Doxorubicin induces expression of multidrug resistance-associated protein 1 in human small cell lung cancer cell lines by the c-jun N-terminal kinase pathway

Abstract

Multidrug resistance (MDR) is a major impediment to successful chemotherapy for lung cancer. Overexpression of multidrug resistance-associated protein 1 (MRP1) appears to be involved in MDR development in lung cancer cells. A number of chemotherapeutic agents including doxorubicin (DOX) were reported to induce MRP1 expression in human lung cancer cells. In our study, we investigated the mechanism by which DOX induces MRP1 expression in human small cell lung cancer (SCLC) cell lines, GLC4 and NCI-H82. These cells expressed MRP1 protein at low levels and were sensitive to DOX. Doxorubicin at 50 nM induced a marked increase in MRP1 expression in 24 hr, and stimulated c-jun N-terminal kinase (JNK) activity. Treatment with a JNK inhibitor, SP600125, significantly inhibited MRP1 induction. Furthermore, transfection with JNK1 and JNK2 antisense oligonucleotides markedly inhibited DOX-induced MRP1 expression. Chromatin immunoprecipitation assays revealed an enhanced recruitment of phosphorylated c-jun to the MRP1 promoter containing the AP-1 site upon DOX stimulation, which was inhibited by pretreatment with SP600125. Surprisingly, GLC4 cells exposed to DOX for 24 hr maintained increased MRP1 expression and resistance to DOX for at least 3 weeks. Pretreatment with SP600125 before DOX stimulation blocked the appearance of the MDR phenotype as well as MRP1 induction in GLC4 cells. These findings suggest that JNK activation may play an essential role for the induction of MRP1 protein in human SCLC cells by chemotherapeutic agents and that combined treatment of a JNK inhibitor with anticancer drugs may prevent the development of MDR by the abrogation of MRP1 induction. ' 2005 Wiley-Liss, Inc.Key words: MRP1; JNK; doxorubicin; small-cell lung cancer; multidrug resistance Small cell lung cancer (SCLC) accounts for 15-25% of all lung cancer patients.1,2 Although up to 90% of SCLC tumors initially respond to chemotherapy, patients with SCLC often relapse with multidrug resistant state. There are several types of multidrug resistance (MDR) to anticancer agents, and some of them were identified in human carcinoma cell lines in vitro. One of the mechanisms of MDR is decreased accumulation of drug within cells because of reduced inward transport or increased drug efflux, such as overexpression of adenosine triphosphate (ATP)-binding cassette (ABC) transporters. 3 P-glycoprotein (P-gp) that was cloned from KB-C 2.5 cells was the first ABC transporter identified. 4 Overexpression of P-gp has been detected in many multidrug resistant tumor cell lines and a variety of tumors from patients with both acquired and inherent drug resistance.5 Although P-gp is implicated in drug resistance in a number of tumor types, it is infrequently expressed in lung cancer cells. Another ABC transporter was cloned from the doxorubicin (DOX) resistant H69/AR SCLC cell line by Cole et al. 6 The transporter was designated as multidrug resistance-associated protein 1 (MRP1). The MRP family is now composed of 9 related ABC transporters that are able to transp...

Regulation of Multidrug Resistance Protein 1 by Tumor Necrosis Factor α in Cultured Glial Cells: Involvement of Nuclear Factor-κB and c-Jun N-Terminal Kinase Signaling Pathways

“…MAPK signaling cascades (i.e., JNK) are complex and may affect the expression of ABC transporter genes by the recruitment of transcription factors (i.e., AP-1, c-Jun) (Shinoda et al, 2005;Zhou et al, 2006;Hartz et al, 2008). Using chromatin immunoprecipitation, increased c-Jun binding to the MRP1 promoter was observed in human small-cell lung cancer cell lines treated with the anticancer drug doxorubicin (Shinoda et al, 2005). In addition, this study showed that SP600125 inhibited c-Jun binding, confirming the involvement of JNK signaling in MRP1 regulation.…”

“…Although conducted in cancerous human cell culture systems, the hypotheses proposed in this study can be tested in healthy rodent cell culture systems because similar signaling pathways and/or transcription factors are expressed in both models. Some of these similarities include expression of JNK/AP-1 (Nair et al, 2008), JNK/c-Jun (Shinoda et al, 2005), and Nrf2 (Song et al, 2009). Furthermore, human MRP1 and rat Mrp1 are both regulated by a highly conserved 100-nucleotide sequence in the promoter region, suggesting that regulatory mechanisms for both genes may be structurally and functionally similar (Muredda et al, 2003).…”

Portrait of multifaceted transporter, the multidrug resistance-associated protein 1 (MRP1/ABCC1)

“…Numerous chemotherapeutic agents, including doxorubicin [144] and vinblastine [141], as well as heavy metals such as arsenite, cadmium, and mercury [58,70], have been reported to induce MRP1 expression. In addition, reactive oxygen species (ROS) have been found to regulate the expression of MRP1.…”

“…One possibility is that these putative cis-acting elements mediate the oxidative stress-induced expression by interacting with redox-sensitive transcription factors. On the other hand, it has recently been demonstrated that doxorubicin upregulates the transcription of MRP1 gene via doxorubicin-activated c-jun N-terminal kinase (JNK), which enhances the association of the activated form of c-jun with the AP-1 site in the MRP1 promoter [144]. Moreover, stimulation of JNK activity has been observed upon depletion of intracellular GSH.…”

Loss of Kv and MaxiK currents associated with increased MRP1 expression in small cell lung carcinoma

“…Downregulation of these two channels as a consequence of drug resistance has not been previously reported. In contrast, drug resistance mediated by doxorubicin-induced increase in MRP1 expression is associated with increased c-jun N-terminal kinase activity in small-cell lung cancer cells (Shinoda et al, 2005) and increased phosphorylated cjun in H69AR as compared with H69 cells (Kurz et al, 2001). Overexpression of c-jun depresses expression of Kv channels in smooth muscle (Yu et al, 2001).…”