Epigenetic alterations have contributed greatly to human carcinogenesis. Conventional epigenetic studies have predominantly focused on DNA methylation, histone modifications, and chromatin ...remodeling. Recently, diverse and reversible chemical modifications of RNAs have emerged as a new layer of epigenetic regulation. N6‐methyladenosine (m6A) is the most abundant chemical modification of eukaryotic messenger RNA (mRNA) and is important for the regulation of mRNA stability, splicing, and translation. Using transcriptome sequencing, we discovered that methyltransferase‐like 3 (METTL3), a major RNA N6‐adenosine methyltransferase, was significantly up‐regulated in human hepatocellular carcinoma (HCC) and multiple solid tumors. Clinically, overexpression of METTL3 is associated with poor prognosis of patients with HCC. Functionally, we proved that knockdown of METTL3 drastically reduced HCC cell proliferation, migration, and colony formation in vitro. Knockout of METTL3 remarkably suppressed HCC tumorigenicity and lung metastasis in vivo. On the other hand, using the CRISPR/dCas9‐VP64 activation system, we demonstrated that overexpression of METTL3 significantly promoted HCC growth both in vitro and in vivo. Through transcriptome sequencing, m6A sequencing, and m6A methylated RNA immuno‐precipitation quantitative reverse‐transcription polymerase chain reaction, we identified suppressor of cytokine signaling 2 (SOCS2) as a target of METTL3‐mediated m6A modification. Knockdown of METTL3 substantially abolished SOCS2 mRNA m6A modification and augmented SOCS2 mRNA expression. We also showed that m6A‐mediated SOCS2 mRNA degradation relied on the m6A reader protein YTHDF2‐dependent pathway. Conclusion: METTL3 is frequently up‐regulated in human HCC and contributes to HCC progression. METTL3 represses SOCS2 expression in HCC through an m6A‐YTHDF2‐dependent mechanism. Our findings suggest an important mechanism of epigenetic alteration in liver carcinogenesis. (Hepatology 2018;67:2254‐2270).
Interaction between tumor cells and immune cells in the tumor microenvironment is important in cancer development. Immune cells interact with the tumor cells to shape this process. Here, we use ...single-cell RNA sequencing analysis to delineate the immune landscape and tumor heterogeneity in a cohort of patients with HBV-associated human hepatocellular carcinoma (HCC). We found that tumor-associated macrophages suppress tumor T cell infiltration and TIGIT-NECTIN2 interaction regulates the immunosuppressive environment. The cell state transition of immune cells towards a more immunosuppressive and exhaustive status exemplifies the overall cancer-promoting immunocellular landscape. Furthermore, the heterogeneity of global molecular profiles reveals co-existence of intra-tumoral and inter-tumoral heterogeneity, but is more apparent in the latter. This analysis of the immunosuppressive landscape and intercellular interactions provides mechanistic information for the design of efficacious immune-oncology treatments in hepatocellular carcinoma.
Abstract
Exosomes belong to a subpopulation of extracellular vesicles secreted by the dynamic multistep endocytosis process and carry diverse functional molecular cargoes, including proteins, lipids, ...nucleic acids (DNA, messenger and noncoding RNA), and metabolites to promote intercellular communication. Proteins and noncoding RNA are among the most abundant contents in exosomes; they have biological functions and are selectively packaged into exosomes. Exosomes derived from tumor, stromal and immune cells contribute to the multiple stages of cancer progression as well as resistance to therapy. In this review, we will discuss the biogenesis of exosomes and their roles in cancer development. Since specific contents within exosomes originate from their cells of origin, this property allows exosomes to function as valuable biomarkers. We will also discuss the potential use of exosomes as diagnostic and prognostic biomarkers or predictors for different therapeutic strategies for multiple cancers. Furthermore, the applications of exosomes as direct therapeutic targets or engineered vehicles for drugs are an important field of exosome study. Better understanding of exosome biology may pave the way to promising exosome-based clinical applications.
