論文 - 時野　隆至

多発性骨髄腫におけるDNA繰り返し配列の低メチル化と染色体異常の関連(The relationship between DNA hypomethylation of repetitive elements and chromosomal aberrations in multiple myeloma)

青木 由佳, 野島 正寛, 安井 寛, 丸山 玲緒, 鈴木 拓, 石田 禎夫, 麻奥 英毅, 時野 隆至, 豊田 実, 篠村 恭久

日本癌学会総会記事 ( (一社)日本癌学会 )  70回   137 - 137  2011年09月

多発性骨髄腫におけるDNAメチロームと染色体異常 メチル化DNA結合蛋白を利用した次世代シーケンシングによる検討(DNA methylome and chromosomal aberrations in multiple myeloma: an integrated analysis based on MBD-sequencing)

野島 正寛, 青木 由佳, 安井 寛, 丸山 玲緒, 鈴木 拓, 石田 禎夫, 時野 隆至, 篠村 恭久, 豊田 実

日本癌学会総会記事 ( (一社)日本癌学会 )  70回   47 - 47  2011年09月

Cysteine-rich intestinal protein 2 (CRIP2) acts as a repressor of NF-kappaB-mediated proangiogenic cytokine transcription to suppress tumorigenesis and angiogenesis.

Cheung AK, Ko JM, Lung HL, Chan KW, Stanbridge EJ, Zabarovsky E, Tokino T, Kashima L, Suzuki T, Kwong DL, Chua D, Tsao SW, Lung ML

Proceedings of the National Academy of Sciences of the United States of America   108 ( 20 ) 8390 - 5  2011年05月  [査読有り]  [国際誌]

　概要を見る

Chromosome 14 was transferred into tumorigenic nasopharyngeal carcinoma and esophageal carcinoma cell lines by a microcell-mediated chromosome transfer approach. Functional complementation of defects present in the cancer cells suppressed tumor formation. A candidate tumor-suppressor gene, cysteine-rich intestinal protein 2 (CRIP2), located in the hot spot for chromosomal loss at 14q32.3, was identified as an important candidate gene capable of functionally suppressing tumor formation. Previous studies have shown that CRIP2 is associated with development. To date, no report has provided functional evidence supporting a role for CRIP2 in tumor development. The present study provides unequivocal evidence that CRIP2 can functionally suppress tumorigenesis. CRIP2 is significantly down-regulated in nasopharyngeal carcinoma cell lines and tumors. CRIP2 reexpression functionally suppresses in vivo tumorigenesis and angiogenesis; these effects are induced by its transcription-repressor capability. It interacts with the NF-κB/p65 to inhibit its DNA-binding ability to the promoter regions of the major proangiogenesis cytokines critical for tumor progression, including IL6, IL8, and VEGF. In conclusion, we provide compelling evidence that CRIP2 acts as a transcription repressor of the NF-κB-mediated proangiogenic cytokine expression and thus functionally inhibits tumor formation and angiogenesis.

DOI PubMed

Epigenetic alteration of DNA in mucosal wash fluid predicts invasiveness of colorectal tumors.

Kamimae S, Yamamoto E, Yamano HO, Nojima M, Suzuki H, Ashida M, Hatahira T, Sato A, Kimura T, Yoshikawa K, Harada T, Hayashi S, Takamaru H, Maruyama R, Kai M, Nishiwaki M, Sugai T, Sasaki Y, Tokino T, Shinomura Y, Imai K, Toyota M

Cancer prevention research (Philadelphia, Pa.)   4 ( 5 ) 674 - 83  2011年05月  [査読有り]  [国際誌]

