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Flavopiridol pharmacogenetics: clinical and functional evidence for the role of SLCO1B1/OATP1B1 in flavopiridol disposition.

Ni W, Ji J, Dai Z, Papp A, Johnson AJ, Ahn S, Farley KL, Lin TS, Dalton JT, Li X, Jarjoura D, Byrd JC, Sadee W, Grever MR, Phelps MA - PLoS ONE (2010)

Bottom Line: Polymorphisms in ABCC2, ABCG2, UGT1A1, UGT1A9, and SLCO1B1 were found to significantly correlate with flavopiridol PK in univariate analysis.To validate these findings, a second set of data with 51 patients was evaluated, and overall trends for associations between PK and PGx were found to be consistent.Polymorphisms in transport genes were found to be associated with flavopiridol disposition and outcomes.

View Article: PubMed Central - PubMed

Affiliation: Division of Pharmaceutics, College of Pharmacy, The Ohio State University, Columbus, Ohio, United States of America.

ABSTRACT

Background: Flavopiridol is a cyclin-dependent kinase inhibitor in phase II clinical development for treatment of various forms of cancer. When administered with a pharmacokinetically (PK)-directed dosing schedule, flavopiridol exhibited striking activity in patients with refractory chronic lymphocytic leukemia. This study aimed to evaluate pharmacogenetic factors associated with inter-individual variability in pharmacokinetics and outcomes associated with flavopiridol therapy.

Methodology/principal findings: Thirty-five patients who received single-agent flavopiridol via the PK-directed schedule were genotyped for 189 polymorphisms in genes encoding 56 drug metabolizing enzymes and transporters. Genotypes were evaluated in univariate and multivariate analyses as covariates in a population PK model. Transport of flavopiridol and its glucuronide metabolite was evaluated in uptake assays in HEK-293 and MDCK-II cells transiently transfected with SLCO1B1. Polymorphisms in ABCC2, ABCG2, UGT1A1, UGT1A9, and SLCO1B1 were found to significantly correlate with flavopiridol PK in univariate analysis. Transport assay results indicated both flavopiridol and flavopiridol-glucuronide are substrates of the SLCO1B1/OATP1B1 transporter. Covariates incorporated into the final population PK model included bilirubin, SLCO1B1 rs11045819 and ABCC2 rs8187710. Associations were also observed between genotype and response. To validate these findings, a second set of data with 51 patients was evaluated, and overall trends for associations between PK and PGx were found to be consistent.

Conclusions/significance: Polymorphisms in transport genes were found to be associated with flavopiridol disposition and outcomes. Observed clinical associations with SLCO1B1 were functionally validated indicating for the first time its relevance as a transporter of flavopiridol and its glucuronide metabolite. A second 51-patient dataset indicated similar trends between genotype in the SLCO1B1 and other candidate genes, thus providing support for these findings. Further study in larger patient populations will be necessary to fully characterize and validate the clinical impact of polymorphisms in SLCO1B1 and other transporter and metabolizing enzyme genes on outcomes from flavopiridol therapy.

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Related in: MedlinePlus

Uptake of flavopiridol, flavo-G, SN-38 and lenalidomide in OATP1B1 transfected cells.The bar graph indicates means + SD for triplicate determinations of 10 µM flavopiridol, flavo-G, SN-38 and lenalidomide uptake rates in cell lines (HEK-293 or MDCK-II) transfected with empty control vector or vector cloned with the OATP1B1 gene (SCLO1B1). Incubations with flavo-G were for 30 min., and all other drugs were for 10 min. Transport rates are expressed as percentages normalized to empty vector control. * p<.05, ** p<0.001,Student's t-test.
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pone-0013792-g003: Uptake of flavopiridol, flavo-G, SN-38 and lenalidomide in OATP1B1 transfected cells.The bar graph indicates means + SD for triplicate determinations of 10 µM flavopiridol, flavo-G, SN-38 and lenalidomide uptake rates in cell lines (HEK-293 or MDCK-II) transfected with empty control vector or vector cloned with the OATP1B1 gene (SCLO1B1). Incubations with flavo-G were for 30 min., and all other drugs were for 10 min. Transport rates are expressed as percentages normalized to empty vector control. * p<.05, ** p<0.001,Student's t-test.

