Expression level of TFCP2L1 in liver tissue is inversely correlated with HCC patient survival

To investigate the impact of TFCP2L1 expression on disease prognosis, we analyzed tumor RNA expression data from the TCGA database. Our findings indicated that HCC patients with high TFCP2L1 expression had significantly shorter overall survival compared to those with low expression (Fig. 1A). Moreover, high TFCP2L1 expression showed a trend towards shorter disease-specific survival times (Fig. 1B), suggesting that high TFCP2L1 expression in HCC tissues is associated with poor prognosis in patients. Furthermore, we also investigated TFCP2L1 gene expression in HCC tissues compared to adjacent normal liver tissues from the TCGA dataset. Our findings consistently showed significantly higher levels of TFCP2L1 expression in cancerous tissues. Moreover, paired samples from the same patients confirmed increased TFCP2L1 expression in tumor tissues compared to adjacent non-cancerous liver tissues, highlighting its potential role in HCC progression.

Fig. 1: Expression level of TFCP2L1 in liver tissue is inversely correlated with the survival of HCC patients.

A Kaplan–Meier curve analysis depicting overall survival (OS; p = 0.022) in HCC patients based on high and low TFCP2L1 expression levels in TCGA cohorts. B Kaplan–Meier curve analysis illustrating disease-specific survival (p = 0.085) in HCC patients stratified by high and low TFCP2L1 expression using TCGA cohorts. C Immunohistochemical staining showing TFCP2L1 expression in tumor and adjacent normal liver tissues. D Representative images demonstrating high and low TFCP2L1 expression in tumor tissues. E, F Kaplan-Meier curve analysis presenting overall survival (OS; p = 0.001) and progression-free survival (PFS; p = 0.014) in HCC patients based on high and low TFCP2L1 protein expression levels in immunohistochemical staining (Red arrowhead indicates low nuclear expression; blue arrowhead indicates high nuclear expression). G Quantitative analysis of TFCP2L1 mRNA expression levels in liver tissues from HCC patients by qRT-PCR analysis (n = 16 paired samples, p = 0.0053). H Western blot analysis showing representative TFCP2L1 protein levels in adjacent normal tissues and tumor tissues from 10 HCC patients.

Further analysis of patients with poor prognostic indicators, such as those who had passed away or underwent palliative surgical resection, revealed significantly elevated TFCP2L1 expression in their tumor tissues. To delve deeper into the prognostic implications, we utilized TMAs to assess TFCP2L1 protein expression in 124 surgically treated HCC patients. Consistently, TFCP2L1 protein levels were found to be markedly higher in cancerous tissues compared to adjacent normal liver tissues (Fig. 1C, D). Importantly, patients with high TFCP2L1 protein expression exhibited significantly reduced overall survival and disease-free survival times compared to those with low expression levels (Fig. 1E, F). Multivariate analysis revealed TFCP2L1 as an independent prognostic factor for overall survival in HCC, with its expression also correlating with larger tumor size (Table S1 and S2). Validation studies using HCC tumor tissues and adjacent tissues further confirmed elevated TFCP2L1 expression in cancerous versus non-cancerous tissues (Fig. 1G, H). These findings collectively suggest TFCP2L1’s crucial role in the development and progression of HCC.

TFCP2L1 expression increased in HCC tissues and cells

CSCs are characterized by their ability to self-renew, metastasize, and initiate tumors. To explore the role of TFCP2L1 in liver CSCs, we first analyzed its protein and mRNA expression levels in various HCC cell lines, including Huh7, Hep3B, 97 L, 97H, and SK-Hep-1 (Fig. 2A, B). Subsequently, we assessed the stem cell properties of each cell line by quantifying sphere formation, limiting dilution assay, and examining the expression of stem cell surface markers on sphere cells. Our findings demonstrated a correlation between TFCP2L1 expression levels and the number of spheres formed, as well as an increase in the proportion of CD133/CD90 double-positive cells within the spheres. Specifically, cell lines such as 97 L and SK-Hep-1, which exhibited high TFCP2L1 expression, formed significantly more spheres and higher frequency of stem cells compared to Huh7 cells with low TFCP2L1 expression (Fig. S1A, B). Moreover, the proportion of CD133/CD90 double-positive cells in spheres derived from 97L and SK-Hep-1 cells increased by 6.55 and 5.17 times, respectively, compared to adherent cells, whereas Huh7 sphere cells only increased by 1.8 times (Figs. 2C and S1C–G). These results suggest that elevated TFCP2L1 expression enhances the stemness of HCC cells.

