Publications by authors named "Shumpei Ishikawa"

95 Publications

Helicobacter pylori CagA elicits BRCAness to induce genome instability that may underlie bacterial gastric carcinogenesis.

Cell Host Microbe 2021 06 13;29(6):941-958.e10. Epub 2021 May 13.

Department of Microbiology, Graduate School of Medicine, the University of Tokyo, Tokyo 113-0033, Japan. Electronic address:

Infection with CagA-producing Helicobacter pylori plays a causative role in the development of gastric cancer. Upon delivery into gastric epithelial cells, CagA deregulates prooncogenic phosphatase SHP2 while inhibiting polarity-regulating kinase PAR1b through complex formation. Here, we show that CagA/PAR1b interaction subverts nuclear translocation of BRCA1 by inhibiting PAR1b-mediated BRCA1 phosphorylation. It hereby induces BRCAness that promotes DNA double-strand breaks (DSBs) while disabling error-free homologous recombination-mediated DNA repair. The CagA/PAR1b interaction also stimulates Hippo signaling that circumvents apoptosis of DNA-damaged cells, giving cells time to repair DSBs through error-prone mechanisms. The DSB-activated p53-p21 axis inhibits proliferation of CagA-delivered cells, but the inhibition can be overcome by p53 inactivation. Indeed, sequential pulses of CagA in TP53-mutant cells drove somatic mutation with BRCAness-associated genetic signatures. Expansion of CagA-delivered cells with BRCAness-mediated genome instability, from which CagA-independent cancer-predisposing cells arise, provides a plausible "hit-and-run mechanism" of H. pylori CagA for gastric carcinogenesis.
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http://dx.doi.org/10.1016/j.chom.2021.04.006DOI Listing
June 2021

High levels of human epididymis protein 4 mRNA and protein expression are associated with chemoresistance and a poor prognosis in pancreatic cancer.

Int J Oncol 2021 01 12;58(1):57-69. Epub 2020 Nov 12.

Department of Clinical Diagnostic Oncology, Clinical Research Institute for Clinical Pharmacology and Therapeutics, Showa University, Tokyo 157‑8577, Japan.

Pancreatic cancer is associated with an exceedingly poor prognosis, warranting the development of novel therapeutic strategies and discovery of prognostic predictors. Given that chemoresistance‑related molecules are reportedly associated with the poor prognosis of pancreatic cancer, the present study aimed to identify molecules that could be efficacious therapeutic targets for pancreatic cancer. First, 10 patient‑derived xenografts (PDXs) were established from patients with pancreatic cancer. Subsequently, after treating tumor tissue generated from the PDXs with standard drugs, next‑generation sequencing (NGS) was performed using these tissues. The results of NGS analysis and immunohistochemical analysis on 80 pancreatic cancer tissues revealed that human epididymis protein 4 (HE4) expression in the anticancer drug‑treated PDX group was higher than that in the untreated PDXs. In addition, chemoresistance ability was observed in tumor cell lines overexpressing HE4. Furthermore, Kaplan‑Meier analysis of tumor tissues from 80 patients with pancreatic cancer was performed and it was found that patients with a high HE4 expression level had a poor survival rate compared with those who had a low HE4 expression level. Multivariate analysis also indicated the high expression level of HE4 was an independent poor prognostic biomarker. Thus, it was concluded that high gene and protein expression levels of HE4 mediate chemoresistance and are independent prognostic factors for pancreatic cancer.
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http://dx.doi.org/10.3892/ijo.2020.5147DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7721086PMC
January 2021

Focal adhesion ribonucleoprotein complex proteins are major humoral cancer antigens and targets in autoimmune diseases.

Commun Biol 2020 10 16;3(1):588. Epub 2020 Oct 16.

Department of Preventive Medicine, Graduate School of Medicine, the University of Tokyo, Tokyo, Japan.

Despite the accumulating evidences of the significance of humoral cancer immunity, its molecular mechanisms have largely remained elusive. Here we show that B-cell repertoire sequencing of 102 clinical gastric cancers and molecular biological analyses unexpectedly reveal that the major humoral cancer antigens are not case-specific neo-antigens but are rather commonly identified as ribonucleoproteins (RNPs) in the focal adhesion complex. These common antigens are shared as autoantigens with multiple autoimmune diseases, suggesting a direct molecular link between cancer- and auto-immunity on the focal adhesion RNP complex. This complex is partially exposed to the outside of cancer cell surfaces, which directly evokes humoral immunity and enables functional bindings of antibodies to cancer cell surfaces in physiological conditions. These findings shed light on humoral cancer immunity in that it commonly targets cellular components fundamental for cytoskeletal integrity and cell movement, pointing to a novel modality of immunotherapy using humoral immunological reactions to cancers.
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http://dx.doi.org/10.1038/s42003-020-01305-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7567837PMC
October 2020

Clinicopathological and molecular analyses of linearly expanded epithelial cells with β-catenin alterations, "β-catenin signature", in the normal fallopian tube.

Histopathology 2020 Dec 17;77(6):880-889. Epub 2020 Oct 17.

Department of Cellular and Organ Pathology, Graduate School of Medicine, Akita University, Akita, Japan.

Aims: Recent advances in next-generation sequencing have made it clear that clonal expansion of cells harbouring driver gene mutations occurs in physiologically normal epithelium. Molecular analysis of tubal epithelium has been almost exclusively confined to the TP53 pathway, which is involved in serous carcinogenesis. Other oncogenic events have not been explored in detail. Here, we report the linear expansion of fallopian tubal epithelial cells exhibiting an altered β-catenin profile (β-catenin signature). Through molecular analyses, we determined the incidence and clinicopathological significance of β-catenin signatures.

Methods And Results: We evaluated 64 specimens of surgically removed bilateral fallopian tubes. Thirty-three β-catenin signatures were identified in 13 cases (20.3%); these patients were significantly younger than those without β-catenin signatures (median ages of 44 and 57 years, respectively, P = 0.0317). No correlation between β-catenin signature and any clinical factor was observed. CTNNB1 mutations were detected in three of eight β-catenin signatures when tissues were microdissected and subjected to Sanger sequencing in two representative cases.

