Biomedical subjects

Johji Inazawa

Publications and source records attributed to Johji Inazawa.

71 records · Page 4Linked to original sources

PubMed2003

Association of 17q21-q24 gain in ovarian clear cell adenocarcinomas with poor prognosis and identification of PPM1D and APPBP2 as likely amplification targets.

PURPOSE: Although tumor stage is considered a prognosticfeature for ovarian clear cell adenocarcinomas (OCCAs), it is not likely to fully account for the clinical and biological variability characteristic of the disease. The aim of this study was to investigate aberrations of DNA copy number in OCCA tumors and identify genetic markers that would increase our understanding of the pathogenesis of OCCA and assist in more accurately predicting the outcome for an individual patient with this disease. EXPERIMENTAL DESIGN: We determined copy number aberrations among 20 primary OCCA tumors by means of comparative genomic hybridization and investigated their relationship to clinicopathological data. We also measured expression levels of candidate target genes within critical regions by quantitative real-time reverse transcription-PCRs and compared those data with copy number status and patient outcomes. RESULTS: We identified several nonrandom chromosomal aberrations among the 20 primary OCCA tumors examined. Among them, gain of DNA at 17q21-q24 showed significantly negative correlation with disease-free and overall survival (P = 0.0012 and 0.0039, respectively, log-rank test). This correlation held even for patients with stage I tumors. Among 15 candidate genes within the 17q21-q24 region, we found significantly elevated expression of PPM1D and APPBP2, and their heightened expression correlated negatively with disease-free survival (P = 0.0090, log-rank test adjusted for multiple comparisons). CONCLUSIONS: Information gained from our relatively large panel of OCCA tumors suggested that 17q21-q24 gain and consequent overexpression of two potential targets, PPM1D and APPBP2, are associated with malignant phenotypes of this tumor and may be useful predictors for prognosis.

Adenocarcinoma, Clear Cell↗

PubMed2002

Expression of cIAP1, a target for 11q22 amplification, correlates with resistance of cervical cancers to radiotherapy.

Inhibition of, or increased resistance to, apoptosis is a common property of cancer cells. This means that a constitutive activation of antiapoptotic molecules via genetic or epigenetic mechanisms, including gene amplification, may well be involved in carcinogenesis. Recently we reported that cIAP1, an inhibitor of apoptosis, is overexpressed through 11q22 amplification in cell lines derived from esophageal squamous cell carcinomas and is associated with resistance of esophageal squamous cell carcinomas to drug-induced apoptosis (I. Imoto et al. Cancer Res., 61: 6629-6634, 2001). Because amplification of 11q22 has been implicated in other malignancies also, including cervical squamous cell carcinomas (CSCCs), we attempted to correlate amplification and overexpression of cIAP1 with radiation sensitivity in CSCC-derived cell lines and primary CSCC tumors. In the nine cell lines we examined, two showed amplification and consistent overexpression of cIAP1, as well as significant resistance to radiation-induced cell death as compared with lines showing no cIAP1 amplification. Immunohistochemical analysis of 70 primary CSCCs from patients treated only with radiotherapy demonstrated that both overall survival and local recurrence-free survival was significantly poorer among patients with tumors showing high levels of nuclear cIAP1 staining than among patients whose tumors revealed little or no nuclear cIAP1. Multivariate analysis showed nuclear cIAP1 staining to be an independent predictive factor for local recurrence-free survival after radiotherapy among patients with CSCC. These findings demonstrate that cIAP1 may play an important role in the development/progression of this disease and that cIAP1 could be a novel predictive marker for resistance to radiotherapy in individual CSCC patients.

Adult↗

PubMed2002

Damaged epithelia regenerated by bone marrow-derived cells in the human gastrointestinal tract.

