Cancer cells incorporate and remodel exogenous palmitate into structural and oncogenic signaling lipids - PubMed (original) (raw)

Figure 2. Mapping exogenous isotopic FFA-derived lipid metabolism in human cancer cells

(A-E) Shown on the left are all ions detected in 231MFP, M4, SKOV3, PC3, or C8161 aggressive human cancer cells. For the volcano plot, points that are to the left of the dotted line (p<0.05) represent ions that were not statistically altered in levels between d0-C16:0 FFA versus d4-C16:0 FFA-labeled cells. All points to the right of the dotted line (p<0.05) represent ions that had significantly higher ion intensity in the d4-C16:0 FFA labeled group compared to d0-C16:0 FFA labeled group, i.e. d4-incorporated lipids. In total, at least ~5,000-10,000 ions were detected and analyzed between targeted and untargeted analysis comparing d0-C16:0 FFA labeled to d4-C16:0 FFA labeled M4, 231MFP, C8161, SKOV3, or PC3 cells. The y-axis denotes fold-change between raw integrated values of isotopically-incorporated ions by either targeted or untargeted analysis between d0 versus d4-C16:0 FFA-labeled samples. For the ions that exhibited no background peak corresponding to the m/z of the d4-lipid in the d0-C16:0 FFA-treated cells, we considered this value to be 1 to obtain a fold-change value compared to the raw integration values of d4-C16:0 FFA-treated cells. For the ions for which there was a background peak, we obtained a fold-change value by dividing the ion intensity for the d4-C16:0 FFA compared to d0-C16:0 FFA groups. The heat-map on the right shows relative levels of d4-C16:0 FFA-incorporated lipids in non-aggressive (MCF7, OVCAR3, LNCaP, MUM2C) or non-transformed (MCF10A) cells compared to aggressive (231MFP, M2T, M4, SKOV3, PC3, C8161) or transformed (M2) cells. In the heat-map, relative levels of each d4-incorporated lipid metabolite are shown (darker blue shading corresponds to higher level of metabolite). The lipid designations next to the heat map are color-coded red for significantly higher, blue for significantly lower, and grey for unchanged d4-metabolites in aggressive cancer cells (231MFP, M4, SKOV3, PC3, and C8161) compared to non-aggressive (MCF7, MCF10A, OVCAR3, LNCaP, and MUM2C, respectively) cells (p<0.05). **(F)** Shown are lipids species that exhibited significant d4-C16:0 FFA incorporation _in vivo_ in mice bearing a tumor xenograft from M4 cells. Mice were subcutaneously injected with 2 × 106 M4 cells and tumors were grown out to ~800-1000 mm3. Mice were treated with vehicle (polyethylene glycol 300 (PEG300)) or d4-C16:0 FFA (100 mg/kg in PEG) by oral gavage (4 h). Tumors were harvested and lipids were extracted and analyzed by SRM-based metabolomics. For **A-F**, those metabolites where there was a background peak for the d4-lipid m/z in the d0-C16:0 FFA-treated cells, the average of the background ion intensity was subtracted from both d0 and d4-C16:0 FFA-treated groups. For all lipid shown here, any background peak for a d4-lipid detected in d0-C16:0 FFA-treated cells was assumed to either be a coeluting isobaric metabolite or natural isotopic abundance of the lipid. We have only presented here the d4-incorporated lipids that showed >5-fold significantly (p<0.05) higher ion intensity in the d4-C16:0 FFA-treated group compared to the d0-C16:0 FFA-treated group. All data from A-E is shown in Supplemental Table 1 and certain lipids are quantified in Figure 3. Data in (A-E) are average values of n=4-6 biological replicates. Data in (F) are mean ± standard error of n=4-6 biological replicates. Significance in (F) is represented as *p<0.05 in d4-C16:0 FFA-treated mice compared with vehicle-treatment.