Okayama University Medical School Acta Medica Okayama 0386-300X 74 2 2020 Association between Histological Types and Enhancement of Dynamic CT for Primary Lung Cancer 129 135 EN Shogo Fukuma Department of Radiology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Takayoshi Shinya Department of Radiology, Okayama University Hospital Junichi Soh Department of General Thoracic Surgery, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Ryuichiro Fukuhara Department of Pediatric Radiology, Okayama University Hospital Nanako Ogawa Department of Radiology, Okayama University Hospital Fumiyo Higaki Department of Radiology, Okayama City General Medical Center Takehiro Tanaka Department of Pathology, Okayama University Hospital Eiki Ichihara Department of Allergy and Respiratory Medicine, Okayama University Hospital Takao Hiraki Department of Radiology, Okayama University Hospital Shinichi Toyooka Department of General Thoracic Surgery, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Susumu Kanazawa Department of Radiology, Okayama University Hospital Original Article 10.18926/AMO/58271 The aim of this study was to explore enhancement patterns of different types of primary lung cancers on 2-phase dynamic computed tomography (CT). This study included 217 primary lung cancer patients (141 adenocarcinomas [ADs], 48 squamous cell carcinomas [SCCs], 20 small cell lung carcinomas [SCLCs], and 8 others) who were examined using a 2-phase dynamic scan. Regions of interest were identified and mean enhancement values were calculated. After excluding the 20 SCLCs because these lesions had different clinical stages from the other cancer types, the mean attenuation values and subtractions between phases were compared between types of non-small cell lung carcinomas (NSCLCs) using the Kruskal–Wallis test. Late phase attenuation and attenuation of the late minus unenhanced phase (LMU) of SCCs were significantly higher than those of ADs (p<0.05). To differentiate SCC and AD in the late phase, a threshold of 80.21 Hounsfield units (HU) gave 52.9% accuracy. In LMU, a threshold of 52.16 HU gave 59.3% accuracy. Dynamic lung CT has the potential to aid in differentiating among NSCLC types. No potential conflict of interest relevant to this article was reported.

Wiley Open Access Acta Medica Okayama 1347-9032 110 8 2019 Acquired resistance mechanisms to afatinib in HER2-amplified gastric cancer cells 2549 2557 EN Takahiro Yoshioka Departments of Gastroenterological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Kazuhiko Shien General Thoracic Surgery, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Tatsuaki Takeda Department of Clinical Pharmacy, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Yuta Takahashi General Thoracic Surgery, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Eisuke Kurihara General Thoracic Surgery, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Yusuke Ogoshi General Thoracic Surgery, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Kei Namba General Thoracic Surgery, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Hidejiro Torigoe General Thoracic Surgery, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Hiroki Sato General Thoracic Surgery, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Shuta Tomida Bioinformatics, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Hiromasa Yamamoto General Thoracic Surgery, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Junichi Soh General Thoracic Surgery, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Toshiyoshi Fujiwara Departments of Gastroenterological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Shinichi Toyooka General Thoracic Surgery, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Cancer treatment, especially that for breast and lung cancer, has entered a new era and continues to evolve, with the development of genome analysis technology and the advent of molecular targeted drugs including tyrosine kinase inhibitors. Nevertheless, acquired drug resistance to molecular targeted drugs is unavoidable, creating a clinically challenging problem. We recently reported the antitumor effect of a pan-HER inhibitor, afatinib, against human epidermal growth factor receptor 2 (HER2)-amplified gastric cancer cells. The purpose of the present study was to identify the mechanisms of acquired afatinib resistance and to investigate the treatment strategies for HER2-amplified gastric cancer cells. Two afatinib-resistant gastric cancer cell lines were established from 2 HER2-amplified cell lines, N87 and SNU216. Subsequently, we investigated the molecular profiles of resistant cells. The activation of the HER2 pathway was downregulated in N87-derived resistant cells, whereas it was upregulated in SNU216-derived resistant cells. In the N87-derived cell line, both MET and AXL were activated, and combination treatment with afatinib and cabozantinib, a multikinase inhibitor that inhibits MET and AXL, suppressed the cell growth of cells with acquired resistance both in vitro and in vivo. In the SNU216-derived cell line, YES1, which is a member of the Src family, was remarkably activated, and dasatinib, a Src inhibitor, exerted a strong antitumor effect in these cells. In conclusion, we identified MET and AXL activation in addition to YES1 activation as novel mechanisms of afatinib resistance in HER2-driven gastric cancer. Our results also indicated that treatment strategies targeting individual mechanisms of resistance are key to overcoming such resistance. No potential conflict of interest relevant to this article was reported.

