Evaluation of drug-drug interactions at the level of hepatic excretion

Stanislav Mičuda

Department of Pharmacology, Charles University in PragueFaculty of Medicine in Hradec Králové

55. Česko-Slovenské Farmakologické dny30.8.-1.9.2005, Hradec Králové

55. Farmakologické dnyHepatic

drug processing

MDR1

MDR3

NTCP

MRP2 MRP2

NTCP

BCRP

OCT1

MRP3

Phase IPhase II

BILE SALTS BULKY ORGANIC

COMPOUNDSORGANICANIONS

SMALL ORGANICCATIONS

OATs

BSEP

OATP2

Inhibition – MDR1

Shon JH; Clin Pharmacol Ther 2005;78(2):191-201 Kovarik JM; Clin Pharmacol Ther 1999;66(4):391-400

Itraconazole

Digoxin 0.125 mg +valspodar 400 mg 7-11 day

Greiner B; J Clin Invest 1999;104(2):147-53.

Induction – MDR1

In vivo

Single and Repeat Dose PharmacokineticsDrug Interaction StudiesMass Balance Excretion Studies Imaging StudiesIn Situ Tissue Perfusions

Preclinical -Clinical Studies

Mai I; Clin Pharmacol Ther 2004;76(4):330-40.Sasongko L; Clin Pharmacol Ther 2005;77(6):503-14.

99mTc-Sestamibi Scan following XR-9576

– a synthetic corticosteroid, (t1/2 = 36-54 h), eliminated mainly in the unchanged form in urine – induction of mrp2 protein expression?

SPC: 86698 DEXAMETHAZON LÉČIVA TBL 20X0.5MG-BLISTR LEX CZ

Dexamethasone

The aim of study

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influence of dexamethasone on expression and activity of mrp2

Methotrexate (MTX) as a selective substrate for mrp2

Probenecid as an inhibitor of mrp2

In vivo pharmacokinetics

Wistar male rats (Konárovice) - 3 groups (N = 6) Dex (25 mg/kg daily 4 days, p.o.),Control (Oliv. oil 4 days, p.o.), Probenecid 70 mol/kg 3.33 mol/min.kg-1

simultaneously with MTX

Clearance study left jugular v. (MTX 22 mol/kg 164 nmol/kg.h-1 4 ml/min) right a. carotis (blood sampling - since 55‘ - 10 min) bladder (urine 0-90 min 10 min) bile duct (bile 0-90 min 10 min)

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MTX concentrations HPLC method with fluorescent detection *

Farmacokinetic analysis

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In vivo pharmacokinetics

CLtotal = Rinf / Css

CLbil = Cbil . Vbil / Css

CLren = CU.VU / Css

BE = Cbil . Vbil

UE = CU.VU

CLRratio = CLren / GFR

CLRratioU = CLren / (fU .GFR)

CLRS = (CLren / fU) - GFR

* Chladek J; J Chromatogr B Biomed Sci Appl 2000;744(2):307-13.

Pharmacokinetics of MTX in steady-state

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60 70 80 90 100

0

10

20

30

40Probenecid

Control

Dexamethasone

Time (min)

Me

tho

tre

xa

te (

uM

)

Pharmacokinetics of MTX in steady-state

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Control DEX 25 mg po Probenecid DIF