Studies have shown that bone marrow cells have the potential to differentiate into a variety of cell types. Here we show that bone marrow cells can repopulate the epithelia of the human gastrointestinal tract. Epithelial cells of male donor origin were distributed in every part of the gastrointestinal tract of female bone marrow transplant recipients. Donor-derived epithelial cells substantially repopulated the gastrointestinal tract during epithelial regeneration after graft-versus-host disease or ulcer formation. Regeneration of gastrointestinal epithelia with donor-derived cells in humans shows a potential clinical application of bone marrow-derived cells for repairing severely damaged epithelia, not only in the gastrointestinal tract but also in other tissues.

Adult↗

PubMed2002

cDNA cloning, genomic structure and chromosomal mapping of the mouse glucuronyltransferase-S involved in the biosynthesis of the HNK-1 carbohydrate epitope.

The HNK-1 carbohydrate epitope is expressed on a series of cell adhesion molecules and some glycolipids in the nervous system. Two glucuronyltransferases (GlcAT-P and GlcAT-S) are involved in the biosynthesis of the HNK-1 carbohydrate epitope. In this study, we isolated cDNA and genomic clones encoding the mouse glucuronyltransferase-S involved in the biosynthesis of the HNK-1 carbohydrate epitope and determined the structural organization of the gene. The deduced amino acid sequence of mouse GlcAT-S consists of 324 amino acids and has a type II membrane topology. The predicted amino acid sequence of mouse GlcAT-S is 98.1% identical to that of rat GlcAT-S. Northern blot analysis revealed that the mouse GlcAT-S transcript is specifically expressed in the nervous system. Moreover, the mouse GlcAT-S gene is composed of four exons spanning over more than 25 kilobase pairs. Southern blot analysis and chromosomal mapping indicated that the mouse GlcAT-S gene is a single copy gene and it was mapped to the A4-B region of mouse chromosome 1.

5' Flanking Region↗

PubMed2002

The Xq22 inversion breakpoint interrupted a novel Ras-like GTPase gene in a patient with Duchenne muscular dystrophy and profound mental retardation.

A male patient with profound mental retardation, athetosis, nystagmus, and severe congenital hypotonia (Duchenne muscular dystrophy [DMD]) was previously shown to carry a pericentric inversion of the X chromosome, 46,Y,inv(X)(p21.2q22.2). His mother carried this inversion on one X allele. The patient's condition was originally misdiagnosed as cerebral palsy, and only later was it diagnosed as DMD. Because the DMD gene is located at Xp21.2, which is one breakpoint of the inv(X), and because its defects are rarely associated with severe mental retardation, the other clinical features of this patient were deemed likely to be associated with the opposite breakpoint at Xq22. Our precise molecular-cytogenetic characterization of both breakpoints revealed three catastrophic genetic events that had probably influenced neuromuscular and cognitive development: deletion of part of the DMD gene at Xp21.2, duplication of the human proteolipid protein gene (PLP) at Xq22.2, and disruption of a novel gene. The latter sequence, showing a high degree of homology to the Sec4 gene of yeast, encoded a putative small guanine-protein, Ras-like GTPase that we have termed "RLGP." Immunocytochemistry located RLGP at mitochondria. We speculate that disruption of RLGP was responsible for the patient's profound mental retardation.

Adolescent↗

PubMed2002

An alternatively spliced isoform of transcriptional repressor ATF3 and its induction by stress stimuli.

Activating transcription factor 3 (ATF3) is a member of the ATF/CREB family of transcription factors and its expression is increased by various pathophysiological conditions and in several cancer cells. In this study, we describe two alternatively spliced ATF3DeltaZip mRNAs: ATF3DeltaZip2a and ATF3DeltaZip2b. Both variants encoded the same truncated protein of 135 amino acids, which lacked the leucine zipper domain and was incapable of binding to the ATF/CRE motif. The ATF3DeltaZip2 protein was shown to be localized in the nuclei and counteracted the transcriptional repression by the full-length ATF3. Western blot analysis showed that ATF3DeltaZip2 was expressed in cells exposed to A23187. Further study showed that, similar to the full-length ATF3, the expression of ATF3DeltaZip2 was induced by a wide range of stress stimuli. However, its expression was not detectable in cancer cells that constitutively over-expressed ATF3. Taken together, our results suggest that ATF3DeltaZip2, a protein derived from alternatively spliced mRNAs, is induced by various stress signals and may modulate the activity of the full-length ATF3 protein during stress response.

