Takehisa Suzuki

Yokohama City University, Yokohama, Kanagawa, Japan

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Publications (12)37.34 Total impact

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    ABSTRACT: An autopsy case of leptomeningeal melanomatosis associated with neurocutaneous melanosis (NCM) involving a 44-year-old male is reported. The autopsy showed that the leptomeningeal surface of the brain and the spinal cord were covered with a diffuse black lesion. A histological examination detected diffusely distributed, proliferating, melanin-containing cells and demonstrated that the lesion consisted of three different components; i.e. regions of melanomatosis, melanocytosis, and melanocyte hyperplasia. In the leptomeningeal melanomatosis component, tumor cells with pleomorphic nuclei and prominent nucleoli had infiltrated into the cerebral parenchyma via Virchow–Robin spaces. The Ki-67 labeling index and the nuclear accumulation of p53 and p16 protein were immunohistochemically examined in each component. The Ki-67 labeling indices of the melanomatosis, melanocytosis, and melanocyte hyperplasia components were 8.7%, 0.8%, and 0%, respectively. Immunostaining of nuclear p16 produced a negative result in the melanomatosis component, but positive results in the melanocytosis and melanocyte hyperplasia components, whereas nuclear p53 expression was not detected in any of the components. This case suggests that p16INK4/CDKN2 may play a significant role in progression of leptomeningeal melanocytic neoplasms. We also reviewed previously reported cases of leptomeningeal neoplasms associated with NCM and discussed the relationship between the biological behavior and proliferative activity of such lesions.
    Pathology International 01/2015; · 1.72 Impact Factor
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    ABSTRACT: This study investigated the proteome modulated by oncogenic KRAS in immortalized airway epithelial cells. Chloride intracellular channel protein 4 (CLIC4), S100 proteins (S100A2 and S100A11), tropomyosin 2, cathepsin L1, integrinsα3, eukaryotic elongation factor 1, vimentin, and others were discriminated. We here focused on CLIC4 to investigate its potential involvement in carcinogenesis in the lung because previous studies suggested that some chloride channels and chloride channel regulators could function as tumor suppressors. CILC4 protein levels were reduced in some lung cancer cell lines. The restoration of CLIC4 in lung cancer cell lines in which CLIC4 expression was reduced attenuated their growth activity. The immunohistochemical expression of the CLIC4 protein was weaker in primary lung cancer cells than in non-tumorous airway epithelial cells and was occasionally undetectable in some tumors. CLIC4 protein levels were significantly lower in a subtype of mucinous ADC than in others, and were also significantly lower in KRAS-mutated ADC than in EGFR-mutated ADC. These results suggest that the alteration in CLIC4 could be involved in restrictedly the development of a specific fraction of lung adenocarcinomas. The potential benefit of the proteome modulated by oncogenic KRAS to lung cancer research has been demonstrated.
    PLoS ONE 01/2014; 9(2):e87193. · 3.53 Impact Factor
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    ABSTRACT: Our previous studies identified important molecules involved in lung carcinogenesis through a comprehensive search for the downstream targets of oncogenic KRAS, and these findings suggested that an investigation into the downstream targets of oncogenic KRAS might represent a useful strategy for elucidating the common molecular bases of lung cancer. Among the downstream targets of oncogenic KRAS, a focus was placed on HDAC9, a member of the histone deacetylase family, in the present study because epigenetic modification of DNA or the histone proteins is known to play an important role in carcinogenesis. The immunohistochemical expression of HDAC9 was examined in surgically resected primary lung cancers (130 adenocarcinoma, 49 squamous cell carcinomas, one large cell carcinoma, and 6 small cell carcinomas) and potential associations between its expression level and pathologic factors were analyzed. The results showed that HDAC9 expression levels were lower in lung cancer cells than in non-tumor epithelial cells, and were also significantly lower in adenocarcinomas among the histological types. Moreover, HDAC9 expression levels were significantly lower in adenocarcinomas with lymphatic canal involvement. The restoration of HDAC9 in lung cancer cells losing its expression severely attenuated their growth activity in vitro. These results suggest that HDAC9 may be a suppressor and its downregulation might promote the progression process, especially in lung adenocarcinomas.
