Emerging Role of Sirtuins-Mediated Ferroptosis in Hepatocellular Carcinoma Progression: Mechanisms and Therapeutic Perspectives

Abstract

Hepatocellular carcinoma (HCC) ranks as the most common type of primary liver cancer, characterized by rapid tumor growth and therapeutic resistance. Evading cell death is a key characteristic of multiple cancers, including HCC. Many programmed cell death (PCD) processes, comprising apoptosis, autophagy, necroptosis, and ferroptosis, have been identified to affect tumor growth and recurrence of HCC. Ferroptosis is an emerging iron-dependent PCD mode characterized by lipid peroxidation and iron accumulation. It has emerged as a critical regulatory mechanism in HCC progression. Sirtuins (SIRTs), a class III histone deacetylases (HDACs) that require NAD⁺ as a cofactor, exhibit exclusive and poised functions in the pathophysiological processes of cancers by mediating ferroptosis. However, a summary of the mechanisms of SIRT-mediated ferroptosis in HCC development and associated therapeutic strategies is limited. This article provides an outline of recent developments in the role of SIRTs in HCC proliferation, metastasis, and the development of multidrug resistance. We highlight the roles and mechanisms of SIRTs in mediating ferroptosis in HCC, aiming to provide a comprehensive and novel perspective for developing diagnostic biomarkers and therapeutic strategies for HCC, thereby advancing the understanding of HCC pathogenesis and treatment.

Introduction

Primary liver cancer is the sixth most common malignant tumor globally, with its mortality rate ranking third among all cancers. Primary liver cancer is particularly prevalent in the Asia-Pacific region, accounting for nearly 75% of all LC cases and three-quarters of global LC deaths.^1,2 Primary liver cancer is mainly classified into hepatocellular carcinoma (HCC), intrahepatic cholangiocarcinoma (ICC), and mixed HCC-ICC, with HCC being the most common, constituting 90% of liver cancer patients. HCC typically originates from the malignant transformation of hepatocytes in the context of diseased liver tissue.³ By the time HCC patients are diagnosed, the disease has often reached an intermediate or advanced stage. At this point, treatment methods such as surgical resection, radiotherapy, and chemotherapy are ineffective and may cause side effects (organ damage, bone marrow suppression, etc). The cure rate for patients with HCC is 25%, with a 5-year survival rate of only 18%.⁴ In recent years, with the rising incidence of liver-related metabolic diseases, the incidence of HCC has also continued to increase, posing a significant threat to human life and health. Therefore, identifying appropriate treatment methods to improve the survival rate and early recovery potential of cancer patients is crucial.

Cell death is a crucial process in human growth and metabolism and plays a key role in human health and disease.⁵ As a hallmark of cancer, resisting cell death affects cancer progression at various stages. Cell death can be classified into accidental cell death (ACD) and programmed cell death (PCD).⁶ PCDs, including classical apoptosis and other novel forms (such as necroptosis, pyroptosis, autophagy, ferroptosis), rely on specialized molecular machinery. Ferroptosis, a unique form of PCD, was proposed in 2012 by the laboratory of Brent R. Stockwell.⁷ It often exhibits environment-dependent duality. Under physiological conditions, ferroptosis participates in maintaining tissue homeostasis and normal physiological activities; however, its dysregulation is closely associated with the occurrence and progression of various diseases, including cancer, either by inhibiting tumor growth through the clearance of abnormal cells or by promoting cancer cell survival and metastasis to drive tumor progression.⁸ The hallmarks of ferroptosis mainly include iron overload, accumulation of lethal lipid peroxides in the cell membrane (ultimately leading to cell death⁹), and ROS accumulation. Ferroptosis is often featured by depletion of intracellular reduced glutathione (GSH), inactivation of glutathione peroxidase 4 (GPX4), and abnormal accumulation of phospholipids containing polyunsaturated fatty acids (PUFAs).^10,11 GSH is a small peptide composed of three amino acids (glutamic acid, cysteine, and glycine) residues and exists in two forms: reduced GSH and oxidized glutathione (GSSG). GSH is the most abundant intracellular antioxidant and is capable of scavenging ROS within cells under the catalysis of GPX4. GSH is negatively correlated with ferroptosis, while GSSG is positively correlated with ferroptosis. The three main regulatory mechanisms of ferroptosis are as follows: 1) Core suppressive pathways of ferroptosis include the GPX4-dependent antioxidant system, such as the System xc⁻-GSH and GPX4 axes; GPX4-independent mechanisms, such as the FSP1-CoQ10 and GCH1-BH4 axes. 2) Regulation of the labile iron pool: This involves modulation of ferritin (FTH/FTL), ferritinophagy mediated by NCOA4, and Fe²⁺ uptake via transferrin and its receptor. 3) Metabolic regulation of membrane polyunsaturated fatty acid phospholipids (PUFA-PLs) as substrates for lipid peroxidation: Key enzymes involved are ACSL3, ACSL4, LPCAT3, and lipoxygenases (LOXs).^12–16

Ferroptosis acts as a “double-edged sword” in cancer, exerting context-dependent dual functions. It is one of the inhibitory factors in HCC. Numerous studies have shown that ferroptosis suppresses the growth and proliferation of HCC cells in in vitro xenograft models.^17,18 Liang et al conducted a TCGA cohort analysis on HCC patient samples and reported that more than 70% of ferroptosis-related genes were differentially expressed in HCC tissues and peri-tumoral tissues compared with normal tissues.¹⁹ In specific contexts, the activation of ferroptosis may promote cancer progression. Dai et al found that in mouse models of pancreatic cancer harboring the oncogene Kras, GPX4 depletion or feeding with a high-iron diet accelerates the formation of pancreatic intraepithelial neoplasia (PanIN) and shortens mouse survival. However, treatment with the ferroptosis inhibitor liproxstatin-1 significantly suppresses the accelerated tumor progression induced by Gpx4 deficiency or a high-iron diet, thereby extending mouse survival. Ferroptosis activates the STING pathway by releasing oxidation-damaged associated molecules (such as 8-OHG), inducing macrophage infiltration and inflammatory responses, thereby creating a favorable microenvironment for tumorigenesis.²⁰ This dual role highlights the crucial hub position of ferroptosis in cancer progression and treatment.

Sirtuins (SIRTs), known as “longevity proteins”, are widely distributed in cells and belong to the class III histone deacetylases (HDACs) dependent on NAD⁺. Mammals have seven subtypes of SIRTs, including SIRT1-7, which are classified based on their subcellular localization: 1) SIRT1, SIRT6, and SIRT7 are located primarily in the nucleus; 2) SIRT2 is distributed mainly in the cytoplasm; 3) SIRT3, SIRT4, and SIRT5 are found predominantly in the mitochondrial matrix.²¹ The primary function of SIRTs is to deacetylate and modify proteins, maintain their functionality and stability, and participate in various cellular processes, such as cell cycle regulation and metabolic regulation. They play significant roles in energy metabolism, apoptosis, and aging.^22,23 The regulatory roles of SIRTs in ferroptosis differ in liver disorders. In non-alcoholic fatty liver disease (NAFLD), treatment with C. indicum inhibits SIRT1 expression, reduces lipid accumulation and Fe²⁺ content, promotes GPX4 expression, and therefore suppresses ferroptosis. Inhibiting ferroptosis is beneficial in NAFLD as it alleviates hepatic steatosis.²⁴ Targeting SIRT1 shows a converse outcome in HCC. SIRT1 knockdown significantly increases reactive oxygen species (ROS), MDA, and Fe²⁺ levels, while markedly reducing HepG2 cell viability. In this context, SIRT1 inhibition promotes ferroptosis, which shows beneficial effects by preventing HCC progression.²⁵ In summary, these findings suggest that SIRT-mediated ferroptosis regulates liver disease progression in an environment-dependent manner.

Although existing studies have confirmed the regulatory role of ferroptosis in HCC and the involvement of SIRTs in ferroptosis modulation, lacking a comprehensive summary of the regulatory mechanisms of the entire SIRT family in ferroptosis during HCC progression. This review summarizes the classification of SIRTs, the functions and mechanisms of distinct SIRT isoforms in regulating ferroptosis during tumor development, and focuses on the impact of SIRT‑mediated ferroptosis modulation on HCC progression. We also discuss the context‑dependent duality of ferroptosis in HCC and the potential application value of targeting the SIRT‑ferroptosis pathway in the diagnosis and treatment of HCC, aiming to provide a novel theoretical basis and research direction for HCC therapy.

