ABSTRACT
Nilotinib, a second-generation tyrosine kinase inhibitor (TKI), has been approved for first-line chronic myeloid leukemia (CML) treatment. The improved clinical response of nilotinib over that of the first generation TKI, imatinib, has been thought to be a result of its high potency of inhibition of BCR-ABL kinase. This study aimed to characterize differences between nilotinib and imatinib in the intracellular accumulation and cytotoxic effect on the CML cell line K562. Accumulation of nilotinib in K562 cells was from 4.7- to 9.0-fold higher than that of imatinib. The cytotoxic effect of nilotinib on K562 cells was 14.2-fold higher than that of imatinib. Inhibition experiments in K562 cells, and examination of the cellular uptake using influx transporter-transfected human embryonic kidney (HEK) 293 cells, suggested that the influx transporters OCT1 and OATP1A2, which have been reported to mediate accumulation of imatinib in CML cells, contributed little to the uptake of nilotinib. Nilotinib was found to accumulate in imatinib-resistant K562 (K562/IM) cells overexpressing the efflux transporter P-glycoprotein (P-gp), although cytotoxic assays showed that K562/IM cells displayed 20000-fold greater resistance to nilotinib over the parent K562 cells. In conclusion, the present findings suggest that intracellular accumulation of nilotinib in CML cells contributes to its clinical response and efficacy in CML patients. Although nilotinib has been reported to be effective against imatinib-resistant ABL kinase mutants, the drug could not overcome imatinib resistance acquired by P-gp-overexpression. These results imply that classification of mechanisms of drug resistance is important for suitable strategies to treat imatinib-resistant CML patients.
Subject(s)
ATP Binding Cassette Transporter, Subfamily B, Member 1/metabolism , Antineoplastic Agents/pharmacology , Benzamides/pharmacology , Piperazines/pharmacology , Protein Kinase Inhibitors/pharmacology , Pyrimidines/pharmacology , Biological Transport/drug effects , Cell Survival/drug effects , HEK293 Cells , Humans , Imatinib Mesylate , K562 Cells , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/drug therapy , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/metabolismABSTRACT
A precise and convenient high-performance liquid chromatography (HPLC) method has been established to assay nilotinib in human plasma. Chromatographic separation of nilotinib was performed on a LiChrosphere(®)100 RP-18(e) column (250 mm×4.0 mm, 5 µm) using a mixture of acetonitrile and 0.01 M phosphate buffer (pH 3.0) (42 : 58, v/v) under isocratic conditions at a flow rate of 1.0 ml/min with ultraviolet (UV) detection at 266 nm. The calibration curve showed linearity at concentrations between 250 ng/ml and 5000 ng/ml (r(2)>0.999). The mean±S.D. absolute recovery of nilotinib from plasma was 99.2±3.3%. The coefficients of variation of both intra- and inter-day precision were below 9.1%. These results indicate that this new HPLC-based quantification may be useful for therapeutic drug monitoring of nilotinib to help manage treatment in patients with chronic myeloid leukemia in clinical practice.
Subject(s)
Chromatography, High Pressure Liquid/methods , Protein Kinase Inhibitors/blood , Protein-Tyrosine Kinases/antagonists & inhibitors , Pyrimidines/blood , Humans , Spectrophotometry, UltravioletABSTRACT
This study explored the association of 14 single nucleotide polymorphisms in three genes coding for influx transporters (SLC22A1, SLCO1B1, and SLCO1B3), two genes coding for efflux transporters (ABCB1 and ABCG2), and four genes coding for enzymes (CYP2C9, CYP2C19, CYP2D6, and CYP3A5) with the pharmacokinetics of imatinib in Japanese patients with chronic myeloid leukemia. Pharmacokinetic parameters were estimated by a population pharmacokinetic analysis based on 622 plasma samples from 34 patients at steady state. Approximately 4.6-fold variability in individual clearance was observed (range, 3.4-15.5 L/hr). The individual estimated clearance was significantly increased in patients with the SLCO1B3 334GG genotype (median value ± standard deviation, 9.5 ± 3.1 L/hr; n = 19) compared with SLCO1B3 334TT and TG genotypes (7.0 ± 3.1 L/hr; n = 15) (P = 0.019). Patients with the ABCB1 3435CC genotype had significantly higher imatinib clearance (12.7 ± 3.0 L/hr; n = 7) compared with patients with ABCB1 3435CT and TT genotypes (7.9 ± 2.7 L/hr; n = 27) (P = 0.035). In conclusion, the present study suggests that single nucleotide polymorphisms of the influx transporter SLCO1B3 and the efflux transporter ABCB1 were functionally associated with individual variability of imatinib pharmacokinetics in Japanese patients with chronic myeloid leukemia.
Subject(s)
ATP Binding Cassette Transporter, Subfamily B, Member 1/genetics , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/drug therapy , Membrane Transport Proteins/genetics , Organic Anion Transporters, Sodium-Independent/genetics , Piperazines/pharmacokinetics , Polymorphism, Single Nucleotide , Protein Kinase Inhibitors/pharmacokinetics , Pyrimidines/pharmacokinetics , ATP Binding Cassette Transporter, Subfamily B , ATP Binding Cassette Transporter, Subfamily B, Member 2 , ATP-Binding Cassette Transporters/genetics , Adult , Aged , Benzamides , Cytochrome P-450 CYP3A/genetics , Cytochrome P-450 Enzyme System/genetics , Female , Genotype , Humans , Imatinib Mesylate , Leukemia, Myelogenous, Chronic, BCR-ABL Positive/genetics , Liver-Specific Organic Anion Transporter 1 , Male , Membrane Transport Proteins/metabolism , Middle Aged , Organic Anion Transporters/genetics , Organic Cation Transporter 1/genetics , Piperazines/blood , Piperazines/therapeutic use , Protein Kinase Inhibitors/blood , Protein Kinase Inhibitors/therapeutic use , Pyrimidines/blood , Pyrimidines/therapeutic use , Solute Carrier Organic Anion Transporter Family Member 1B3 , Young AdultABSTRACT
Intracellular concentration of imatinib in leukemic cells is thought to affect the clinical efficacy of this drug in patients with chronic myeloid leukemia (CML); however, there is no report that directly indicates the relationship between intracellular concentration and clinical outcome and/or, plasma concentration. In addition, the impacts of genetic variations of drug transporters, which mediate leukocyte concentration of imatinib, are unknown. In the present study, we investigated the correlation between intracellular imatinib concentrations in leukocytes, plasma imatinib levels, and genotypes of drug transporters, including ATP binding cassette B1 (ABCB), ABCG2, solute carrier 22A1 (SLC22A1), solute carrier organic anion transporter family members 1B1 (SLCO1B1) and SLCO1B3. The imatinib levels in leukocytes were determined using HPLC in 15 patients with chronic phase CML. No significant correlation between intracellular and plasma concentrations of imatinib was observed. The intracellular concentration was comparable in both patients with or without complete cytogenetic response. The intracellular imatinib concentration was significantly higher in patients with SLCO1B3 334TT than in those with 334TG/GG (p=0.0188). Plasma concentrations were similar in both SLCO1B3 genotypes (p=0.860), thereby resulting in the intracellular to plasma concentration ratio being higher in patients with SLCO1B3 334TT than those with 334 TG/GG (p=0.0502). These results suggested that the SLCO1B3 334T>G polymorphism could have a significant impact on the intracellular concentration of imatinib in leukocytes as a promising biomarker for personalized treatment of CML patients.
