The exosomal circRNAs derived from HCC cells and adjacent non-cancerous cells might have different mechanisms of action (Shi et al., 2020). Hsa-circ-0051443, which is highly expressed in normal cells, can be transferred to HCC cells via exosomes. It can competitively bind to miR-331-3p leading to decreased expression of BCL2 antagonist/killer 1(BAK1), thus inhibiting HCC cell growth and promoting apoptosis. Moreover, the level of hsa-circ-0051443 could differentiate between patients with HCC and non-HCC individuals, with an area under the curve (AUC) of 0.8089 (Chen et al., 2020). However, further clinical validation is required for accurate diagnosis.

Macrophages play a well-established role in HCC, and understanding their mechanism is essential for HCC treatment (Tian et al., 2019). Recent studies have demonstrated that exosomes derived from macrophages with high expression of recombination signal-binding protein Jk (RBPJ) can effectively inhibit the progression of HCC, and further investigation has revealed that exosomes carry hsa-circ-0004658, which is abundantly expressed in macrophages overexpressing RBPJ. This circRNA sponges miR-499b-5p, leading to an upregulation of junctional adhesion molecule 3 (JAM3). As a result, HCC cell proliferation is inhibited, and apoptosis is promoted (Zhang et al., 2022a). However, the results still need to be validated in a clinical cohort. In addition, it should be noted that not all exosomal circRNAs derived from non-cancerous cells have inhibitory effects on HCC. CircWDR25 facilitated HCC cell proliferation and invasion by regulating the circWDR25/miR-4474-3p/arachidonate 15-lipoxygenase (ALOX15) and EMT axes, in which it enhances the expression of cytotoxic T-lymphocyte associated protein 4 (CTLA-4) in HSCs and programmed cell death 1 ligand 1 (PD-L1) in HCC cells, thereby influencing the immune response in the TME and ultimately affecting HCC prognosis and recurrence (Liu et al., 2022).

Understanding the mechanisms of cell cycle regulation is crucial in studying the transformation of normal cells into malignant tumors, because abnormal cell proliferation plays a vital role in this process (Fernández et al., 2002). Researchers have discovered that prolonged exposure to arsenic trioxide increases the exosome levels of circRNA-100284 and facilitates its transfer to normal LO-2 cells, in which the circRNA interacts with miRNA-217, leading to accelerated cell cycle progression and promotion of hepatocellular malignancy (Dai et al., 2018). Similarly, circ-002136 is upregulated in HCC tissues and cells and can be transferred via cancer cell-derived exosomes. It promotes HCC cell proliferation, migration, and invasion via the miR-19a-3p/Rab GTPase YPT1 homolog (RAB1A) pathway, is associated with poor prognosis, and correlates positively with disease staging (Yuan et al., 2022). However, it's worth noting that the study lacked clinical validation.

Exosomal has-circ-0061395 is also significantly upregulated in the serum of patients with HCC, and silencing its expression led to cell cycle arrest and promotion of apoptosis. It interacts with the 3′untranslated region of miR-877-5p, resulting in increased expression of phosphatidylinositol 3-kinase regulatory subunit gamma (PIK3R3), thereby promoting HCC cell proliferation, migration, invasion, and malignant behavior (Yu et al., 2021). Circ-0072088 is widely enriched in exosomes derived from HCC cells and can be secreted into surrounding normal cells through blood circulation, promoting cell transfer. It has a high diagnostic value in patients with HCC (AUC: 0.899), and high levels of circ-0072088 are associated with poor prognosis. Knocking out circ-0072088 abolished the sponging of miR-375 and downregulated matrix metalloproteinase 16 (MMP-16) expression, thus inhibiting HCC cell invasion and metastasis (Lin et al., 2021). CircANTXR1 is highly abundant in exosomes isolated from the serum of patients with HCC and plays a crucial role in promoting HCC cell proliferation, migration, invasion, and tumorigenicity. It directly interacts with miR-532-5p, which in turn regulates the expression of X-ray repair cross-complementing 5 (XRCC5) to facilitate HCC development and worsen prognosis. Inhibiting circANTXR1 could potentially serve as a novel therapeutic approach for HCC and as an early diagnostic marker (Huang et al., 2021). Matrix metalloproteinase 2 (MMP2) is closely associated with metastasis and malignant tumor dissemination (Zhu et al., 2017a). A recent study found that exosomal circMMP2 (has-circ-0039411) can be transmitted to normal liver cell line L02, acting as a ceRNA to enhance the expression of MMP2 by sequestering miR-136-5p, thereby promoting the metastasis of HCC cells (Liu et al., 2020). Conversely, suppressing the expression of circMMP2 reversed the aggressive behavior of L02 cells, and its increased expression is associated with a lower overall survival rate in patients. Exosomal circ-0006602 is specifically expressed in the plasma of patients with HCC and enhances the expression of proteins associated with tumor proliferation in HCC cell lines; its diagnostic performance surpasses that of AFP, with an AUC of 0.907 compared with 0.694 (Guo et al., 2021). Combining circ-0006602 and AFP greatly improved the early detection rate of HCC (AUC: 0.942).