    International journal of clinical and experimental pathology 01/2013; 7(1):213-20. · 2.24 Impact Factor
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    ABSTRACT: Vascular endothelial growth factor-A (VEGF-A) is crucial for angiogenesis, vascular permeability, and metastasis during tumor development. We demonstrate here that early growth response-1 (EGR-1), which is induced by the extracellular signal-regulated kinase (ERK) pathway activation, activates VEGF-A in lung cancer cells. Increased EGR-1 expression was found in adenocarcinoma cells carrying mutant K-RAS or EGFR genes. Hypoxic culture, siRNA experiment, luciferase assays, chromatin immunoprecipitation, electrophoretic mobility shift assays, and quantitative RT-PCR using EGR-1-inducible lung cancer cells demonstrated that EGR-1 binds to the proximal region of the VEGF-A promoter, activates VEGF-A expression, and enhances hypoxia inducible factor 1alpha (HIF-1alpha)-mediated VEGF-A expression. The EGR-1 modulator, NAB-2, was rapidly induced by increased levels of EGR-1. Pathology samples of human lung adenocarcinomas revealed correlations between EGR-1/HIF-1alpha and VEGF-A expressions and relative elevation of EGR-1 and VEGF-A expression in mutant K-RAS- or EGFR-carrying adenocarcinomas. Both EGR-1 and VEGF-A expression increased as tumors dedifferentiated, whereas HIF-1alpha expression did not. Although weak correlation was found between EGR-1 and NAB-2 expressions on the whole, NAB-2 expression decreased as tumors dedifferentiated, and inhibition of DNA methyltransferase/histone deacetylase increased NAB-2 expression in lung cancer cells despite no epigenetic alteration in the NAB-2 promoter. These findings suggest that EGR-1 plays important roles on VEGF-A expression in lung cancer cells, and epigenetic silencing of transactivator(s) associated with NAB-2 expression might also contribute to upregulate VEGF-A expression.
    American Journal Of Pathology 07/2010; 177(1):70-83. · 4.60 Impact Factor
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    ABSTRACT: Small cell lung cancer (SCLC) exhibits insulin-like growth factor-dependent growth. SCLC is the most aggressive among known in vivo lung cancers, whereas in vitro growth of SCLC is paradoxically slow as compared with that of non-SCLC (NSCLC). In this study, we demonstrate that SCLC cells overexpress insulin-like growth factor binding protein (IGFBP)-2 via NeuroD, a neuroendocrine cell-specific transcription factor. Chromatin immunoprecipitation, electrophoretic mobility shift, and IGFBP-2 promoter assays all revealed that NeuroD binds to the E-box in the 5'-untranslated region of IGFBP-2. A NeuroD transgene in both airway epithelial and NSCLC cells up-regulated the transcription of IGFBP-2 and retarded cell growth. Recombinant IGFBP-2 repressed the growth of both airway epithelial and NSCLC cells in a dose-dependent manner. A NeuroD-specific small interfering RNA repressed IGFBP-2 expression in SCLC, and neutralization of IGFBP-2 and an IGFBP-2-specific small interfering RNA increased SCLC cell growth. Pathological samples of SCLC also expressed IGFBP-2 abundantly, as compared with NSCLC, and showed only rare (8%) IGFBP-2 promoter methylation, whereas the IGFBP-2 promoter was methylated in 71% of adenocarcinomas and 29% of squamous cell carcinomas. These findings suggest that 1) SCLC has an IGFBP-2 overexpression mechanism distinct from NSCLC, 2) secreted IGFBP-2 contributes to the slow growth of SCLC in vitro, and 3) the epigenetic alterations in the IGFBP-2 promoter contribute to the striking differences in IGFBP-2 expression between SCLC and NSCLC in vivo.
