Sorafenib Uptake Research Articles

Abstract 4425: Characterisation of cellular transport and accumulation of six clinically approved tyrosine kinase inhibitors (TKIs) in colon cancer cells.

Abstract TKIs are a loosely characterised family of small molecules which inhibit the ATP driven phosphorylation of signalling proteins that normally activate transduction cascades. Aberrations in signal cascades have been linked to the development of tumours and their survival pathways; TKIs are targeted at these pathways. However, promising preclinical studies on structurally different TKIs often did not translate into clinical success due to cellular resistance, cascade mutations and poor bioavailability. TKIs tend to be alkali in nature with a correspondingly high pKa (5-8) while absorption properties have been linked to both ABC and Organic Cation Transporters (OCT). Previously we demonstrated in a gut epithelium model that absorption of TKIs is relatively poor, while a correspondingly high negative flow was observed. In this study we aimed to characterize tumour cell uptake and accumulation of TKIs using specific transporter inhibitors. We used a sensitive and specific LC-MS-MS method to characterize the role of the transport inhibitors Ko143 (BCRP inhibitor), amantadine (Am;hOCT1&2), cimetidine (Ci;hOCT1&2&3), desipramine (Ds;hOCT1&2&3), β-estradiol (Be; hOCT1&3) and verapamil (Ve; hOCT1&P-gp). Uptake of TKIs at 4°C and 37°C in CaCo2 and WiDr colon cancer cells showed large differences; uptake of Gefitinib (G - 10μM), Erlotinib (E - 10 μM) & Dasatinib (D - 1μM) was partially active since after 2hr uptake at 4°C was lower compared to 37°C. Uptake of Sunitinib (Su - 10μM), Sorafenib (So - 10 μM) and Crizotinib (Cr - 10 μM) was predominantly passive since values were comparable at 4°C and 37°C. Absolute accumulation over 24hr (37°C) was very different with E and D reaching 20-150 ng/mg protein, while G was 20 fold higher. Su, So and Cr accumulation were even higher reaching 2 to 12 μg/mg protein. This pattern matched accumulation of some TKIs observed in the clinical setting. Accumulation for Su and Cr showed a similar pattern suggesting that Cr accumulated in lysosomes similar to Su. Bafilomycin, a lysosomal inhibitor, decreased total accumulation of Su 14.1-fold, and that of Cr 2.4-fold indicating lysosomal accumulation. The hOCT inhibitor Ds consistently (25-55%) decreased accumulation of G, So Su and Cr; Ci, Be and Am had little effect on cellular accumulation for all TKIs tested. Ve, De and Ci increased accumulation of E, and Ve that of D, suggesting inhibition of efflux via P-gp, MRP or BCRP. Ko143 increased accumulation for G, E, So and Su in WiDr but only for Su in CaCo2 cells indicating a differential role for BCRP depending on TKI and tumour type. In conclusion, several transporters appear to be involved to differing degrees for each molecule, indicating TKIs pharmacokinetics should be investigated on an individual basis. Citation Format: Richard J. Honeywell, Sarina Hitzerd, Ietje Kathmann, Elisa Giovannetti, Henk Verheul, Frits Peters. Characterisation of cellular transport and accumulation of six clinically approved tyrosine kinase inhibitors (TKIs) in colon cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4425. doi:10.1158/1538-7445.AM2013-4425

Sorafenib Hepatobiliary Disposition: Mechanisms of Hepatic Uptake and Disposition of Generated Metabolites

Sorafenib is an orally active tyrosine kinase inhibitor used in the treatment of renal and hepatocellular carcinoma. This study was designed to establish whether transport proteins are involved in the hepatic uptake of sorafenib and to determine the extent of biliary excretion of sorafenib and its metabolites in human hepatocytes. Initial uptake was assessed in freshly isolated, suspended human hepatocytes in the presence of inhibitors and modulators. [(14)C]Sorafenib (1 µM) uptake at 4°C was reduced by about 61-63% of the uptake at 37°C, suggesting a high degree of passive diffusion. Hepatocyte uptake of [(14)C]sorafenib was not Na(+) dependent or influenced by the organic anion transporter 2 inhibitor ketoprofen. However, initial [(14)C]sorafenib hepatocyte uptake was reduced by 46 and 30% compared with control values in the presence of the organic anion transporting polypeptide inhibitor rifamycin SV and the organic cation transporter (OCT) inhibitor decynium 22, respectively. [(14)C]Sorafenib (0.5-5 µM) uptake was significantly higher in hOCT1-transfected Chinese hamster ovary cells compared with mock cells, and inhibited by the general OCT inhibitor, 1-methyl-4-phenylpryidinium. OCT1-mediated uptake was saturable with a Michaelis-Menten constant of 3.80 ± 2.53 µM and a V(max) of 116 ± 42 pmol/mg/min. The biliary excretion index and in vitro biliary clearance of sorafenib (1 µM) in sandwich-cultured human hepatocytes were low (∼11% and 11 ml/min/kg, respectively). Results suggest that sorafenib uptake in human hepatocytes occurs via passive diffusion, by OCT1, and by organic anion transporting polypeptide(s). Sorafenib undergoes modest biliary excretion, predominantly as a glucuronide conjugate(s).

Interaction of the Multikinase Inhibitors Sorafenib and Sunitinib with Solute Carriers and ATP-Binding Cassette Transporters

To compare side-by-side the uptake of sorafenib and sunitinib in vitro by human uptake solute carriers of the SLC22A and SLCO families, the transport by and inhibition of efflux ATP-binding cassette (ABC) transporters, and the role of ABCB1 in the plasma pharmacokinetics and brain penetration of these agents. Uptake of [(3)H]sorafenib or [(3)H]sunitinib was assessed in Xenopus laevis oocytes or mammalian cells transfected with cDNAs coding for human OATP1A2, OATP1B1, OATP1B3, OCT1, OAT2, OAT3, OCTN1, or OCTN2. Efflux and inhibition experiments were conducted in cells transfected with human ABCB1, ABCG2, ABCC2, or ABCC4. In vivo pharmacokinetic studies were done in knockout mice lacking Abcb1-type transporters. Intracellular uptake was not appreciably affected by any of the studied solute carriers and was minute relative to the respective prototypical substrates. Sorafenib and sunitinib showed concentration-dependent (1 and 10 micromol/L), low to moderate affinity for ABCB1 but were not affected by the other ABC transporters. Both agents inhibited all tested ABC transporters. The absence of Abcb1 had no affect on plasma pharmacokinetics, but brain penetration was moderately increased by 1.9- and 2.9-fold for sorafenib and sunitinib, respectively, in knockout animals versus controls. Unlike other tyrosine kinase inhibitors, sorafenib and sunitinib do not appear to rely on active transport to enter the cell nor are they high-affinity substrates for ABC efflux transporters. Based on these characteristics, these two drugs may be less susceptible to transporter-mediated alterations in systemic exposure and transporter-related resistance mechanisms.