Background: N⁶-Methyladenosine (m⁶A) modification has been implicated in many biological processes. It is important for the regulation of messenger RNA (mRNA) stability, splicing, and translation. However, its role in cancer has not been studied in detail. Here we investigated the biological role and underlying mechanism of m⁶A modification in hepatoblastoma (HB). Methods: We used Reverse transcription quantitative real-time PCR (RT-qPCR) and Western blotting to determine the expression of m⁶A related factors. And we clarified the effects of these factors on HB cells using cell proliferation assay, colony formation, apoptotic assay. Then we investigated of methyltransferase-like 13 (METTL3) and its correlation with clinicopathological features and used xenograft experiment to check METTL3 effect in vivo. m⁶A-Seq was used to profiled m⁶A transcriptome-wide in hepatoblastoma tumor tissue and normal tissue. Finally, methylated RNA immunoprecipitation (MeRIP) assay, RNA remaining assay to perform the regulator mechanism of MEETL3 on the target CTNNB1 in HB. Results: In this research, we discovered that m⁶A modifications are increased in hepatoblastoma, and METTL3 is the main factor involved with aberrant m⁶A modification. We also profiled m⁶A across the whole transcriptome in hepatoblastoma tumor tissues and normal tissues. Our findings suggest that m⁶A is highly expressed in hepatoblastoma tumors. Also, m⁶A is enriched not only around the stop codon, but also around the coding sequence (CDS) region. Gene ontology analysis indicates that m⁶A mRNA methylation contributes significantly to regulate the Wnt/β-catenin pathway. Reduced m⁶A methylation can lead to a decrease in expression and stability of the CTNNB1. Conclusion: Overall our findings suggest enhanced m⁶A mRNA methylation as an oncogenic mechanism in hepatoblastoma, METTL3 is significantly up-regulated in HB and promotes HB development. And identify CTNNB1 as a regulator of METTL3 guided m⁶A modification in HB.

The N6-methyladenosine (m⁶A) RNA modification can regulate autophagy to modulate the growth and development of tumors, but the mechanism of m6A modification for the regulation of autophagy in hepatocellular carcinoma cells (HCC) remains unclear. In the study, the knockdown of the Wilms’ tumor 1-associating protein (WTAP) was made in HCC to study the correlation between m6A modification and autophagy. A fluorescent confocal microscopy analysis showed that the knockdown of WTAP could facilitate the autophagy of HCC. A Western blot analysis showed that the level of p-AMPK was decreased in WTAP-knockdown HCC cells. Additionally, LKB1, the upstream kinase of AMPK, was regulated by WTAP and it could mediate the phosphorylation of AMPK in an m6A-dependent manner. Further studies revealed that the knockdown of WTAP could reduce the level of LKB1 mRNA with m6A. This could result in the increased stability of LKB1 mRNA to promote its expression. The knockdown of WTAP could upregulate the level of autophagy and inhibit HCC proliferation. However, the overexpression of WTAP could resist autophagic cell death.

Background: Hepatoblastoma is a common hepatic tumor occurring in children between 0-5 years. Accumulating studies have shown lncRNA's potential role in distinct cancer progression and development, including hepatoblastoma. SnoRNA host gene 9 (SNHG9) is associated with the progression of distinct human cancers, but, its specific molecular mechanisms in hepatoblastoma is not unknown.

