Cooperation Between the NRF2 Pathway and Oncogenic β‐catenin During HCC Tumorigenesis

Abstract

CTNNB1 (catenin beta 1)-mutated hepatocellular carcinomas (HCCs) account for a large proportion of human HCCs. They display high levels of respiratory chain activity. As metabolism and redox balance are closely linked, tumor cells must maintain their redox status during these metabolic alterations. We investigated the redox balance of these HCCs and the feasibility of targeting this balance as an avenue for targeted therapy. We assessed the expression of the nuclear erythroid 2 p45-related factor 2 (NRF2) detoxification pathway in an annotated human HCC data set and reported an enrichment of the NRF2 program in human HCCs with CTNNB1 mutations, largely independent of NFE2L2 (nuclear factor, erythroid 2 like 2) or KEAP1 (Kelch-like ECH-associated protein 1) mutations. We then used mice with hepatocyte-specific oncogenic β-catenin activation to evaluate the redox status associated with β-catenin activation in preneoplastic livers and tumors. We challenged them with various oxidative stressors and observed that the β-catenin pathway activation increased transcription of Nfe2l2, which protects β-catenin-activated hepatocytes from oxidative damage and supports tumor development. Moreover, outside of its effects on reactive oxygen species scavenging, we found out that Nrf2 itself contributes to the metabolic activity of β-catenin-activated cells. We then challenged β-catenin activated tumors pharmacologically to create a redox imbalance and found that pharmacological inactivation of Nrf2 was sufficient to considerably decrease the progression of β-catenin-dependent HCC development. Conclusion:These results demonstrate cooperation between oncogenic β-catenin signaling and the NRF2 pathway in CTNNB1mediated HCC tumorigenesis, and we provide evidence for the relevance of redox balance targeting as a therapeutic strategy in CTNNB1-mutated HCC. (Hepatology Communications 2021;0:1-17).T umorigenesis is a complex process involving multiple modifications promoting proliferation and preventing cell death. The maintenance of neoplastic cell proliferation and growth requires a metabolic rewiring to provide essential macromolecules and energy. (1) Since the first observations that tumor cells displayed higher glucose consumption through a glycolytic pathway, it has been shown that there are numerous metabolic pathways, from enhanced glutaminolysis to increased fatty acid oxidation, that can be altered to favor tumor survival and progression. (2)(3)(4) Although altering their metabolic activity, tumor cells also have to maintain a redox status compatible with survival, as metabolism and redox balance are closely linked-mostly through the production

Nrf2 Mutation/Activation Is Dispensable for the Development of Chemically Induced Mouse HCC

“… 16 In support of cooperation between the Keap1–Nrf2 pathway and β-catenin signaling, a very recent article reported an enrichment of the NRF2 program in human HCCs with b-catenin (CTBBB1) mutations, largely independent of NRF2 or KEAP1 mutations. 27 The same study also showed that mice with hepatocyte-specific oncogenic β-catenin activation increased Nrf2 activation, most likely to protect β-catenin–activated hepatocytes from oxidative damage, thus favoring tumor development, and proposed cooperation between oncogenic β-catenin signaling and the NRF2 pathway in CTNNB1-mediated HCC. 27 Furthermore, an independent work showed co-activation of β-catenin and NRF2 in 9% of all human HCCs, and discovered that co-expression of mutated CTNNB1 with mutant NRF2 , but not wild-type NRF2 , led to rapid HCC development and mortality.…”

“… 27 The same study also showed that mice with hepatocyte-specific oncogenic β-catenin activation increased Nrf2 activation, most likely to protect β-catenin–activated hepatocytes from oxidative damage, thus favoring tumor development, and proposed cooperation between oncogenic β-catenin signaling and the NRF2 pathway in CTNNB1-mediated HCC. 27 Furthermore, an independent work showed co-activation of β-catenin and NRF2 in 9% of all human HCCs, and discovered that co-expression of mutated CTNNB1 with mutant NRF2 , but not wild-type NRF2 , led to rapid HCC development and mortality. 28 Thus, it appears that the frequency of Nrf2 mutations greatly vary among HCCs occurring in different species ( Figure 7 ).…”

“…Recent work on the mouse and human HCCs characterized by β-catenin mutations has shown that the Keap1–Nrf2 pathway is activated independently from NRF2 mutations, suggesting cooperation between the oncogenic β-catenin and Nrf2 pathways in Ctnnb1 -mediated HCC tumorigenesis. 27 , 28 Because HCCs generated by DEN + TCPOBOP or TCPOBOP alone display from 80% to 90% of Ctnnb1 mutations, 29 we wished to assess whether activation of the Keap1–Nrf2 pathway could occur concomitantly with β-catenin mutation, but independently from Nrf2 mutation.…”

“… 22 In this context, it should be noted that NRF2 mutations in rats occur in HCCs generated by a regimen almost identical to the experimental protocol 3 of the present study, consisting of a single dose of DEN followed by a choline-deficient diet. 22 Furthermore, in light of the recent works showing cooperation between β-catenin and Nrf2 in promoting liver cancer, 27 , 28 it is even more puzzling to find a complete absence of Nrf2 mutation and/or activation of this transcription factor in HCCs developed by DEN + TCPOBOP or TCPOBOP alone that displayed up to 80%–90% of mutations of Ctnnb1 . 29 …”

“…This study aimed to investigate whether Nrf2 mutation/activation occurs in distinct mouse models of hepatocarcinogenesis. In addition, because it recently was reported that in human HCCs characterized by β-catenin mutations the KEAP1–NRF2 pathway is activated independently from mutations of these genes, 27 , 28 we also wished to determine whether Nrf2 mutations could co-occur in β-catenin mutated HCC. Finally, we also investigated whether metabolic reprogramming, a hallmark of cancer cells, takes place in these experimental models.…”