Neurotoxicity fingerprinting of venoms using on-line microfluidic AChBP profiling

Slagboom, Julien; Otvos, Reka A.; Cardoso, Fernanda C.; Iyer, Janaki Krishnamoorthy; Visser, Jeroen C.; Doodewaerd, Bjorn R. van; McCleary, Ryan J.R.; Niessen, W.M.A.; Somsen, Govert W.; Lewis, Richard J.; Kini, R. Manjunatha; Smit, August B.; Casewell, Nicholas R.; Kool, Jeroen

doi:10.1016/j.toxicon.2018.04.022

Cited by 20 publications

(22 citation statements)

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“…The methodology applied here facilitated the rapid identification and fractionation of coagulopathic toxins, which resulted in the identification of anticoagulant venom activities previously being masked by net procoagulant effects of the crude venom, and also detected a number of anticoagulant PLA 2 s not previously known to exhibit anticoagulant activities. Success here, alongside the recent development of other small-scale medium-throughput bioassays focused on characterizing other relevant toxin activities (20, 83–86) means that such assays could be used interchangeably for understanding venom function and pathology – ultimately providing a ‘bioassay toolkit’ for deconvoluting venom mixtures. These approaches will undoubtedly facilitate new research focused on improving snakebite therapy.…”

Section: Resultsmentioning

confidence: 99%

High throughput screening and identification of coagulopathic snake venom proteins and peptides using nanofractionation and proteomics approaches

Slagboom

Mladic

Xie³

et al. 2019

Preprint

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17Snakebite is a neglected tropical disease that results in a variety of systemic and local pathologies in 18 envenomed victims and is responsible for around 138,000 deaths every year. Many snake venoms cause 19 severe coagulopathy that makes victims vulnerable to suffering life-threating haemorrhage. The 20 mechanisms of action of coagulopathic snake venom toxins are diverse and can result in both anticoagulant 21 and procoagulant effects. However, because snake venoms consist of a mixture of numerous protein and 22 peptide components, high throughput characterizations of specific target bioactives is challenging. In this 23 study, we applied a combination of analytical and pharmacological methods to identify snake venom toxins 24 from a wide diversity of snake species that perturb coagulation. To do so, we used a high-throughput 25 screening approach consisting of a miniaturised plasma coagulation assay in combination with a venom 26 nanofractionation approach. Twenty snake venoms were first separated using reversed-phase liquid 27 chromatography, and a post-column split allowed a small fraction to be analyzed with mass spectrometry, 28 while the larger fraction was collected and dispensed onto 384-well plates before direct analysis using a 29 plasma coagulation assay. Our results demonstrate that many snake venoms simultaneously contain both 30 procoagulant and anticoagulant bioactives that contribute to coagulopathy. In-depth identification analysis 31 from seven medically-important venoms, via mass spectrometry and nanoLC-MS/MS, revealed that 32 phospholipase A 2 toxins are frequently identified in anticoagulant venom fractions, while serine protease 33 and metalloproteinase toxins are often associated with procoagulant bioactivities. The nanofractionation 34 and proteomics approach applied herein seems likely to be a valuable tool for the rational development of 35 next-generation snakebite treatments by facilitating the rapid identification and fractionation of 36 coagulopathic toxins, thereby enabling specific targeting of these toxins by new therapeutics such as 37 monoclonal antibodies and small molecule inhibitors. Classified Personnel Information 39Author summary 40 Snakebite is a neglected tropical disease that results in more than 100,000 deaths every year. 41Haemotoxicity is one of the most common signs of systemic envenoming observed after snakebite, and 42 many snake venoms cause severe impairment of the blood coagulation that makes victims vulnerable to 43 suffering life-threating hemorrhage. In this study, we applied a combination of analytical and 44 pharmacological methods to identify snake venom toxins from a wide diversity of snake species that 45 interfere with blood coagulation. Twenty snake venoms were screened for their effects on the blood 46 coagulation cascade and based on the initial results and the medical relevance of the species, seven 47 venoms were selected for in-depth analysis of the responsible toxins using advanced identification 48 techniques. Our findings reveal a number ...

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Section: Resultsmentioning

confidence: 99%

High throughput screening and identification of coagulopathic snake venom proteins and peptides using nanofractionation and proteomics approaches

Slagboom

Mladic

Xie³

et al. 2019

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

“…The methodology applied here facilitated the rapid identification and fractionation of coagulopathic toxins, which resulted in the identification of anticoagulant venom activities previously being masked by net procoagulant effects of the crude venom, and also detected a number of anticoagulant PLA 2 s not previously known to exhibit anticoagulant activities. Success here, alongside the recent development of other small-scale mediumthroughput bioassays focused on characterizing other relevant toxin activities [21,[84][85][86][87] means that such assays could be used interchangeably for understanding venom function and pathology-ultimately providing a 'bioassay toolkit' for deconvoluting venom mixtures. These approaches will undoubtedly facilitate new research focused on improving snakebite therapy.…”

