👤 Daisuke Okamura

Hepatoid adenocarcinoma (HAD) and adenocarcinoma with enteroblastic differentiation (ACED) are rare gastric cancer types that produce alpha-fetoprotein (AFP) that morphologically resembles the liver or fetal organ. This study investigated the molecular profiling of HAD and ACED compared with common-type gastric adenocarcinoma (CGA). This study enrolled 496 patients with gastric adenocarcinoma who underwent radical gastrectomy. Whole-exome sequencing and gene expression profiling were conducted to compare HAD/ACED and CGA. Immunohistochemistry was performed in 39 patients with HAD/ACED, including 10 who underwent genomic analysis. TP53 (100 %), CSMD3 (30 %), LRP1B, FAT3, TG, APOB, CREBBP, PASK, DROSHA, and STK40 (20 %) were mutated genes with high frequency (>20 %) in HAD/ACED. Gene expression analysis revealed 15 overexpressed genes in HAD/ACED, many of which were associated with hepatocytes and fetal organs. Furthermore, cancer stemness gene overexpression, including LIN28B, IGF2BP1, and HMGA2, which are related to TP53, was observed. Immunohistochemistry revealed positive staining for LIN28B (82 %), IGF2BP1 (94 %), and HMGA2 (72 %), as well as staining for AFP (69 %), GPC3 (75 %), and SALL4 (94 %). Additionally, positive cancer stemness gene staining was observed in CGA mucosa coexisting with HAD/ACED. HAD/ACED demonstrated higher TP53 mutation accumulation and TP53-related cancer stemness gene overexpression, including LIN28B, IGF2BP1, and HMGA2. Therefore, TP53 and these cancer stemness genes might be involved in the occurrence of HAD/ACED. Show less

Futibatinib is the only covalent inhibitor of FGFR1-4 to gain regulatory approval in oncology. In this article, we present genomic analyses of tissue biopsies and circulating tumor DNA (ctDNA) from patients with 1 of nearly 20 tumor types treated with futibatinib in the phase I/II FOENIX study. Eligible patients included those with ctDNA samples collected per protocol at baseline and/or progression on futibatinib in the phase Ib portion of the study for FGF/FGFR-altered advanced solid tumors or the phase II portion of the study for FGFR2 fusion/rearrangement-positive cholangiocarcinoma. Assessments included analytical concordance between tumor and ctDNA analyses for detection of FGFR alterations, association of ctDNA-detected co-occurring genomic alterations with response to futibatinib, and determination of patterns of acquired resistance following progression on futibatinib. Among 300 patients treated with futibatinib, 226 were eligible for this analysis, including 139 (62%) with cholangiocarcinoma. Among patients with known FGFR2 fusions/rearrangements, FGFR1 fusions, FGFR3 fusions, or FGFR2 amplifications per tissue analysis, detection rates in ctDNA for these aberrations were 84%, 0%, 11%, and 59%, respectively. Objective response rates on futibatinib were not significantly different between patients with TP53-altered versus -unaltered solid tumors; progression-free survival was reduced in patients with CDKN2B-altered versus -unaltered cholangiocarcinoma (median 4.8 versus 11.0 months; P = 0.03). Acquired resistance to futibatinib was frequently polyclonal and driven by an array of mutations within the relevant FGFR kinase domain, predominantly V565L, V565F, and N550K variants. In this largest and most systematic analysis of acquired resistance to an FGFR inhibitor from prospective clinical trials, emergence of secondary FGFR2 kinase domain mutations was observed in most patients receiving clinical benefit to futibatinib. ctDNA analysis shows clinically relevant potential as a noninvasive method for assessing genomic profiles, identifying patients who may benefit from FGFR inhibitor treatment, and exploring acquired resistance mechanisms. Show less

Computational modeling indicated that pathological high shear stress (HSS; 100 dyn/cm We used the Ibidi perfusion system to determine whether HSS applied to human PA endothelial cells (ECs) induces EndMT when compared with physiological laminar shear stress (15 dyn/cm EndMT, a feature of PAH not previously attributed to HSS, was observed. HSS did not alter the induction of transcription factors KLF (Krüppel-like factor) 2/4, but an ERG (ETS-family transcription factor) was reduced, as were histone H3 lysine 27 acetylation enhancer-promoter peaks containing ERG motifs. Consequently, there was reduced interaction between ERG and KLF2/4, a feature important in tethering KLF and the chromatin remodeling complex to DNA. In PA ECs under laminar shear stress, reducing ERG by siRNA caused EndMT associated with decreased BMPR2 (bone morphogenetic protein receptor 2), CDH5 (cadherin 5), and PECAM1 (platelet and EC adhesion molecule 1) and increased SNAI1/2 (Snail/Slug) and ACTA2 (smooth muscle α2 actin). In PA ECs under HSS, transfection of ERG prevented EndMT. HSS was then induced in mice by an aortocaval shunt, causing progressive PAH over 8 weeks. An adeno-associated viral vector (AAV2-ESGHGYF) was used to replenish ERG selectively in PA ECs. Elevated PA pressure, EndMT, and vascular remodeling (muscularization of peripheral arteries) in the aortocaval shunt mice were markedly reduced by ERG delivery. Pathological HSS reduced lung EC ERG, resulting in EndMT and PAH. Agents that upregulate ERG could reverse HSS-mediated PAH and occlusive vascular remodeling resulting from high flow or narrowed PAs. Show less