Elsevier Acta Medica Okayama 2213-0071 28 2019 Long-term spontaneous remission with active surveillance in IgG4-related pleuritis: A case report and literature review 100938 EN Go Makimoto Department of Hematology, Oncology and Respiratory Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Kadoaki Ohashi Department of Allergy and Respiratory Medicine, Okayama University Hospital Kohei Taniguchi Department of Pathology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Junichi Soh Department of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Akihiko Taniguchi Department of Allergy and Respiratory Medicine, Okayama University Hospital Nobuaki Miyahara Department of Medical Technology, Okayama University Graduate School of Health Sciences Shinichi Toyooka Department of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Tadashi Yoshino Department of Pathology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Yoshinobu Maeda Department of Hematology, Oncology and Respiratory Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Katsuyuki Kiura Department of Allergy and Respiratory Medicine, Okayama University Hospital Pleural effusion is a relatively rare feature of IgG4-related disease (IgG4-RD). Here, we report a case of a 72-year-old woman who presented with pleural effusion. Although the pleural adenosine deaminase level was increased, surgical biopsy of the pleura and left inguinal lymph node indicated that the effusion was due to IgG4-RD. Active surveillance was initiated because serum IgG4 and pleural effusion naturally decreased and then completely disappeared. The patient has shown no recurrence for >4 years. This case suggests that pleural biopsy can be used to distinguish IgG4-RD from tuberculosis; moreover, some cases with pleural effusion could improve without treatment. No potential conflict of interest relevant to this article was reported.

Okayama University Medical School Acta Medica Okayama 0386-300X 71 2 2017 Diagnostic Value of Dual-time-point F-18 FDG PET/CT and Chest CT for the Prediction of Thymic Epithelial Neoplasms 105 112 EN Takayoshi Shinya Department of Radiology, Okayama University Hospital Takashi Tanaka Department of Radiology, Okayama University Hospital Junichi Soh Department General Thoracic Surgery, Okayama University Hospital Toshi Matsushita Department of Radiology, Okayama University Hospital Shuhei Sato Department of Radiology, Okayama University Hospital Shinichi Toyooka Department General Thoracic Surgery, Okayama University Hospital Tadashi Yoshino Department of Pathology, Okayama University Hospital Shinichiro Miyoshi Department General Thoracic Surgery, Okayama University Hospital Susumu Kanazawa Department of Radiology, Okayama University Hospital Original Article 10.18926/AMO/54978 We retrospectively assessed the dual-time-point (DTP) F-18 FDG PET/CT findings of thymic epithelial neoplasms (TENs) and investigated the diagnostic capacity of PET/CT compared to that of CT for predicting carcinoma. We calculated the ratio of the standardized uptake value of the tumor and that of the aortic arch (T/M ratio) for both the 90-min early scan and the 2-h delayed scan in 56 TEN patients. We used a multivariate logistic regression (MLR) analysis to estimate the CT features of carcinoma. We compared the diagnostic capacities of PET/CT and chest CT using receiver operating characteristic (ROC) analyses. The ROC curve revealed that the appropriate cut-off T/M ratio value for the highest accuracy was 2.39 with 75.0% accuracy. The area under the curve (AUC) was 0.855. The statistical analyses for DTP scans of 35 TEN patients demonstrated 74.3% accuracy and 0.838 AUC for the early scan versus 82.9% and 0.825 for the delayed scan. The MLR analysis indicated that mediastinal fat infiltration was a predictor of carcinoma. The ROC curve obtained for the model yielded an AUC of 0.853. Delayed scanning could improve the diagnostic capacity for carcinoma. The T/M ratio and mediastinal fat infiltration are predictive of carcinoma with moderate diagnostic accuracy. No potential conflict of interest relevant to this article was reported.

