Switch of ELF3 and ATF4 transcriptional axis programs the amino acid insufficiency-linked epithelial-to-mesenchymal transition.

Epithelial-to-mesenchymal transition (EMT) that endows cancer cells with increased invasive and migratory capacity enables cancer dissemination and metastasis. This process is tightly associated with metabolic reprogramming acquired for rewiring cell status and signaling pathways for survival in dietary insufficiency conditions. However, it remains largely unclear how transcription factor (TF)-mediated transcriptional programs are modulated during the EMT process. Here, we reveal that depletion of a key epithelial TF, ELF3 (E74-like factor-3), triggers a transforming growth factor ß (TGF-ß) signaling activation-like mesenchymal transcriptomic profile and metastatic features linked to the aminoacyl-tRNA biogenesis pathway. Moreover, the transcriptome alterations elicited by ELF3 depletion perfectly resemble an ATF4-dependent weak response to amino acid starvation. Intriguingly, we observe an exclusive enrichment of ELF3 and ATF4 in epithelial and TGF-ß-induced or ELF3-depletion-elicited mesenchymal enhancers, respectively, with rare co-binding on altered enhancers. We also find that the upregulation of aminoacyl-tRNA synthetases and some mesenchymal genes upon amino acid deprivation is diminished in ATF4-depleted cells. In sum, the loss of ELF3 binding on epithelial enhancers and the gain of ATF4 binding on the enhancers of mesenchymal factors and amino acid deprivation responsive genes facilitate the loss of epithelial cell features and the gain of TGF-ß-signaling-associated mesenchymal signatures, which further promote lung cancer cell metastasis.

CDK7 inhibition suppresses aberrant hedgehog pathway and overcomes resistance to smoothened antagonists.

The aberrant hedgehog (Hh) pathway plays important roles in multiple cancer types, therefore serving as a promising drug target. Current clinically available hedgehog-targeted drugs act mostly by antagonizing the upstream component smoothened; however, both primary and acquired resistance to FDA-approved smoothened inhibitor (SMOi) drugs have been described. We have recently demonstrated that the BET inhibitor effectively suppresses SMOi-resistant Hh-driven cancers through antagonizing transcription of GLI1 and GLI2, the core transcriptional factors of Hh pathway, suggesting epigenetic or transcriptional targeted therapy represents an anti-Hh therapeutic strategy that can overcome SMOi resistance. Here we performed an unbiased screening of epigenetic or transcriptional targeted small molecules to test their inhibitory effects on GLI1 and GLI2 transcription or cell viability of Hh-driven tumor lines. THZ1, a covalent inhibitor of cyclin-dependent kinase 7 (CDK7), is identified as the top hit in our screening. We then confirmed that antagonizing CDK7 by either small-molecule inhibitors or the CRISPR-Cas9 approach causes substantial suppression of GLI1 and GLI2 transcription, resulting in effective inhibition of Hh-driven cancers in vitro and in vivo. More importantly, antagonizing CDK7 retains inhibitory activity against Hh-driven cancers with almost all so-far described primary or acquired SMOi resistance. Furthermore, we reveal a synergy between CDK7 inhibition and BET inhibition on antagonizing aberrant Hh pathway and Hh-driven cancers that are either responsive or resistant to SMOi. Our results illustrate transcriptional inhibition through targeting CDK7 as a promising therapeutic strategy for treating Hh-driven cancers, especially those with primary or acquired resistance to SMOi drugs.

Protein tyrosine phosphatase receptor type R (PTPRR) antagonizes the Wnt signaling pathway in ovarian cancer by dephosphorylating and inactivating ß-catenin.