(fold) values DIF

(fold) Urine flow rate (l/min) 28.9 4.7 71.4 8.2 C 2.5 27.7 8.5 1.0

UE rate (nmol/ml.kg-1) 92.7 10.6

85.9 5.8 0.9 57.0 13.5 A 0.6

Bile flow rate (l/min) 25.2 1.6 28.0 2.7 1.1 32.7 2.8 A 1.3

BE rate (nmol/ml.kg-1) 98.5 8.2 124.0 9.9 A 1.3 62.0 5.2 B 0.6

Plasma (M) 18.4 1.3 20.2 2.2 1.1 35.4 1.2 C 1.9

CLR (ml/min.kg-1) 5.0 0.4 4.6 0.7 0.9 1.6 0.4 C 0.3

CLBile (ml/min.kg-1) 5.6 0.7 6.4 0.6 1.1 1.8 0.2 C 0.3

CLTOT (ml/min.kg-1) 9.2 0.6 8.7 1.0 0.9 4.7 0.2 C 0.5

CLCR (ml/min.kg-1) 4.8 0.6 4.9 0.4 0.9 3.6 0.4 0.7

fU 0.5 0.02 0.7 0.1 + 1.3 0.6 0.03 B 1.3

CLR/fu 10.0 0.2 6.7 0.2 B 0.7 2.4 0.2 C 0.2

CLR/GFR 1.1 0.9 1.1 0.4 0.9 0.5 0.5 B 0.4

CLRS 5.3 1.0 2.2 0.6 A 0.4 -1.2 0.8 C -0.2

CLR/(fU x GFR) 2.2 0.3 1.6 0.2 + 0.7 0.7 0.2 B 0.4

Values are means SEM A P < 0.05; B P < 0.01; C P < 0.001

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Dexametazon 25 mg/kgOlive oilRat liver - mrp2 – Dex per os

Control Dexamethasone Parameter for image analysis

Centr Port Centr Port

Proportion of visual field area occupied by mrp2 staining

0.13 ± 0.05 2.11 ± 0.772 2.92 ± 0.88 C 3.91 ± 1.14 A

Integral optical densities of mrp2 in visual field

28 ± 11 467 ± 1742 664 ± 202 C 903 ± 266 A

Values are means ± SEM of 6 rats. Centr - pericentral areas; Port - periportal areas; 1P<0.05, 2P<0.001 periportal vs pericentral areas; AP<0.05, CP<0.001 comparison of data from respective locations between control and dexamethasone (25 mg/kg daily, p.o. for 4 days) pretreated animals.

BE 1.3-foldCLbile 1.1-foldHistol 3.1-foldWestern 2.4-fold

Pharmacokinetics of Rho-123

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Influence of DEXon P-gp activity

Control DEX 25 mg po

DIF (fold)

Urine flow rate (l/min) 26.0 4.0 96.0 12.0 C 3.6

UE rate (nmol/ml.kg-1) 0.10 0.04 0.28 0.06 B 2.7

Bile flow rate (l/min) 27.0 2.0 28.0 4.0 1.0

BE rate (nmol/ml.kg-1) 0.26 0.04 0.57 0.14 A 2.2

Plasma (M) 0.12 0.02 0.08 0.003 B 0.6

CLR (ml/min.kg-1) 0.71 0.24 3.70 0.68 B 5.2

CLBile (ml/min.kg-1) 2.22 0.34 7.88 2.29 A 3.5

CLTOT (ml/min.kg-1) 21.68 1.00 29.26 1.18 B 1.4

CLCR (ml/min.kg-1) 1.19 1.01 1.34 1.20 1.1

CLR/GFR 0.59 0.08 2.90 0.10 B 4.9

Values are means SEM A P < 0.05; B P < 0.01; C P < 0.001 Ctrl DEX

BE 2.2-foldCLbile 3.5-foldHistol 2.3-foldWestern 2.8-foldReal RT-PCR 3.1-fold

Conclusion

Drug-drug interactions may be of great clinical importance - A significant number is transport protein-mediated- in vivo studies – a complex view of pharmacokinetics

and mechanisms using model substrates

- in vivo studies – verification of outcome from in vitro models

allowing predictions- interspecies extrapolation

Glucocorticoids and MRP2 vs. MTX- induction of proteins without changes in pharmacokinetics of MTX

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ACKNOWLEDGEMENTS

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Grants GAUK 89/2002/CMŠMT EC-B25.001

Co-workersLeoš FuksaJolana CermanováEva Brčáková Jitka Hájková František ŠtaudPetr Pávek Lucie MundlováJan OsterreicherJaroslav MokrýJaroslav ChládekJiřina Martínková

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