Roles and Mechanisms of SIRTs in Cancer Development Through the Regulation of Ferroptosis

SIRTs are well-known NAD⁺-dependent HDACs and constitute an important family of seven members (SIRT1-7).²⁶ Studies have shown that abnormal expression of SIRTs occurs in almost all cancer types and affects cancer metabolism, genomic stability, and the tumor microenvironment (TME) through various mechanisms.^27,28 Numerous studies have revealed that SIRTs can influence tumor progression by regulating ferroptosis (Figure 1).

Figure 1.

The pathway of ferroptosis mediated by SIRTs in cancer. Abnormal expression of SIRTs is observed in many cancer types, and SIRTs modulate ferroptosis through various mechanisms, influencing cancer progression. By Figdraw.

SIRT1

SIRT1 participates in regulating the progression, invasion, and therapeutic response of cancer cells by binding to the substrate NAD⁺ to remove acetylated lysine residues on target proteins such as p53.²⁹ The inhibition of SLC7A11 and GPX4 activity induces lipid peroxidation and cell death, and the downregulation of SLC7A11 and GPX4 expression can serve as a key marker for inducing ferroptosis. SIRT1 can influence the expression of SLC7A11 and GPX4 to regulate ferroptosis.^30,31 Regulating the expression of SIRT1 can significantly affect tumor progression by influencing ferroptosis. Compared with that in normal lung cell lines, SIRT1 expression is downregulated in various lung cancer cell lines. The SIRT1 activator SRT2104 increases the production of ROS and superoxide anions, reduces the GSH/GSSG ratio, induces lipid peroxidation, and inhibits cell proliferation by activating SIRT1 activity.³² After treatment with the ferroptosis inducer RSL3, Chen et al reported that SIRT1 is activated by upregulating AROS expression, promoting the upregulation and nuclear translocation of ATF3, and inhibiting the activity of ATF3-dependent SLC7A11 and GPX4, thereby exacerbating ferroptosis in glioma cells.³³ After SIRT1 was knocked down, p53 protein levels significantly increased in gastric cancer (GC) cells, but SLC7A11 and GPX4 levels decreased. APE1 overexpression reversed these results, indicating that SIRT1/APE1 inhibits ferroptosis to promote GC cell growth.³⁴ Combined administration of ropivacaine and cisplatin inhibited SIRT1 expression, reduced cell viability and migration, decreased mitochondrial membrane potential, promoted ROS accumulation and apoptosis in LOVO and LOVO/DDP cells, downregulated SLC7A11 and GPX4 expression, suppressed Nrf2/Keap1 pathway activation, and enhanced ferroptosis and antitumor properties. Ropivacaine increases the sensitivity of CRC cells to cisplatin by inhibiting SIRT1 expression to promote ferroptosis.³⁵ In some tumors, such as lung cancer and gliomas, SIRT1 expression is downregulated, and promoting ferroptosis by activating SIRT1 can inhibit tumor progression. In contrast, in GC and colorectal cancer, SIRT1 expression is upregulated, and inhibiting SIRT1 can promote ferroptosis to suppress tumor progression. SIRT1 exerts divergent roles in tumors, and this contradictory effect is attributed to different tumor backgrounds. Additionally, recent studies have shown that SIRT1-mediated ferroptosis also regulates the development of liver cancer, melanoma, acute and chronic myeloid leukemia, head and neck cancer, breast cancer, osteosarcoma, and lymphoma.^36–43

SIRT2

SIRT2 is a crucial member of the sirtuin family that is abnormally expressed in various tumors and plays a significant regulatory role in ferroptosis. In multiple myeloma (MM) patients, SIRT2 expression is markedly elevated. Knockdown of SIRT2 promotes Fe²⁺ generation, increases ROS accumulation, inhibits the protein expression of the ferroptosis markers SLC7A11, GPX4, and FTH1, increases ACSL4 protein expression, and induces ferroptosis to suppress the malignant progression of MM.⁴⁴ Glutamate-cysteine ligase consists of a catalytic subunit (GCLC) and a regulatory subunit and plays a vital role in GSH synthesis. Both acetylation and succinylation modifications can occur at lysine sites on target proteins. In tumor cells, SIRT2 can catalyze the desuccinylation of GCLC, thereby promoting GSH synthesis and inhibiting ferroptosis. SIRT2 knockout induces a reduction in GSH levels, increasing tumor cell susceptibility to ferroptosis.⁴⁵

SIRT3

SIRT3 is localized in mitochondria, but under cellular stress conditions, such as various stimuli, including ultraviolet irradiation and chemotherapy, SIRT3 can be translocated from mitochondria to the nucleus, where it performs NAD+-dependent deacetylation functions.⁴⁶ SIRT3 regulates the metabolism of various mitochondria, including glycolysis and respiration, fatty acid oxidation, and ROS generation.^47–49 SIRT3 exerts tumor-suppressive effects by promoting cancer cell death through the induction of ferroptosis. For example, in patients with gallbladder cancer (GBC), the expression level of SIRT3 is lower in tumor tissues than in adjacent normal tissues, and low SIRT3 expression is associated with poor prognosis in GBC patients. Knockdown of SIRT3 in GBC cell lines promotes mitochondrial respiration and energy metabolism while inhibiting ROS production. Silencing SIRT3 suppresses AKT-dependent ferroptosis. In SIRT3-knockdown cells, AKT phosphorylation levels increase, whereas ACSL4 levels decrease. Inhibiting AKT activity promotes ACSL4 expression and ferroptosis. These results indicate that SIRT3 inhibits AKT-dependent mitochondrial metabolism and EMT, thereby promoting ferroptosis and suppressing tumor progression.⁵⁰ SIRT3 overexpression reduces CRC cell viability; increases intracellular ROS, MDA, and Fe²⁺ levels; suppresses SLC7A11 expression; and activates the AMPK/mTOR pathway. Compared with SIRT3 knockdown alone, restoration of SLC7A11 expression blocked the effects of SIRT3 knockdown on ferroptosis induction and cell viability inhibition. Additionally, the use of AMPK or mTOR inhibitors also suppressed the promoting effects of SIRT3 on cell viability and ferroptosis. SIRT3 triggers SLC7A11-mediated ferroptosis in HCT-116 cells, reducing cell viability and inhibiting CRC progression through activation of the AMPK/mTOR pathway.⁵¹

However, SIRT3 is functioned as an oncogene in certain cancers.⁵² SIRT3 expression is higher in glioblastoma (GBM) tissues than in normal brain tissues. After SIRT3 knockdown, the levels of Fe²⁺ and ROS in mitochondria increase, whereas the levels of GSH, ATF4, and SLC7A11 decrease. All groups were treated with RSL3, and supplementation with SLC7A11 in response to SIRT3 knockdown restored cysteine uptake, increased GSH levels, and enhanced cell survival. Additionally, overexpression of ATF3 in response to SIRT3 knockdown significantly increased SLC7A11 expression. SIRT3 regulates SLC7A11 transcription through ATF4, inhibiting ferroptosis and promoting GBM progression.⁵³ Furthermore, modulating SIRT3 expression can influence breast cancer progression through the regulation of ferroptosis.^54,55

SIRT4

SIRT4 is associated with various mitochondrial dysfunction-related diseases, including nonalcoholic diseases, type 2 diabetes, fatty liver disease, apoptosis, inflammation, vascular diseases, and various cancers.⁵⁶ Compared with that in normal tissues, SIRT4 expression is downregulated in CRC tissues. Through clinicopathological feature correlation analysis, SIRT4 was linked to adverse outcomes, such as tumor invasion depth, lymph node metastasis, lymphatic invasion, and distant metastasis.⁵⁷ In GC, low SIRT4 expression is associated with an increased incidence of malignant tumors.⁵⁸ Additionally, SIRT4 expression is downregulated in various cancers, including prostate cancer, breast cancer, and cervical cancer.^59–61

