The prognostic role of ACSL4 in postoperative adjuvant TACE-treated HCC: implications for therapeutic response and mechanistic insights
Authors: Feng Ji , Bin Jin-Lian , Liao Xi-Wen , Wu Yong , Tang Yue , Lu Pei-Zhi , Zhu Guang-Zhi , Cui Qian-Ru , Dan Yock Young , Yang Guo-Huan , Li Li-Xin , Deng Jing-Huan , Peng Tao , Hooi Shing Chuan , Zhou Jing , Lu Guo-Dong 
Published: November 2024

Abstract
Background
The response of hepatocellular carcinoma (HCC) to transarterial chemoembolization (TACE) treatment and its underlying mechanisms remain elusive. This study investigates the role of enzymes involved in fatty acid activation, specifically Acyl-CoA synthetase long chain 4 (ACSL4), in HCC patients treated with postoperative adjuvant TACE (PA-TACE) and in nutrient-deprived HCC cells.

Methods
We examined the expression of ACSL4 and its family members in HCC clinical samples and cell lines. The clinical significance of ACSL4, particularly regarding the prognosis of patients treated with PA-TACE, was assessed using two independent HCC cohorts. We further explored the role of ACSL4 in glucose starvation-induced cell death in HCC cells and xenograft mouse models.

Results
Among the family members, ACSL4 is the most up-regulated enzyme, associated with poor survival in HCC patients, particularly in post-recurrent TACE-treated patients in a Singapore cohort. ACSL4 is essential for HCC cell survival in response to glucose starvation, rather than to hypoxia or to the combination of hypoxia with doxorubicin or cisplatin. ACSL4-mediated arachidonic acid (AA) metabolism supports mitochondrial β-oxidation and energy production. CCAAT/enhancer binding protein α (CEBPA) transcriptionally regulates ACSL4 by binding 3 motifs (-623 to -613, -1197 to -1187 and -1745 to -1735) of ACSL4 upstream promoter region, enhancing its pro-survival effects. Furthermore, canagliflozin (Cana), a clinical-approved drug for type 2 diabetes, mimics glucose starvation and inhibits the growth of ACSL4-low xenograft tumors. Moreover, high ACSL4 or CEBPA expressions correlate with increased recurrence susceptibility after PA-TACE in the China-Guangxi HCC cohort.

Conclusions
The CEBPA-ACSL4 pathway is critical in protecting HCC cells from glucose starvation-induced cell death, suggesting that ACSL4 and CEBPA could serve as valuable prognostic indicators and potential therapeutic targets in the context of PA-TACE treatment for HCC.

Graphical Abstract
TACE is the first-line treatment for intermediate-stage HCC patients with unresectable tumor and a common postoperative adjuvant (PA) treatment. The present study found that HCC patients with the presence of CEBPA-ACSL4 expression in tumor are more resistant to TACE, susceptible for PA-TACE relapse and poorer survival. Mechanically, ACSL4 is essential for fatty acid activation particularly conversion of arachidonic acid (AA) into AA-CoA, which promotes lipid anabolism in nutrition-replete condition and lipid catabolism in glucose-deplete condition. As a result, ACSL4-high HCC cells, in response to glucose restriction (rather than hypoxia and chemotherapeutic drugs), can donor mitochondrial energy production through β-oxidation and protect HCC cells from mitochondrial membrane impairment and cell death in vitro and in vivo. In addition, CEBPA transcriptionally activates ACSL4 , and knockout of CEBPA aborted ACSL4-mediated lipid metabolism.