Epithelial-mesenchymal transition (EMT) is a cellular process in which epithelial cells lose their epithelial traits and shift to the mesenchymal phenotype, and is associated with various biological events, such as embryogenesis, wound healing and cancer progression. The transcriptional program that promotes phenotype switching is dynamically controlled by transcription factors during EMT, including Snail (SNAI1), twist family bHLH transcription factor (TWIST) and zinc finger E-box binding homeobox 1 (ZEB1). The present study aimed to investigate the molecular mechanisms underlying EMT in squamous epithelial cells. Western blot analysis and immunocytochemical staining identified Slug (SNAI2) as a transcription factor that is induced during transforming growth factor (TGF)-β1-mediated EMT in the human keratinocyte cell line HaCaT. The effect of SNAI2 overexpression and knockdown on the phenotypic characteristics of HaCaT cells was evaluated. Filamentous actin staining and western blot analysis revealed that the overexpression of SNAI2 did not induce the observed EMT-related phenotypic changes. In addition, SNAI2 knockdown demonstrated almost no impact on the EMT phenotypes induced by TGF-β1. Notably, DNA microarray analysis followed by comprehensive bioinformatics analysis revealed that the differentially expressed genes upregulated by TGF-β1 were significantly enriched in cell adhesion and extracellular matrix binding, whereas the genes downregulated in response to TGF-β1 were significantly enriched in the cell cycle. No enriched gene ontology term and biological pathways were identified in the differentially expressed gene sets of SNAI2-overexpressing cells. In addition, the candidates for master transcription factors regulating the TGF-β1-induced EMT were identified using transcription factor enrichment analysis. In conclusion, the results of study demonstrated that SNAI2 does not play an essential role in the EMT of HaCaT cells and identified candidate transcription factors that may be involved in EMT-related gene expression induced by TGF-β1. These findings may enhance the understanding of molecular events in EMT and contribute to the development of a novel therapeutic approach against EMT in cancers and wound healing. Show less

Cholesteryl ester transfer protein (CETP) mediates a step in reverse cholesterol transport, which channels cholesterol from peripheral tissues back to the liver. Mice and rats are CETP-deficient species, which assumedly contribute to rodent atherosclerosis resistance. Both pro- and anti-atherogenic effects have been shown in studies of CETP-transgenic rodent models thus far. As the results of pharmacological studies of CETP modification are largely controversial in humans, further knowledge about the impact of CETP on atherogenic phenotypes is required to evaluate its clinical utility for the prevention of cardiovascular and other organ damage associated with metabolic syndrome. Therefore, we newly generated a human CETP-transgenic (Tg[hCETP]) strain on the genetic background of spontaneously hypertensive rats (SHRs), which are characterized by the spontaneous occurrence of hypertension and insulin resistance. This allowed us to assess the in vivo role of CETP on cardiometabolic phenotypes in combination with hypertension. In Tg[hCETP] SHRs fed normal rat chow, systolic blood pressure was markedly elevated by 20-37 mmHg throughout the study period, and the development of fatty liver was accelerated with appreciable changes in the plasma lipid profile (HDL cholesterol reduction and triglyceride elevation). These phenotypic changes are in accordance with the assumption of proatherogenic effects inducible by the overexpression of CETP. However, with plasma LDL cholesterol levels concomitantly reduced, no apparent progression of atherosclerosis was detected in either the aorta or coronary arteries of Tg[hCETP] SHRs fed a high-fat, high-cholesterol diet. Our data provide new insight into the multifaceted regulation of cardiometabolic phenotypes via the modification of CETP. Show less

The cytokine, interleukin-18 (IL-18), was originally identified as an interferon-γ-inducing proinflammatory factor; however, there is increasing evidence suggesting that it has non-immunological effects on physiological functions. We have previously investigated the potential pathophysiological relationship between IL-18 and dyslipidemia, non-alcoholic fatty liver disease and non-alcoholic steatohepatitis, which were mediated by lipid energy imbalance. Therefore, herein we focused on brown adipocytes (BAs) and brown adipose tissue (BAT) related to energy consumption as non-shivering thermogenesis. Il18 Compared with Il18 This study demonstrated the critical function of IL-18 in differentiation and lipid metabolism in BAs. Furthermore, IL-18 may contribute to novel treatments by improving the energy imbalance. Show less

