SummaryObesity is a major driver of cancer, especially hepatocellular carcinoma (HCC). The prevailing view is that non-alcoholic steatohepatitis (NASH) and fibrosis or cirrhosis are required for HCC in obesity. Here, we report that NASH and fibrosis and HCC in obesity can be dissociated. We show that the oxidative hepatic environment in obesity inactivates the STAT-1 and STAT-3 phosphatase T cell protein tyrosine phosphatase (TCPTP) and increases STAT-1 and STAT-3 signaling. TCPTP deletion in hepatocytes promoted T cell recruitment and ensuing NASH and fibrosis as well as HCC in obese C57BL/6 mice that normally do not develop NASH and fibrosis or HCC. Attenuating the enhanced STAT-1 signaling prevented T cell recruitment and NASH and fibrosis but did not prevent HCC. By contrast, correcting STAT-3 signaling prevented HCC without affecting NASH and fibrosis. TCPTP-deletion in hepatocytes also markedly accelerated HCC in mice treated with a chemical carcinogen that promotes HCC without NASH and fibrosis. Our studies reveal how obesity-associated hepatic oxidative stress can independently contribute to the pathogenesis of NASH, fibrosis, and HCC.

Restoration of tumor suppression is an attractive onco-therapeutic approach. It is particularly relevant when a tumor suppressor is excessively degraded by an overactive oncogenic E3 ligase. We previously discovered that the E6-associated protein (E6AP; as classified in the human papilloma virus context) is an E3 ligase that has an important role in the cellular stress response, and it directly targets the tumor-suppressor promyelocytic leukemia protein (PML) for proteasomal degradation. In this study, we have examined the role of the E6AP-PML axis in prostate cancer (PC). We show that knockdown (KD) of E6AP expression attenuates growth of PC cell lines in vitro. We validated this finding in vivo using cell line xenografts, patient-derived xenografts and mouse genetics. We found that KD of E6AP attenuates cancer cell growth by promoting cellular senescence in vivo, which correlates with restoration of tumor suppression by PML. In addition, we show that KD of E6AP sensitizes cells to radiation-induced death. Overall, our findings demonstrate a role for E6AP in the promotion of PC and support E6AP targeting as a novel approach for PC treatment, either alone or in combination with radiation.

Formalin-fixed (FF) autopsy tissue comprises the bulk of existing Multiple Sclerosis (MSc) pathology archives, providing a rich pool of material for biomarker discovery and disease characterization. Here, we present the development of a heat-induced extraction protocol for the proteomic analysis of FF brain tissue, its application to the study of lesion remyelination and its failure in MSc. A 4-round extraction strategy was optimized using FF tissue leading to a 35% increase in the number of proteins identified compared to a single extraction; and a 65% increase in proteins identified with ≥4 peptides. Histological staining of sections with oil red O and luxol fast blue-periodic acid Schiff, required to characterize MSc lesions was found to have minimal effect on LC-MS/MS. The application of the optimized protocol to chronic demyelinated and remyelinated FF MSc lesions and the adjacent periplaque white matter, isolated through laser guided manual dissection from 3 patients, identified 428 unique proteins (0.2% FDR) using LC-MS/MS. Comparison of the lesion types using iTRAQ and 2-D LC-MS/MS revealed 82 differentially expressed proteins. Protein quantitation by iTRAQ and spectral counting was well-correlated (r(s)= 0.7653; p < 10(-30)). The data generated from this work illustrates the scope of the methodology and provides insights into the pathogenesis of MSc and remyelination.

The tumor suppressor p16, one protein encoded by the INK4/ARF locus, is frequently absent in multiple cancers, including non-small cell lung cancer (NSCLC). Whereas increased methylation of the encoding gene (CDKN2A) accounts for its loss in a third of patients, no molecular explanation exists for the remainder. We unraveled an alternative mechanism for the silencing of the INK4/ARF locus involving the E3 ubiquitin ligase and transcriptional cofactor E6AP (also known as UBE3A). We found that the expression of three tumor suppressor genes encoded in the INK4/ARF locus (p15, p16, and p19) was decreased in E6AP mouse embryo fibroblasts. E6AP induced the expression of the INK4/ARF locus at the transcriptional level by inhibiting CDC6 transcription, a gene encoding a key repressor of the locus. Luciferase assays revealed that E6AP inhibited CDC6 expression by reducing its E2F1-dependent transcription. Chromatin immunoprecipitation analysis indicated that E6AP reduced the amount of E2F1 at the CDC6 promoter. In a subset of NSCLC samples, an E6AP-low/CDC6-high/p16-low protein abundance profile correlated with low methylation of the gene encoding p16 (CDKN2A) and poor patient prognosis. These findings define a previously unrecognized tumor-suppressive role for E6AP in NSCLC, reveal an alternative silencing mechanism of the INK4/ARF locus, and reveal E6AP as a potential prognostic marker in NSCLC.