Intratumoral PD-1high CD8+ T cells correlate with AFP levels in HCC patients: a brief report

Background: To date, the only FDA approved targeted agent in advanced hepatocarcinomas is sorafenib, indicating a critical need for innovative therapeutic options. LY2157299, a selective ATP-mimetic inhibitor of TGF-β receptor (TβR)-I activation, is currently under clinical investigation in HCC patients after sorafenib or in patients ineligible for sorafenib. Our study aimed at investigating the effects of LY2157299 in HCC cell lines and patient samples with various AFP expression levels. Materials and Methods: Antiproliferative effects of LY2157299 were evaluated in a panel of human HCC cells by MTT assay. Baseline and phosphorylated (p-) protein levels were assessed by Western blot analysis and mRNA expressions by qRT-PCR. Invasion assays were done on matrigel and in OptiCell systems. Tumor samples from HCC patients were surgically resected and cut in 300 µm thick slices using a Tissue Slicer. Each slice was randomly exposed to LY2157299 (1µM and 10µM) and sorafenib (5µM) for 48h. At the end of treatment, tumor samples were analyzed by immunohistochemistry (IHC) or immunofluorescence (IF). Results: LY2157299 was evaluated in HEPG2 and HUH-7 AFP-positive and HEP3B and SK-HEP1 AFP negative cells as well as “in lab-established” SK-HEP1-derived cell lines resistant to the tyrosine kinase inhibitors sorafenib (SK-Sora) and sunitinib (SK-Suni). Exogenous stimulation of all HCC cell lines with TGF-β yielded downstream activation of canonical Smad pathway that was potently inhibited with LY2157299 treatment at micromolar concentrations. In contrast, the non-canonical pathways such as MAPK and Akt/mTOR were not activated by TGF-β in most cell lines with the exception of SK-HEP1 cells where LY2157299 restored the basal level of p-ERK1/2, p-AKT and p-S6 kinase phosphorylation. Low concentrations of LY2157299 displayed antiproliferative effects in HEPG2 and HEP3B cells when stimulated by TGF-β but not in other cell lines. LY2157299 yielded potent anti-migratory and anti-invasive properties in invasive SK-HEP1, SK-Suni and SK-Sora cells. Tumor slices from surgically resected advanced HCC tumor from 5 different patients, were exposed ex vivo to LY2157299 or sorafenib for 48h. This method allows the evaluation of novel anticancer agents in whole tumors containing cancer cells and stromal cells. LY2157299 but not sorafenib decreased 2-5 fold the p-Smad2/3. LY2157299 treatment mainly affected PI3K/AKT rather than MAPK signalling pathway. In contrast, sorafenib treatment reduced both PI3K and MAPK pathway activation. IHC analysis of LY2157299 and sorafenib-exposed samples showed a significant decrease (>3 fold) of the proliferative marker Ki67 and increase of the apoptotic marker caspase-3 (3-5 fold). Interestingly, all molecular effects were independent of AFP expression. Conclusion: TGF-β/TβR-I inactivation using LY2157299 inhibits TGF-β-dependent cell signaling in HCC cell lines with either anti-proliferative or anti-invasive effects depending on the model. In tumor samples from patients, inhibition of TGF-β signaling was associated with inhibition of proliferation and apoptosis induction regardless AFP level. Our data suggest that LY2157299 may be useful for patients with HCC. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A228. Citation Format: Marie Serova, Annemilaï Tijeras-Raballand, Celia Dos Santos, Karim A. Benhadji, Valerie Paradis, Eric Raymond, Sandrine Faivre, Armand de Gramont. TGF-beta signaling inhibition using LY2157299 affects proliferation or invasion in hepatocarcinoma cells and patient samples. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A228.

