Toxic epibatidine was structurally modified to image Alzheimer´s disease
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| License | CC Attribution - NoDerivatives 4.0 International: You are free to use, copy, distribute and transmit the work or content in unchanged form for any legal purpose as long as the work is attributed to the author in the manner specified by the author or licensor. | |
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00:00
GesundheitsstörungEpibatidinPoisonWinnowingWine tasting descriptorsComputer animation
00:19
GesundheitsstörungToxinForkhead-GenMeeting/Interview
00:30
GesundheitsstörungPosttranslational modificationProtein subunitNicotinischer AcetylcholinrezeptorToxinPharmacologyMoleculeChemistBinding energyStop codonToxicityMeeting/Interview
01:11
Posttranslational modificationChemistMeeting/Interview
01:19
Wine tasting descriptorsToxinChemical structurePainRadioactive tracerChemical experiment
01:31
ToxinPyridineChemical compoundMolecular geometry
01:41
Organische ChemieCarbon (fiber)Synthetic oilChemical experiment
01:49
Carbon (fiber)Systemic therapyStereoselektive SyntheseChemical compoundChemical experimentMolecular geometry
02:08
ChlorineThermoformingPyridineNomifensineChemical compoundMolecular geometry
02:18
Nicotinischer AcetylcholinrezeptorSunscreenMoleculeFluorineChemical compoundHope, ArkansasGrease interceptorMeeting/Interview
02:39
RadionuclideChemical compoundMolecular geometry
02:48
Combustion chamberHydrogenElectronProtonationCobaltoxideWine tasting descriptorsWaterNickel(II) oxideNiobiumChemical experiment
03:22
WaterCobaltoxideWine tasting descriptorsCombustion chamberProtonationKernproteineTransportChemical experimentMeeting/Interview
03:33
ProtonationEmission spectrumKernproteineCombustion chamberMeeting/Interview
03:44
Combustion chamberWaterBiosynthesisStarvation responseChemical reactionMetabolic pathwayHeiße ZelleChemical experiment
03:59
BiosynthesisModul <Membranverfahren>Chemical reactionStarvation responseContainment buildingChemical reactorSystemic therapyWater purificationChemical experiment
04:09
BimetalChemical reactorAmmoniumSystemic therapyWursthülleWalkingChemical reactionWaterCombine harvesterNucleophilic substitutionPrecursor (chemistry)Functional groupDiseaseWater purificationTopicityMetalChemical experiment
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Chemical reactionDiseaseWater purificationWalkingScreening (medicine)Modul <Membranverfahren>Process (computing)Chemical experiment
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WalkingProcess (computing)Electronic cigaretteRadioactive tracerThermoformingHigh-performance liquid chromatographySaline (medicine)Chemical experiment
05:29
Emission spectrumTodesbescheinigungMoleculeOctane ratingNicotinischer AcetylcholinrezeptorChemical experimentEngineering drawing
05:44
MetabolismRadioactive tracerChemical experiment
05:52
ChromatographyThermoformingAmyloid (mycology)ProteinIceWine tasting descriptorsPhosphorous acidRadioactive tracerThin filmMeeting/Interview
06:07
Phosphorous acidThermoformingAmyloid (mycology)Beta sheetChemical experiment
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ProteinPyroglutamic acidThermoformingInflammationTiermodellDeposition (phase transition)Chemical experiment
06:31
TiermodellInflammationGesundheitsstörungCork taintRedoxChemical experiment
06:49
Chemical experiment
06:59
Phosphorous acidColourantPainProcess (computing)Cell growthChemical experimentComputer animation
07:20
Nicotinischer AcetylcholinrezeptorSetzen <Verfahrenstechnik>Reducing agentComputer animation
07:39
Octane ratingReducing agentNicotinischer AcetylcholinrezeptorComputer animation
07:52
Computer animation
08:04
BiomarkerReaction mechanismPharmaceutical drugChemical experiment
08:22
Emission spectrumIceCombine harvesterMeeting/Interview
08:31
MagnetismChemische VerschiebungTool steelChemical experiment
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Tool steelWine tasting descriptorsPitting corrosionHematiteLeadComputer animation
08:56
Wine tasting descriptorsChemical experiment
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IsotopenmarkierungMoleculeWine tasting descriptorsEnzymkinetikKlinisches ExperimentImpfung <Chemie>Setzen <Verfahrenstechnik>Chemical experiment
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Klinisches ExperimentSetzen <Verfahrenstechnik>Diagram
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DiseaseGesundheitsstörungPharmaceutical drug
Transcript: English(auto-generated)
00:11
May I introduce epipatiditis Antoni ETU? It is a poison arrow frog, which lives in the rainforest of South America. And we want to show you how the toxin from the skin of this frog
00:24
helped us to develop an imaging agent for Alzheimer's disease. In 1992, the toxin was isolated, identified, and later called, according to the frog's name, epipatidine. Pharmacological investigations revealed that the toxin binds
00:43
the various subtypes of nicotinic acetylcholine receptors. Those containing an alpha-1 subunit are mainly important for the toxic action. And those containing an alpha-4 or an alpha-7 subunit are important for Alzheimer's disease.
01:01
To make use of this valuable feature, structural modification of the molecule was needed to avoid its toxic action. Matthias Schäunemann, who is a chemist in our team, will explain us how this modification was achieved.
01:21
We are here in the organic synthetic lab. I would like to show you the small changes based on the epipatidine structure which have been made for our radiotracer development. This structural model shows the toxin as an atzabitsicloroheptane ring, to which a chlorine-substituted pyridine is attached.
