Brazzolotto X


Full name : Brazzolotto Xavier

First name : Xavier

Mail : Institut de Recherche Biomedicale des Armees, 91223 Bretigny sur Orge

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Country : France

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Phone : +33476639738

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References (43)

Title : Warfare Nerve Agents and Paraoxonase-1 as a Potential Prophylactic Therapy against Intoxication - Iyengar_2024_Protein.Pept.Lett__
Author(s) : Iyengar ARS , Khandave PY , Bzdrenga J , Nachon F , Brazzolotto X , Pande AH
Ref : Protein Pept Lett , : , 2024
Abstract : Nerve agents are a class of lethal neurotoxic chemicals used in chemical warfare. In this review, we have briefly discussed a brief history of chemical warfare, followed by an exploration of the historical context surrounding nerve agents. The article explores the classification of these agents, their contemporary uses, their toxicity mechanisms, and the disadvantages of the current treatment options for nerve agent poisoning. It then discusses the possible application of enzymes as prophylactics against nerve agent poisoning, outlining the benefits and drawbacks of paraoxonase-1. Finally, the current studies on paraoxonase-1 are reviewed, highlighting that several challenges need to be addressed in the use of paraoxonase-1 in the actual field and that its potential as a prophylactic antidote against nerve agent poisoning needs to be evaluated. The literature used in this manuscript was searched using various electronic databases, such as PubMed, Google Scholar, Web of Science, Elsevier, Springer, ACS, Google Patent, and books using the keywords chemical warfare agent, Butyrylcholinesterase, enzyme, nerve agent, prophylactic, and paraoxonase- 1, with the time scale for the analysis of articles between 1960 to 2023, respectively. The study has suggested that concerted efforts by researchers and agencies must be made to develop effective countermeasures against NA poisoning and that PON1 has suitable properties for the development of efficient prophylaxis against NA poisoning.
ESTHER : Iyengar_2024_Protein.Pept.Lett__
PubMedSearch : Iyengar_2024_Protein.Pept.Lett__
PubMedID: 38706353

Title : Detoxification of V-nerve agents assisted by a microperoxidase: new pathway revealed by the use of a relevant VX simulant - Baptista_2024_Chembiochem__e202400137
Author(s) : Baptista da Silva V , Mahy JP , Brazzolotto X , Renard PY , Ricoux R , Legros J
Ref : Chembiochem , :e202400137 , 2024
Abstract : The biocatalyzed oxidative detoxification of the V-series simulant PhX, by mean of the microperoxidase AcMP11, affords the corresponding phosphonothioate as the prominent product instead of the classical P-S and P-O bond cleavage. While PhX is structurally very close to the live agent VX (the methyl group is replaced by a phenyl), assessment with other surrogates missing the nucleophilic amino function displayed more resistance under the same conditions with no phosphonothioate observed. These encouraging results highlight 1) the efficacy of AcMP11 microperoxidase to efficiently detoxify V-series organophosphorus nerve agents (OPNA), and 2) the necessity to use representative alkyl or aryl phosphonothioates simulants such as PhX bearing the appropriate side chain as well as the P-O and P-S cleavable bond to mimic accurately the V-series OPNA to prevent false positive or false negative results.
ESTHER : Baptista_2024_Chembiochem__e202400137
PubMedSearch : Baptista_2024_Chembiochem__e202400137
PubMedID: 38591336

Title : Inhibition of Human Cholinesterases and In Vitro beta-Amyloid Aggregation by Rationally Designed Peptides - Sanchis_2023_ChemMedChem__e202200691
Author(s) : Sanchis I , Spinelli R , Dias J , Brazzolotto X , Rietmann A , Aimaretti F , Siano AS
Ref : ChemMedChem , :e202200691 , 2023
Abstract : The multifactorial nature of Alzheimer's disease (AD) is now widely recognized, which has increased the interest in compounds that can address more than one AD-associated targets. Herein, we report the inhibitory activity on the human cholinesterases (acetylcholinesterase, hAChE and butyrylcholinesterase, hBChE) and on the AChE-induced β-amyloid peptide (Aβ) aggregation by a series of peptide derivatives designed by mutating aliphatic residues for aromatic ones. We identified peptide W3 (LGWVSKGKLL-NH(2)) as an interesting scaffold for the development of new anti-AD multitarget-directed drugs. It showed the lowest IC(50)value against hAChE reported for a peptide (0.99 +/- 0.02 micro M) and inhibited 94.2% +/- 1.2 of AChE-induced Aβ aggregation at 10 micro M. Furthermore, it inhibited hBChE (IC(50), 15.44 +/- 0.91 micro M), showed no 'in vivo' toxicity in brine shrimp and had shown moderated radical scavenging and Fe(2+) chelating capabilities in previous studies. The results are in line with multiple reports showing the utility of the indole moiety for the development of cholinesterase inhibitors.
ESTHER : Sanchis_2023_ChemMedChem__e202200691
PubMedSearch : Sanchis_2023_ChemMedChem__e202200691
PubMedID: 36995341

Title : Highly selective butyrylcholinesterase inhibitors related to Amaryllidaceae alkaloids - Design, synthesis, and biological evaluation - Pidany_2023_Eur.J.Med.Chem_252_115301
Author(s) : Pidany F , Kroustkova J , Al Mamun A , Suchankova D , Brazzolotto X , Nachon F , Chantegreil F , Dolezal R , Pulkrabkova L , Muckova L , Hrabinova M , Finger V , Kufa M , Soukup O , Jun D , Jenco J , Kunes J , Novakova L , Korabecny J , Cahlikova L
Ref : Eur Journal of Medicinal Chemistry , 252 :115301 , 2023
Abstract : Butyrylcholinesterase (BChE) is one of the most frequently implicated enzymes in the advanced stage of Alzheimer's disease (AD). As part of our endeavors to develop new drug candidates for AD, we have focused on natural template structures, namely the Amaryllidaceae alkaloids carltonine A and B endowed with high BChE selectivity. Herein, we report the design, synthesis, and in vitro evaluation of 57 novel highly selective human BChE (hBChE) inhibitors. Most synthesized compounds showed hBChE inhibition potency ranging from micromolar to low nanomolar scale. Compounds that revealed BChE inhibition below 100 nM were selected for detailed biological investigation. The CNS-targeted profile of the presented compounds was confirmed theoretically by calculating the BBB score algorithm, these data were corroborated by determining the permeability in vitro using PAMPA-assay for the most active derivatives. The study highlighted compounds 87 (hBChE IC(50) = 3.8 +/- 0.2 nM) and 88 (hBChE IC(50) = 5.7 +/- 1.5 nM) as the top-ranked BChE inhibitors. Compounds revealed negligible cytotoxicity for the human neuroblastoma (SH-SY5Y) and hepatocellular carcinoma (HepG2) cell lines compared to BChE inhibitory potential. A crystallographic study was performed to inspect the binding mode of compound 87, revealing essential interactions between 87 and hBChE active site. In addition, multidimensional QSAR analyses were applied to determine the relationship between chemical structures and biological activity in a dataset of designed agents. Compound 87 is a promising lead compound with potential implications for treating the late stages of AD.
ESTHER : Pidany_2023_Eur.J.Med.Chem_252_115301
PubMedSearch : Pidany_2023_Eur.J.Med.Chem_252_115301
PubMedID: 36996715
Gene_locus related to this paper: human-BCHE

Title : Rapid discovery and crystallography study of highly potent and selective butylcholinesterase inhibitors based on oxime-containing libraries and conformational restriction strategies - Jing_2023_Bioorg.Chem_134_106465
Author(s) : Jing L , Wei W , Meng B , Chantegreil F , Nachon F , Martinez A , Wu G , Zhao H , Song Y , Kang D , Brazzolotto X , Zhan P , Liu X
Ref : Bioorg Chem , 134 :106465 , 2023
Abstract : Butyrylcholinesterase is regarded as a promising drug target in advanced Alzheimer's disease. In order to identify highly selective and potent BuChE inhibitors, a 53-membered compound library was constructed via the oxime-based tethering approach based on microscale synthesis. Although A2Q17 and A3Q12 exhibited higher BuChE selectivity versus acetylcholinesterase, the inhibitory activities were unsatisfactory and A3Q12 did not inhibit Abeta1-42 peptide self-induced aggregation. With A2Q17 and A3Q12 as leads, a novel series of tacrine derivatives with nitrogen-containing heterocycles were designed based on conformation restriction strategy. The results demonstrated that 39 (IC50 = 3.49 nM) and 43 (IC50 = 7.44 nM) yielded much improved hBuChE inhibitory activity compared to the lead A3Q12 (IC50 = 63 nM). Besides, the selectivity indexes (SI = AChE IC50 / BChE IC50) of 39 (SI = 33) and 43 (SI = 20) were also higher than A3Q12 (SI = 14). The results of the kinetic study showed that 39 and 43 exhibited a mixed-type inhibition against eqBuChE with respective Ki values of 1.715 nM and 0.781 nM. And 39 and 43 could inhibit Abeta1-42 peptide self-induced aggregation into fibril. X-ray crystallography structures of 39 or 43 complexes with BuChE revealed the molecular basis for their high potency. Thus, 39 and 43 are deserve for further study to develop potential drug candidates for the treatment of Alzheimer's disease.
ESTHER : Jing_2023_Bioorg.Chem_134_106465
PubMedSearch : Jing_2023_Bioorg.Chem_134_106465
PubMedID: 36933339
Gene_locus related to this paper: human-BCHE

