Liem H. Nguyen

Lin28 enhances tissue repair by reprogramming cellular metabolism

Regeneration capacity declines with age, but why juvenile organisms show enhanced tissue repair remains unexplained. Lin28a, a highly conserved RNA-binding protein expressed during embryogenesis, plays roles in development, pluripotency, and metabolism. To determine whether Lin28a might influence tissue repair in adults, we engineered the reactivation of Lin28a expression in several models of tissue injury. Lin28a reactivation improved hair regrowth by promoting anagen in hair follicles and accelerated regrowth of cartilage, bone, and mesenchyme after ear and digit injuries. Lin28a inhibits let-7 microRNA biogenesis; however, let-7 repression was necessary but insufficient to enhance repair. Lin28a bound to and enhanced the translation of mRNAs for several metabolic enzymes, thereby increasing glycolysis and oxidative phosphorylation (OxPhos). Lin28a-mediated enhancement of tissue repair was negated by OxPhos inhibition, whereas a pharmacologically induced increase in OxPhos enhanced repair. Thus, Lin28a enhances tissue repair in some adult tissues by reprogramming cellular bioenergetics. PAPERCLIP:

Lin28b Is Sufficient to Drive Liver Cancer and Necessary for Its Maintenance in Murine Models

Lin28a/b are RNA-binding proteins that influence stem cell maintenance, metabolism, and oncogenesis. Poorly differentiated, aggressive cancers often overexpress Lin28, but its role in tumor initiation or maintenance has not been definitively addressed. We report that LIN28B overexpression is sufficient to initiate hepatoblastoma and hepatocellular carcinoma in murine models. We also detected Lin28b overexpression in MYC-driven hepatoblastomas, and liver-specific deletion of Lin28a/b reduced tumor burden, extended latency, and prolonged survival. Both intravenous siRNA against Lin28b and conditional Lin28b deletion reduced tumor burden and prolonged survival. Igf2bp proteins are upregulated, and Igf2bp3 is required in the context of LIN28B overexpression to promote growth. Therefore, multiple murine models demonstrate that Lin28b is both sufficient to initiate liver cancer and necessary for its maintenance.

Modular degradable dendrimers enable small RNAs to extend survival in an aggressive liver cancer model

Significance Liver cancer is a leading cause of death and a global health problem. Unfortunately, five small-molecule drugs for hepatocellular carcinoma (HCC) recently failed phase III clinical trials partly because late-stage liver dysfunction amplifies drug toxicity. MicroRNAs (miRNAs) present a promising alternative treatment strategy but require development of delivery vehicles that can avoid this cancer-induced dysfunction, which exacerbates toxicity. We overcame this challenge by developing dendrimer nanoparticles that mediate miRNA delivery to late-stage liver tumors with low hepatotoxicity. An aggressive, MYC-driven transgenic liver cancer model was used to examine let-7g tumor suppressor efficacy, resulting in a significant survival benefit. These dendrimer carriers provide high potency in tumors without negatively affecting normal tissues, solving a critical issue in treating aggressive liver cancer. RNA-based cancer therapies are hindered by the lack of delivery vehicles that avoid cancer-induced organ dysfunction, which exacerbates carrier toxicity. We address this issue by reporting modular degradable dendrimers that achieve the required combination of high potency to tumors and low hepatotoxicity to provide a pronounced survival benefit in an aggressive genetic cancer model. More than 1,500 dendrimers were synthesized using sequential, orthogonal reactions where ester degradability was systematically integrated with chemically diversified cores, peripheries, and generations. A lead dendrimer, 5A2-SC8, provided a broad therapeutic window: identified as potent [EC50 < 0.02 mg/kg siRNA against FVII (siFVII)] in dose–response experiments, and well tolerated in separate toxicity studies in chronically ill mice bearing MYC-driven tumors (>75 mg/kg dendrimer repeated dosing). Delivery of let-7g microRNA (miRNA) mimic inhibited tumor growth and dramatically extended survival. Efficacy stemmed from a combination of a small RNA with the dendrimer’s own negligible toxicity, therefore illuminating an underappreciated complication in treating cancer with RNA-based drugs.

