Search

Various cancer stem cell (CSC) biomarkers have been identified for hepatocellular carcinoma (HCC), but little is known about the implications of heterogeneity and shared molecular networks within the CSC population. Through miRNA profile analysis in an HCC cohort (n = 241) for five groups of CSC+ HCC tissues, i.e., EpCAM+, CD90+, CD133+, CD44+, and CD24+ HCC, we identified a 14-miRNA signature commonly altered among these five groups of CSC+ HCC. miR-192-5p, the top-ranked CSC miRNA, was liver-abundant and -specific and markedly downregulated in all five groups of CSC+ HCC from two independent cohorts (n = 613). Suppressing miR-192-5p in HCC cells significantly increased multiple CSC populations and CSC-related features through targeting PABPC4. Both TP53 mutation and hypermethylation of the mir-192 promoter impeded transcriptional activation of miR-192-5p in HCC cell lines and primary CSC+ HCC. This study reveals the circuit from hypermethylation of the mir-192 promoter through the increase in PABPC4 as a shared genetic regulatory pathway in various groups of primary CSC+ HCC. This circuit may be the driver that steers liver cells toward hepatic CSC cells, leading to hepatic carcinogenesis.Significance:miR-192-5p and its regulatory pathway is significantly abolished in multiple groups of HCC expressing high levels of CSC markers, which may represent a key event for hepatic carcinogenesis.

PDF file - 2.5MB

PDF file - 146K

Supplementary Figures 1-2 from Inhibition of Growth and Metastasis of Human Hepatocellular Carcinoma by Antisense Oligonucleotide Targeting Signal Transducer and Activator of Transcription 3

Purpose: Hepatocellular carcinoma (HCC), a common cancer worldwide, has a dismal outcome partly due to the poor identification of early-stage HCC. Currently, one third of HCC patients present with low serum α-fetoprotein (AFP) levels, the only clinically available diagnostic marker for HCC. The aim of this study was to identify new diagnostic molecular markers for HCC, especially for individuals with low serum AFP.Experimental Design: We used the microarray technique to determine the expression profiles of 218 HCC specimens from patients with either high or low serum AFP. From the microarray study, we selected five candidate genes (i.e., GPC3, PEG10, MDK, SERPINI1, and QP-C), which were overexpressed in HCCs. Using quantitative real-time PCR analyses, we validated the expression of these five genes in 50 AFP-normal and 8 AFP-positive HCC specimens and 36 cirrhotic noncancerous hepatic specimens, which include 52 independent specimens not used in microarray analysis.Results: A significant increase in the expression of the five candidate genes could be detected in most of the HCC samples, including those with normal serum AFP and small tumors. GPC3, MDK, and SERPINI1 encode known serum proteins. Consistently, a significant increase in serum midkine, encoded by MDK, was associated with HCC patients, including those with normal serum AFP. Using prediction analysis of microarray, we showed that a combined score of these five genes can accurately classify noncancerous hepatic tissues (100%) and HCC (71%).Conclusions: We suggest that a diagnostic signature approach using a combined score of these five biomarkers rather than a single marker may improve the prediction accuracy of HCC patients, including those with normal serum AFP and smaller-sized tumors.

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.