Title : 8-Hydroxyquinolylnitrones as multifunctional ligands for the therapy of neurodegenerative diseases - Knez_2023_Acta.Pharmaceutica.Sinica.B_13_2152
Author(s) : Knez D , Diez-Iriepa D , Chioua M , Gottinger A , Denic M , Chantegreil F , Nachon F , Brazzolotto X , Skrzypczak-Wiercioch A , Meden A , Pislar A , Kos J , Zakelj S , Stojan J , Salat K , Serrano J , Patricia Fernandez AP , Sanchez-Garcia A , Martinez-Murillo R , Binda C , Lopez-Munoz F , Gobec S , Marco-Contelle J
Ref : Acta Pharmaceutica Sinica B , 13 :2152 , 2023
Abstract : We describe the development of quinolylnitrones (QNs) as multifunctional ligands inhibiting cholinesterases (ChEs: acetylcholinesterase and butyrylcholinesterase -hBChE) and monoamine oxidases (hMAO-A/B) for the therapy of neurodegenerative diseases. We identified QN 19, a simple, low molecular weight nitrone, that is readily synthesized from commercially available 8-hydroxyquinoline-2-carbaldehyde. Quinolylnitrone 19 has no typical pharmacophoric element to suggest ChE or MAO inhibition, yet unexpectedly showed potent inhibition of hBChE (IC50 = 1.06 +/- 0.31 nmol/L) and hMAO-B (IC50 = 4.46 +/- 0.18 micromol/L). The crystal structures of 19 with hBChE and hMAO-B provided the structural basis for potent binding, which was further studied by enzyme kinetics. Compound 19 acted as a free radical scavenger and biometal chelator, crossed the blood-brain barrier, was not cytotoxic, and showed neuroprotective properties in a 6-hydroxydopamine cell model of Parkinson's disease. In addition, in vivo studies showed the anti-amnesic effect of 19 in the scopolamine-induced mouse model of AD without adverse effects on motoric function and coordination. Importantly, chronic treatment of double transgenic APPswe-PS1deltaE9 mice with 19 reduced amyloid plaque load in the hippocampus and cortex of female mice, underscoring the disease-modifying effect of QN 19
ESTHER : Knez_2023_Acta.Pharmaceutica.Sinica.B_13_2152
PubMedSearch : Knez_2023_Acta.Pharmaceutica.Sinica.B_13_2152
PubMedID: 37250172
Gene_locus related to this paper: human-BCHE

Title : Discovery of new, highly potent and selective inhibitors of BuChE - Design, synthesis, in vitro and in vivo evaluation and crystallography studies - Panek_2023_Eur.J.Med.Chem_249_115135
Author(s) : Panek D , Pasieka A , Latacz G , Zareba P , Szczech M , Godyn J , Chantegreil F , Nachon F , Brazzolotto X , Skrzypczak-Wiercioch A , Walczak M , Smolik M , Salat K , Hofner G , Wanner K , Wieckowska A , Malawska B
Ref : Eur Journal of Medicinal Chemistry , 249 :115135 , 2023
Abstract : The symptomatic and disease-modifying effects of butyrylcholinesterase (BuChE) inhibitors provide an encouraging premise for researching effective treatments for Alzheimer's disease. Here, we examined a series of compounds with a new chemical scaffold based on 3-(cyclohexylmethyl)amino-2-hydroxypropyl, and we identified a highly selective hBuChE inhibitor (29). Based on extensive in vitro and in vivo evaluations of the compound and its enantiomers, (R)-29 was identified as a promising candidate for further development. Compound (R)-29 is a potent hBuChE inhibitor (IC50 = 40 nM) with selectivity over AChE and relevant off-targets, including H1, M1, alpha1A and beta1 receptors. The compound displays high metabolic stability on human liver microsomes (90% of the parent compound after 2 h of incubation), and its safety was confirmed through examining the cytotoxicity on the HepG2 cell line (LC50 = 2.85 microM) and hERG inhibition (less than 50% at 10 microM). While (rac)-29 lacked an effect in vivo and showed limited penetration to the CNS in pharmacokinetics studies, compound (R)-29 exhibited a procognitive effect at 15 mg/kg in the novel object recognition task in scopolamine-treated mice.
ESTHER : Panek_2023_Eur.J.Med.Chem_249_115135
PubMedSearch : Panek_2023_Eur.J.Med.Chem_249_115135
PubMedID: 36696766
Gene_locus related to this paper: human-BCHE

Title : Grid-type Quaternary Metallosupramolecular Compounds Inhibit Human Cholinesterases through Dynamic Multivalent Interactions - Nachon_2022_Chembiochem_23_e202200456
Author(s) : Nachon F , Brazzolotto X , Dias J , Courageux C , Drozdz W , Cao XY , Stefankiewicz AR , Lehn JM
Ref : Chembiochem , 23 :e202200456 , 2022
Abstract : We report herein the implementation of coordination complexes containing two types of cationic moieties, i.e. pyridinium and ammonium quaternary salt, as potential inhibitors of human cholinesterase enzymes. Utilization of ligands containing NNO-coordination site and binding zinc metal ion allowed obtaining mono- and tetra-nuclear complexes with corner and grid structural type respectively, thus affecting the overall charge of the compounds (from +1 to +8). It enabled for the first time to examine the multivalency effect of metallosupramolecular species on their inhibitory abilities towards acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). Importantly, resolution of the crystal structures of the obtained enzyme-substrate complexes provided a better understanding of the inhibition process at the molecular level.
ESTHER : Nachon_2022_Chembiochem_23_e202200456
PubMedSearch : Nachon_2022_Chembiochem_23_e202200456
PubMedID: 36193860
Gene_locus related to this paper: human-BCHE

Title : From tryptophan-based amides to tertiary amines: Optimization of a butyrylcholinesterase inhibitor series - Meden_2022_Eur.J.Med.Chem_230_114248
Author(s) : Meden A , Knez D , Brazzolotto X , Nachon F , Dias J , Svete J , Stojan J , Groselj U , Gobec S
Ref : Eur Journal of Medicinal Chemistry , 230 :114248 , 2022
Abstract : Lead optimization of a series of tryptophan-based nanomolar butyrylcholinesterase (BChE) inhibitors led to tertiary amines as highly potent, achiral, sp rich analogues with better synthetic accessibility and high selectivity over acetylcholinesterase (one to ten thousandfold) (Chierrito et al., 2018).. Taking it one step further, the introduction of a carbamate warhead on the well-explored reversible scaffold allowed conversion to pseudoirreversible inhibitors that bound covalently to BChE and prolonged the duration of inhibition (half-life of 14.8 h for compound 45a-carbamoylated enzyme). Additionally, N-hydroxyindole was discovered as a novel leaving group chemotype. The covalent mechanism of action was confirmed by time-dependency experiments, progress curve analysis, and indirectly by co-crystallization with the human recombinant enzyme. Two crystal structures of BChE-inhibitor complexes were solved and coupled with the supporting molecular dynamics simulations increased our understanding of the structure-activity relationship, while also providing the neccessary structural infromation for future optimization of this series. Overall, this research demonstates the high versatility and potential of this series of BChE inhibitors.
ESTHER : Meden_2022_Eur.J.Med.Chem_230_114248
PubMedSearch : Meden_2022_Eur.J.Med.Chem_230_114248
PubMedID: 35299116
Gene_locus related to this paper: human-BCHE

Title : Pseudo-irreversible butyrylcholinesterase inhibitors: Structure-activity relationships, computational and crystallographic study of the N-dialkyl O-arylcarbamate warhead - Meden_2022_Eur.J.Med.Chem_247_115048
Author(s) : Meden A , Knez D , Brazzolotto X , Modeste F , Perdih A , Pislar A , Zorman M , Zorovic M , Denic M , Pajk S , Zivin M , Nachon F , Gobec S
Ref : Eur Journal of Medicinal Chemistry , 247 :115048 , 2022
Abstract : Alongside reversible butyrylcholinesterase inhibitors, a plethora of covalent butyrylcholinesterase inhibitors have been reported in the literature, typically pseudo-irreversible carbamates. For these latter, however, most cases lack full confirmation of their covalent mode of action. Additionally, the available reports regarding the structure-activity relationships of the O-arylcarbamate warhead are incomplete. Therefore, a follow-up on a series of pseudo-irreversible covalent carbamate human butyrylcholinesterase inhibitors and the structure-activity relationships of the N-dialkyl O-arylcarbamate warhead are presented in this study. The covalent mechanism of binding was tested by IC(50) time-dependency profiles, and sequentially and increasingly confirmed by kinetic analysis, whole protein LC-MS, and crystallographic analysis. Computational studies provided valuable insights into steric constraints and identified problematic, bulky carbamate warheads that cannot reach and carbamoylate the catalytic Ser198. Quantum mechanical calculations provided further evidence that steric effects appear to be a key factor in determining the covalent binding behaviour of these carbamate cholinesterase inhibitors and their duration of action. Additionally, the introduction of a clickable terminal alkyne moiety into one of the carbamate N-substituents and in situ derivatisation with azide-containing fluorophore enabled fluorescent labelling of plasma human butyrylcholinesterase. This proof-of-concept study highlights the potential of this novel approach and for these compounds to be further developed as clickable molecular probes for investigating tissue localisation and activity of cholinesterases.
ESTHER : Meden_2022_Eur.J.Med.Chem_247_115048
PubMedSearch : Meden_2022_Eur.J.Med.Chem_247_115048
PubMedID: 36586299
Gene_locus related to this paper: human-BCHE

Title : Pseudo-irreversible butyrylcholinesterase inhibitors: SAR, kinetic, computational, and crystallographic study of the N-dialkyl O-arylcarbamate warhead - Meden_2022_Chemrxiv__
Author(s) : Meden A , Knez D , Brazzolotto X , Modeste F , Perdih A , Pislar A , Zorman M , Denic M , Pajk S , Nachon F , Gobec S
Ref : Chemrxiv , : , 2022
Abstract : Beside reversible butyrylcholinesterse inhibitors (BChEIs), a plethora of covalent ones, typically pseudo-irreversible carbamates, have been reported in literature. For the latter, however, in most cases the proper confirmation of their covalent mode of action is lacking. Additionally, the available reports on the structure-activity relationships of the O-arylcarbamate warhead are incomplete. Therefore, a follow-up on a series of pseudo-irreversible covalent carbamate human butyrylcholinesterase inhibitors (hBChEIs) and the structure-activity relationships of the N-dialkyl O-arylcarbamate warhead is presented. The covalent mechanism of binding was tested by IC50 time-dependency profiles, and sequentially and increasingly confirmed by kinetic analysis, whole protein LC-MS, and crystallographic evidence. The computational studies provided valuable insights into the steric constraints and identified problematic, bulky carbamate warheads that could not reach and carbamoylate the catalytic Ser198. QM calculations lent further evidence that the steric effects seemed to be a key factor in determining the covalent binding behaviour of these carbamate ChEIs and their duration of action. Furthermore, the introduction of a clickable terminal alkyne moiety into one of the carbamate N-substituents and in situ derivatization with an azide-containing fluorophore enabled fluorescent labelling of plasma hBChE. This proof-of-concept study highlighted the potential of this novel approach and these compounds to be further developed as clickable molecular probes for investigating tissue localization and activity of ChEs
ESTHER : Meden_2022_Chemrxiv__
PubMedSearch : Meden_2022_Chemrxiv__
Gene_locus related to this paper: human-BCHE