　概要を見る

Although conventional colonoscopy is considered the gold standard for detecting colorectal tumors, accurate staging is often difficult because advanced histology may be present in small colorectal lesions. We collected DNA present in mucosal wash fluid from patients undergoing colonoscopy and then assessed the methylation levels of four genes frequently methylated in colorectal cancers to detect invasive tumors. We found that methylation levels in wash fluid were significantly higher in patients with invasive than those with noninvasive tumors. Cytologic and K-ras mutation analyses suggested that mucosal wash fluid from invasive tumors contained greater numbers of tumor cells than wash fluid from noninvasive tumors. Among the four genes, levels of mir-34b/c methylation had the greatest correlation with the invasion and showed the largest area under the receiver operating characteristic curve (AUC = 0.796). Using cutoff points of mir-34b/c methylation determined by efficiency considerations, the sensitivity/specificity were 0.861/0.657 for the 13.0% (high sensitivity) and 0.765/0.833 for the 17.8% (well-balanced) cutoffs. In the validation test set, the AUC was also very high (0.915), the sensitivity/specificity were 0.870/0.875 for 13.0% and 0.565/0.958 for 17.8%. Using the diagnostic tree constructed by an objective algorithm, the diagnostic accuracy of the invasiveness of colorectal cancer was 91.3% for the training set and 85.1% for the test set. Our results suggest that analysis of the methylation of DNA in mucosal wash fluid may be a good molecular marker for predicting the invasiveness of colorectal tumors.

DOI PubMed

Apoptosis induction in cancer cells by the simultaneous transduction of p53 and artificial miRNAs based on the screening of genome-wide shRNA library

Masashi Idogawa, Yasushi Sasaki, Lisa Kashima, Yasuhisa Shinomura, Kohzoh Imai, Takashi Tokino

CANCER RESEARCH ( AMER ASSOC CANCER RESEARCH )  71  2011年04月  [査読有り]

DOI

The functional relationship between CHFR and PARP-1 controls the antephase checkpoint and tumor development

Lisa Kashima, Hiroaki Mita, Masashi Idogawa, Minoru Toyota, Yasushi Sasaki, Takashi Tokino

CANCER RESEARCH ( AMER ASSOC CANCER RESEARCH )  71  2011年04月  [査読有り]

DOI

Identification and characterization of early growth response 2, a zinc-finger transcription factor, as a p53-regulated proapoptotic gene.

Yokota I, Sasaki Y, Kashima L, Idogawa M, Tokino T

International journal of oncology   37 ( 6 ) 1407 - 16  2010年12月  [査読有り]  [国際誌]

　概要を見る

Tumor suppressor p53 is a transcription factor that induces growth arrest and/or apoptosis in response to cellular stress. In recent years, many genes have been identified as p53-regulated genes; however, no single target gene has been shown to be required for the apoptotic effect. Using microarray analysis, we have identified the transcription factor early growth response 2 (EGR2) as a target of the p53 family, specifically p53, p63 and p73. EGR2 expression was up-regulated by DNA damage-induced p53 activity, as well as by overexpression of p53 family genes. Furthermore, we identified a responsive element to p53, TAp63, and TAp73 within the EGR2 gene. This response element is highly conserved between human and rodents. We also found that overexpression of EGR2 induced apoptosis when combined with anticancer agents. Conversely, inactivation of EGR2 attenuated p53-mediated apoptosis. The results presented here suggest that EGR2 is a direct transcriptional target of p53 family that can in part mediate the p53-dependent apoptotic pathway.

DOI PubMed

Methylation-associated silencing of microRNA-34b/c in gastric cancer and its involvement in an epigenetic field defect.