Suppression of the SWI/SNF Component Arid1a Promotes Mammalian Regeneration

Mammals have partially lost the extensive regenerative capabilities of some vertebrates, possibly as a result of chromatin-remodeling mechanisms that enforce terminal differentiation. Here, we show that deleting the SWI/SNF component Arid1a substantially improves mammalian regeneration. Arid1a expression is suppressed in regenerating tissues, and genetic deletion of Arid1a increases tissue repair following an array of injuries. Arid1a deficiency in the liver increases proliferation, reduces tissue damage and fibrosis, and improves organ function following surgical resection and chemical injuries. Hepatocyte-specific deletion is also sufficient to increase proliferation and regeneration without excessive overgrowth, and global Arid1a disruption potentiates soft tissue healing in the ear. We show that Arid1a loss reprograms chromatin to restrict promoter access by transcription factors such as C/ebpα, which enforces differentiation, and E2F4, which suppresses cell-cycle re-entry. Thus, epigenetic reprogramming mediated by deletion of a single gene improves mammalian regeneration and suggests strategies to promote tissue repair after injury.

Precise let-7 expression levels balance organ regeneration against tumor suppression

The in vivo roles for even the most intensely studied microRNAs remain poorly defined. Here, analysis of mouse models revealed that let-7, a large and ancient microRNA family, performs tumor suppressive roles at the expense of regeneration. Too little or too much let-7 resulted in compromised protection against cancer or tissue damage, respectively. Modest let-7 overexpression abrogated MYC-driven liver cancer by antagonizing multiple let-7 sensitive oncogenes. However, the same level of overexpression blocked liver regeneration, while let-7 deletion enhanced it, demonstrating that distinct let-7 levels can mediate desirable phenotypes. let-7 dependent regeneration phenotypes resulted from influences on the insulin-PI3K-mTOR pathway. We found that chronic high-dose let-7 overexpression caused liver damage and degeneration, paradoxically leading to tumorigenesis. These dose-dependent roles for let-7 in tissue repair and tumorigenesis rationalize the tight regulation of this microRNA in development, and have important implications for let-7 based therapeutics. DOI: http://dx.doi.org/10.7554/eLife.09431.001

Lin28 and let-7 in cell metabolism and cancer

Malignant cells exhibit major metabolic alterations. The regulatory gene networks that regulate metabolism and the impact of these alterations on overall cellular fitness deserve further exploration. The let-7 microRNAs and their antagonists, the Lin28 RNA-binding proteins, are well-known for controlling the timing of embryonic development. This pathway has recently been shown to regulate glucose metabolism in adult mice and to reprogram metabolism during tissue injury and repair. In addition, many lines of evidence have established that Lin28 is an oncogene that drives tumorigenesis in part by suppressing let-7. The metabolic underpinnings of this oncogenic program are just beginning to be uncovered. Here, we will review the current understanding of how Lin28 exerts regenerative and oncogenic effects through metabolic mechanisms.

Arid1a Has Context-Dependent Oncogenic and Tumor Suppressor Functions in Liver Cancer

ARID1A, an SWI/SNF chromatin-remodeling gene, is commonly mutated in cancer and hypothesized to be tumor suppressive. In some hepatocellular carcinoma patients, ARID1A was highly expressed in primary tumors but not in metastatic lesions, suggesting that ARID1A can be lost after initiation. Mice with liver-specific homozygous or heterozygous Arid1a loss were resistant to tumor initiation while ARID1A overexpression accelerated initiation. In contrast, homozygous or heterozygous Arid1a loss in established tumors accelerated progression and metastasis. Mechanistically, gain of Arid1a function promoted initiation by increasing CYP450-mediated oxidative stress, while loss of Arid1a within tumors decreased chromatin accessibility and reduced transcription of genes associated with migration, invasion, and metastasis. In summary, ARID1A has context-dependent tumor-suppressive and oncogenic roles in cancer.

Knockdown of Anillin Actin Binding Protein Blocks Cytokinesis in Hepatocytes and Reduces Liver Tumor Development in Mice Without Affecting Regeneration

