Although DNA hypermethylation within promoter CpG islands is highly correlated with tumorigenesis, it has not been established whether DNA hypermethylation within a specific tumor suppressor gene (TSG) is sufficient to fully transform a somatic stem cell. In this study, we addressed this question using a novel targeted DNA methylation technique to methylate the promoters of HIC1 and RassF1A, two well-established TSGs, along with a two-component reporter system to visualize successful targeting of human bone marrow-derived mesenchymal stem cells (MSC) as a model cell system. MSCs harboring targeted promoter methylations of HIC1/RassF1A displayed several features of cancer stem/initiating cells including loss of anchorage dependence, increased colony formation capability, drug resistance, and pluripotency. Notably, inoculation of immunodeficient mice with low numbers of targeted MSC resulted in tumor formation, and subsequent serial xenotransplantation and immunohistochemistry confirmed the presence of stem cell markers and MSC lineage in tumor xenografts. Consistent with the expected mechanism of TSG hypermethylation, treatment of the targeted MSC with a DNA methyltransferase inhibitor reversed their tumorigenic phenotype. To our knowledge, this is the first direct demonstration that aberrant TSG hypermethylation is sufficient to transform a somatic stem cell into a fully malignant cell with cancer stem/initiating properties. Cancer Res; 71(13); 4653-63. Ó2011 AACR.

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Hepatocellular carcinoma (HCC) is a leading cause of cancer-related death worldwide. Early diagnosis and treatment of HCC remain a key goal for improving patient survival. Chronic hepatitis B virus (HBV) infection is a major risk factor for HCC development. Pre-S mutants harboring deletions in HBV large surface antigen have been well demonstrated as HBV oncoproteins that dysregulate multiple signaling pathways in hepatocytes, leading to HCC formation. The presence of pre-S mutants in plasma represents important predictive and prognostic markers for HCC in patients with chronic HBV infection. However, the method to detect pre-S mutants remains to be optimized. In this study, we developed a platform, based on the next-generation sequencing (NGS) technology, for detection of pre-S mutants in plasma of HBV-related HCC patients. Compared to the current TA cloning-based analysis, the NGS-based analysis could detect pre-S deletion quantitatively, and the detection rate was significantly more sensitive in 49 plasma analyzed (McNemar’s paired proportion test, P value < 0.0001; simple kappa coefficient, κ = 0.29 (95% CI, 0.12 to 0.46)). Our data suggest that the NGS-based platform may hold a promise for improving the clinical application of pre-S mutants in serving as predictive and prognostic markers for HBV-related HCC.

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Evidence has emerged that stem cells represent a promising therapeutic tool for tissue engineering and regenerative medicine. Thus, identifying functional markers for selecting stem cells capable of superior self-renewal and pluripotency (or multipotency) and maintaining stem cell identity under appropriate culture conditions are critical for guiding the use of stem cells toward clinical applications. Many investigations have implicated the insulin-like growth factor 1 receptor (IGF1R) signaling in maintenance of stem cell characteristics and enhancement of stem cell therapy efficacy. IGF1R-expressing stem cells display robust pluripotent or multipotent properties. In this review, we summarize the essential roles of IGF1R signaling in self-renewal, pluripotency (or multipotency), and therapeutic efficacy of stem cells, including human embryonic stem cells, neural stem cells, cardiac stem cells, bone marrow mesenchymal stem cells, placental mesenchymal stem cells, and dental pulp mesenchymal stem cells. Modifying IGF1R signaling may thus provide potential strategies for maintaining stem cell properties and improving stem-cell-based therapeutic applications.

Backgrounds/AimsThe protective effect of everolimus (EVR) in hepatocellular carcinoma (HCC) patients who receive liver transplantation in terms of reducing the recurrence has not been sufficiently investigated in clinical trials. In this second stage of our ongoing study, we intend to analyze the effects of EVR as an immunosuppressant, when it is started in the early phase after living donor liver transplantation (LDLT), on HCC recurrence in patients with HCC within the University of California at San Francisco (UCSF) criteria.MethodsFrom January 2011 to June 2013, a total of 250 patients underwent LDLT for HCC at our institute. The patients with HCC within the UCSF criteria were included in the study and divided in two groups depending upon the postoperative immunosuppression. Group A: HCC patients that received EVR+TAC based immunosuppressive regimen (n=37). Group B: HCC patients that received standard TAC based immunosuppressive regimen without EVR (n=29). The target trough level for EVR was 3 to 5 ng/ml while for TAC it was 8–10 ng/ml.ResultsFor group A patients, the mean trough level of the EVR was 3.47±1.53 ng/ml (range, 1.5–11.2) with a daily dose of 1.00±0.25 mg/day. For group A and B, the average TAC trough levels were 6.97±3.98 ng/ml (range, 2.50 to 11.28 ng/ml) and 6.93±2.58 (range, 2–16.30), respectively. The 1-year, 3-year and 4-year overall survival achieved for Group A patients was 94.95%, 86.48% and 86.48%, respectively while for Group B patients it was 82.75%, 68.96%, and 62.06%, respectively (p=0.0217).ConclusionsEVR use in liver transplant recipients in the early stage after transplantation reduces the HCC recurrence rates in HCC patients within the UCSF criteria.