Background and ObjectivesHepatocellular carcinoma (HCC) is the fourth leading cause of cancer associated death globally. Serum micro RNAs are full of potential as noninvasive biomarkers. Here, we aim to assess the performance of serum MicroRNA-155 and MicroRNA-665 as diagnostic biomarker for HCC comparing to AFP.MethodsSerum samples were collected from 200 subjects (40 healthy control, 80 chronic hepatitis C patients with cirrhosis and without HCC (LC) and 80 HCC patients currently infected by hepatitis C infection and didn’t start the treatment). The HCC patients didn’t include alcoholic liver disease, nonalcoholic fatty liver disease nor autoimmune liver disease. MicroRNA-155 and MicroRNA-665 expression were measured by real-time quantitative PCR (RT-qPCR), while AFP level was assessed by ELISA method.ResultsBoth miR-155 and miR-665 were significantly elevated in HCC group as compared to both control and LC groups. The comparison between LC and HCC patients revealed that the serum level of miR-155 was a significant increase in HCC patients compared to LC patients; however, the serum level of miR-665 didn’t show any significant difference between the same two groups. MiR-665 expression level showed a direct correlation with tumor size in HCC patients.ConclusionsUsing measurement against AFP level in serum, miR-665 is considered a promising serum biomarker for the diagnosis of HCC patients among the LC patients without HCC. MiR-155 didn’t provide a better performance than serum AFP as a diagnostic biomarker among the same group. MiR-665 may serve as a good indicator for HCC prognosis.

DCP is a useful HCC tumor marker, which reflects a defect in vitamin K metabolism. We tested the hypothesis that vitamin K treatment of HCC patients might suppress this marker and possibly AFP also. HCC patients who had both elevated AFP and DCP were included. A phase I cohort was treated with escalating vitamin K1 intravenous weekly doses and a 27-patient phase II cohort was then treated with a fixed oral daily dose. A maximum tolerated dose was not reached up to 100-fold the normal vitamin K1 dose. No toxicities were found up to 1,000 mg/infusion. In the phase II cohort, 93% of patients had tumor marker responses by decreased DCP levels, but only 22% had responses by decreased AFP levels. CT scans showed 11% of patients had PRs, 59% had stable tumors and 29.6% had tumor progression. Mechanism studies showed that vitamin K1 induced phosphorylation of JNK and c-Jun and caspase-mediated apoptosis. Vitamin K1 was non-toxic at high doses, strongly inhibited plasma DCP levels, but weakly suppressed AFP levels. The results provide evidence that the two tumor markers are not directly linked and that DCP levels may not reflect HCC cell growth, as DCP levels were decreased in patients without AFP change, and were suppressed in vitro at 1% of the vitamin K1 concentration needed to inhibit AFP.

<p>PDF file - 589K, Supplementary Fig. S1 Elevated MDK levels in the culture medium of HCC cell lines compared with normal liver cell lines. Supplementary Fig. S2 The prognostic values of AFP and MDK levels for HCC patients. Supplementary Fig. S3.Expression of serum MDK in another independent cohort (cohort B) from Egypt. Supplementary Fig. S4. Logistic Regression Model to combine information of MDK and AFP for prediction of HCC in the learning cohort A (n=252): Comparison of AUROC for diagnosis HCC through logist regression model I, serum MDK and AFP. Supplementary Fig. S5. Positive predictive value (PPV) and negative predictive value (NPV) for identifying HCC patients through serum MDK according to different prevalence. Supplementary Fig. S6. Correlation between serum AFP and MDK levels in HCC patients (n=252). Supplementary Fig. S7. Serum MDK is elevated in different HCC subsets, No significant difference in serum MDK levels was found between the learning cohort A and the external validation cohort of HCC patients(P=0.398), or between the early stage HCC patients in learning and validation cohort (P=0.295). Table S1. Relationship between MDK expression and clinicopathological features of HCC patients. Table S2. Clinical and biochemical characteristics of liver cirrhosis (LC) patients in cohort A. Table S3. Baseline characteristics of the hospital controls with different etiologies from learning Cohort A. Table S4. Clinical characteristics of HCC patients according to different cohorts in this study. Table S5. MDK protein levels in cancer-free cirrhotic liver and HCC tissues. Table S6. Comparison of AFP and MDK with clinicopathological parameters in learning cohort A. Table S7．Univariate Cox-regression Analyses of Factors Associated With OS and TTR. Table S8. Sensitivities and specificities of AFP and MDK according to various cutoff values. Table S9. Cross-validation of the diagnostic ability of serum MDK. Table S10. Comparison of MDK with clinicopathological parameters in validation cohort C.</p>