A previous study reported that cytoplasmic circRNA Cdr1as functions as an oncogene by targeting the miR-7/epidermal growth factor receptor (EGFR) axis to promote HCC development (Yang et al., 2017). Further investigations revealed that Cdr1as is highly abundant in exosomes derived from HCC cells, acting as a ceRNA to promote HCC cell proliferation and migration by sequestering miR-1270 and upregulating AFP expression. In vitro experiments also demonstrated that exosomes derived from HCC cells overexpressing Cdr1as can directly transfer to adjacent normal cells, enhancing their proliferation and invasion abilities (Su et al., 2019). Other animal experiments showed that exosomal circular RNA tubulin tyrosine ligase-like family member 5 (circTTLL5) promotes mouse hepatocellular carcinoma cell proliferation and metastasis through the miR-136-5p/KIAA1522 axis, thereby suppressing tumor growth. This suggests that blocking exosome-mediated circTTLL5 transfer might be a therapeutic target for HCC (Liu et al., 2023a). CircTMEM45A (has-circ-0066659) acts as a sponge for miR-665, regulating the expression of downstream insulin-like growth factor 2 (IGF2) to promote cell migration in vitro and the occurrence of HCC in vivo, and its expression correlates with tumor size, TNM staging, and vascular invasion. Patients with low circTMEM45A expression have longer survival (Zhang et al., 2020c). Another study has revealed that circPTGR1 is abundant in exosomes released by highly metastatic HCC cells, and the presence of high levels of circPTGR1 enhances EMT processes in low or non-metastatic cells through its interaction with miR-449a, thereby disrupting TME homeostasis, and is closely related to the clinical staging of HCC (Wang et al., 2019a). Has-circ-0004001, has-circ-0004123, and has-circ-0075792 were found to be upregulated in HCC serum exosomes according to bioinformatic analysis, and they demonstrated high sensitivity and specificity in distinguishing between healthy controls and patients with HCC. When combined, these three circRNAs achieved an AUC of 0.885 for HCC diagnosis, indicating their potential as biomarkers (Sun et al., 2020). Furthermore, exosomal circAKT3 is associated with a high risk of HCC recurrence and death. Patients with high circAKT3 expression have lower overall survival and disease-free survival rates, suggesting a poor prognosis (Luo et al., 2020).

EMT plays a crucial role in HCC metastasis, and exosomal circRNAs have been found to promote HCC progression by influencing EMT processes (Liu et al., 2022). Overexpression of HUR, an RNA-binding protein involved in maintaining mRNA stability, has been associated with poor prognosis in HCC (Song et al., 2020). Has-circ-0074854 is highly expressed in both HCC cells and tissues, and downregulation of its expression affects the stability of the HUR protein. This attenuates zinc finger E-box binding homeobox 1 (ZEB1) signal transduction in HCC cells, which suppresses malignant behaviors, including cell proliferation, migration, and EMT, and can induce M2 polarization in macrophages, thereby promoting HCC progression (Wang et al., 2021a). There are experiments have revealed that has-circ-0004277 competes to bind with HUR, leading to reduced expression of zona occludens-1 (ZO-1) and upregulation of the EMT-related transcription factor ZEB1 to promote HCC progression (Zhu et al., 2020). Notably, exosomal has-circ-0004277 can be used to diagnose HCC, with a diagnostic area AUC reaching 0.816 (Zhu et al., 2020).