Fig. 2: TFCP2L1 expression is increased in HCC cell spheres.

A Western blot analysis of TFCP2L1 protein expression levels in different HCC cell lines, with β-ACTIN used as a loading control. B qRT-PCR analysis of TFCP2L1 mRNA expression levels in different HCC cell lines. C Quantification of CD133 and CD90 double-positive cells by flow cytometry after culturing different HCC cell lines in tumor sphere-forming medium for 12 days. D–G qRT-PCR analysis showing mRNA expression levels of OCT4, NANOG, c-MYC, and TFCP2L1 in various types of HCC spheres across different passages. H Western blot analysis depicting c-MYC and TFCP2L1 protein expression levels in different HCC cell lines, with GAPDH used as a loading control. I Fold change in TFCP2L1 mRNA expression levels in liver tissues of HCC patients with varying expression levels of CSC genes. The data are presented as mean ± SD of three independent experiments. *p < 0.05; **p < 0.01; ns no significance, compared with the control group.

Next, we further investigated the expression of NANOG, OCT4, and c-MYC, key regulators associated with CSC induction and maintenance, alongside TFCP2L1. Our results indicated significantly higher mRNA levels of these genes in spheres compared to adherent cells, with their enrichment becoming more pronounced over successive passages (Fig. 2D–G). Additionally, we observed increased sphere formation ability and sphere cell diameter with passages (Fig. S2A, B). Protein analysis confirmed elevated levels of TFCP2L1 and c-MYC in spheres (Fig. 2H). To validate our findings in clinical samples, we extracted mRNA from 70 HCC tissue samples and conducted qRT-PCR to assess the expression changes of CSC genes (NANOG, OCT4, c-MYC) and TFCP2L1 relative to normal liver tissues. Samples showing CSC gene mRNA expression more than two times higher than the control group (normal liver tissue) were categorized as the high expression group, while others were classified as the low expression group. Comparison of TFCP2L1 mRNA fold change in tumor tissue versus normal liver tissue revealed significantly higher expression in the high CSC gene expression group compared to the low expression group, with statistical significance (Fig. 2I). Collectively, our findings indicate a positive correlation between TFCP2L1 expression and the stemness of HCC cells, suggesting its involvement in the maintenance and self-renewal of HCC CSCs.

TFCP2L1 overexpression increased HCC cell proliferation, clonogenicity, sphere formation, and invasion abilities

To explore the potential role of TFCP2L1 in HCC tumorigenesis in vitro, we transfected HCC cell lines Hep3B and 97L with a PiggyBac (PB) transposon-based vector expressing TFCP2L1 (Fig. 3A). Our findings revealed that TFCP2L1 overexpression significantly enhanced cell proliferation compared to control cells, as assessed by a CCK8 assay (Fig. 3B). Additionally, TFCP2L1 overexpression in Hep3B and 97 L cells increased clonogenicity, as demonstrated by cell colony formation assays. To evaluate the impact of TFCP2L1 on stem cell-like properties, we conducted sphere formation assays using transfected cells. Our results showed a marked increase in both the number and size of spheres formed by TFCP2L1-overexpressing Hep3B and 97L cells compared to controls (Fig. 3C–F). Furthermore, limiting dilution assays (LDA) indicated that TFCP2L1 overexpression elevated the probability of sphere formation (Fig. S2C). We further investigated the effect of TFCP2L1 on tumor cell invasiveness using transwell assays. The number of migrated cells significantly increased in TFCP2L1-overexpressing Hep3B and 97 L cells (Fig. 3G, H). Given that tumor stemness is often associated with drug resistance, we evaluated the half-maximal inhibitory concentrations (IC50) of Sorafenib in HCC cells overexpressing TFCP2L1. Our results demonstrated a notable increase in the IC50 of Sorafenib in cells overexpressing TFCP2L1 compared to control cells (Fig. 3I).

Fig. 3: TFCP2L1 overexpression promotes the proliferation and clonogenicity of HCC cells.