Okayama University Medical School Acta Medica Okayama 0386-300X 70 6 2016 Usefulness of Thoracoscopic Debridement for Chronic Empyema after an Extrapleural Pneumonectomy 507 510 EN Hidejiro Torigoe Department of Thoracic Surgery, Okayama University Hospital Shinichi Toyooka Department of Thoracic Surgery, Okayama University Hospital Hiromasa Yamamoto Department of Thoracic Surgery, Okayama University Hospital Junichi Soh Department of Thoracic Surgery, Okayama University Hospital Shinichiro Miyoshi Department of Thoracic Surgery, Okayama University Hospital Case Report 10.18926/AMO/54816 We present the case of a 65-year-old Japanese man diagnosed with chronic empyema (without a bronchopleural fistula) that occurred 7 months after he underwent an extrapleural pneumonectomy for right malignant pleural mesothelioma (MPM). Following thoracic drainage and irrigation for 1 month, we performed surgery by a thoracoscopic approach, in light of his general condition. We performed debridement and removal of the Gore-Tex polytetrafluoroethylene (PTFE) patch that had been used for the reconstruction of the diaphragm and the pericardium. The empyema had not relapsed when he died from recurrence of the MPM at 4 months after the thoracoscopic surgery. This patientʼs case suggests that thoracoscopic debridement and patch removal can be a therapeutic option for not only early-stage (exudative or fibrinopurulent) empyema but also late-stage (organized and chronic) empyema without a bronchopleural fistula, particularly for patients in poor general condition. No potential conflict of interest relevant to this article was reported.

Okayama University Medical School Acta Medica Okayama 0386-300X 70 4 2016 Study about the Efficacy of Metformin to Immune Function in Cancer Patients 327 330 EN Mototsugu Watanabe Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Hiromasa Yamamoto Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Shingo Eikawa Department of Immunology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Kazuhiko Shien Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Tadahiko Shien Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Junichi Soh Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Katsuyuki Hotta Center for Innovative Clinical Medicine, Okayama University Hospital Jun Wada Department of Nephrology, Rheumatology, Endocrinology and Metabolism, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Shiro Hinotsu Center for Innovative Clinical Medicine, Okayama University Hospital Toshiyoshi Fujiwara Department of Gastroenterological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Katsuyuki Kiura Department of Hematology, Oncology and Respiratory Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Hiroyoshi Doihara Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Shinichiro Miyoshi Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Heiichiro Udono Department of Immunology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Shinichi Toyooka Department of Thoracic, Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Clinical Study Protocols 10.18926/AMO/54514 A study to evaluate the effect of metformin on the immune system was commenced in July 2014. Metformin is one of the most commonly prescribed drugs for type 2 diabetes, and previous studies have reported that metformin has an anti-tumor effect. The aim of this study is to evaluate the efficacy of metformin on the immune system in human cancer patients in vivo. The primary outcome parameter will be the rate change in the population of CD8＋ T cells, which produce multiple cytokines. No potential conflict of interest relevant to this article was reported.

Okayama University Medical School Acta Medica Okayama 0386-300X 68 4 2014 Drug Resistance to EGFR Tyrosine Kinase Inhibitors for Non-small Cell Lung Cancer 191 200 EN Kazuhiko Shien Hiromasa Yamamoto Junichi Soh Shinichiro Miyoshi Shinichi Toyooka Review 10.18926/AMO/52785 Non-small cell lung cancer (NSCLC) harboring an activating mutation within the epidermal growth factor receptor (EGFR) was defined as a clinically distinct molecular group. These lesions show oncogene addiction to EGFR and dramatic responses to the EGFR tyrosine kinase inhibitors (TKIs). Several large Phase III trials have shown that EGFR-TKIs improved the progression-free survival of patients with EGFR mutant NSCLC compared to conventional chemotherapy. However, the long-term effectiveness of EGFR-TKIs is usually limited because of acquired drug resistance. To overcome this resistance to EGFR-TKIs, it will be essential to identify the specific mechanisms underlying the resistance. Many investigators have attempted to identify the mechanisms using preclinical models and drug-resistant clinical samples. As a result, several mechanisms have been showed to be responsible for the resistance, but not all of the relevant mechanisms have been uncovered. In this review, we provide an overview of mechanisms underlying drug-resistance to EGFR-TKIs, focusing on results obtained with preclinical models, and we present some possible strategies to overcome the EGFR-TKI resistance. No potential conflict of interest relevant to this article was reported.