Inducing neovascularization is a crucial step in the invasion and metastasis of HCC, and exosomes released by cancer cells play a significant role in stimulating the formation of new blood vessels through signaling between HCC cells and endothelial cells (Morse et al., 2019). In the stimulation of tumor angiogenesis, exosomes can directly interact with vascular endothelial growth factor (VEGF)/VEGF receptors or act as carriers to transport circRNAs that target downstream molecules, indirectly contributing to the generation of new blood vessels (Zhang et al., 2022b). Huang et al. discovered that circRNA-100338 is highly abundant in exosomes of HCC cells with high metastatic potential, and the mechanism is that it can promote the proliferation of human umbilical vein endothelial cells (HUVECs) and regulates angiogenesis by interacting with neuro-oncological ventral antigen 2 (NOVA2), an RNA-binding protein associated with vascular development and lumen formation. Thus, the level of circRNA-100338 in postoperative HCC serum can predict the occurrence of postoperative metastasis (Huang et al., 2020). Another study reported that exosomal circCMTM3 directly binds to miR-3619-5p and targets the downstream SRY-box transcription factor 9 (SOX9) to promote the proliferation, migration, invasion, and angiogenesis of HUVECs, enhancing their survival capacity and inducing tumor growth. The level of exosomal circCMTM3 also correlates positively with tumor stage and lymph node metastasis (Hu et al., 2021). Besides, exosomal circPAK1 has also been found to promote angiogenesis and metastasis in HCC cells (Hao et al., 2022).

Cytokine Storm is involved in the occurrence and development of many diseases and may be one of the important causes of poor prognosis (Fajgenbaum and June 2020). Studies have found that circRNAs may be involved in the development of cytokine storms, for example, has-circ-0004812 acts on the JAK/STAT, STAT3 signaling pathway through hsa-mir-1287-5p/IL6R, RIG-I axis, and causes a in COVID-19 inflammatory responses (Mohammadisoleimani et al., 2022). Similarly, circRNAs may also contribute to HCC by promoting inflammatory cytokine storm processes. Tumor cells can modify their surface antigens and change the TME of the surrounding tissue, enabling them to evade recognition and attack by immune cells, which ultimately leads to tumor immune escape (Villalba et al., 2013). Programmed cell death receptor 1 (PD-1) is an inhibitory receptor expressed on the surface of active lymphocytes such as T cells, B cells, and NK cells (Greten and Sangro, 2017). Overexpression of PD-1 can lead to dysfunction of these cells and facilitate tumor immune evasion (Gao et al., 2009). In recent years, immune checkpoint blockade therapy, particularly anti-PD-1 therapy targeting the PD-1/PD-L1 axis and immune cell exhaustion, has been extensively used to treat various malignancies, including HCC. It has also been approved as a second-line treatment for advanced HCC (El-Khoueiry et al., 2017). However, less than 20% of patients with advanced HCC respond effectively to anti-PD-1 treatment because of innate or acquired resistance (Zhu et al., 2018). The specific mechanisms underlying resistance to anti-PD-1 immune therapy in HCC are still unclear. Nevertheless, some studies have demonstrated the involvement of exosomal circRNAs in regulating cancer immune evasion, promoting exhaustion of NK cells and CD8⁺ effector T cells, expanding regulatory T cells (Tregs), driving cancer progression, and inducing resistance to anti-PD-1 therapy (Chen et al., 2019).