Purpose: To evaluate the value of serum midkine (MDK) as a diagnostic biomarker in hepatocellular carcinoma, particularly for those with negative alpha-fetoprotein (AFP) and at an early stage.Experimental Design: MDK expression in tumors was assessed by immunohistochemistry from 105 patients with hepatocellular carcinomas or liver cirrhosis. Serum MDK levels were detected by ELISA in 933 participants including hepatocellular carcinomas and hospital controls from different medical centers. Sensitivities and specificities of serum MDK in diagnosing hepatocellular carcinoma according to AFP level and Barcelona Clinic Liver Cancer (BCLC) stage were analyzed.Results: MDK levels were significantly elevated in hepatocellular carcinoma tissues as well as serum samples. The sensitivity of serum MDK for hepatocellular carcinoma diagnosis was much higher than that of AFP (86.9% vs. 51.9%) with similar specificities (83.9% vs. 86.3%). Notably, serum MDK had an outstanding performance in distinguishing AFP-negative hepatocellular carcinomas from different controls: In those AFP-negative hepatocellular carcinomas, the sensitivity could reach as high as 89.2%. Moreover, receiver operating characteristic (ROC) curve analysis also showed that serum MDK had a better performance compared with AFP in distinguishing early-stage hepatocellular carcinomas as well as small hepatocellular carcinomas. Even in very early-stage hepatocellular carcinomas, MDK showed an obviously higher sensitivity compared with AFP (80% vs. 40%). Furthermore, serum MDK level was significantly decreased in patients with hepatocellular carcinomas after curative resection and re-elevated when tumor relapse occurred.Conclusions: Serum MDK is significantly elevated in most hepatocellular carcinomas, including those with negative AFP and at an early stage, which may serve as a novel diagnostic marker in early diagnosis and postoperative monitoring of hepatocellular carcinomas. Clin Cancer Res; 19(14); 3944–54. ©2013 AACR.

Purpose: To investigate the role of macrophages in tumor progression under sorafenib treatment and to explore whether combination of drugs that deplete macrophages improved the antitumor effect of sorafenib.Experimental Design: Tumor growth, lung metastasis, and tumor angiogenesis were observed in HCCLM3-R and SMMC7721, two human hepatocellular carcinoma xenograft nude mouse models, when treated with sorafenib (30 mg/kg daily, n = 6 per group) or a vehicle as control. Macrophage infiltration was measured in the peripheral blood and in sorafenib-treated tumor by immunohistochemistry and flow cytometry with F4/80 antibody and CD11b antibody. The effect of macrophage depletion on tumor angiogenesis and metastasis after sorafenib treatment, using two drug target macrophages, zoledronic acid (ZA) and clodrolip, was measured in the two models of hepatocellular carcinoma.Results: Although sorafenib significantly inhibited tumor growth and lung metastasis, it induced a significant increase in peripheral recruitment and intratumoral infiltration of F4/80- and CD11b-positive cells, which was accompanied with elevation of colony-stimulating factor-1, stromal-derived factor 1α, and vascular endothelial growth factor in the tumor and elevation of plasma colony-stimulating factor-1 and mouse vascular endothelial growth factor in peripheral blood, suggesting the role of macrophages in tumor progression under sorafenib treatment. Depletion of macrophages by clodrolip or ZA in combination with sorafenib significantly inhibited tumor progression, tumor angiogenesis, and lung metastasis compared with mice treated with sorafenib alone. ZA was more effective than clodrolip.Conclusions: Macrophages may have an important role in tumor progression under sorafenib treatment. ZA is promising when combined with sorafenib to enhance its antitumor effect. Clin Cancer Res; 16(13); 3420–30. ©2010 AACR.

PDF file - 4.1MB

It includes additional materials and methods, i.e., Plasmids, siRNAs, and LY294002 treatment; assays to examine cell malignancy features; qPCR and Western blot.

Supplementary Data from Human Hepatocellular Carcinoma Tumor–derived Endothelial Cells Manifest Increased Angiogenesis Capability and Drug Resistance Compared with Normal Endothelial Cells

Supplementary Text, Figures 1-3, Tables 1-6 from EpCAM and α-Fetoprotein Expression Defines Novel Prognostic Subtypes of Hepatocellular Carcinoma

Supplementary Figures 1-3, Table 1 from A Unique Metastasis Gene Signature Enables Prediction of Tumor Relapse in Early-Stage Hepatocellular Carcinoma Patients