Conclusions: This is the first report of the CTNNB1 mutation in clusters of morphologically bland tubal epithelial cells. The results of this study indicate that β-catenin signatures are common, and they may be a part of diverse molecular alterations occurring in normal tubal epithelium.
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http://dx.doi.org/10.1111/his.14227DOI Listing
December 2020

Accumulation of Molecular Aberrations Distinctive to Hepatocellular Carcinoma Progression.

Cancer Res 2020 09 8;80(18):3810-3819. Epub 2020 Jul 8.

Genome Science Division, RCAST, University of Tokyo, Tokyo, Japan.

Cancer develops through the accumulation of genetic and epigenetic aberrations. To identify sequential molecular alterations that occur during the development of hepatocellular carcinoma (HCC), we compared 52 early and 108 overt HCC samples by genome sequencing. Gene mutations in the p53/RB1 pathway, WNT pathway, MLL protein family, SWI/SNF complexes, and AKT/PI3K pathway were common in HCC. In the early phase of all entities, was the most frequently upregulated gene owing to diverse mechanisms. Despite frequent somatic mutations in driver genes, including and , early HCC was a separate molecular entity from overt HCC, as each had a distinct expression profile. Notably, WNT target genes were not activated in early HCC regardless of mutation status because β-catenin did not translocate into the nucleus due to the E-cadherin/β-catenin complex at the membrane. Conversely, WNT targets were definitively upregulated in overt HCC, with mutation associated with downregulation of and hypomethylation of CpG islands in target genes. Similarly, cell-cycle genes downstream of the p53/RB pathway were upregulated only in overt HCC, with or gene mutations associated with chromosomal deletion of 4q or 16q. HCC was epigenetically distinguished into four subclasses: normal-like methylation, global-hypomethylation (favorable prognosis), stem-like methylation (poor prognosis), and CpG island methylation. These methylation statuses were globally maintained through HCC progression. Collectively, these data show that as HCC progresses, additional molecular events exclusive of driver gene mutations cooperatively contribute to transcriptional activation of downstream targets according to methylation status. SIGNIFICANCE: In addition to driver gene mutations in the WNT and p53 pathways, further molecular events are required for aberrant transcriptional activation of these pathways as HCC progresses.
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http://dx.doi.org/10.1158/0008-5472.CAN-20-0225DOI Listing
September 2020

High expression levels of polymeric immunoglobulin receptor are correlated with chemoresistance and poor prognosis in pancreatic cancer.

Oncol Rep 2020 Jul 11;44(1):252-262. Epub 2020 May 11.

Department of Clinical Diagnostic Oncology, Clinical Research Institute for Clinical Pharmacology and Therapeutics, Showa University, Tokyo 157‑8577, Japan.

Pancreatic cancer has extremely poor prognosis, warranting the discovery of novel therapeutic and prognostic markers. The expression of polymeric immunoglobulin receptor (pIgR), a key component of the mucosal immune system, is increased in several cancers. However, its clinical relevance in pancreatic cancer remains unclear. In the present study, the prognostic value of pIgR in pancreatic cancer patients after surgical resection was assessed and it was determined that the expression of pIgR was correlated with poor prognosis. Ten pancreatic cancer patient‑derived xenograft (PDX) lines were established, followed by next‑generation sequencing of tumor tissues from these lines after standard chemotherapy. Immunohistochemical analysis of chemoresistance‑related molecules using 77 pancreatic cancer tissues was also performed. The expression of pIgR mRNA in the PDX group treated with anticancer drugs was higher than in the untreated group. High pIgR expression in tissue specimens from 77 pancreatic cancer patients was significantly associated with poor prognosis and was revealed to be an independent prognostic factor, predicting poor outcomes. High pIgR mRNA and protein levels were independent prognostic factors, indicating that pIgR could be a novel predictor for poor prognosis of pancreatic cancer patients.
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http://dx.doi.org/10.3892/or.2020.7610DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7251687PMC
July 2020

Miclxin, a Novel MIC60 Inhibitor, Induces Apoptosis via Mitochondrial Stress in β-Catenin Mutant Tumor Cells.

ACS Chem Biol 2020 08 10;15(8):2195-2204. Epub 2020 Jul 10.

Department of Biosciences and Informatics, Faculty of Science and Technology, Keio University, Yokohama 223-8522, Japan.

The Wnt signaling pathway regulates diverse cellular processes. β-Catenin is one of the major components of this pathway, in which it plays a main role. Although it has been established that β-catenin is mutated in a wide variety of tumors, there are currently no effective therapeutic agents that target β-catenin. In this study, we searched for the compound that targets mutant β-catenin and found DS37262926 (miclxin). Miclxin exhibited β-catenin-dependent apoptosis in β-catenin-mutated HCT116 cells and isogenic HCT116 ( Δ45/-) cells; however, this effect was not observed in isogenic HCT116 ( +/-) cells. Using miclxin-immobilized beads, MIC60, one of the major components of the mitochondrial contact site and cristae organizing system (MICOS) complex, was identified as a target protein of miclxin. We revealed that MIC60 dysfunction caused by miclxin induced a mitochondrial stress response in a mutant β-catenin-dependent manner. Activation of the mitochondrial stress response was responsible for the downregulation of Bcl-2, leading to severe loss of mitochondrial membrane potential and subsequent apoptosis-inducing factor-dependent apoptosis. Our findings suggest that targeting MIC60 is a potential strategy with which tumor cells can be killed through induction of severe mitochondrial damage in a mutant β-catenin-dependent manner.
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http://dx.doi.org/10.1021/acschembio.0c00381DOI Listing
August 2020

Defined lifestyle and germline factors predispose Asian populations to gastric cancer.

Sci Adv 2020 May 6;6(19):eaav9778. Epub 2020 May 6.

Genome Science Division, Research Center for Advanced Science and Technology (RCAST), The University of Tokyo, Tokyo, Japan.