Mentions: Although associations between PK and SNPs in ABCC2 may have been expected due to known in vitro interactions of MRP2 and flavopiridol, no such evidence existed prior to this study for the role of SLCO1B1/OATP1B1 in flavopiridol transport. To determine if the observed associations between flavopiridol PK and SLCO1B1 PGx were functionally relevant for flavopiridol disposition, we measured uptake of flavopiridol and flavo-G in cells transfected with SLCO1B1. Transfection efficiencies were estimated at approximately 60% using GFP-containing control vectors. Mean uptake velocities were 261±12 fmol/mg protein/10 min and 38±10 fmol/mg protein/30 min for flavopiridol and flavo-G, respectively, in MDCK-II cells. Flavopiridol transport rates in HEK-293 cells were approximately 2–3 fold higher than in MDCK-II cells suggesting its transport may be affected by the different membrane and transporter compositions in the two cell lines. Flavo-G transport rates were similar in both cell lines. Figure 3 shows normalized uptake velocities of flavopiridol and flavo-G in both HEK293 and MDCK-II cells transfected with either SLCO1B1 or empty vector. Expression of the transfected SLCO1B1 gene was verified with real-time PCR (data not shown). Functional expression of OATP1B1 was verified by evaluating uptake of a positive control substrate, SN-38 [41]. A second agent, lenalidomide, was used as a negative control substrate. Total intracellular accumulation and Initial transport velocities of SN-38, flavopiridol, and flavo-G were significantly increased in HEK293 and MDCK-II cells transiently transfected with SLCO1B1, compared to empty control vectors, whereas no increased uptake was shown for lenalidomide.


Flavopiridol pharmacogenetics: clinical and functional evidence for the role of SLCO1B1/OATP1B1 in flavopiridol disposition.

Ni W, Ji J, Dai Z, Papp A, Johnson AJ, Ahn S, Farley KL, Lin TS, Dalton JT, Li X, Jarjoura D, Byrd JC, Sadee W, Grever MR, Phelps MA - PLoS ONE (2010)

Uptake of flavopiridol, flavo-G, SN-38 and lenalidomide in OATP1B1 transfected cells.The bar graph indicates means + SD for triplicate determinations of 10 µM flavopiridol, flavo-G, SN-38 and lenalidomide uptake rates in cell lines (HEK-293 or MDCK-II) transfected with empty control vector or vector cloned with the OATP1B1 gene (SCLO1B1). Incubations with flavo-G were for 30 min., and all other drugs were for 10 min. Transport rates are expressed as percentages normalized to empty vector control. * p<.05, ** p<0.001,Student's t-test.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2967470&req=5

