, UK UK , Aurelia Bioscience Ltd, BioCity Nottingham Nottingham BioCity Ltd, Bioscience Aurelia
Email: [email protected] Tel : Web: www.aureliabio.com www.aureliabio.com Web: : Tel [email protected] Email: +44 (0) 115 8370503 8370503 115 (0) +44
*Please turn over for examples of available kinases kinases available of examples for over turn *Please
• This technique is applicable to a wide range of kinases kinases of range wide a to applicable is technique This •
• The approach can be used to look at allosteric inhibitors in addition to ATP competitive inhibitors inhibitors competitive ATP to addition in inhibitors allosteric at look to used be can approach The •
• Examines the K-on (association rate) and K-off (dissociation rate) of compounds in live cells cells live in compounds of rate) (dissociation off - K and rate) (association on - K the Examines •
pseudo biochemical binding assay assay binding biochemical pseudo
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full-length length - full • Expression of kinase target in cells cells in target kinase of Expression •
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assay assay
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Advantages Advantages : :
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• This quantitative analysis can be performed in cultured cells cells cultured in performed be can analysis quantitative This •
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Bioscience has implemented target engagement engagement target implemented has Bioscience
Using Promega’s NanoBRET™ technology, Aurelia Aurelia technology, ™ NanoBRET Promega’s Using •
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K and rate) (association on - K the Examines • • • TheExaminesapproachthecanK-beon used(associationto look atrate)allostericand K-inhibitorsoff (dissociationin additionrate)toofATPcompoundscompetitivein liveinhibitorscells kinases of range wide a to applicable is technique This • cells cells in in target target kinase kinase length length - - full full of of Expression Expression • • • TheTheapproachapproachcancanbebeusedusedtotolooklookatatallostericallostericinhibitorsinhibitorsininadditionadditionto• toATPATPcompetitivecompetitiveinhibitorsinhibitors inhibitors competitive ATP to addition in inhibitors allosteric at look to used be can approach The • • • ThisThetechniqueapproachiscanapplicablebe usedtotoa lookwideatrangeallostericof kinasesinhibitors in addition to ATP competitive inhibitors a a as as performed performed be be can can assay assay the the and and cell cell the the from from content content cellular cellular release release to to digitonin) digitonin) (with (with disrupted disrupted be be can can Cells Cells • • • This•ThisThistechniquetechniquetechniqueis isapplicableisapplicableapplicabletotoatowideaawidewiderangerangerangeofofkinasesofkinaseskinases • kinases of range wide a to applicable is technique This • assay assay binding binding biochemical biochemical pseudo pseudo cells cells live live in in compounds compounds of of rate) rate) (dissociation (dissociation off off - - K K and and rate) rate) (association (association on on - - K K the the Examines Examines • • inhibitors inhibitors competitive competitive ATP ATP to to addition addition in in inhibitors inhibitors allosteric allosteric at at look look to to used used be be can can approach approach The The • • kinases kinases of of range range wide wide a a to to applicable applicable is is technique technique This This • *Please turn over for examples of available kinases • *Please*Please*Please turn turn turn over over over for for examplesfor examples examples of of availableof available available kinases kinases kinases kinases available of examples for over turn *Please kinases available of examples for over turn *Please Email: [email protected] Tel : +44 (0) 115 8370503 Web: www.aureliabio.com Email:Email:Email: [email protected] [email protected] [email protected] TelTelTel : +44 : :+44 +44 (0) (0) (0) 115 115 115 8370503 8370503 8370503 Web:Web:Web: www.aureliabio.com www.aureliabio.com www.aureliabio.com Web: 8370503 115 (0) +44 : Tel [email protected] Email: Web: www.aureliabio.com Web: 8370503 115 (0) +44 : Tel [email protected] Email: Aurelia Bioscience Ltd, BioCity Nottingham , UK *Please turn over for examples of available kinases kinases available available of of examples examples for for over over turn turn *Please *Please AureliaAureliaAurelia Bioscience Bioscience Bioscience Ltd, Ltd, Ltd, BioCity BioCity BioCity Nottingham Nottingham Nottingham, UK, UK, UK UK , Nottingham BioCity Ltd, Bioscience Aurelia , UK , Nottingham BioCity Ltd, Bioscience Aurelia Web: www.aureliabio.com www.aureliabio.com Web: Web: 8370503 8370503 115 115 (0) (0) +44 +44 : : Tel Tel [email protected] [email protected] Email: Email:
, UK UK , , Nottingham Nottingham BioCity BioCity Ltd, Ltd, Bioscience Bioscience Aurelia Aurelia
, UK UK , Aurelia Bioscience Ltd, BioCity Nottingham Nottingham BioCity Ltd, Bioscience Aurelia
Email: [email protected] Tel : Web: www.aureliabio.com www.aureliabio.com Web: : Tel [email protected] Email: +44 (0) 115 8370503 8370503 115 (0) +44
*Please turn over for examples of available kinases kinases available of examples for over turn *Please
