Targeting β‐catenin in hepatocellular cancers induced by coexpression of mutant β‐catenin and K‐Ras in mice

Abstract

Recently we have shown that co-expression of hMet and mutant-β-catenin using sleeping beauty transposon/transposase leads to HCC in mice that represents around 10% of human HCC. In the current study, we investigate if Ras activation, which can occur downstream of Met signaling, is sufficient to cause HCC in association with mutant-β-catenin. We also tested therapeutic efficacy of targeting β-catenin in HCC model. We show that mutant-K-Ras (G12D), which leads to Ras activation, cooperates with β-catenin mutants (S33Y, S45Y) to yield HCC in mice. Affymetrix microarray shows >90% similarity in gene expression in mutant-K-Ras-β-catenin and Met-β-catenin HCC. K-Ras-β-catenin tumors showed upregulation of β-catenin targets like Glutamine Synthetase (GS), Lect2, Regucalcin and Cyclin-D1 and of K-Ras effectors including p-ERK, p-AKT, p-mTOR, p-EIF4E, p-4E-BP1 and p-S6 Ribosomal protein. Inclusion of dominant-negative TCF4 at the time of K-Ras-β-catenin injection prevented HCC and downstream β-catenin and Ras signaling. To address if targeting β-catenin has any benefit post-establishment of HCC, we administered K-Ras-β-catenin mice with EnCore lipid nanoparticle (LNP) loaded with a Dicer substrate siRNA targeting CTNNB1 (CTNNB1-LNP), scrambled sequence (Scr-LNP) or PBS for multiple cycles. A significant decrease in tumor burden was evident in CTNNB1-LNP group versus all controls, which was associated with dramatic decreases in β-catenin targets and some K-Ras effectors, leading to reduced tumor cell proliferation and viability. Intriguingly, in few mice, non-GS-positive tumors, which were evident as a small subset of overall tumor burden, were not affected by β-catenin suppression. In conclusion, we show that Ras activation downstream of c-Met is sufficient to induce clinically relevant HCC in cooperation with mutant β-catenin. β-Catenin suppression by a clinically relevant modality is effective in treatment of β-catenin-positive, GS-positive HCCs.

β-Catenin mRNA Silencing and MEK Inhibition Display Synergistic Efficacy in Preclinical Tumor Models

“…Wnt/β-catenin is one of several signaling networks that regulate its expression and activity (50), and therefore RNAi-mediated silencing of CTNNB1 is highly effective in HCC preclinical models (51, 52). Interestingly, MEK inhibition has been shown to destabilize MYC protein by regulating its ERK-dependent phosphorylation (53).…”

Wnt/β-Catenin Signaling in Liver Development, Homeostasis, and Pathobiology

“…As this signaling pathway can promote both liver injury and regeneration, therapies targeting Wnt/β-catenin signaling must be carefully evaluated to consider both the disease context and the targeted cell type. For example, targeting Wnt/β-catenin signaling may be a viable therapeutic option for liver cancer with oncogenic β-catenin signaling (119, 120). However, targeting Wnt/β-catenin signaling in HSCs to reduce ECM deposition in liver fibrosis may potentially be effective (186–188), but other studies indicate that it could potentially exacerbate fibrogenesis by promoting HSC activation (184, 185).…”

“…These mice developed HCC with gene expression profiles that displayed high correlation with the gene profiles of a subset of human HCC patients with both CTNNB1 mutations and Met activation signatures (118). To address if Ras activation downstream of Met could be contributing to Met-β-catenin HCC, G12D- KRAS and mutant-β-catenin were expressed using SB-HTVI, which also yielded HCC with approximately 90% molecular similarity to Met-β-catenin HCC (119). In fact, treatment of these mice with lipid nanoparticles containing small interfering RNA (siRNA) targeting CTNNB1 yielded a significant decrease in tumor burden (119).…”

“…To address if Ras activation downstream of Met could be contributing to Met-β-catenin HCC, G12D- KRAS and mutant-β-catenin were expressed using SB-HTVI, which also yielded HCC with approximately 90% molecular similarity to Met-β-catenin HCC (119). In fact, treatment of these mice with lipid nanoparticles containing small interfering RNA (siRNA) targeting CTNNB1 yielded a significant decrease in tumor burden (119). Thus, targeting β-catenin in HCC may prove to be part of a viable treatment strategy, as a course of antisense-mediated β-catenin suppression treatment led to a complete therapeutic response of β-catenin-activated HCC in mice (120).…”

Glutamine synthetase promotes tumor invasion in hepatocellular carcinoma through mediating epithelial–mesenchymal transition

“…GS is a target of the Wnt/beta‐catenin signal pathway in the liver, and the expression of GS is regulated by translocation of beta‐catenin from the cytoplasm to the cell nucleus, which indicates the activation of the Wnt/beta‐catenin signal pathway . Recently, studies have shown that activation of this pathway contributes to carcinogenesis and epithelial–mesenchymal transition (EMT) in HCC . Therefore, it is logical to speculate that GS might play a critical role in HCC progression.…”