Kentaro Kitano

Genome structure-based screening identified epigenetically silenced microRNA associated with invasiveness in non-small-cell lung cancer†

MicroRNA (miRNA) expression is frequently altered in human cancers. To search for epigenetically silenced miRNAs in non‐small‐cell lung cancer (NSCLC), we mapped human miRNAs on autosomal chromosomes and selected 55 miRNAs in silico. We treated six NSCLC cell lines with the DNA methylation inhibitor 5‐aza‐2′‐deoxycytidine (5‐aza‐CdR) and determined the expressions of the 55 miRNAs. Fourteen miRNAs were decreased in the cancer cell lines and were induced after 5‐aza‐CdR treatment. After a detailed DNA methylation analysis, we found that mir‐34b and mir‐126 were silenced by DNA methylation. Mir‐34b was silenced by the DNA methylation of its own promoter, whereas mir‐126 was silenced by the DNA methylation of its host gene, EGFL7. A chromatin immunoprecipitation assay revealed H3K9me2 and H3K9me3 in mir‐34b and EGFL7, and H3K27me3 in EGFL7. The overexpression of mir‐34b and mir‐126 decreased the expression of c‐Met and Crk, respectively. The 5‐aza‐CdR treatment of lung cancer cell line resulted in increased mir‐34b expression and decreased c‐Met protein. We next analyzed the DNA methylation status of these miRNAs using 99 primary NSCLCs. Mir‐34b and mir‐126 were methylated in 41 and 7% of all the cases, respectively. The DNA methylation of mir‐34b was not associated with c‐Met expression determined by immunohistochemistry, but both mir‐34b methylation (p = 0.007) and c‐Met expression (p = 0.005) were significantly associated with lymphatic invasion in a multivariate analysis. The DNA methylation of mir‐34b can be used as a biomarker for an invasive phenotype of lung cancer.

CpG island methylation of microRNAs is associated with tumor size and recurrence of non-small-cell lung cancer

We investigated whether the CpG island methylation of certain microRNAs was associated with the clinicopathological features and the prognosis of non‐small‐cell lung cancer. The methylation of mir‐152, ‐9‐3, ‐124‐1, ‐124‐2, and ‐124‐3 was analyzed in 96 non‐small‐cell lung cancer specimens using a combined bisulfite restriction analysis. The median observation period was 49.5 months. The methylation of mir‐9‐3, ‐124‐2, and ‐124‐3 was individually associated with an advanced T factor independent of age, sex, and smoking habit. Moreover, the methylation of multiple microRNA loci was associated with a poorer progression‐free survival in a univariate analysis. Our result enlightens the accumulation of aberrant DNA methylation which occurs in concordance with the tumor progression. (Cancer Sci 2011; 102: 2126–2131)

Outcome and survival analysis of pulmonary metastasectomy for hepatocellular carcinoma

OBJECTIVE The lung is the most common site for extrahepatic metastasis from hepatocellular carcinoma (HCC). We previously reported in a series of 20 patients that pulmonary metastasectomy for HCC is feasible in selected patients. The objective of this study was to re-evaluate the long-term outcomes and prognostic factors with an additional 25 patients. METHODS We retrospectively analyzed the records of 45 consecutive patients who underwent pulmonary metastasectomy due to HCC at our institution between 1990 and 2010. RESULTS Thirty-nine patients underwent hepatectomy or liver transplantation, whereas six patients underwent locoregional therapy for primary liver lesions. Twenty-seven patients died during a median 17.6-month follow-up period. The 2-year disease-free survival (DFS) was 19.5%. The 5-year overall survival (OS) was 40.9%. History of recurrence and serum des-gamma-carboxy prothrombin (DCP) level >40 mAU ml(-1) at initial pulmonary resection were unfavorably associated with OS in univariate analysis. CONCLUSIONS Pulmonary metastasectomy for HCC in selected patients resulted in relatively good outcomes with regard to OS. History of recurrence and serum DCP levels were shown to be candidates of prognostic factors for OS.

Survival after extended thymectomy for thymoma.

OBJECTIVES Thymoma is a relatively rare tumour and is uniquely associated with autoimmune diseases such as myasthenia gravis (MG). However, the factors involved in the prognosis of thymoma remain under discussion. METHODS We retrospectively reviewed 162 patients who underwent extended thymectomy via median sternotomy for thymoma at our institute from 1976 to 2009. The histological subtype was classified according to the World Health Organization (WHO) histological classification system. Survival analysis was performed using the Kaplan-Meier method and the Cox proportional hazards model. RESULTS Tumours comprised 7 Type A tumours, 38 Type AB, 49 Type B1, 45 Type B2 and 23 Type B3. Various types of autoimmune diseases were comorbid in 66 patients. The median follow-up period was 94 months, and 14 patients experienced recurrence. Seven patients died of recurrent tumour, and 18 patients died of causes other than thymoma. The 10-year overall survival was 85.7%, and the 10-year disease-free survival (DFS) was 76.8%. The 10-year DFS was 62.5% for Type A, 86.3% for Type AB, 91.5% for Type B1, 77.1% for Type B2 and 26.3% for Type B3. In multivariate analysis, age, Type B3 and MG were determined as prognostic factors for survival. On the other hand, Masaokas stage did not influence survival. CONCLUSIONS Type B3 classified by the WHO histological classification system is a poor prognostic factor for survival of thymoma after extended thymectomy. Association with MG is possibly an indicator of poor survival. Age is an independent prognostic factor, suggesting favourable prognosis of thymoma after surgical treatment. Considering thymoma is a rare tumour, it would be necessary to build a multi-institutional database as soon as possible.

