Oncopig Soft-Tissue Sarcomas Recapitulate Key Transcriptional Features of Human Sarcomas

Abstract

Human soft-tissue sarcomas (STS) are rare mesenchymal tumors with a 5-year survival rate of 50%, highlighting the need for further STS research. Research has been hampered by limited human sarcoma cell line availability and the large number of STS subtypes, making development of STS cell lines and animal models representative of the diverse human STS subtypes critical. Pigs represent ideal human disease models due to their similar size, anatomy, metabolism, and genetics compared to humans. The Oncopig encodes inducible KRAS ^G12D and TP53 ^R167H transgenes, allowing for STS modeling in a spatial and temporal manner. This study utilized Oncopig STS cell line (fibroblast) and tumor (leiomyosarcoma) RNA-seq data to compare Oncopig and human STS expression profiles. Altered expression of 3,360 and 7,652 genes was identified in Oncopig STS cell lines and leiomyosarcomas, respectively. Transcriptional hallmarks of human STS were observed in Oncopig STS, including altered TP53 signaling, Wnt signaling activation, and evidence of epigenetic reprogramming. Furthermore, master regulators of Oncopig STS expression were identified, including FOSL1, which was previously identified as a potential human STS therapeutic target. These results demonstrate the Oncopig STS model's ability to mimic human STS transcriptional profiles, providing a valuable resource for sarcoma research and cell line development.

Figure 1

Expression of KRAS and TP53 across Oncopig STS cell lines and leiomyosarcoma tumors. Expression of (a) total, (b) wild type (WT), and (c) mutant KRAS and TP53 in Oncopig primary fibroblasts, AdGFP treated fibroblasts, and STS cell lines. Expression of total, WT, and mutant (d) KRAS and (e) TP53 in Oncopig skeletal muscle and leiomyosarcomas (LMS). Samples represent biological replicates. Expression values are presented as fragments per kilobase of transcript per million fragments mapped (FPKM). *denotes p < 0.05, **denotes p < 0.01, ***denotes q < 0.05.

Figure 2

Total number of DEGs and their expression levels across biological replicates. Total number of DEGs, including those displaying elevated and reduced expression in (a) Oncopig STS cell lines compared to controls and (b) leiomyosarcoma tumors compared to controls. (c) Heatmap of the normalized expression level of the 3,360 DEGs for each cell line, represented as z-scores. (d) Heatmap of the normalized expression level of the 7,625 DEGs for each in vivo sample, represented as z-scores. LMS = leiomyosarcoma. Dendrograms represent relationships between samples based on complete linkage clustering.

Figure 3

Map of DEGs in Oncopig leiomyosarcomas and their functions within the TP53 signaling pathway. Adopted from the KEGG hsa04115 p53 signaling pathway. Green ovals represent genes with elevated expression, red ovals represent genes with reduced expression, and grey ovals represent genes with no expression change in Oncopig leiomyosarcomas compared to controls. Black bars represent inhibition, black arrows represent activation, and blue arrows represent indirect effects.

Figure 4

Map of DEGs in Oncopig STS cell lines and their functions within the TP53 signaling pathway. Adopted from the KEGG hsa04115 p53 signaling pathway. Green ovals represent genes with elevated expression, red ovals represent genes with reduced expression, and grey ovals represent genes with no expression change in Oncopig STS cell lines compared to controls. Black bars represent inhibition, black arrows represent activation, and blue arrows represent indirect effects.

Figure 5

Differential expression of genes involved in Wnt signaling in Oncopig STS. (a) Expression of WNTs in Oncopig leiomyosarcomas and STS cell lines relative to controls, represented as the log2 fold change. (b) DEGs involved in the Wnt receptor signaling pathway in the Oncopig STS cell lines, represented as the log2 fold change relative to control. (c) DEGs involved in the regulation of Wnt receptor signaling pathway in the Oncopig leiomyosarcomas, represented as the log2 fold change relative to control. *denotes q-value < 0.05.

Figure 6

Independent validation confirms RNA-seq results. Log2 fold changes assessed using PCR and RNA-seq for 5 genes displaying elevated and reduced expression in the (a) Oncopig STS cell lines relative to controls and (b) leiomyosarcomas relative to controls. (c) Log2 fold changes assessed using qPCR and RNA-seq were highly correlated (Spearman’s Rho 0.992, p = 2.2 × 10⁻¹⁶).