Excretion of Aristolochic Acid I in the IPK Page 1 Mariana Babayeva MD PhD Touro College of Pharmacy New York NY USA Aristolochic Acid I Page 2 Compound extracted from plant ID: 363530
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Renal Excretion of Aristolochic Acid I in the IPK
Page 1
Mariana Babayeva MD, PhD
Touro College of Pharmacy,
New York, NY, USASlide2
Aristolochic Acid IPage 2Compound extracted from plant AristolochiaNephrotoxin and carcinogenSlide3
Adverse Effects of AA-IPage 3AA-I is an organic anion eliminated by the kidneyProduces nephrotoxic effect to S3 segment of renal proximal tubule Slide4
Renal Excretion Page 4Glomerular Filtration GFR: glomerular filtration rateTubular Secretion Active transport Secretion transporters: OATs, OCTs, etc.Tubular Reabsorption Active transportReabsorptive transportersPassive transport
Glomerular Filtration (1)
Tubular Secretion (2)
Tubular Reabsorption (3)
Renal Excretion (4)Slide5
Renal Organic Anion Transport (OAT)Page 5OAT system plays an important role in tubular secretion and reabsorption of compounds (organic anions)Slide6
ObjectivesPage 6The overall goal of the research was to assess transport mechanism of renal excretion of AA-I. Further identify potential strategies to mitigate drug toxicity by reducing renal uptakeSlide7
MethodsStep 1 Step 2 Step 3Slide8
Isolated Perfused Kidney (IPK) Model Assessment of renal drug excretion mechanism- Dose-linearity- Inhibition studiesDrug interaction screeningModel for nephrotoxicityProbing renal drug metabolismGender differences in renal function and drug excretion
Correlation between drug excretion and membrane transporter expression
Model for aging
Studies in mutant strains (genetic “knockout” animals)Slide9
IPK Surgical Procedurecannulate forurine collectioncannulate forkidney perfusionSlide10
IPK Apparatus Slide11
Perfusate CompositionKrebs-Henseleit buffer (KHS buffer)Bovine serum albumin (BSA)DextranGlucoseInulinAmino acidsMixture of 20 amino acids Slide12
IPK Study GroupsTreatment GroupCompound(s)(Concentration)
Justification
Control Perfusion
None
Establish viability of preparation and allow for evaluating of drug effects on kidney function
AA-I Excretion
Aristolochic acid I
(20uM)
Obtain baseline parameter values of renal excretion and of AA-I
AA-I Transport Inhibition
Aristolochic
acid I
(20uM)
+
Probenecid
(1mM)
Study mechanisms of AA-I renal transport Slide13
IPK Experimental Design Slide14
IPK Viability Criteria Viability ParametersMinimum Acceptable ValueGFR
> 0.5 ml/min
Glucose Reabsorption (FR
Glu
)
> 90%
Sodium Reabsorption (FR
Na
)
> 85%
Urine Flow Rate
> 0.03 ml/minSlide15
Renal Excretion Parameters dXU/dt = urinary drug excretion rate Cp = perfusate drug concentration XR = excretion ratio CLr = clearance fu = fraction unbound GFR = glomerular filtration rate
XR > 1 = net secretion process
XR < 1 = net reabsorption processSlide16
ResultsProtein Binding of AA-IIPK Viability ParametersAA-I Renal Excretion ParametersSlide17
Protein Binding StudiesThere were no significant differences in protein binding among the different concentrations of AA-I (ANOVA, p >0.05) Ultrafiltration technique was used for the protein binding studiesTable 1. Summary of AAI Protein Binding Studies in Perfusate Slide18
Protein Binding StudiesThere was no significant difference in protein binding between the two study groups (ANOVA, p >0.05) Table 2. Effect of Probenecid on AAI Protein Binding in Perfusate Slide19
Viability of the Perfused Kidney Kidney function was well maintained across all study groupsThe IPK technique has been successfully applied Slide20
Renal Excretion StudiesTable 5. Summary of AAI Renal Excretion Parameters in IPKThe renal excretion parameters ~ 2-fold higher in studies with PBCSlide21
Plot of Cumulative Amount of AA-I Excreted in Urine vs Time. Effect of Probenecid on AA-I Excretion in the IPK.Slide22
ResultsPage 22XR of AA-I < 1 (0.08 and 0.17) for both study groups The results suggest net reabsorptionXR of AA-I was more than 2-fold higher in the presence of probenecidProbenecid inhibited the tubular reabsorption of AA-I most probably by interaction with renal organic anion transport systemThe amount of unrecovered AA-I in Phase I studies was 3-fold higher than in Phase IIProbenecid decreased accumulation of AA-I in the kidney cellsSlide23
DiscussionPage 23Bidirectional Transport: Oat2 / Oat5Slide24
DiscussionPage 24Slide25
ConclusionPage 25The present study described the transport mechanism of Aristolochic acid I renal excretionThe observation suggested that the renal apical transporters (Oat2 and Oat5) may function as reabsorptive pathway during renal elimination of AA-I.Inhibition of
AA-I reabsorption can decrease nephrotoxicity of AA-I.
This assumption requires further investigationSlide26