Title : Discovery of 1-(phenylsulfonyl)-1H-indole-based multifunctional ligands targeting cholinesterases and 5-HT(6) receptor with anti-aggregation properties against amyloid-beta and tau - Wichur_2021_Eur.J.Med.Chem_225_113783
Author(s) : Wichur T , Pasieka A , Godyn J , Panek D , Goral I , Latacz G , Honkisz-Orzechowska E , Bucki A , Siwek A , Gluch-Lutwin M , Knez D , Brazzolotto X , Gobec S , Kolaczkowski M , Sabate R , Malawska B , Wieckowska A
Ref : Eur Journal of Medicinal Chemistry , 225 :113783 , 2021
Abstract : Multifunctional ligands as an essential variant of polypharmacology are promising candidates for the treatment of multi-factorial diseases like Alzheimer's disease. Based on clinical evidence and following the paradigm of multifunctional ligands we have rationally designed and synthesized a series of compounds targeting processes involved in the development of the disease. The biological evaluation led to the discovery of two compounds with favorable pharmacological characteristics and ADMET profile. Compounds 17 and 35 are 5-HT(6)R antagonists (K(i) = 13 nM and K(i) = 15 nM respectively) and cholinesterase inhibitors with distinct mechanisms of enzyme inhibition. Compound 17, a tacrine derivative is a reversible inhibitor of acetyl- and butyrylcholinesterase (IC(50) = 8 nM and IC(50) = 24 nM respectively), while compound 35 with rivastigmine-derived phenyl N-ethyl-N-methylcarbamate fragment is a selective, pseudo-irreversible inhibitor of butyrylcholinesterase (IC(50) = 455 nM). Both compounds inhibit aggregation of amyloid beta in vitro (75% for compound 17 and 68% for 35 at 10 microM) moreover, compound 35 is a potent tau aggregation inhibitor in cellulo (79%). In ADMET in vitro studies both compounds showed acceptable metabolic stability on mouse liver microsomes (28% and 60% for compound 17 and 35 respectively), no or little effect on CYP3A4 and 2D6 up to a concentration of 10 microM and lack of toxicity on HepG2 cell line (IC(50) values of 80 and 21 microM, for 17 and 35 respectively). Based on the pharmacological characteristics and favorable pharmacokinetic properties, we propose compounds 17 and 35 as an excellent starting point for further optimization and in-depth biological studies.
ESTHER : Wichur_2021_Eur.J.Med.Chem_225_113783
PubMedSearch : Wichur_2021_Eur.J.Med.Chem_225_113783
PubMedID: 34461507

Title : Discovery of drug-like acetylcholinesterase inhibitors by rapid virtual screening of a 6.9 million compound database - Miles_2021_Chem.Biol.Drug.Des__
Author(s) : Miles JA , Ng JH , Sreenivas BY , Courageux C , Igert A , Dias J , McGeary RP , Brazzolotto X , Ross BP
Ref : Chemical Biology Drug Des , : , 2021
Abstract : Cholinesterase inhibitors remain the mainstay of Alzheimer's disease treatment, and the search for new inhibitors with better efficacy and side effect profiles is ongoing. Virtual screening (VS) is a powerful technique for searching large compound databases for potential hits. This study used a sequential VS workflow combining ligand-based VS, molecular docking and physicochemical filtering to screen for CNS drug-like acetylcholinesterase inhibitors (AChEIs) amongst the 6.9 million compounds of the CoCoCo database. Eleven in silico hits were initially selected, resulting in the discovery of an AChEI with a K(i) of 3.2 microM. In vitro kinetics and in silico molecular dynamics experiments informed the selection of an additional seven analogues. This led to the discovery of two further AChEIs, with K(i) values of 2.9 microM and 0.65 microM. All three compounds exhibited reversible, mixed inhibition of AChE. Importantly, the in silico physicochemical filter facilitated the discovery of CNS drug-like compounds, such that all three inhibitors displayed high in vitro blood-brain barrier model permeability.
ESTHER : Miles_2021_Chem.Biol.Drug.Des__
PubMedSearch : Miles_2021_Chem.Biol.Drug.Des__
PubMedID: 33455074

Title : Discovery of a Potent Dual Inhibitor of Acetylcholinesterase and Butyrylcholinesterase with Antioxidant Activity that Alleviates Alzheimer-like Pathology in Old APP\/PS1 Mice - Viayna_2021_J.Med.Chem_64_812
Author(s) : Viayna E , Coquelle N , Cieslikiewicz-Bouet M , Cisternas P , Oliva CA , Sanchez-Lopez E , Ettcheto M , Bartolini M , De Simone A , Ricchini M , Rendina M , Pons M , Firuzi O , Perez B , Saso L , Andrisano V , Nachon F , Brazzolotto X , Garcia ML , Camins A , Silman I , Jean L , Inestrosa NC , Colletier JP , Renard PY , Munoz-Torrero D
Ref : Journal of Medicinal Chemistry , 64 :812 , 2021
Abstract : The combination of the scaffolds of the cholinesterase inhibitor huprine Y and the antioxidant capsaicin results in compounds with nanomolar potencies toward human acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) that retain or improve the antioxidant properties of capsaicin. Crystal structures of their complexes with AChE and BChE revealed the molecular basis for their high potency. Brain penetration was confirmed by biodistribution studies in C57BL6 mice, with one compound (5i) displaying better brain/plasma ratio than donepezil. Chronic treatment of 10 month-old APP/PS1 mice with 5i (2 mg/kg, i.p., 3 times per week, 4 weeks) rescued learning and memory impairments, as measured by three different behavioral tests, delayed the Alzheimer-like pathology progression, as suggested by a significantly reduced Abeta42/Abeta40 ratio in the hippocampus, improved basal synaptic efficacy, and significantly reduced hippocampal oxidative stress and neuroinflammation. Compound 5i emerges as an interesting anti-Alzheimer lead with beneficial effects on cognitive symptoms and on some underlying disease mechanisms.
ESTHER : Viayna_2021_J.Med.Chem_64_812
PubMedSearch : Viayna_2021_J.Med.Chem_64_812
PubMedID: 33356266
Gene_locus related to this paper: human-ACHE

Title : Diisopropylfluorophosphate-induced status epilepticus drives complex glial cell phenotypes in adult male mice - Maupu_2021_Neurobiol.Dis__105276
Author(s) : Maupu C , Enderlin J , Igert A , Oger M , Auvin S , Hassan-Abdi R , Soussi-Yanicostas N , Brazzolotto X , Nachon F , Bo GD , Dupuis N
Ref : Neurobiol Dis , :105276 , 2021
Abstract : Organophosphate pesticides and nerve agents (OPs), are characterized by cholinesterase inhibition. In addition to severe peripheral symptoms, high doses of OPs can lead to seizures and status epilepticus (SE). Long lasting seizure activity and subsequent neurodegeneration promote neuroinflammation leading to profound pathological alterations of the brain. The aim of this study was to characterize neuroinflammatory responses at key time points after SE induced by the OP, diisopropylfluorophosphate (DFP). Immunohistochemistry (IHC) analysis and RT-qPCR on cerebral tissue are often insufficient to identity and quantify precise neuroinflammatory alterations. To address these needs, we performed RT-qPCR quantification after whole brain magnetic-activated cell-sorting (MACS) of CD11B (microglia/infiltrated macrophages) and GLAST (astrocytes)-positive cells at 1, 4, 24 h and 3 days post-SE. In order to compare these results to those obtained by IHC, we performed, classical Iba1 (microglia/infiltrated macrophages) and GFAP (astrocytes) IHC analysis in parallel, focusing on the hippocampus, a brain region affected by seizure activity and neurodegeneration. Shortly after SE (1-4 h), an increase in pro-inflammatory (M1-like) markers and A2-specific markers, proposed as neurotrophic, were observed in CD11B and GLAST-positive isolated cells, respectively. Microglial cells successively expressed immuno-regulatory (M2b-like) and anti-inflammatory (M2a-like) at 4 h and 24 h post-SE induction. At 24 h and 3 days, A1-specific markers, proposed as neurotoxic, were increased in isolated astrocytes. Although IHC analysis presented no modification in terms of percentage of marked area and cell number at 1 and 4 h after SE, at 24 h and 3 days after SE, microglial and astrocytic activation was visible by IHC as an increase in Iba1 and GFAP-positive area and Iba1-positive cells in DFP animals when compared to the control. Our work identified sequential microglial and astrocytic phenotype activation. Although the role of each phenotype in SE cerebral outcomes requires further study, targeting specific markers at specific time point could be a beneficial strategy for DFP-induced SE treatment.
ESTHER : Maupu_2021_Neurobiol.Dis__105276
PubMedSearch : Maupu_2021_Neurobiol.Dis__105276
PubMedID: 33529768