Springer Acta Medica Okayama 0171-5216 138 5 2012 DNA methylation status of REIC/Dkk-3 gene in human malignancies 799 809 EN Tatsuro Hayashi Hiroaki Asano Shinichi Toyooka Kazunori Tsukuda Junichi Soh Tadahiko Shien Naruto Taira Yuho Maki Norimitsu Tanaka Hiroyoshi Doihara Yasutomo Nasu Nam-ho Huh Shinichiro Miyoshi The REIC (reduced expression in immortalized cells)/Dkk-3 is down-regulated in various cancers and considered to be a tumor suppressor gene. REIC/Dkk-3 mRNA has two isoforms (type-a,b). REIC type-a mRNA has shown to be a major transcript in various cancer cells, and its promoter activity was much stronger than that of type-b. In this study, we examined the methylation status of REIC/Dkk-3 type-a in a broad range of human malignancies. We examined REIC/Dkk-3 type-a methylation in breast cancers, non-small-cell lung cancers, gastric cancers, colorectal cancers, and malignant pleural mesotheliomas using a quantitative combined bisulfite restriction analysis assay and bisulfate sequencing. REIC/Dkk-3 type-a and type-b expression was examined using reverse transcriptional PCR. The relationships between the methylation and clinicopathological factors were analyzed. The rate of REIC/Dkk-3 type-a methylation ranged from 26.2 to 50.0% in the various primary tumors that were examined. REIC/Dkk-3 type-a methylation in breast cancer cells was significantly heavier than that in the other cell lines that we tested. REIC/Dkk-3 type-a methylation was inversely correlated with REIC/Dkk-3 type-a expression. There was a correlation between REIC/Dkk-3 type-a and type-b mRNA expression. REIC/Dkk-3 type-a expression was restored in MDA-MB-231 cells using 5-aza-2'-deoxycytidine treatment. We found that estrogen receptor-positive breast cancers were significantly more common among the methylated group than among the non-methylated group. REIC/Dkk-3 type-a methylation was frequently detected in a broad range of cancers and appeared to play a key role in silencing REIC/Dkk-3 type-a expression in these malignancies. No potential conflict of interest relevant to this article was reported.

Okayama University Medical School Acta Medica Okayama 0386-300X 68 1 2014 Preclinical Evaluation of MicroRNA-34b/c Delivery for Malignant Pleural Mesothelioma 23 26 EN Tsuyoshi Ueno Shinichi Toyooka Takuya Fukazawa Takafumi Kubo Junichi Soh Hiroaki Asano Takayuki Muraoka Norimitsu Tanaka Yuho Maki Kazuhiko Shien Masashi Furukawa Masakiyo Sakaguchi Hiromasa Yamamoto Kazunori Tsukuda Shinichiro Miyoshi Original Article 10.18926/AMO/52140 The microRNA-34s (miR-34s) have p53 response elements in their 5ʼ-flanking regions and demonstrate tumor-suppressive functions. In malignant pleural mesothelioma (MPM), we previously reported that expression of miR-34b and miR-34c (miR-34b/c) was frequently downregulated by methylation in MPM cell lines and primary tumors. The forced overexpression of miR-34b/c showed significant antitumor effects with the induction of apoptosis in MPM cells. In this study, we examined the in vivo antitumor effects of miR-34b/c using adenovirus vector on MPM. We subcutaneously transplanted NCI-H290, a human MPM cell line, into BALB/C mice and injected adenovirus vector expressing miR-34b/c, luciferase driven by the cytomegalovirus promoter (Ad-miR-34b/c or Ad-Luc), or PBS control into tumors over 5mm in diameter. A statistically significant growth inhibition of the tumor volume was observed in the Ad-miR-34b/c group from day 6 onward compared to the Ad-Luc group. The inhibition rate of Ad-miR-34b/c, compared to the tumor volume treated with Ad-Luc, was 58.6% on day 10 and 54.7% on day13. Our results indicate that adenovirus-mediated miR-34b/c gene therapy could be useful for the clinical treatment of MPM. No potential conflict of interest relevant to this article was reported.

