CanagliflozinisaselectiveSGLT2inhibitorwithIC₅₀of2nM,3.7nM,and4.4nMformSGLT2,rSGLT2,andhSGLT2inCHOKcells,respectively.

IC50:2/3.7/4.4nM(m/r/hSGLT2,inCHOKcells)^[1]

Canagliflozinisasodiumglucoseco-transporter(SGLT)2inhibitor.Inaconcentration-dependentfashion,CanagliflozininhibitsNa⁺-dependent¹⁴C-AMGuptakeinCHO-hSGLT2cells,withanIC₅₀of4.4±1.2nM.SimilarIC₅₀valuesareobtainedinCHO-rSGLT2andCHO-mSGLT2cells(IC₅₀=3.7and2.0nMforratandmouseSGLT2,respectively).Canagliflozininhibits¹⁴C-AMGuptakeinCHO-hSGLT1andmSGLT1cellswithIC₅₀of684±159nMand>1,000nM,respectively.At10µM,Canagliflozininhibitsthefacilitative(non-Na⁺-linked)GLUT-mediated3H-2-DGuptakeinL6myoblastsbylessthan50%^[1].

Canagliflozintreatment(1mg/kg)notablylowersrenalthresholdforglucoseexcretion(RT_G)inZuckerdiabeticfatty(ZDF)ratsto94±10mg/dL.Inthesecondstudy,aninsulininfusionisgiventolowerbloodglucose(BG)toapproximately25mg/dL,andthenthegradedglucoseinfusion(GGI)isgiventoslowlyraiseBGtoapproximately300mg/dL.InZDFratstreatedwithCanagliflozin(1mg/kg),therelationshipbetweenBGandurinaryglucoseexcretion(UGE)remainswell-describedbyathresholdrelationshipwithnegligIBLeUGEoccurringwhenBG<90 mg/dl="" and="" uge="" increases="" in="" proportion="" to="" bg="" at="" higher="" bg="" levels.="" in="" db/db="" mice,="" single="" doses="" of="" canagliflozin="" dose-dependently="" reduces="" non-fasting="" bg="" concentrations.="" the="" onset="" of="" the="" bg-lowering="" effect="" after="" a="" single="" dose="" is="" rapid,="" and="" bg="" levels="" in="" canagliflozin-treated="" mice="" (at="" 1="" and="" 10="" mg/kg="" doses)="" are="" significantly="" different="" from="" those="" of="" vehicle-treated="" mice="" at="" 1="" hour="" after="" treatment="" (22%="" and="" 36%="" reduction="" of="" bg="" levels="" compared="" with="" that="" in="" vehicle-treated="" mice,="">^[1].

• [1].LiangY,etal.Effectofcanagliflozinonrenalthresholdforglucose,glycemia,andbodyweightinnormalanddiabeticanimalmodels.PLoSOne.2012;7(2):e30555.

  [2].KobuchiS,etal.AvalidatedLC-MS/MSmethodforthedeterminationofcanagliflozin,asodium-glucoseco-transporter2(SGLT-2)inhibitor,inalowervolumeofratplasma:applicationtopharmacokineticstudiesinrats.BiomedChromatogr.2016Oct;30(10):1549-55.

CanagliflozinisdissolvedinDMSOandstored,andthendilutedwithappropriatemedia(DMSO0.1%)beforeuse^[1].

Sodium-dependentglucoseuptakeinChineseHamsterOvary(CHO)cellsexpressingSGLT1andSGLT2co-transportersparentalCHO-K(CHOK)cells(commonlyusedmammaliancellsforgeneoverexpressionstudies)expressinghumanormouseSGLT1andSGLT2areutilizedinthisstudy.Cellsareseededinto96-wellplates.Cellsarethenwashedonetimewith0.15mLassaybuffer(137mMNaCl,5mMKCl,1mMCaCl₂,1mMMgCl₂,50mMHEPES,pH7.4)at37°C.Aftertheassaybufferisremoved,50µLoffreshassaybufferwith5µLofCanagliflozin(0.3-300nM)isadded,followedby10minutesofincubation.Then,5µLof6mMalpha-methyl-D-glucopyranoside(AMG,aselectiveSGLT1/2substrate)/¹⁴C-AMG(0.07µCi)isaddedtothecellsandincubatedat37°Cfor2hours.Next,thecellsarewashed3timeswith0.15mLice-coldphosphate-bufferedsaline(PBS).Afterthefinalwashisaspirated,50µLofmicroscint20isadded.TheplateiscountedbyTopCount^[1].MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly.

Canagliflozinisformulatedin0.5%hydroxypropylmethylcellulose^[1].

MiceandRat^[1]
Fourrodentmodelsareusedintheseexperiments:(1)maleC57BL/6Jmicefedwithahigh-fatdiet(D-12492with60kcal%fat)(diet-inducedobese,insulinresistantmice[DIO]);(2)maleC57BL/ksj-db/dbhyperglycemicmice;(3)maleZuckerfatty(ZF)obese,insulinresistantrats;and(4)maleZDFobese,hyperglycemicrats.ToexaminetheeffectofCanagliflozinonhyperglycemia,singledosesofCanagliflozin(0.1,1,and10mg/kg)areadministeredtoovernight-fasteddb/dbmice.BGlevelsaremonitoredat0,0.5,1,3,6,and24hoursafterdosing.CanagliflozinisalsoadministeredtoZDFratsatvaryingdoses(3-30mg/kg)for4weekstoevaluateitseffectonBGcontrolandpancreaticbeta-cellfunction.BGlevelsaremonitoredweekly,andHbA1c,plasmaglucose,andinsulinlevelsaredeterminedattheendofthe4-weektreatment.Anoralglucosetolerancetest(OGTT)(2mg/kgofbodyweight,givenbygavage)isconductedinZDFratsafter4weeksoftreatment.Bloodissampledat0,30,60,and120minutesafterglucosechallengefromthetailveinformeasurementofBGlevelsusingaglucometerandplasmainsulinusingELISAmethod.MCEhasnotindependentlyconfirmedtheaccuracyofthesemethods.Theyareforreferenceonly.

• [1].LiangY,etal.Effectofcanagliflozinonrenalthresholdforglucose,glycemia,andbodyweightinnormalanddiabeticanimalmodels.PLoSOne.2012;7(2):e30555.

  [2].KobuchiS,etal.AvalidatedLC-MS/MSmethodforthedeterminationofcanagliflozin,asodium-glucoseco-transporter2(SGLT-2)inhibitor,inalowervolumeofratplasma:applicationtopharmacokineticstudiesinrats.BiomedChromatogr.2016Oct;30(10):1549-55.

444.52

C₂₄H₂₅FO₅S

842133-18-0

RoomtemperatureincontinentalUS;mayvaryelsewhere

10mMinDMSO

Canagliflozinispreparedin1%carboxymethyl-cellulosesodiumindistilledwater^[2].

*"<1 mg/ml"="" means="" slightly="" soluble="" or="" insoluble.="" "≥"="" means="" soluble,="" but="" saturation="">

• [1].LiangY,etal.Effectofcanagliflozinonrenalthresholdforglucose,glycemia,andbodyweightinnormalanddiabeticanimalmodels.PLoSOne.2012;7(2):e30555.

  [2].KobuchiS,etal.AvalidatedLC-MS/MSmethodforthedeterminationofcanagliflozin,asodium-glucoseco-transporter2(SGLT-2)inhibitor,inalowervolumeofratplasma:applicationtopharmacokineticstudiesinrats.BiomedChromatogr.2016Oct;30(10):1549-55.