Title : Characterization of four BCHE mutations associated with prolonged effect of suxamethonium - Brazzolotto_2021_Pharmacogenomics.J_21_165
Author(s) : Brazzolotto X , Courcelle S , Sauvanet C , Guillon V , Igert A , Kononchik J , Nachon F , Ceppa F , Delacour H
Ref : Pharmacogenomics J , 21(2):165-173 : , 2021
Abstract : Butyrylcholinesterase (BChE) deficiency is characterized by prolonged apnea after the use of muscle relaxants (suxamethonium or mivacurium) in patients who have mutations in the BCHE gene. Here, we report the characterization of four BCHE mutations associated with prolonged effect of suxamethonium (amino acid numbering based on the matured enzyme): p.20delValPheGlyGlyThrValThr, p.Leu88His, p.Ile140del and p.Arg386Cys. Expression of recombinant BCHE mutants, kinetic analysis and molecular dynamics were undertaken to understand how these mutations induce BChE deficiency. Three of the mutations studied (p.20delValPheGlyGlyThrValThr, p.Ile140del and p.Arg386Cys) lead to a "silent" BChE phenotype. Recombinant BCHE expression studies for these mutants revealed BChE activity levels comparable to untransfected cells. Only the last one (hBChE-L88H) presented BChE activity in the transfected cell culture medium. This BChE mutant (p.Leu88His) is associated with a lower k(cat) value compare to the wild-type enzyme. Molecular dynamics simulations analyses suggest that a destabilization of a structure implicated in enzyme activity (-loop) can explain the modification of the kinetic parameter of the mutated protein.
ESTHER : Brazzolotto_2021_Pharmacogenomics.J_21_165
PubMedSearch : Brazzolotto_2021_Pharmacogenomics.J_21_165
PubMedID: 33024248

Title : Development and crystallography-aided SAR studies of multifunctional BuChE inhibitors and 5-HT6R antagonists with beta-amyloid anti-aggregation properties - Wichur_2021_Eur.J.Med.Chem_225_113792
Author(s) : Wichur T , Godyn J , Goral I , Latacz G , Bucki A , Siwek A , Gluch-Lutwin M , Mordyl B , Sniecikowska J , Walczak M , Knez D , Jukic M , Salat K , Gobec S , Kolaczkowski M , Malawska B , Brazzolotto X , Wieckowska A
Ref : Eur Journal of Medicinal Chemistry , 225 :113792 , 2021
Abstract : The lack of an effective treatment makes Alzheimer's disease a serious healthcare problem and a challenge for medicinal chemists. Herein we report interdisciplinary research on novel multifunctional ligands targeting proteins and processes involved in the development of the disease: BuChE, 5-HT6 receptors and beta-amyloid aggregation. Structure-activity relationship analyses supported by crystallography and docking studies led to the identification of a fused-type multifunctional ligand 50, with remarkable and balanced potencies against BuChE (IC50 = 90 nM) and 5-HT6R (Ki = 4.8 nM), and inhibitory activity against Abeta aggregation (53% at 10 microM). In in vitro ADME-Tox and in vivo pharmacokinetic studies compound 50 showed good stability in the mouse liver microsomes, favourable safety profile and brain permeability with the brain to plasma ratio of 6.79 after p.o. administration in mice, thus being a promising candidate for in vivo pharmacology studies and a solid foundation for further research on effective anti-AD therapies.
ESTHER : Wichur_2021_Eur.J.Med.Chem_225_113792
PubMedSearch : Wichur_2021_Eur.J.Med.Chem_225_113792
PubMedID: 34530376
Gene_locus related to this paper: human-BCHE

Title : A Thermophilic Bacterial Esterase for Scavenging Nerve Agents: A Kinetic, Biophysical and Structural Study - Bzdrenga_2021_Molecules_26_657
Author(s) : Bzdrenga J , Trenet E , Chantegreil F , Bernal K , Nachon F , Brazzolotto X
Ref : Molecules , 26 :657 , 2021
Abstract : Organophosphorous nerve agents (OPNA) pose an actual and major threat for both military and civilians alike, as an upsurge in their use has been observed in the recent years. Currently available treatments mitigate the effect of the nerve agents, and could be vastly improved by means of scavengers of the nerve agents. Consequently, efforts have been made over the years into investigating enzymes, also known as bioscavengers, which have the potential either to trap or hydrolyze these toxic compounds. We investigated the previously described esterase 2 from Thermogutta terrifontis (TtEst2) as a potential bioscavenger of nerve agents. As such, we assessed its potential against G-agents (tabun, sarin, and cyclosarin), VX, as well as the pesticide paraoxon. We report that TtEst2 is a good bioscavenger of paraoxon and G-agents, but is rather slow at scavenging VX. X-ray crystallography studies showed that TtEst2 forms an irreversible complex with the aforementioned agents, and allowed the identification of amino-acids, whose mutagenesis could lead to better scavenging properties for VX. In conjunction with its cheap production and purification processes, as well as a robust structural backbone, further engineering of TtEst2 could lead to a stopgap bioscavenger useful for in corpo scavenging or skin decontamination.
ESTHER : Bzdrenga_2021_Molecules_26_657
PubMedSearch : Bzdrenga_2021_Molecules_26_657
PubMedID: 33513869
Gene_locus related to this paper: 9bact-TtEst2

Title : Fine-Tuning the Biological Profile of Multitarget Mitochondriotropic Antioxidants for Neurodegenerative Diseases - Chavarria_2021_Antioxidants.(Basel)_10_329
Author(s) : Chavarria D , Da Silva O , Benfeito S , Barreiro S , Garrido J , Cagide F , Soares P , Remiao F , Brazzolotto X , Nachon F , Oliveira PJ , Dias J , Borges F
Ref : Antioxidants (Basel) , 10 :329 , 2021
Abstract : Neurotransmitter depletion and mitochondrial dysfunction are among the multiple pathological events that lead to neurodegeneration. Following our previous studies related with the development of multitarget mitochondriotropic antioxidants, this study aims to evaluate whether the Pi-system extension on the chemical scaffolds of AntiOXCIN2 and AntiOXCIN3 affects their bioactivity and safety profiles. After the synthesis of four triphenylphosphonium (TPP(+)) conjugates (compounds 2-5), we evaluated their antioxidant properties and their effect on neurotransmitter-metabolizing enzymes. All compounds were potent equine butyrylcholinesterase (eqBChE) and moderate electric eel acetylcholinesterase (eeAChE) inhibitors, with catechols 4 and 5 presenting lower IC(50) values than AntiOXCIN2 and AntiOXCIN3, respectively. However, differences in the inhibition potency and selectivity of compounds 2-5 towards non-human and human cholinesterases (ChEs) were observed. Co-crystallization studies with compounds 2-5 in complex with human ChEs (hChEs) showed that these compounds exhibit different binging modes to hAChE and hBChE. Unlike AntiOXCINs, compounds 2-5 displayed moderate human monoamine oxidase (hMAO) inhibitory activity. Moreover, compounds 4 and 5 presented higher ORAC-FL indexes and lower oxidation potential values than the corresponding AntiOXCINs. Catechols 4 and 5 exhibited broader safety windows in differentiated neuroblastoma cells than benzodioxole derivatives 2 and 3. Compound 4 is highlighted as a safe mitochondria-targeted antioxidant with dual ChE/MAO inhibitory activity. Overall, this work is a contribution for the development of dual therapeutic agents addressing both mitochondrial oxidative stress and neurotransmitter depletion.
ESTHER : Chavarria_2021_Antioxidants.(Basel)_10_329
PubMedSearch : Chavarria_2021_Antioxidants.(Basel)_10_329
PubMedID: 33672269
Gene_locus related to this paper: human-ACHE , human-BCHE

Title : Discovery of multifunctional anti-Alzheimers agents with a unique mechanism of action including inhibition of the enzyme butyrylcholinesterase and gamma-aminobutyric acid transporters - Pasieka_2021_Eur.J.Med.Chem_218_113397
Author(s) : Pasieka A , Panek D , Jonczyk J , Godyn J , Szalaj N , Latacz G , Tabor J , Mezeiova E , Chantegreil F , Dias J , Knez D , Lu J , Pi R , Korabecny J , Brazzolotto X , Gobec S , Hofner G , Wanner K , Wieckowska Q , Malawska B
Ref : Eur Journal of Medicinal Chemistry , 218 :113397 , 2021
Abstract : Looking for an effective anti-Alzheimer's agent is very challenging; however, a multifunctional ligand strategy may be a promising solution for the treatment of this complex disease. We herein present the design, synthesis and biological evaluation of novel hydroxyethylamine derivatives displaying unique, multiple properties that have not been previously reported. The original mechanism of action combines inhibitory activity against disease-modifying targets: beta-secretase enzyme (BACE1) and amyloid beta (Abeta) aggregation, along with an effect on targets associated with symptom relief - inhibition of butyrylcholinesterase (BuChE) and gamma-aminobutyric acid transporters (GATs). Among the obtained molecules, compound 36 exhibited the most balanced and broad activity profile (eeAChE IC50 = 2.86 microM; eqBuChE IC50 = 60 nM; hBuChE IC50 = 20 nM; hBACE1 IC50 = 5.9 microM; inhibition of Abeta aggregation = 57.9% at 10 microM; mGAT1 IC50 = 10.96 microM; and mGAT2 IC50 = 19.05 microM). Moreover, we also identified 31 as the most potent mGAT4 and hGAT3 inhibitor (IC50 = 5.01 microM and IC50 = 2.95 microM, respectively), with high selectivity over other subtypes. Compounds 36 and 31 represent new anti-Alzheimer agents that can ameliorate cognitive decline and modify the progress of disease.
ESTHER : Pasieka_2021_Eur.J.Med.Chem_218_113397
PubMedSearch : Pasieka_2021_Eur.J.Med.Chem_218_113397
PubMedID: 33838585

Title : Rapid discovery of a selective butyrylcholinesterase inhibitor using structure-based virtual screening - Miles_2020_Bioorg.Med.Chem.Lett__127609
Author(s) : Miles JA , Kapure JS , Singh Deora G , Courageux C , Igert A , Dias J , McGeary RP , Brazzolotto X , Ross BP
Ref : Bioorganic & Medicinal Chemistry Lett , :127609 , 2020
Abstract : Acetylcholinesterase inhibitors are the mainstay of Alzheimer's disease treatments, despite only short-term symptomatic benefits and severe side effects. Selective butyrylcholinesterase inhibitors (BuChEIs) may be more effective treatments in late-stage Alzheimer's disease with fewer side effects. Virtual screening is a powerful tool for identifying potential inhibitors in large digital compound databases. This study used structure-based virtual screening combined with physicochemical filtering to screen two the InterBioScreen and Maybridge databases for novel selective BuChEIs. The workflow rapidly identified 22 potential hits in silico, resulting in the discovery of a human BuChEI with low-micromolar potency in vitro (IC(50) 2.4 muM) and high selectivity for butyrylcholinesterase over acetylcholinesterase. The compound was a rapidly reversible BuChEI with mixed-model in vitro inhibition kinetics. The binding interactions were investigated using in silico molecular dynamics, and by developing structure-activity relationships using nine analogues. The compound also displayed high permeability in an in vitro model of the blood-brain barrier.
ESTHER : Miles_2020_Bioorg.Med.Chem.Lett__127609
PubMedSearch : Miles_2020_Bioorg.Med.Chem.Lett__127609
PubMedID: 33039562