Section: Plos Neglected Tropical Diseasesmentioning

confidence: 99%

High throughput screening and identification of coagulopathic snake venom proteins and peptides using nanofractionation and proteomics approaches

et al. 2020

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Snakebite is a neglected tropical disease that results in a variety of systemic and local pathologies in envenomed victims and is responsible for around 138,000 deaths every year. Many snake venoms cause severe coagulopathy that makes victims vulnerable to suffering life-threating haemorrhage. The mechanisms of action of coagulopathic snake venom toxins are diverse and can result in both anticoagulant and procoagulant effects. However, because snake venoms consist of a mixture of numerous protein and peptide components, high throughput characterizations of specific target bioactives is challenging. In this study, we applied a combination of analytical and pharmacological methods to identify snake venom toxins from a wide diversity of snake species that perturb coagulation. To do so, we used a high-throughput screening approach consisting of a miniaturised plasma coagulation assay in combination with a venom nanofractionation approach. Twenty snake venoms were first separated using reversed-phase liquid chromatography, and a post-column split allowed a small fraction to be analyzed with mass spectrometry, while the larger fraction was collected and dispensed onto 384-well plates. After fraction collection, any solvent present in the wells was removed by means of freeze-drying, after which it was possible to perform a plasma coagulation assay in order to detect coagulopathic activity. Our results demonstrate that many snake venoms simultaneously contain both procoagulant and anticoagulant bioactives that contribute to coagulopathy. In-depth identification analysis from seven medically-important venoms, via mass spectrometry and nanoLC-MS/MS, revealed that phospholipase A 2 toxins are frequently identified in anticoagulant venom fractions, while serine protease and metalloproteinase toxins are often associated with procoagulant bioactivities. The nanofractionation and proteomics approach applied herein seems likely to be a valuable tool for the rational development of next-generation snakebite treatments by facilitating the rapid identification and fractionation of coagulopathic toxins, thereby enabling PLOS NEGLECTED TROPICAL DISEASES

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“…Analytical at-line nanofractionation techniques, which couple high-throughput functional screening with compound identification, have proven to be useful tools for identifying bioactives from complex mixtures [27,28]. Improvements in NMR and ambient mass spectrometry provide the potential for the detection and identification of toxins in foodstuff (see Food and Water Biosafety/ Security Sectors).…”

Section: Glossarymentioning

confidence: 99%

Friends or Foes? Emerging Impacts of Biological Toxins

Clark

Casewell

Elliott

et al. 2019

Trends in Biochemical Sciences

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Toxins are substances produced from biological sources (e.g., animal, plants, microorganisms) that have deleterious effects on a living organism. Despite the obvious health concerns of being exposed to toxins, they are having substantial positive impacts in a number of industrial sectors. Several toxin-derived products are approved for clinical, veterinary, or agrochemical uses. This review sets out the case for toxins as 'friends' that are providing the basis of novel medicines, insecticides, and even nucleic acid sequencing technologies. We also discuss emerging toxins ('foes') that are becoming increasingly prevalent in a range of contexts through climate change and the globalisation of food supply chains and that ultimately pose a risk to health. Toxins and Toxinology In the natural world, toxins are employed for diverse purposes, from the prey-incapacitating molecules found in snake, spider, and cone snail venom, to the defensive compounds harboured by numerous poisonous plant and animal species. While the consequences of human intoxication can be severe (Box 1), the functional diversity of biological toxins has led to their frequent use as experimental tools for studying physiological and pharmacological mechanisms (e.g., synaptic transmission, ion channel subtypes) (Figure 1). Toxinology involves the identification, characterisation, production, and engineering of biological toxins along with their application or repurposing as research tools and clinical products. As such, toxinology encompasses the study of the evolution, chemistry, biology, and clinical effects of toxins and includes potential biotechnological and/or therapeutic applications. Experimental applications of toxins as tools for physiologists go back a long time, including Claude Bernard's experiments in the 1800s with curare to demonstrate the existence of chemical signalling between nerves and muscles [2], and Henry Dale's use of muscarine and nicotine to show different subtypes of receptors for acetylcholine [3]. Snake toxins were critical for the first isolation of a receptor for a neurotransmitter [a-bungarotoxin and nicotinic acetylcholine receptors (nAChRs)] [4]. Recently, peptide toxins have been very useful in teasing out the functional importance of different subtypes of ion channels that are critical for neuronal function and cellular signalling [5-7]. An increasing number of toxins are now transitioning from the laboratory to the clinic and the balance of research in this aspect of toxinology is shifting from the classical development of anti-toxins and anti-venoms towards drug discovery [8,9]. The attraction of toxins for drug discovery lies in their often unique selectivity of their biological effects coupled with high potency. Toxic plant alkaloids were the first source of toxin-derived therapeutics, notably

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Neurotoxicity fingerprinting of venoms using on-line microfluidic AChBP profiling

Cited by 20 publications

References 37 publications

High throughput screening and identification of coagulopathic snake venom proteins and peptides using nanofractionation and proteomics approaches

High throughput screening and identification of coagulopathic snake venom proteins and peptides using nanofractionation and proteomics approaches

High throughput screening and identification of coagulopathic snake venom proteins and peptides using nanofractionation and proteomics approaches

Friends or Foes? Emerging Impacts of Biological Toxins

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