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. Author manuscript; available in PMC: 2022 Mar 1.
Published in final edited form as: J Thromb Haemost. 2021 Jan 22;19(3):689–700. doi: 10.1111/jth.15209

Figure 2. Characterization of VK-M-COT as an inhibitor for VKD carboxylation in HEK293 reporter cells.

Figure 2

(A) Inhibition of VKD carboxylation by VK-M-COT (ο) and warfarin (◻) in HEK293 cells. FIXgla-PC/HEK293 cells were cultured with increasing concentrations of VK-M-COT or warfarin in cell culture medium containing 5 μM KO for the cell-based activity assay, as described in the legend of Figure 1C. (B) Inhibition potency of warfarin and VK-M-COT for VKD carboxylation. Half-maximal inhibition concentrations (IC50) of VK-M-COT and warfarin on VKD carboxylation were determined from Figure 2A using GraphPad Prism 8. (C) The vitamin K redox cycle. When VKD proteins are carboxylated by GGCX, the cofactor reduced vitamin K (KH2) is oxidized to vitamin K epoxide (KO), which needs to be reduced back to KH2 by a two-step reduction of KO to vitamin K and vitamin K to KH2. (D) Inhibition of VKD carboxylation by VK-M-COT in HEK293 and DGKO reporter cells. FIXgla-PC/HEK293 or DGKO reporter cells were cultured with increasing concentrations of VK-M-COT in cell culture medium containing 5 μM KO (• KOHEK293) or vitamin K1 (♦ KDGKO), respectively, for the cell-based activity assay and cell viability assays (■ viability).