Title : Structure-activity relationship study of tryptophan-based butyrylcholinesterase inhibitors - Meden_2020_Eur.J.Med.Chem_208_112766
Author(s) : Meden A , Knez D , Malikowska-Racia N , Brazzolotto X , Nachon F , Svete J , Salat K , Groselj U , Gobec S
Ref : Eur Journal of Medicinal Chemistry , 208 :112766 , 2020
Abstract : A series of tryptophan-based selective nanomolar butyrylcholinesterase (BChE) inhibitors was designed and synthesized. Compounds were optimized in terms of potency, selectivity, and synthetic accessibility. The crystal structure of the inhibitor 18 in complex with BChE revealed the molecular basis for its low nanomolar inhibition (IC50 = 2.8 nM). The favourable in vitro results enabled a first-in-animal in vivo efficacy and safety trial, which demonstrated a positive impact on fear-motivated and spatial long-term memory retrieval without any concomitant adverse motor effects. Altogether, this research culminated in a handful of new lead compounds with promising potential for symptomatic treatment of patients with Alzheimers disease.
ESTHER : Meden_2020_Eur.J.Med.Chem_208_112766
PubMedSearch : Meden_2020_Eur.J.Med.Chem_208_112766
PubMedID: 32919297
Gene_locus related to this paper: human-BCHE

Title : N-alkylpiperidine carbamates as potential anti-Alzheimer's agents - Kosak_2020_Eur.J.Med.Chem_197_112282
Author(s) : Kosak U , Strasek N , Knez N , Jukic M , Zakelj S , Zahirovic A , Pislar A , Brazzolotto X , Nachon F , Kos J , Gobec S , Knez D
Ref : Eur Journal of Medicinal Chemistry , 197 :112282 , 2020
Abstract : Compounds capable of interacting with single or multiple targets involved in Alzheimer's disease (AD) pathogenesis are potential anti-Alzheimer's agents. In our aim to develop new anti-Alzheimer's agents, a series of 36 new N-alkylpiperidine carbamates was designed, synthesized and evaluated for the inhibition of cholinesterases [acetylcholinesterase (AChE) and butyrylcholinesterase (BChE)] and monoamine oxidases [monoamine oxidase A (MAO-A and monoamine oxidase B (MAO-B)]. Four compounds are very promising: multiple AChE (IC50 = 7.31 muM), BChE (IC50 = 0.56 muM) and MAO-B (IC50 = 26.1 muM) inhibitor 10, dual AChE (IC50 = 2.25 muM) and BChE (IC50 = 0.81 muM) inhibitor 22, selective BChE (IC50 = 0.06 muM) inhibitor 13, and selective MAO-B (IC50 = 0.18 muM) inhibitor 16. Results of enzyme kinetics experiments showed that despite the carbamate group in the structure, compounds 10, 13, and 22 are reversible and non-time-dependent inhibitors of AChE and/or BChE. The resolved crystal structure of the complex of BChE with compound 13 confirmed the non-covalent mechanism of inhibition. Additionally, N-propargylpiperidine 16 is an irreversible and time-dependent inhibitor of MAO-B, while N-benzylpiperidine 10 is reversible. Additionally, compounds 10, 13, 16, and 22 should be able to cross the blood-brain barrier and are not cytotoxic to human neuronal-like SH-SY5Y and liver HepG2 cells. Finally, compounds 10 and 16 also prevent amyloid beta1-42 (Abeta1-42)-induced neuronal cell death. The neuroprotective effects of compound 16 could be the result of its Abeta1-42 anti-aggregation effects.
ESTHER : Kosak_2020_Eur.J.Med.Chem_197_112282
PubMedSearch : Kosak_2020_Eur.J.Med.Chem_197_112282
PubMedID: 32380361
Gene_locus related to this paper: human-BCHE

Title : Development of potent reversible selective inhibitors of butyrylcholinesterase as fluorescent probes - Pajk_2020_J.Enzyme.Inhib.Med.Chem_35_498
Author(s) : Pajk S , Knez D , Kosak U , Zorovic M , Brazzolotto X , Coquelle N , Nachon F , Colletier JP , Zivin M , Stojan J , Gobec S
Ref : J Enzyme Inhib Med Chem , 35 :498 , 2020
Abstract : Brain butyrylcholinesterase (BChE) is an attractive target for drugs designed for the treatment of Alzheimer's disease (AD) in its advanced stages. It also potentially represents a biomarker for progression of this disease. Based on the crystal structure of previously described highly potent, reversible, and selective BChE inhibitors, we have developed the fluorescent probes that are selective towards human BChE. The most promising probes also maintain their inhibition of BChE in the low nanomolar range with high selectivity over acetylcholinesterase. Kinetic studies of probes reveal a reversible mixed inhibition mechanism, with binding of these fluorescent probes to both the free and acylated enzyme. Probes show environment-sensitive emission, and additionally, one of them also shows significant enhancement of fluorescence intensity upon binding to the active site of BChE. Finally, the crystal structures of probes in complex with human BChE are reported, which offer an excellent base for further development of this library of compounds.
ESTHER : Pajk_2020_J.Enzyme.Inhib.Med.Chem_35_498
PubMedSearch : Pajk_2020_J.Enzyme.Inhib.Med.Chem_35_498
PubMedID: 31914836
Gene_locus related to this paper: human-BCHE

Title : Enantioseparation, in vitro testing, and structural characterization of triple-binding reactivators of organophosphate-inhibited cholinesterases - Marakovic_2020_Biochem.J_477_2771
Author(s) : Marakovic N , Knezevic A , Roncevic I , Brazzolotto X , Kovarik Z , Sinko G
Ref : Biochemical Journal , 477 :2771 , 2020
Abstract : The enantiomers of racemic 2-hydroxyimino-N-(azidophenylpropyl)acetamide-derived triple-binding oxime reactivators were separated, and tested for inhibition and reactivation of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibited with tabun (GA), cyclosarin (GF), sarin (GB), and VX. Both enzymes showed the greatest affinity toward the methylimidazole derivative (III) of 2-hydroxyimino-N-(azidophenylpropyl)acetamide (I). The crystal structure was determined for the complex of oxime III within human BChE, confirming that all three binding groups interacted with active site residues. In the case of BChE inhibited by GF, oximes I (kr=207M-1min-1) and III (kr=213M-1min-1) showed better reactivation efficiency than the reference oxime 2-PAM. Finally, the key mechanistic steps in the reactivation of GF-inhibited BChE with oxime III were modeled using the PM7R6 method, stressing the importance of proton transfer from Nsigma of His438 to Ogamma of Ser203 for achieving successful reactivation.
ESTHER : Marakovic_2020_Biochem.J_477_2771
PubMedSearch : Marakovic_2020_Biochem.J_477_2771
PubMedID: 32639532
Gene_locus related to this paper: human-BCHE

Title : Efficacy Assessment of an Uncharged Reactivator of NOP-Inhibited Acetylcholinesterase Based on Tetrahydroacridine Pyridine-Aldoxime Hybrid in Mouse Compared to Pralidoxime - Calas_2020_Biomolecules_10_
Author(s) : Calas AG , Hanak AS , Jaffre N , Nervo A , Dias J , Rousseau C , Courageux C , Brazzolotto X , Villa P , Obrecht A , Goossens JF , Landry C , Hachani J , Gosselet F , Dehouck MP , Yerri J , Kliachyna M , Baati R , Nachon F
Ref : Biomolecules , 10 : , 2020
Abstract : (1) Background: Human exposure to organophosphorus compounds employed as pesticides or as chemical warfare agents induces deleterious effects due to cholinesterase inhibition. One therapeutic approach is the reactivation of inhibited acetylcholinesterase by oximes. While currently available oximes are unable to reach the central nervous system to reactivate cholinesterases or to display a wide spectrum of action against the variety of organophosphorus compounds, we aim to identify new reactivators without such drawbacks. (2) Methods: This study gathers an exhaustive work to assess in vitro and in vivo efficacy, and toxicity of a hybrid tetrahydroacridine pyridinaldoxime reactivator, KM297, compared to pralidoxime. (3) Results: Blood-brain barrier crossing assay carried out on a human in vitro model established that KM297 has an endothelial permeability coefficient twice that of pralidoxime. It also presents higher cytotoxicity, particularly on bone marrow-derived cells. Its strong cholinesterase inhibition potency seems to be correlated to its low protective efficacy in mice exposed to paraoxon. Ventilatory monitoring of KM297-treated mice by double-chamber plethysmography shows toxic effects at the selected therapeutic dose. This breathing assessment could help define the No Observed Adverse Effect Level (NOAEL) dose of new oximes which would have a maximum therapeutic effect without any toxic side effects.
ESTHER : Calas_2020_Biomolecules_10_
PubMedSearch : Calas_2020_Biomolecules_10_
PubMedID: 32512884