A Western blot analysis showing the effect of TFCP2L1 overexpression in Hep3B and 97L cell lines. B Cell viability of Hep3B and 97L cells overexpressing TFCP2L1 assessed by CCK-8 assay at indicated time points. C Colony formation assay demonstrating increased clonogenicity of Hep3B and 97 L cell lines with TFCP2L1 overexpression. D Representative images of sphere formation assay in Hep3B and Huh7 cells with TFCP2L1 overexpression. Scale bar = 100 μm. E, F Quantitative analysis of colony formation (E) and sphere formation (F) assays. G Representative images of transwell invasion assay showing enhanced invasion of Hep3B and 97L cells overexpressing TFCP2L1 compared to empty vector control. Scale bar = 100 μm. H Quantitative analysis of cell invasion. I IC50 values of Sorafenib in indicated HCC cell lines after 48 h, evaluated using Compusyn. The data are presented as mean ± SD of three independent experiments. *p < 0.05; **p < 0.01; ns no significance, compared with the control group.

TFCP2L1 knockdown decreased HCC cell proliferation, clonogenicity, sphere formation, and invasiveness

To assess the impact of TFCP2L1 knockdown in HCC cells, we used lentiviral vectors encoding three specific short hairpin RNAs (shRNAs) targeting TFCP2L1 mRNA to infect Hep3B and 97L cells. After selection with appropriate drugs, we confirmed the stable reduction of both TFCP2L1 mRNA and protein levels (Fig. 4A, B). Among the tested shRNAs, TFCP2L1-sh#2 and TFCP2L1-sh#3 demonstrated high knockdown efficiency and were chosen for subsequent experiments. TFCP2L1 knockdown in Hep3B and 97L cells resulted in fewer colonies formed and a reduced proliferation rate compared to the scramble control group (Fig. 4C–E). Moreover, TFCP2L1 knockdown led to decreased numbers and sizes of spheres in the sphere formation assay (Fig. 4F, G). Limiting dilution assays (LDA) further confirmed a lower probability of sphere formation following TFCP2L1 knockdown (Fig. S2D). Additionally, migration assays showed a decrease in the number of migrated cells upon TFCP2L1 knockdown in both cell lines (Fig. 4H, I). These findings collectively indicate that TFCP2L1 plays a critical role in promoting the proliferation, sphere formation, and invasiveness of HCC cells. Moreover, TFCP2L1 emerges as a potential biomarker and promising therapeutic target in HCC. Furthermore, the reduced IC50 of Sorafenib observed in HCC cells following TFCP2L1 knockdown suggests a potential strategy to enhance drug sensitivity in HCC treatment (Fig. 4J).

Fig. 4: TFCP2L1 knockdown decreases the proliferation and clonogenicity of HCC cells.

A qRT-PCR analysis showing the effect of TFCP2L1 knockdown in Hep3B and 97 L cell lines. B Western blot analysis confirming TFCP2L1 knockdown in Hep3B and 97 L cell lines. C Cell viability of Hep3B and 97 L cells with TFCP2L1 knockdown assessed by CCK-8 assay at indicated time points. D Colony formation assay demonstrating reduced clonogenic potential of Hep3B and 97L cell lines with TFCP2L1 knockdown. E Quantitative analysis of colony formation assay. F Representative images of sphere formation assay in Hep3B and 97L cells with TFCP2L1 knockdown. Scale bar = 100 μm. G Quantitative analysis of sphere formation assay. H Representative images of transwell invasion assay showing decreased invasion of Hep3B and 97 L cells with TFCP2L1 knockdown compared to scramble RNA control. Scale bar = 100 μm. I Quantitative analysis of cell invasion assay. J IC50 values of Sorafenib in indicated HCC cell lines after 48 h, evaluated using Compusyn. K Photograph of xenograft tumors derived from modified Hep3B cells at the final time point after subcutaneous injection into nude mice. L Statistical analysis of the final weight of xenograft tumors derived from indicated Hep3B cells at the endpoint. The data are presented as mean ± SD of three independent experiments. *p < 0.05; **p < 0.01; ns no significance, compared with the control group.

TFCP2L1 regulates tumor growth in vivo

In vivo experiments confirmed the role of TFCP2L1 in tumor growth regulation. Subcutaneous implantation of 5 × 106 Hep3B cells overexpressing TFCP2L1 and control (vector) cells into nude mice revealed that after 6 weeks, mice injected with TFCP2L1-overexpressing Hep3B cells developed significantly larger tumors compared to those injected with control cells. Conversely, the tumor volume in the TFCP2L1-knockdown group was notably reduced compared to the scramble control group (Fig. 4K and L). These findings underscore the regulatory impact of TFCP2L1 in vivo.