    American Journal Of Pathology 09/2009; 175(3):976-87. · 4.60 Impact Factor
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    ABSTRACT: The authors' previous study demonstrated that oncogenic KRAS modulates the shape and motility of airway epithelial cells. To explore detailed mechanism mediating these events, the possible involvement of phosphatidylinositides (PIP) was investigated. The intracellular localization of PIP was visualized with a pleckstrin homology domain-enhanced green fluorescent protein (EGFP) construct. PIP accumulated at the leading edges of polarizing epithelial cells, while they co-localized with cortical actin at cell-cell contacts, suggesting that PIP play important roles in the cytoskeletal organization. Transduction of oncogenic KRAS induced multiple pseudopodia and disrupted cortical actin, enhancing motility. A mitogen activated protein kinase kinase (MEK) inhibitor reduced the accumulation of PIP at membranes and development of pseudopodia, and restored stable cortical actin, reducing the motility. A phosphoinositide 3-kinase (PI3K) inhibitor also reduced accumulation of PIP at membranes, formation of pseudopodia and motility, but its effect on cortical actin was indistinct. The KRAS V12/S35 mutant, activating only the MEK pathway, induced multiple pseudopodia and disrupted the cortical actin. The KRAS V12/C40 mutant, activating only the PI3K pathway, also induced pseudopodia, but its effect on cortical actin was obscure. Taken together, oncogenic KRAS could cause the accumulation of PIP via the PI3K and MEK pathways and modulate the cell shape and migration.
    Pathology International 02/2009; 59(1):28-37. · 1.72 Impact Factor
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    ABSTRACT: Gain-of-function point mutations in K-ras affect early events in pulmonary bronchioloalveolar carcinoma. We investigated altered mRNA expression on K-Ras activation in human peripheral lung epithelial cells (HPL1A) using oligonucleotide microarrays. Mutated K-Ras stably expressed in HPL1A accelerated cell growth and induced the expression of insulin-like growth factor (IGF)-binding protein (IGFBP)-4 and IGFBP-2, which modulate cell growth via IGF. Other lung epithelial cell lines (NHBE and HPL1D) revealed the same phenomena as HPL1A by mutated K-ras transgene. Lung cancer cell growth was also accelerated by mutated K-ras gene transduction, whereas IGFBP-4/2 induction was weaker compared with mutated K-Ras-expressing lung epithelial cells. To understand the differences in IGFBP-4/2 inducibility via K-Ras-activated signaling between nonneoplastic lung epithelia and lung carcinoma, we addressed the mechanisms of IGFBP-4/2 transcriptional activation. Our results revealed that Egr-1, which is induced on activation of Ras-mitogen-activated protein kinase signaling, is crucial for transactivation of IGFBP-4/2. Furthermore, IGFBP-4 and IGFBP-2 promoters were often hypermethylated in lung carcinoma, yielding low basal expression/weak induction of IGFBP-4/2. These findings suggest that continuous K-Ras activation accelerates cell growth and evokes a feedback system through IGFBP-4/2 to prevent excessive growth. Moreover, this growth regulation is disrupted in lung cancers because of promoter hypermethylation of IGFBP-4/2 genes.
