Hua Xiao - US grants

tissue /cell culture

DESCRIPTION (provided by applicant): Deregulation of c-Myc expression is implicated in pathogenesis of many human cancers including breast cancer. Estrogen receptor alpha can increase the rate of c-Myc transcription through the recruitment of a variety of cofactors to the c-Myc promoter, yet the precise roles of these cofactors in transcription and tumorigenesis are largely unknown. We have identified a putative tumor suppressor, TIP30, also called CC3, that can specifically regulate ER alpha-mediated c-Myc transcription. Our objective is to investigate the role of TIP30 in the regulation of transcription of ER alpha-target genes and tumorigenesis. The specific aims of this proposal are to: 1) investigate whether deletion of TIP30 increases expression of ER alpha-target genes in the epithelial cells of the mammary glands in vivo. 2) investigate whether breast cancers from patients harbor mutations in the TIP30 gene. 3) evaluate the susceptibility of TIP30-mutant mice to mammary tumorigenesis. These studies will have a significant impact on three important aspects. They will provide: 1) a molecular basis for a TIP30-mediated regulatory pathway in mammary gland development, 2) direct evidence for the role of a transcription cofactor in breast carcinogenesis, and 3) a system for studying biologically relevant factors and events responsible for the pathogenesis of breast cancers. Therefore, this proposal will not only yield insights into the mechanisms for regulation of gene expression intumorigenesis but also provide a potential target for the diagnosis and treatment of human cancers.

DESCRIPTION (provided by applicant): The incidence of hepatocellular carcinoma (HCC) is two to four times higher in men than in women and the progression of chronic liver diseases to cirrhosis is much faster in men than in women. These suggest an important role of sex hormones in hepatocarcinogenesis. However, the molecular mechanism underlying the gender disparity in HCC still remains poorly understood. And there have been controversies on the use of hormonal therapies for patients with HCC. Hence, a better understanding of the genetic and epigenetic alterations that contribute to HCC development, especially to the gender disparity of HCC, may lead to a discovery of new etiologic, prognostic or therapeutic targets. The long- term goal is to understand how the genetic alterations in cells lead to hepatocarcinogenesis. The objective of this proposal is to determine the mechanism by which down regulation of nuclear receptor coactivator 5 (Ncoa5) contributes to the development of HCC. The central hypothesis is that Ncoa5 deficiency preferentially promotes hepatocarcinogenesis in males through inhibition of estrogen receptor 1 (ER?) activity and upregulation of androgen receptor (AR) expression in the liver. This hypothesis has been formulated on the basis of our recent preliminary data. The rationale for the proposed research is that understanding of the mechanism underlying Ncoa5-mediated suppression of HCC will reveal a novel regulatory pathway and new targets for developing innovative strategies to the prevention and treatment of HCC. This hypothesis will be tested by the following two specific aims: 1). Determine the effects of estrogen on Ncoa5 deficiency induced hepatocarcinogenesis using a novel genetically-engineered mouse model;and 2) Determine the molecular mechanism by which Ncoa5 exerts its effect on Kupffer cells and hepatocytes. The approach is innovative, because it utilizes a novel genetically-engineered mouse model of spontaneous HCC to prove a critical role of Ncoa5 in the suppression of HCC development. The proposed research is significant, because it is expected to demonstrate Ncoa5 as a novel mouse tumor suppressor with haploinsufficiency that links up ER? and AR mediated regulatory pathways to HCC. This will not only advance our understanding of the etiological mechanisms of HCC, but also provide the fields of basic and preclinical studies with a new mouse model that mimics the initiation and progression of human HCC. PUBLIC HEALTH RELEVANCE: The proposed research is relevant to public health because the development of an animal model of hepatocellular carcinoma (HCC) and discovery of the mechanism underlying the gender disparity of HCC will not only contribute to a more comprehensive picture of the roles of sex hormone in Hepatocarcinogenesis, but also lead to the discovery of new etiologic, prognostic or therapeutic targets. Thus, the proposed research is relevant to the part of NIH's mission that pertains to developing the prevention and/or treatment of liver cancer.

DESCRIPTION (provided by applicant): Nonalcoholic fatty liver disease (NAFLD) and type-2 diabetes (T2D) are associated with increased risk of hepatocellular carcinoma (HCC), and individuals with T2D treated with metformin have a reduced risk of HCC incidence and mortality. Although several factors have been reported to probably mediate the preventive effect of metformin on the risk of several types of cancers, a precise mechanism by which metformin reduces HCC risk remains largely unknown. Specifically, it is not known what cellular factors, targeted by metformin, are critical in the development of HCC in diabetic patients, and which sub- population of patients may be more responsive to metformin treatment. Our long-term goal is to understand the molecular basis of HCC. The objective of this proposal is to unveil the mechanism by which metformin inhibits spontaneous HCC development in mice bearing a heterozygous deletion of the nuclear receptor coactivator-5 (Ncoa5) gene and improve current tools utilizing identified genes and signatures to predict prognosis and response to metformin in patients with HCC. The central hypothesis is that reduced Ncoa5 expression is a key risk factor for HCC development in the context of a diabetic condition, and metformin protects mice against HCC development by reducing hepatic insulin resistance and steatosis, in part by positively regulating the expression of Ncoa5. The rationale for the proposed research is that therapeutic strategies, aimed at increasing Ncoa5 expression or targeting Ncoa5 deficiency driven oncogenic pathways, are potentially effective in protecting against HCC. This hypothesis will be tested by two specific aims: 1) determine the effect of metformin on HCC development and Ncoa5 expression in the liver using a novel genetically-engineered mouse model (Ncoa5+/- mice) of HCC, and 2) identify and translate critical targets and pathways, in metformin-mediated protection against HCC, from a mouse model of HCC to human HCC. The approach is innovative because it utilizes a novel genetically-engineered Ncoa5+/- mouse model of HCC to analyze the effects of metformin on hepatocarcinogenesis and identifies new genes and gene signatures for predicting the prognosis of patients with HCC. The proposed research is significant because it is expected to establish a new concept that metformin inhibits an Ncoa5 deficiency-induced pathogenic pathway that is commonly shared by NAFLD, T2D and HCC. This will not only advance our understanding of the etiological mechanisms of HCC, but also provide better biomarkers for predicting the risk of recurrence and response to metformin related therapy in patients with HCC.