Elsevier Ireland Ltd. Acta Medica Okayama 0169-5002 82 3 2013 The degree of microRNA-34b/c methylation in serum-circulating DNA is associated with malignant pleural mesothelioma 485 490 EN Takayuki Muraoka Junichi Soh Shinichi Toyooka Keisuke Aoe Nobukazu Fujimoto Shinsuke Hashida Yuho Maki Norimitsu Tanaka Kazuhiko Shien Masashi Furukawa Hiromasa Yamamoto Hiroaki Asano Kazunori Tsukuda Takumi Kishimoto Takemi Otsuki Shinichiro Miyoshi Objectives: Malignant pleural mesothelioma (MPM) is an aggressive tumor with a poor prognosis. microRNA-34b/c (miR-34b/c), which plays an important role in the pathogenesis of MPM, is frequently downregulated by DNA methylation in approximately 90% of MPM cases. In this study, we estimated the degree of miR-34b/c methylation in serum-circulating DNA using a digital methylation specific PCR assay (MSP). Materials and methods: A real-time MSP assay was performed using the SYBR Green method. The melting temperature (Tm) of each PCR product was examined using a melting curve analysis. For a digital MSP assay, 40 wells were analyzed per sample. A total of 110 serum samples from 48 MPM cases, 21 benign asbestos pleurisy (BAP) cases, and 41 healthy volunteers (HVs) were examined. Results: Positive range of Tm value for miR-34b/c methylation was defined as 77.71-78.79 degrees C which was the mean 3 standard deviations of 40 wells of a positive control. The number of miR-34b/c methylated wells was counted per sample according to this criterion. The number of miR-34b/c methylated wells in MPM cases was significantly higher than that in BAP cases (P = 0.03) or HVs (P < 0.001). Advanced MPM cases tended to have higher number of miR-34b/c methylated wells than early MPM cases. Receiver-operating characteristic (ROC) curve analysis revealed that three number of miR-34b/c methylated wells per sample was the best cut-off of positivity of MPM with a 67% of sensitivity and a 77% specificity for prediction. The area under the ROC curve was 0.77. Conclusions: Our digital MSP assay can quantify miR-34b/c methylation in serum-circulating DNA. The degree of miR-34b/c methylation in serum-circulating DNA is associated with MPM, suggesting that this approach might be useful for the establishment of a new detection system for MPM. No potential conflict of interest relevant to this article was reported.

Acta Medica Okayama 1021-335X 29 1 2013 Impact of GLUT1 and Ki-67 expression on early-stage lung adenocarcinoma diagnosed according to a new international multidisciplinary classification 133 140 EN Yuho Maki Junichi Soh Kouichi Ichimura Kazuhiko Shien Masashi Furukawa Takayuki Muraoka Norimitsu Tanaka Tsuyoshi Ueno Hiromasa Yamamoto Hiroaki Asano Kazunori Tsukuda Shinichi Toyooka Shinichiro Miyoshi High expression levels of glucose transporter isoform 1 (GLUT1) and Ki-67 are reportedly associated with malignancy-related clinicopathological factors in malignant tumors. Recently, a new histological IASLC/ATS/ERS classification for lung adenocarcinoma was proposed. In this study, we investigated the clinicopathological impact of GLUT1 and Ki-67 expression on early-stage lung adenocarcinoma classified according to the IASLC/ATS/ERS classification. One hundred and five patients with completely resected stage IA lung adenocarcinoma were retrospectively classified into two groups, a 'non-invasive type' (n=31) or an 'invasive type' (n=74), based on the IASLC/ATS/ERS classification. GLUT1 and Ki-67 expression status was evaluated using immunohistochemistry. The epidermal growth factor receptor (EGFR) and KRAS mutation status was determined using PCR-based assays. Positive GLUT1 and Ki-67 expression and EGFR and KRAS mutations were detected in 28 (27%), 33 (31%), 51 (49%) and 5 (8%) cases, respectively. Positive GLUT1 expression was significantly associated with a wild-type EGFR and mutant KRAS status. A multivariate analysis revealed that positive GLUT1 expression was independently associated with the 'invasive type'. In multivariate analyses for overall survival (OS) and disease-free survival (DFS), positive Ki-67 and GLUT1 expression was the only independent factor for a poor OS (P=0.012) and DFS (P=0.040), respectively. In addition, when stratified according to the GLUT1 and Ki-67 status, double-positive cases had the poorest DFS and OS times, compared with the other categories. Positive GLUT1 expression is associated with the invasive character of early-stage lung adenocarcinoma and with early disease relapse. Our results strongly suggest that GLUT1 and Ki-67 play important roles in acquiring biological malignant potential in early-stage lung adenocarcinoma. No potential conflict of interest relevant to this article was reported.

