薬剤代謝酵素の遺伝子多型と胃癌感受性についての研究

取得学位：博士(医学)，学位授与番号：医博甲第1838号，学位授与年月日：平成13年3月22日,学位授与年：200

肺癌に対する分子標的薬剤併用療法適応拡大の検討

金沢大学医学系本年度は,肺癌におけるEGFR(epidermal growth factor receptor)蛋白の過剰発現,EGFR遺伝子増幅,EGFR遺伝子変異,PDGF(platelet-derived growth factor)の発現について検索した.1.肺癌におけるEGFR蛋白過剰発現の免疫染色による検索159例の肺癌のホルマリン固定・パラフィン包埋組織で抗EGFR抗体を用いて免疫染色を行ったところ,EGFRの過剰発現は61例(38%)の症例にみられた.2.FISH(fluorescence in situ hybridization)法によるEGFR遺伝子増幅の検索上記EGFR過剰発現を示す61症例に対してEGFR(7p12)と染色体7セントロメアを認識する2色プローブを用いた2色FISH法を行ったところ,11例でEGFR遺伝子増幅,11例で7番染色体polysomyがみられた.3.ダイレクトシークエンス法によるEGFR遺伝子変異の検索159例の肺癌凍結標本から抽出したDNAを用いて,EGFR遺伝子のexon18-21のダイレクトシークエンス法を行ったところ,60例(38%)においてEGFR遺伝子変異がみられた.EGFR遺伝子コピー数の増加を示さない群(137例)の37%,EGFR遺伝子増幅を示す群(11例)の55%,polysomyを有する群(11例)の33%の症例で変異がみられた.4.肺癌におけるPDGF蛋白発現の免疫染色による検索PDGFの発現は全体の90%の症例でみられ,さらに24%の症例において中等度以上の染色強度がみられた.上記1-4.で得られた結果の一部は,他の実験結果とともに学術雑誌上および学会において報告した(下記「雑誌論文」と「学会発表」の項目を参照).研究課題/領域番号:22790340, 研究期間(年度):2010-04-01 – 2013-03-31出典：研究課題「肺癌に対する分子標的薬剤併用療法適応拡大の検討」課題番号22790340（KAKEN：科学研究費助成事業データベース（国立情報学研究所）） （https://kaken.nii.ac.jp/ja/grant/KAKENHI-PROJECT-22790340/）を加工して作

Roles of epithelial-mesenchymal transition in squamous cell carcinoma of the temporal bone

金沢大学医薬保健研究域医学系Objectives: The authors recently demonstrated that extensive bone erosion correlated with a worse prognosis of the squamous cell carcinoma (SCC) of the temporal bone. The present objective was to investigate the expressions and the roles of epithelial-mesenchymal transition (EMT) in SCC of the temporal bone. Study Design: Retrospective patient file review and immunohistochemical study of tissues of patients with SCC of the temporal bone. Patients: The authors examined the expressions of vimentin in 16 patients with SCC of the temporal bone by immunohistochemical analysis. Results: As expected, vimentin was expressed on SCC of the temporal bone. We regard the patient who has vimentin-positive cells as EMT-positive patient because gain of vimentin is a hallmark in the EMT reported in several types of cancer. EMT positive tumors significantly correlated with extensive bone erosion, and extensive bone erosion correlated with a worse prognosis of the SCC of the temporal bone. Conclusion: The study results suggest that EMT in tumor cells becomes an important indicator of poor prognosis in SCC of the temporal bone. Copyright © 2011 Otology & Neurotology, Inc. Unauthorized reproduction of this article is prohibited

Cytological Features of a Variant NUT Midline Carcinoma of the Lung Harboring the NSD3-NUT

Background. Nuclear protein in testis (NUT) midline carcinoma (NMC) is a very rare and aggressive malignancy. In more than two-thirds of these NMC cases, a fusion between NUT and BRD4 or BRD3 has been documented; other variants are rare. The cytology of NMC itself has been sparsely documented and that of variant NMC has never been reported. Case Presentation. A 36-year-old woman was admitted because of a rapidly progressing lung tumor with metastases to the breast and bone. We recently reported this patient as the first case of a variant NMC of the lung harboring an NSD3-NUT fusion, based on immunohistochemical and genetic analyses. Cytological material was available for the present review. A highly cellular smear contained a predominantly noncohesive pattern of monomorphic cells with diameters 2–2.5 times greater than those of small lymphocytes, with a round-to-oval nucleus, slightly irregular nuclear contours, variably prominent nucleoli, scant cytoplasm, and identifiable mitotic figures. Foci of stratification and overt pearl formation, including a dyskeratocyte, were occasionally observed. The necrotic background contained naked nuclei, karyorrhectic debris, apoptotic cells, and macrophages phagocytizing karyorrhectic debris; nuclear crushing was noted. Conclusion. The cytological features of a variant NMC of the lung are described for the first time