<p>PDF file - 589K, Supplementary Fig. S1 Elevated MDK levels in the culture medium of HCC cell lines compared with normal liver cell lines. Supplementary Fig. S2 The prognostic values of AFP and MDK levels for HCC patients. Supplementary Fig. S3.Expression of serum MDK in another independent cohort (cohort B) from Egypt. Supplementary Fig. S4. Logistic Regression Model to combine information of MDK and AFP for prediction of HCC in the learning cohort A (n=252): Comparison of AUROC for diagnosis HCC through logist regression model I, serum MDK and AFP. Supplementary Fig. S5. Positive predictive value (PPV) and negative predictive value (NPV) for identifying HCC patients through serum MDK according to different prevalence. Supplementary Fig. S6. Correlation between serum AFP and MDK levels in HCC patients (n=252). Supplementary Fig. S7. Serum MDK is elevated in different HCC subsets, No significant difference in serum MDK levels was found between the learning cohort A and the external validation cohort of HCC patients(P=0.398), or between the early stage HCC patients in learning and validation cohort (P=0.295). Table S1. Relationship between MDK expression and clinicopathological features of HCC patients. Table S2. Clinical and biochemical characteristics of liver cirrhosis (LC) patients in cohort A. Table S3. Baseline characteristics of the hospital controls with different etiologies from learning Cohort A. Table S4. Clinical characteristics of HCC patients according to different cohorts in this study. Table S5. MDK protein levels in cancer-free cirrhotic liver and HCC tissues. Table S6. Comparison of AFP and MDK with clinicopathological parameters in learning cohort A. Table S7．Univariate Cox-regression Analyses of Factors Associated With OS and TTR. Table S8. Sensitivities and specificities of AFP and MDK according to various cutoff values. Table S9. Cross-validation of the diagnostic ability of serum MDK. Table S10. Comparison of MDK with clinicopathological parameters in validation cohort C.</p>

The prognosis of HCC remains poor mainly because of the lack of diagnosis biomarkers especially in patients with cirrhosis background. To identify serum biomarkers for HCC, we use cleavable stable isotope labeling (cICAT) combined with LC-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) to compare the serum proteome between liver cirrhosis and HCC patients. Sera from nine liver cirrhosis patients and nine HCC patients were selected for screening study. Then the alpha-1-acid glycoprotein (AGP) was chosen for validation by western blot. AGP was measured in the separate validation study including 52 HCC patients and 40 liver cirrhosis patients by rate nephelometry. Its value for HCC diagnosis from liver cirrhosis was also appreciated through receiver operating curve (ROC). We quantified and identified 31 different proteins which include AGP, complement C4, haptoglobin, alpha-1-antitrypsin precursor, alpha-2-macroglobulin precursor, prothrombin precursor and ubiquitin carboxyl-terminal hydrolase 44 (USP44), etc. The serum concentration of AGP in HCC patients (n = 52) was significantly higher than in liver cirrhosis (n = 40) (P < 0.005) in validation study. AGP was useful for discrimination of the HCC cases from LC patients when the AFP levels were below 500 ng/ml. The area under curve (AUC) of the AGP and the combination of AGP and AFP were 0.834 (P < 0.0005, 95% CI: 0.734-0.934) and 0.880 (P < 0.0005, 95% CI, 0.789-0.970) separately, which are higher than AFP alone (0.538, P = 0.604, 95% CI: 0.386-0.689). cICAT combined with LC-MS/MS-based serum proteome analysis can be useful in the screening of serum biomarkers for HCC. Alpha-1-acid glycoprotein combining AFP could aid the diagnosis of HCC.