• This technique is applicable to a wide range of kinases kinases of range wide a to applicable is technique This •
• The approach can be used to look at allosteric inhibitors in addition to ATP competitive inhibitors inhibitors competitive ATP to addition in inhibitors allosteric at look to used be can approach The •
• Examines the K-on (association rate) and K-off (dissociation rate) of compounds in live cells cells live in compounds of rate) (dissociation off - K and rate) (association on - K the Examines •
pseudo biochemical binding assay assay binding biochemical pseudo
• Cells can be disrupted (with digitonin) to release cellular content from the cell and the assay can be performed as a a as performed be can assay the and cell the from content cellular release to digitonin) (with disrupted be can Cells •
full-length length - full • Expression of kinase target in cells cells in target kinase of Expression •
Intracellular physiological ATP ATP physiological Intracellular • examination of compound binding to target kinase at concentrations in live cells cells live in concentrations at kinase target to binding compound of examination •
assay assay
• First biophysical technique to profile compound binding for intracellular targets – truly functional pharmacology pharmacology functional truly – targets intracellular for binding compound profile to technique biophysical First •
Advantages Advantages : :
during compound SAR development development SAR compound during
(B) and the residence time (C) of compounds at the target kinase allows kinetic binding parameter evaluation evaluation parameter binding kinetic allows kinase target the at compounds of (C) time residence the and (B)
Competitive inhibition (A) of compound binding to DDR1 kinase in living cells. Studying both the association rate rate association the both tudying S cells. living in kinase DDR1 to binding compound of (A) inhibition Competitive
A A B B
C C
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for the target kinase. . kinase target the for
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dependent decrease in NanoBRET™ signal, which allows allows which signal, ™ NanoBRET in decrease dependent
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energy transfer probe introduced to the assay medium. . medium assay the to introduced probe transfer energy
Nluc)-tagged kinase and a cell-permeable fluorescent fluorescent permeable - cell a and kinase tagged - ) Nluc
via energy transfer between a 19-kDa luciferase (NanoLuc, , NanoLuc ( luciferase kDa - 19 a between transfer energy via
• This quantitative analysis can be performed in cultured cells cells cultured in performed be can analysis quantitative This •
of compounds on kinase targets in intact living cells. . cells living intact in targets kinase on compounds of
kinase assays designed to study the interaction interaction the study to designed assays kinase intracellular intracellular Profiling Compound Kinase
Bioscience has implemented target engagement engagement target implemented has Bioscience
Using Promega’s NanoBRET™ technology, Aurelia Aurelia technology, ™ NanoBRET Promega’s Using •
• discovery” drug kinase your energise - “Re How it works: : works it How
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S 2, tudyingS SNARKtudying both both the the association association rate rate rate association the both tudying S cells. living in kinase DDR1 to binding compound of (A) inhibition Competitive C C Competitive inhibition (A) of compound binding to DDR1 kinase in living cells. Studying both the association rate : Advantages B B A A (B)(B) and and the the residence residenceRIOK2 time time (C) (C) of ofcompounds compoundsAtypical at atthe the target target kinase kinase allows allows kinetic kinetic binding binding parameter parameter evaluation evaluation evaluation parameter binding kinetic allows kinase target the at compounds of (C) time residence the and (B) (B) and the residence time (C) of compounds at the target kinase allows kinetic binding parameter evaluation RIPK2 duringTKLduringduring compound compound compoundRIP2, SAR SAR SAR developmentGIG30, development development CARDIAK, RICK, CCK development SAR compound during pharmacology functional truly – targets intracellular for binding compound profile to technique biophysical First • both the association rate rate rate association association the the both both tudying tudying S S cells. cells. living living in in kinase kinase DDR1 DDR1 to to binding binding compound compound of of (A) (A) inhibition inhibition Competitive Competitive SIK1 CAMK SIK, MSK assay binding parameter evaluation evaluation evaluation parameter parameter binding binding kinetic kinetic allows allows kinase kinase target target the the at at compounds compounds of of (C) (C) time time residence residence the the and and (B) (B) SIK3 CAMK QSK, L19, KIAA0999 during compound SAR development development SAR SAR compound compound during during cells live in concentrations ATP physiological at kinase target to binding compound of examination Intracellular • AdvantagesAdvantages:SRC: TK SRC1, ASV : Advantages Advantages: cells in target kinase length - 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