A population of stromal cells, myeloid‐derived suppressor cells (MDSCs), is present in tumors. Though studies have gradually revealed the protumorigenic functions of MDSCs, the molecular mechanisms ...guiding MDSC recruitment remain largely elusive. Hypoxia, O2 deprivation, is an important factor in the tumor microenvironment of solid cancers, whose growth often exceeds the growth of functional blood vessels. Here, using hepatocellular carcinoma as the cancer model, we show that hypoxia is an important driver of MDSC recruitment. We observed that MDSCs preferentially infiltrate into hypoxic regions in human hepatocellular carcinoma tissues and that hypoxia‐induced MDSC infiltration is dependent on hypoxia‐inducible factors. We further found that hypoxia‐inducible factors activate the transcription of chemokine (C‐C motif) ligand 26 in cancer cells to recruit chemokine (C‐X3‐C motif) receptor 1‐expressing MDSCs to the primary tumor. Knockdown of chemokine (C‐C motif) ligand 26 in cancer cells profoundly reduces MDSC recruitment, angiogenesis, and tumor growth. Therapeutically, blockade of chemokine (C‐C motif) ligand 26 production in cancer cells by the hypoxia‐inducible factor inhibitor digoxin or blockade of chemokine (C‐X3‐C motif) receptor 1 in MDSCs by chemokine (C‐X3‐C motif) receptor 1 neutralizing antibody could substantially suppress MDSC recruitment and tumor growth. Conclusion: This study unprecedentedly reveals a novel molecular mechanism by which cancer cells direct MDSC homing to primary tumor and suggests that targeting MDSC recruitment represents an attractive therapeutic approach against solid cancers. (Hepatology 2016;64:797‐813)
Sorafenib is the standard treatment for advanced hepatocellular carcinoma (HCC). However, the development of drug resistance is common. By using genome-wide CRISPR/Cas9 library screening, we identify ...phosphoglycerate dehydrogenase (PHGDH), the first committed enzyme in the serine synthesis pathway (SSP), as a critical driver for Sorafenib resistance. Sorafenib treatment activates SSP by inducing PHGDH expression. With RNAi knockdown and CRISPR/Cas9 knockout models, we show that inactivation of PHGDH paralyzes the SSP and reduce the production of αKG, serine, and NADPH. Concomitantly, inactivation of PHGDH elevates ROS level and induces HCC apoptosis upon Sorafenib treatment. More strikingly, treatment of PHGDH inhibitor NCT-503 works synergistically with Sorafenib to abolish HCC growth in vivo. Similar findings are also obtained in other FDA-approved tyrosine kinase inhibitors (TKIs), including Regorafenib or Lenvatinib. In summary, our results demonstrate that targeting PHGDH is an effective approach to overcome TKI drug resistance in HCC.
Cancer stemness, referring to the stem-cell-like phenotype of cancer cells, has been recognised to play important roles in different aspects of hepatocarcinogenesis. A number of well-established ...cell-surface markers already exist for liver cancer stem cells, with potential new markers of liver cancer stem cells being identified. Both genetic and epigenetic factors that affect various signalling pathways are known to contribute to cancer stemness. In addition, the tumour microenvironment-both physical and cellular-is known to play an important role in regulating cancer stemness, and the potential interaction between cancer stem cells and their microenvironment has provided insight into the regulation of the tumour-initiating ability as well as the cellular plasticity of liver CSCs. Potential specific therapeutic targeting of liver cancer stemness is also discussed. With increased knowledge, effective druggable targets might be identified, with the aim of improving treatment outcome by reducing chemoresistance.
Hepatocellular carcinoma (HCC) cells exploit an aberrant transcriptional program to sustain their infinite growth and progression. Emerging evidence indicates that the continuous and robust ...transcription of oncogenes in cancer cells is often driven by super‐enhancers (SEs). In this study, we systematically compared the SE landscapes between normal liver and HCC cells and revealed that the cis‐acting SE landscape was extensively reprogrammed during liver carcinogenesis. HCC cells acquired SEs at multiple prominent oncogenes to drive their vigorous expression. We identified sphingosine kinase 1 (SPHK1) as an SE‐associated oncogene, and we used this gene as an example to illustrate the impact of SEs on the activation of oncogenes in HCC. Concurrently, we also showed that the critical components of the trans‐acting SE complex, namely, cyclin‐dependent kinase 7 (CDK7), bromodomain‐containing protein 4 (BRD4), E1A binding protein P300 (EP300), and mediator complex subunit 1 (MED1), were frequently overexpressed in human HCCs and were associated with the poor prognosis of patients with HCC. Using the CRISPR/Cas9 gene‐editing system and specific small‐molecule inhibitors, we further demonstrated that HCC cells were highly sensitive to perturbations of the SE complex. The inactivation of CDK7, BRD4, EP300, and MED1 selectively repressed the expression of SE‐associated oncogenes in HCC. Finally, we demonstrated that THZ1, which is a small‐molecule inhibitor of CDK7, exerted a prominent anticancer effect in both in vitro and in vivo HCC models. Conclusion: The SE landscape and machinery were significantly altered in human HCCs. HCC cells are highly susceptible to perturbations of the SE complex due to the resulting selective suppression of SE‐associated oncogenes. Our results suggest that targeting SE complex is a promising therapeutic strategy for HCC treatment.
