L-Trpisanessentialaminoacidnecessaryforproteinsynthesisinmammaliancells,andtheL-Trptokynurenine(Kyn)pathwayisfirmlyestablishedasakeyregulatorofinnateandadaptiveimmunity.CatabolismofL-TrptoKynmaintainsanimmunosuppressivemicroenvironmentbystarvingimmunecellsofL-TrpandreleasingdegradationproductsofL-Trpthathaveimmunosuppressivefunctions.Indoleamine2,3-dioxygenases(IDO1&IDO2),twooftheratelimitingenzymesinthispathway,areupregulatedinmanytumors,providingcancercellswithanavenueforimmuneevasion.
InhibitorsofIDO1canbindeitherirreversIBLy(covalentbindingtotheprotein)orreversibly(non-covalentassociation)totheenzyme.TheIDO1InhibitorMechanismofActionAssayKitallowsresearcherstodeterminethemechanismofIDO1inhibitorbinding.Athighconcentrations,e.g.,10xtheIC50concentrationoftheinhibitor,bothreversibleandirreversibleinhibitorswillinterferewithIDO1activity.However,aftersufficientdilution,e.g.to0.3xtheIC50concentration,reversibleinhibitorswilldissociate,relievinginhibitionoftheIDO1enzyme.Irreversibleinhibitorswillnotdissociate,andwillcontinuetoinhibittheIDO1enzymeatthesamelevel.Therefore,bymonitoringenzymaticactivityafterinhibitordilution,themechanismofbindingtotheIDO1proteincanbedetermined.
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