Clinicopathological significance of platelet-derived growth factor (PDGF)-B and vascular endothelial growth factor-A expression, PDGF receptor-β phosphorylation, and microvessel density in gastric cancer

Background: Angiogenesis is important in the growth and metastasis of various kinds of solid tumors, including gastric cancers. The angiogenic process is triggered by several key growth factors, including vascular endothelial growth factor (VEGF)-A and platelet-derived growth factor (PDGF)-B, that are secreted by tumors. Our aim was to define: i) the expression pattern of VEGF-A and PDGF-B in tumor cells and the activation of PDGF receptor (PDGFR)-β tyrosine kinase in stromal cells of human gastric adenocarcinomas; and ii) the relationship between VEGF-A and PDGF-B expression and microvessel density (MVD), to determine if there is a rationale for a new therapeutic strategy.Methods: A series of 109 gastric adenocarcinoma cases that had undergone surgical resection was examined immunohistochemically using antibodies against VEGF-A, PDGF-B, and CD34, followed by further examination of PDGFR-β phosphorylation by immunoblotting analysis.Results: MVD was higher in diffuse-type than intestinal-type cancers (p < 0.001). VEGF-A overexpression correlated to PDGF-B overexpression in both the intestinal-type (p < 0.005) and diffuse-type (p < 0.0001) groups, indicating that VEGF-A and PDGF-B are secreted simultaneously in the same tumor, and may thus play important roles together in angiogenesis. However, several differences between intestinal-type and diffuse-type cancers were observed. In the diffuse-type cancer group, higher MVD was related to the PDGF-B proportion (p < 0.05) and VEGF-A overexpression (p < 0.05), but not to PDGF-B overexpression or the VEGF-A proportion. On the other hand, in the intestinal-type cancer group, higher MVD was correlated to overexpression (p < 0.005), intensity (p < 0.05), and proportion (p < 0.05) of PDGF-B, but not of VEGF-A. In addition, phosphorylation of PDGFR-β was correlated with depth of cancer invasion at statistically significant level.Conclusions: Our results indicate that PDGF-B, which is involved in the maintenance of microvessels, plays a more important role in angiogenesis in intestinal-type gastric carcinomas than VEGF-A, which plays a key role mainly in the initiation of new blood vessel formation. In contrast, VEGF-A has a critical role for angiogenesis more in diffuse-type cancers, but less in those of intestinal type. Thus, a therapy targeting the PDGF-B signaling pathway could be effective for intestinal-type gastric carcinoma, whereas targeting VEGF-A or both VEGF-A and PDGF-B signaling pathways could be effective for diffuse-type gastric carcinomas. © 2010 Suzuki et al; licensee BioMed Central Ltd

Gene amplification of ERBB2 and EGFR in adenocarcinoma in situ and intramucosal adenocarcinoma of Barrett\u27s esophagus

金沢大学医薬保健研究域医学系We examined 11 cases of carcinoma arising from Barrett\u27s esophagus consisting of two adenocarcinomas in situ (ACIS), two intramucosal adenocarcinomas, and seven overt invasive adenocarcinomas. Overexpression of p53 (implying a mutation of the p53 gene), ERBB2, and EGFR was measured by immunohistochemistry, and gene amplification of ERBB2 and EGFR was measured by fluorescence in situ hybridization (FISH). In all cases of ACIS and the intramucosal adenocarcinomas, almost all cancer cells overexpressed p53, however the populations overexpressing ERBB2 and EGFR varied in different cases: in one ACIS, ERBB2 was coexpressed in all the cancer cells, in the other ACIS and one intramucosal adenocarcinoma, ERBB2 was overexpressed in about 50% and only 10% of the p53-positive cells respectively. EGFR was co-expressed in 20% in the other intramucosal adenocarcinoma. Protein overexpression of ERBB2 or EGFR corresponded to the amplification of their respective genes on a cell by cell basis. These gene amplifications, however, were not found in the seven invasive adenocarcinomas. Thus we speculate that the gene amplification occurred late in the dysplasia-carcinoma sequence probably after the mutation of p53. Furthermore, new clonal expansion accompanied by tumor invasion might have extinguished the originally amplified genes in these tumors. © 2010 Japanese Society of Pathology and Blackwell Publishing Asia Pty Ltd