Germline and environmental effects on the development of gastric cancers (GC) and their ethnic differences have been poorly understood. Here, we performed genomic-scale trans-ethnic analysis of 531 GCs (319 Asian and 212 non-Asians). There was one distinct GC subclass with clear alcohol-associated mutation signature and strong Asian specificity, almost all of which were attributable to alcohol intake behavior, smoking habit, and Asian-specific defective allele. Alcohol-related GCs have low mutation burden and characteristic immunological profiles. In addition, we found frequent (7.4%) germline variants among Japanese GCs, most of which were attributed to a few recurrent single-nucleotide variants shared by Japanese and Koreans, suggesting the existence of common ancestral events among East Asians. Specifically, approximately one-fifth of diffuse-type GCs were attributable to the combination of alcohol intake and defective allele or to variants. These results revealed uncharacterized impacts of germline variants and lifestyles in the high incidence areas.
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http://dx.doi.org/10.1126/sciadv.aav9778DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7202881PMC
May 2020

Reciprocal expression of trefoil factor-1 and thyroid transcription factor-1 in lung adenocarcinomas.

Cancer Sci 2020 Jun 30;111(6):2183-2195. Epub 2020 Apr 30.

Division of Integrative Pathology, Jichi Medical University, Shimotsuke, Japan.

Molecular targeted therapies against EGFR and ALK have improved the quality of life of lung adenocarcinoma patients. However, targetable driver mutations are mainly found in thyroid transcription factor-1 (TTF-1)/NK2 homeobox 1 (NKX2-1)-positive terminal respiratory unit (TRU) types and rarely in non-TRU types. To elucidate the molecular characteristics of the major subtypes of non-TRU-type adenocarcinomas, we analyzed 19 lung adenocarcinoma cell lines (11 TRU types and 8 non-TRU types). A characteristic of non-TRU-type cell lines was the strong expression of TFF-1 (trefoil factor-1), a gastric mucosal protective factor. An immunohistochemical analysis of 238 primary lung adenocarcinomas resected at Jichi Medical University Hospital revealed that TFF-1 was positive in 31 cases (13%). Expression of TFF-1 was frequently detected in invasive mucinous (14/15, 93%), enteric (2/2, 100%), and colloid (1/1, 100%) adenocarcinomas, less frequent in acinar (5/24, 21%), papillary (7/120, 6%), and solid (2/43, 5%) adenocarcinomas, and negative in micropapillary (0/1, 0%), lepidic (0/23, 0%), and microinvasive adenocarcinomas or adenocarcinoma in situ (0/9, 0%). Expression of TFF-1 correlated with the expression of HNF4-α and MUC5AC (P < .0001, P < .0001, respectively) and inversely correlated with that of TTF-1/NKX2-1 (P < .0001). These results indicate that TFF-1 is characteristically expressed in non-TRU-type adenocarcinomas with gastrointestinal features. The TFF-1-positive cases harbored KRAS mutations at a high frequency, but no EGFR or ALK mutations. Expression of TFF-1 correlated with tumor spread through air spaces, and a poor prognosis in advanced stages. Moreover, the knockdown of TFF-1 inhibited cell proliferation and soft-agar colony formation and induced apoptosis in a TFF-1-high and KRAS-mutated lung adenocarcinoma cell line. These results indicate that TFF-1 is not only a biomarker, but also a potential molecular target for non-TRU-type lung adenocarcinomas.
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http://dx.doi.org/10.1111/cas.14403DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7293082PMC
June 2020

The first case of gastric carcinoma with NTRK rearrangement: identification of a novel ATP1B-NTRK1 fusion.

Gastric Cancer 2020 09 18;23(5):944-947. Epub 2020 Mar 18.

Department of Pathology, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan.

NTRK gene rearrangements occur in a wide spectrum of tumors and are actionable events predictive of response to TRK inhibitor. We report the first case of gastric carcinoma harboring a NTRK fusion in a 79-year-old man. The tumor was composed predominantly of poorly cohesive carcinoma with focal tubular differentiation. Solid sheet-like or nested pattern of large oxyphilic cells was also noted in 10% of tumor. Pan-Trk immunohistochemistry demonstrated Trk expression with a diffuse cytoplasmic and dot-like staining only in the solid component. Extensive lymphatic invasion and multiple nodal metastases were noted and were predominated by Trk-positive component. A novel ATP1B1-NTRK1 fusion was detected by RNA-seq using fresh frozen sample. The patient died of the disease, despite surgery and chemotherapy. Although extremely rare, NTRK rearrangement does occur in gastric carcinoma and might be associated with aggressive phenotype as well as histologic features like solid growth with extensive lymphatic invasion.
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http://dx.doi.org/10.1007/s10120-020-01061-9DOI Listing
September 2020

Post-mortem Plasma Cell-Free DNA Sequencing: Proof-of-Concept Study for the "Liquid Autopsy".

Sci Rep 2020 02 7;10(1):2120. Epub 2020 Feb 7.

Department of Cellular and Organ Pathology, Graduate School of Medicine, Akita University, Akita, Japan.

Recent genomic studies on cancer tissues obtained during rapid autopsy have provided insights into the clonal evolution and heterogeneity of cancer. However, post-mortem blood has not been subjected to genetic analyses in relation to cancer. We first confirmed that substantial quantities of cell-free DNA were present in the post-mortem plasma of 12 autopsy cases. Then, we focused on a pilot case of prostate cancer with multiple metastases for genetic analyses. Whole-exome sequencing of post-mortem plasma-derived cell-free DNA and eight frozen metastatic cancer tissues collected during rapid autopsy was performed, and compared their mutational statuses. The post-mortem plasma cell-free DNA was successfully sequenced and 344 mutations were identified. Of these, 160 were detected in at least one of the metastases. Further, 99% of the mutations shared by all metastases were present in the plasma. Sanger sequencing of 30 additional formalin-fixed metastases enabled us to map the clones harboring mutations initially detected only in the plasma. In conclusion, post-mortem blood, which is usually disposed of during conventional autopsies, can provide valuable data if sequenced in detail, especially regarding cancer heterogeneity. Furthermore, post-mortem plasma cell-free DNA sequencing (liquid autopsy) can be a novel platform for cancer research and a tool for genomic pathology.
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http://dx.doi.org/10.1038/s41598-020-59193-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7005783PMC
February 2020

High expression of olfactomedin-4 is correlated with chemoresistance and poor prognosis in pancreatic cancer.

PLoS One 2020 10;15(1):e0226707. Epub 2020 Jan 10.

Department of Clinical Diagnostic Oncology, Clinical Research Institute for Clinical Pharmacology & Therapeutics, Showa University, Tokyo, Japan.