CD24 is reported to be up-regulated in the tissues of HCC patients when compared with normal liver tissues. We aim to determine whether CD24 protein is also overexpressed in the plasma of HCC patients, and its diagnostic value for HCC. Plasma levels of CD24 protein and AFP were measured by enzyme linked immunosorbent assay (ELISA) in the plasma of 90 patients with hepatocellular carcinoma and 30 healthy controls. The sensitivity and specificity were calculated and the relationship between the expression of CD24 and clinical pathological parameters was analyzed. Both plasma CD24 protein and AFP levels in HCC patients were higher than those in healthy controls (p<0.05). There was no correlation between plasma levels of AFP and CD24 in 90 patients with HCC (r=-0.084, p=0.430). The best cut-off value of CD24 was 3.31ng/mL, which yielded a sensitivity and specificity of 83.3% and 93.3%, respectively, for screening HCC; and plasma CD24 level was not associated with gender, age, hepatitis infection status, tumor size and histological differentiation and TNM stage (p>0.05). Plasma CD24 protein might serve as a novel tumor marker in differentiating HCC patients from normal individuals as well as monitor HCC status in AFP negative HCC patients.

e15157 Background: Cirrhosis is often associated with thrombocytopenia and predisposes to HCC. Platelets are known to produce several HCC growth factors. Methods: A cohort of 4,139 Chinese HCC patients from Taiwan was interrogated for baseline clinical, tumor and liver function characteristics in relation to blood platelet counts. Results: Trichotomization of the cohort by tumor size after log-linearization, revealed significant relationships of tumor size to blood platelet, AFP and GGTP levels. Patients with higher platelets had larger tumors, more tumor nodules and higher AFP and GGTP levels than those with lower platelet levels. 80% of patients with larger tumors had normal platelet counts, compared to 50% of patients with smaller tumors. Analysis of the cohort according to platelet sub-sets, revealed significant relationships of platelet counts to tumor size and AFP levels. By Spearman’s correlation coefficient, platelet counts correlated moderately with tumor size (rho=0.411), but weakly with tumor numbers, PVT, AST or AFP levels. Sub-set analysis of the cohort according to platelet levels showed that 38% of patients had thrombocytopenia and they had the smallest tumors, lowest tumor numbers, lowest percent with PVT and lowest AFP levels. Conclusions: There was a significant correlation between HCC size and blood platelet counts; patients with larger tumors had higher platelet levels.

Correction to: Intratumoral PD-1high CD8+T cells correlate with AFP levels in HCC patients: a brief report.

IntroductionHepatocellular Carcinoma (HCC) is a primary tumor of the liver; it is one of the most common cancers worldwide. Osteopontin (OPN) is a phosphorylated glycoprotein which is implicated in enhancing invasive and metastatic progression of many tumors. Midkine (MDK) is a 13-kDa small heparin-binding growth factor which plays a significant role in carcinogenesis related activities. The aim of this study was to assess the efficacy of serum Midkine and Osteopontin levels as diagnostic biomarkers of Hepatocellular Carcinoma.MethodsThis study was carried out from January 2014 to January 2016 in Internal Medicine, Clinical Oncology and Tropical Medicine Departments of Tanta University Hospital (Egypt). One hundred forty subjects were enrolled in our study, they were divided into four groups: Group I included 35 patients presented with HCV without cirrhosis; Group II included 35 patients presented with HCV with liver cirrhosis; Group III included 35 patients presented with HCC on top of cirrhosis; and Group IV included 35 apparently healthy subjects as a control group. The studied groups were age and sex matched. Routine and specific (OPN and MDK) laboratory investigations were performed in all included subjects.ResultsThe main finding of the present work was that the mean serum levels of OPN and MDK were significantly elevated in HCC patients either by comparing HCC patients vs. HCV patients without cirrhosis, HCC patients vs. HCV patients with cirrhosis or HCC patients vs. healthy subjects. Interestingly, by performing a ROC analysis, serum MDK levels had better sensitivity and specificity than OPN and AFP levels in the diagnosis of HCC (98.4 %, 97.1% and 97%) and (96.2%, 95.3% and 95%) for MDK, OPN and AFP respectively.ConclusionSerum MDK and OPN levels are comparable to AFP levels and could be used as potential diagnostic biomarkers of HCC in HCV patients with liver cirrhosis and in the prediction of liver cirrhosis in HCV patients without cirrhosis.