Platelet-derived growth factor receptor-β, carrying the activating mutation D849N, accelerates the establishment of B16 melanoma

<p>Abstract</p> <p>Background</p> <p>Platelet-derived growth factor (PDGF)-BB and PDGF receptor (PDGFR)-β are mainly expressed in the developing vasculature, where PDGF-BB is produced by endothelial cells and PDGFR-β is expressed by mural cells, including pericytes. PDGF-BB is produced by most types of solid tumors, and PDGF receptor signaling participates in various processes, including autocrine stimulation of tumor cell growth, recruitment of tumor stroma fibroblasts, and stimulation of tumor angiogenesis. Furthermore, PDGF-BB-producing tumors are characterized by increased pericyte abundance and accelerated tumor growth. Thus, there is a growing interest in the development of tumor treatment strategies by blocking PDGF/PDGFR function. We have recently generated a mouse model carrying an activated PDGFR-β by replacing the highly conserved aspartic acid residue (D) 849 in the activating loop with asparagine (N). This allowed us to investigate, in an orthotopic tumor model, the role of increased stromal PDGFR-β signaling in tumor-stroma interactions.</p> <p>Methods</p> <p>B16 melanoma cells lacking PDGFR-β expression and either mock-transfected or engineered to express PDGF-BB, were injected alone or in combination with matrigel into mice carrying the activated PDGFR-β (D849N) and into wild type mice. The tumor growth rate was followed and the vessel status of tumors, i.e. total vessel area/tumor, average vessel surface and pericyte density of vessels, was analyzed after resection.</p> <p>Results</p> <p>Tumors grown in mice carrying an activated PDGFR-β were established earlier than those in wild-type mice. In this early phase, the total vessel area and the average vessel surface were higher in tumors grown in mice carrying the activated PDGFR-β (D849N) compared to wild-type mice, whereas we did not find a significant difference in the number of tumor vessels and the pericyte abundance around tumor vessels between wild type and mutant mice. At later phases of tumor progression, no significant difference in tumor growth rate was observed between wild type mice and mutant mice, although the pericyte coverage was higher around tumor vessels from mutant mice.</p> <p>Conclusion</p> <p>Our findings suggest that the activated PDGFR-β (D849N) in the host animal increased the total vessel area and the average vessel surface even in PDGF-negative tumors, resulting in a shorter lag phase during tumor establishment.</p

Two genetic variants of CD38 in subjects with autism spectrum disorder and controls

金沢大学医薬保健研究域医学系The neurobiological basis of autism spectrum disorder (ASD) remains poorly understood. Given the role of CD38 in social recognition through oxytocin (OT) release, we hypothesized that CD38 may play a role in the etiology of ASD. Here, we first examined the immunohistochemical expression of CD38 in the hypothalamus of post-mortem brains of non-ASD subjects and found that CD38 was colocalized with OT in secretory neurons. In studies of the association between CD38 and autism, we analyzed 10 single nucleotide polymorphisms (SNPs) and mutations of CD38 by re-sequencing DNAs mainly from a case-control study in Japan, and Caucasian cases mainly recruited to the Autism Genetic Resource Exchange (AGRE). The SNPs of CD38, rs6449197 (p 70; designated as high-functioning autism (HFA)) in the U.S. 104 AGRE family trios, but not with Japanese 188 HFA subjects. A mutation that caused tryptophan to replace arginine at amino acid residue 140 (R140W; (rs1800561, 4693C>T)) was found in 0.6-4.6% of the Japanese population and was associated with ASD in the smaller case-control study. The SNP was clustered in pedigrees in which the fathers and brothers of T-allele-carrier probands had ASD or ASD traits. In this cohort OT plasma levels were lower in subjects with the T allele than in those without. One proband with the T allele who was taking nasal OT spray showed relief of symptoms. The two variant CD38 poloymorphysms tested may be of interest with regard of the pathophysiology of ASD. © 2010 Elsevier Ireland Ltd and the Japan Neuroscience Society