Pancreatic cancer has an extremely poor prognosis, and identification of novel predictors of therapeutic efficacy and prognosis is urgently needed. Chemoresistance-related molecules are correlated with poor prognosis and may be effective targets for cancer treatment. Here, we aimed to identify novel molecules correlated with chemoresistance and poor prognosis in pancreatic cancer. We established 10 patient-derived xenograft (PDX) lines from patients with pancreatic cancer and performed next-generation sequencing (NGS) of tumor tissues from PDXs after treatment with standard drugs. We established a gene-transferred tumor cell line to express chemoresistance-related molecules and analyzed the chemoresistance of the established cell line against standard drugs. Finally, we performed immunohistochemical (IHC) analysis of chemoresistance-related molecules using 80 pancreatic cancer tissues. From NGS analysis, we identified olfactomedin-4 (OLFM4) as having high expression in the PDX group treated with anticancer drugs. In IHC analysis, OLFM4 expression was also high in PDXs administered anticancer drugs compared with that in untreated PDXs. Chemoresistance was observed by in vitro analysis of tumor cell lines with forced expression of OLFM4. In an assessment of tissue specimens from 80 patients with pancreatic cancer, Kaplan-Meier analysis showed that patients in the low OLFM4 expression group had a better survival rate than patients in the high OLFM4 expression group. Additionally, multivariate analysis showed that high expression of OLFM4 was an independent prognostic factor predicting poor outcomes. Overall, our study revealed that high expression of OLFM4 was involved in chemoresistance and was an independent prognostic factor in pancreatic cancer. OLFM4 may be a candidate therapeutic target in pancreatic cancer.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0226707PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6953839PMC
April 2020

Novel targets identified by integrated cancer-stromal interactome analysis of pancreatic adenocarcinoma.

Cancer Lett 2020 01 24;469:217-227. Epub 2019 Oct 24.

Kanagawa Cancer Center Research Institute, Yokohama, Japan.

The pancreatic cancer microenvironment is crucial in cancer development, progression and drug resistance. Cancer-stromal interactions have been recognized as important targets for cancer therapy. However, identifying relevant and druggable cancer-stromal interactions is challenging due to the lack of quantitative methods to analyze the whole cancer-stromal interactome. Here we studied 14 resected pancreatic cancer specimens (8 pancreatic adenocarcinoma (PDAC) patients as a cancer group and 6 intraductal papillary-mucinous adenoma (IPMA) patients as a control). Shotgun proteomics of the stromal lesion dissected with laser captured microdissection (LCM) was performed, and identified 102 differentially expressed proteins in pancreatic cancer stroma. Next, we obtained gene expression data in human pancreatic cancer and normal pancreatic tissue from The Cancer Genome Atlas database (n = 169) and The Genotype-Tissue Expression database (n = 197), and identified 1435 genes, which were differentially expressed in pancreatic cancer cells. To identify relevant and druggable cancer-stromal-interaction targets, we applied these datasets to our in-house ligand-receptor database. Finally, we identified 9 key genes and 8 key cancer-stromal-interaction targets for PDAC patients. Furthermore, we examined FN1 and ITGA3 protein expression in pancreatic cancer tissues using tissue microarrays (TMAs) of 271 PDAC cases, and demonstrated that FN1-ITGA3 had unfavorable prognostic impact for PDAC patients.
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http://dx.doi.org/10.1016/j.canlet.2019.10.031DOI Listing
January 2020

In vivo effects of mutant RHOA on tumor formation in an orthotopic inoculation model.

Oncol Rep 2019 Nov 3;42(5):1745-1754. Epub 2019 Sep 3.

Department of Research, Forerunner Pharma Research Co., Ltd., Komaba Open Laboratory, The University of Tokyo, Tokyo 153‑8904, Japan.

Ras homolog family member A (RHOA) mutations are driver genes in diffuse‑type gastric cancers (DGCs), and we previously revealed that RHOA mutations contribute to cancer cell survival and cell migration through their dominant negative effect on Rho‑associated kinase (ROCK) signaling in vitro. However, how RHOA mutations contribute to DGC development in vivo is poorly understood. In the present study, the contribution of RHOA mutations to tumor morphology was investigated using an orthotopic xenograft model using the gastric cancer cell line MKN74, in which wild‑type (WT) or mutated (Y42C and Y42S) RHOA had been introduced. When we conducted RNA sequencing to distinguish between the genes expressed in human tumor tissues from those in mouse stroma, the expression profiles of the tumors were clearly divided into a Y42C/Y42S group and a mock/WT group. Through gene set enrichment analysis, it was revealed that inflammation‑ and hypoxia‑related pathways were enriched in the mock/WT tumors; however, cell metabolism‑ and cell cycle‑related pathways such as Myc, E2F, oxidative phosphorylation and G2M checkpoint were enriched in the Y42C/Y42S tumors. In addition, the gene set related to ROCK signaling inhibition was enriched in the RHOA‑mutated group, which indicated that a series of events are related to ROCK inhibition induced by RHOA mutations. Histopathological analysis revealed that small tumor nests were more frequent in RHOA mutants than in the mock or WT group. In addition, increased blood vessel formation and infiltration of macrophages within the tumor mass were observed in the RHOA mutants. Furthermore, unlike mock/WT, the RHOA‑mutated tumor cells had little antitumor host reaction in the invasive front, which is similar to the pattern of mucosal invasion in clinical RHOA‑mutated DGC. These transcriptome and pathological analyses revealed that mutated RHOA functionally contributes to the acquisition of DGC features, which will accelerate our understanding of the contribution of RHOA mutations in DGC biology and the development of further therapeutic strategies.
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http://dx.doi.org/10.3892/or.2019.7300DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6775816PMC
November 2019

Machine learning approaches for pathologic diagnosis.

Virchows Arch 2019 Aug 20;475(2):131-138. Epub 2019 Jun 20.

Department of Preventive Medicine, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan.

