Cancer Letters

Cancer Letters

Volume 138, Issues 1–2, 26 April 1999, Pages 13-22
Cancer Letters

An enhanced active efflux of CPT-11 and SN-38 in cisplatin-resistant human KB carcinoma cells

https://doi.org/10.1016/S0304-3835(98)00367-XGet rights and content

Abstract

Cisplatin-resistant KCP-4 cells were 12.4- and 31.6-fold more resistant to CPT-11 and SN-38 than parental KB-3-1 cells, respectively. We studied the mechanism of cross-resistance to CPT-11 and SN-38. Our previous study showed that multidrug resistance protein (MRP), canalicular multispecific organic anion transporter (cMOAT) and P-glycoprotein (P-gp) were not expressed in KCP-4 cells (Chen, Z.-S. et al., Exp. Cell Res., 240 (1998) 312–320, and Chuman, Y. et al., Biochem. Biophys. Res. Commun., 226 (1996) 158–165). The accumulation of both CPT-11 and SN-38 in KCP-4 cells was lower than that in KB-3-1 cells. The ATP-dependent efflux of CPT-11 and SN-38 from KCP-4 cells was enhanced compared with that from KB-3-1 cells. DNA topoisomerase (topo) I expression, topo I activity, topo I-mediated cleavable complex, and the sensitivity to SN-38 of DNA topo I in KCP-4 were similar to those in KB-3-1 cells. Furthermore, the conversion of CPT-11 to SN-38 in the two cell lines was also similar. The transport of LTC4 in KCP-4 membrane vesicles was competitively inhibited by bis-(glutathionato)-platinum (II) (GS-Pt), CPT-11 and SN-38. These findings suggested that an unknown transporter distinct from P-gp, MRP or cMOAT is expressed in KCP-4 cells and transports CPT-11 and SN-38.

Introduction

It is very important to elucidate the mechanism for resistance to camptothecin (CPT) and its analog, CPT-11, since the resistance to CPT and CPT-11 in tumor cells reduces the success of chemotherapy. Many cell lines resistant to CPT analogs have been isolated in vitro and some mechanisms of resistance to CPT analogs have been determined. These mechanisms of resistance include decreased conversion of CPT-11 to SN-38 [1], [2], reduced sensitivity of topoisomerase (topo) I to CPT-11 [3], and decreased expression of topo I and/or topo II [4], [5], [6]. Cells selected for resistance to cisplatin [1], melphalan [7], mAMSA [8], or mitoxantrone [9] developed cross-resistance to CPT analogs.

In the present study, we found that cisplatin-resistant KCP-4 cells derived from human epidermoid carcinoma KB-3-1 cells are cross-resistant to CPT-11 and SN-38. The mechanism for the cross-resistance was examined. Our results suggest that an active efflux system for CPT-11 and SN-38 exists in KCP-4 cells.

Section snippets

Chemicals

CPT-11 and SN-38 were obtained from Daiichi Seiyaku (Tokyo, Japan). Minimal essential medium (MEM) was purchased from Nissui Seiyaku Co. (Tokyo, Japan). Newborn calf serum was from Cell Culture Laboratories (Cleveland, OH). 14,15,19,20-3H[N]Leukotriene C4 ([3H]LTC4) (110.5 Ci/mmol) was obtained from Du Pont NEN (Boston, MA). PAK-104P was obtained from Nissan Chemical Industries (Chiba, Japan). Cisplatin, BSO and other agents were obtained from Sigma Chemical Co. (St. Louis, MO).

Cell culture and cell lines.

The human

Cross-resistance to CPT-11 and SN-38 in KB sublines

As shown in Table 1, IC50 values for CPT-11 of KB-3-1 and KCP-4 cells were 3.5 and 43.4 μM, respectively. KCP-4 cells were 12.4-fold more resistant to CPT-11 than the parental KB-3-1 cells. IC50 values for SN-38 of KB-3-1 and KCP-4 cells were 0.008 and 0.253 μM, respectively. KCP-4 cells were 31.6-fold more resistant to SN-38 than the parental KB-3-1 cells. An incomplete revertant, KCP-4R, isolated from KCP-4 cells was 4.4-, 2.3- and 2.9-fold more resistant to cisplatin, CPT-11 and SN-38 than

Discussion

Cancer cells treated with a certain anticancer agent acquire cross-resistance to other structurally unrelated anticancer agents. We have established and characterized a cisplatin resistant KCP-4 cell line [11], [24], [25]. The KCP-4 cells were highly resistant to cisplatin and cross-resistant to carboplatin, 254-S, melphalan and methotrexate, but not resistant to adriamycin (ADM) and vincristine (VCR) [11]. In the present study, KCP-4 cells were cross-resistant to CPT-11 and SN-38, and we

Acknowledgements

This work was supported by grants from the Ministry of Education, Science and Culture, the Ministry of Health and Welfare, and Sasakawa Health Science Foundation, Japan. We thank Ms. Etuko Sudou for technical assistance and Ms. Hiromi Kakura for secretarial assistance.

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