Induction of MDR1 gene expression by anthracycline analogues in a human drug resistant leukaemia cell line

Abstract

SummaryThe effects of 4-demethoxydaunorubicin (idarubicin, IDA) and MX2, a new morpholino-anthracycline, on up-regulation of the MDR1 gene in the low-level multidrug resistant (MDR) cell line CEM/A7R were compared at similar concentrations (IC 10 , IC 50 and IC 90 ) over a short time exposure (4 and 24 h). The chemosensitivity of each drug was determined by a 3-day cell growth inhibition assay. Compared with epirubicin (EPI), IDA and MX2 were 17-and eightfold more effective in the CEM/A7R line respectively. No cross-resistance to 5-FU was seen in the CEM/A7R line. Verapamil (5 µM) and PSC 833 (1 µM), which dramatically reversed resistance to EPI in the CEM/A7R line, had no sensitizing effect on the resistance of this line to MX2, but slightly decreased resistance to IDA. The sensitivity to 5-FU was unchanged by these modulators. The induction of MDR1 mRNA expression by IDA, MX2 and 5-FU was analysed by Northern blotting and semiquantitatively assessed by scanning Northern blots on a phosphorimager. The relative level of MDR1 expression was expressed as a ratio of MDR1 mRNA to the internal RNA control glyceraldehyde-3-phosphate dehydrogenase (GAPDH). IDA, MX2 and 5-FU differentially up-regulated MDR1 mRNA in the CEM/A7R line in a dose-dependent manner. Both IDA and MX2 induced MDR1 expression within 4 h. 5-FU up-regulated MDR1 expression only when drug exposure was prolonged to 24 h. Based on MRK 16 binding, flow cytometric analysis of P-glycoprotein (Pgp) expression paralleled the increase in MDR1 mRNA levels. For the three anthracyclines, the increase in MDR1 expression was stable in cells grown in the absence of drug for more than 3 weeks after drug treatment. The induction of MDR1 expression by 5-FU was transient, associated with a rapid decrease in the increased Pgp levels which returned to baseline 72 h after the removal of 5-FU. This study demonstrates that MDR1 expression can be induced by analogues of anthracyclies not pumped by Pgp, and that this induction appears to be stable despite a 3-week drug-free period.

Anti-Cripto Mab inhibit tumour growth and overcome MDR in a human leukaemia MDR cell line by inhibition of Akt and activation of JNK/SAPK and bad death pathways

“…A MDR variant CEM/A7R cell line, developed by DOX selection of T-cell lymphoblastic leukaemia CCRF-CEM cell line (Zalcberg et al, 1994), has been successfully used as a model to study the molecular mechanisms underlying regulation of drug resistance. A rapid upregulation of MDR1 gene in CEM/A7R cell line was induced by anthracyclines and its analogues (Hu et al, 1995(Hu et al, , 1999b, and prevented by CyA and its analogue PSC 833 (Hu et al, 1996). These results represent the clinical development of Pgp-mediated drug resistance during chemotherapy (Hu et al, 1999a).…”

Epigenetic changes to the MDR1 locus in response to chemotherapeutic drugs

“…Evidence from cell lines and in vivo studies of patients undergoing chemotherapy demonstrate that MDR1 can be rapidly upregulated following exposure to chemotherapeutic drugs (Chaudhary and Roninson, 1993;Hu et al, 1995Hu et al, , 1999aAbolhoda et al, 1999), supporting the activation model. For example, ex vivo treatment of AML blasts isolated from newly diagnosed patients prior to induction chemotherapy resulted in the rapid upregulation of MDR1 expression, concomitant with increases in Pgp expression within 16 h of treatment, and in one AML patient increased Pgp expression in vivo was observed after only 4 h of chemotherapy (Hu et al, 1999a).…”

“…For example, ex vivo treatment of AML blasts isolated from newly diagnosed patients prior to induction chemotherapy resulted in the rapid upregulation of MDR1 expression, concomitant with increases in Pgp expression within 16 h of treatment, and in one AML patient increased Pgp expression in vivo was observed after only 4 h of chemotherapy (Hu et al, 1999a). Furthermore, cells isolated from five patients undergoing lung perfusion with doxorubicin were shown to increase in MDR1 expression within as little as 50 min after treatment commencement (Abolhoda et al, 1999).…”

“…In the first model 'selection and expansion', proposed almost two decades ago, tumors overexpressing Pgp develop following the drug-induced selection of a small population of Pgp high cells that subsequently becomes expanded and eventually overgrows the sensitive Pgp Àve /Pgp low cell population (Goldie and Coldman, 1984). Alternatively, according to the 'activation model', chemotherapeutic drugs can directly activate the multidrug resistance 1 (MDR1) gene, leading to Pgp overexpression and drug resistance (Abolhoda et al, 1999;Hu et al, 1999a). More recently, drug-induced increases in MDR1 expression in leukemic cell lines have also been attributed to increases in mRNA stability (Yague et al, 2003).…”

Ecteinascidin 743, a transcription-targeted chemotherapeutic that inhibits MDR1 activation

“…Our laboratory has shown that, in five of five patients with metastatic sarcoma, tumor MDR1 RNA levels increased up to 10-fold within 50 min of exposure to the DNA-damaging agent doxorubicin (5). In a separate study, an increase in Pgp expression was observed in a patient with acute myeloid leukemia after 4 and 12 h of administration of daunorubicin͞AraC (19). Thus, inducible expression of MDR1 by toxic agents such as chemotherapy and radiation may play a major role in the development of clinical drug resistance, and ET-743 is the first agent identified with the potential for blocking this activation.…”