Elsevier Ireland Ltd. Acta Medica Okayama 0169-5002 76 1 2012 Strong anti-tumor effect of NVP-AUY922, a novel Hsp90 inhibitor, on non-small cell lung cancer 26 31 EN Tsuyoshi Ueno Kazunori Tsukuda Shinichi Toyooka Midori Ando Munenori Takaoka Junichi Soh Hiroaki Asano Yuho Maki Takayuki Muraoka Norimitsu Tanaka Kazuhiko Shien Masashi Furukawa Tomoki Yamatsuji Katsuyuki Kiura Yoshio Naomoto Shinichiro Miyoshi The anti-tumor activity of a newly developed Hsp90 inhibitor, NVP-AUY922 (AUY922), against non-small cell lung cancer (NSCLC) was examined. Twenty-one NSCLC cell lines were used, the somatic alterations of which were characterized. Cell proliferation was analyzed using a modified MTS assay. Expression of the client proteins was assessed using Western blotting. The cell cycle was analyzed using flow cytometry. The IC50 value of AUY922 for the NSCLC cell lines ranged from 5.2 to 860 nM (median, 20.4 nM). Based on previous data, cells with an IC50 of less than 50 nM were classified as sensitive cells and 19 of the 21 NSCLC cell lines were judged to be sensitive. The IC50 of five malignant pleural mesothelioma (MPM) cell lines revealed that the MPM cells had a significantly higher IC50 value (median, 89.2 nM; range, 22.2-24, 100 nM) than the NSCLC cells (p = 0.015). There was significant depletion of both the total and phosphorylated client proteins - EGFR, MET, HERZ and ART - at low drug concentrations (50-100 nM) in drug-sensitive cell lines. Cell-cycle analysis was performed for two sensitive cell lines, H1975 and H838. Following AUY922 treatment, an increase in the sub-G(0)-G(1) cell population, as well as appearance of cleaved PARP expression, indicated the induction of apoptosis. In conclusion, AUY922 was effective against most NSCLC cell lines, independent of the type of known molecular alteration, and appears to be a promising new drug for the treatment of NSCLC. (C) 2011 Elsevier Ireland Ltd. All rights reserved. No potential conflict of interest relevant to this article was reported.

Elsevier Ireland Ltd. Acta Medica Okayama 0169-5002 76 1 2012 Frequent methylation and oncogenic role of microRNA-34b/c in small-cell lung cancer 32 38 EN Norimitsu Tanaka Shinichi Toyooka Junichi Soh Takafumi Kubo Hiromasa Yamamoto Yuho Maki Takayuki Muraoka Kazuhiko Shien Masashi Furukawa Tsuyoshi Ueno Hiroaki Asano Kazunori Tsukuda Keisuke Aoe Shinichiro Miyoshi Small-cell lung cancer (SCLC) is an aggressive tumor with a dismal prognosis among primary lung cancers. MicroRNAs (miRNAs) can act as oncogenes or tumor-suppressor genes in human malignancy. The miR-34 family is comprised of tumor-suppressive miRNAs, and its reduced expression by methylation has been reported in various cancers, including non-small cell lung cancer (NSCLC). In this study, we investigated the alteration and tumor-suppressive impact of miR-34s in SCLC. The methylation of miR-34a and miR-34b/c was observed in 4 (36%) and 7 (64%) of 11 SCLC cell lines, respectively. Among the 27 SCLC clinical specimens, miR-34a and miR-34b/c were methylated in 4(15%) and 18 (67%), respectively. In contrast, 13 (28%) miR-34a methylated cases and 12 (26%) miR-34b/c methylated cases were found in 47 NSCLC primary tumors. The frequency of miR-34b/c methylation was significantly higher in SCLC than in NSCLC (p < 0.001). The expressions of miR-34s were reduced in methylated cell lines and tumors and restored after 5-aza-2'-deoxycytidine treatment, indicating that methylation was responsible for the reduced expression of miR-34s. Because the frequency of methylation was higher in miR-34b/c, we focused on miR-34b/c for a functional analysis. We examined the effect of miR-34b/c introduction on cell proliferation, migration and invasion. The transfection of miR-34b/c to two SCLC cell lines (H1048 and SBC5) resulted in the significant inhibition of cell growth, migration, and invasion, compared with control transfectants. Our results indicate that the aberrant methylation of miR-34b/c plays an important role in the pathogenesis of SCLC, implying that miR-34b/c may be a useful therapeutic target for SCLC. No potential conflict of interest relevant to this article was reported.

