Abstract
To investigate the possible involvement of P-glycoprotein (P-gp), multidrug resistance-associated protein (MRP), and/or other glutathione S-conjugate export pump (GS-X pump) family members on the active efflux of irinotecan [(7-ethyl-10-[4-(1-piperidino)-1-pipertidino)-1-piperidino]carbonyloxy camptothecin (CPT-11)] and its metabolites, as well as their contribution to the acquisition of resistance, we studied the uptake of CPT-11, its active metabolite SN-38, and glucuronide conjugate (SN38-Glu) using membrane vesicles from human epidermoid KB-3-1-derived cell lines. These lines included KB-C2, C-A500, and KCP-4, which overexpress P-gp, MRP, and the unidentified GS-X pump, respectively. The carboxylate form of SN-38 exhibited significant ATP-dependent transport, with a Michaelis constant of 17 μM, into membrane vesicles from C-A500 but not from other cell lines. Among these KB-derived cells, significant ATP-dependent uptake of the carboxylate form of CPT-11 was only observed in KB-C2 vesicles. In addition, the uptake of the lactone and carboxylate forms of SN38-Glu into membrane vesicles from C-A500 and KB-C2, but not KCP-4, was ATP dependent, although the transport activity in C-A500 was much higher than that in KB-C2. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay revealed that the resistance of KB-C2 to CPT-11 andSN-38, compared with that of KB-3-1, was 6.3- and 6.8-fold, respectively; the corresponding figures for C-A500 were 12- and 27-fold, respectively, whereas those for KCP-4 were 2.3- and 20-fold, respectively. These results suggest that MRP and P-gp are involved in the active efflux of SN-38 and CPT-11, respectively, from human KB-derived cells. In addition, a difference in substrate specificity among GS-X pump members was demonstrated.
Footnotes
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Send reprint requests to: Yuichi Sugiyama, Ph.D., Graduate School of Pharmaceutical, Sciences, The University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113-0033, Japan. E-mail:sugiyama{at}seizai.f.u-tokyo.ac.jp
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↵1 This work was supported in part by a grant-in-aid from the Ministry of Education, Science, Sports and Culture of Japan, and the Core Research for Evolutional Sciences and Technology of Japan Sciences and Technology Corporation.
- Abbreviations:
- CMV
- canalicular membrane vesicles
- CPT
- camptothecin
- CPT-11
- (7-ethyl-10-[4-(1-piperidino)-1-pipertidino)-1-piperidino] carbonyloxy camptothecin
- SN38-Glu
- SN38-glucuronide
- P-gp
- P-glycoprotein
- MRP
- multidrug resistance-associated protein
- GS-X pump
- ATP-dependent glutathione S-conjugate export pump, cMOAT, canalicular multispecific organic anion transporter
- MTT
- 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide
- Received June 24, 1998.
- Accepted September 3, 1998.
- The American Society for Pharmacology and Experimental Therapeutics
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