Title : X-ray structures of human bile-salt activated lipase conjugated to nerve agents surrogates - Touvrey_2019_Toxicology_411_15
Author(s) : Touvrey C , Courageux C , Guillon V , Terreux R , Nachon F , Brazzolotto X
Ref : Toxicology , 411 :15 , 2019
Abstract : The efficiency of human butyrylcholinesterase (BChE) as a stoichiometric bioscavenger of nerve agents is well established. However, wide use is currently limited by production and purification costs. Aiming at identifying an alternative human protein bioscavenger, we looked for an original scaffold candidate by virtual screening of the Protein Data Bank for functional similarity using the "Surfing the Molecules" software ( and a search model based on the BChE active site topology. Besides the expected acetylcholinesterase and butyrylcholinesterase, we identified a set of bile salt activated lipases structures, among which the human pancreatic lipase (hBAL) that shares 34% identity with BChE. We produced the recombinant enzyme in mammalian cells, purified it, and measured the inhibition constants for paraoxon and surrogates of VX, sarin and tabun. We solved the X-ray structure of apo hBAL and conjugates with paraoxon and the surrogates at resolutions in the 2-A range. These structures allow the assessment of hBAL for scavenging nerve agents. They revealed that hBAL has inverted stereoselectivity for the surrogates of nerve agent compared to human cholinesterases. We observed a remarkable flip of the catalytic histidine driven by the chelation of Zn(2+). Dealkylation of the conjugate, aka aging, was solely observed for paraoxon.
ESTHER : Touvrey_2019_Toxicology_411_15
PubMedSearch : Touvrey_2019_Toxicology_411_15
PubMedID: 30359675
Gene_locus related to this paper: human-CEL

Title : Rivastigmine and metabolite analogues with putative Alzheimer's disease-modifying properties in a Caenorhabditis elegans model. - Dighe_2019_Commun.Chem_2_35
Author(s) : Dighe SN , De la Mora E , Chan S , Kantham S , McColl G , Miles JA , Veliyath SK , Sreenivas BY , Nassar ZD , Silman I , Sussman JL , Weik M , McGeary RP , Parat MO , Brazzolotto X , Ross BP
Ref : Communications chemistry , 2 :35 , 2019
Abstract : The development of polyphenols as drugs for Alzheimer's disease (AD) is thwarted by their meagre brain availability due to instability and poor druglikeness. Here we describe the successful development of stable, druglike polyphenolic analogues of the current AD drug rivastigmine, that have high apparent blood-brain barrier permeabilities and multifunctional properties for AD treatment. The compounds inhibit cholinesterases and amyloid beta (Abeta) fibrillation, protect against Abeta42-induced toxicity in vitro, and demonstrate efficacy in vivo in a transgenic Caenorhabditis elegans model expressing Abeta42, with potencies similar to rivastigmine and natural polyphenols. The results suggest that a tertiary amine substituent is amenable for developing water-soluble, membrane-permeable polyphenols, and its incorporation adjacent to a hydroxy group is favourable for intramolecular hydrogen bonding that facilitates membrane permeability. Carbamylation of one hydroxy group protects the polyphenols from degradation and mostly improves their membrane permeability. These design strategies may assist in the development of polyphenol-based drugs.
ESTHER : Dighe_2019_Commun.Chem_2_35
PubMedSearch : Dighe_2019_Commun.Chem_2_35
Gene_locus related to this paper: human-BCHE , torca-ACHE

Title : Novel tacrine-tryptophan hybrids: Multi-target directed ligands as potential treatment for Alzheimer's disease - Chalupova_2019_Eur.J.Med.Chem_168_491
Author(s) : Chalupova K , Korabecny J , Bartolini M , Monti B , Lamba D , Caliandro R , Pesaresi A , Brazzolotto X , Gastellier AJ , Nachon F , Pejchal J , Jarosova M , Hepnarova V , Jun D , Hrabinova M , Dolezal R , Karasova JZ , Mzik M , Kristofikova Z , Misik J , Muckova L , Jost P , Soukup O , Benkova M , Setnicka V , Habartova L , Chvojkova M , Kleteckova L , Vales K , Mezeiova E , Uliassi E , Valis M , Nepovimova E , Bolognesi ML , Kuca K
Ref : Eur Journal of Medicinal Chemistry , 168 :491 , 2019
Abstract : A combination of tacrine and tryptophan led to the development of a new family of heterodimers as multi-target agents with potential to treat Alzheimer's disease. Based on the in vitro biological profile, compound S-K1035 was found to be the most potent inhibitor of human acetylcholinesterase (hAChE) and human butyrylcholinesterase (hBChE), demonstrating balanced IC50 values of 6.3 and 9.1nM, respectively. For all the tacrine-tryptophan heterodimers, favorable inhibitory effect on hAChE as well as on hBChE was coined to the optimal spacer length ranging from five to eight carbon atoms between these two pharmacophores. S-K1035 also showed good ability to inhibit Abeta42 self-aggregation (58.6+/-5.1% at 50muM) as well as hAChE-induced Abeta40 aggregation (48.3+/-6.3% at 100muM). The X-ray crystallographic analysis of TcAChE in complex with S-K1035 pinpointed the utility of the hybridization strategy applied and the structures determined with the two K1035 enantiomers in complex with hBChE could explain the higher inhibition potency of S-K1035. Other in vitro evaluations predicted the ability of S-K1035 to cross blood-brain barrier and to exert a moderate inhibition potency against neuronal nitric oxide synthase. Based on the initial promising biochemical data and a safer in vivo toxicity compared to tacrine, S-K1035 was administered to scopolamine-treated rats being able to dose-dependently revert amnesia.
ESTHER : Chalupova_2019_Eur.J.Med.Chem_168_491
PubMedSearch : Chalupova_2019_Eur.J.Med.Chem_168_491
PubMedID: 30851693
Gene_locus related to this paper: torca-ACHE

Title : Tryptophan-derived butyrylcholinesterase inhibitors as promising leads against Alzheimer's disease - Meden_2019_Chem.Commun.(Camb)_55_3765
Author(s) : Meden A , Knez D , Jukic M , Brazzolotto X , Grsic M , Pislar A , Zahirovic A , Kos J , Nachon F , Svete J , Gobec S , Groselj U
Ref : Chem Commun (Camb) , 55 :3765 , 2019
Abstract : We have identified tryptophan-based selective nanomolar butyrylcholinesterase (BChE) inhibitors. They are defined according to their chemical modularity, novel binding mode revealed by five solved crystal structures with human BChE, low cytotoxicity, and predicted permeability of the blood-brain barrier. Altogether, these factors indicate their potential as unique lead compounds for symptomatic therapy against Alzheimer's disease.
ESTHER : Meden_2019_Chem.Commun.(Camb)_55_3765
PubMedSearch : Meden_2019_Chem.Commun.(Camb)_55_3765
PubMedID: 30864579
Gene_locus related to this paper: human-BCHE

Title : An evolutionary perspective on the first disulfide bond in members of the cholinesterase-carboxylesterase (COesterase) family: Possible outcomes for cholinesterase expression in prokaryotes - Chatonnet_2019_Chem.Biol.Interact_13ChEPon_308_179
Author(s) : Chatonnet A , Brazzolotto X , Hotelier T , Lenfant N , Marchot P , Bourne Y
Ref : Chemico-Biological Interactions , 308 :179 , 2019
Abstract : Within the alpha/beta hydrolase fold superfamily of proteins, the COesterase group (carboxylesterase type B, block C, cholinesterases ...) diverged from the other groups through simultaneous integration of an N-terminal, first disulfide bond and a significant increase in the protein mean size. This first disulfide bond ties a large Cys loop, which in the cholinesterases is named the omega loop and forms the upper part of the active center gorge, essential for the high catalytic activity of these enzymes. In some non-catalytic members of the family, the loop may be necessary for heterologous partner recognition. Reshuffling of this protein portion occurred at the time of emergence of the fungi/metazoan lineage. Homologous proteins with this first disulfide bond are absent in plants but they are found in a limited number of bacterial genomes. In prokaryotes, the genes coding for such homologous proteins may have been acquired by horizontal transfer. However, the cysteines of the first disulfide bond are often lost in bacteria. Natural expression in bacteria of CO-esterases comprising this disulfide bond may have required compensatory mutations or expression of new chaperones. This disulfide bond may also challenge expression of the eukaryote-specific cholinesterases in prokaryotic cells. Yet recently, catalytically active human cholinesterase variants with enhanced thermostability were successfully expressed in E. coli. The key was the use of a peptidic sequence optimized through the Protein Repair One Stop Shop process, an automated structure- and sequence-based algorithm for expression of properly folded, soluble and stable eukaryotic proteins. Surprisingly however, crystal structures of the optimized cholinesterase variants expressed in bacteria revealed co-existing formed and unformed states of the first disulfide bond. Whether the bond never formed, or whether it properly formed then broke during the production/analysis process, cannot be inferred from the structural data. Yet, these features suggest that the recently acquired first disulfide bond is difficult to maintain in E. coli-expressed cholinesterases. To explore the fate of the first disulfide bond throughout the cholinesterase relatives, we reanalyzed the crystal structures of representative COesterases members from natural prokaryotic or eukaryotic sources or produced as recombinant proteins in E. coli. We found that in most cases this bond is absent.
ESTHER : Chatonnet_2019_Chem.Biol.Interact_13ChEPon_308_179
PubMedSearch : Chatonnet_2019_Chem.Biol.Interact_13ChEPon_308_179
PubMedID: 31100280

Title : Purification of recombinant human butyrylcholinesterase on Hupresin(R) - Lockridge_2018_J.Chromatogr.B.Analyt.Technol.Biomed.Life.Sci_1102-1103_109
Author(s) : Lockridge O , David E , Schopfer LM , Masson P , Brazzolotto X , Nachon F
Ref : Journal of Chromatography B Analyt Technol Biomed Life Sciences , 1102-1103 :109 , 2018
Abstract : Affinity chromatography on procainamide-Sepharose has been an important step in the purification of butyrylcholinesterase (BChE) and acetylcholinesterase (AChE) since its introduction in 1978. The procainamide affinity gel has limitations. In the present report a new affinity gel called Hupresin(R) was evaluated for its ability to purify truncated, recombinant human butyrylcholinesterase (rHuBChE) expressed in a stably transfected Chinese Hamster Ovary cell line. We present a detailed example of the purification of rHuBChE secreted into 3940mL of serum-free culture medium. The starting material contained 13,163units of BChE activity (20.9mg). rHuBChE was purified to homogeneity in a single step by passage over 82mL of Hupresin(R) eluted with 0.1M tetramethylammonium bromide in 20mM TrisCl pH7.5. The fraction with the highest specific activity of 630units/mg contained 11mg of BChE. Hupresin(R) is superior to procainamide-Sepharose for purification of BChE, but is not suitable for purifying native AChE because Hupresin(R) binds AChE so tightly that AChE is not released with buffers, but is desorbed with denaturing solvents such as 50% acetonitrile or 1% trifluoroacetic acid. Procainamide-Sepharose will continue to be useful for purification of AChE.
ESTHER : Lockridge_2018_J.Chromatogr.B.Analyt.Technol.Biomed.Life.Sci_1102-1103_109
PubMedSearch : Lockridge_2018_J.Chromatogr.B.Analyt.Technol.Biomed.Life.Sci_1102-1103_109
PubMedID: 30384187