01:42
Thereby, an asymmetric carbon and hence a chiral center is created, which is always a synthetic challenge for organic chemists. We accomplished insertion of an additional carbon into the atzabitsiclorine system.
02:01
By utilizing a highly axial-selective and asymmetric keto-DLR direction, the chlorine in this position of the pyridine was replaced by the fluorine, for later radiolabeling, with fluorine-18. Thereby, we got two enantiomeric forms of our derivative called flubatine.
02:23
The compound is now highly selective for the alpha-4 beta-2 nicotinic receptor subtype. And about a factor 100 less toxic than epipatidine itself. The fluorine in the molecule can be replaced by the fluorine-18 isotope. In order to do so, we need to produce this positron-emitting radioisotope
02:45
with the help of our cyclotron. This cyclotron accelerates about 200 trillion of negatively charged hydrogen ions per second in a horizontal plane on a spiral trajectory. After 300 turns, the accelerated ions reach their maximum velocity
03:04
of almost a fifth of the speed of light. Then, the two electrons of the hydrogen ions are removed and the so-formed protons change their trajectory, leave the cyclotron and crash into the oxygen-enriched water target.
03:21
The target water is encased in this small, water-cooled niobium chamber. It will be filled with about 3 milliliters of the oxygen-18-enriched water. And with this isotope, nuclear reaction occurs in the chamber by insertion of a proton into the nucleus and emission of a neutron producing fluorine-18.
03:45
A remote system empties the target chamber and transports the enriched water containing the produced fluorine-18 ions along this line. We follow now the path into lead-shielded hot cells for further handling.
04:00
So in these hot cells, we have special synthesis modules which allow us to perform a radiochemical reaction in a remote mode. You can see here the containers for the reagents, two reactors where the labeling takes place and a small HPC system for the purification.
04:21
However, the first step in a radiochemical reaction is to get rid of the enriched water by using a combination of a special cartridge and azeotropic trine. This step is necessary because the fluorine-18 ions need to be completely free of water to allow them to react with a precursor with a good leaving group
04:44
such as trimetal ammonium in case of our flubertine permits nucleophilic substitution. The supply of the reagents, the reaction conditions and the different purification steps are all controlled by this computer here. It shows us the complete reaction process on the screen as it happens in the module
05:04
and allows simultaneously a handling of the different steps. At the end of the process, the radio tracer has to be converted into an injectable form by using a solvent which is well tolerated by animals and humans such as isotonic saline.
05:23
The quality of the radio tracer has to be checked by various analytical procedures such as high-performance liquid chromatography. Now that we got our radio labeled flubertine molecule, we make use of its positron emission to characterize its biological features
05:43
such as binding to the various nicotinic receptor subtypes, biodistribution and metabolism. We call it now a radio tracer. Here we have a phosphor imager which allows us to obtain two-dimensional images of the distribution of our radio tracer in the whole body,
06:02
brain slices or in thin layer chromatographs respectively. We have used a phosphor imager that we have just seen to perform a beta-auto radio fee on mouse brain slices. These are transgenic mice expressing a truncated and mutant form of human amyloid beta.
06:21
And in the mouse brain, this amyloid protein is enzymatically transformed and the resulting pyroglutamate A-beta aggregates very rapidly and forms neurotoxic deposits as you can see here. Neuronal loss and inflammatory processes are the main reason for the development of the behavioral phenotype of these transgenic animals.
06:44
And altogether this animal model represents the key characteristics of human Alzheimer's disease and was therefore chosen to perform our future imaging studies. The brain slices for those studies were obtained with such a cryostat microtome. And now I'm going to show you the result of our autoradiographic studies.
07:04
What we see here is an autoradiographic image of a brain slice of a mouse brain that we obtained from a wild-type mouse which was incubated with the radiolabeled flubertin and afterwards processed with the phosphor imager.
07:20
And here you can see the autoradiograph of a brain of a transgenic animal. And by directly comparing the color-coded autoradiographs of both animals, we can clearly see a decrease in the binding of the radiolabeled flubertin in the transgenic animal in comparison to the wild-type animal.
07:44
And this indicates a reduction in the alpha-4 beta-2 subtype of nicotinic receptors in the transgenic animals. And in particular the decrease in the cortical and hippocampal regions corresponds to the observed neuropathological phenotype of the transgenic animals.
08:05
Now that we have evidence that flubertin might be a biomarker for Alzheimer's disease, we need to prove that the mechanism which works on brain slices also works in vivo. This is our medicinal physicist Matthias Krantz who will tell us how this was achieved.
08:22
Here we have a small animal PET MRI. This device combines a one-tesla MRI based on a permanent magnet with a positron emission tomograph. The MRI has a resolution of about 100 micrometers and a PET of about one millimeter. This unique device has the advantage of zero magnetic fringe fields due to the internal shielding.
08:43
As you can see over here, one can easily work with paramagnetic tools like this narcosis device in the close proximity to the scanner without any further precautions. On this PET image one can easily delineate the brain, the liver, the kidneys and the urinary bladder.
09:03
PET provides images of the radiodrazor distribution in different parts of the body. An overlay using the anatomical information from the MRI, as you can see over here, allows us to investigate and to quantify the amount of radiolabeled molecules in different parts of the body.
09:21
For example, this diagram over here shows the kinetics of fluvodine uptake in the mouse brain. Only if such type of animal experiment shows positive results, permission for clinical studies may be granted. Our new radiodrazor is now used by our colleagues from the Department of Nuclear Medicine
09:40
and Psychiatry of the Leipzig University to better understand how Alzheimer's disease develops and how it can be distinguished from similar diseases.