The N-terminus and CP2-like domain are required for the regulatory function of TFCP2L1

To identify the essential domain of TFCP2L1 required for its regulatory function, we generated four mutant TFCP2L1 proteins lacking the N-terminus (ΔNT), CP2-like domain (ΔCP2), SAM-like domain (ΔSAM), or C-terminus (ΔCT) (Fig. 5A), as previously described. These flag-tagged mutant forms of TFCP2L1 were successfully overexpressed in both Hep3B and 97 L cells (Fig. 5B). Functional assays demonstrated that TFCP2L1ΔSAM and TFCP2L1ΔCT could mimic the effects of full-length TFCP2L1, enhancing migration, colony formation, and sphere formation compared to the vector control group. In contrast, TFCP2L1ΔNT and TFCP2L1ΔCP2 failed to induce clonogenicity and invasiveness in Hep3B and 97 L cells (Fig. 5C–E). These results indicate that TFCP2L1’s regulatory role in HCC cells relies on its N-terminus and CP2-like domain. Moreover, LDA experiments demonstrated that overexpression of TFCP2L1ΔNT and TFCP2L1ΔCP2, but not TFCP2L1ΔSAM and TFCP2L1ΔCT, resulted in loss of the ability seen with full-length TFCP2L1 to increase the probability of spheroid formation (Fig. S2E, F). The statistical analyses of the invasion assay, colony formation assay, and sphere formation assay are shown in Fig. 5F–H.

Fig. 5: The N-terminal domain and CP2 domain are required to promote TFCP2L1 function.

A Schematic representation of TFCP2L1 domains. B Western blot analysis showing overexpression of different TFCP2L1 mutants in Hep3B and 97L cell lines. C Representative images of transwell invasion assay in indicated cell lines, with empty vector as the control group. Scale bar = 100 μm. D Colony formation assay of indicated cell lines, with empty vector as the control group. E Representative images of sphere formation assay in indicated cell lines. Scale bar = 100 μm. F–H Quantitative analysis of cell invasion (F), colony formation (G), and sphere formation (H) assays. The data are presented as mean ± SD of three independent experiments. *p < 0.05; **p < 0.01; ns no significance, compared with the control group.

TFCP2L1 regulates HCC cell proliferation and invasion partially by the NANOG/STAT3 signaling pathway

To elucidate the regulatory mechanism of TFCP2L1 in HCC cells, we conducted western blot analysis to assess the expression of stemness-related proteins and the phosphorylation status of STAT3, AKT, and ERK. TFCP2L1-overexpressing cells demonstrated significantly elevated levels of phosphorylated STAT3 and AKT compared to the control group, while total STAT3 and AKT levels remained unchanged. Conversely, TFCP2L1 knockdown markedly attenuated STAT3 and AKT phosphorylation (Fig. 6A–D). Notably, no significant changes were observed in ERK phosphorylation throughout this process. Furthermore, we observed a concurrent increase in NANOG expression upon TFCP2L1 overexpression, which decreased correspondingly following TFCP2L1 downregulation (Fig. 6E, F). However, the expression levels of CTNNB and c-MYC did not show significant alterations under these conditions, and OCT4 was undetectable by Western blotting in these cells.

Fig. 6: Potential mechanism for the stem cell-promotion activity of TFCP2L1.

A Western blot analysis of total and phosphorylated levels of STAT3, AKT, and ERK after TFCP2L1 overexpression in Hep3B and 97L cells. B Western blot analysis of total and phosphorylated levels of STAT3, AKT, and ERK after TFCP2L1 knockdown in Hep3B and 97L cells. C Quantitative analysis of phosphorylation levels in panel A. D Quantitative analysis of phosphorylation levels in panel B. E Western blot analysis showing expression levels of CTNNB1, c-MYC, and NANOG after TFCP2L1 overexpression in Hep3B and 97L cells. F Western blot analysis showing expression levels of CTNNB1, c-MYC, and NANOG after TFCP2L1 knockdown in Hep3B and 97L cells. G Promoter activity of the NANOG gene measured using Dual-luciferase reporter assay. The cells were transfected with pGL3-basic or various lengths of the 5′-flanking region of the NANOG gene, as indicated. The data are presented as mean ± SD of three independent experiments. *p < 0.05; **p < 0.01; ns no significance, compared with the control group.