SIRT4 has been found to be a ferroptosis modulator in several non-tumor diseases. SIRT4 is a critical regulator of ferroptosis and modulates cytokine expression and oxidative stress levels to control ferroptosis. For example, SIRT4 regulated inflammation and oxidative stress through the HIF-1α/HO-1 pathway, inhibiting ferroptosis and alleviating severe acute pancreatitis.⁶² In LIRI mice and AML12 cells subjected to hypoxia–reoxygenation injury, SIRT4 expression is downregulated. Knocking out SIRT4 led to repressed expression of GPX4 and SLC7A11 but elevated expression of ACSL4. Furthermore, SIRT4 knockout significantly increased the level of acetylated PRDX3, whereas SIRT4 overexpression produced the opposite effects. This study suggested that SIRT4 inhibits ferroptosis by suppressing PRDX3 acetylation, thereby alleviating LIRI.²⁴

Although the regulatory effects of SIRT4-mediated ferroptosis in cancers remain unclear, several studies have found that SIRT4 plays a role in regulating glutamine metabolism in several cancers, including prostate cancer, breast cancer, and cervical cancer.^59–61 For example, SIRT4 overexpression disturbed proliferation, migration, and invasion abilities and contributed to apoptosis in prostate cancer cells. Meanwhile, SIRT4 restrained glutamine metabolism in tumor cells.⁵⁹ In breast cancer, SIRT4 knockout promoted self-renewal of tumor cells by enhancing glutamine metabolism.⁶⁰ Similarly, SIRT4 exerted inhibitory effects on glutamine metabolism in cervical cancer by inhibiting glutaminase 1 (GLS1) expression.⁶¹ Considering that disturbed glutamine metabolism often contributes to ferroptosis in cancers,^12–16 it is believed that SIRT4 potentially regulates cancer development by affecting ferroptosis.

SIRT5

SIRT5 is present in almost all human tissues and organs.²⁸ SIRT5 is located primarily in the mitochondrial matrix and is expressed at low levels in the mitochondrial membrane, nucleus, cytoplasm, and peroxisomes.^63,64 SIRT5 plays dual roles in cancer, either as a tumor suppressor or as a tumor promoter. In GC, METTL14 expression was reduced. Its overexpression inhibited the malignancies of GC cells and enhanced the stability of SIRT5 mRNA through m6A modification. Meanwhile, METTL14 overexpression increased ROS, Fe²⁺, and MDA levels, but inhibited the GSH content, SOD activity, and GPX4 expression. Those results suggested that METTL14 induced ferroptosis in GC cells. Knockdown of SIRT5 partially reversed METTL14-mediated ferroptosis in GC cells. Collectively, METTL14 promotes ferroptosis and inhibits GC progression through SIRT5 upregulation.⁶⁵ Wang et al⁶⁶ designed a succinylated D-type neuropeptide integrated with an iron-based probe, which can disrupt the mitochondrial membrane potential and electron transport through SIRT5-mediated desuccinylation. Hence, this functional material promoted apoptosis, inflammation, and ferroptosis in tumor cells. This study suggests that SIRT5 is an antitumor target.

However, SIRT5 can also promote cancer development. SIRT5 expression was upregulated in gliomas. SIRT5-mediated desuccinylation of BCAT1 at lysine K39 inhibited its interaction with E3 ubiquitin ligases, thereby preventing BCAT1 degradation via the ubiquitin‒proteasome system. Therefore, the SIRT5/BCAT1 axis promotes glioma progression and reduces sensitivity to ferroptosis.⁶⁷

SIRT6

SIRT6 is primarily localized in the nucleus and plays a critical role in metabolism, inflammation, DNA repair, and the regulation of cancer development.^68–70 Notably, SIRT6 influences tumor progression and chemotherapy resistance by regulating ferroptosis. NCOA4, an autophagy-related receptor for ferritin, binds to ferritin and promotes its autophagic degradation. The overexpression of SIRT6 induced NCOA4-dependent ferritinophagy, thereby increasing Fe²⁺ levels and promoting ferroptosis in anaplastic thyroid cancer cells.⁷¹ Compared with that in non-tumor cells, the expression of SIRT6 in colorectal cancer (CRC) cells was reduced. Following SIRT6 overexpression, total iron content and MDA, 4-HNE, and ROS levels increased, whereas expressions of GPX4, FTH1, and SLC7A11 were decreased. The overexpression of SIRT6 also reduced the phosphorylation levels of mTOR and STAT3. SIRT6 enhanced autophagy-dependent ferroptosis in colorectal cancer cells by inhibiting the mTOR/STAT3 pathway.⁷² In nasopharyngeal carcinoma (NPC), SIRT6 knockdown increased cell viability, reduced the levels of MDA, ROS, total iron, and Fe²⁺, and increased the level of GSH. Concurrently, SIRT6 knockdown upregulated the expression of the ferroptosis markers FTH1 and GPX4 while downregulating ACSL4 and NCOA4 expression. In NPC, SIRT6 knockdown suppressed ferroptosis and promotes tumor progression.⁷³ These findings suggest that SIRT6 can inhibit tumor progression by promoting ferroptosis.

SIRT6 can also promote tumor progression by inhibiting ferroptosis. For example, in thyroid cancer, SIRT6 knockdown downregulated GPX4 expression. Overexpression of SIRT6 upregulated GPX4 expression, which inhibited ferroptosis.⁷⁴ In lung cancer cells, SIRT6 knockdown reduced the survival rate of drug-resistant cells, with the concentrations of intracellular Fe²⁺, MDA, and ROS significantly increasing, while the levels of GSH and NRF2 and GPX4 expression were decreased. Overexpression of SIRT6 upregulated NRF2 and GPX4 expression. Therefore, those results indicated that SIRT6 regulates ferroptosis in lung cancer cells through NRF2/GPX4 activity and reverses drug resistance.⁷⁵ SIRT6 was upregulated in sorafenib-resistant GC cells. SIRT6 knockdown significantly reduced the survival rate of sorafenib-resistant GC cells, promoted sorafenib-induced accumulation of ROS, Fe²⁺, and MDA, and downregulated the expression of GSH and GPX4. Additionally, SIRT6 knockdown significantly upregulated Keap1 expression and downregulated Nrf2 expression. It could be concluded that downregulating SIRT6 promotes GPX4 downregulation, enhances ferroptosis, and suppresses sorafenib resistance in GC cells by regulating Keap1/Nrf2.⁷⁶

SIRT7

SIRT7, the most recently identified member of the sirtuin family, is primarily localized in the nucleus. A distinct feature that differentiates it from other SIRTs is its unique ability to participate in global RNA transcription, splicing, and stability.⁷⁷ SIRT7 can regulate metabolism and stress responses.⁷⁸ H2O2 significantly induces melanocyte death, whereas SIRT7 expression increases in melanocytes in response to H2O2 stimulation. SIRT7 may play a role in resistance to oxidative stress-induced ferroptosis, and its activation in vitiligo melanocytes might be impaired. After SIRT7 knockdown, SLC7A11 and SLC3A2 expression is downregulated, ACSL1 and TFR1 expression is upregulated, and cell mortality is significantly increased. SIRT7 inhibits ferroptosis to protect melanocyte survival in vitiligo.⁷⁹ SIRT7 alleviates renal fibrosis, injury, and dysfunction by regulating the KLF15/Nrf2 signaling pathway, thereby mitigating renal cell ferroptosis.⁸⁰ These results indicate that SIRT7 is among the key regulators of ferroptosis.

SIRT7 is upregulated in liver, pancreatic, and colorectal tumors (compared with that in healthy individuals).^81–83 Autophagy in cancer cells protects them from various stresses, and when autophagy is inhibited, the accumulation of ROS leads to mitochondrial damage. After SIRT7 was knocked down, the mitochondrial membrane potential in cervical squamous cell carcinoma (CSCC) cells significantly decreased, and ROS levels significantly increased. SIRT7 inhibits ROS accumulation in CSCC cells by promoting autophagy.⁸⁴ Following SIRT7 overexpression, ROS levels in human lung cancer A549 cells are reduced.⁸⁵ ROS content is closely related to ferroptosis.¹³ SIRT7 may influence tumor development by mediating ferroptosis, but the specific mechanisms require further investigation.

