2.28E E AKT/CAT P-3-4:

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1 Supplemental Table 1: IPA network analysis of the microarray data presented in Figure 6 showing the most significant molecular and cellular function classifications along with the respective number of molecules and corresponding range of p-values. Name p-value Molecules AKT/CAT P-0: Amino Acid Metabolism 9.45E E Small Molecule Biochemistry 9.45E E Lipid Metabolism 6.14E E Cell Cycle 1.08E E Cell Death 2.28E E AKT/CAT P-3-4: Lipid Metabolism 3.02E E Molecular Transport 3.02E E Small Molecule Biochemistry 3.02E E Amino Acid Metabolism 4.73E E Cellular Growth and Proliferation 3.35E E MET/CAT P-0: Cell-To-Cell Signaling and Interaction 1.02E E Cellular Movement 1.81E E Amino Acid Metabolism 4.34E E Small Molecule Biochemistry 4.34E E Cellular Growth and Proliferation 4.40E E

2 Supplemental Table 2A: Sub-networks of the lipid metabolism functional category (Supplemental Table 1) derived by IPA network analysis of the AKT/CAT P-0 model. Functional networks are annotated and shown along with their respective number of molecules and corresponding p-values. Networks involved in lipid breakdown are highlighted in yellow. Function Function Annotation p-value # Molecules production production of 12-hydroxyeicosatetraenoic acid 6.14E-06 5 production production of 12(S)-hydroxyeicosatetraenoic acid 1.27E-04 3 production production of 6-keto-prostaglandin F1 alpha 2.26E-03 3 production production of eicosanoid 2.98E production production of estrogen 7.12E-03 4 production production of progesterone 7.66E-03 7 production production of lipid 8.94E production production of prostaglandin E2 9.82E-03 9 conversion conversion of fatty acid 4.27E-05 9 conversion conversion of eicosanoid 4.50E-04 6 conversion conversion of prostaglandin h2 6.54E-04 4 conversion conversion of lipid 1.22E biosynthesis biosynthesis of acyl-coenzyme A 1.14E-04 5 biosynthesis biosynthesis of acetyl-coenzyme A 1.05E-03 4 modification modification of prostaglandin h2 1.14E-04 5 modification modification of prostaglandin 2.45E-04 6 modification modification of fatty acid 1.65E modification modification of acyl-coenzyme A 2.62E-03 5 modification modification of eicosanoid 2.98E-03 7 modification modification of lipid 3.91E quantity quantity of fatty acid 2.79E quantity quantity of prostaglandin E2 3.41E-04 8 quantity quantity of eicosanoid 9.98E quantity quantity of aldosterone 1.58E-03 5 quantity quantity of vitamin E 2.53E-03 2 quantity quantity of lipid 3.48E quantity quantity of 6-keto-prostaglandin F1 alpha 8.49E-03 3 quantity quantity of glycosylceramide 1.17E-02 3 quantity quantity of lysophosphatidylcholine 1.17E-02 3 release release of epoprostenol 4.89E-04 3 secretion secretion of prostaglandin 8.74E-04 5 secretion secretion of corticosterone 3.16E-03 4 secretion secretion of prostaglandin E2 5.56E-03 4 steroidogenesis steroidogenesis of cells 1.18E-03 3 steroidogenesis steroidogenesis of Leydig cells 2.53E-03 2 metabolism metabolism of fatty acid 1.19E metabolism metabolism of lipid 6.55E-03 46

3 Supplemental Table 2A (continued): Function Function Annotation p-value # Molecules binding binding of phosphatidic acid 1.11E-02 4 catabolism catabolism of glucocorticoid 2.53E-03 2 induction induction of lipid 3.30E-03 5 induction induction of prostaglandin 5.56E-03 4 metabolic process metabolic process of lipid 3.41E stimulation stimulation of prostaglandin E2 3.81E-03 3 reduction reduction of fatty acid 4.25E-03 4 formation formation of testosterone 7.33E-03 2 inhibition inhibition of prostaglandin 7.33E-03 2 inhibition inhibition of lipid 1.17E-02 3 oxidation oxidation of lipid 1.16E-02 14