Title : Multi-target-directed ligands for treating Alzheimer's disease: Butyrylcholinesterase inhibitors displaying antioxidant and neuroprotective activities - Knez_2018_Eur.J.Med.Chem_156_598
Author(s) : Knez D , Coquelle N , Pislar A , Zakelj S , Jukic M , Sova M , Mravljak J , Nachon F , Brazzolotto X , Kos J , Colletier JP , Gobec S
Ref : Eur Journal of Medicinal Chemistry , 156 :598 , 2018
Abstract : The limited clinical efficacy of current symptomatic treatment and minute effect on progression of Alzheimer's disease has shifted the research focus from single targets towards multi-target-directed ligands. Here, a potent selective inhibitor of human butyrylcholinesterase was used as the starting point to develop a new series of multifunctional ligands. A focused library of derivatives was designed and synthesised that showed both butyrylcholinesterase inhibition and good antioxidant activity as determined by the DPPH assay. The crystal structure of compound 11 in complex with butyrylcholinesterase revealed the molecular basis for its low nanomolar inhibition of butyrylcholinesterase (Ki=1.09+/-0.12nM). In addition, compounds 8 and 11 show metal-chelating properties, and reduce the redox activity of chelated Cu(2+) ions in a Cu-ascorbate redox system. Compounds 8 and 11 decrease intracellular levels of reactive oxygen species, and are not substrates of the active efflux transport system, as determined in Caco2 cells. Compound 11 also protects neuroblastoma SH-SY5Y cells from toxic Abeta1-42 species. These data indicate that compounds 8 and 11 are promising multifunctional lead ligands for treatment of Alzheimer's disease.
ESTHER : Knez_2018_Eur.J.Med.Chem_156_598
PubMedSearch : Knez_2018_Eur.J.Med.Chem_156_598
PubMedID: 30031971
Gene_locus related to this paper: human-BCHE

Title : The Magic of Crystal Structure-Based Inhibitor Optimization: Development of a Butyrylcholinesterase Inhibitor with Picomolar Affinity and in Vivo Activity - Kosak_2018_J.Med.Chem_61_119
Author(s) : Kosak U , Brus B , Knez D , Zakelj S , Trontelj J , Pislar A , Sink R , Jukic M , Zivin M , Podkowa A , Nachon F , Brazzolotto X , Stojan J , Kos J , Coquelle N , Salat K , Colletier JP , Gobec S
Ref : Journal of Medicinal Chemistry , 61 :119 , 2018
Abstract : The enzymatic activity of butyrylcholinesterase (BChE) in the brain increases with the progression of Alzheimer's disease, thus classifying BChE as a promising drug target in advanced Alzheimer's disease. We used structure-based drug discovery approaches to develop potent, selective, and reversible human BChE inhibitors. The most potent, compound 3, had a picomolar inhibition constant versus BChE due to strong cation-pi interactions, as revealed by the solved crystal structure of its complex with human BChE. Additionally, compound 3 inhibits BChE ex vivo and is noncytotoxic. In vitro pharmacokinetic experiments show that compound 3 is highly protein bound, highly permeable, and metabolically stable. Finally, compound 3 crosses the blood-brain barrier, and it improves memory, cognitive functions, and learning abilities of mice in a scopolamine model of dementia. Compound 3 is thus a promising advanced lead compound for the development of drugs for alleviating symptoms of cholinergic hypofunction in patients with advanced Alzheimer's disease.
ESTHER : Kosak_2018_J.Med.Chem_61_119
PubMedSearch : Kosak_2018_J.Med.Chem_61_119
PubMedID: 29227101
Gene_locus related to this paper: human-BCHE

Title : Comparison of the Binding of Reversible Inhibitors to Human Butyrylcholinesterase and Acetylcholinesterase: A Crystallographic, Kinetic and Calorimetric Study - Rosenberry_2017_Molecules_22_
Author(s) : Rosenberry TL , Brazzolotto X , Macdonald IR , Wandhammer M , Trovaslet-Leroy M , Darvesh S , Nachon F
Ref : Molecules , 22 : , 2017
Abstract : Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) hydrolyze the neurotransmitter acetylcholine and, thereby, function as coregulators of cholinergic neurotransmission. Although closely related, these enzymes display very different substrate specificities that only partially overlap. This disparity is largely due to differences in the number of aromatic residues lining the active site gorge, which leads to large differences in the shape of the gorge and potentially to distinct interactions with an individual ligand. Considerable structural information is available for the binding of a wide diversity of ligands to AChE. In contrast, structural data on the binding of reversible ligands to BChE are lacking. In a recent effort, an inhibitor competition approach was used to probe the overlap of ligand binding sites in BChE. Here, we extend this study by solving the crystal structures of human BChE in complex with five reversible ligands, namely, decamethonium, thioflavin T, propidium, huprine, and ethopropazine. We compare these structures to equivalent AChE complexes when available in the protein data bank and supplement this comparison with kinetic data and observations from isothermal titration calorimetry. This new information now allows us to define the binding mode of various ligand families and will be of importance in designing specific reversible ligands of BChE that behave as inhibitors or reactivators.
ESTHER : Rosenberry_2017_Molecules_22_
PubMedSearch : Rosenberry_2017_Molecules_22_
PubMedID: 29186056
Gene_locus related to this paper: human-BCHE

Title : N-Propargylpiperidines with naphthalene-2-carboxamide or naphthalene-2-sulfonamide moieties: Potential multifunctional anti-Alzheimer's agents - Kosak_2017_Bioorg.Med.Chem_25_633
Author(s) : Kosak U , Knez D , Coquelle N , Brus B , Pislar A , Nachon F , Brazzolotto X , Kos J , Colletier JP , Gobec S
Ref : Bioorganic & Medicinal Chemistry , 25 :633 , 2017
Abstract : In the brains of patients with Alzheimer's disease, the enzymatic activities of butyrylcholinesterase (BChE) and monoamine oxidase B (MAO-B) are increased. While BChE is a viable therapeutic target for alleviation of symptoms caused by cholinergic hypofunction, MAO-B is a potential therapeutic target for prevention of neurodegeneration in Alzheimer's disease. Starting with piperidine-based selective human (h)BChE inhibitors and propargylamine-based MAO inhibitors, we have designed, synthesized and biochemically evaluated a series of N-propargylpiperidines. All of these compounds inhibited hBChE with good selectivity over the related enzyme, acetylcholinesterase, and crossed the blood-brain barrier in a parallel artificial membrane permeation assay. The crystal structure of one of the inhibitors (compound 3) in complex with hBChE revealed its binding mode. Three compounds (4, 5, 6) showed concomitant inhibition of MAO-B. Additionally, the most potent hBChE inhibitor 7 and dual BChE and MAO-B inhibitor 6 were non-cytotoxic and protected neuronal SH-SY5Y cells from toxic amyloid beta-peptide species.
ESTHER : Kosak_2017_Bioorg.Med.Chem_25_633
PubMedSearch : Kosak_2017_Bioorg.Med.Chem_25_633
PubMedID: 27908752
Gene_locus related to this paper: human-BCHE

Title : Bacterial Expression of Human Butyrylcholinesterase as a Tool for Nerve Agent Bioscavengers Development - Brazzolotto_2017_Molecules_22_
Author(s) : Brazzolotto X , Igert A , Guillon V , Santoni G , Nachon F
Ref : Molecules , 22 : , 2017
Abstract : Human butyrylcholinesterase is a performant stoichiometric bioscavenger of organophosphorous nerve agents. It is either isolated from outdated plasma or functionally expressed in eukaryotic systems. Here, we report the production of active human butyrylcholinesterase in a prokaryotic system after optimization of the primary sequence through the Protein Repair One Stop Shop process, a structure- and sequence-based algorithm for soluble bacterial expression of difficult eukaryotic proteins. The mutant enzyme was purified to homogeneity. Its kinetic parameters with substrate are similar to the endogenous human butyrylcholinesterase or recombinants produced in eukaryotic systems. The isolated protein was prone to crystallize and its 2.5-A X-ray structure revealed an active site gorge region identical to that of previously solved structures. The advantages of this alternate expression system, particularly for the generation of butyrylcholinesterase variants with nerve agent hydrolysis activity, are discussed.
ESTHER : Brazzolotto_2017_Molecules_22_
PubMedSearch : Brazzolotto_2017_Molecules_22_
PubMedID: 29077024
Gene_locus related to this paper: human-BCHE

Title : Development of an in-vivo active reversible butyrylcholinesterase inhibitor - Kosak_2016_Sci.Rep_6_39495
Author(s) : Kosak U , Brus B , Knez D , Sink R , Zakelj S , Trontelj J , Pislar A , Slenc J , Gobec M , Zivin M , Tratnjek L , Perse M , Salat K , Podkowa A , Filipek B , Nachon F , Brazzolotto X , Wieckowska A , Malawska B , Stojan J , Rascan IM , Kos J , Coquelle N , Colletier JP , Gobec S
Ref : Sci Rep , 6 :39495 , 2016
Abstract : Alzheimer's disease (AD) is characterized by severe basal forebrain cholinergic deficit, which results in progressive and chronic deterioration of memory and cognitive functions. Similar to acetylcholinesterase, butyrylcholinesterase (BChE) contributes to the termination of cholinergic neurotransmission. Its enzymatic activity increases with the disease progression, thus classifying BChE as a viable therapeutic target in advanced AD. Potent, selective and reversible human BChE inhibitors were developed. The solved crystal structure of human BChE in complex with the most potent inhibitor reveals its binding mode and provides the molecular basis of its low nanomolar potency. Additionally, this compound is noncytotoxic and has neuroprotective properties. Furthermore, this inhibitor moderately crosses the blood-brain barrier and improves memory, cognitive functions and learning abilities of mice in a model of the cholinergic deficit that characterizes AD, without producing acute cholinergic adverse effects. Our study provides an advanced lead compound for developing drugs for alleviating symptoms caused by cholinergic hypofunction in advanced AD.
ESTHER : Kosak_2016_Sci.Rep_6_39495
PubMedSearch : Kosak_2016_Sci.Rep_6_39495
PubMedID: 28000737
Gene_locus related to this paper: human-BCHE

