Purpose: Tryptophan 2,3-dioxygenase (TDO), encoded by the gene TDO2, is an enzyme that catalyses the first and rate-limiting step of tryptophan (Try) degradation in the kynurenine (Kyn) pathway in the liver. Recently, TDO has been demonstrated to be expressed in various human tumours, especially hepatocellular carcinoma (HCC). However, the role of TDO in HCC is still not very clear. Here, we studied the role of TDO in HCC. Methods: We demonstrated that TDO is overexpressed in human HCC tissues and is significantly correlated with malignant phenotype characteristics, including tumour size, tumour differentiation, vascular invasion, etc. Kaplan–Meier analysis showed a poor overall survival rate in patients with TDO-overexpressing tumours. In addition, the effects of TDO on HCC tumour growth and metastasis were detected both in vivo and in vitro. TDO overexpression facilitated HCC cell growth, invasion and migration. Conclusion: Our results suggest that TDO positively regulates HCC proliferation and invasion and acts as a new prognostic biomarker of HCC.

Background: Cuproptosis has been studied in various aspects as a new form of cell death.Aims: We hope to explore the molecular patterns and genes related to cuproptosis in evaluating and predicting the prognosis of hepatocellular carcinoma (HCC), as well as the impact of tumor immune microenvironment.Methods and Results: Sixteen cuproptosis related gene (CRGs) and cuproptosis related molecular and gene characteristics were comprehensively analyzed from 492 HCC samples. Cuproptosis related molecular patterns were generated by consensus clustering algorithm, including cuproptosis clusters, cuproptosis gene clusters (CGC) and cuproptosis score (CS). The characteristics of tumor microenvironment (TME) and tumor immune cells were described by the ssGSEA and ESTIMATE algorithms. Cuproptosis score was established to assess the clinical characteristics, prognostic and immunotherapy. The role and mechanism of CRG (ATP7A) in HCC, as well as its relationship with TME and immune checkpoints, have been further explored. The results of somatic mutation, copy number variations (CNV), and CRGs expression in HCC suggested the CRGs might participate in the HCC oncogenesis. The cuproptosis clusters were closely related to the clinical pathological characteristics, biological processes, and prognosis of HCC. The three CGC was revealed to be consistent with the three immune infiltration characterizations, including immune‐high, immune‐mid, and immune‐low subtypes. Higher CS was characterized by decreased TMB, activated immunity, higher immune cell proportion score (IPS) and better overall survival (OS), which indicated higher CS was immune‐high type and with better treatment effect and prognosis. The ATP7A had the highest hazard ratio (HR = 1.465, p < .001), was high expression in HCC tissues and with a shorter 5‐year OS. Knocking down ATP7A could enhance intracellular copper concentration, cause a decrease in DLAT expression, and induce cuproptosis and inhibit cell proliferation and migration. ATP7A was also positively correlated with most cancer immune cells and immune checkpoints.Conclusion: Taken together, this research revealed the cuproptosis related molecular patterns and genes associated with the clinical pathological characteristics, TME phenotype and prognosis of HCC. The CS will further deepen our understanding of the TME characteristics of HCC, and the involvement of ATP7A in cuproptosis will provide new ideas for predicting HCC prognosis and immunotherapy.

Background: The correlations between cuproptosis and long noncoding RNAs (lncRNAs) with the tumor microenvironment (TME), immunotherapy, and some other characteristics of hepatocellular carcinoma (HCC) remain unclear.Methods: Sixteen cuproptosis regulators and 356 cuproptosis-related lncRNAs (CRLnc) were identified from 374 HCC profiles in The Cancer Genome Atlas (TCGA) database. Six differentially expressed CRLnc were selected, and a prognostic risk model based on the CRLnc signature (CRLncSig) was constructed. The prognostic power of the model was verified. Moreover, a cuproptosis-related gene cluster (CRGC) was generated based on six lncRNAs and differentially expressed genes. The relationship between immune cell infiltration in the TME, immunotherapy, CRLncSig, and CRGC was demonstrated through various algorithms, Tumor Immune Dysfunction and Exclusion (TIDE), tumor mutational burden (TMB), etc. Potential drugs and sensitivity to those agents were evaluated for the risk model. LncRNA AL158166.1 was selected and verified in HCC tissues and cell lines, the impact of its knockdown and overexpression in HCC cells was examined, and the copper (Cu) concentration and the cuproptosis-related gene expression were detected.Results: A CRLncSig prognostic risk model with good predictive ability was constructed. The low-risk group had a longer overall survival (OS), lower tumor purity, more extensive immune cell infiltration, higher immune score, enrichment in immune-activated pathways, and more positive response to immunotherapy versus the high-risk group. CRGC-B exhibited the best OS and the lowest tumor stage; the immune cell infiltration analysis was similar to the low-risk group in CRLncSig. CRGC-B belonged to the “immune-high” group of the TME. The low-risk group had a higher TIDE score and susceptibility to antitumor drugs. The lncRNA AL158166.1 had the highest hazard ratio. The levels of AL158166.1 were higher in HCC tissues versus healthy tissues. Knockdown of AL158166.1 could lead to an increase in intracellular Cu concentration, induce DLAT low expression, and inhibit the proliferation and migration of HCC cells, whereas overexpression of AL158166.1 exerted the reverse effect.Conclusion: Overall, a new CRLncSig prognostic risk model and a cuproptosis-related molecular signature were constructed and evaluated. The model and signature were associated with the prognosis, immune infiltration, and immunotherapy of HCC. Inhibiting the lncRNA AL158166.1 may induce cuproptosis and showed potential for the inhibition of tumors. Evaluation of the CRLnc, CRLncSig, and CRGC may enhance our understanding of the TME, determine the effectiveness of immunotherapy, and act as a marker for the prognosis of HCC.