Katsunobu Kawahara

A Polycistronic MicroRNA Cluster, miR-17-92, Is Overexpressed in Human Lung Cancers and Enhances Cell Proliferation

MicroRNAs (miRNAs) are small noncoding RNAs, thought to be involved in physiologic and developmental processes by negatively regulating expression of target genes. We have previously reported frequent down-regulation of the let-7 miRNA family in lung cancers and, in the present study, assessed alteration in a panel of 19 lung cancer cell lines. As a result, we found for the first time that the miR-17-92 cluster, which comprises seven miRNAs and resides in intron 3 of the C13orf25 gene at 13q31.3, is markedly overexpressed in lung cancers, especially with small-cell lung cancer histology. Southern blot analysis revealed the presence of increased gene copy numbers of the miRNA cluster in a fraction of lung cancer cell lines with overexpression. In addition, we were able to show predominant localization of C13orf25 transcripts within the nucleus and introduction of the expression construct of the miR-17-92 cluster, but not the putative open reading frame of C13orf25, enhancing lung cancer cell growth. These findings clearly suggest that marked overexpression of the miR-17-92 cluster with occasional gene amplification may play a role in the development of lung cancers, especially in their most aggressive form, small-cell lung cancer, and that the C13orf25 gene may well be serving as a vehicle in this regard.

Video-assisted thoracoscopic esophagectomy for esophageal cancer

AbstractBackground: The Ivor-Lewis procedure is a radical, invasive, and effective procedure for the resection of most esophageal cancers. To minimize invasiveness, we performed thoracoscopic and video-assisted esophagectomy and mediastinal dissection for esophageal cancer. Methods: From November 1995 to June 1997, 23 patients with intrathoracic esophageal cancer, excluding T4 cancers, underwent thoracoscopic and video-assisted esophagectomy. Bilateral cervical dissections were performed as well as preparation of the gastric tube and transhiatal dissection of the lower esophagus. The cervical esophagus was cut using a stapler knife, and esophageal reconstruction was performed through the retrosternal route or anterior chest wall. Next, thoracoscopic mediastinal dissection and esophagectomy were performed. Results: The mean volume of blood loss was 163 ± 122 ml; mean thoracoscopic surgery duration, 111 ± 24 min; mean postoperative day for patients to start eating, 8 ± 3 days; and mean hospital stay, 26 ± 8 days. No patient developed systemic inflammatory response syndrome postoperatively. Tracheal injury occurred and was repaired during the thoracoscopic approach in one patient. No patients died within 30 days after surgery. Postoperative complications included transient recurrent nerve palsy in five patients, pulmonary secretion retention requiring tracheotomy in two, and chylothorax in one. Five patients died of cancer recurrence within 1 year of surgery. Conclusions: Our surgical experience with thoracoscopic and video-assisted esophagectomy indicate that it is a feasible and useful procedure.

Genome-wide analysis of DNA copy number alterations and gene expression in gastric cancer.

Genomic copy number aberrations (CNAs) are believed to play a major role in the development and progression of human cancers. Although many CNAs have been reported in gastric cancer, their genome‐wide transcriptional consequences are poorly understood. In this study, to reveal the impact of CNAs on genome‐wide expression in gastric cancer, we analysed 30 cases of gastric cancers for their CNAs by array comparative genomic hybridization (array CGH) and 24 of these 30 cases for their expression profiles by oligonucleotide‐expression microarray. We found that with the application of laser microdissection, most CNAs were detected at higher frequency than in previous studies. Notably, gain at 20q13 was detected in almost all cases (97%), suggesting that this may play an important role in the pathogenesis of gastric cancer. By comparing the array CGH data with expression profiles of the same samples, we showed that both genomic amplification and deletion strongly influence the expression of genes in altered genomic regions. Furthermore, we identified 125 candidate genes, consisting of 114 up‐regulated genes located in recurrent regions (>10%) of amplification and 11 down‐regulated genes located in recurrent regions of deletion. Up‐regulation of several candidate genes, such as CDC6, SEC61G, ANP32E, BYSL and FDFT1, was confirmed by immunohistochemistry. Interestingly, some candidate genes were localized at genomic loci adjacent to well‐known genes such as EGFR, ERBB2 and SMAD4, and concordantly deregulated by genomic alterations. Based on these results, we propose that our list of candidate genes may contain novel genes involved in the pathogenesis of advanced gastric cancer. Copyright

Genetic alterations in 102 primary gastric cancers by comparative genomic hybridization: gain of 20q and loss of 18q are associated with tumor progression.