4 Supplemental Table 2B: Sub-networks of the lipid metabolism functional category (Supplemental Table 1) derived by IPA network analysis of the AKT/CAT serial passage (P-3-4) model. Functional networks are annotated and shown along with their respective number of molecules and corresponding p-values. Networks involved in lipid breakdown are highlighted in yellow. Function Function Annotation p-value # Molecules quantity quantity of lipid 3.02E quantity quantity of cholesterol 1.49E quantity quantity of acylglycerol 6.91E quantity quantity of triacylglycerol 8.10E quantity quantity of steroid 1.74E quantity quantity of fatty acid 7.38E quantity quantity of prostaglandin 3.32E quantity quantity of phospholipid 3.32E quantity quantity of bile acid 6.23E-04 6 metabolic process metabolic process of lipid 1.52E metabolic process metabolic process of fatty acid 8.53E metabolism metabolism of lipid 1.98E metabolism metabolism of fatty acid 7.86E metabolism metabolism of terpenoid 4.48E modification modification of lipid 4.94E modification modification of fatty acid 1.34E modification modification of prostaglandin E2 1.41E-03 4 oxidation oxidation of lipid 5.43E oxidation oxidation of fatty acid 2.27E oxidation oxidation of retinol 1.41E-03 4 synthesis synthesis of lipid 2.31E synthesis synthesis of acylglycerol 2.17E synthesis synthesis of cholesterol 8.36E synthesis synthesis of triacylglycerol 1.05E-03 9 accumulation accumulation of lipid 3.38E secretion secretion of bile acid 1.11E-04 4 moiety attachment moiety attachment of lipid 4.22E degradation degradation of lipid 4.25E transport transport of lipid 7.44E beta-oxidation beta-oxidation of fatty acid 1.03E binding binding of phosphatidic acid 1.37E-03 7

5 Supplemental Figure Legends Supplemental Figure 1. Schematic of transposonable cassettes and the transposase expression vector. Direct repeats/inverted repeats (DIR) are recognized by transposase and delineates the borders of the integrated transposable element. hmet, Myr-AKT, Δ90N β-catenin, and HSB2 are the expressed genes. Myr is the myristolation signal from v-src fused to the N-terminus of AKT. Supplemental Figure 2. Histochemical analysis of lipid and glycogen distribution in the AKT/CAT and MET/CAT models. Panel A: Oil red-o staining (red) demonstrates large lipid droplets coinciding with vacuoles characteristic of steatosis (100X, bar =100um). Top row shows the incidence of lipid droplets decreases with increased passage in the AKT/CAT model. Middle row shows MET/CAT tumor contains very little lipid and is not significantly different than background (T3, lowest panel). Each specimen is framed to show uninvolved liver tissue at the left margin for reference. Panel B: PAS staining (magenta) does not selectively associate with any tumor cell or feature in either the AKT/CAT or MET/CAT models (100X, bar =100um). Sections are made from the same specimen as Panel A. The AKT/CAT passage induced increase in cytoplasmic granularity is not due to accumulation of glycogen (top row). The MET/CAT model shows nearly equivalent staining to background (T3, lowest panel). Supplemental Figure 3. Histochemical analysis of fibrosis in the AKT/CAT and MET/CAT models. Masson trichrome staining does not reveal the presence of a

6 fibrotic network associated with either the AKT/CAT or MET/CAT models (100X, bar =100um). Bottom panel (T3) shows collagen staining (blue) of cells associated with the portal vein. Supplemental Figure 4. Hierarchical cluster analysis of integrated human HCC and mouse models. Panel A: Hierarchical cluster analysis of integrated 53 human HCC and 11 MET/CAT derived mouse tumors. A list of 2403 orthologous genes between the two species presented on both platforms was identified Clustering by transcriptional similarity forms a heat map with a format showing individual gene expression levels (red induced, green repressed relative to non-tumorous liver) in each column and the rows reflecting the tumor samples. The resulting dendrogram is presented in Figure 5B with stratification. Panel B: Hierarchical cluster analysis of integrated 53 human HCC and 9 AKT/CAT derived mouse tumors. A list of 833 orthologous genes between the two species presented on both platforms was identified. A clustered heat map was arranged as described above and the dendrogram is presented in Figure 5C. Supplemental Figure 5. Cumulative survival analysis. Plotting the survival of the two primary subdivisions generated by integration with MET/CAT (Panel A) or AKT/CAT (Panel B) mouse tumor followed by hierarchical clustering, identified clinically relevant distinct subgroups of human HCC. Kaplan-Meier plots and Mantel-Cox statistical analysis were applied in the survival analysis (p<0.04).

7 Supplemental Figure 6. Network comparison by Ingenuity Pathway Analysis. Panel A: Network comparison of primary AKT/CAT and MET/CAT tumor gene expression found Liver steatosis as the most statistically significant (AKT/CAT, p<0.004; MET/CAT, p<0.09) pathway that is also selectively expressed in AKT/CAT. Some of the genes involved in liver steatosis appear to be solely expressed in AKT/CAT but not in MET/CAT although below the threshold of p<0.05 (dashed line). Panel B: Network comparison of primary (P-0) and passaged (P-3-4) AKT/CAT tumor gene expression identified lipid metabolism as the most significant pathway in common. The relative significance of this pathway is shown in Supplemental Table 1. The dotted line represents the statistical significance threshold of p<0.05.