SIRTs Regulate Ferroptosis to Influence the Progression of Liver Diseases

The liver is a vital metabolic organ. When liver metabolism is abnormal or severely damaged and difficult to recover, severe steatosis and hepatocyte injury occur. Prolonged poor control can further evolve into inflammation and fibrosis, ultimately leading to cirrhosis, HCC, and end-stage liver disease.^86,87 In recent years, ferroptosis has been shown to participate in the pathogenesis and progression of metabolic diseases and even liver fibrosis through elevated levels of peroxidation biomarkers such as MDA and 4-HNE, as well as changes in iron content.^88,89 The SIRT family influences iron metabolism and lipid peroxidation homeostasis to mediate ferroptosis, thereby regulating liver diseases.

SIRTs-Mediated Ferroptosis and Liver Injury

SIRTs regulate ferroptosis to influence liver injury. For example, compared with normal mice, ethanol-fed mice exhibit elevated hepatic MPO activity. MPO is an enzyme present in neutrophils and monocytes and is involved in killing pathogens and regulating inflammatory responses. After intestine-specific SIRT1 deficiency, hepatic MPO activity was decreased, and the expression of the neutrophil marker Ly6G and the inflammatory markers TNF-α, iNOS, and IL-1β were all repressed in the liver. Additionally, SIRT1 deficiency led to reduced hepatic MDA, enhanced GSH contents, and downregulated ferroptosis-associated genes in the liver, which included Ncoa4, Acsl4, DPP4, Cot1, Rpl8, and ATP5G3. This study suggested that targeting SIRT1 in the intestine alleviated ethanol-induced liver damage by inhibiting ferroptosis.⁷ SIRT1 also shows protective effects in liver injuries. Compared with that in the normal group, SIRT1 expression in the acetaminophen (APAP)-induced liver injury group was downregulated. Treatment with forsythiaside A upregulated SIRT1 expression, increased expression of SLC7A11 and GPX4, but decreased ACSL4 expression. Higher SIRT1 expression increased the intracellular GSH content and reduced the MDA, Fe²⁺, and ROS levels. SIRT1 enhances cellular antioxidant capacity, reduces lipid peroxide accumulation, and inhibits ferroptosis. High SRIT1 expression decreases AST and ALT levels. SIRT1 mitigates APAP-induced liver injury by reducing lipid peroxide accumulation and inhibiting ferroptosis.⁹⁰

In addition to SIRT1, other SIRTs have also been found to mediated ferroptosis in liver injury. In the thioacetamide (TAA)-induced liver failure model, SIRT2 expression is upregulated. Inhibition of SIRT2 upregulates the expression of MFN2 and GPX4, reduces intracellular Fe²⁺ and ROS levels, inhibits ferroptosis, and alleviates liver injury.⁹¹ In liver injury models, downregulation of SIRT3 promotes ferroptosis. Activation of SIRT3 upregulates the expression of FTH1, FTL, GPX4, and SLC7A11, while downregulating TFRC expression, thereby reducing intracellular Fe²⁺ and ROS levels and mitigating liver injury.⁹² Furthermore, SIRT4 inhibits ferroptosis by suppressing PRDX3 acetylation, thereby alleviating LIRI.²²

SIRTs, Ferroptosis, Liver Fibrosis and Metabolism-Associated Liver Disease

SIRTs regulate ferroptosis to influence liver fibrosis and metabolism-associated liver diseases. For example, SIRT2 deacetylates USP2 at Lys447 to inhibit its activity. TGFβ1 and ferroptosis are key drivers of organ fibrosis. Specifically, USP2 is a deubiquitinating enzyme, where the USP2a isoform deubiquitinates TGFBR, recruiting SMAD2/3, which thereby promotes TGF-B signaling. SIRT2-mediated inhibition of USP2 expression suppresses ferroptosis and alleviates liver fibrosis.⁹³ Persistent injury or oxidative stress triggers the EMT in hepatocytes, which transform them into myofibroblasts, leading to excessive ECM deposition and ultimately liver fibrosis.⁹⁴ Gong et al reported that in the context of induced nonalcoholic steatohepatitis (NASH) combined with type 2 diabetes (T2D), the expression of the ferroptosis markers GPX4, SLC7A11, and FTH decreased, whereas that of ferroptosis increased. However, after ferroptosis was inhibited with Fer-1, EMT in hepatocytes is reduced as evidenced by an increased expression of E-cad and a reduced expression of canonical mesenchymal markers (vimentin and a-SMA), and the AST and ALT levels decreased, alleviating NASH- and T2D-induced liver fibrosis. Compared with that in the normal group, SIRT4 expression in the group with NASH combined with T2D was reduced. SIRT4 functions as an endogenous analogue of Fer-1 by inhibiting ferroptosis and reversing EMT, thereby alleviating persistent liver injury, ECM deposition and liver fibrosis.⁹⁵ To validate the association between SIRT1 and metabolism-associated fatty liver disease (MAFLD), a high-fat, high-fructose diet (MAFLD) model was established. Compared with a normal diet, a HFHFD resulted in reduced expression of SIRT1 and PGC-1A, along with elevated levels of MDA and Fe²⁺. Akkermansia muciniphila (A. muc) is a beneficial bacterium. Compared with the HFHFD, A. muc treatment increased SIRT1 and PGC-1A expression, elevated intestinal and liver SCFA levels, and upregulated the SCFA receptors Ffar2 and Ffar3. Concurrently, the expression of the ferroptosis markers GPX4 and SLC7A11 increased, whereas the expression of MDA and Fe²⁺ decreased, indicating that ferroptosis was suppressed. A. muc alleviates ferroptosis by inhibiting PUFA synthesis via SIRT1/PGC-1A, improving MAFLD.⁹⁶

Effects of SIRTs on HCC Progression and Treatment

HCC is among the most common malignant tumors worldwide, and its development and progression are closely related to abnormalities in epigenetic regulation.⁹⁷ Histone deacetylation is one of the modes of epigenetic modification. SIRTs play a significant role in metabolic reprogramming, genomic stability, and stress responses in HCC through the effects of deacetylation. In HCC, SIRTs influence the proliferation and growth, migration, invasion and metastasis, and cancer stemness—malignant characteristics of HCC (Figure 2 and Table 1).

Figure 2.

Effect of SIRT1-7 on the Malignant Phenotype of HCC. In HCC, SIRTs influence the proliferation and growth, migration, infiltration, and metastasis of HCC, as well as the cancer cell dryness. By Figdraw.

Table 1.

Expression of SIRTs in HCC

SIRTs	Control Group	Experimental Group	Expression	Function	Prognosis	References
SIRT1	Normal	HCC patients	High	Prognosis, proliferation and growth, migration, invasion, and metastasis, cancer stemness, therapy resistance	Poor	[83,90,94,98]
SIRT2	Normal tissue	Human HCC samples	High	Prognosis, proliferation and growth, migration, invasion, and metastasis, therapy resistance	Poor	[84,91,95]
SIRT3	Normal tissue	Human HCC samples	Down	Prognosis, proliferation and growth, migration, invasion, and metastasis, therapy resistance	Poor	[85,92,96]
	Early-stage HCC patients	Late-stage HCC patients
SIRT4	Normal tissue	Human HCC samples	Down	Prognosis, proliferation and growth, Migration, invasion, and metastasis	NA	[86,93]
	Adjacent paracancerous tissues	HCC tissues	High	Invasion, cancer stemness	Poor	[99]
SIRT5	Normal tissue and cell lines	HCC tissues and cell lines	High	Prognosis, proliferation, migration, invasion, therapy resistance	Poor	[87,100]
	Normal tissue	Human HCC samples	Down	Prognosis, proliferation and growth, migration, invasion, and metastasis	Poor	[94]
SIRT6	Normal tissue	Human HCC samples	Down	Prognosis, proliferation, migration, invasion	NA	[88]
	HCC cell lines	HCC cell lines (Drug-resistant)	High	Proliferation, therapy resistance	Poor	[101]
SIRT7	Normal tissue	Human HCC samples	High	Prognosis, proliferation, migration, invasion, and metastasis, therapy resistance	Poor	[98,102,103]