Activating Transcription Factor 3↗

PubMed2002

Isolation, characterization and mapping of the mouse and human RB1CC1 genes.

RB1CC1 (RB1-inducible Coiled-Coil 1), a putative transcription factor implicated in the regulation of RB1 (retinoblastoma 1) expression, was recently identified in a screen for genes involved in multi-drug resistance to anticancer agents. Information about the RB1CC1 gene is limited, however, and its biological function is not determined. Here we report the isolation, characterization and mapping of the mouse RB1CC1 gene (Rb1cc1), together with further characterization of the human RB1CC1 gene. Mouse Rb1cc1 encodes 1588 amino acids, sharing 89% identity and key sequence motifs with its human counterpart. Rb1cc1 is expressed abundantly in heart and testis, with lower levels detected in lung and spleen. Immunohistochemical analysis revealed the Rb1cc1 and Rb1 proteins are co-localized in the cell nuclei of NIH3T3-3 cell and various mouse tissues. The human and mouse RB1CC1 genes, both of which contain 24 exons, span 74 kb on chromosome 8q11.2 and 57 kb on chromosome 1A2-4, respectively. Conserved sequence motifs and nuclear localization suggest that the RB1CC1 proteins function as transcription factors.

3T3 Cells↗

PubMed2002

Causal relationship between the loss of RUNX3 expression and gastric cancer.

Runx3/Pebp2alphaC null mouse gastric mucosa exhibits hyperplasias due to stimulated proliferation and suppressed apoptosis in epithelial cells, and the cells are resistant to growth-inhibitory and apoptosis-inducing action of TGF-beta, indicating that Runx3 is a major growth regulator of gastric epithelial cells. Between 45% and 60% of human gastric cancer cells do not significantly express RUNX3 due to hemizygous deletion and hypermethylation of the RUNX3 promoter region. Tumorigenicity of human gastric cancer cell lines in nude mice was inversely related to their level of RUNX3 expression, and a mutation (R122C) occurring within the conserved Runt domain abolished the tumor-suppressive effect of RUNX3, suggesting that a lack of RUNX3 function is causally related to the genesis and progression of human gastric cancer.

Animals↗

PubMed2002

MYEOV, a gene at 11q13, is coamplified with CCND1, but epigenetically inactivated in a subset of esophageal squamous cell carcinomas.

DNA amplifications at 11q13 are frequently observed in esophageal squamous cell carcinoma (ESC) and correlate with a malignant phenotype. Although this amplicon spans a region of several megabases and harbors numerous genes, CCND1 and EMS1 are thought to be the relevant candidates in ESC. We investigated whether the putative transforming gene MYEOV, mapping 360 kb centromeric to CCND1 and activated concomitantly with CCND1 in a subset of t(11;14)(q13;q32) positive multiple myeloma cell lines, represents a target of 11q13 amplification in ESC. To evaluate the role of MYEOV in ESC, we tested 31 ESC cell lines and 48 primary tumors for copy number levels of MYEOVand demonstrated that MYEOV was always coamplified with CCND1. However, MYEOV expression levels correlated only inconsistently with DNA amplification data. Treatment with the demethylating agent 5-aza-2'-deoxycytidine restored MYEOV expression in a subset of cell lines exhibiting DNA amplification without high MYEOV expression, suggesting that MYEOV is transcriptionally silenced by a DNA methylation mechanism in most of the latter cell lines. Our results indicate that MYEOV is a coamplified gene with CCND1 at 11q13, but its activation is sometimes inhibited by an epigenetic mechanism.

Azacitidine↗

PubMed2002

A novel target gene, SKP2, within the 5p13 amplicon that is frequently detected in small cell lung cancers.

