Yuan Zhou1,2# , Haijuan Gu3#, Bingfeng Shao1, Suqing Zhang1, Harpreet Pall4, Renata D?Alpino Peixoto5, Shaffer R. S. Mok6, Guodong Zhu7

Background: Recent research suggests that dihydrolipoamide acetyltransferase (DLAT), which is a copper induced cell death-related gene, is involved in multiple biological events in tumors. This study sought to investigate the relationship between DLAT and hepatocellular carcinoma (HCC). Methods: In the Cancer Genome Atlas (TCGA) database, we first identified the differentially expressed gene (i.e., DLAT), then confirmed DLAT expression, and found a link between it and the prognosis of HCC patients. An internal validation nomogram was built based on a multivariate Cox regression analysis. Data from the Tumor Immune Estimation Resource (TIMER) database was used to examine the association between DLT and immunological cells. A gene set enrichment analysis (GSEA) was conducted to investigate the probable mechanism of action. Finally, in vitro cytological research was conducted to further examin the involvement of DLAT in HCC-related unfavorable biological events. Results: The database screenings showed that DLAT was a differentially expressed molecule; that is, DLAT was more highly expressed in the cancer tissues than normal tissues. TCGA results and Kaplan Meier-plotter data sets showed that HCC patients with reduced DLAT expression had greater disease specific survival (DSS), overall survival (OS), and progression-free interval (PFI). The prediction model had a concordance index of 0.659 (0.614-0.704), which indicates high accuracy. According to the TIMER database, tumor cells in the HCC microenvironment may be able to bypass the immune system due to the expression of DLAT. The in vitro cytological tests showed that DLAT knockdown significantly decreased the proliferation and invasion of the HCC cells. It also inhibited the activity of the phosphatidylinositol-4,5bisphosphate 3-kinase (PI3K)/protein kinase B (Akt) and Wnt/13-catenin signaling pathways. Conclusions: Decreased DLAT expression significantly prolongs the OS, PFI, and DSS of HCC patients. DLAT may be employed as a new predictive biomarker for HCC, and may be linked to the immune system in HCC patients. The tumor microenvironment (TME) may have a significant effect on the ability of tumor cells to evade the immune system. The PI3K/Akt and Wnt/13-catenin signaling pathways may affect the prognosis of HCC by interfering with DLAT. Given these findings, HCC may be an ideal target for the development of anti-cancer therapies.