Machine learning techniques, especially deep learning techniques such as convolutional neural networks, have been successfully applied to general image recognitions since their overwhelming performance at the 2012 ImageNet Large Scale Visual Recognition Challenge. Recently, such techniques have also been applied to various medical, including histopathological, images to assist the process of medical diagnosis. In some cases, deep learning-based algorithms have already outperformed experienced pathologists for recognition of histopathological images. However, pathological images differ from general images in some aspects, and thus, machine learning of histopathological images requires specialized learning methods. Moreover, many pathologists are skeptical about the ability of deep learning technology to accurately recognize histopathological images because what the learned neural network recognizes is often indecipherable to humans. In this review, we first introduce various applications incorporating machine learning developed to assist the process of pathologic diagnosis, and then describe machine learning problems related to histopathological image analysis, and review potential ways to solve these problems.
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http://dx.doi.org/10.1007/s00428-019-02594-wDOI Listing
August 2019

Capturing the differences between humoral immunity in the normal and tumor environments from repertoire-seq of B-cell receptors using supervised machine learning.

BMC Bioinformatics 2019 May 28;20(1):267. Epub 2019 May 28.

Department of Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, 113-0033, Japan.

Background: The recent success of immunotherapy in treating tumors has attracted increasing interest in research related to the adaptive immune system in the tumor microenvironment. Recent advances in next-generation sequencing technology enabled the sequencing of whole T-cell receptors (TCRs) and B-cell receptors (BCRs)/immunoglobulins (Igs) in the tumor microenvironment. Since BCRs/Igs in tumor tissues have high affinities for tumor-specific antigens, the patterns of their amino acid sequences and other sequence-independent features such as the number of somatic hypermutations (SHMs) may differ between the normal and tumor microenvironments. However, given the high diversity of BCRs/Igs and the rarity of recurrent sequences among individuals, it is far more difficult to capture such differences in BCR/Ig sequences than in TCR sequences. The aim of this study was to explore the possibility of discriminating BCRs/Igs in tumor and in normal tissues, by capturing these differences using supervised machine learning methods applied to RNA sequences of BCRs/Igs.

Results: RNA sequences of BCRs/Igs were obtained from matched normal and tumor specimens from 90 gastric cancer patients. BCR/Ig-features obtained in Rep-Seq were used to classify individual BCR/Ig sequences into normal or tumor classes. Different machine learning models using various features were constructed as well as gradient boosting machine (GBM) classifier combining these models. The results demonstrated that BCR/Ig sequences between normal and tumor microenvironments exhibit their differences. Next, by using a GBM trained to classify individual BCR/Ig sequences, we tried to classify sets of BCR/Ig sequences into normal or tumor classes. As a result, an area under the curve (AUC) value of 0.826 was achieved, suggesting that BCR/Ig repertoires have distinct sequence-level features in normal and tumor tissues.

Conclusions: To the best of our knowledge, this is the first study to show that BCR/Ig sequences derived from tumor and normal tissues have globally distinct patterns, and that these tissues can be effectively differentiated using BCR/Ig repertoires.
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http://dx.doi.org/10.1186/s12859-019-2853-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6537402PMC
May 2019

Artificial Smooth Muscle Model Composed of Hierarchically Ordered Microtubule Asters Mediated by DNA Origami Nanostructures.

Nano Lett 2019 06 3;19(6):3933-3938. Epub 2019 May 3.

Department of Chemistry and Materials Engineering , Kansai University , Osaka 564-8680 , Japan.

DNA has been well-known for its applications in programmable self-assembly of materials. Nonetheless, utility of DNA origami, which offers more opportunity to realize complicated operations, has been very limited. Here we report self-assembly of a biomolecular motor system, microtubule-kinesin mediated by DNA origami nanostructures. We demonstrate that a rodlike DNA origami motif facilitates self-assembly of microtubules into asters. A smooth-muscle like molecular contraction system has also been realized using the DNA origami in which self-assembled microtubules exhibited fast and dynamic contraction in the presence of kinesins through an energy dissipative process. This work provides potential nanotechnological applications of DNA and biomolecular motor proteins.
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http://dx.doi.org/10.1021/acs.nanolett.9b01201DOI Listing
June 2019

Keratinocyte Proline-Rich Protein Deficiency in Atopic Dermatitis Leads to Barrier Disruption.

J Invest Dermatol 2019 09 21;139(9):1867-1875.e7. Epub 2019 Mar 21.

Department of Dermatology, the University of Tokyo Graduate School of Medicine, Tokyo, Japan.

Atopic dermatitis is a common inflammatory skin disease caused by the interaction of genetic and environmental factors. By allelic copy number analysis at missense single-nucleotide polymorphisms on 26 genes with copy number variation, we identified a significant association between atopic dermatitis and human KPRP. Human KPRP expression, which was localized to the upper granular layer of epidermis, was significantly decreased in atopic dermatitis compared with normal skin. KPRP was histologically colocalized with loricrin and was mainly detected in cytoskeleton fractions of human keratinocytes. To further investigate the role of KPRP in skin, Kprp-knockout mice were generated. Heterozygous knockout (Kprp) mice exhibited reduced KPRP expression to level a similar to that of human AD lesional skin. Kprp mice showed abnormal desmosome structure and detachment of lower layers of the stratum corneum. Percutaneous inflammation by topical application of croton oil or oxazolone was enhanced, and epicutaneous immunization with ovalbumin induced a high level of IgE in Kprp mice. Our study, started from allelic copy number analysis in human AD, identified the importance of KPRP, the decrease of which leads to barrier dysfunction.
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http://dx.doi.org/10.1016/j.jid.2019.02.030DOI Listing
September 2019

Molecular Taxonomy of Interstitial Cystitis/Bladder Pain Syndrome Based on Whole Transcriptome Profiling by Next-Generation RNA Sequencing of Bladder Mucosal Biopsies.

J Urol 2019 08 8;202(2):290-300. Epub 2019 Jul 8.

Japanese Red Cross Medical Center , Tokyo.

Purpose: We systematically characterized gene expression, inflammation and neovascularization in patients with interstitial cystitis/bladder pain syndrome to obtain biological evidence supporting diagnosis and classification.

Materials And Methods: We sequenced RNA obtained from bladder mucosal biopsies of 33 patients with 3 subtypes of interstitial cystitis/bladder pain syndrome, including Hunner lesions in 12, no Hunner lesions in 11 but with glomerulations and neither Hunner lesions nor glomerulations in 10, and 9 controls. Differentially expressed genes of each subtype were searched to identify subtype specific biological pathways and candidate genes important for pathogenesis. Candidate genes were validated by quantitative polymerase chain reaction and immunohistochemistry. Digital immunohistochemical quantification was performed to assess subepithelial lymphoplasmacytic cell and microvessel density. Relationships between candidate gene over expression and symptom severity were explored.

