Objective:To establish an LC-MS/MS and HPLC assay for the determination of linezolid in human plasma to be used for the therapeutic drug monitoring ( TDM) and pharmacokinetic study. Methods:Acetontrile containing furazolidone ( internal stand-ard) as the protein precipitation agent was added to100 μl human plasma, and then vibrated and centrifuged for the precipitation of plasma protein. ① The supernatant was eluted on an Eclipse XDB-C18 (100mm × 2. 1mm,3. 5μm) column with acetontrile and water (80 :20) as the mobile phase at the flow rate of 0. 3 ml·min-1. The electrospray ionization (ESI) source was applied and operated in the positive ion mode. The multiple reaction monitoring (MRM) modes with the transition of m/z338. 1→296. 2 (linezolid) and m/z226.1→122.0 (furazolidone) were used for the quantification. ② The supernatant was eluted on an Eclipse Eclipse XDB -C18(250 mm × 4. 6 mm, 5μm) column with acetontrile and 0. 1% formic acid (20 :80) as the mobile phase at the flow rate of 1. 0 ml·min-1 and detected at 254 nm. The established assays were used for the determination of linezolid in the plasma samples after the administra-tion. Results:Linezolid was linear within the range of 0. 05-30 μg·ml-1 for LC-MS/MS, and 0. 25-30 μg·ml-1 for HPLC ( r2 >0. 999). The extraction recovery and the matrix effect respectively was 82. 1%-91. 3% and 74. 0%-82. 3%. The relative recovery of LC-MS/MS and HPLC was 91. 2%-106. 4% and 100. 1%-111. 6%, respectively. The intra-and inter-day RSDs were both lower than 20%. There was a good correlation between LC-MS/MS and HPLC. The trough concentration of 12 patients was (1. 77 ± 1. 23) g· ml-1 and the plasma concentration of 5 patients 2h after linezolid adminstration was (13. 36 ± 2. 63) g·ml-1 , respectively. Conclu-sion:The established assays are simple, rapid, specific, sensitive and accurate, which are suitable for the TDM and pharmacokinetic study of linezolid.

Objective:To establish an LC-MS/MS and HPLC assay for the determination of linezolid in human plasma to be used for the therapeutic drug monitoring ( TDM) and pharmacokinetic study. Methods:Acetontrile containing furazolidone ( internal stand-ard) as the protein precipitation agent was added to100 μl human plasma, and then vibrated and centrifuged for the precipitation of plasma protein. ① The supernatant was eluted on an Eclipse XDB-C18 (100mm × 2. 1mm,3. 5μm) column with acetontrile and water (80 :20) as the mobile phase at the flow rate of 0. 3 ml·min-1. The electrospray ionization (ESI) source was applied and operated in the positive ion mode. The multiple reaction monitoring (MRM) modes with the transition of m/z338. 1→296. 2 (linezolid) and m/z226.1→122.0 (furazolidone) were used for the quantification. ② The supernatant was eluted on an Eclipse Eclipse XDB -C18(250 mm × 4. 6 mm, 5μm) column with acetontrile and 0. 1% formic acid (20 :80) as the mobile phase at the flow rate of 1. 0 ml·min-1 and detected at 254 nm. The established assays were used for the determination of linezolid in the plasma samples after the administra-tion. Results:Linezolid was linear within the range of 0. 05-30 μg·ml-1 for LC-MS/MS, and 0. 25-30 μg·ml-1 for HPLC ( r2 >0. 999). The extraction recovery and the matrix effect respectively was 82. 1%-91. 3% and 74. 0%-82. 3%. The relative recovery of LC-MS/MS and HPLC was 91. 2%-106. 4% and 100. 1%-111. 6%, respectively. The intra-and inter-day RSDs were both lower than 20%. There was a good correlation between LC-MS/MS and HPLC. The trough concentration of 12 patients was (1. 77 ± 1. 23) g· ml-1 and the plasma concentration of 5 patients 2h after linezolid adminstration was (13. 36 ± 2. 63) g·ml-1 , respectively. Conclu-sion:The established assays are simple, rapid, specific, sensitive and accurate, which are suitable for the TDM and pharmacokinetic study of linezolid.

Metformin is currently a strong candidate anti-tumor agent in multiple cancers. However, its anti-tumor effectiveness varies among different cancers or subpopulations, potentially due to tumor heterogeneity. It thus remains unclear which hepatocellular carcinoma (HCC) patient subpopulation(s) can benefit from metformin treatment. Here, through a genome-wide CRISPR-Cas9-based knockout screen, we find that DOCK1 levels determine the anti-tumor effects of metformin and that DOCK1 is a synthetic lethal target of metformin in HCC. Mechanistically, metformin promotes DOCK1 phosphorylation, which activates RAC1 to facilitate cell survival, leading to metformin resistance. The DOCK1-selective inhibitor, TBOPP, potentiates anti-tumor activity by metformin in vitro in liver cancer cell lines and patient-derived HCC organoids, and in vivo in xenografted liver cancer cells and immunocompetent mouse liver cancer models. Notably, metformin improves overall survival of HCC patients with low DOCK1 levels but not among patients with high DOCK1 expression. This study shows that metformin effectiveness depends on DOCK1 levels and that combining metformin with DOCK1 inhibition may provide a promising personalized therapeutic strategy for metformin-resistant HCC patients.
Animals ; Antineoplastic Agents/therapeutic use* ; Carcinoma, Hepatocellular/metabolism* ; Cell Line, Tumor ; Clustered Regularly Interspaced Short Palindromic Repeats ; Genome ; Humans ; Liver Neoplasms/metabolism* ; Metformin/therapeutic use* ; Mice ; Phosphorylation ; Synthetic Lethal Mutations ; Transcription Factors/metabolism* ; rac GTP-Binding Proteins/metabolism*