Given previous reports indicating that NANOG can independently activate the STAT3 and AKT pathways in various cell types, we hypothesized that TFCP2L1 might regulate NANOG gene transcription in HCC cells by interacting with its promoter. To investigate this hypothesis, we cloned different lengths of the NANOG promoter region into the pGL3 vector (−6k/+1, −2342/+1, −1.5k/+1, and −322/+1) and conducted dual-luciferase reporter assays. Our results indicated that the maximum luciferase activity was observed with the pGL-322 construct, suggesting that the core regulatory site of TFCP2L1 may reside within the −322/+1 region of the NANOG promoter (Fig. 6G). Together, these findings suggest that TFCP2L1 plays a crucial role in activating the STAT3 and AKT pathways, potentially through upregulation of NANOG expression in HCC cells.

Specific inhibitor of TFCP2L1 and Sorafenib combination synergistically targets HCC cells in vitro

To identify a small molecule capable of selectively inhibiting TFCP2L1 function, we conducted a virtual screening of a molecular library containing over 200,000 compounds using Schrödinger software (Fig. 7A). From this screening, we selected the top nine potential compounds for further experimentation (Table S3). Hep3B and 97 L cells were individually treated with these nine compounds, and the expression levels of NANOG and OCT4 were assessed by qRT-PCR (Fig. S3). Among them, compound number 3 (termed Ti3) exhibited significant suppressive effects, reducing both the number and size of spheres compared to the control group (Fig. S4).

Fig. 7: TFCP2L1 specific inhibitor promotes the anti-tumor effects of Sorafenib by suppressing the STAT3/NANOG pathway.

A Structural formula of Ti3, a specific inhibitor of TFCP2L1. B IC50 values of Sorafenib in the indicated HCC cell lines for 48 h with or without Ti3. C IC50 values of Ti3 in the indicated HCC cell lines for 48 h were evaluated using Compusyn. D Cell apoptosis of HCC cell lines treated with Ti3, Sorafenib, and their combination at different concentrations as assessed by flow cytometry. E Quantitative analysis of apoptotic cells in panel D. F Expression of cleaved-PARP, PARP, cleaved-Caspase3, and Caspase3 after treatment with Ti3 or Sorafenib at indicated concentrations in HCC cell lines detected by western blotting. G Representative image of sphere formation assay in indicated cell lines treated with Ti3, Sorafenib, and their combination. Scale bar = 100 μm. H Quantitative analysis of sphere formation in panel G. I Expression of p-STAT3, STAT3, p-AKT, AKT, TFCP2L1, and NANOG after treatment with Ti3 or Sorafenib at indicated concentrations in HCC cell lines detected by western blotting. The data are presented as mean ± SD of three independent experiments. *p < 0.05; **p < 0.01; ns no significance, compared with the control group.

Interestingly, we observed that Sorafenib, a standard drug for HCC, exhibited enhanced anti-cancer activity when combined with Ti3. Our data showed that the half-maximal inhibitory concentrations (IC50) of Sorafenib on HCC cell lines were reduced in the presence of Ti3. Although both Hep3B and 97L cells were sensitive to Sorafenib alone, they demonstrated increased sensitivity when treated in combination with Ti3 (Fig. 7B, C).

To explore the effects of this combination on cell viability inhibition, we subjected HCC cells to different treatments followed by Annexin V-FITC/PI double-staining and flow cytometry analysis. The results showed that the combination treatment group had more Annexin V-positive cells compared to the Sorafenib-only group. Importantly, treatment with Ti3 alone did not significantly increase the number of Annexin V-positive cells compared to the control group (Fig. 7D, E). Annexin V-positive cells include apoptotic and some necrotic cells, suggesting that Ti3 enhances Sorafenib’s inhibition of cell viability, possibly through the combined induction of apoptotic pathways.

