Differential sex-related pattern on intestinal P-glycoprotein activity: impact on ivermectin secretion along the digestive tract

LIFSCHITZ, A.; BALLENT, M.; VIRKEL, G.; SALLOVITZ, J.; CERIANI, C.; SOLANA, H.; LANUSSE, C.

Torino, Italia

Simposio; 10th International Congress of The European Association for Veterinary Pharmacology and Toxicology; 2006

Introduction: The role of P-glycoprotein (P-gp) on the pharmacokinetic disposition of different drugs has been demonstrated. Differences on P-gp function among animal species have also been shown (1). The antiparasitic drug ivermectin (IVM) is a P-gp substrate and the involvement of P-gp on the IVM intestinal excretion has been recently demonstrated (2, 3). Sex-related differences on the disposition of IVM in cattle (4) and selamectin in dogs (5) have been observed. A suitable approach to assess the in vivo P-gp function/activity is the evaluation of the effects of known P-gp inhibitors on drug disposition. The ratio between IVM concentrations in the intestinal wall and luminal content may be used as an estimator of the capacity of inhibition of P-gp modulator agents on the in vivo gastrointestinal (GI) secretion. The aim of the current study was to evaluate in vivo the influence of animal gender and P-gp modulation on the in vivo GI secretion of IVM using a laboratory animal model. Complementary work included the comparative assessment of P-gp expression in liver tissue and along different sections of the GI tract in female and male sheep. Material and Methods: The trial was conducted in two experimental phases. Animal’s management was in agreement with institutional and internationally accepted welfare guidelines. Phase 1: Seventy five (75) Wistar rats (45 males, 30 females) weighing 350-400 g were used. Rats were separated by sex and randomly allocated into five (5) experimental groups of fifteen (15) animals. Animals received IVM (200 µg/kg) by the subcutaneous (SC) route either alone (Group A: IVM alone in females; Group B: IVM alone in males) or co-administered with the P-gp modulators itraconazole (ITZ, 3 x 5 mg doses every 12 h) (Group C: IVM+ITZ in females; Group D: IVM+ITZ in males) and PSC833 (3 x 8.6 mg doses every 12 h) (Group E: IVM+PSC833 in males). Rats were sacrificed (between 6 and 72 h pt) and GI tissues and lumen contents were collected. Tissue and fluid digestive samples were cleaned up and extracted to measure IVM concentrations by HPLC as previously reported (6, 7). Phase II: Six adult untreated Corriedale sheep (3 males, 3 females) were sacrificed. Liver tissue and mucosal samples from duodenum, ileum and colon were collected. Western Blot analyses were performed on protein samples from liver and intestinal homogenates using the C219 monoclonal antibody (8) to assess P-gp tissue expression. Results: The ratios between IVM peak concentrations (Cmax) measured in the wall tissue and luminal content (wall/content ratio) of the different GI sections were estimated. The Cmax wall/content ratio obtained in the proximal section of the GI tract (stomach-jejunum) was 3.25-fold higher (P<0.05) compared to that measured at the caudal region of the gut (ileum-colon). A significant enhancement on the wall/content Cmax ratio in the different sections of the GI tract was obtained after the IVM+PSC833 treatment (0.99 ± 0.30) compared to both IVM+ITZ (0.35 ± 0.05) and IVM alone (0.20 ± 0.04) treatments. An evident gender difference was observed on the concentration profiles measured along the GI tract. The mean Cmax wall/content ratios were 69 % higher in female (P< 0.05) compared to male rats. The ITZ-induced modulation of the P-gp-mediated IVM secretion was markedly greater in male rats. Whereas the ratio wall/content increased a 297 % in the presence of ITZ in males, the enhancement in female animals reached a 93% (P< 0.05). Additionally, the use of the C219 monoclonal antibody successfully permitted the assessment of the P-gp expression along the GI tract in sheep. The Western Blot analysis showed that P-gp expression in the digestive tract tends to be higher in male compared to female sheep. Discussion: The Cmax wall/content ratio was a useful tool to estimate the IVM active intestinal transport. The higher ratios obtained in the proximal section of the GI tract agrees with previous reports (9) indicating a higher P-gp activity in ileum and distal colon in mice. The comparative evaluation between both inhibitory agents indicated that PSC833 is more effective than ITZ to modulate the P-gp-mediated intestinal secretion of IVM. Altogether, the results presented here may suggest that male animals may have an enhanced intestinal P-gp activity, which would explain the observed sex-related differences on the IVM GI disposition. Consistently, a higher P-gp expression along the GI tract was observed in male compared to female sheep. Further work on this therapeutically relevant area is required.