    American Journal Of Pathology 12/2006; 169(5):1550-66. · 4.60 Impact Factor
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    ABSTRACT: The expression of vascular endothelial growth factors (VEGFs) in tumors including lung cancer is considered to be associated with tumor development via capillary and lymph vessel neogenesis. Dissemination of the tumor cells to the pleura or regional lymph nodes is a critical poor prognostic factor for lung cancer patients. To investigate how VEGFs expressed in the intrathoracic infiltrating lung cancer cells participate in disease progression, we established stably VEGF-A-, VEGF-C-, VEGF-D-, VEGF-A and VEGF-C-, and VEGF-A and VEGF-D-expressing large cell lung cancer clones (TKB5/VEGF-A, TKB5/VEGF-C, TKB5/VEGF-D, TKB5/VEGF-A/C, and TKB5/VEGF-A/D), orthotopically inoculated these into the right thoracic cavity (i.t.) of nude mice, and evaluated the subsequent development of lung lesion, pleural effusion, pleural dissemination, and lymph node metastasis. While there were no significant differences either in culture or in subcutaneous tumor cell growth between the empty vector-transfected group (TKB5/empty) and each transfectant, the i.t. model demonstrated significantly different biological properties between the transfectants. TKB5/empty-inoculated mice frequently developed a large tumor on the pleura without pleural effusion, dissemination, or lymph node (LN) metastasis. In contrast, VEGF-A promoted a bloody pleural effusion (6/14), and VEGF-A and VEGF-D frequently generated pleural dissemination (11/14 and 9/11, respectively). Although both VEGF-C and VEGF-D generated LN metastasis (6/10 and 8/11, respectively), the locations of the metastasized LNs were quite different. TKB5/VEGF-C metastasized on the same side of axillary LNs as i.t. (right axillary LNs), whereas TKB5/VEGF-D metastasized to the mediastinal and left axillary and/or cervical LNs. Since the TKB5/VEGF-A/C or TKB5/VEGF-A/D co-transfectants revealed overlapping tumor progression patterns of VEGF-A and VEGF-C or VEGF-D, the metastatic LNs had abundant new capillaries and were larger than those of TKB5/VEGF-C or TKB5/VEGF-D-inoculated mice. Our results clearly demonstrate that VEGF-A secreted from intrathoracic lung cancer cells plays important roles in producing pleural effusion, dissemination, and capillary neogenesis, that VEGF-C is involved in LN metastasis, and VEGF-D in pleural dissemination and LN metastasis. It is most likely, however, that the mechanisms by which VEGF-C promotes LN metastasis are different from those of VEGF-D. The regulation of the expression of VEGFs in intrathoracic lung cancer cells might be a useful therapeutic approach to inhibiting tumor development and improving patient prognosis.
    Lung Cancer 10/2004; 45(3):325-37. · 3.39 Impact Factor
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    ABSTRACT: Point mutations of the K-ras gene, which are found in 10 to 30% of lung adenocarcinomas, are regarded as being an early event during the carcinogenesis. Autonomous vigorous motility of neoplastic cells, as well as growth and survival advantages, are considered to be necessary for cancer development and progression. The present study describes the contributions of the K-ras gene mutation and its downstream pathway via phosphatidylinositol 3-OH kinase (PI3K)-Akt to the cell motility in an immortalized human peripheral airway epithelial cell (HPL1D) and lung adenocarcinoma cells (A549, H820, TKB6, and TKB14). We have also evaluated the relationship between pathological events and the K-ras-Akt pathway using surgically resected lung tumors. The HPL1D cells transfected with the mutated K-ras gene (HPL-V12) showed a significant increase in cell motility compared to those transfected with empty vector (HPL-E) or wild-type K-ras gene (HPL-K). The enhanced motility in the HPL-V12 cells was markedly reduced by either treatment with inhibitors of ras, PI3K, and/or MEK, or by transfection with the dominant-negative mutant Akt (dnAkt). The lung adenocarcinoma cells bearing the K-ras gene mutation (A549 and H820) showed consistently higher levels of cell motilities than those without the mutation (TKB6 and TKB14), and the motility of A549 and H820 cells were significantly inhibited by dnAkt transfection. These results suggest that the K-ras gene mutation could enhance the motility of neoplastic cells through a pathway involving PI3K-Akt. Actually, among the surgically resected lung tumors, the adenocarcinomas with the K-ras gene mutation tended to show a higher frequency and intensity of immunoreactivity for phosphorylated Akt (p-ser473Akt) than those without the mutation, supporting the in vitro observation that the mutated K-ras can activate the PI3K-Akt pathway. Immunoreactivity for p-ser473Akt was also seen in the pre-malignant and early lesions at a frequency similar to that in the advanced lung adenocarcinomas,. No correlation was seen between p-ser473Akt immunoreactivity and lymphatic/organ metastasis or prognosis. These results taken together suggest that the K-ras-Akt pathway might facilitate the motility of neoplastic cells during the early period of carcinogenesis in lung adenocarcinomas, and may contribute to their non-invasive expansion along the alveolar septa, rather than invasion or metastasis.