The heterogeneous nature of hepatocellular carcinoma (HCC) and the lack of appropriate biomarkers have hampered patient prognosis and treatment stratification. Recently, we have identified that a hepatic stem cell marker, epithelial cell adhesion molecule (EpCAM), may serve as an early biomarker of HCC because its expression is highly elevated in premalignant hepatic tissues and in a subset of HCC. In this study, we aimed to identify novel HCC subtypes that resemble certain stages of liver lineages by searching for EpCAM-coexpressed genes. A unique signature of EpCAM-positive HCCs was identified by cDNA microarray analysis of 40 HCC cases and validated by oligonucleotide microarray analysis of 238 independent HCC cases, which was further confirmed by immunohistochemical analysis of an additional 101 HCC cases. EpCAM-positive HCC displayed a distinct molecular signature with features of hepatic progenitor cells including the presence of known stem/progenitor markers such as cytokeratin 19, c-Kit, EpCAM, and activated Wnt-β-catenin signaling, whereas EpCAM-negative HCC displayed genes with features of mature hepatocytes. Moreover, EpCAM-positive and EpCAM-negative HCC could be further subclassified into four groups with prognostic implication by determining the level of α-fetoprotein (AFP). These four subtypes displayed distinct gene expression patterns with features resembling certain stages of hepatic lineages. Taken together, we proposed an easy classification system defined by EpCAM and AFP to reveal HCC subtypes similar to hepatic cell maturation lineages, which may enable prognostic stratification and assessment of HCC patients with adjuvant therapy and provide new insights into the potential cellular origin of HCC and its activated molecular pathways. [Cancer Res 2008;68(5):1451–61]

A new oxygenated lanostane-type triterpenoid, 20S,24S-epoxy-lanosta-7,9(11)-dien-3β,15α,25R,26-tetraol (1), together with three known compounds (2–4) were isolated from the fruiting bodies of Ganoderma weberianum. Extensive NMR spectrometry and HRESIMS analysis, as well as NMR and ECD calculations elucidated the structure of the new compound. 27-nor-3β-hydroxylanosta-7,9(11),23E-trien-25-one (2) showed superior α-glucosidase inhibitory activity with IC50 value of 122.1 μM to that of positive control acarbose (304.6 μM).

Additional file 10: Table S2. Comparisons among different CTC enrichment techniques

Background Combined treatment with tyrosine kinase inhibitors (TKI) plus anti-PD-1 antibodies showed high anti-tumor efficacy and made conversion resection possible for patients with unresectable hepatocellular carcinoma (HCC). However, long-term survival has not been reported. Methods A cohort of consecutive patients who received combined TKI/anti-PD-1 antibodies as first-line treatment for initially unresectable HCC at the authors’ hospital between August 2018 and September 2020 was eligible for this study. Patients who were responding to systemic therapy and met the criteria for hepatectomy underwent liver resection with curative intention. The study also investigated the association of clinical factors with successful conversion resection and postoperative recurrence. Results The study enrolled 101 patients including 24 patients (23.8 %) who underwent R0 resection a median of 3.9 months (interquartile range: 2.5–5.9 months) after initiation of systemic therapy. Patients with an Eastern cooperative oncology group performance status of 0, fewer intrahepatic tumors, or a radiographic response to systemic therapy were more likely to be able to receive curative resection. After a median follow-up period of 21.5 months, hepatectomy was independently associated with a favorable overall survival (hazard ratio [HR], 0.050; 95 % confidence interval [CI], 0.007–0.365; P = 0.003). For the 24 patients who underwent surgery, the 12-month recurrence-free survival and overall survival rates were respectively 75% and 95.8%. Achieving a pathologic complete response (n = 10) to systemic therapy was associated with a favorable recurrence-free survival after resection, with a trend toward significance (HR, 0.345; 95% CI, 0.067–1.785; P = 0.187). Conclusions Selected patients with initially unresectable HCC can undergo hepatectomy after systemic therapy with combined TKI/anti-PD-1 antibodies. In this study, conversion resection was associated with a favorable prognosis.