Proliferation and Growth

SIRTs regulate the proliferation of HCC cells and tumor size. SIRT1-overexpressing and knockdown-transfected cells were subcutaneously implanted into nude mice to establish xenograft tumor models. After SIRT1 overexpression, tumor weight and volume significantly increased. Conversely, tumor weight and volume significantly decreased in HCC mice with SIRT1 knockdown. Additionally, flow cytometry revealed an increased proportion of HCC cells in the G0/G1 phase, a decreased proportion in the G2/M phase, and a corresponding reduction in the proportion of S-phase cells. In functional experiments, SIRT1 knockdown significantly impaired the proliferative capacity of HCC cells.¹⁰⁴ In the TCCG HCC sample cohort, compared with that in normal liver tissues, SIRT2 expression was significantly elevated in HCC tumor samples, and patients with higher SIRT2 expression had poorer prognoses. In both the MET/CAT- and AKT/NRAS-induced HCC models, SIRT2 knockdown mice exhibited improved survival rates, reduced liver volume, and fewer tumors. SIRT2 knockdown increased the survival rate of mice with HCC and inhibited tumor growth.¹⁰⁰ SIRT3 overexpression attenuated the proliferative activity of HepG2 cells. SIRT3 can inhibit the proliferation of HCC cells.¹⁰¹ SIRT4 knockdown enhanced the proliferative activity of HCC cells. Compared with the low SIRT4 expression group, the high SIRT4 expression group had a reduced tumor volume. The high SIRT4 expression group also exhibited decreased Ki67 positivity and significantly decreased tumor proliferation levels.¹⁰⁵ SIRT5 is upregulated in HCC tissues and cell lines, and higher SIRT5 expression indicates lower overall survival. In functional experiments, SIRT5 overexpression weakened the proliferative capacity of Hep3B cells. SIRT5 knockdown enhanced the proliferative capacity of Hep3B cells. SIRT5 can inhibit the proliferation of HCC cells.⁹⁸ SIRT6 overexpression reduced the proliferative capacity of HepG2 cells. Conversely, SIRT6 knockdown significantly enhanced the proliferative capacity of HepG2 cells. Further validation in revealed that SIRT6 knockdown resulted in increased tumor volume in mouse experiments.¹⁰⁶ SIRT7 knockout attenuated the formation of transplanted tumors, with tumor volume significantly reduced.⁹⁹ SIRTs play complex regulatory roles in HCC, with SIRT1, SIRT2, and SIRT7 promoting tumor proliferation, whereas SIRT3, SIRT4, SIRT5, and SIRT6 primarily suppress cell proliferation and restrict tumor growth.

Migration, Invasion, and Metastasis

SIRTs regulate the migration, invasion, and metastasis of HCC cells. After treatment with the SIRT1 inhibitor EX527, SIRT1 expression was decreased, and the profiles of p53, acetylated p53, Bax, cleaved-caspase 3, cleaved-caspase 9, and cytochrome c were increased. The migratory capacity of HepG2 cells was significantly inhibited after EX527 treatment. This inhibition was accompanied by a reduction in the levels of migration-associated MMP-2, uPA, and MMP-9, as well as increased expression of E-cadherin, which are associated with tumor invasion. By contrast, treatment with the SIRT1 activator SRT1720 significantly increased SIRT1 levels and promoted the migration, invasion, and metastasis of HCC cells.¹⁰² Salermide, a SIRT2 inhibitor, significantly inhibited the migration of HepG2 and Hep3B cells. Salermide treatment or SIRT2 knockdown markedly promoted the expression of E-cadherin.¹⁰⁷ Those two studies indicate that inhibiting SIRT1 or SIRT2 can disturb the migration/invasion of HCC cells.

Paxillin, a substrate of JNK and one of the main kinases involved in focal adhesion dynamics, mediates enhanced metastasis in tumors. SIRT3 overexpression inhibited JNK activation through repressing Paxillin phosphorylation. Functional experiments demonstrated that SIRT3 overexpression suppressed HCC cell migration and invasion. These data indicate that SIRT3 suppresses HCC cell invasion and migration by regulating the production of active paxillin and cell motility through the JNK pathway.¹⁰⁸ In HCC cells, SIRT4 deficiency significantly enhanced cell migration and spheroid formation ability. Moreover, SIRT4 deficiency in mice markedly promoted tumor formation, increased tumor weight and volume, and significantly shortened the lifespan of HCC-bearing mice. Conversely, SIRT4 overexpression significantly reduced cell proliferation, migration, spheroid formation ability, and tumor size while increasing mouse survival.^105,109 SIRT5 overexpression inhibited migration and invasion in HCC cells. In HCC-bearing mice, SIRT5 overexpression reduced lung metastasis and suppressed pulmonary metastasis. Additionally, SIRT5 knockdown enhanced the invasiveness of the MHCC-97H cell line.¹¹⁰ SIRT6 knockdown resulted in the formation of numerous metastatic nodules in the lungs and pancreas of mice. In orthotopic primary tumors from HCC-bearing mice, E-cadherin expression was downregulated, while vimentin and fibronectin expressions were upregulated. SIRT6 knockdown increased tumor invasion and metastatic susceptibility.¹⁰⁶ SIRT7 expression was elevated in HCC tissues and positively correlated with disease progression at all stages except stage IV. Functional experiments revealed that SIRT7 knockdown significantly reduced the migratory ability of Huh7 and HepG2 cells. SIRT7 knockdown markedly inhibited lung metastasis in HCC-bearing mice. Conversely, SIRT7 overexpression led to a significant increase in lung metastasis. In HCC cells, SIRT7 overexpression induces significant downregulation of E-cadherin expression and upregulation of vimentin expression, promoting HCC migration and metastasis.¹¹¹ Multiple studies have indicated that SIRTs play critical regulatory roles in the malignant processes of metastasis, migration, and invasion in HCC. Changes in SIRT activity or expression levels can significantly affect downstream signaling pathways and effector proteins, providing potential molecular targets for targeted therapy in HCC.

Therapy Resistance

Although multiple chemotherapeutic agents have demonstrated significant antitumor effects during the initial treatment phase of HCC, a substantial proportion of advanced HCC patients fail to derive long-term benefits from systemic therapy because of primary and acquired drug resistance,¹¹² whereas SIRTs can influence drug resistance in HCC cells. For example, in functional experiments, upregulation of SIRT1 expression promotes HCC cell proliferation. To further validate the correlation between SIRT1 expression and doxorubicin resistance, all cells were treated with doxorubicin. After SIRT1 overexpression, the survival rate of HCC cells significantly increased, and the proportion of cells in the subG1 fraction significantly decreased. SIRT1 overexpression enhances chemoresistance by inhibiting apoptosis.¹¹³ Donafenib reduces SIRT2 phosphorylation by downregulating the cyclin E/CDK2 complex, downregulates p300, reduces SIRT2 autoacetylation, and further activates SIRT2 activity. SIRT2 leads to decreased mTOR phosphorylation and activity, thereby promoting enhanced autophagic flux and the formation of protective autophagy, resulting in donafenib resistance. The combination of the SIRT2-specific inhibitor TM with donafenib enhances the inhibitory effect of donafenib on HCC cells, increases the expression of cleaved PARP and caspase-3, suppresses protective autophagy, and increases the sensitivity of HCC cells to donafenib.¹¹⁴ After sorafenib treatment, SIRT3 expression decreases in HCC cells. SIRT3 overexpression significantly reduces the proliferation rate of Huh7 cells. SIRT3 knockdown significantly increases the proliferation rate, and SIRT3 knockdown significantly reduces the sensitivity of HCC cells to sorafenib.¹¹⁵ SIRT5 knockdown exacerbates sorafenib-induced apoptosis in HepG2 cells, whereas SIRT5 overexpression inhibits cell death. SIRT5 promotes autophagy by catalyzing the deacetylation of LDHB. After SIRT5 knockdown, the LC3II/LC3I ratio decreased, p62 expression increased, and LDHB acetylation levels increased. SIRT5 knockdown inhibits autophagy and increases HCC cell sensitivity to sorafenib. SIRT5 enhances HCC cell resistance to sorafenib by activating autophagy.¹⁰³ Treatment with chemotherapeutic agents (doxorubicin, cisplatin, and sorafenib) significantly increased SIRT6 expression. All cells were treated with chemotherapeutic agents. After SIRT6 knockdown, the proliferation rate of HCC cells significantly decreases, and their sensitivity to chemotherapeutic agents increases. Additionally, flow cytometry has shown that the apoptosis rate of HCC cells significantly increases after SIRT6 knockdown.¹¹⁶ SIRT7 expression is upregulated in sorafenib-resistant HCC cells. SIRT7 knockdown alone only moderately reduced cell survival and growth, but the combined use of SIRT7 and sorafenib significantly inhibited the survival and proliferation of sorafenib-resistant HCC cells. In mice with HCC, the combination of sorafenib and a SIRT7 inhibitor significantly suppressed tumor growth rate and mass. The combination of sorafenib and a SIRT7 inhibitor reduces Ki-67 and AFP positivity rates.¹¹⁷ These results indicate that inhibition of SIRT1, SIRT3, SIRT6, and SIRT7 activity or expression enhances the sensitivity of HCC cells to treatments such as sorafenib, whereas SIRT2 and SIRT5 mediate HCC cell resistance through mechanisms such as the promotion of protective autophagy. Targeted specific interventions against SIRTs may serve as a potential strategy to improve the efficacy of targeted therapies for liver cancer.