Elsevier Ireland Ltd. Acta Medica Okayama 0169-5002 77 1 2012 Prognostic impact of cancer stem cell-related markers in non-small cell lung cancer patients treated with induction chemoradiotherapy 162 167 EN Kazuhiko Shien Shinichi Toyooka Kouichi Ichimura Junichi Soh Masashi Furukawa Yuho Maki Takayuki Muraoka Norimitsu Tanaka Tsuyoshi Ueno Hiroaki Asano Kazunori Tsukuda Masaomi Yamane Takahiro Oto Katsuyuki Kiura Shinichiro Miyoshi The expression of several cancer stem cell (CSC)-related markers has been confirmed in non-small cell lung cancer (NSCLC). The aim of this study was to clarify the clinical role of CSC-related markers in patients with NSCLC undergoing induction chemoradiotherapy (CRT). Fifty patients with clinically diagnosed N2 or N3 NSCLC who underwent induction CRT with docetaxel and cisplatin concurrently with thoracic radiation followed by surgery were examined in this study. The expressions of CSC related markers (CD133, ALDH1, ABCG2, and Bmi-1) were examined using immunohistochemical staining in surgically resected specimens. Among the 50 patients, 20 patients had no residual tumor cells in the resected specimen when examined pathologically; CSC-related marker expressions and their correlation to survival were evaluated in the other 30 patients. After a median follow-up period of 72 months, the 5-year overall survival rate of the patients with CD133-positive or ALDH1-positive specimens was significantly worse than that of the patients with both CD133-negative and ALDH1-negative expressions (449% vs. 90.0%, respectively; P=0.042). In a multivariate analysis. CD133 and ALDH1 negativity (P=0.047) and cN2-3 single station metastasis (P=0.03) were significant independent prognostic factors for prolonged survival. The expressions of CSC-related markers after CRT were significantly correlated with a poor prognosis in patients with NSCLC. The development of therapeutic strategies including adjuvant therapy that take CSC-related marker positivity into consideration is likely to be a key factor in further improvements of the prognosis of patients undergoing trimodality therapy. No potential conflict of interest relevant to this article was reported.

Okayama University Medical School Acta Medica Okayama 0386-300X 67 1 2013 Silenced Expression of NFKBIA in Lung Adenocarcinoma Patients with a Never-smoking History 19 24 EN Masashi Furukawa Junichi Soh Hiromasa Yamamoto Kouichi Ichimura Kazuhiko Shien Yuho Maki Takayuki Muraoka Norimitsu Tanaka Tsuyoshi Ueno Hiroaki Asano Kazunori Tsukuda Shinichi Toyooka Shinichiro Miyoshi Original Article 10.18926/AMO/49253 Nuclear factor of κ-light polypeptide gene enhancer in B cells inhibitor α (NFKBIA), which is a tumor suppressor gene, was found to be silenced in lung adenocarcinomas. We examined NFKBIA expression, mutations in the EGFR and K-ras genes, and EML4-ALK fusion in 101 resected lung adenocarcinoma samples from never-smokers. NFKBIA expression was evaluated using immunohistochemistry. NFKBIA expression was negative in 16 of the 101 samples (15.8%). EGFR and K-ras mutations and EML4-ALK fusion were detected in 61 (60.5%), 1 (1.0%), and 2 (2.0%) of the 101 samples, respectively, in a completely mutually exclusive manner. Negative NFKBIA expression was observed significantly more frequently among the tumors with none of the three genetic alterations compared to those with such alterations (p＝0.009). In addition, negative NFKBIA expression was significantly more frequent among the EGFR-wild type samples compared to the EGFR-mutant samples (p＝0.013). In conclusion, NFKBIA expression was silenced in adenocarcinomas without EGFR/K-ras mutations or EML4-ALK fusion, suggesting that the silencing of NFKBIA may play an important role in the carcinogenesis of adenocarcinomas independent of EGFR/K-ras mutations or EML4-ALK fusion. No potential conflict of interest relevant to this article was reported.

Okayama University Medical School Acta Medica Okayama 0386-300X 66 4 2012 Ectopic Cervical Thymoma:A Case Report with 18F-fluorodeoxyglucose Positron Emission Tomography Findings 357 361 EN Kazuhiko Shien Tadahiko Shien Junichi Soh Hirokuni Ikeda Tomohiro Nogami Naruto Taira Hiroyoshi Doihara Shinichiro Miyoshi Case Report 10.18926/AMO/48691 Ectopic thymoma is considered to arise from ectopic thymus tissue deposited as a result of the abnormal mislocalization of thymus tissue during the embryonic stage. An 86-year-old man visited our hospital with chief complaints of hoarseness and a mass in his anterior neck. A preoperative needle biopsy of the mass did not yield a definitive diagnosis. A positron emission tomography (PET) study revealed heterogeneous accumulation of ¹⁸F-fluorodeoxyglucose (FDG) in the tumor. The tumor, affecting the left sternocleidomastoid muscle, the recurrent laryngeal nerve, the internal carotid vein, and the brachiocephalic vein, was resected using a combination of a collar incision in the neck and a median incision in the sternum. Immunohistochemically, the tumor was diagnosed as an ectopic thymoma of the neck. To date, only a few cases of ectopic thymoma presenting with FDG accumulation have been reported. Our experience indicates that ectopic thymoma should be kept in mind during the differential diagnosis of neck tumors with FDG accumulation appearing on PET images. No potential conflict of interest relevant to this article was reported.