BACKGROUND & AIMS Cytokinesis can fail during normal postnatal liver development, leading to polyploid hepatocytes. We investigated whether inhibiting cytokinesis in the liver slows tumor growth without compromising the health of normal hepatocytes. We inhibited cytokinesis in cancer cells by knocking down ANLN, a cytoskeletal scaffolding protein that regulates cytokinesis and might promote tumorigenesis, in mice with liver disease. METHODS We analyzed clinical and gene expression data from The Cancer Genome Atlas, Oncomine, PrognoScan, and a hepatocellular carcinoma (HCC) tissue microarray. We knocked down ANLN with small interfering RNAs (siRNAs) in H2.35 liver cells and performed image analyses of cells undergoing cytokinesis. siRNAs were delivered to LAP-MYC mice, which develop hepatoblastoma, using lipid nanoparticles. H2.35 cells with knockdown of ANLN or control cells were injected into FRG mice, which develop chronic liver damage, and tumor growth was monitored. We also developed mice with inducible expression of transgenes encoding small hairpin RNAs (shRNAs) against Anln messenger RNA and studied liver tumorigenesis after administration of diethylnitrosamine and carbon tetrachloride. siRNAs against Anln messenger RNA were conjugated to N-acetylgalactosamine to reduce toxicity and increase hepatocyte tropism; their effects were studied in mouse models of liver cancer and regeneration. RESULTS Levels of ANLN messenger RNA were increased in human HCC tissues compared to non-tumor liver tissues. siRNA knockdown of ANLN blocked cytokinesis in H2.35 liver cells. Administration of siRNA against ANLN increased survival times of LAP-MYC mice, compared to mice given a control siRNA. H2.35 liver cells with shRNA knockdown of ANLN formed tumors more slowly in FRG mice than control H2.35 cells. Mice with inducible expression of shRNAs against Anln mRNA developed fewer liver tumors after administration of diethylnitrosamine and carbon tetrachloride than control mice. Knockdown of ANLN did not affect liver regeneration after acute and chronic liver injuries. CONCLUSIONS Knockdown of ANLN in liver cells blocks cytokinesis and inhibits development of liver tumors in mice. Agents that inhibit ANLN in the liver might be effective for prevention or treatment of HCC.

Erratum: Arid1a Has Context-Dependent Oncogenic and Tumor Suppressor Functions in Liver Cancer (Cancer Cell (2017) 32(5) (574–589.e6)(S1535610817304609)(10.1016/j.ccell.2017.10.007))

A. Gordon Robertson, Juliann Shih, Christina Yau, Ewan A. Gibb, Junna Oba, Karen L. Mungall, Julian M. Hess, Vladislav Uzunangelov, Vonn Walter, Ludmila Danilova, Tara M. Lichtenberg, Melanie Kucherlapati, Patrick K. Kimes, Ming Tang, Alexander Penson, Ozgun Babur, Rehan Akbani, Christopher A. Bristow, Katherine A. Hoadley, Lisa Iype, Matthew T. Chang, TCGA Research Network, Andrew D. Cherniack, Christopher Benz, Gordon B. Mills, Roel G.W. Verhaak, Klaus G. Griewank, Ina Felau, Jean C. Zenklusen, Jeffrey E. Gershenwald, Lynn Schoenfield, Alexander J. Lazar, Mohamed H. Abdel-Rahman, Sergio Roman-Roman, Marc-Henri Stern, Colleen M. Cebulla, Michelle D. Williams, Martine J. Jager, Sarah E. Coupland, Bita Esmaeli,* Cyriac Kandoth,* and Scott E. Woodman* *Correspondence: besmaeli@mdanderson.org (B.E.), kandothc@mskcc.org (C.K.), swoodman@mdanderson.org (S.E.W.) https://doi.org/10.1016/j.ccell.2017.12.013

SWI/SNF component ARID1A restrains pancreatic neoplasia formation

Objective ARID1A is commonly mutated in pancreatic ductal adenocarcinoma (PDAC), but the functional effects of ARID1A mutations in the pancreas are unclear. Understanding the molecular mechanisms that drive PDAC formation may lead to novel therapies. Design Concurrent conditional Arid1a deletion and Kras activation mutations were modelled in mice. Small-interfering RNA (siRNA) and CRISPR/Cas9 were used to abrogate ARID1A in human pancreatic ductal epithelial cells. Results We found that pancreas-specific Arid1a loss in mice was sufficient to induce inflammation, pancreatic intraepithelial neoplasia (PanIN) and mucinous cysts. Concurrent Kras activation accelerated the development of cysts that resembled intraductal papillary mucinous neoplasm. Lineage-specific Arid1a deletion confirmed compartment-specific tumour-suppressive effects. Duct-specific Arid1a loss promoted dilated ducts with occasional cyst and PDAC formation. Heterozygous acinar-specific Arid1a loss resulted in accelerated PanIN and PDAC formation with worse survival. RNA-seq showed that Arid1a loss induced gene networks associated with Myc activity and protein translation. ARID1A knockdown in human pancreatic ductal epithelial cells induced increased MYC expression and protein synthesis that was abrogated with MYC knockdown. ChIP-seq against H3K27ac demonstrated an increase in activated enhancers/promoters. Conclusions Arid1a suppresses pancreatic neoplasia in a compartment-specific manner. In duct cells, this process appears to be associated with MYC-facilitated protein synthesis.