We investigated DNA copy-number aberrations in 22 cell lines derived from small cell lung cancers (SCLCs) using comparative genomic hybridization. A minimal common region at 5p13, within the 5p11-p13 amplicon that was most frequently involved, harbored the CDH6, PC4, and SKP2 genes. These three genes showed amplification and consequent overexpression in the SCLC cell lines. SKP2 positively regulates progression of cell cycle by targeting several regulators, such as the cell-cycle inhibitor p27(KIP1), for ubiquitin-mediated degradation. SKP2 was amplified in 7 (44%) of 16 primary SCLC tumors, and consequently overexpressed in 10 (83%) of the 12 of those tumors we examined. Expression levels of SKP2 protein were cell cycle-dependent in SCLC cells as well as in normal cells, and were correlated with the DNA copy-number of the gene. There was an inverse correlation between the expression of SKP2 and p27(KIP1) proteins. Down-regulation of SKP2 using an anti-sense oligonucleotide remarkably suppressed the growth of SCLC cells. Our results indicate that SKP2 is likely to be a target of the 5p13 amplification and to play an important role in the growth of SCLC cells.

Carcinoma, Small Cell↗

PubMed2002

Identification and mutational analysis of candidate genes for juvenile myoclonic epilepsy on 6p11-p12: LRRC1, GCLC, KIAA0057 and CLIC5.

Juvenile myoclonic epilepsy (JME) is one of the most frequent hereditary epilepsies characterized by myoclonic and tonic-clonic convulsions beginning at 8-20 years of age. Genetic studies have revealed four major chromosomal loci on 6p21.3, 6p11-12, 6q24, and 15q14 as candidate regions harboring genes responsible for JME. Previously we reported the region on 6p11-p12 (EJM1), and here we report the identification and mutational analysis of candidate genes for EJM1. One of those is a leucine-rich repeat-containing 1 (LRRC1) gene that is composed of 14 exons and codes for 524 amino acid residues. In Northern analysis, 7 kb transcripts of LRRC1 gene were detected in multiple tissues, most strongly, in heart, lung, and kidney. Mutation analysis of LRRC1 gene in 20 JME patients from ten families revealed one nucleotide substitution that lead to amino acid exchange (c.577 A>G; Ile193Val). This variation, however, did not co-segregate with the disease phenotype. We further performed mutational analyses of CLIC5, KIAA0057 and GCLC genes in or flank to the EJM1 region. These analyses did not provide any evidences that these genes are responsible for the JME phenotype, and suggested that these may not be the EJM1 gene.

Base Sequence↗

PubMed2002

TFDP1, CUL4A, and CDC16 identified as targets for amplification at 13q34 in hepatocellular carcinomas.

We carried out molecular cytogenetic characterization of 11 cell lines derived from hepatocellular carcinomas (HCCs) and 51 primary HCCs. Comparative genomic hybridization (CGH) revealed frequent amplification at 13q34, where we had detected amplification in several other types of tumor, including esophageal squamous cell carcinomas (ESC). Previously, we suggested possible involvement of TFDP1, encoding a transcription factor DP-1, in the 13q34 amplification observed in a primary ESC. Therefore, we investigated amplifications and expression levels of 5 genes mapped on the amplified region, including TFDP1, for exploring amplification targets at 13q34 in HCCs. 3 of those genes, TFDP1, CUL4A (cullin 4A), and CDC16 (cell division cycle 16), showed distinct amplification and consequent over-expression in some cell lines. Moreover, each was amplified in 3 or 4 of the 51 primary HCCs, and all 3 were amplified in 2 tumors, in which their expression patterns correlated with amplification patterns. To elucidate the functional role of TFDP1 in HCC, we examined expression levels of genes downstream of TFDP1 with real-time quantitative polymerase chain reaction (PCR). Expression of cyclin E gene (CCNE1) correlated closely with that of TFDP1 in not only cell lines, but also primary tumors. Treatment of HCC cells with the antisense oligonucleotide targeting TFDP1 resulted in down-regulation of CCNE1, suggesting that TFDP1 overexpression led to up-regulation of CCNE1 that encoded a positive regulator for cell cycle G1/S transition. In conclusion, our findings suggest that TFDP1, CUL4A, and CDC16 are probable targets of an amplification mechanism and therefore may be involved, together or separately, in development and/or progression of some HCCs.