DESCRIPTION (provided by applicant): Hepatocellular carcinoma (HCC) is the fifth most common and the third most lethal cancer worldwide, with increasing incidence in developed countries including the United States. It is estimated that 15-50% of HCC patients develop HCC in the absence of eminent etiological factors such as hepatitis viral infection and alcohol abuse. Emerging evidence has indicated that metabolic disorders, such as nonalcoholic fatty liver disease (NAFLD) and type-2 diabetes (T2D), are linked to HCC development, which may account for the increasing incidence of HCC in developed countries. However, mechanisms underlying the connection between these disorders and HCC remain largely unknown. Our laboratory has recently demonstrated that reduced nuclear receptor coactivator-5 (Ncoa5) expression is associated with 40% of human HCC specimens, and that haploinsufficiency of NCOA5 in heterozygous knockout male mice results in glucose intolerance, NAFLD and subsequent HCC. These suggest that a NCOA5 deficiency-driven pathogenic pathway is commonly shared by NAFLD, T2D and HCC. Therefore, we hypothesize that NCOA5 acts as a haploinsufficient tumor suppressor by controlling temporal expression of genes encoding key proinflammatory cytokines and lipogenic enzymes in the liver. Thus, dysfunction of NCOA5 induces gene expression programs promoting hepatic inflammation and lipogenesis, leading to HCC development. Our objective is to provide evidence that NCOA5 is a key regulator controlling hepatic inflammation and lipid metabolism and impairment of its function results in HCC development. This hypothesis will be tested by three specific aims: 1). Illustrate the molecular links between hepatic inflammation and HCC using cell specific Ncoa5 knockout mice; 2) Determine the molecular connections between NCOA5 deficiency-driven NAFLD and HCC; and 3) Determine the role of aberrant NCOA5 expression in human HCCs and establish a molecular relationship between NCOA5 deficiency-driven mouse and human HCCs. The approach is innovative, because it combines the use of genetics to control metabolic phenotypes and system-level bioinformatic methods with novel mouse models of HCC to understand the underlying oncogenic pathway and hidden key factors in hepatocarcinogenesis. The proposed research is significant because these studies will not only provide unique insight into the reciprocal relationship between metabolic diseases and HCC, but also new targets for the development of preventive and therapeutic approaches against HCC.

DNA ligases are critical enzymes for virtually all DNA transactions, including DNA replication, repair and recombination. Vertebrates have three DNA ligase genes: Lig1, Lig3 and Lig4; none can be directly deleted from the mouse germline. The Lig3 gene encodes two isoforms: mitochondrial Lig3 (which is cell essential) and the more abundant nuclear Lig3. Recently, we established a mutant mouse strain harboring a knock-in mutation that specifically ablates nuclear Lig3 using a one-step CRISPR/Cas9- mediated genome editing strategy in mouse embryos. Nuclear Lig3 is widely considered to be the primary ligase for DNA single strand break (SSB) repair due to its strong interaction with an essential SSBR factor called X-ray cross complementation factor 1 (XRCC1). Our unique mouse model will allow for the first time, a thorough dissection of the function of Lig3 in nuclear DNA repair in vivo. Experiments proposed in this application will provide the initial characterization of these mice. A comprehensive pathological phenotype analysis will be performed. Because DNA repair defects are often associated with genomic instability and neurodegenerative diseases, additional experiments will focus on the impact of Lig3 deficiency on spontaneous tumor development and neuropathology. Successful completion of the proposed studies will provide unprecedented insight into Lig3's normal physiologic role. Conversely, Lig3 overexpression is found in a number of tumor cell lines and tumor samples from human patients; this impacts DNA double strand break (DSB) repair such that the Lig4-dependent canonical non-homologous end-joining (NHEJ) is, to varying extents, replaced by the Lig3-dependent alternative end-joining (A-EJ), which has been implicated in chromosomal translocations. We will address the impact of nuclear Lig3 deficiency on IgH/c-myc translocation that occurs during immunoglobulin (Ig) heavy (H) chain class switch recombination (CSR). These particular translocations are critical to the development of many human and mouse B cell tumors. Furthermore, characterizing the dependence of these translocations on Lig3 may be relevant to many other tumor types. Successful completion of these studies may elucidate a pathological role of Lig3 in tumorigenesis.