Cancer Stemness

Cancer stem cells (CSCs) possess tumorigenic, self-renewal, metastatic, and chemoresistant capabilities and are widely recognized as driving factors in HCC progression. Understanding the molecular mechanisms that drive cancer stemness in HCC is crucial for developing targeted therapies that can effectively eliminate these aggressive cancer cells and improve patient outcomes. CSC markers such as CD90, EPCAM, CD133, NANOG, SOX2, OCT4, and CD44 are highly expressed in HCC and are associated with stemness.¹¹⁸ The identification and characterization of specific markers associated with HCC CSCs provide opportunities for the development of novel diagnostic and therapeutic strategies. By targeting these specific markers, it may be possible to selectively inhibit CSCs, preventing tumor recurrence and metastasis.¹¹⁹ After SIRT1 knockdown, the proportion of NANOG-positive liver CSCs decreased. Following SIRT1 knockdown, the expression of the stemness markers NANOG, SOX2, OCT4, CD13, CD44, and EpCAM significantly decreased, and the number of CSCs in the S phase markedly decreased. CSCs can form large colonies, and this clonogenic activity is considered a key indicator of an undifferentiated state. In SIRT1-knockdown CSCs, both the size and the number of colonies were reduced, and the spheroid formation capacity was significantly inhibited. SIRT1 knockdown in CSCs suppressed their self-renewal and tumorigenic capabilities.¹²⁰ Tumor-initiating cells (TICs) resemble stem cells and are capable of self-renewal and generating a hierarchical organization of heterogeneous cancer cells. These cells are resistant to conventional chemotherapy and radiotherapy and are thus responsible for sustaining tumor growth and recurrence.¹²¹ After SIRT4 knockdown, the expression of TIC-associated molecular markers, including ABCG2, BMI1, and NANOG, was significantly suppressed, and their spheroid-forming ability and tumorigenicity markedly decreased. Conversely, SIRT4 overexpression led to increased expression of the TIC-associated markers ABCG2, BMI1, and NANOG but significantly increased spheroid formation capacity.¹²² The maintenance and progression of CSC/TIC stemness depend on key regulatory molecules such as SIRT1 and SIRT4. Inhibiting SIRT1 and SIRT4 expression downregulates stemness marker expression, significantly impairing the self-renewal and tumorigenic abilities of CSCs/TICs. Targeting SIRTs offers a potential novel therapeutic strategy for selectively eliminating CSCs and suppressing HCC recurrence and metastasis.

Regulatory Role of SIRTs in Ferroptosis in HCC

The occurrence and development of HCC are closely related to complex metabolic reprogramming. Ferroptosis is an Fe²⁺ dependent, novel form of programmed cell death characterized by excessive accumulation of ROS and plays a critical role in the progression and therapeutic resistance of HCC. In recent years, an increasing number of studies have revealed that SIRTs influence HCC progression by regulating ferroptosis (Figure 3).

Figure 3.

The regulatory role of SIRTs in ferroptosis in HCC. SIRTs regulate cellular ferroptosis through pathways such as SIRT1/YY1/GPX4, PCDH20/SIRT1/NRF2, SIRT6/ACSL5, and SIRT5/ACSL4/ALOX15, thereby inhibiting the malignant progression of HCC. By Figdraw.

Du et al discovered through TCGA screening that YY1 is a transcriptional activator of GPX4. The SIRT1 inhibitor Ex-527 increased the protein levels of YY1 and GPX4. Previous studies have shown that YY1 can be deacetylated by SIRT1.¹²³ In functional experiments, Ex-527 inhibited SIRT1 expression, increased cell colony formation, and reduced ROS levels. In mouse in vivo experiments, upregulation of SIRT1 expression led to decreased tumor weight and volume, as well as downregulation of YY1, Ac-YY1, and GPX4 expression. SIRT1 can deacetylate YY1, leading to its protein degradation, reducing GPX4 transcriptional levels, promoting ferroptosis, and thereby inhibiting HCC progression.³⁶ The results of Jun et al are opposite to those of Du et al, where SIRT1 acts as an oncogene that inhibits ferroptosis. After SIRT1 overexpression, the expression of GPX4 and NRF2 in SUN-387 cells increased, and the levels of intracellular iron, MDA, and DCF decreased. Concurrently, the survival rate and proliferative capacity of HCC cells were enhanced. Coimmunoprecipitation confirmed that SIRT1 deacetylates NRF2 to increase its activity. NRF2 can also induce the expression of iron metabolism-related genes, thereby upregulating the expression of downstream targets such as ferritin to maintain iron balance.^124,125 This finding is consistent with the increased expression of SLC7A11 observed after SIRT1 overexpression. These results suggest that SIRT1 may increase NRF2 activity through acetylation, inhibit ferroptosis, and promote HCC progression.¹²⁴

Based on current literature, we suggest that SIRT1 is not a simple tumor suppressor or oncogene in HCC, but rather an environment- and substrate-dependent bifunctional regulatory protein. SIRT1 exerts tumor suppressor activity by promoting ferroptosis and inhibiting HCC through the YY1-GPX4 axis; conversely, it exerts oncogenic activity by inhibiting ferroptosis and promoting tumor growth through the NRF2-SLC7A11/GPX4 axis. However, current research on this topic is limited, and further studies are needed to validate the functions of SIRT1.

Through immunofluorescence experiments, Xue et al reported that ACSL4 signaling was enhanced and SIRT5 signaling was elevated, with the upregulation of SIRT5 positively correlated with ACSL4 induction. Coimmunoprecipitation experiments confirmed the endogenous interaction between SIRT5 and ACSL4 in Hepa1-6 cells. After treatment with the SIRT5 inhibitor ET-29, the expression of SIRT5 decreased. The SIRT5 inhibitor ET-29 suppressed the expression of ACSL4, LPCAT3, and ALOX15; increased the expression of GPX4 and SLC7A11. SIRT5 promoted and stabilized the expression of ACSL4, which mediated LPCAT3/ALOX15 to trigger ferroptosis and inhibit the progression of HCC.¹²⁶

Ma et al reported that ACSL5 promotes increases in lipid ROS, MDA, and Fe²⁺ contents. SIRT6 can deacetylate H3K9 and H3K56, which are crucial for regulating genes involved in various biological processes, including lipid metabolism.^127,128 After SIRT6 overexpression, the mRNA level of ACSL5 was suppressed. Conversely, SIRT6 knockdown reduced the deacetylation of H3K9 and H3K56, thereby increasing their enrichment on the ACSL5 promoter and upregulating ACSL5 mRNA expression. These results demonstrate that SIRT6-mediated deacetylation of H3K9 and H3K56 inhibits ACSL5 transcription and reduces susceptibility to ferroptosis.^127–129

The specific functions of SIRTs are associated with their subcellular location. SIRT1 and SIRT6 are localized in the nucleus, while SIRT5 is located in the mitochondria. The specific subcellular localization directly determines their substrate spectrum and functional specificity. Nuclear-localized SIRTs primarily target histones and nuclear transcription factors, thereby regulating chromatin structure and gene transcription. In this section, we found that SIRT1 and SIRT6 exert deacetylase activity in the nucleus, modulating the transcriptional levels of downstream genes to regulate ferroptosis. For example, SIRT1 deacetylates YY1, leading to its protein degradation and reduced transcriptional level of GPX4. SIRT1 deacetylates NRF2 to enhance its activity, inducing the expression of iron metabolism-related genes. SIRT6 deacetylates H3K9 and H3K56, inhibiting ACSL5 transcription. SIRTs localized in the cytoplasm and mitochondria primarily regulate metabolic enzymes and signaling molecules, participating in cellular metabolism and stress responses. For instance, SIRT5 promotes and stabilizes the expression of ACSL4, mediating the expression of LPCAT3/ALOX15.