Suzuki H, Yamamoto E, Nojima M, Kai M, Yamano HO, Yoshikawa K, Kimura T, Kudo T, Harada E, Sugai T, Takamaru H, Niinuma T, Maruyama R, Yamamoto H, Tokino T, Imai K, Toyota M, Shinomura Y

Carcinogenesis   31 ( 12 ) 2066 - 73  2010年12月  [査読有り]  [国際誌]

　概要を見る

Altered expression of microRNA (miRNA) is strongly implicated in cancer, and recent studies have shown that the silencing of some miRNAs is associated with CpG island hypermethylation. To identify epigenetically silenced miRNAs in gastric cancer (GC), we screened for miRNAs induced by treatment with 5-aza-2'-deoxycytidine and 4-phenylbutyrate. We found that miR-34b and miR-34c are epigenetically silenced in GC and that their downregulation is associated with hypermethylation of the neighboring CpG island. Methylation of the miR-34b/c CpG island was frequently observed in GC cell lines (13/13, 100%) but not in normal gastric mucosa from Helicobacter pylori-negative healthy individuals. Transfection of a precursor of miR-34b and miR-34c into GC cells induced growth suppression and dramatically changed the gene expression profile. Methylation of miR-34b/c was found in a majority of primary GC specimens (83/118, 70%). Notably, analysis of non-cancerous gastric mucosae from GC patients (n = 109) and healthy individuals (n = 85) revealed that methylation levels are higher in gastric mucosae from patients with multiple GC than in mucosae from patients with single GC (27.3 versus 20.8%; P < 0.001) or mucosae from H. pylori-positive healthy individuals (27.3 versus 20.7%; P < 0.001). These results suggest that miR-34b and miR-34c are novel tumor suppressors frequently silenced by DNA methylation in GC, that methylation of miR-34b/c is involved in an epigenetic field defect and that the methylation might be a predictive marker of GC risk.

DOI PubMed

p53 family network and human cancer

Sasaki Y, Idogawa M, Yokota I, Kashima L, Tokino T

Tumor Research   45   33 - 44  2010年12月  [査読有り]

CHFRとPARP-1の機能的関連が有糸分裂チェックポイントおよび腫瘍発生をコントロールする(The functional relationship between CHFR and PARP-1 controls the mitotic checkpoint and tumor development)

鹿島 理沙, 見田 裕章, 井戸川 雅史, 豊田 実, 佐々木 泰史, 時野 隆至

日本生化学会大会・日本分子生物学会年会合同大会講演要旨集 ( (公社)日本生化学会 )  83回・33回   3T4 - 4  2010年12月

アデノウイルスを用いたアポトーシス阻害的p53標的遺伝子に対する人工miRNAとp53の導入による癌細胞でのアポトーシス誘導(Apoptosis induction in cancer cells by the adenovirus-mediated transduction of p53 and artificial miRNAs targeting anti-apoptotic p53 target genes)

杉坂 淳, 井戸川 雅史, 佐々木 泰史, 鹿島 理沙, 篠村 恭久, 今井 浩三, 時野 隆至

日本生化学会大会・日本分子生物学会年会合同大会講演要旨集 ( (公社)日本生化学会 )  83回・33回   4P - 0967  2010年12月

p53ファミリーの新規標的ICAM2の同定 がん細胞の接着・遊走能への関与(Identification and characterization of the intercellular adhesion molecule-2 gene as a novel p53 family target)

平野 雄大, 佐々木 泰史, 横田 育子, 杉坂 淳, 鹿島 理紗, 井戸川 雅史, 時野 隆至

日本生化学会大会・日本分子生物学会年会合同大会講演要旨集 ( (公社)日本生化学会 )  83回・33回   3P - 0982  2010年12月

A novel correlation between LINE-1 hypomethylation and the malignancy of gastrointestinal stromal tumors.