Title : Discovery and Structure-Activity Relationships of a Highly Selective Butyrylcholinesterase Inhibitor by Structure-Based Virtual Screening - Dighe_2016_J.Med.Chem_59_7683
Author(s) : Dighe SN , Deora GS , De la Mora E , Nachon F , Chan S , Parat MO , Brazzolotto X , Ross BP
Ref : Journal of Medicinal Chemistry , 59 :7683 , 2016
Abstract : Structure-based virtual screening of two libraries containing 567981 molecules was used to discover novel, selective BuChE inhibitors, which are potentially superior symptomatic treatments in late-stage Alzheimer's disease. Compound 16 was identified as a highly selective submicromolar inhibitor of BuChE (huBuChE IC50 = 0.443 muM) with high permeability in the PAMPA-BBB model. The X-ray crystal structure of huBuChE in complex with 16 revealed the atomic-level interactions and offers opportunities for further development of the series.
ESTHER : Dighe_2016_J.Med.Chem_59_7683
PubMedSearch : Dighe_2016_J.Med.Chem_59_7683
PubMedID: 27405689
Gene_locus related to this paper: human-BCHE

Title : Structure-based development of nitroxoline derivatives as potential multifunctional anti-Alzheimer agents - Knez_2015_Bioorg.Med.Chem_23_4442
Author(s) : Knez D , Brus B , Coquelle N , Sosic I , Sink R , Brazzolotto X , Mravljak J , Colletier JP , Gobec S
Ref : Bioorganic & Medicinal Chemistry , 23 :4442 , 2015
Abstract : Tremendous efforts have been dedicated to the development of effective therapeutics against Alzheimer's disease, which represents the most common debilitating neurodegenerative disease. Multifunctional agents are molecules designed to have simultaneous effects on different pathological processes. Such compounds represent an emerging strategy for the development of effective treatments against Alzheimer's disease. Here, we report on the synthesis and biological evaluation of a series of nitroxoline-based analogs that were designed by merging the scaffold of 8-hydroxyquinoline with that of a known selective butyrylcholinesterase inhibitor that has promising anti-Alzheimer properties. Most strikingly, compound 8g inhibits self-induced aggregation of the amyloid beta peptide (Abeta1-42), inhibits with sub-micromolar potency butyrylcholinesterase (IC50=215 nM), and also selectively complexes Cu(2+). Our study thus designates this compound as a promising multifunctional agent for therapeutic treatment of Alzheimer's disease. The crystal structure of human butyrylcholinesterase in complex with compound 8g is also solved, which suggests ways to further optimize compounds featuring the 8-hydroxyquinoline scaffold.
ESTHER : Knez_2015_Bioorg.Med.Chem_23_4442
PubMedSearch : Knez_2015_Bioorg.Med.Chem_23_4442
PubMedID: 26116179
Gene_locus related to this paper: human-BCHE

Title : Progress in the development of enzyme-based nerve agent bioscavengers - Nachon_2013_Chem.Biol.Interact_206_536
Author(s) : Nachon F , Brazzolotto X , Trovaslet M , Masson P
Ref : Chemico-Biological Interactions , 206 :536 , 2013
Abstract : Acetylcholinesterase is the physiological target for acute toxicity of nerve agents. Attempts to protect acetylcholinesterase from phosphylation by nerve agents, is currently achieved by reversible inhibitors that transiently mask the enzyme active site. This approach either protects only peripheral acetylcholinesterase or may cause side effects. Thus, an alternative strategy consists in scavenging nerve agents in the bloodstream before they can reach acetylcholinesterase. Pre- or post-exposure administration of bioscavengers, enzymes that neutralize and detoxify organophosphorus molecules, is one of the major developments of new medical counter-measures. These enzymes act either as stoichiometric or catalytic bioscavengers. Human butyrylcholinesterase is the leading stoichiometric bioscavenger. Current efforts are devoted to its mass production with care to pharmacokinetic properties of the final product for extended lifetime. Development of specific reactivators of phosphylated butyrylcholinesterase, or variants with spontaneous reactivation activity is also envisioned for rapid in situ regeneration of the scavenger. Human paraoxonase 1 is the leading catalytic bioscavenger under development. Research efforts focus on improving its catalytic efficiency toward the most toxic isomers of nerve agents, by means of directed evolution-based strategies. Human prolidase appears to be another promising human enzyme. Other non-human efficient enzymes like bacterial phosphotriesterases or squid diisopropylfluorophosphatase are also considered though their intrinsic immunogenic properties remain challenging for use in humans. Encapsulation, PEGylation and other modifications are possible solutions to address this problem as well as that of their limited lifetime. Finally, gene therapy for in situ generation and delivery of bioscavengers is for the far future, but its proof of concept has been established.
ESTHER : Nachon_2013_Chem.Biol.Interact_206_536
PubMedSearch : Nachon_2013_Chem.Biol.Interact_206_536
PubMedID: 23811386

Title : Human butyrylcholinesterase produced in insect cells: huprine-based affinity purification and crystal structure - Brazzolotto_2012_Febs.J_279_2905
Author(s) : Brazzolotto X , Wandhammer M , Ronco C , Trovaslet M , Jean L , Lockridge O , Renard PY , Nachon F
Ref : Febs J , 279 :2905 , 2012
Abstract : Butyrylcholinesterase is a serine hydrolase present in all mammalian tissues. It can accommodate larger substrates or inhibitors than acetylcholinesterase, the enzyme responsible for hydrolysis of the neurotransmitter acetylcholine in the central nervous system and neuromuscular junctions. AChE is the specific target of organophosphorous pesticides and warfare nerve agents, while BChE is their stoichiometric bioscavenger. Conversion of BChE into a catalytic bioscavenger by rational design or designing reactivators specific to BChE required structural data obtained with a recombinant low-glycosylated human BChE expressed in Chinese hamster ovary cells. This expression system yields about 1 mg of pure enzyme per liter of cell culture. Here, we report an improved expression system with 4-fold higher yield for truncated human BChE with all glycosylation sites present using insect cells. We developed a fast purification protocol of the recombinant protein using a huprine-based affinity chromatography superior to the classical procainamide-based affinity. The purified BChE crystallized in different conditions and space group than those for the recombinant low-glycosylated protein produced in Chinese hamster ovary cells. The crystals diffracted to 2.5 A. The overall monomer structure is similar to the low-glycosylated structure but for the presence of the additional glycans. Remarkably, the carboxylic acid molecule systematically bound to the catalytic serine in the low-glycosylated structure is also present in this new structure, despite the different expression system, purification protocol and crystallization conditions.
ESTHER : Brazzolotto_2012_Febs.J_279_2905
PubMedSearch : Brazzolotto_2012_Febs.J_279_2905
PubMedID: 22726956
Gene_locus related to this paper: human-BCHE

Title : Organophosphate hydrolases as catalytic bioscavengers of organophosphorus nerve agents - Trovaslet-Leroy_2011_Toxicol.Lett_206_14
Author(s) : Trovaslet-Leroy M , Musilova L , Renault F , Brazzolotto X , Misik J , Novotny L , Froment MT , Gillon E , Loiodice M , Verdier L , Masson P , Rochu D , Jun D , Nachon F
Ref : Toxicol Lett , 206 :14 , 2011
Abstract : Bioscavengers are molecules able to neutralize neurotoxic organophosphorus compounds (OP) before they can reach their biological target. Human butyrylcholinesterase (hBChE) is a natural bioscavenger each molecule of enzyme neutralizing one molecule of OP. The amount of natural enzyme is insufficient to achieve good protection. Thus, different strategies have been envisioned. The most straightforward consists in injecting a large dose of highly purified natural hBChE to increase the amount of bioscavenger in the bloodstream. This proved to be successful for protection against lethal doses of soman and VX but remains expensive. An improved strategy is to regenerate prophylactic cholinesterases (ChE) by administration of reactivators after exposure. But broad-spectrum efficient reactivators are still lacking, especially for inhibited hBChE. Cholinesterase mutants capable of reactivating spontaneously are another option. The G117H hBChE mutant has been a prototype. We present here the Y124H/Y72D mutant of human acetylcholinesterase; its spontaneous reactivation rate after V-agent inhibition is increased up to 110 fold. Catalytic bioscavengers, enzymes capable of hydrolyzing OP, present the best alternative. Mesophilic bacterial phosphotriesterase (PTE) is a candidate with good catalytic efficiency. Its enantioselectivity has been enhanced against the most potent OP isomers by rational design. We show that PEGylation of this enzyme improves its mean residence time in the rat blood stream 24-fold and its bioavailability 120-fold. Immunogenic issues remain to be solved. Human paraoxonase 1 (hPON1) is another promising candidate. However, its main drawback is that its phosphotriesterase activity is highly dependent on its environment. Recent progress has been made using a mammalian chimera of PON1, but we provide here additional data showing that this chimera is biochemically different from hPON1. Besides, the chimera is expected to suffer from immunogenic issues. Thus, we stress that interest for hPON1 must not fade away, and in particular, the 3D structure of the hPON1 eventually in complex with OP has to be solved.
ESTHER : Trovaslet-Leroy_2011_Toxicol.Lett_206_14
PubMedSearch : Trovaslet-Leroy_2011_Toxicol.Lett_206_14
PubMedID: 21683774