title: Influx and efflux transport as determinants of melphalan cytotoxicity: Resistance to melphalan in MDR1 overexpressing tumor cell lines.
abstract: There is a considerable variation in efficacy of melphalan therapy in multiple myeloma (MM) and other hematopoietic tumors. We hypothesized that this may be due to variations in the expression of influx and efflux transporters of melphalan. We measured the expression of the influx transporters LAT1, LAT2, and TAT1 and the efflux transporters MDR1, MRP1 and BCRP by quantitative RT-PCR and related their expression to the intracellular accumulation and cytotoxicity of melphalan in 7 MM and 21 non-MM hematopoietic tumor cell lines. Variation in the intracellular accumulation accounted for nearly half of the variation in the cytotoxicity of melphalan in MM cell lines (r(2)=0.47, P=0.04). High expression of the efflux transporter MDR1 was associated with low intracellular accumulation and low cytotoxicity of melphalan (r(2)=0.56, P=0.03 and r(2)=0.62, P=0.02, respectively). The effect was reversed by the MDR1 inhibitor cyclosporine. In addition, the MDR1 overexpressing HL-60 cell line showed 10-fold higher resistance to melphalan than the non-MDR1 expressing one. Again, the resistance was reversed by cyclosporine and by MDR1-specific shRNA. LAT1 was the major influx transporter in tumor cell lines with 4000-fold higher expression than LAT2. Down-regulation of LAT1 by siRNA reduced the melphalan uptake by 58% and toxicity by 3.5-fold, but natural variation in expression between the tumor cell lines was not associated with accumulation or cytotoxicity of melphalan. In conclusion, tumor-specific variations in the expression of the efflux transporter MDR1, but not of the influx transporter LAT1, affect the intracellular accumulation of melphalan and thus determine its cytotoxicity.
authors.affiliation: Department of Clinical Pharmacology, University Medical Center, University of Göttingen, Robert-Koch-Str. 40, D-37075 Göttingen, Germany.
authors.forename: Annett
authors.initials: A
authors.lastname: Kühne
authors.forename: Mladen Vassilev
authors.initials: MV
authors.lastname: Tzvetkov
authors.forename: Yohannes
authors.initials: Y
authors.lastname: Hagos
authors.forename: Hermann
authors.initials: H
authors.lastname: Lage
authors.forename: Gerhard
authors.initials: G
authors.lastname: Burckhardt
authors.forename: Jürgen
authors.initials: J
authors.lastname: Brockmöller
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000187:drug effects / Q000235:genetics*; D000818:Animals; D018906:Antineoplastic Agents, Alkylating / Q000493:pharmacokinetics / Q000633:toxicity; D001692:Biological Transport; D045744:Cell Line, Tumor / Q000187:drug effects; D003131:Combined Modality Therapy; D019008:Drug Resistance, Neoplasm / Q000187:drug effects; D006801:Humans; D008558:Melphalan / Q000493:pharmacokinetics* / Q000633:toxicity*; D009101:Multiple Myeloma / Q000188:drug therapy / Q000235:genetics / Q000601:surgery; D012334:RNA, Neoplasm / Q000235:genetics; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D033581:Stem Cell Transplantation
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D018906:Antineoplastic Agents, Alkylating; D012334:RNA, Neoplasm; D008558:Melphalan