Igarashi S, Suzuki H, Niinuma T, Shimizu H, Nojima M, Iwaki H, Nobuoka T, Nishida T, Miyazaki Y, Takamaru H, Yamamoto E, Yamamoto H, Tokino T, Hasegawa T, Hirata K, Imai K, Toyota M, Shinomura Y

Clinical cancer research : an official journal of the American Association for Cancer Research   16 ( 21 ) 5114 - 23  2010年11月  [査読有り]  [国際誌]

　概要を見る

PURPOSE: Gastrointestinal stromal tumors (GIST) are the most important mesenchymal tumors of the gastrointestinal tract. The vast majority of GISTs exhibit activating mutations of KIT or PDGFRA, but epigenetic alteration of GISTs is largely unknown. In this study, we aimed to clarify the involvement of DNA methylation in GIST malignancy. EXPERIMENTAL DESIGN: A total of 106 GIST specimens were studied. Levels of LINE-1 methylation were analyzed using bisulfite pyrosequencing. In addition, methylation of three other repetitive sequences (Alu Yb8, Satellite-α, and NBL2) was similarly analyzed, and CpG island hypermethylation was analyzed using MethyLight. Array-based comparative genomic hybridization (array CGH) was carried out in 25 GIST specimens. RESULTS: LINE-1 hypomethylation was significantly correlated with risk, and high-risk GISTs exhibited significantly lower levels of LINE-1 methylation than low-risk (61.3% versus 53.2%; P = 0.001) or intermediate-risk GISTs (60.8% versus 53.2%; P = 0.002). Hypomethylation of Satellite-α and NBL2 was also observed in high-risk GISTs. By contrast, promoter hypermethylation was relatively infrequent (CDH1, 11.2%; MLH1, 9.8%; SFRP1, 1.2%; SFRP2, 11.0%; CHFR, 9.8%; APC, 6.1%; CDKN2A, 0%; RASSF1A, 0%; RASSF2, 0%) and did not correlate with LINE-1 methylation or risk. Array CGH analysis revealed a significant correlation between LINE-1 hypomethylation and chromosomal aberrations. CONCLUSIONS: Our data suggest that LINE-1 hypomethylation correlates significantly with the aggressiveness of GISTs and that LINE-1 methylation could be a useful marker for risk assessment. Hypomethylation may increase the malignant potential of GISTs by inducing accumulation of chromosomal aberrations.

DOI PubMed

Genomic screening for genes upregulated by demethylation revealed novel targets of epigenetic silencing in breast cancer.

Fujikane T, Nishikawa N, Toyota M, Suzuki H, Nojima M, Maruyama R, Ashida M, Ohe-Toyota M, Kai M, Nishidate T, Sasaki Y, Ohmura T, Hirata K, Tokino T

Breast cancer research and treatment   122 ( 3 ) 699 - 710  2010年08月  [査読有り]  [国際誌]

　概要を見る

Breast cancer arises through the accumulation of multiple genetic alterations and epigenetic changes such as methylation, which silences gene expression in a variety of cancers. In the present study, we applied genomic screening to identify genes upregulated by the demethylating agent 5-aza-2'-deoxycytidine (DAC) in a human breast cancer cell line (MCF7). We identified 288 genes upregulated and 29 genes downregulated more than fivefold after treatment with DAC, and gene ontology analyses revealed the genes to be involved in immune responses, apoptosis, and cell differentiation. In addition, real-time PCR analysis of ten genes silenced in MCF7 cells confirmed that they are upregulated by DAC, while bisulfite-pyrosequencing analysis confirmed that nine of those genes were silenced by methylation. We also found that treating MCF7 cells with DAC restored induction of DFNA5 by p53, as well as by two other p53 family genes, p63gamma and p73beta. Introduction of NTN4 into MCF7 cells suppressed cell growth, indicating that NTN4 has tumor suppressive activity. In primary breast cancers, we detected cancer-specific methylation of NTN4, PGP9.5, and DKK3, suggesting that methylation of these genes could be useful markers for diagnosis of breast cancer. Thus, DNA methylation appears to be a common event in breast cancer, and the genes silenced by methylation could be useful targets for both diagnosis and therapy.

