Tarcomnicu Isabela, Iacob Simona, Anuta Valentina, Neaga Emil, Otelea Dan
National Institute for Infectious Diseases "Prof. Dr. Matei Bals", 021105 Bucharest, Romania.
Infectious Diseases Department, Faculty of Medicine, "Carol Davila" University of Medicine and Pharmacy, 020021 Bucharest, Romania.
Pharmaceuticals (Basel). 2025 Jun 16;18(6):899. doi: 10.3390/ph18060899.
: Sustained drug exposure is a key factor in the treatment of patients infected with human immunodeficiency virus (HIV) or hepatitis B virus (HBV) in order to achieve the intended virological response. Although influenced also by other parameters, adherence to the treatment scheme is the most important for adequate drug exposure. This can be assessed by therapeutic drug monitoring (TDM). Tenofovir (TFV) is a nucleotide analogue used in the treatment of both HIV and HBV. Although various analytical methods for the quantification of tenofovir prodrugs have been published, there is limited literature on methods for simultaneous TFV and its active metabolite, tenofovir diphosphate (TFVDP) direct determination. : In this study, we describe a novel micro-liquid-chromatography-mass spectrometry (micro-LC-MS/MS) method for TDM of TFV and TFVDP in biological matrices (whole blood, plasma). The challenging separation of the high-polarity analytes was resolved on an amino stationary phase, eluted in HILIC (hydrophilic interaction liquid chromatography) mode. The sample preparation included a clean-up step with hexane for the removal of lipophilic compounds and then protein precipitation with organic solvent. : The achieved low limits of quantification in blood were 0.25 ng/mL for TFV, and 0.5 ng/mL for TFVDP. Linearity, accuracy (91.63-109.18%), precision (2.48-14.08), and stability were validated for whole blood matrix, meeting the guidelines performance criteria. Samples collected from treated patients were analyzed, with results being in accordance with the reported pharmacokinetics. : The new method is adequate for analyzing samples in a clinical set-up. The measurement of both TFV and TFVDP improves clinical decision by an in-depth evaluation of long-term adherence, and together with viral load and resistance data helps guiding the treatment towards the intended virological suppression.
持续的药物暴露是治疗感染人类免疫缺陷病毒（HIV）或乙型肝炎病毒（HBV）患者以实现预期病毒学反应的关键因素。尽管还受其他参数影响，但坚持治疗方案对于实现足够的药物暴露最为重要。这可以通过治疗药物监测（TDM）来评估。替诺福韦（TFV）是一种用于治疗HIV和HBV的核苷酸类似物。尽管已发表了多种用于定量替诺福韦前药的分析方法，但关于同时直接测定TFV及其活性代谢物替诺福韦二磷酸（TFVDP）的方法的文献有限。
在本研究中，我们描述了一种用于生物基质（全血、血浆）中TFV和TFVDP的TDM的新型微液相色谱 - 质谱联用（micro-LC-MS/MS）方法。高极性分析物的挑战性分离在氨基固定相上得以解决，以亲水相互作用液相色谱（HILIC）模式洗脱。样品制备包括用己烷进行净化步骤以去除亲脂性化合物，然后用有机溶剂进行蛋白沉淀。
在血液中实现的低定量限为TFV 0.25 ng/mL，TFVDP 0.5 ng/mL。对全血基质的线性、准确度（91.63 - 109.18%）、精密度（2.48 - 14.08）和稳定性进行了验证，符合指南性能标准。对接受治疗患者采集的样本进行了分析，结果与报道的药代动力学一致。
新方法适用于临床环境中的样本分析。TFV和TFVDP的测定通过对长期依从性的深入评估改善临床决策，并与病毒载量和耐药性数据一起有助于指导治疗以实现预期的病毒学抑制。
