Development and validation of a rapid and simple LC-MS/MS method for quantification of vemurafenib in human plasma: application to a human pharmacokinetic study.

BACKGROUND: Vemurafenib (Zelboraf) is a new tyrosine kinase inhibitor that selectively targets activated BRAF V600E gene and is indicated for the treatment of advanced BRAF mutation-positive melanoma. We developed a simple method for vemurafenib quantification using liquid chromatography-tandem mass spectrometry. A stability study of vemurafenib in human plasma was also performed. METHODS: (13)C(6)-vemurafenib was used as the internal standard. A single-step protein precipitation was used for plasma sample preparation. Chromatography was performed on an Acquity UPLC system (Waters) with chromatographic separation by the use of an Acquity UPLC BEH C18 column (2.1 × 50 mm, 1.7-mm particle size; Waters). Quantification was performed using the monitoring of multiple reactions of following transitions: m/z 488.2 → 381.0 for vemurafenib and m/z 494.2 → 387.0 for internal standard. RESULTS: This method was linear over the range from 1.0 to 100.0 mcg/mL. The lower limit of quantification was 0.1 mcg/mL for vemurafenib in plasma. Vemurafenib remained stable for 1 month at all levels tested, when stored indifferently at room temperature (20 °C), at +4 °C, or at -20 °C. This method was used successfully to perform a plasma pharmacokinetic study of vemurafenib in a patient after oral administration at a steady state. CONCLUSIONS: This liquid chromatography-tandem mass spectrometry method for vemurafenib quantification in human plasma is simple, rapid, specific, sensitive, accurate, precise, and reliable.

RAD51 haploinsufficiency causes congenital mirror movements in humans.

Congenital mirror movements (CMM) are characterized by involuntary movements of one side of the body that mirror intentional movements on the opposite side. CMM reflect dysfunctions and structural abnormalities of the motor network and are mainly inherited in an autosomal-dominant fashion. Recently, heterozygous mutations in DCC, the gene encoding the receptor for netrin 1 and involved in the guidance of developing axons toward the midline, have been identified but CMM are genetically heterogeneous. By combining genome-wide linkage analysis and exome sequencing, we identified heterozygous mutations introducing premature termination codons in RAD51 in two families with CMM. RAD51 mRNA was significantly downregulated in individuals with CMM resulting from the degradation of the mutated mRNA by nonsense-mediated decay. RAD51 was specifically present in the developing mouse cortex and, more particularly, in a subpopulation of corticospinal axons at the pyramidal decussation. The identification of mutations in RAD51, known for its key role in the repair of DNA double-strand breaks through homologous recombination, in individuals with CMM reveals a totally unexpected role of RAD51 in neurodevelopment. These findings open a new field of investigation for researchers attempting to unravel the molecular pathways underlying bimanual motor control in humans.