pone-0013792-g003: Uptake of flavopiridol, flavo-G, SN-38 and lenalidomide in OATP1B1 transfected cells.The bar graph indicates means + SD for triplicate determinations of 10 µM flavopiridol, flavo-G, SN-38 and lenalidomide uptake rates in cell lines (HEK-293 or MDCK-II) transfected with empty control vector or vector cloned with the OATP1B1 gene (SCLO1B1). Incubations with flavo-G were for 30 min., and all other drugs were for 10 min. Transport rates are expressed as percentages normalized to empty vector control. * p<.05, ** p<0.001,Student's t-test.
Mentions: Although associations between PK and SNPs in ABCC2 may have been expected due to known in vitro interactions of MRP2 and flavopiridol, no such evidence existed prior to this study for the role of SLCO1B1/OATP1B1 in flavopiridol transport. To determine if the observed associations between flavopiridol PK and SLCO1B1 PGx were functionally relevant for flavopiridol disposition, we measured uptake of flavopiridol and flavo-G in cells transfected with SLCO1B1. Transfection efficiencies were estimated at approximately 60% using GFP-containing control vectors. Mean uptake velocities were 261±12 fmol/mg protein/10 min and 38±10 fmol/mg protein/30 min for flavopiridol and flavo-G, respectively, in MDCK-II cells. Flavopiridol transport rates in HEK-293 cells were approximately 2–3 fold higher than in MDCK-II cells suggesting its transport may be affected by the different membrane and transporter compositions in the two cell lines. Flavo-G transport rates were similar in both cell lines. Figure 3 shows normalized uptake velocities of flavopiridol and flavo-G in both HEK293 and MDCK-II cells transfected with either SLCO1B1 or empty vector. Expression of the transfected SLCO1B1 gene was verified with real-time PCR (data not shown). Functional expression of OATP1B1 was verified by evaluating uptake of a positive control substrate, SN-38 [41]. A second agent, lenalidomide, was used as a negative control substrate. Total intracellular accumulation and Initial transport velocities of SN-38, flavopiridol, and flavo-G were significantly increased in HEK293 and MDCK-II cells transiently transfected with SLCO1B1, compared to empty control vectors, whereas no increased uptake was shown for lenalidomide.

Bottom Line: Polymorphisms in ABCC2, ABCG2, UGT1A1, UGT1A9, and SLCO1B1 were found to significantly correlate with flavopiridol PK in univariate analysis.To validate these findings, a second set of data with 51 patients was evaluated, and overall trends for associations between PK and PGx were found to be consistent.Polymorphisms in transport genes were found to be associated with flavopiridol disposition and outcomes.

View Article: PubMed Central - PubMed

Affiliation: Division of Pharmaceutics, College of Pharmacy, The Ohio State University, Columbus, Ohio, United States of America.

ABSTRACT

Background: Flavopiridol is a cyclin-dependent kinase inhibitor in phase II clinical development for treatment of various forms of cancer. When administered with a pharmacokinetically (PK)-directed dosing schedule, flavopiridol exhibited striking activity in patients with refractory chronic lymphocytic leukemia. This study aimed to evaluate pharmacogenetic factors associated with inter-individual variability in pharmacokinetics and outcomes associated with flavopiridol therapy.

Methodology/principal findings: Thirty-five patients who received single-agent flavopiridol via the PK-directed schedule were genotyped for 189 polymorphisms in genes encoding 56 drug metabolizing enzymes and transporters. Genotypes were evaluated in univariate and multivariate analyses as covariates in a population PK model. Transport of flavopiridol and its glucuronide metabolite was evaluated in uptake assays in HEK-293 and MDCK-II cells transiently transfected with SLCO1B1. Polymorphisms in ABCC2, ABCG2, UGT1A1, UGT1A9, and SLCO1B1 were found to significantly correlate with flavopiridol PK in univariate analysis. Transport assay results indicated both flavopiridol and flavopiridol-glucuronide are substrates of the SLCO1B1/OATP1B1 transporter. Covariates incorporated into the final population PK model included bilirubin, SLCO1B1 rs11045819 and ABCC2 rs8187710. Associations were also observed between genotype and response. To validate these findings, a second set of data with 51 patients was evaluated, and overall trends for associations between PK and PGx were found to be consistent.

Conclusions/significance: Polymorphisms in transport genes were found to be associated with flavopiridol disposition and outcomes. Observed clinical associations with SLCO1B1 were functionally validated indicating for the first time its relevance as a transporter of flavopiridol and its glucuronide metabolite. A second 51-patient dataset indicated similar trends between genotype in the SLCO1B1 and other candidate genes, thus providing support for these findings. Further study in larger patient populations will be necessary to fully characterize and validate the clinical impact of polymorphisms in SLCO1B1 and other transporter and metabolizing enzyme genes on outcomes from flavopiridol therapy.

Show MeSH
Related in: MedlinePlus