Rower Joseph E, Jimmerson Leah C, Chen Xinhui, Zheng Jia-Hua, Hodara Ariel, Bushman Lane R, Anderson Peter L, Kiser Jennifer J
University of Colorado, Skaggs School of Pharmacy and Pharmaceutical Sciences, Aurora, Colorado, USA.
University of Colorado, Skaggs School of Pharmacy and Pharmaceutical Sciences, Aurora, Colorado, USA
Antimicrob Agents Chemother. 2015 Dec;59(12):7671-9. doi: 10.1128/AAC.01693-15. Epub 2015 Sep 28.
Sofosbuvir (SOF) is a highly efficacious and well-tolerated uridine nucleotide analog that inhibits the hepatitis C virus (HCV) NS5B polymerase enzyme. SOF is administered as a prodrug, which undergoes intracellular phosphorylation by host enzymes to a monophosphate, diphosphate, and finally a pharmacologically active triphosphate. In order to fully understand the clinical pharmacology of SOF, there is a great need to determine the intracellular phosphate concentrations of the drug. We describe the validation and utilization of a method to characterize SOF's disposition into various in vivo cell types, including hepatocytes, peripheral blood mononuclear cells (PBMC), and red blood cells (RBC). Standard bioanalytical validation criteria were applied to lysed cellular matrices, with a validated linear range of 50 to 50,000 fmol/sample for each phosphate moiety. The assay was utilized to collect the first data demonstrating concentrations of phosphorylated anabolites formed in PBMC, hepatocytes, and RBC in vivo during SOF therapy. Median concentrations in PBMC were 220 (range, 51.5 to 846), 70.2 (range, 25.8 to 275), and 859 (range, 54.5 to 6,756) fmol/10(6) cells in the monophosphate, diphosphate, and triphosphate fractions, respectively. In contrast, RBC triphosphate concentrations were much lower than those of PBMC, as the median concentration was 2.91 (range, 1.14 to 10.4) fmol/10(6) cells. The PBMC triphosphate half-life was estimated at 26 h using noncompartmental approaches, while nonlinear mixed-effect modeling was used to estimate a 69 h half-life for this moiety in RBC. The validated method and the data it generates provide novel insight into the cellular disposition of SOF and its phosphorylated anabolites in vivo.
索磷布韦（SOF）是一种高效且耐受性良好的尿苷核苷酸类似物，可抑制丙型肝炎病毒（HCV）NS5B聚合酶。SOF作为前药给药，通过宿主酶在细胞内磷酸化生成一磷酸、二磷酸，最终生成具有药理活性的三磷酸。为了全面了解SOF的临床药理学，非常需要确定该药物在细胞内的磷酸盐浓度。我们描述了一种用于表征SOF在各种体内细胞类型（包括肝细胞、外周血单核细胞（PBMC）和红细胞（RBC））中分布情况的方法的验证和应用。将标准生物分析验证标准应用于裂解的细胞基质，每个磷酸盐部分的验证线性范围为50至50,000 fmol/样品。该测定法用于收集首批数据，证明在SOF治疗期间体内PBMC、肝细胞和RBC中形成的磷酸化代谢产物的浓度。PBMC中一磷酸、二磷酸和三磷酸部分的中位浓度分别为220（范围，51.5至846）、70.2（范围，25.8至275）和859（范围，54.5至6,756）fmol/10(6)细胞。相比之下，RBC三磷酸浓度远低于PBMC，中位浓度为2.91（范围，1.14至10.4）fmol/10(6)细胞。使用非房室方法估计PBMC三磷酸半衰期为26小时，而使用非线性混合效应模型估计该部分在RBC中的半衰期为69小时。经过验证的方法及其产生的数据为SOF及其磷酸化代谢产物在体内的细胞分布提供了新的见解。