BackgroundHepatocellular carcinoma (HCC) is the sixth prevailing cancer globally and is the second greatest reason for cancer-linked deaths in males, surpassed only by lung cancer. A significant number of HCC cases experiencing advanced stages are due to the frequent absence or inconspicuousness of early symptoms. Consequently, discovering reliable early diagnostic indicators and innovative treatment targets is essential to improve survival and overall outcomes for HCC patients. Herein, we aimed to evaluate the diagnostic potential of APEX1 in Egyptian HCC patients and explore its prognostic significance before treatment initiation.Study design and methodologyThis research was conducted as a prospective comparative cohort study involving 60 Egyptian participants, age range more than 18 years selected from internal medicine and hepatology outpatient clinics and inpatient wards at Ain Shams University hospitals from January 2024 to July 2024, after informed consent was taken from the patients, who were allocated into Group A (Cirrhotic) comprising 12 liver cirrhosis patients but without HCC, acting as the control group; Group B (HCC) included 48 patients diagnosed with HCC. Group B was further subdivided into four subgroups: Subgroup 1: 12 patients receiving palliative (supportive) care; Subgroup 2: 12 patients treated with Sorafenib, Subgroup 3: 12 patients treated with trans-arterial chemoembolization (TACE); and Subgroup 4: 12 patients treated with radiofrequency ablation (RFA). Data collection included anthropometric measurements, laboratory tests, radiological findings, Child–Pugh scores, BCLC scores, and patient performance metrics, gathered both before and after treatment. The data were analyzed utilizing IBM SPSS (ver. 27).ResultsThe study identified a serum APEX1 level cut-off of > 13.72 ng/mL as a biomarker to differentiate between cirrhotic patients and those with HCC (sensitivity = 92.31%, specificity = 100.0%). The threshold for AFP was 20 ng/mL (sensitivity = 76.92%, specificity = 100.0%). Notably, significant differences in AFP and APEX1 levels were found between cirrhotic patients and HCC patients in the RF and TACE subgroups at baseline (P-value 0.004, < 0.001 for AFP and APEX1, respectively), indicating that APEX1 may serve as a more sensitive biomarker for early HCC detection.Both AFP and APEX1 exhibited a positive predictive value (PPV) of 100%.Furthermore, the study revealed that an AFP level exceeding 280 ng/mL at baseline could predict a poor prognosis (sensitivity = 57.1%, specificity = 81.8%, AUC = 0.679). In contrast, an APEX1 level greater than 9.96 ng/mL at baseline was found to be a noteworthy predictor of poor prognosis (specificity = 86.4%, sensitivity = 92.9%, AUC = 0.857).ConclusionsAPEX1 has proven to be a more sensitive and accurate serum biomarker than AFP for diagnosing HCC.-Serum APEX1 levels revealed a significant rise in HCC patients, even during the early stages (candidates for radiological/therapeutic intervention), making it a valuable tool for screening and early detection in surveillance programs.-APEX1 demonstrated the ability to predict patient prognosis even prior to the initiation of treatment.