Despite a lack of mutations, accumulating evidence supports an important role for the Wnt/ß-catenin pathway in ovarian tumorigenesis. However, the molecular mechanism that contributes to the aberrant activation of the Wnt signaling cascade in ovarian cancer has not been fully elucidated. Here, we found that protein tyrosine phosphatase receptor type R (PTPRR) suppressed the activation of the Wnt/ß-catenin pathway in ovarian cancer. We performed an shRNA-based biochemical screen, which identified PTPRR as being responsible for tyrosine dephosphorylation of ß-catenin on Tyr-142, a key site controlling the transcriptional activity of ß-catenin. Of note, PTPRR was down-regulated in ovarian cancers, and ectopic PTPRR re-expression delayed ovarian cancer cell growth both in vitro and in vivo Using a proximity-based tagging system and RNA-Seq analysis, we identified a signaling nexus that includes PTPRR, α-catenin, ß-catenin, E-cadherin, and AT-rich interaction domain 3C (ARID3C) in ovarian cancer. Immunohistochemistry staining of human samples further suggested that PTPRR expression is inversely correlated with disease prognosis. Collectively, our findings indicate that PTPRR functions as a tumor suppressor in ovarian cancer by dephosphorylating and inactivating ß-catenin. These results suggest that PTPRR expression might have utility as a prognostic marker for predicting overall survival.

Swc4 protects nucleosome-free rDNA, tDNA and telomere loci to inhibit genome instability.

In the baker's yeast Saccharomyces cerevisiae, NuA4 and SWR1-C, two multisubunit complexes, are involved in histone acetylation and chromatin remodeling, respectively. Eaf1 is the assembly platform subunit of NuA4, Swr1 is the assembly platform and catalytic subunit of SWR1-C, while Swc4, Yaf9, Arp4 and Act1 form a functional module, and is present in both NuA4 and SWR1 complexes. ACT1 and ARP4 are essential for cell survival. Deletion of SWC4, but not YAF9, EAF1 or SWR1 results in a severe growth defect, but the underlying mechanism remains largely unknown. Here, we show that swc4Δ, but not yaf9Δ, eaf1Δ, or swr1Δ cells display defects in DNA ploidy and chromosome segregation, suggesting that the defects observed in swc4Δ cells are independent of NuA4 or SWR1-C integrity. Swc4 is enriched in the nucleosome-free regions (NFRs) of the genome, including characteristic regions of RDN5s, tDNAs and telomeres, independently of Yaf9, Eaf1 or Swr1. In particular, rDNA, tDNA and telomere loci are more unstable and prone to recombination in the swc4Δ cells than in wild-type cells. Taken together, we conclude that the chromatin associated Swc4 protects nucleosome-free chromatin of rDNA, tDNA and telomere loci to ensure genome integrity.

Induction of senescence-associated secretory phenotype underlies the therapeutic efficacy of PRC2 inhibition in cancer.

The methyltransferase Polycomb Repressive Complex 2 (PRC2), composed of EZH2, SUZ12, and EED subunits, is associated with transcriptional repression via tri-methylation of histone H3 on lysine 27 residue (H3K27me3). PRC2 is a valid drug target, as the EZH2 gain-of-function mutations identified in patient samples drive tumorigenesis. PRC2 inhibitors have been discovered and demonstrated anti-cancer efficacy in clinic. However, their pharmacological mechanisms are poorly understood. MAK683 is a potent EED inhibitor in clinical development. Focusing on MAK683-sensitive tumors with SMARCB1 or ARID1A loss, we identified a group of PRC2 target genes with high H3K27me3 signal through epigenomic and transcriptomic analysis. Multiple senescence-associated secretory phenotype (SASP) genes, such as GATA4, MMP2/10, ITGA2 and GBP1, are in this group besides previously identified CDKN2A/p16. Upon PRC2 inhibition, the de-repression of SASP genes is detected in multiple sensitive models and contributes to decreased Ki67+, extracellular matrix (ECM) reorganization, senescence associated inflammation and tumor regression even in CDKN2A/p16 knockout tumor. And the combination of PRC2 inhibitor and CDK4/6 inhibitor leads to better effect. The genes potential regulated by PRC2 in neuroblastoma samples exhibited significant enrichment of ECM and senescence associated inflammation, supporting the clinical relevance of our results. Altogether, our results unravel the pharmacological mechanism of PRC2 inhibitors and propose a combination strategy for MAK683 and other PRC2 drugs.