Resection of solitary pulmonary lesion is beneficial to patients with a history of malignancy.

BACKGROUND Solitary pulmonary lesion poses a diagnostic challenge, especially in patients with a history of malignancy. The purpose of this study was to evaluate the characteristics of solitary pulmonary lesions and the outcome of surgical resection. METHODS We retrospectively analyzed 243 patients with a history of cancer who underwent surgery for new-found solitary pulmonary lesion between January 1998 and December 2007. RESULTS The diagnosis was primary lung cancer in 92 patients, metastasis in 133, and benign lesions in 18. The 5-year survival rate was 67.9% in all patients, 74.6% in those with primary lung cancer, 62.8% in those with metastasis, and 79.9% in those with benign lesions (p = 0.56). In metastasis patients, history of extrapulmonary recurrence and larger diameter lesion were risk factors for recurrence by multivariate analysis. History of cancers other than colorectal and bone and soft tissue sarcoma and shorter disease-free interval were indicators of poor prognosis. Pathologic stage was the only indicator of prognosis for primary lung cancer, and none of the factors concerning antecedent cancer influenced prognosis. CONCLUSIONS Surgical resection of solitary pulmonary lesion is essential in patients with a history of cancer because substantial numbers of benign lesions are included. In the case of malignancy, metastasectomy had a life-prolonging effect for selected patients, and prognosis of primary lung cancer was no worse than for the general population if treated appropriately. It is important not to hesitate to take a surgical approach for a diagnosis and to treat with standard therapy for primary lung cancer.

Analysis for the Combination Expression of CK20, FABP1 and MUC2 is Sensitive for the Prediction of Peritoneal Recurrence in Gastric Cancer

Prediction of peritoneal recurrence in gastric cancer patients is important for application of adjuvant chemotherapy. After surgery, occasional patients have peritoneal recurrence despite negative cytology of the peritoneal washings. Thus, molecular detection of a subliminal number of cancer cells in peritoneal washings may overcome the sensitivity limitation of conventional cytology. In this study, expressions of five specific marker genes, namely, TFF1, TFF2, CK20, FABP1 and MUC2, were evaluated for their usefulness as markers of micro-dissemination. It was found that reverse transcriptase-polymerase chain reaction for these five genes yielded results highly specific for the depth of invasion and disease stage. Furthermore, the expression of CK20, FABP1 and MUC2 was a reliable prognostic indicator of peritoneal metastasis. Our results suggest that evaluation of the expression of CK20, FABP1 and MUC2 in peritoneal washings is a useful tool for identifying patients at high risk of peritoneal recurrence who may need adjuvant chemotherapy.

Invasive thymoma disseminated into the pleural cavity: mid-term results of surgical resection

OBJECTIVES The optimal strategy for pleural dissemination of advanced thymoma remains controversial, while a potential benefit from macroscopic clearance of disseminations has been reported. In this study, we review our mid-term results of surgical resection of pleural disseminations of invasive thymoma. METHODS Data from patients with pleural dissemination synchronously or metachronously to primary invasive thymoma who underwent surgical resection from 1991 to 2012 at our institute were retrospectively reviewed. RESULTS Of 136 thymoma patients who underwent surgery during the study period, 13 consecutive patients with pleural dissemination (synchronous: 7, metachronous: 6) with a median age of 49 years (range: 27-78 years) at the time of dissemination resection were identified. No patients presented with haematogenous metastases. Operative procedures included the thorough resection of visible disseminated nodules in 11 patients and extrapleural pneumonectomy (EPP) in 2 patients. The median number of resected nodules was 6 (range: 1-52). The median follow-up was 948 days (range: 38-4025 days). One patient died of postoperative bleeding, but there were no tumour-related deaths during the study period. Pleural recurrence was found in 9 cases, including 2 EPP cases, and among them, 3 underwent repeated resection. The overall survival and the recurrence-free survival ratio at 5 years was 92.3 and 33.3%, respectively. Five patients, including 2 repeated resection cases, remained tumour-free at the final observation. CONCLUSIONS Resection of pleural dissemination of invasive thymoma can be performed in selected patients and may offer optimal local control as part of a multimodal strategy.

