4)  RELEVANCE OF THE ISOLATED PERFUSED RAT LIVER TO EXAMINE THE BILIARY EXCRETION OF SPECIFIC SUBSTRATE OF THE C-MOAT.

Pierre Gires*1, Alain Grelier, Daniel Poulet and Yves Archimbaud.

Abstract of a poster presented in St Andrew  ISSX meeting  (Scotland)  in June 2000.

The hepatocyte is a polarized secretory epithelial cell with specific transporters localized either to the sinusoidal (basolateral) and canalicular (apical) domains. To secrete bile and to excrete metabolites of toxic substances, hepatocytes must transport bile salts, phospholipid and other solutes from blood to bile. In recent years, various transporters for uptake or excretion have been cloned and characterized. The canalicular multispecifc organic anion transporter, or c-MOAT has a substrate specificity for the ATP-dependent transport of glutathion and glucuronide conjugates and is responsible for the biliary clearance of these metabolites, particulary BSP-glutathione, p-nitrophenyl glucuronide and Estradiol-17b -D-glucuronide (E217G).

The aim of the present study was to use the isolated perfused rat liver technique to examine the influence of a c-MOAT specific substrate as the antitubercular drug Rifampin on the hepatobiliary transport of BSP. It was shown that Rifampin caused a dose-dependent inhibition of BSP removal from the perfusate and excretion in the bile. On the contrary, BSP had any influence on the biliary elimination of 3H Rifampin. This could suggest that a saturation process occurred for the BSP-glutathione transport.Furthermore, using TR- rats (c-MOAT deficient), it was shown a significant decreasing of the biliary elimination of both compounds. It confirms that BSP-glutathione and some products of Rifampin should be eliminated in bile via this transporter. The use of this experimental model has been useful in evaluating the mechanisms of liver-specific drug effect without interfering with other organs.

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5)  Interest of the IPRL used in preclinical studies and experimental surgery

Isolated perfused rat liver is a powerful and versatile model used mainly to support pharmacokinetics  and metabolism during the preclinical phase of the development of a new drug. Also, this model is able to get relevant and predictive information about toxicology and to support preformulation. Briefly, it’s a recirculating system enclose in a thermostated cabinet as shown below.  

A semi-synthetic medium is commonly used. It's consisted of blood rat diluted in krebs buffer supplemented in BSA (5%). The final hematocrit is about 13% and the pH is maintained at 7.4. This medium is continuously oxygenated to maintain a good viability of the parenchyma. In the standard conditions, bile flow rate is about stable over 3 hours at 1.2 µl/g/min. To assume the oxygen transport to the parenchyma, hepatic flow rate is about 3 fold higher than in vivo. Moreover, a constant infusion of bile salts assumes the enterohepatic recirculation to mimic the in vivo situation. Numerous studies in drug disposition and in toxicology are usually carried out using this model. Furthermore, this technology is currently applied in Experimental Surgery.  

Currently, numerous articles are focused about the preservation effect of on the liver using of  CMU-1 solution or  UW solution. 

 

Hepatic extraction, metabolism, and biliary excretion of irinotecan in the isolated perfused rat liver.

Farabos C, Haaz MC, Gires P, Robert J.

Laboratoire de Pharmacologie des Medicaments Anticancereux Institut Bergonie, 180 rue de Saint-Genes 33076 BORDEAUX-cedex, France.

Irinotecan (CPT-11) is a semisynthetic derivative of camptothecine that has proved activity in the treatment of colorectal carcinoma. The metabolites identified in humans include SN-38, SN-38 glucuronide, and several CYP3A-derived metabolites. We have studied the hepatic extraction, metabolism, and biliary excretion of irinotecan in the isolated perfused rat liver. After injection of a bolus dose of 5 micromol in the reservoir, irinotecan lactone disappeared from the perfusate following a two-exponential decay with half-lives of 3.5 and 120 min and a total clearance of 1.54 +/- 0.07 mL/min per gram of liver. The area under the curve (AUC) ratio lactone/total drug was 0.212 +/- 0.098 and the half-life of interconversion was 5.02 +/- 0.10 min. Bolus administrations of 2.5, 5, and 25 micromol of irinotecan gave AUCs proportional to the doses administered, indicating that no saturation occurred during dose increase. However, the relative formation of SN-38 and SN-38 glucuronide decreased at the high dose. This result was not the case for the CYP3A metabolites, which had identical metabolic ratios at all three doses. Infusions of 30 and 90 min of a dose of 5 micromol led to the same AUCs and metabolic ratios as a bolus of the same dose. Biliary elimination of irinotecan and metabolites represented 18-22% of the dose administered at 2.5 and 5 micromol but only 7-9% at 25 micromol, suggesting a saturation of this process. These data indicate that the hepatic disposition of irinotecan may vary at high dose, both at the level of biliary excretion and of activation to SN-38. Copyright 2001 Wiley-Liss, Inc.

PMID: 11357175 [PubMed - indexed for MEDLINE]

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