Gastric cancer is one of the most common cancers. Molecular events in the carcinogenesis of gastric cancer remain, however, largely undefined. We investigated changes in DNA copy number in 102 gastric cancers by CGH. We found changes in DNA copy number in all cases, with frequent (≧30% of patients) gains at 20q, 8q, 20p, 7q, 17q, 5p, and 13q. Frequent (≧20%) losses were found at 19p, 18q, 5q, 21q, 4p, 4q, 15q, and 17p. The mean number of total alterations was significantly lower in grade 3 and scirrhous-type carcinomas (10.81 in grade 3 vs 13.98 in grade 1 and grade 2, 9.31 in scirrhous-type vs 13.18 in medullary- and intermediate-type). The mean number of losses and total alterations were higher in tumors at pT2, pT3 and pT4 (4.68 and 12.77 in pT2, pT3, and pT4 vs 2.55 and 9.22 in pT1). The mean number of losses was higher in carcinomas with lymph node metastasis (4.83). The mean number of gains and total alterations were higher in carcinomas with venous invasion (8.44 and 13.28). Several chromosomal alterations were linked in a statistically significant manner to specific clinicopathological parameters. Gain of 17q, 20p, and 20q and loss of 4p were associated with the pattern of the cancer–stroma relationship; loss of 18q was associated with pT category; gain of 5p was associated with pN category; loss of 4q and loss of 21q were associated with lymphatic invasion; gain of 7p and loss of 4q and 18q were associated with venous invasion; and loss of 18q was associated with pathological stage. These data suggest that gain of 20q and loss of 18q might play an important role in the development and progression of gastric cancer. Moreover, some genes on 20q and 18q might be target genes of gastric cancer.

The number of lymph node metastases influences survival in esophageal cancer

Background and Objectives: Lymph node involvement adversely affects the survival of patients with esophageal cancer. We retrospectively investigated whether the number of involved lymph nodes and the degree of lymph node dissection affect survival.

E-cadherin expression in patients with esophageal squamous cell carcinoma: promoter hypermethylation, Snail overexpression, and clinicopathologic implications.

Hypermethylation in the E-cadherin promoter region and expression of the transcription factor Snail were analyzed in 41 cases of esophageal squamous cell carcinoma (ESCC) and paired normal squamous epithelium by methylation-specific polymerase chain reaction (PCR) and reverse transcription-polymerase chain reaction (RT-PCR) to clarify the mechanism regulating E-cadherin deletion; 93 cases of ESCC were analyzed immunohistochemically to determine the clinicopathologic impact of E-cadherin deletion. Hypermethylation of the E-cadherin promoter and Snail overexpression were detected in 25 cases (61%) by methylation-specific PCR and 34 cases (83%) by RT-PCR, respectively. Reduced E-cadherin expression, observed immunohistochemically in 55 cases (59%), correlated with hypermethylation (P = .0011) but not Snail overexpression (P = .685). Hypermethylation and Snail overexpression correlated significantly with E-cadherin deletion (P = .0018). Snail overexpression was unrelated to clinicopathologic factors. Reduced E-cadherin expression correlated with tumor invasion (P = .019) and vascular invasion (P = .052) but not other factors. E-cadherin deletion had prognostic impact in univariate (P = .023) and multivariate (P = .034) analyses. E-cadherin deletion was regulated by hypermethylation and Snail expression. Examination of reduced E-cadherin expression is important for assessing biologic behavior, including clinical outcome, in patients with ESCC.

Video-assisted thoracoscopic indocyanine green fluorescence imaging system shows sentinel lymph nodes in non-small-cell lung cancer.