Rapid Cancer Fluorescence Imaging Using A γ-Glutamyltranspeptidase-Specific Probe For Primary Lung Cancer

BACKGROUND: We set out to examine the activity of γ-glutamyltranspeptidase (GGT) in lung cancer and the validity of γ-glutamyl hydroxymethyl rhodamine green (gGlu-HMRG) for intraoperative imaging of primary lung cancer. METHODS: GGT activities and mRNA expression levels of GGT1 (one of the GGT subtypes) in five human lung cancer cell lines were examined by fluorescence imaging and quantitative reverse transcription polymerase chain reaction. In vivo imaging of an orthotopic A549 xenograft model in nude mice was performed to confirm its applicability to intraoperative imaging. Furthermore, ex vivo imaging of 73 specimens from lung cancer patients were performed and analyzed to calculate the sensitivity/specificity of gGlu-HMRG for lung cancer diagnosis. RESULTS: GGT activities and mRNA expression levels of GGT1 are diverse depending on cell type; A549, H441, and H460 showed relatively high GGT activities and expression levels, whereas H82 and H226 showed lower values. In the in vivo mouse model study, tiny pleural dissemination and hilar/mediastinal lymph node metastasis (less than 1 mm in diameter) were clearly detected 15 minutes after topical application of gGlu-HMRG. In the ex vivo study of specimens from patients, the sensitivity and specificity of gGlu-HMRG were calculated to be 43.8% (32/73) and 84.9% (62/73), respectively. When limited to female patients, never smokers, and adenocarcinomas, these values were 78.9% (15/19) and 73.7% (14/19), respectively. CONCLUSIONS: Although GGT activity of lung cancer cells vary, gGlu-HMRG can serve as an intraoperative imaging tool to detect small foci of lung cancer when such cells have sufficient GGT activity.

Results of Bony Chest Wall Reconstruction with Expanded Polytetrafluoroethylene Soft Tissue Patch

PURPOSE The early and long-term outcomes of bony chest wall reconstruction with expanded polytetrafluoroethylene (Gore-Tex) soft tissue patch remain a concern. No clinical study has reported the shrinkage of Gore-Tex following reconstruction to date. METHODS Thirty-seven patients who underwent bony chest wall reconstruction from 1994 to 2012 were retrospectively reviewed. Postoperative chest computed tomography images of 17 patients were examined, and shrinkage of reconstruction materials was measured and compared. RESULTS Gore-Tex was used for reconstruction in 18 patients, autologous materials were used in 14, Marlex mesh was used in four, and Medifit felt was used in one. No surgery-related deaths were observed. Twenty patients experienced early postoperative complications. Four patients experienced local infection. One patient with Marlex-mesh experienced empyema 33 days postoperatively. Chest drainage time in the Gore-Tex patients was significantly lower than in patients with other types of prosthetic reconstruction. No dislocation or dehiscence was found. Shrinkage of Gore-Tex was absent in 4 patients and acceptable in seven patients. No granulation formation was evident around the Gore-Tex, No significant difference in shrinkage was seen between the different materials used. CONCLUSION Chest wall reconstruction with Gore-Tex was feasible with favorable early and long-term results.

Prognostic impact of the current Japanese nodal classification on outcomes in resected non-small cell lung cancer.

BACKGROUND The prognosis of N2 non-small cell lung cancer (NSCLC) has been reported to be heterogeneous. The recently revised Japanese nodal classification subcategorizes N2 disease according to the tumor-bearing lobe. We evaluated the prognostic impact of the Japanese nodal classification and its ability to define favorable N2 disease in resected NSCLC. METHODS A total of 496 patients with NSCLC who underwent lobectomy with systematic lymph node dissection between 1998 and 2009 were analyzed retrospectively. N2 status was subdivided into N2a-1 and N2a-2, according to the Japanese nodal classification. Overall survival (OS), disease-free survival (DFS), and clinicopathologic features were compared between the two groups. RESULTS There were 67 cases with N2 disease. The outcome of resected N2a-2 NSCLC was far poorer than that of the N2a-1 group (5-year OS, 28% vs 62%, P < .001; 5-year DFS, 5% vs 35%, P < .001). Multivariate analysis revealed that pathologic N2a-2 was an independent prognostic factor (hazard ratio, 2.86; P < .05). Patients in the N2a-2 group showed more involved nodes and stations, less skip metastasis, and more locoregional recurrence than did patients in the N2a-1 group. The outcome of the N2a-1 group was satisfactory, and there was no significant difference in OS and DFS between N1 and N2a-1. CONCLUSIONS The Japanese nodal classification is able to identify a favorable N2 subgroup in resected NSCLC. Nodal staging by the Japanese system should be considered when a clinical trial of N2 disease is designed.