BackgroundHepatocellular carcinoma (HCC) is a major health problem worldwide. However, the popular tumor marker, AFP, lacks sensitivity although its specificity is high. Tissue biopsy is an invasive operation and may increase the risk of needle-track metastases. Heat shock protein 90 (HSP90) is a potential biomarker for tumor diagnosis and prognosis. This study aims to determine whether levels of plasma HSP90α in HCC patients can be used as a cost-effective and simple test for the initial diagnosis of the disease.MethodsPlasma samples were collected from 659 HCC patients, 114 secondary hepatic carcinoma (SHC) patients, 28 hepatic hemangioma patients and 230 healthy donors. The levels of HSP90α were measured by ELISA.ResultsThe levels of plasma HSP90α in HCC patients were significantly higher than in healthy donors and in patients with hepatic hemangioma or SHC (144.08 ± 4.98, 46.81 ± 1.11, 61.56 ± 8.20 and 111.96 ± 10.08 ng/mL, respectively; p < 0.05 in all cases). The levels were associated with age (p = 0.001), BCLC stage (p < 0.001), levels of AFP (p < 0.001), tumor size (p < 0.001), tumor number (p < 0.001), PVTT (p < 0.001), EHM (p < 0.001) and Child-Pugh stage in the HCC cohort. In addition, the levels of plasma HSP90α showed an upward trend along with the progression of the BCLC stage. ROC curve analysis showed that compared to AFP (AUC 0.922, 95%CI 0.902–0.938) or HSP90α (AUC 0.836, 95%CI 0.810–0.860), the combination of HSP90α and AFP (AUC0.943, 95%CI 0.925–0.957) significantly improved the diagnostic efficiency for HCC patients.ConclusionThe results suggest that plasma Hsp90 α levels can be used as an initial diagnosis for patients with HCC in both rural and cosmopolitan settings.

Background Hepatocellular carcinoma (HCC) is the fifth most common cancer worldwide and the most common form of primary liver cancer. Aim of the study to study the level of serum vitronectin (VTN) compared to AFP in diagnosis and prognosis of hepatocellular carcinoma on top of HCV-related liver diseases. Patients and Methods This prospective observational study included a total number of 60 subjects who were divided into 4 groups: Group 1: include 10 normal persons. Group 2: included 10 patients who have Hepatitis C Virus infection. Group 3: included 20 cirrhotic patients. Group 4: included 20 patients with HCC (on top of hepatitis C virus related cirrhosis), in which we tested them for vitronectin before first intervention and after intervention within 3 months. Results The mean age of the cases showed high statistically significant difference between the study groups (P &amp;lt; 0.001).There were a total number of 29 males and 31 females with significant difference in the sex distribution between the study groups. 80% of the cases in HCC group were males. This sex distribution can be attributed to high prevalence of risk factors like smoking, DM and HCV in males in addition to possible role of sex hormones. On analyzing laboratory characters of the studied groups, we found statistically significant difference between cirrhosis group and non-cirrhotic groups with low platelets count, high serum creatinine, high INR, lower serum albumin, high bilirubin and higher AST. The median level of AFP in group 4 (HCC patients) was 110 IU/ml which was higher than its median value in the other study groups with high level of significance between the study groups. The median level of vitronectin levels in the different study groups didn’t reveal a statistically significant difference between the different study groups. Serum level of vitronectin in group 3 (cirrhotic patients) has no statistically significant difference between the three subgroups according to child's classification. By the analysis of serum level of AFP and vitronectin level in group IV (HCC patients) before and after treatment, the median level of AFP was significantly lower than its level before intervention, while the decrease in vitronectin level was statistically insignificant after intervention. Conclusions Chronic HCV infection and its serious complicatons specially HCC are still representing a health challenge in Egypt, reflecting wide HCV prevalence and late diagnosis. Serum AFP alone has an unreliable role in HCC surveillance as it has low sensitivity because it may be normal in up to 40% of HCC cases especially early stages of the tumor, and low specificity as its levels may be elevated in conditions other than HCC as cirrhosis or exacerbation of chronic hepatitis or even in some cases of cholangiocarcinoma. AFP is useful in clinical practice in screening and diagnosis of HCC in association with US and CT. According to our study, we can't use Vitronectin alone in diagnosis and prognosis of HCC on top of cirrhosis related to HCV +ve infection. It didn’t reveal a statistically significant difference between the different study groups ,so it may require further study and research.