FER-mediated phosphorylation and PIK3R2 recruitment on IRS4 promotes AKT activation and tumorigenesis in ovarian cancer cells.

Tyrosine phosphorylation, orchestrated by tyrosine kinases and phosphatases, modulates a multi-layered signaling network in a time- and space-dependent manner. Dysregulation of this post-translational modification is inevitably associated with pathological diseases. Our previous work has demonstrated that non-receptor tyrosine kinase FER is upregulated in ovarian cancer, knocking down which attenuates metastatic phenotypes. However, due to the limited number of known substrates in the ovarian cancer context, the molecular basis for its pro-proliferation activity remains enigmatic. Here, we employed mass spectrometry and biochemical approaches to identify insulin receptor substrate 4 (IRS4) as a novel substrate of FER. FER engaged its kinase domain to associate with the PH and PTB domains of IRS4. Using a proximity-based tagging system in ovarian carcinoma-derived OVCAR-5 cells, we determined that FER-mediated phosphorylation of Tyr779 enables IRS4 to recruit PIK3R2/p85ß, the regulatory subunit of PI3K, and activate the PI3K-AKT pathway. Rescuing IRS4-null ovarian tumor cells with phosphorylation-defective mutant, but not WT IRS4 delayed ovarian tumor cell proliferation both in vitro and in vivo. Overall, we revealed a kinase-substrate mode between FER and IRS4, and the pharmacological inhibition of FER kinase may be beneficial for ovarian cancer patients with PI3K-AKT hyperactivation.

Phylogenetic Tree Inference: A Top-Down Approach to Track Tumor Evolution.

Recently, an increasing number of studies sequence multiple biopsies of primary tumors, and even paired metastatic tumors to understand heterogeneity and the evolutionary trajectory of cancer progression. Although several algorithms are available to infer the phylogeny, most tools rely on accurate measurements of mutation allele frequencies from deep sequencing, which is often hard to achieve for clinical samples (especially FFPE samples). In this study, we present a novel and easy-to-use method, PTI (Phylogenetic Tree Inference), which use an iterative top-down approach to infer the phylogenetic tree structure of multiple tumor biopsies from same patient using just the presence or absence of somatic mutations without their allele frequencies. Therefore PTI can be used in a wide range of cases even when allele frequency data is not available. Comparison with existing state-of-the-art methods, such as LICHeE, Treeomics, and BAMSE, shows that PTI achieves similar or slightly better performance within a short run time. Moreover, this method is generally applicable to infer phylogeny for any other data sets (such as epigenetics) with a similar zero and one feature-by-sample matrix.

Treatment of femoral shaft fractures by closed reduction and interlocking intramedullary nailing / 中国骨伤

<p><b>OBJECTIVE</b>To discuss the clinical effect of closed reduction and interlocking intramedullary nailing in the treatment of femoral shaft fracture.</p><p><b>METHODS</b>From March 2006 to December 2011,103 patients with femoral shaft fracture were treated by closed reduction and interlocking intramedullary nailing including 76 males and 27 females with an average age of 36 years old ranging from 19 to 55 years old. According to AO classification,there were 64 cases with type A,27 with type B, 12 with type C. Thirteen cases were open fractures including 5 cases with Gustilo type I , 8 with Gustilo type II . The time of bone healing were observed after operation, the knee function recovery was evaluated by HSS scoring standard at 1 year after operation.</p><p><b>RESULTS</b>The intraoperative complications included femoral neck fracture in 1 case and proximal femoral fracture in 1 case,both of the patients were treated with reconstructive intramedullary interlocking nail and the fractures healed postoperatively. One patient was suffered from common peroneal nerve injury,which were fully recovered at 4 months later after medical treatment. All the patients were followed up from 12 to 28 months (averaged 22 months). All of the fractures were healed well and the average healing time was 3 to 9 months (averaged 5 months). All the hip joints were recovered to normal function. The average HSS was 90.89±5.06 at 1 year after operation.</p><p><b>CONCLUSION</b>Interlocking intramedullary nailing is the preferred treatment for patients with femoral shaft fracture. Carefully operating and closed reduction can reduce the complications.</p>