Results: Patients with Hunner lesions showed a distinct gene expression profile associated with significant up-regulation of biological processes involving immune responses and infection, and an increase in subepithelial lymphoplasmacytic cell and microvessel density. Over expression of 2 candidate genes, VEGF and BAFF, correlated with symptom severity. Meanwhile, the gene expression profiles of patients with the 2 subtypes without Hunner lesions were similar to those of controls. No difference in biological pathways or subepithelial lymphoplasmacytic cell and microvessel density were detected between these 2 subtypes and controls.

Conclusions: Interstitial cystitis/bladder pain syndrome with Hunner lesions shows distinct genomic and histological features associated with immune responses and infection. In addition, VEGF and BAFF are potential disease biomarkers and therapeutic targets. This subtype should be considered separate from the syndrome.
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http://dx.doi.org/10.1097/JU.0000000000000234DOI Listing
August 2019

Functional genomics screen with pooled shRNA library and gene expression profiling with extracts of identify potential pathways for therapeutic targets in head and neck squamous cell carcinoma.

PeerJ 2019 1;7:e6464. Epub 2019 Mar 1.

Ganit Labs, Bio-IT Centre, Institute of Bioinformatics and Applied Biotechnology, Bangalore, India.

Tumor suppression by the extracts of (neem) works via anti-proliferation, cell cycle arrest, and apoptosis, demonstrated previously using cancer cell lines and live animal models. However, very little is known about the molecular targets and pathways that neem extracts and their associated compounds act through. Here, we address this using a genome-wide functional pooled shRNA screen on head and neck squamous cell carcinoma cell lines treated with crude neem leaf extracts, known for their anti-tumorigenic activity. We analyzed differences in global clonal sizes of the shRNA-infected cells cultured under no treatment and treatment with neem leaf extract conditions, assayed using next-generation sequencing. We found 225 genes affected the cancer cell growth in the shRNA-infected cells treated with neem extract. Pathway enrichment analyses of whole-genome gene expression data from cells temporally treated with neem extract revealed important roles played by the TGF-β pathway and -related gene network. Our results indicate that neem extract affects various important molecular signaling pathways in head and neck cancer cells, some of which may be therapeutic targets for this devastating tumor.
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http://dx.doi.org/10.7717/peerj.6464DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6398373PMC
March 2019

Interleukin-13 receptor α2 is a novel marker and potential therapeutic target for human melanoma.

Sci Rep 2019 02 4;9(1):1281. Epub 2019 Feb 4.

Laboratory of Oncology, School of Life Sciences, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan.

Malignant melanoma is one of the untreatable cancers in which conventional therapeutic strategies, including chemotherapy, are hardly effective. Therefore, identification of novel therapeutic targets involved in melanoma progression is urgently needed for developing effective therapeutic methods. Overexpression of interleukin-13 receptor α2 (IL13Rα2) is observed in several cancer types including glioma and pancreatic cancer. Although IL13Rα2 is implicated in the progression of various types of cancer, its expression and roles in the malignant melanoma have not yet been elucidated. In the present study, we showed that IL13Rα2 was expressed in approximately 7.5% melanoma patients. While IL13Rα2 expression in human melanoma cells decreased their proliferation in vitro, it promoted in vivo tumour growth and angiogenesis in melanoma xenograft mouse model. We also found that the expression of amphiregulin, a member of the epidermal growth factor (EGF) family, was correlated with IL13Rα2 expression in cultured melanoma cells, xenograft tumour tissues and melanoma clinical samples. Furthermore, expression of amphiregulin promoted tumour growth, implicating causal relationship between the expression of IL13Rα2 and amphiregulin. These results suggest that IL13Rα2 enhances tumorigenicity by inducing angiogenesis in malignant melanoma, and serves as a potential therapeutic target of malignant melanoma.
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http://dx.doi.org/10.1038/s41598-019-39018-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6362032PMC
February 2019

Viral loads correlate with upregulation of PD-L1 and worse patient prognosis in Epstein-Barr Virus-associated gastric carcinoma.

PLoS One 2019 29;14(1):e0211358. Epub 2019 Jan 29.

Department of Pathology, Graduate School of Medicine, the University of Tokyo, Tokyo, Japan.

Epstein-Barr virus (EBV)-associated gastric carcinoma (EBVaGC), one of four major gastric cancer types, consists of clonal growth of EBV-infected epithelial cells. However, the significance of viral loads in each tumor cell has not been evaluated. EBV-DNA is stably maintained in episomal form in the nucleus of each cancer cell. To estimate EBV copy number per genome (EBV-CN), qPCR of viral EBNA1 and host GAPDH, standardized by Namalwa DNA (one copy/genome), was applied to the formalin-fixed paraffin embedded (FFPE) surgically resected EBVaGC specimens (n = 43) and EBVaGC cell lines (SNU-719 and NCC-24). In surgical specimens, the cancer cell ratio (CCR) was determined with image analysis, and EBV-CN was obtained by adjusting qPCR value with CCR. Fluorescent in situ hybridization (FISH) was also applied to the FFPE sections using the whole EBV-genome as a probe. In surgical specimens, EBV-CN obtained by qPCR/CCR was between 1.2 and 185 copies with a median of 9.9. EBV-CN of SNU-719 and NCC-24 was 42.0 and 1.1, respectively. A linear correlation was observed with qPCR/CCR data up to 20 copies/genome (40 signals/nucleus), the limit of FISH analysis. In addition, substantial variation in the number of EBV foci was observed. Based on qPCR/CCR, high EBV-CN (>10 copies) correlated with PD-L1 expression in cancer cells (P = 0.015), but not with other pathological indicators. Furthermore, EBVaGC with high EBV-CN showed worse disease-specific survival (P = 0.041). Our findings suggest that cancer cell viral loads may contribute to expression of the immune checkpoint molecule and promotion of cancer progression in EBVaGC.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0211358PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6350976PMC
October 2019

Difference in morphology and interactome profiles between orthotopic and subcutaneous gastric cancer xenograft models.

J Toxicol Pathol 2018 Oct 13;31(4):293-300. Epub 2018 Aug 13.

Forerunner Pharma Research Co., Ltd., Komaba Open Laboratory, The University of Tokyo, 4-6-1 Komaba, Meguro-ku, Tokyo 153-8904, Japan.