    American Journal Of Pathology 02/2004; 164(1):91-100. · 4.60 Impact Factor
  • American Journal of Pathology - AMER J PATHOL. 01/2004; 164(1):91-100.
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    ABSTRACT: We report an extremely rare case of primary lung cancer showing various histological elements diagnosed as the collision of an adenosquamous carcinoma and a large cell neuroendocrine carcinoma by loss of heterozygosity (LOH) analysis of the human androgen receptor (AR) and phosphoglycerate kinase (PGK-1) genes. The tumor exhibited a tiny ground-glass opaque shadow suggesting atypical adenomatous hyperplasia 18 months prior to surgery. However, the tumor grew rapidly, and the resected tumor consisted of two closely located nodules. The larger nodule was composed of well-differentiated adenocarcinomatous and moderately to poorly differentiated squamous cell carcinomatous elements, while the smaller nodule consisted of a large cell neuroendocrine carcinomatous element with partial squamoid differentiation having focal continuity with the adenocarcinomatous element. Both the adenocarcinomatous and squamous cell carcinomatous elements revealed transitional features and LOH of AR and PGK-1 genes, while the large cell neuroendocrine carcinomatous element showed a monoclonal pattern but possessed both alleles of AR and PGK-1 genes. From these clinical and pathological results, the parental cell of the large cell neuroendocrine carcinomatous element was considered to be different from that of the adenosquamous carcinomatous element.
    Pathology International 02/2003; 53(1):58-65. · 1.72 Impact Factor
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    ABSTRACT: Small cell lung cancer (SCLC) and neuroblastoma (NB), the most aggressive adult and infant neuroendocrine cancers, respectively, are immunologically characterized by a severe reduction in major histocompatibility complex (MHC) that is indispensable for anti-tumor immunity. We had reported that the severe reduction of MHC in SCLC was caused by a deficient interferon (IFN)-gamma-inducible expression of class II transactivator (CIITA) that is known as a very important transcription factor for IFN-gamma-inducible class II and class I MHC expression (Yazawa T, Kamma H, Fujiwara M, Matsui M, Horiguchi H, Satoh H, Fujimoto M, Yokohama K, Ogata T: Lack of class II transactivator causes severe deficiency of HLA-DR expression in small cell lung cancer. J Pathol 1999, 187:191-199). Here, we demonstrate that the reduction of MHC in NB was also caused by a deficient IFN-gamma-inducible expression of CIITA and that the deficiency in SCLC and NB was caused by similar mechanisms. Human achaete-scute complex homologue (HASH)-1, L-myc, and N-myc, which are specifically overexpressed in SCLC and NB, bound to the E-box in CIITA promoter IV and reduced the transcriptional activity. Anti-sense oligonucleotide experiments revealed that overexpressed L-myc and N-myc lie upstream in the regulatory pathway of HASH-1 expression. The expression of HASH-1 was also up-regulated by IFN-gamma. Our results suggest that SCLC and NB have complicated mechanisms of IFN-gamma-inducible CIITA transcription deficiency through the overexpressed HASH-1, L-myc, and N-myc. These complicated mechanisms may play an important role in the escape from anti-tumor immunity.
    American Journal Of Pathology 08/2002; 161(1):291-300. · 4.60 Impact Factor