NK cells play an important role in the antitumor immune response, and T cell immunoglobulin and mucin domain-containing protein 3 (TIM-3) is expressed on the surface of NK cells as an inhibitory molecule. Increased expression of TIM-3 is associated with NK cell dysfunction and death (Zheng et al., 2019). Exosomal circUHRF1 (has-circ-0048677) derived from HCC cells can be delivered to NK cells in the body fluid circulation where it induces NK cell dysfunction and exhaustion by upregulating the expression of downstream target gene TIM3 through adsorption of miR-449c-5p. This promotes cancer immune escape and drives resistance to anti-PD-1 immune therapy in patients with HCC (Zhang et al., 2020a). Therefore, analyzing the expression levels of circUHRF1 can identify patients who are resistant to PD-1 therapy and improve the clinical efficacy in patients with HCC.

CD8⁺ effector T cells also play a key role in the immune system (Greten and Sangro, 2017). Recent studies have shown that circCCAR1 is significantly enriched in exosomes derived from HCC and it promotes immune evasion of cancer cells and resistance to anti-PD-1 immune therapy by acting on the miR-127-5p/WT1 associated protein (WTAP) axis, increasing the stability of PD-1 protein, and causing dysfunction of CD8⁺ T cells (Hu et al., 2023). Additionally, circRNAs in exosomes derived from HCC cells, specifically circGSE1, act on the miR-324-5p/transforming growth factor beta receptor 1 (TGFBR1)/SMAD family member 3 (Smad3) axis to induce expansion of regulatory Tregs and facilitate immune evasion of HCC cells (Huang et al., 2022).

Macrophages can also influence the liver immune response in HCC (Tian et al., 2019). Exosomal circRNAs mediate intercellular communication between macrophages and HCC cells, thereby mediating HCC progression (Han et al., 2019). Depending on the stimuli, macrophages can polarize into two phenotypes: classical (M1) with anti-tumor properties or alternative (M2), with pro-tumor properties. During HCC progression, tumor cells often induce M2 polarization of macrophages to promote tumor growth (Komohara et al., 2016). Research has identified the overexpression of circtMEM181 in HCC tissues, particularly in patients with a poor PD-1 response. The mechanism involves HCC cells delivering circtMEM181 to macrophages via exosomes, where it binds to miR-488-3p and upregulates CD39 expression. This cooperative action between macrophages and HCC cells activates the ATP-adenosine pathway in the HCC TME, which interferes with the proliferation of CD8⁺ T cells, inducing CD8⁺ T cell exhaustion, and promoting resistance to PD-1 antibodies (Lu et al., 2021). As a result, the responsiveness to PD-1 therapy is limited. These findings highlight the potential of exosomal circRNAs in HCC immunotherapy and their potential as therapeutic targets. Furthermore, the ratio of Tregs to CD8⁺ T cells can serve as a predictive marker for the anti-tumor response to PD-1/PD-L1 inhibitors (Huang et al., 2022).

Malignant tumors rely on aerobic glycolysis to generate the substantial amount of energy required for the rapid proliferation of tumor cells; thus, glycolysis plays a significant role in the accelerated progression of malignant tumors (Lunt and Vander Heiden, 2011). Exosomal circRNAs have been found to influence the glycolytic process in HCC. The level of CircFBLIM1 is significantly elevated in the serum exosomes of patients with HCC and HCC cells. It can be transferred to HCC cells through exosomes and acts on the miR-338/low-density lipoprotein receptor-related protein 6 (LRP6) axis, inducing the glycolytic process in HCC (Lai et al., 2023). Another study discovered that in HCC, exosomal circ-ZNF652 can adsorb miR-29a-3p, leading to increased expression of the downstream target gene GUCD1. This, in turn, promotes cancer cell glycolysis and affects malignant phenotypes like proliferation, migration, and invasion. Silencing the expression of circ-ZNF652 could inhibit HCC progression (Li et al., 2020).