In xenograft models, orthotopic (ORT) engraftment is thought to provide a different tumor microenvironment compared with subcutaneous (SC) engraftment. We attempted to characterize the biological difference between OE19 (adenocarcinoma of the gastroesophageal junction) SC and ORT models by pathological analysis and CASTIN (CAncer-STromal INteractome) analysis, which is a novel method developed to analyze the tumor-stroma interactome framework. In SC models, SCID mice were inoculated subcutaneously with OE19 cells, and tumor tissues were sampled at 3 weeks. In ORT models, SCID mice were inoculated under the serosal membrane of the stomach wall, and tumor tissues were sampled at 3 and 6 weeks after engraftment. Results from the two models were then compared. Histopathologically, the SC tumors were well circumscribed from the adjacent tissue, with scant stroma and the formation of large ductal structures. In contrast, the ORT tumors were less circumscribed, with small ductal structures invading into abundant stroma. Then we compared the transcriptome profiles of human tumor cells with the mouse stromal cells of each model by species-specific RNA sequencing. With CASTIN analysis, we successfully identified several interactions that are known to affect the tumor microenvironment as being selectively enhanced in the ORT model. In conclusion, pathological analysis and CASTIN analysis revealed that ORT models of OE19 cells have a more invasive character and enhanced interaction with stromal cells compared with SC models.
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http://dx.doi.org/10.1293/tox.2018-0020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6206286PMC
October 2018

EVI1 expression is associated with aggressive behavior in intrahepatic cholangiocarcinoma.

Virchows Arch 2019 Jan 23;474(1):39-46. Epub 2018 Oct 23.

Department of Pathology, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-0033, Japan.

Ecotropic virus integration site 1 protein homolog (EVI1), a well-known oncogenic transcriptional factor of hematopoietic cells, contributes to pancreatic cancer oncogenicity through increased expression of KRAS. Because EVI1 was upregulated in cholangiocarcinoma by referring The Cancer Genome Atlas, we investigated the importance of EVI1 in intrahepatic cholangiocarcinoma (ICC) which has been regarded as a heterogeneous group of cancers. Immunohistochemical analysis results demonstrated that EVI1 was overexpressed in about half of ICC (53/101, 52.5%). Moreover, all intraductal papillary neoplasms of the bile duct cases expressed EVI1 regardless of histological grading and subtypes such as gastric, intestinal, pancreatobiliary, or oncocytic (20/20, 100%). EVI1-positive ICC showed higher frequencies of aggressive pathological indicators such as periductal infiltrative growth (p = 0.022), hilar invasion (p = 0.041), advanced UICC stage (p = 0.026), major vascular invasion (p = 0.026), and perineural invasion (p = 0.007) than EVI1-negative ICC. Patients with EVI1-positive ICC showed worse overall survival and recurrence-free survival in all resected cases and in curative resected cases. Recently, we proposed type 1/2 (large/small duct types) classification of ICC based on mucin productivity and immunophenotypes (S100P, N-cadherin, and NCAM). Type 1 predominantly consisted of EVI1-positive ICC (33/42 cases, 79%), and the frequency was significantly higher than type 2 (18/55 cases, 32.7%) (p < 0.0001). EVI1-positive ICC was likely to express stomach-specific claudin CLDN18 (correlation coefficient r = 0.55373) and mucin MUC5AC (r = 0.42718). EVI1-positive ICC is an aggressive ICC showing both large-duct and/or gastric phenotypes. Consequently, a transcriptional factor EVI1 is associated with aggressive behavior in ICC and can be a therapeutic target molecule, while EVI1 might be a key molecule for the development of intraductal papillary neoplasms of the bile duct.
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http://dx.doi.org/10.1007/s00428-018-2476-0DOI Listing
January 2019

Machine Learning Methods for Histopathological Image Analysis.

Comput Struct Biotechnol J 2018 9;16:34-42. Epub 2018 Feb 9.

Department of Genomic Pathology, Medical Research Institute, Tokyo Medical and Dental University, Tokyo, Japan.

Abundant accumulation of digital histopathological images has led to the increased demand for their analysis, such as computer-aided diagnosis using machine learning techniques. However, digital pathological images and related tasks have some issues to be considered. In this mini-review, we introduce the application of digital pathological image analysis using machine learning algorithms, address some problems specific to such analysis, and propose possible solutions.
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http://dx.doi.org/10.1016/j.csbj.2018.01.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6158771PMC
February 2018

Generation of an anti-desmoglein 3 antibody without pathogenic activity of pemphigus vulgaris for therapeutic application to squamous cell carcinoma.

J Biochem 2018 Dec;164(6):471-481

Forerunner Pharma Research Co., Ltd., Komaba Open Laboratory, The University of Tokyo, 4-6-1 Komaba, Meguro-ku, Tokyo, Japan.

It is ideal for the target antigen of a cytotoxic therapeutic antibody against cancer to be cancer-specific, but such antigens are rare. Thus an alternative strategy for target selection is necessary. Desmoglein 3 (DSG3) is highly expressed in lung squamous cell carcinoma, while it is well-known that anti-DSG3 antibodies cause pemphigus vulgaris, an autoimmune disease. We evaluated DSG3 as a novel target by selecting an epitope that exerts efficacy against cancer with no pathogenic effects in normal tissues. Pathogenic anti-DSG3 antibodies induce skin blisters by inhibiting the cell-cell interaction in a Ca2+-dependent manner. We screened anti-DSG3 antibodies that bind DGS3 independent of Ca2+ and have high antibody-dependent cell cytotoxicity (ADCC) activity against DSG3-expressing cells. These selected antibodies did not inhibit cell-cell interaction and showed ADCC activity against squamous cell carcinoma cell lines. Furthermore, one of the DSG3 antibodies showed anti-tumour activity in tumour mouse models but did not induce adverse effects such as blister formation in the skin. Thus it was possible to generate an antibody against DSG3 by using an appropriate epitope that retained efficacy with no pathogenicity. This approach of epitope selection may expand the variety of druggable target molecules.
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http://dx.doi.org/10.1093/jb/mvy074DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6267343PMC
December 2018

HER2 Heterogeneity Is Associated with Poor Survival in HER2-Positive Breast Cancer.