DOI PubMed

p53ファミリーの標的遺伝子EGR2の同定と機能解析(Identification and characterization of early growth response gene-2 as a p53-regulated proapoptotic gene)

横田 育子, 佐々木 泰史, 安保 直樹, 鹿島 理沙, 井戸川 雅史, 時野 隆至

日本癌学会総会記事 ( (一社)日本癌学会 )  69回   303 - 304  2010年08月

がん抑制分子CHFRおよびPARP-1相互作用による細胞周期制御(Functional association between CHFR and PARP-1 controls the mitotic checkpoint)

鹿島 理沙, 見田 裕章, 井戸川 雅史, 佐々木 泰史, 時野 隆至

日本癌学会総会記事 ( (一社)日本癌学会 )  69回   312 - 312  2010年08月

p53とアポトーシス阻害的標的に対するmiRNAの単一アデノウイルス発現による癌治療(Cancer Therapy by single adenovirus-mediated expression of p53 and microRNAs targeting anti-apoptotic p53 target genes)

井戸川 雅史, 佐々木 泰史, 鹿島 理沙, 篠村 恭久, 今井 浩三, 時野 隆至

日本癌学会総会記事 ( (一社)日本癌学会 )  69回   322 - 322  2010年08月

p53により発現抑制される分泌性増殖因子HDGFの同定(Negative regulation of hepatoma-derived growth factor by p53)

佐々木 泰史, 根岸 秀明, 横田 育子, 鹿島 理沙, 井戸川 雅史, 鈴木 拓, 豊田 実, 今井 浩三, 篠村 恭久, 時野 隆至

日本癌学会総会記事 ( (一社)日本癌学会 )  69回   303 - 303  2010年08月

次世代シークエンサーを用いた遺伝子転写開始点予測とエピゲノム解析への応用(Prediction of transcription start sites using next-generation sequencer and its application to epigenome analysis)

豊田 実, 鈴木 拓, 野島 正寛, 佐々木 泰史, 篠村 恭久, 時野 隆至, 今井 浩三

日本癌学会総会記事 ( (一社)日本癌学会 )  69回   417 - 417  2010年08月

DNA methylation of interferon regulatory factors in gastric cancer and noncancerous gastric mucosae.

Yamashita M, Toyota M, Suzuki H, Nojima M, Yamamoto E, Kamimae S, Watanabe Y, Kai M, Akashi H, Maruyama R, Sasaki Y, Yamano H, Sugai T, Shinomura Y, Imai K, Tokino T, Itoh F

Cancer science   101 ( 7 ) 1708 - 16  2010年07月  [査読有り]  [国際誌]

　概要を見る

Interferon regulatory factors (IRFs) are transcription factors known to play key roles in innate and adaptive immune responses, cell growth, apoptosis, and development. Their function in tumorigenesis of gastric cancer remains to be determined, however. In the present study, therefore, we examined epigenetic inactivation of IRF1-9 in a panel of gastric cancer cell lines. We found that expression of IRF4, IRF5, and IRF8 was frequently suppressed in gastric cancer cell lines; that methylation of the three genes correlated with their silencing; and that treating the cells with the demethylating agent 5-aza-2'-deoxycytidine (DAC) restored their expression. Expression of IRF5 in cancer cells was enhanced by the combination of DAC treatment and adenoviral vector-mediated expression of p53, p63, or p73. Interferon-gamma-induced expression of IRF8 was also enhanced by DAC. Moreover, treating gastric cancer cells with DAC enhanced the suppressive effects of interferon-alpha, interferon-beta, and interferon-gamma on cell growth. Among a cohort of 455 gastric cancer and noncancerous gastric tissue samples, methylation of IRF4 was frequently observed in both gastric cancer specimens and noncancerous specimens of gastric mucosa from patients with multiple gastric cancers, which suggests IRF4 methylation could be a useful molecular marker for diagnosing recurrence of gastric cancers. Our findings indicate that epigenetic IRF inactivation plays a key role in tumorigenesis of gastric cancer, and that inhibition of DNA methylation may restore the antitumor activity of interferons through up-regulation of IRFs.

DOI PubMed