Okayama University Medical School Acta Medica Okayama 0386-300X 65 3 2011 Aberrant Methylation of p21 Gene in Lung Cancer and Malignant Pleural Mesothelioma 179 184 EN Hirotake Teramen Kazunori Tsukuda Norimitsu Tanaka Tsuyoshi Ueno Takafumi Kubo Midori Ando Junichi Soh Hiroaki Asano Harvery I. Pass Shinichi Toyooka Shinichiro Miyoshi Original Article 10.18926/AMO/46629 Suppression of p21 has been implicated in the genesis and progression of many human malignancies. DNA methylation is an important mechanism of gene silencing in human malignancies. In this study, we examined the expression status and aberrant methylaion of p21 in lung cancers and malignant pleural mesotheliomas (MPM). We used 12 small cell lung cancer (SCLC) cell lines, 13 non-small cell lung cancer (NSCLC) cell lines, 50 primary NSCLCs, 6 MPM cell lines and 10 primary MPMs. The expression and methylation of p21 was examined by reverse transcription-PCR (RT-PCR), Western blotting and methylation-specific PCR (MSP) assay. Loss of p21 protein expression was observed in 7 SCLC cell lines (58.3%), 5 NSCLC cell lines (38.5%) and 3 MPM cell lines (50%) while mRNA expression was lost in 2 SCLC cell lines (16.7%), 2 NSCLC cell lines (15.4%) and none of the MPM cell lines. Aberrant methylation of p21 was found in 8.3% of SCLC cell lines, 30.2% of NSCLCs and 6.3% of MPMs. Among primary NSCLCs, methylation in adenocarcinomas was significantly more frequent than in squamous cell carcinomas. Loss of p21 expression was frequently observed in lung cancers and MPMs and aberrant methylation was one of the mechanisms of suppression of p21, especially in NSCLCs. No potential conflict of interest relevant to this article was reported.

岡山医学会 Acta Medica Okayama 0030-1558 120 3 2008 EGFR 変異と Uracil-Tegafur による肺腺癌術後補助療法の関連性についての検討 265 269 EN Hiroshi Suehisa Shinichi Toyooka Katsuyuki Hotta Akiko Uchida Junichi Soh Yoshiro Fujiwara Keitaro Matsuo Mamoru Ouchida Minoru Takata Katsuyuki Kiura Hiroshi Date No potential conflict of interest relevant to this article was reported.

Acta Medica Okayama 2007 Usefulness of EGFR mutation screening in pleural fluid to predict the clinical outcome of gefitinib treated patients with lung cancer EN Jun-ichi So The importance of epidermal growth factor receptor (EGFR) gene mutation has been recognized in nonsmall cell lung cancer (NSCLC), requiring the standardization of mutation screening system including the kind of samples. Here, we examined the EGFR mutation status in 61 pleural fluid samples from NSCLC cases using direct sequencing, nonenriched PCR, mutant-enriched PCR and peptide nucleic acid-locked nucleic acid (PNA-LNA) PCR clamp assay. The mutant-enriched PCR assay detected 16 mutant cases. Among them, the nonenriched PCR assay failed to detect 3 mutant cases. Regarding the discrepancy between mutant-enriched PCR and PNA-LNA PCR clamp assays, 3 cases of exon19-deletions were detected only by mutant-enriched PCR assay and no difference at the L858R mutation. There was no difference in results between direct sequencing and nonenriched PCR assay. We also correlated the EGFR mutation with clinical outcome of gefitinib-treated 29 cases. EGFR mutations were present in 10 cases, revealing 7 partial response and 3 no change (NC). In EGFR wild-type cases, 10 revealed NC and 9 progressive disease. The responders were significantly more frequent among the EGFR mutant cases than among the wild-type (p < 0.0001). Overall survival (p = 0.0092) and progression-free survival (p = 0.018) were significantly longer among the EGFR mutant cases than among the wild-type. In summary, we evaluated the utility of EGFR mutation screening in pleural fluid using 4 assays that showed some discrepancies arising from the designs of the assays. As clinical importance, the EGFR mutation status in pleural fluid can be a biomarker for the favorable outcome of gefitinib-treated NSCLC cases. No potential conflict of interest relevant to this article was reported.