Int J Mol Sci 2018 Jul 24;19(8). Epub 2018 Jul 24.

Department of Breast Surgery and Oncology, Tokyo Medical University, 6-7-1 Nishishinjuku, Shinjuku-ku, Tokyo 160-0023, Japan.

Intratumoral human epidermal growth factor receptor 2 (HER2) heterogeneity has been reported in 16⁻36% of HER2-positive breast cancer and its clinical impact is under discussion. We examined the biological effects of HER2-heterogeneity on mouse models and analyzed metastatic brains by RNA sequence analysis. A metastatic mouse model was developed using 231-Luc (triple negative cells) and 2 HER2-positive cell lines, namely, HER2-60 and HER2-90 which showed heterogeneous and monotonous HER2 expressions, respectively. Metastatic lesions developed in 3 weeks in all the mice injected with HER2-60 cells, and in 69% of the mice injected with HER2-90 and 87.5% of the mice injected with 231-Luc. The median survival days of mice injected with 231-Luc, HER2-60, and HER2-90 cells were 29 ( = 24), 24 ( = 22) and 30 ( = 13) days, respectively. RNA sequence analysis showed that and its related genes were significantly downregulated in metastatic brain tumors with HER2-60 cells. The low expression of caspase-1 could be a new prognostic biomarker for early relapse in HER2-positive breast cancer.
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http://dx.doi.org/10.3390/ijms19082158DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6121890PMC
July 2018

Frequent fusion in highly metastatic diffuse-type gastric cancer with relatively early onset.

Oncotarget 2018 Jun 29;9(50):29336-29350. Epub 2018 Jun 29.

Department of Pathology, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.

CLDN18-ARHGAP26/6 fusions have been identified in gastric cancers, with a predominance in diffuse-type gastric cancers (DGCs). Although experiments have suggested an oncogenic role for fusions, the exact frequencies and clinicopathological characteristics of the fusion-positive cases are poorly understood. We analyzed 254 cases of gastric cancer (172 diffuse-type and 82 intestinal-type) using RT-PCR and FISH, and also analyzed TCGA transcriptome datasets to identify genes that are related to the aggressive behaviors of fusion-positive cancers. Our assays identified 26 fusion-positive cases, 22 of which were DGCs (22/172, 12.8%). Unlike fusion-negative DGCs, almost all fusion-positive DGCs retained E-cadherin expression (P = 0.036). Fusion-positive DGCs also showed a higher prevalence of lymphatic and distant organ metastases, and these trends were only significant in the younger age group (< 60 years). In this group, the majority of cases with distant organ metastases (4 of 6 cases) were fusion-positive, and the multivariate regression analysis revealed that fusion status was an independent predictive marker for distant organ metastases (P = 0.002). In the TCGA dataset analysis, carbonic anhydrase 9 was postulated to be a potential modulator of the age-specific effects of the fusion protein, compatible with the immunohistochemical analysis of our cohort. Therefore, CLDN18-ARHGAP26/6 fusion-positive DGCs are considered biologically distinct entities that will require more advanced therapeutic options.
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http://dx.doi.org/10.18632/oncotarget.25464DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6047683PMC
June 2018

DGC-specific mutations maintained cancer cell survival and promoted cell migration via ROCK inactivation.

Oncotarget 2018 May 1;9(33):23198-23207. Epub 2018 May 1.

Genome Science Division, Research Center for Advanced Science and Technology, The University of Tokyo, Tokyo, Japan.

missense mutations exist specifically in diffuse type gastric cancers (DGC) and are considered one of the DGC driver genes, but it is not fully understood how mutations contribute to DGC development. Here we examined how mutations affect cancer cell survival and cell motility. We revealed that cell survival was maintained by specific mutation sites, namely G17, Y42, and L57. Because these functional mutations suppressed MLC2 phosphorylation and actin stress fiber formation, we realized they act in a dominant-negative fashion against the ROCK pathway. Through the same inactivating mechanism that maintained cell survival, mutations also increased cell migration activity. Cell survival and migration studies on () fusions, which are known to be mutually exclusive to mutations, showed that fusions complemented cell survival under knockdown condition and also induced cell migration. Site-directed mutagenesis analysis revealed the importance of the GAP domain and indicated that fusions maintained RHOA in the inactive form. Taken together, these findings show that the inactivation of ROCK would be a key step in DGC development, so ROCK activation might provide novel therapeutic opportunities.
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http://dx.doi.org/10.18632/oncotarget.25269DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5955407PMC
May 2018

A benchmark for comparing precision medicine methods in thyroid cancer diagnosis using tissue microarrays.

Bioinformatics 2018 05;34(10):1767-1773

Department of Surgery, Mackay Memorial Hospital, Taipei, Taiwan.

Motivation: The aim of precision medicine is to harness new knowledge and technology to optimize the timing and targeting of interventions for maximal therapeutic benefit. This study explores the possibility of building AI models without precise pixel-level annotation in prediction of the tumor size, extrathyroidal extension, lymph node metastasis, cancer stage and BRAF mutation in thyroid cancer diagnosis, providing the patients' background information, histopathological and immunohistochemical tissue images.

Results: A novel framework for objective evaluation of automatic patient diagnosis algorithms has been established under the auspices of the IEEE International Symposium on Biomedical Imaging 2017- A Grand Challenge for Tissue Microarray Analysis in Thyroid Cancer Diagnosis. Here, we present the datasets, methods and results of the challenge and lay down the principles for future uses of this benchmark. The main contributions of the challenge include the creation of the data repository of tissue microarrays; the creation of the clinical diagnosis classification data repository of thyroid cancer; and the definition of objective quantitative evaluation for comparison and ranking of the algorithms. With this benchmark, three automatic methods for predictions of the five clinical outcomes have been compared, and detailed quantitative evaluation results are presented in this paper. Based on the quantitative evaluation results, we believe automatic patient diagnosis is still a challenging and unsolved problem.

Availability And Implementation: The datasets and the evaluation software will be made available to the research community, further encouraging future developments in this field. (http://www-o.ntust.edu.tw/cvmi/ISBI2017/).

Contact: [email protected]

Supplementary Information: Supplementary data are available at Bioinformatics online.
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http://dx.doi.org/10.1093/bioinformatics/btx838DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5946957PMC
May 2018