Apc6 Subunit, Anaphase-Promoting Complex-Cyclosome↗

PubMed2002

Genomic organization and fine-mapping of the human leucine zipper-bearing kinase (LZK) gene.

Leucine zipper-bearing kinase (LZK) is a new member of the mixed lineage protein kinase family. We previously cloned a cDNA encoding LZK from a human cerebellum cDNA library. The following studies indicated that LZK serves as a MAPKKK in the JNK/SAPK pathway in cells, and a scaffold protein, JIP-1, enhances LZK-induced JNK/SAPK pathway activation via physical association. Here we report characterization of the gene structure and fine chromosomal mapping of the human LZK gene. Polymerase chain-reaction (PCR) studies indicated that the human LZK coding sequence is composed of 13 exons, and that all the splice acceptor and donor sequences obey the GT-AG rule. Chromosomal localization studies involving FISH mapping demonstrated that the human LZK gene is located at 3q27.

Amino Acid Sequence↗

PubMed2002

Novel targets for the 18p11.3 amplification frequently observed in esophageal squamous cell carcinomas.

Amplification of DNA in certain chromosomal regions, with consequent over-expression of specific genes within these amplicons, plays a crucial role in the development and progression of human cancer. Since our previous comparative genomic hybridization (CGH) study revealed frequent amplifications at 18p in esophageal squamous cell carcinomas (ESC) cell lines, we focused on the identification of genetic target(s) within the 18p amplicon. In four cell lines having remarkable copy-number amplification with homogeneously staining region (HSR) pattern by fluorescence in situ hybridization (FISH), the smallest common region of overlapping covered approximately 3.5 Mb at 18p11.3. We screened 29 ESC cell lines to discern amplifications and expression levels of 14 known genes and 21 uncharacterized transcripts within the amplicon. Only four known genes, YES1, TYMS, HEC and TGIF showed amplification and consequent over-expression. These genes were amplified in several of primary ESCs. Moreover, resistance to transforming growth factor beta (TGFbeta)-induced growth inhibition was enhanced in four cell lines with amplification and expression of TGIF, which encodes the repressor for TGFbeta-activated transcription, appears to be involved in the progression of ESC. Taken together, these results suggest that YES1, TYMS, HEC and TGIF are likely to be candidate targets for 18p11.3 amplification and be associated with esophageal tumorigenesis.

Carcinoma, Squamous Cell↗

PubMed2002

Molecular cloning and genomic analysis of mouse glucuronyltransferase involved in biosynthesis of the HNK-1 epitope.

cDNA and genomic clones encoding the mouse glucuronyltransferase (GlcAT-P) involved in biosynthesis of the HNK-1 carbohydrate epitope were isolated and the structural organization of the gene was determined. The predicted amino acid sequence of mouse GlcAT-P is 96.2 and 98.2% identical to those of the rat and human enzymes, respectively. Alternatively spliced isoforms of mouse GlcAT-P are present in the brain and encode two proteins that are identical throughout their length except for an additional 13 amino acids in the N-terminal cytoplasmic domain of the major form. The coding region of GlcAT-P is composed of 5 exons spanning approximately 6 kb, and the GlcAT-P gene was mapped to the A4 region of mouse chromosome 9. Upstream of the transcriptional start site, no typical TATA or CCAAT box was found, but binding sites for several known transcription factors including Sp1 and Krox-20 were identified. Transient transfection of luciferase reporter constructs demonstrated that a 207 bp fragment of the 5'-upstream region acts as a strong promoter in PC-12 cells, which express the HNK-1 epitope, but not in COS-1 cells. Thus, this minimal promoter region of GlcAT-P is suggested to be associated with the regulation of HNK-1 expression.