In-depth elucidation of the regulatory effects of SIRT family proteins on ferroptosis not only provides new perspectives for understanding the pathophysiological processes of HCC but also lays the foundation for the development of novel anticancer strategies targeting the SIRT-ferroptosis axis.

Strategies Targeting SIRTs for HCC Treatment

Strategies targeting the SIRT family for HCC treatment are now hot research topics in precision therapy for liver cancer. SIRTs play crucial roles in the occurrence and development of liver cancer by regulating the acetylation status of proteins, such as by controlling cancer cell proliferation, migration, and metastasis and maintaining the “stemness” of cancer cells. By specifically modulating the activity of specific SIRT subtypes and maintaining epigenetic stability, the malignant progression of HCC can be effectively suppressed. For example, small-molecule compounds such as resveratrol and metformin can influence the activity of SIRT subtypes and inhibit HCC (Table 2).

Table 2.

Drugs Targeting SIRTs for the Treatment of HCC

Drug	Type	SIRTs	Mechanism	Possible Pathway	References
Resveratrol	Polyphenolic	SIRT1	Inhibit HCC cell proliferation and migration, increase the content of free iron and reactive oxygen species (ROS), and suppress GPX4 expression	SIRT1-ferroptosis	[129,130]
Oxymatrine	Alkaloids	SIRT1	Inhibit HCC cell proliferation, increase Fe2+, ROS, and MDA levels, and suppress the expression of SLC7A11 and GPX4	SIRT1/YY1/GPX4	[40,131]
Arachidonic Acid	Omega-6 polyunsaturated fatty acids	SIRT5	Inhibited the proliferation and invasion of HCC cells, increased the levels of Fe2+, ROS, and MDA, decreased the GSH level, and reduced the expression of GPX4 and SLC7A11.	SIRT5/ACSL4/LPCAT3/ALOX15	[124,132,133]
Metformin	Chemical synthesis category	SIRT1	Inhibit HCC cell proliferation, invasion, and migration; reduce GPX4 expression; promote ACSL4 expression; and increase ROS levels in cells	AMPK/SIRT1	[134,135]
Quercetin	Flavonoids	SIRT1	Inhibit HCC cell proliferation, reduce the expression of GPX4, FTL, and FTH, and increase ROS and free iron content	SIRT1/AMPK	[136,137]

Resveratrol is an activator of SIRT1. Studies have shown that the histone deacetylase SIRT1 regulates EMT by activating ZEB1 function.¹³⁰ SIRT1 knockdown or Ex-527 treatment reduced the expression of ZEB1 and vimentin but increased the expression of E-cadherin. Resveratrol treatment produced the opposite results, significantly reducing cell viability and increasing free iron and ROS levels. Resveratrol activates SIRT1, promotes ferroptosis, and inhibits the proliferation of head and neck cancer cells.³⁸ Resveratrol can upregulate the expression and enzymatic activity of SIRT1, inhibit the proliferation and migration of HepG2 cells, suppress the PI3K/AKT pathway, reduce FoxO3a phosphorylation, and induce apoptosis.¹³⁸ During cancer progression, the levels of antioxidant enzymes such as catalase, glutathione peroxidase, glutathione reductase (GR), the antioxidant GSH, and detoxification pathway enzymes such as GSH S-transferase decrease. Resveratrol restored catalase and GSH peroxidase levels to normal levels in alcohol-aflatoxin B1-induced HCC. Resveratrol regulates the activity of antioxidant enzymes by modulating SIRT1 expression, thereby inhibiting HCC.¹³⁹ The therapeutic mechanism of Resveratrol in HCC may involve SIRT1-mediated ferroptosis.

Oxymatrine (OMT) is an HCC inhibitor. OMT can inhibit tumor growth. Immune checkpoint inhibitors (ICIs), such as anti-PD-L1 and anti-PD-1 antibodies, can enhance antitumor immunity.¹³¹ Aberrantly upregulated PD-L1 promotes immune escape by inhibiting the tumor-killing effect of cytotoxic T lymphocytes. The regulation of PD-L1 involves multiple mechanisms, among which IFN-γ induction has been demonstrated to be a major source of PD-L1 in many tumors.¹⁴⁰ OMT reverses the inhibitory effects of IFN-γ on Fe²⁺, ROS, and MDA in HepG2 cells, suppresses the expression of SLC7A11 and GPX4, promotes the occurrence of ferroptosis, and inhibits HCC.¹⁴¹ OMT is also an activator of SIRT1. By activating SIRT1, OMT promotes cell death and inhibits proliferation in HCC cells while downregulating YY1 and GPX4 levels. Upon inhibition of SIRT1, OMT-induced ferroptosis is reversed. Furthermore, OMT suppresses HCC growth through the SIRT1/YY1/GPX4 axis. These results indicate that OMT inhibits cell viability and induces ferroptosis in HCC cells, involving the regulatory mechanism of the SIRT1/YY1/GPX4 axis.³⁶

Arachidonic acid (AA) is a typical ω-6 polyunsaturated fatty acid.¹³² The peroxidation of AA-containing phospholipids (PLs) has been shown to induce ferroptosis in tumor cells. Lipid peroxidation can be triggered by ROS and lipoxygenases (such as ALOX15). ALOX15 is considered to be associated with lipid ROS generation and serves as a marker of ferroptosis.¹³³ Wen et al reported that OIT3 overexpression increased Fe2, MDA, and ROS levels, promoting ferroptosis. Arachidonic acid can act as a substrate for ferroptosis. OIT3 overexpression significantly increased AA content and ALOX15 expression, thereby activating the arachidonic acid metabolic pathway. OIT3 overexpression markedly suppressed the growth of HepG2 cells and reduced tumor weight and volume in HCC mice but also decreased GPX4 levels in HepG2 cells. By upregulating ALOX15 expression and increasing AA content, OIT3 activated arachidonic acid metabolism, promoted ferroptosis, and inhibited HCC.¹³⁴ ACSL4 is an enzyme responsible for esterifying long-chain PUFAs into phospholipids. Liu et al reported that in HCC, after FSP1 inhibition, ACSL4 phosphorylation increased, leading to a significant increase in oxidized products of arachidonic acid and promoting AA oxidation. The exogenous addition of AA increased FSP1 inhibitor-induced ferroptosis, as AA is the primary substrate for lipid peroxidation. Increased lipid peroxidation promoted ferroptosis. ACSL4 knockdown reduced lipid ROS production and cell death, thus suppressing ferroptosis. The inhibition of FSP1 activated ACSL4, promoted AA oxidation, accelerated ferroptosis, and inhibited HCC.¹³⁵ AA treatment inhibited HCC cell proliferation and invasion; induced ferroptosis; increased Fe²⁺, ROS, and MDA levels; and reduced GSH levels. Additionally, AA upregulated the expression of SIRT5, ACSL4, LPCAT3, and ALOX15. AA promoted ferroptosis in HCC cells. AA treatment dose-dependently increased SIRT5, ACSL4, LPCAT3, and ALOX15 levels but decreased GPX4 and SLC7A11 levels. Quantitative fluorescence analysis revealed that SIRT5 upregulation was positively correlated with ACSL4 induction. ET-29 cotreatment reversed the AA-induced changes and alleviated the ferroptosis phenotype caused by AA treatment. AA administration resulted in significant dose-dependent suppression of tumor growth. AA promotes ferroptosis in HCC cells by activating the SIRT5/ACSL4/LPCAT3/ALOX15 pathway.¹²⁶

Metformin is a ferroptosis activator. After metformin treatment, the proliferation and survival rates of HCC cells, as well as their invasion and migration, significantly decreased, the half-maximal inhibitory concentration (IC50) of sorafenib significantly decreased, and the sensitivity to sorafenib increased. The expression of the drug resistance-related proteins ABCG2 and P-GP was significantly reduced. Moreover, after metformin treatment, the expression of GPX4 decreased, promoting the expression of ACSL4 and leading to increased ROS levels and elevated lipid peroxidation in cells. These results indicate that metformin promotes ferroptosis and inhibits HCC.¹³⁶ In NAFLD, metformin treatment increased AMPK phosphorylation and SIRT1 expression in hepatocytes. Further validation of the regulatory relationship between AMPK and SIRT1 revealed that combined treatment with the AMPK inhibitor CC resulted in reduced AMPK phosphorylation and decreased SIRT1 expression. Combined treatment with EX527 did not significantly alter AMPK phosphorylation but did reduce SIRT1 expression. Metformin activates SIRT1 activity by promoting AMPK phosphorylation.¹³⁷ SIRT1 is associated with ferroptosis.³¹ Metformin may mediate ferroptosis through the AMPK/SIRT1 pathway, thereby inhibiting HCC.

