Puhlmann Ulrike, Ziemann Christina, Ruedell Gudrun, Vorwerk Hagen, Schaefer Dirk, Langebrake Claudia, Schuermann Peter, Creutzig Ursula, Reinhardt Dirk
AML-BFM Study, Department of Pediatric Hematology and Oncology, University Children's Hospital Muenster, Albert-Schweitzer-Str. 33, 48129 Muenster, Germany.
J Pharmacol Exp Ther. 2005 Jan;312(1):346-54. doi: 10.1124/jpet.104.071571. Epub 2004 Oct 22.
Multidrug resistance (MDR), a challenge in treating childhood acute myeloid leukemia (AML), is frequently associated with decreased drug accumulation caused by multidrug transporter MDR1. Doxorubicin, an important anti-AML drug, is a known MDR1 substrate and inducer. Its cytostatic efficacy is thus limited by MDR1 overexpression. A recent study demonstrated cyclooxygenase-2-dependent, prostaglandin E(2) (PGE(2))-mediated regulation of mdr1b expression in primary rat hepatocyte cultures. Cyclooxygenase-2 expression is increased in several malignancies and considered a negative prognostic factor. Our study focused on cyclooxygenase system's impact on drug-induced MDR1 overexpression in AML cells. As a prerequisite, coexpression of MDR1 and cyclooxygenase-2 mRNA in HL-60 cells and primary AML blasts was demonstrated by Northern blot. Interestingly, incubation of AML cells with doxorubicin not only induced functionally active MDR1 overexpression but also mediated increased cyclooxygenase-2 mRNA and protein expressions with subsequent PGE(2) release (determined by flow cytometry, rhodamine123 efflux assay, reverse transcription-polymerase chain reaction, and enzyme-linked immunosorbent assay). After preincubation and subsequent parallel treatment with the cyclooxygenase-2-preferential inhibitor meloxicam, doxorubicin-induced MDR1 overexpression and function were reduced (maximally at 0.1-0.5 microM meloxicam), whereas cytostatic efficacy of doxorubicin in 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide assays was significantly increased by up to 78 (HL-60) and 30% (AML blasts) after 72 h of doxorubicin treatment. In HL-60 cells, meloxicam-dependent effect on doxorubicin cytotoxicity was neutralized by PGE(2) preincubation. In conclusion, the cyclooxygenase system, especially the cyclooxygenase-2 isoform, might be involved in regulating doxorubicin-induced MDR1 overexpression in AML cells, with PGE(2) seeming to be a mediating factor. Cyclooxygenase inhibitors thus bear promise to overcome MDR in AML and improve therapy.
多药耐药性（MDR）是儿童急性髓系白血病（AML）治疗中的一项挑战，它常常与多药转运蛋白MDR1导致的药物蓄积减少有关。阿霉素是一种重要的抗AML药物，是已知的MDR1底物和诱导剂。因此，其细胞生长抑制功效受到MDR1过表达的限制。最近一项研究表明，在原代大鼠肝细胞培养物中，环氧化酶-2依赖性的前列腺素E2（PGE2）介导了mdr1b表达的调控。环氧化酶-2在多种恶性肿瘤中表达增加，被认为是一个负性预后因素。我们的研究聚焦于环氧化酶系统对AML细胞中药物诱导的MDR1过表达的影响。作为前提条件，通过Northern印迹法证实了HL-60细胞和原代AML母细胞中MDR1和环氧化酶-2 mRNA的共表达。有趣的是，用阿霉素孵育AML细胞不仅诱导了具有功能活性的MDR1过表达，还介导了环氧化酶-2 mRNA和蛋白表达增加，随后释放PGE2（通过流式细胞术、罗丹明123外排试验、逆转录-聚合酶链反应和酶联免疫吸附试验测定）。在用环氧化酶-2选择性抑制剂美洛昔康进行预孵育并随后进行平行处理后，阿霉素诱导的MDR1过表达和功能降低（在美洛昔康浓度为0.1 - 0.5 microM时最大），而在阿霉素处理72小时后，阿霉素在3-（4,5-二甲基噻唑-2-基）-2,5-二苯基-2H-四氮唑溴盐试验中的细胞生长抑制功效显著增加，在HL-60细胞中增加高达78%，在AML母细胞中增加30%。在HL-60细胞中，PGE2预孵育可中和美洛昔康对阿霉素细胞毒性的依赖性作用。总之，环氧化酶系统，尤其是环氧化酶-2亚型，可能参与调控AML细胞中阿霉素诱导的MDR1过表达，PGE2似乎是一个介导因子。因此，环氧化酶抑制剂有望克服AML中的MDR并改善治疗。