The significance of the current study was to determine normative levels of PIVKA-II and AFP in patients with unresectable HCC and healthy participants. The second goal was to assess the roles of PIVKA-II and AFP in predicting radiological response after loco-regional therapy. This prospective cohort study enrolled consecutive samples of HCC patients and healthy controls. Venous blood samples were obtained at baseline and after interventions to determine serum levels of PIVKA-II and AFP using the chemiluminescent microparticle immunoassay method. Radiologic responses were determined based on the WHO criteria. Fifty-four HCC patients (mean age 58.9 years, 49 males) and 40 healthy controls (mean age 33.5 years, 26 males) were recruited. The median serum levels of PIVKA-II and AFP in HCC vs. healthy controls were 988.4 vs. 24.2 mAU/ml and 13.6 vs. 1.7 ng/ml, respectively (both p<0.001). With ROC curve analysis, the area under the curve (AUC) for PIVKA-II was 0.95 95% CI [0.90-0.99], and for AFP it was 0.98, 95% CI [0.95-1.0]). The cut-off value for PIVKA-II was 41.4 mAU/ml, and AFP was 4.8 ng/ml. PIVKA-II levels correlated significantly with radiological responses (r=0.64, p=0.02) but not AFP (r=0.09, p=0.2). PIVKA-II and AFP levels are distinctive between unresectable HCC and healthy controls. However, PIVKA-II, not AFP, can predict the radiological response after loco-regional therapy.

BackgroundHBV infection is the leading risk factor for HCC. HBV infection has been confirmed to be associated with the exhaustion status of CD8+ T cells and immunotherapeutic efficacy in HCC. In this study, we aimed to investigate the prognostic value of the CD8+ T-cell exhaustion signature and immunotherapy response in patients with HBV-related HCC.MethodsWe identified different clusters of HBV-related HCC cells by single-cell RNA sequencing (scRNA-seq) and identified CD8+ T-cell exhaustion-related genes (TERGs) by pseudotime analysis. We conducted differential expression analysis and LASSO Cox regression to detect genes and construct a CD8+ T-cell exhaustion index (TEI). We next combined the TEI with other clinicopathological factors to design a prognostic nomogram for HCC patients. We also analysed the difference in the TEI between the non-responder and responder groups during anti-PD-L1 therapy. In addition, we investigated how HBV induces CD8+ T lymphocyte exhaustion through the inhibition of tyrosine metabolism in HCC using gene set enrichment analysis and RT‒qPCR.ResultsA CD8+ T-cell exhaustion index (TEI) was established with 5 TERGs (EEF1E1, GAGE1, CHORDC1, IKBIP and MAGOH). An AFP level > 500 ng, vascular invasion, histologic grade (G3-G4), advanced TNM stage and poor five-year prognosis were related to a higher TEI score, while HBV infection was related to a lower TEI score. Among those receiving anti-PD-L1 therapy, responders had lower TEIs than non-responders did. The TEI also serves as an independent prognostic factor for HCC, and the nomogram incorporating the TEI, TNM stage, and vascular invasion exhibited excellent predictive value for the prognosis in HCC patients. RT‒qPCR revealed that among the tyrosine metabolism-associated genes, TAT (tyrosine aminotransferase) and HGD (homogentisate 1,2 dioxygenase) were expressed at lower levels in HBV-HCC than in non-HBV HCC.ConclusionGenerally, we established a novel TEI model by comprehensively analysing the progression of CD8+ T-cell exhaustion, which shows promise for predicting the clinical prognosis and potential immunotherapeutic efficacy in HBV-related HCC patients.