Quercetin belongs to the flavonoid family and is among the most important dietary antioxidants. It has been shown to exhibit anticancer effects in various tumors.^142–144 Quercetin stimulates SIRT1 deacetylase activity approximately 25-fold at a concentration of 2 µM, indicating significant activation of SIRT1.¹⁴⁵ After quercetin treatment, the protein levels of SIRT1 and phosphorylated AMPK significantly increased in A549 cells. The SIRT1 inhibitor EX527 was combined with quercetin to treat A549 and H1299 cells. Compared with that in cells treated with quercetin alone, the activity of the SIRT1/AMPK pathway in cells treated with quercetin alone was significantly inhibited. Quercetin can activate the SIRT1/AMPK pathway.¹⁴³ Quercetin promotes HepG2 and Hep3B cancer cell death in a dose-dependent manner but has no significant inhibitory effect on the survival of normal liver cells (L-02). After quercetin treatment, the expression of GPX4, FTL, and FTH decreased, whereas SLC27A4 expression increased. Concurrently, ROS and free iron levels increase. Quercetin promotes ferroptosis-induced HCC cell death.¹⁴⁴ SIRT1/AMPK may play a critical role in the inhibition of ferroptosis by quercetin in HCC.

Limitations

This article systematically summarizes the regulatory role of SIRTs-mediated ferroptosis in HCC. However, it still has several limitations: (1) Existing studies have validated the mechanisms of SIRTs on GPX4/System Xc⁻, but neglecting their non-classical pathways in lipid metabolic reprogramming and mitochondrial function regulation, as well as the role of SIRTs-mediated ferroptosis in tumor stem cells and the TME, along with different immune cell subsets, which remains to be elucidated. (2) Most current research evidence derives from preclinical studies, such as mouse subcutaneous xenograft models and HCC cell lines, lacking clinical translation evidence. Considering the inherent tumoral heterogeneity in HCC, the regulatory roles of SIRTs-mediated ferroptosis in HCC progression and associated therapeutic strategies for HCC therapy have not been verified in patient-derived xenograft models and primary liver cancer models. Additionally, the correlation between SIRTs expression, ferroptosis, and HCC progression have not been validated through prospective cohort studies. Future research should integrate clinical translation data to deeply clarify the regulatory mechanisms of SIRTs in the heterogeneous background of HCC, thereby providing a more solid theoretical basis for HCC treatment targeting SIRTs-mediated ferroptosis.

Conclusion and Future Perspectives

SIRTs are abnormally expressed in various tumors, and their ability to mediate ferroptosis can play a key regulatory role in multiple tumors. In HCC, SIRTs have essential roles in regulating malignant behaviors, angiogenesis, and immunosuppressive TME through multiple mechanisms. This review summarizes the roles of SIRT-mediated ferroptosis in HCC. Strategies targeting SIRTs can alleviate liver damage and liver fibrosis and suppress the progression of HCC. These findings highlight the broad prospects of SIRT-mediated ferroptosis in HCC therapy.

Although ferroptosis mediated by SIRTs provides novel therapeutic targets for HCC, current research still faces challenges. Firstly, the malignant characteristics of HCC also include high cellular heterogeneity in the TME, particularly among tumor cells and immune cells. Tumor-associated macrophages (TAMs) are the most abundant immune cells in the TME; M1-type macrophages exhibit anti-tumor properties, whereas M2-type macrophages promote tumor progression.¹⁴⁶ The M2 polarization of TAMs can influence GPX4-dependent ferroptosis through fatty acid metabolic reprogramming.¹⁴⁷ SIRT5 is downregulated in human primary HCC samples. In SIRT5-deficient mice, SIRT5 deficiency synergizes with oncogenes to stimulate nuclear receptors via the bile acid pathway, promoting M2-like macrophage polarization and facilitating tumor progression.¹⁴⁸ However, the role of SIRT-mediated ferroptosis in the TME of HCC remains unclear. It is necessary to study the complex interactions between different immune cell subsets and tumor cells, as well as context-dependent switching mechanisms, to overcome the limitations of immunotherapy in HCC.

Secondly, the functions and mechanisms of SIRT-targeted small-molecule compounds in HCC have not been fully elucidated. For example, whether resveratrol (a Sirt1 activator) exerts its anti-cancer effects through mediating ferroptosis requires further experimental validation. During the clinical translation of SIRT-targeted natural products or small-molecule compounds, challenges remain due to their poor bioavailability, rapid metabolism, the first-pass effect potential drug interactions. Therefore, developing strategies for the precise delivery of SIRT-targeted chemicals is of great importance.

Moreover, we should pay special attention to the toxic side effects of traditional ferroptosis-inducing drugs. Zhao et al injected Erastin into normal mice and observed altered blood indices, leading to mild cerebral infarction and increased glomerular volume in the kidneys.¹⁴⁹ Moreover, intrinsic and acquired ferroptosis resistance in HCC cells severely undermines the therapeutic efficacy of single-agent tumor drugs, resulting in drug resistance and poor patient prognosis. Combining conventional therapies (chemotherapeutic agents/immunotherapeutic agents) with ferroptosis inducers (eg, erastin) and SIRT modulators (eg, SIRT 1, 3, 5, 7 agonists/antagonists) can promote ferroptosis in tumor cells, overcome tumor drug resistance, reduce drug dosages, and mitigate toxic side effects. Furthermore, developing functionally engineered materials based on SIRT modulators, such as designing selective SIRT activators or inhibitors that act only in specific HCC cell subsets or TME microenvironments and are targeted for delivery to tumor sites, could provide more references for precise targeted therapy of HCC.

Funding Statement

This study is supported by the Self-funded Project of Science and Technology Plan of Baoding (No.2241ZF319) and Research Fund of Affiliated Hospital of Hebei University (No. 2025Q14).

Abbreviation

AA, Arachidonic Acid; A. Muc, Akkermansia muciniphila; APAP, N-acetyl-para-aminophenol; CSCs, Cancer stem cells; GBC, Gallbladder Cancer; GBM, Glioblastoma; GC, Gastric Cancer; GCLC, Glutamate-cysteine ligase catalytic; GR, Glutathione Reductase; GSH, Reduced glutathione; GSSG, Oxidized glutathione; HCC, Hepatocellular carcinoma; HDACs, Histone deacetylases; HFHFD, High fat and high fructose diet; LncRNA, Long noncoding RNA; MAFLD, Metabolic-associated fatty liver disease; miRNA, microRNA; MM, Multiple Myeloma; NASH, Non-alcoholic steatohepatitis; NPC, Nasopharyngeal carcinoma; OMT, Oxymatrine; PanIN, Pancreatic intraepithelial neoplasia; PUFAs, Polyunsaturated fatty acids; ROS, Reactive oxygen species; SIRTs, Sirtuins; TAA, Thioacetamide; TAM, Tumor-Associated Macrophages; T2D, Type 2 Diabetes; TIC, Tumor-initiating cell; TME, Tumor microenvironment.

Data Sharing Statement

The labeled dataset used to support the findings of this study are available from the corresponding authors upon request.

Author Contributions

All authors made a significant contribution to the work reported, whether that is in the conception, study design, execution, acquisition of data, analysis and interpretation, or in all these areas; took part in drafting, revising or critically reviewing the article; gave final approval of the version to be published; have agreed on the journal to which the article has been submitted; and agreed to be accountable for all aspects of the work.

Disclosure

The authors declare that they have no competing interests in this work.