Keika Zen

Defective expression of polarity protein PAR-3 gene ( PARD3 ) in esophageal squamous cell carcinoma

The partition-defective 3 (PAR-3) protein is implicated in the formation of tight junctions at epithelial cell–cell contacts. We investigated DNA copy number aberrations in human esophageal squamous cell carcinoma (ESCC) cell lines using a high-density oligonucleotide microarray and found a homozygous deletion of PARD3 (the gene encoding PAR-3). Exogenous expression of PARD3 in ESCC cells lacking this gene enhanced the recruitment of zonula occludens 1 (ZO-1), a marker of tight junctions, to sites of cell–cell contact. Conversely, knockdown of PARD3 in ESCC cells expressing this gene caused a disruption of ZO-1 localization at cell–cell borders. A copy number loss of PARD3 was observed in 15% of primary ESCC cells. Expression of PARD3 was significantly reduced in primary ESCC tumors compared with their nontumorous counterparts, and this reduced expression was associated with positive lymph node metastasis and poor differentiation. Our results suggest that deletion and reduced expression of PARD3 may be a novel mechanism that drives the progression of ESCC.

SOX2 identified as a target gene for the amplification at 3q26 that is frequently detected in esophageal squamous cell carcinoma

SOX2 is a transcription factor with a high-mobility group DNA-binding domain that functions as a master regulator during embryogenesis and organogenesis. We investigated DNA copy number aberrations in esophageal squamous cell carcinoma (ESCC) cell lines using a high-density oligonucleotide microarray and found frequent amplification at the chromosomal region 3q26. The estimated extent of the minimal overlapping region of amplification was 1.3 Mb. This chromosomal region includes a single gene, SOX2. The SOX2 protein was overexpressed in cell lines in which the gene was amplified. Knockdown experiments showed that SOX2 promotes proliferation of ESCC cells. Genes potentially modulated by SOX2 were determined by expression array analyses combined with small interfering RNA cell-transfection studies. A copy number gain of SOX2 (>2-fold) was observed in 6 of the 40 primary ESCCs (15%). Immunohistochemical study revealed that expression of the SOX2 protein was significantly elevated in 62 of the 89 ESCC tumors (70%), compared with their nontumorous counterparts, and that upregulated expression of SOX2 was associated with poor differentiation of ESCC. Our results suggest that SOX2 is likely to be a target of the 3q26 amplification and may therefore be involved in the development or progression of ESCC.

ERK5 is a target for gene amplification at 17p11 and promotes cell growth in hepatocellular carcinoma by regulating mitotic entry

Using high‐density oligonucleotide microarrays, we investigated DNA copy‐number aberrations in cell lines derived from hepatocellular carcinomas (HCCs) and detected a novel amplification at 17p11. To identify the target of amplification at 17p11, we defined the extent of the amplicon and examined HCC cell lines for expression of all seven genes in the 750‐kb commonly amplified region. Mitogen‐activated protein kinase (MAPK) 7, which encodes extracellular‐regulated protein kinase (ERK) 5, was overexpressed in cell lines in which the gene was amplified. An increase in MAPK7 copy number was detected in 35 of 66 primary HCC tumors. Downregulation of MAPK7 by small interfering RNA suppressed the growth of SNU449 cells, the HCC cell line with the greatest amplification and overexpression of MAPK7. ERK5, phosphorylated during the G2/M phases of the cell cycle, regulated entry into mitosis in SNU449 cells. In conclusion, our results suggest that MAPK7 is likely the target of 17p11 amplification and that the ERK5 protein product of MAPK7 promotes the growth of HCC cells by regulating mitotic entry.

Activation of B-Myb by E2F1 in hepatocellular carcinoma

Aim:  Deregulation of E2F1 transcriptional activity is observed in a variety of cancers, including hepatocellular carcinoma (HCC). The aim of the present study is to identify transcriptional target genes of E2F1 in HCC.

Alterations of the SWI/SNF chromatin remodelling subunit-BRG1 and BRM in hepatocellular carcinoma.

The SWI/SNF chromatin remodelling complex, which contains either brahma‐related gene‐1 (BRG1) or brahma (BRM) as the catalytic ATPase, functions as a master regulator of gene expression.

A novel amplification target, ARHGAP5, promotes cell spreading and migration by negatively regulating RhoA in Huh-7 hepatocellular carcinoma cells

RhoA, a member of the Rho family of small GTPases, directs the organization of the actin cytoskeleton and is involved in regulating cell shape and movement. Its activity is negatively regulated by p190-B RhoGAP (GTPase-activating protein). We investigated DNA copy number aberrations in human hepatocellular carcinoma and esophageal squamous cell carcinoma cell lines using a high-density oligonucleotide microarray and found a novel amplification at chromosomal region 14q12. We identified ARHGAP5 (the gene encoding p190-B RhoGAP) as a probable target for the amplification at 14q12, and our results showed that p190-B RhoGAP promotes cells spreading and migration by negatively regulating RhoA activity in Huh-7 hepatocellular carcinoma cells.

