simulations. The compounds were characterized in terms of their binding affinity towards the aforementioned binding site by applying the UNC0642 MS Binding Assays and in terms of their muscle force reactivation in rat diaphragm myography. More potent compounds were identified compared to MB327, as some of them showed a higher affinity towards MB327-PAM-1 and also a higher recovery of neuromuscular transmission at lower compound concentrations. To improve the treatment of organophosphate poisoning, direct targeting of nAChRs with appropriate compounds is a key step, and this study is an important contribution to this research.

Quantitative T2 mapping-based longitudinal assessment of brain injury and therapeutic rescue in the rat following acute organophosphate intoxication.

Acute intoxication with organophosphate (OP) cholinesterase inhibitors poses a significant public health risk. While currently approved medical countermeasures can improve survival rates, they often fail to prevent chronic neurological damage. Therefore, there is need to develop effective therapies and quantitative metrics for assessing OP-induced brain injury and its rescue by these therapies. In this study we used a rat model of acute intoxication with the OP, diisopropylfluorophosphate (DFP), to test the hypothesis that T2 measures obtained from brain magnetic resonance imaging (MRI) scans provide quantitative metrics of brain injury and therapeutic efficacy. Adult male Sprague Dawley rats were imaged on a 7T MRI scanner at 3, 7 and 28 days post-exposure to DFP or vehicle (VEH) with or without treatment with the standard of care antiseizure drug, midazolam (MDZ); a novel antiseizure medication, allopregnanolone (ALLO); or combination therapy with MDZ and ALLO (DUO). Our results show that mean T2 values in DFP-exposed animals were: (1) higher than VEH in all volumes of interest (VOIs) at day 3; (2) decreased with time; and (3) decreased in the thalamus at day 28. Treatment with ALLO or DUO, but not MDZ alone, significantly decreased mean T2 values relative to untreated DFP animals in the piriform cortex at day 3. On day 28, the DUO group showed the most favorable T2 characteristics. This study supports the utility of T2 mapping for longitudinally monitoring brain injury and highlights the therapeutic potential of ALLO as an adjunct therapy to mitigate chronic morbidity associated with acute OP intoxication.

Comparative analysis of organophosphorus versus carbamate pesticide poisoning: a case study.

Organophosphorus poisoning is a critical condition that can cause central nervous system depression, respiratory failure, and death early on. As its clinical manifestations closely resemble those of carbamate pesticide poisoning, the aim of this case study is to present a case of misdiagnosis, initially identifying carbofuran poisoning as organophosphate in a patient suspect of a heatstroke. We also present a case of intentional self-poisoning with organophosphate dichlorvos to underline the likelihood of pesticide poisoning in patients exhibiting acute cholinergic symptoms when the ingested substance is not known. In such cases, empirical treatment with atropine and oxime can be started pending timely differential diagnosis to adjust treatment as necessary.