In differentiated HepaRG cells, 24-hour lomitapide exposure did not produce lipid accumulation above the imaging assay threshold. The highest tested concentration (12 µM) reduced the mitochondrial membrane-potential signal; apoptotic nuclear morphology increased, while no oxidative-stress change was reported. The negative lipid result is limited to this short-duration assay. [2]
Inhibition of purified human MTP lipid-transfer activity
Biochemical assay · Purified FLAG-tagged human MTP expressed in Cos-7 cells; fluorescent donor/acceptor vesicle assays [1]
Lomitapide inhibited both phospholipid (PL) and triacylglycerol (TAG) transfer, with a reported IC50 of 10 nM for each. This is biochemical potency under the reported assay conditions.
Literature-Reported Treatment Conditions
- Treatment concentration
- Lomitapide concentration series; Fig. 4C–D
- Exposure duration
- 10 min preincubation at room temperature; assay at room temperature. Methods describe fluorescence monitoring over 4 h; the exact endpoint time used in Fig. 4 is not specified.
- Method
- Incubation of purified MTP (25 ng) with inhibitor before lipid-transfer assay
- Reported limits
- Inhibitor solvent/vehicle and independent-experiment n for Fig. 4 are not specified. PL assay Methods state triplicate measurement; no biological n or IC50 uncertainty is assigned.
Methods and source locations
Compound reported in the paper: Lomitapide; Compound label in the paper: Sigma SML1385-5MG
Reported activity of the named Sigma reagent; does not validate Adooq material or establish in vivo potency.
Fig. 4C–D and caption; Materials and Methods: Materials, Measurement of the PL transfer activity of MTP; Results: Effects of different protein and substrate concentrations; Discussion assay caveats
Purified human MTP phospholipid-transfer activity: NBD-labeled phospholipid transfer between donor and acceptor vesicles, measured by fluorescence; compared with uninhibited control. — Fig. 4C, caption and associated Results; Materials and Methods: Measurement of the PL transfer activity of MTP
Purified human MTP triacylglycerol-transfer activity: Fluorescent TAG-transfer assay; compared with uninhibited control. — Fig. 4D, caption and associated Results; Materials and Methods assay paragraph
Mixed cellular responses after 24-hour lomitapide exposure in HepaRG cells
Cell-based study · Differentiated human HepaRG cultures; population-level high-content imaging [2]
The assay did not detect increased lipid accumulation with lomitapide after 24 h. Mitochondrial membrane potential decreased at the highest concentration, apoptotic nuclear morphology increased, and no oxidative-stress change was reported. This short-duration negative lipid result does not establish absence of longer-term steatogenic effects.
Literature-Reported Treatment Conditions
- Treatment concentration
- Concentration series below the WST-1 EC50; Results identify 12 µM as the highest lomitapide concentration. Full dose series is not transcribed from the plot.
- Exposure duration
- 24 h
- Method
- Exposure of differentiated HepaRG cultures followed by high-content fluorescence imaging
- Vehicle
- Treatment medium contained 0.5% DMSO and 2% FBS; Fig. 3 identifies the DMSO control.
- Reported limits
- No Adooq product/lot is reported. No exact fold changes, onset concentration for nuclear morphology, or long-duration lomitapide response are inferred from the curves.
Methods and source locations
Compound reported in the paper: Lomitapide; Compound label in the paper: Sigma-Aldrich; CAS 182431-12-5 in Supplementary table 4
Cellular responses to the reported research reagent; no equivalence to a clinical formulation or Adooq lot is asserted.
Fig. 3 (lomitapide column) and caption; Materials and methods: Chemicals and reagents, Cell culture and chemical exposure, High-content imaging assay, Automated quantitative image analysis, Dose–response data and model fitting; Results reference chemicals; Discussion; Supplementary table 4
Lipid-droplet count normalized to cell number and untreated cells: Bodipy staining and population-level imaging; positive hit defined by fitted response exceeding ±2 SD of the two lowest concentrations. — Fig. 3 lomitapide/lipid-accumulation panel and caption; Results reference chemicals; Dose–response data and model fitting; Discussion
Mitochondrial membrane potential proxy: Mitotracker Orange CM-H2TMROS fluorescence, normalized to cell number and untreated cells. — Fig. 3 lomitapide/mitochondrial-membrane-potential panel; Results reference chemicals; High-content imaging assay
Apoptotic nuclear morphology: Hoechst nuclear imaging with supervised classification; apoptotic cells normalized to total cells and untreated cells. — Fig. 3 lomitapide/nuclear-morphology panel and caption; Results reference chemicals; Automated quantitative image analysis
Oxidative-stress fluorescence: CellROX fluorescence, normalized to cell number and untreated cells. — Fig. 3 lomitapide/oxidative-stress panel; Results reference chemicals; High-content imaging assay