Background & Aims: Recent efforts in stem cell biology suggest that tumors are organized in a hierarchy of heterogeneous cell populations and that the capability to maintain tumor formation/growth ...specifically resides in a small population of cells called cancer stem cells (CSCs). The aim of this study is to identify, isolate, and characterize the CSC population that drives and maintains hepatocellular carcinoma (HCC) growth and metastasis. Methods: Normal stem cells involved in liver regeneration were identified using a severe partial hepatectomy model. Purified HCC cells, with or without expression of the identified normal stem cell phenotype, were evaluated, based on their tumorigenic potential and exhibition of defined stem/progenitor cell-like properties, to determine whether liver CSCs can be or partly be identified by this surface marker. Results: We report the identification and isolation of a population of CSCs expressing a CD133 surface phenotype from human liver cell lines. CD133+ cells possess a greater colony-forming efficiency, higher proliferative output, and greater ability to form tumor in vivo. These cells are endowed with characteristics similar to those of progenitor cells including the expression of “stemness” genes, the ability to self-renew, and the ability to differentiate into nonhepatocyte-like lineages. Furthermore, CD133 is found to represent only a minority of the tumor cell population in human HCC specimens. Conclusions: We report the identification of a CSC population in HCC characterized by their CD133 phenotype. The identification of tumorigenic liver CSCs could provide new insight into the HCC tumorigenic process and possibly bear great therapeutic implications.
Like normal stem cells, tumor-initiating cells (T-ICs) are regulated extrinsically within the tumor microenvironment. Because HCC develops primarily in the context of cirrhosis, in which there is an ...enrichment of activated fibroblasts, we hypothesized that cancer-associated fibroblasts (CAFs) would regulate liver T-ICs. We found that the presence of α-SMA(+) CAFs correlates with poor clinical outcome. CAF-derived HGF regulates liver T-ICs via activation of FRA1 in an Erk1,2-dependent manner. Further functional analysis identifies HEY1 as a direct downstream effector of FRA1. Using the STAM NASH-HCC mouse model, we find that HGF-induced FRA1 activation is associated with the fibrosis-dependent development of HCC. Thus, targeting the CAF-derived, HGF-mediated c-Met/FRA1/HEY1 cascade may be a therapeutic strategy for the treatment of HCC.
Display omitted
•The presence of α-SMA(+) CAFs correlates with poor prognosis of HCC•CAFs regulate liver T-ICs through paracrine secretion of HGF•FRA1 is required for HGF/c-Met-mediated T-IC phenotypes•The HGF-mediated c-Met/FRA1/HEY1 cascade represents a therapeutic target for HCC
Similar to normal stem cells, tumor-initiating cells are regulated extrinsically within their microenvironment. Lau et al. find that cancer-associated fibroblasts play a crucial role in the regulation of liver T-ICs through HGF-mediated activation of the c-Met/FRA1/HEY1 cascade. Targeting this pathway may be a promising approach for cancer treatment.
Cancer metastasis is a multistep process that involves a series of tumor‐stromal interaction, including extracellular matrix (ECM) remodeling, which requires a concerted action of multiple ...proteolytic enzymes and their endogenous inhibitors. This study investigated the role of tissue inhibitor of metalloproteinases (TIMP) 2 in the context of hepatocellular carcinoma (HCC) metastasis. We found that TIMP2 was the most significantly down‐regulated member among the TIMP family in human HCCs. Moreover, TIMP2 underexpression was frequent (41.8%; 23 of 55) in human HCCs and was significantly associated with liver invasion and poorer survival outcomes of HCC patients. Furthermore, stable silencing of TIMP2 in HCC cell lines enhanced cell invasive ability and ECM degradation associated with formation of invadopodia‐like feature, suggesting that TIMP2 is a negative regulator of HCC metastasis. Using an orthotopic tumor xenograft model, we demonstrated that ectopic expression of TIMP2 open reading frame in the highly metastatic HCC cell line, MHCC‐97L, significantly reduced HCC progression as well as pulmonary metastasis. Mechanistically, TIMP2 suppression, in a hypoxic environment, was induced through a regulatory feedback circuit consisting of hypoxia‐inducible factor (HIF) 1 alpha, microRNA‐210 (miR‐210), and HIF‐3α. Conclusion: TIMP2 is frequently down‐regulated in human HCCs and its down‐regulation is associated with aggressive tumor behavior and poorer patient outcome. Its suppression is under the regulation of a novel feedback circuit consisting of HIF‐1α/miR‐210/HIF‐3α. TIMP2 is an important regulator of ECM degradation and HCC metastasis. (Hepatology 2016;64:473‐487)
PDF