Cisplatin is an effective anticancer drug used to treat many types of cancer, including non-small cell lung carcinoma (NSCLCs), but development of resistance is the primary impediment in cancer treatment. Insulin-like growth factor-binding protein 7 (IGFBP7) is a secreted tumor suppressor that is inactivated in human lung cancer. IGFBP7 is known to alter sensitivity to interferon-based anticancer therapy, and here, we examined loss of IGFBP7 as a potential contributor to chemo-resistance to cisplatin. The transcriptional level of IGFBP7 was decreased in cisplatin-resistant human cancer cell lines and NSCLC xenografts. IGFBP7 knock-down increased cellular resistance to cisplatin and increased the level of mitogen-activated protein kinase phosphatases (MKP) 3 levels. The expression of MKP3 increased in a cisplatin-resistant NSCLC cell line and lung xenografts. MKP3 knock-down increased IGFBP7 level, indicating that MKP3 regulates IGFBP7. These findings suggest a novel molecular mechanism responsible for the tumor suppressive function of IGFBP7 in cisplatin-resistant human lung cancer and could lead to the development of IGFBP7 as a cisplatin-sensitizing agent. Show less

In mouse embryos, some primordial germ cells (PGCs) are eliminated by apoptosis, but the molecular pathways that lead to PGC survival versus apoptosis have not been fully characterized. Here, we found that REST (repressor element 1-silencing transcription factor), a transcription factor that binds a conserved regulatory element, NRSE/RE1, played a role in PGC survival. REST expression was higher in PGCs than in surrounding somatic cells. Moreover, in mouse embryos with a PGC-specific conditional REST mutation, the PGC population experienced more apoptosis and was significantly smaller than that in control embryos; these findings indicated that REST functioned in a cell-autonomous fashion that was critical for PGC survival. Several anti-apoptotic genes were among the previously identified REST-target gene candidates; moreover, some of these genes were downregulated in the REST-deficient PGCs. Mek5, which encodes a component in the a MAP kinase cascade, was one of these downregulated REST-target gene candidates, and a Mek5 mutation, like the REST mutation, caused an increase in PGC apoptosis; these finding suggested that REST promoted PGC survival via regulation of the Mek5 expression. Importantly, there were a normal number of PGCs in the REST mutants at birth, and both the male and female REST-mutant adults were fertile; these final observations revealed that the PGC population was very robust and could recover from a genetically induced reduction in cell number. Show less

We identified a novel gene fusion of ANKRD28 (ankyrin repeat domain 28) on 3p25 to NUP98 on 11p15 in a patient with adult myelodysplastic syndrome/acute myelogenous leukemia. A partially cryptic 3-way translocation, t(3;5;11)(p25;q35;p15), that had initially been supposed to be t(3;5)(p25;q35) was revealed by precise breakpoint mapping via fluorescence in situ hybridization analysis with bacterial artificial chromosome clones. This translocation produces the expression of 2 in-frame fusion transcripts, the novel ANKRD28-NUP98 and NUP98-NSD1, and 1 out-of-frame NSD1-ANKRD28 transcript. Transient overexpression of ANKRD28-NUP98 in NIH/3T3 cells, but not the C-terminal deletion mutant of ANKRD28 (DeltaC-ANKRD28), caused significantly increased focus formation compared with mock-transfectant controls. ANKRD28-NUP98 was localized in the nucleolus and cytoplasm, whereas ANKRD28 and DeltaC-ANKRD28 were found exclusively in the cytoplasm. Alteration of the subcellular localization of ANKRD28 might have contributed to the leukemogenesis in this case. This report is the first of ANKRD28 as an NUP98 fusion partner, and this case implies that this fusion may be responsible for hematologic malignancies. Show less

Signalling by the Wnt family of secreted lipoproteins has essential functions in development and disease. The canonical Wnt/beta-catenin pathway requires a single-span transmembrane receptor, low-density lipoprotein (LDL)-receptor-related protein 6 (LRP6), whose phosphorylation at multiple PPPSP motifs is induced upon stimulation by Wnt and is critical for signal transduction. The kinase responsible for LRP6 phosphorylation has not been identified. Here we provide biochemical and genetic evidence for a 'dual-kinase' mechanism for LRP6 phosphorylation and activation. Glycogen synthase kinase 3 (GSK3), which is known for its inhibitory role in Wnt signalling through the promotion of beta-catenin phosphorylation and degradation, mediates the phosphorylation and activation of LRP6. We show that Wnt induces sequential phosphorylation of LRP6 by GSK3 and casein kinase 1, and this dual phosphorylation promotes the engagement of LRP6 with the scaffolding protein Axin. We show further that a membrane-associated form of GSK3, in contrast with cytosolic GSK3, stimulates Wnt signalling and Xenopus axis duplication. Our results identify two key kinases mediating Wnt co-receptor activation, reveal an unexpected and intricate logic of Wnt/beta-catenin signalling, and illustrate GSK3 as a genuine switch that dictates both on and off states of a pivotal regulatory pathway. Show less