New molecular mechanisms in cholangiocarcinoma: signals triggering interleukin-6 production in tumor cells and KRAS co-opted epigenetic mediators driving metabolic reprogramming

Abstract

Background Cholangiocarcinoma (CCA) is still a deadly tumour. Histological and molecular aspects of thioacetamide (TAA)-induced intrahepatic CCA (iCCA) in rats mimic those of human iCCA. Carcinogenic changes and therapeutic vulnerabilities in CCA may be captured by molecular investigations in bile, where we performed bile proteomic and metabolomic analyses that help discovery yet unknown pathways relevant to human iCCA. Methods Cholangiocarcinogenesis was induced in rats (TAA) and mice (JnkΔhepa + CCl4 + DEN model). We performed proteomic and metabolomic analyses in bile from control and CCA-bearing rats. Differential expression was validated in rat and human CCAs. Mechanisms were addressed in human CCA cells, including Huh28-KRASG12D cells. Cell signaling, growth, gene regulation and [U-13C]-D-glucose-serine fluxomics analyses were performed. In vivo studies were performed in the clinically-relevant iCCA mouse model. Results Pathways related to inflammation, oxidative stress and glucose metabolism were identified by proteomic analysis. Oxidative stress and high amounts of the oncogenesis-supporting amino acids serine and glycine were discovered by metabolomic studies. Most relevant hits were confirmed in rat and human CCAs (TCGA). Activation of interleukin-6 (IL6) and epidermal growth factor receptor (EGFR) pathways, and key genes in cancer-related glucose metabolic reprogramming, were validated in TAA-CCAs. In TAA-CCAs, G9a, an epigenetic pro-tumorigenic writer, was also increased. We show that EGFR signaling and mutant KRASG12D can both activate IL6 production in CCA cells. Furthermore, phosphoglycerate dehydrogenase (PHGDH), the rate-limiting enzyme in serine-glycine pathway, was upregulated in human iCCA correlating with G9a expression. In a G9a activity-dependent manner, KRASG12D promoted PHGDH expression, glucose flow towards serine synthesis, and increased CCA cell viability. KRASG12D CAA cells were more sensitive to PHGDH and G9a inhibition than controls. In mouse iCCA, G9a pharmacological targeting reduced PHGDH expression. Conclusions In CCA, we identified new pro-tumorigenic mechanisms: Activation of EGFR signaling or KRAS mutation drives IL6 expression in tumour cells; Glucose metabolism reprogramming in iCCA includes activation of the serine-glycine pathway; Mutant KRAS drives PHGDH expression in a G9a-dependent manner; PHGDH and G9a emerge as therapeutic targets in iCCA.

Crenigacestat blocking notch pathway reduces liver fibrosis in the surrounding ecosystem of intrahepatic CCA viaTGF-β inhibition

“…These MOC allow tumor cells to circumvent the potentially harmful effects of chemotherapy [42]. Recent findings highlight the crucial role of the inflammatory milieu, because of EGFR-RAS-MAPK axis activation and pro-carcinogenic cytokine IL6 production, in iCCA progression [43]. Molecular mechanisms underlying resistance to treatment are still unknown; nevertheless, iCCA is characterized histologically by a strong desmoplastic reaction [9,44,45].…”

Pancreatic Cancer Research beyond DNA Mutations

“…Recent studies indicate that in intrahepatic cholangiocarcinoma (iCCA), which can be influenced by surrounding cells in the liver. Epidermal growth factor receptor (EGFR) signaling and KRAS mutation (G12D) can activate interleukin 6 (IL6) production in iCCA cells [ 57 ]. This condition induces upregulation of phosphoglycerate dehydrogenase (PHGDH), the rate-limiting enzyme in the serine-glycine pathway in human iCCA, which correlates with euchromatic histone lysine methyltransferase 2 ( EHMT2 / G9A ) expression [ 57 ].…”

“…Epidermal growth factor receptor (EGFR) signaling and KRAS mutation (G12D) can activate interleukin 6 (IL6) production in iCCA cells [ 57 ]. This condition induces upregulation of phosphoglycerate dehydrogenase (PHGDH), the rate-limiting enzyme in the serine-glycine pathway in human iCCA, which correlates with euchromatic histone lysine methyltransferase 2 ( EHMT2 / G9A ) expression [ 57 ]. In a G9A activity-dependent manner, KRAS mutation (G12D) promotes the PHGDH expression and glucose flow toward serine synthesis.…”

“…In a G9A activity-dependent manner, KRAS mutation (G12D) promotes the PHGDH expression and glucose flow toward serine synthesis. It increases CCA cell viability [ 57 ], suggesting that the KRAS mutation can elicit metabolic reprogramming under inflammatory conditions in TME. Reportedly, the critical role of this KRAS-dependent serine biosynthesis pathway was demonstrated in PDAC, suggesting that targeting the serine biosynthesis pathway by inhibiting PHGDH is a potential therapeutic approach to eliminate PDAC cells in nutrient-deprived TME [ 58 ] ( Figure 6 ).…”

Signaling and molecular networks related to development and inflammation involved in CCA initiation and progression

“…In iCCA, Nintedanib impaired the activity and proliferation of CAFs and the production of cancer-promoting cytokines. Thus, administered together with gemcitabine might result in a new promising therapy for refractory iCCA [134] .…”