PEG10 is a probable target for the amplification at 7q21 detected in hepatocellular carcinoma

DNA copy number aberrations in human hepatocellular carcinoma (HCC) cell lines were investigated using a high-density oligonucleotide microarray, and a novel amplification at the chromosomal region 7q21 was detected. Molecular definition of the amplicon indicated that PEG10 (paternally expressed gene 10), a paternally expressed imprinted gene, was amplified together with CDK14 (cyclin-dependent kinase 14; previously PFTAIRE protein kinase 1, PFTK1) and CDK6 (cyclin-dependent kinase 6). An increase in PEG10 copy number was detected in 14 of 34 primary HCC tumors (41%). PEG10, but not CDK14 or CDK6, was significantly overexpressed in 30 of 41 tumors (73%) from HCC patients, compared with their nontumorous counterparts. These results suggest that PEG10 is a probable target, acting as a driving force for amplification of the 7q21 region, and may therefore be involved in the development or progression of HCCs.

Complete cytogenetic and molecular response to treatment with imatinib mesylate for philadelphia chromosome positive acute myeloid leukemia with multilineage dysplasia

The Philadelphia (Ph) chromosome positive acute myeloid leukemia (AML) is an uncommon disorder with poor prognosis when treated with conventional chemotherapy [1,2]. Here, we report long-lasting complete molecular remission (CMR) induced by imatinib mesylate monotherapy in a patient of Phþ AML with multilineage dysplasia. A 64-year-old female presented in November 2002 with asymptomatic anemia and neutropenia without a significant past medical history. The initial WBC count was 2.7610/L without a circulating myeloblast. Hb level and platelet count were 9.7 g/dl and 334610/L, respectively. The NAP score was 284 and serum vitamin B12 was 810 pg/ml. There was no splenomegaly or lymphadenopathy. Bone marrow (BM) aspiration showed a hypercellular marrow with immature blasts of 22.8% of all nucleated cells and morphological dysplasia including micromegakaryocytes, hypogranulated pseudo Pelger-Hüet neutrophils and megaloblastoid erythroblasts. Cytogenetic analysis of cultured BM cells identified 46,XX,t(9;22)(q34;q11) karyotype in all twenty analysed metaphases. The bcr-abl fusion signal was positive in 37 of 100 analyzed interphase peripheral mononuclear cells by double color fluorescence in situ hybridization (FISH) analysis using BCR/ ABL ES dual color translocation probe (Vysis, USA) following the manufacturer’s protocol. The blast cells were positive for myeloid antigens comprising CD13, CD33, HLA-DR and MPO. Lymphoid markers were negative. Lack of myeloproliferative features and existence of morphological dysplasia did not meet the criteria for chronic myeloid leukemia (CML); consequently, the patient was diagnosed with de novo Phþ AML with multilineage dysplasia. She was initially treated with oral vitamin D3, as is used for myelodysplastic syndrome (MDS). Eight months after the initial diagnosis, the initial treatment had no effect on either the WBC or the Hb level and the number of blasts increased to reach 50.4% with the cytogenetic findings of the same as the initial abnormal karyotype. The major bcr-abl transcription was detected by means of quantitative reverse transcription polymerase chain reaction (RT-PCR), and identified 230 copies/mg RNA in BM cells. Instead of conventional chemotherapy, imatinib was solely administered at a daily dose of 400 mg/ body after written informed consent was obtained because of the patient’s age and the existence of multilineage dysplasia, which could be associated with poor sensitivity for chemotherapeutic agents [1,2]. After the start of imatinib administration, NCI-CTC grade IV leukocytopenia and thrombocytopenia occurred and imatinib was transiently ceased between day 30 and 42 (Figure 1). Therafter, imatinib was restarted at a daily dose of 300 mg/body. On day 59, complete hematologic remission (CHR) was achieved with 4.8% blasts in BM without a circulating blast,

HEPATOBILIARY SARCOIDOSIS: TYPICAL APPEARANCE IN LAPAROSCOPY

We present a case of systemic sarcoidosis. A 72‐year‐old Japanese woman, who was diagnosed with asymptomatic enlarged abdominal para‐aortic lymph nodes in April 2000, was admitted to our hospital in December 2003 because of multiple mass lesions of the liver and spleen. She had hepatosplenomegaly and asymptomatic pulmonary fibrosis. Blood examination revealed cholestasis and progressing liver synthetic dysfunction. In laparoscopic photographs, various sizes of whitish flat lesions were scattered on the surface of the liver. Liver biopsy showed non‐caseating epithelioid cell granulomas. These lesions of the spleen disappeared on contrast‐enhanced computed tomography imaging, and serum liver dysfunction was improved rapidly after steroid therapy. Laparoscopy may be useful for the diagnosis of granulomatous liver disease, such as hepatobiliary sarcoidosis.