5' Flanking Region↗

PubMed2002

Differentially regulated genes as putative targets of amplifications at 20q in ovarian cancers.

Frequent amplification of DNA at 20q or part of 20q has been demonstrated by comparative genomic hybridization in ovarian cancer (OC), but the genetic target(s) of these amplification events remain unknown. We examined copy-number changes with respect to six candidate genes, E2F1 (20q11.2), TGIF2 (20q11.2), AIB1 (20q12), PTPN1 (20q13.1), ZNF217 (20q13.2), and BTAK (20q13), and then measured transcription levels of each candidate in 18 OC cell lines. Three distinct cores of amplification were identified: 20q11.2, harboring E2F1 and TGIF2 (region I; 1 of 18 cell lines, 5.6%); 20q13.1, harboring PTPN1 (region II; 5 lines, 27.8%); and 20q13.2, harboring ZNF217 and BTAK (region III; 6 lines, 33.3%). Among the six genes examined, expression levels of PTPN1 and ZNF217 were significantly correlated with absolute copy-number, and those of PTPN1 and TGIF2 were significantly correlated with copy-number relative to the centromere of chromosome 20 (20cen). Among 19 primary OCs examined, moreover, we observed amplification of TGIF2, PTPN1 and ZNF217 in five (26.3%), ten (52.6%), and twelve (63.2%) tumors, respectively. Expression levels of PTPN1 and ZNF217 were significantly correlated with their copy-numbers in those primary OCs. Our results suggest that 20q amplifications in OCs can be extensive and complex, probably due to synergistic or non-synergistic amplification of separate regions of 20q, involving multiple, independently amplified targets.

Chromosomes, Human, Pair 20↗

PubMedDate not supplied

Chromosome arm 20q gains and other genomic alterations in esophageal squamous cell carcinoma, as analyzed by comparative genomic hybridization and fluorescence in situ hybridization.

BACKGROUND/AIMS: Our recent analysis of gastric cancers and colorectal cancers using comparative genomic hybridization revealed a novel, high frequent copy number increases the long arm of chromosome 20 in association with possible involvement of liver metastases and poor prognosis. This led to further comparative genomic hybridization analysis of chromosomal aberrations in primary tumors of esophageal squamous cell carcinoma. The aim of the study presented here was to analyze the chromosomal aberrations and to determine the numbers of copies of AIB1, BTAK, DcR3 and E2F1 as putative target genes on chromosome 20q as well as their expression and relation to clinicopathological features in 41 primary tumors of esophageal squamous cell carcinoma. METHODOLOGY: We used comparative genomic hybridization to screen 41 primary tumors of esophageal squamous cell carcinoma for changes in the number of copies of DNA sequences. To further characterize the gain of DNA sequences at 20q, we also performed fluorescence in situ hybridization analysis. We examined the relationship between these changes and clinicopathological factors. RESULTS: Gains in chromosome arm 20q were detected (34.1%) as well as a high level of gain in 20q12-13 (4.8%). AIB1 amplification was observed in 4.9% (2/41), BTAK amplification in 9.8% (4/41), DcR3 amplification was in 4.9% (2/41), and E2F1 amplification in 7.3% (3/41). The survival of patients with BTAK or E2F1 amplification was significantly lower than that of patients without these abnormalities. CONCLUSIONS: These findings provide evidence for a number of previously unknown genomic aberrations in esophageal squamous cell carcinoma, suggesting the existence of target regions relevant to its progression. Esophageal squamous cell carcinoma with 20q gain showed extensive lung metastases, pleural effusion and liver metastases and poorer prognosis compared to cases without 20q gain. Our results suggest that amplification of BTAK or E2F1 are likely to lead to an increase in the number of malignant phenotypes of esophageal squamous cell carcinoma and that these aberrations can be expected to be useful as markers of poor prognosis.

Aged↗