Background: Accumulating studies have shown that La‐related protein 1 (LARP1) is involved in the occurrence and development of various tumours. However, the expression pattern and biological role of LARP1 in hepatoblastoma (HB) remain unclear so far.Methods: LARP1 expression level in HB and adjacent normal liver tissues was analysed by qRT‐PCR, Western blotting and immunohistochemistry assays. The prognostic significance of LARP1 was evaluated by Kaplan–Meier method and multivariate Cox regression analysis. In vitro and in vivo functional assays were implemented to clarify the biological effects of LARP1 on HB cells. Mechanistically, the regulatory roles of O‐GlcNAcylation and circCLNS1A in LARP1 expression were investigated by co‐immunoprecipitation (co‐IP), immunofluorescence, RNA immunoprecipitation (RIP), RNA pull‐down and protein stability assays. Moreover, RNA‐sequencing, co‐IP, RIP, mRNA stability and poly(A)‐tail length assays were performed to investigate the association between LARP1 and DKK4. The expression and diagnostic significance of plasma DKK4 protein in multi‐centre cohorts were evaluated by ELISA and ROC curves.Results: LARP1 mRNA and protein levels were remarkably elevated in HB tissues and associated with worse prognosis of HB patients. LARP1 knockdown abolished cell proliferation, triggered cell apoptosis in vitro as well as prohibited tumour growth in vivo, whereas LARP1 overexpression incited HB progression. Mechanistically, O‐GlcNAcylation of LARP1 Ser672 by O‐GlcNAc transferase strengthened its binding to circCLNS1A and then protected LARP1 from TRIM‐25‐mediated ubiquitination and proteolysis. LARP1 upregulation subsequently led to DKK4 mRNA stabilisation by competitively interacting with PABPC1 to prevent DKK4 mRNA from B‐cell translocation gene 2‐dependent deadenylation and degradation, thus facilitating β‐catenin protein expression and nuclear import.Conclusion: This study indicates that upregulated protein level of O‐GlcNAcylated LARP1 mediated by circCLNS1A promotes the tumorigenesis and progression of HB through LARP1/DKK4/β‐catenin axis. Hence, LARP1 and DKK4 are promising therapeutical target and diagnostic/prognostic plasma biomarker for HB.

Background: The incidence rate of hepatoblastoma (HB), which is the most prevalent malignant tumour among children, rises each year. According to recent studies, a number of neoplastic disorders and ferroptosis are intimately connected. This study aims to identify key ferroptosis-related genes in HB and explore new directions for the diagnosis and treatment of HB. Methods: Differentially expressed ferroptosis-related genes were identified using the Gene Expression Omnibus datasets. The functional annotation of candidate genes was evaluated through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. Machine learning and receiver operating characteristic (ROC) curves revealed protein kinase AMP-activated catalytic subunit alpha 2 (PRKAA2), tribbles homolog 2 (TRIB2), and liver-type glutaminase (GLS2) as potential diagnostic genes of HB. By using quantitative reverse transcription polymerase chain reaction (qRT-PCR) and immunohistochemistry, relative expression of PRKAA2 was examined. The effect of PRKAA2 on proliferation, apoptosis, and ferroptosis of HB cells was verified in vitro and in vivo. Fisher’s exact test was used to evaluate the clinical significance of PRKAA2 in HB. Results: The prognostic indicators had a substantial correlation with PRKAA2 expression, which rose dramatically in HB tissues. PRKAA2 promotes proliferation and inhibits ferroptosis in HB cells. PRKAA2 plays a role in ferroptosis by regulating hypoxia-inducible factor 1α (HIF-1α) and transferrin receptor 1 (TFR1). Conclusions: PRKAA2 functions as a tumor-promoting factor in HB by promoting cell proliferation and prohibiting ferroptosis. Ferroptosis-related genes PRKAA2 is a potential diagnostic and prognostic marker for HB as well as a novel therapeutic target in the future.

Hepatoblastoma, the most aggressive childhood liver tumor, poses significant challenges due to limited knowledge of its pathogenesis, particularly in poorly differentiated advanced tumors where the prognosis is dismal. Single‐cell sequencing provides an in‐depth exploration at the single‐cell level and offers a deep understanding of tumor heterogeneity. Herein, single‐cell transcriptomics analysis is used to identify a unique malignant‐hepatoblast (HB)‐like cell subpopulation as the possible origin of poorly differentiated hepatoblastoma. These cells are associated with an unfavorable clinical prognosis in hepatoblastoma patients. The malignant‐HB‐like cell subpopulation generated insulin‐like growth factor 2 (IGF2) to sustain stem‐like features by promoting abnormal cholesterol accumulation via SREBF2. IGF2 also stimulated fibroblast 2 to secrete collagen 1, intensifying tumor malignancy via the collagen 1/integrin α1 signaling pathway. This suggests that targeting malignant HB‐like cells by inhibiting IGF2‐induced pathways can lead to promising treatments for hepatoblastoma. Additionally, serum IGF2 levels may serve as a diagnostic biomarker for advanced hepatoblastoma. In summary, these findings provide valuable insight into the genesis and malignancy of hepatoblastoma and a foundation for more effective diagnostic tools and therapeutic strategies for this challenging disease.