Moreover, to further validate these findings, we assessed the expression of apoptosis-related proteins following treatment with Ti3, Sorafenib, or their combination. Our results demonstrated that co-treatment with Ti3 and Sorafenib significantly increased the expression of Cleaved-PARP and cleaved-Caspase-3 in HCC cells compared to treatment with Sorafenib alone. Notably, Ti3 alone did not induce activation of these apoptotic proteins (Fig. 7F). Cleaved-PARP and cleaved-Caspase-3 are active forms of the apoptosis-related proteins PARP and caspase-3, indicating activation of the apoptotic pathway leading to cell apoptosis. These results suggest that Ti3 enhances Sorafenib-induced apoptosis. Additionally, the combined treatment of Ti3 and Sorafenib significantly reduced the number of liver cancer cell spheres compared to treatment with either Sorafenib or Ti3 alone (Fig. 7G, H), indicating that Ti3 sensitizes HCC cells to Sorafenib.

To investigate the underlying mechanism of this synergy, Western blot analysis was performed. The results demonstrated marked inhibition of STAT3 and AKT phosphorylation, as well as reduced expression levels of TFCP2L1 and NANOG in the combination treatment group compared to the Sorafenib-only group (Fig. 7I). These findings suggest that Ti3 enhances the therapeutic efficacy of Sorafenib by suppressing the NANOG/STAT3 signaling pathway and downregulating TFCP2L1 expression in HCC cells.

Sorafenib combined with Ti3 gained better tumor inhibitory efficiency for HCC compared with the monotherapy

To investigate the synergistic impact of combining Ti3 with Sorafenib against HCC in vivo, we established a subcutaneous xenograft model using Hep3B-Ctrl, Hep3B-TFCP2L1-KO, and Hep3B-TFCP2L1-OE cells. Three weeks post-injection, tumor-bearing mice were orally administered Ti3 and/or Sorafenib twice weekly. As depicted in Fig. 8A–F, the combined treatment of Ti3 (10 mg/kg) and Sorafenib (25 mg/kg) led to an 84% inhibition of tumor growth in the Hep3B-Ctrl group, whereas Sorafenib alone inhibited growth by 61%. No significant difference was observed between the vehicle group and the Ti3 alone group. Notably, Sorafenib alone inhibited tumor growth by 85% in the TFCP2L1-KO group, with minimal additional benefit observed in the combined treatment group (83%) (Fig. 8E, F). Conversely, in Hep3B-TFCP2L1-OE tumor-bearing mice, Sorafenib monotherapy achieved only a 25% suppression rate, while the combination restored the inhibition rate to 83%. These results underscore TFCP2L1’s significant role in modulating tumor cell sensitivity to Sorafenib. Overall, combining Sorafenib with Ti3 demonstrates superior tumor inhibitory efficacy in HCC treatment.

Fig. 8: Effects of Sorafenib and TFCP2L1 inhibitor combination treatment on Hep3B xenografts.

A–C Photographs of Hep3B-Ctrl (A), Hep3B TFCP2L1-OE (B), and Hep3B TFCP2L1-KO (C) xenograft tumors at the endpoint after treatment with the indicated drug combinations. D–F Statistical analysis of the final weight of xenograft tumors at the endpoint. *p < 0.05; **p < 0.01; ns represents no significance compared with the control group. G–I Immunofluorescence staining of Caspase3 and TFCP2L1 expression in xenograft tumors, along with TUNEL staining to assess tumor cell apoptosis. Representative images are shown. DAPI was used to stain the nuclei. Scale bar = 100 μm. J Statistical analysis of TUNEL-positive cells per field from panels G–I. *p < 0.05; **p < 0.01; ns represents no significance compared with the control group. K Graphical abstract of the research findings. TFCP2L1 drives NANOG expression in HCC cells, activating the JAK/STAT3 signaling pathway to promote CSC stemness and HCC pathogenesis. The small molecule Ti3, which targets the active domain of TFCP2L1, can enhance the efficacy of Sorafenib treatment in hepatoma cells.

To elucidate the underlying inhibitory mechanism, tumor tissues were embedded and subjected to immunofluorescence for Caspase-3 expression and TUNEL staining. As shown in Fig. 8G–J, Caspase-3 expression (including precursor Caspase-3 and cleaved-Caspase-3) and the number of TUNEL-positive cells were consistently highest in the Sorafenib and Ti3 combination group, indicating significant apoptosis induction. TFCP2L1 knockout enhanced apoptosis predominantly with Sorafenib alone, whereas TFCP2L1 overexpression attenuated Sorafenib-induced apoptosis.