Drug resistance is a significant factor contributing to the poor prognosis of HCC, and the dysregulation of extracellular vesicle circRNAs has been identified to play a role in tumor drug resistance (Li et al., 2019). Sorafenib, a Food and Drug Administration (FDA)-approved first-line treatment for advanced HCC, has shown reduced efficacy in recent years because of the emergence of drug resistance, thus limiting its clinical utility (Wen et al., 2022; Zhu et al., 2017b). A study has revealed that circRNA-SORE interacts with Y-box binding protein 1 (YBX1) in the cytoplasm, inhibiting the translocation of YBX1 to the cell nucleus, preventing pre-mRNA processing factor 19 (PRP19)-mediated ubiquitination and nuclear degradation of YBX1, and consequently affecting the expression of downstream target genes controlled by YBX1 (including AKT, RAF1 (encoding Raf-1 proto-oncogene), ERK, MYC (encoding c-Myc), and TGFB1 (encoding transforming growth factor beta 1)). Silencing the expression of circRNA-SORE was demonstrated to enhance the efficacy of sorafenib (Xu et al., 2020a). Lenvatinib, a tyrosine kinase inhibitor, has been approved by the FDA as a targeted therapy drug for advanced HCC alongside sorafenib; however, drug resistance remains a significant obstacle to its clinical use (Personeni et al., 2019). In studies investigating lenvatinib resistance mechanisms, it was discovered that the levels of circPAK1 were significantly increased in extracellular vesicles derived from lenvatinib-resistant HCC cells. The primary mechanism involves circPAK1 competitively binding to the cytoskeletal protein 14-3-3ζ, which promotes the nuclear translocation and inactivation of yes associated protein (YAP) and the Hippo signaling pathway (Hao et al., 2022). The Hippo signaling pathway is an inhibitory pathway primarily responsible for regulating cell size and tumor volume, with YAP serving as a downstream effector molecule. YAP, considered an oncoprotein, is regulated by phosphorylation and its subcellular localization. Phosphorylated YAP interacts with 14-3-3ζ, exits the cell nucleus, and undergoes ubiquitination and degradation mediated by beta-transducin repeat containing E3 ubiquitin protein ligase (β-Trcp). However, when the Hippo signaling pathway is inhibited, YAP remains non-phosphorylated, enters the cell nucleus through the nuclear membrane, interacts with transcription factors, and initiates the transcription of downstream target genes (Zhao et al., 2007). Furthermore, HCC-derived circPAK1 can transfer from extracellular vesicles to lenvatinib-sensitive HCC cells, thereby reducing their drug sensitivity (Hao et al., 2022).

Cisplatin (DDP) is commonly used as a first-line chemotherapy drug for HCC; however, resistance to DDP has a significant impact on the prognosis of patients with HCC (Zhou et al., 2019). Previous studies have demonstrated that CircZFR promotes the progression of HCC by influencing the miR-511/AKT1 axis (Yang et al., 2019). Further investigation into its resistance mechanism revealed that CircZFR is highly expressed in HCC-associated fibroblasts and is abundant in their extracellular vesicles. Through these extracellular vesicles, CircZFR can be transferred to HCC cells, where it inhibits the signal transducer and activator of transcription 3 (STAT3)/nuclear factor kappa B (NF-kβ) signaling pathway, ultimately promoting HCC progression and chemotherapy resistance (Zhou et al., 2022). Transarterial chemoembolization (TACE) is a treatment option for patients with advanced HCCs (Elderkin et al., 2023). In a recent study, researchers examined the variation in the circRNA contents of exosomes during TACE treatment and found that the abundance of exosomal circ-G004213 increased after TACE treatment compared to before treatment. This circRNA acts as a sponge for miR-513b-5p and enhances the expression of pre-mRNA processing factor 39 (PRPF39), thus improving the sensitivity of HepG2 cells to cisplatin and improved survival in vivo. Besides, the circ-G00421 expression level correlated positively with post-TACE prognosis, it may be used as an indicator for predicting the efficacy of TACE (Qin et al., 2021). This research provides a new potential combination treatment strategy to overcome chemotherapy resistance in HCC.