OBJECTIVE The primary objective was to assess the feasibility and accuracy of intraoperative sentinel lymph node mapping by using a video-assisted thoracoscopic indocyanine green fluorescence imaging system in patients with clinical stage I non-small-cell lung cancer. METHODS Thirty-one patients who underwent operation between January 2009 and September 2009 were investigated for sentinel node biopsy. Indocyanine green fluorescence imaging was applied by an infrared light charge-coupled device, and sentinel nodes were identified intraoperatively and dissected. Histologic examination by hematoxylin-eosin staining was used to evaluate metastases. RESULTS Sentinel lymph nodes were identified by segmentectomy in 11 of 14 patients (78.5%) and by lobectomy in 14 of 17 patients (82.4%). The total identification rate was 80.7% (25/31 patients), the false-negative rate was 0% (0/24 patients), and the overall accuracy rate was 80.7% (25/31 patients). CONCLUSION Video-assisted thoracoscopic indocyanine green fluorescence image-guided surgery is feasible for sentinel node biopsy and may be a powerful tool to eliminate unnecessary lymph node dissection in patients with lung cancer.

Aberrant methylation of DPYD promoter, DPYD expression, and cellular sensitivity to 5-fluorouracil in cancer cells.

Purpose: Dihydropyrimidine dehydrogenase (DPD), the initial rate-limiting enzyme in the degradation of 5-fluorouracil (5-FU), is known to be a principal factor in clinical responses to the anticancer agent 5-FU, and various reports have clearly demonstrated that DPD activity is closely correlated to mRNA levels. However, the regulatory mechanisms of DPD gene (DPYD) expression remain unclear. In this study, the regulatory mechanisms have been intensively studied. Experimental Design and Results: A subcloned 3.0-kb fragment of the 5′ region of DPYD contains a total of 60 CpG sites, suggesting that methylation status may affect the repression of DPYD. The clone showed various promoter activities that were largely correlated with mRNA levels in most cell lines, except HSC3 and HepG2. Bisulfite sequencing analysis revealed that various CpG sites around the transcription start site were abnormally methylated in cells with low DPYD expression: Reversal of hypermethylation by 5-azacytidine treatment significantly increased DPYD expression in HSC3 and HepG2 cells that showed strong promoter activity. In HepG2, in vitro methylation of the DPYD promoter directly decreased promoter activity, and 5-azacytidine treatment restored higher DPYD expression in a dose- and time-dependent manner, along with decreased sensitivity to 5-FU. Conclusions: We found that DPD activity was controlled, at least in part, at the transcription level of DPYD and that aberrant methylation of the DPYD promoter region acted as one of the repressors of DPYD expression and affected sensitivity to 5-FU in cancer cells. Our new results could lead to a more precise understanding of the molecular basis of 5-FU response.

Management of anastomotic complications after sleeve lobectomy for lung cancer

One hundred twelve patients (102 male and 10 female) underwent sleeve lobectomy for lung cancer from January 1969 to December 1991. Bronchopleural fistula occurred in 6 (5.6%), bronchovascular fistula in 2 (1.8%), pulmonary arterial occlusion in 2 (1.9%), anastomotic stricture or stenosis in 7 (6.3%), and local recurrence in 7 patients (6.3%). Early repair of bronchopleural fistula combined with an omentopexy achieved permanent closure of the fistula. Two patients who underwent a completion pneumonectomy for a pulmonary arterial occlusion died of respiratory failure. Two patients experienced uncontrollable bleeding into the bronchial tree through a bronchovascular fistula and sudden death. Completion pneumonectomy is indicated for a stricture due to scar formation. If pneumonectomy is precluded by poor pulmonary reserve, endoscopic excision using biopsy forceps is an alternative. Endoscopic resection is the treatment of choice for suture granulomas. Complications associated with bronchial or vascular anastomoses are serious and frequently fatal.

Video-assisted thoracoscopic surgery (VATS) segmentectomy for small peripheral lung cancer tumors: intermediate results

BackgroundWe investigated the feasibility and suitability of video-assisted thoracoscopic surgery (VATS) segmentectomy for curing selected non–small cell lung cancer (NSCLC) with this less invasive technique.MethodsWe performed VATS segmentectomy for small (<20 mm) peripherally located tumors and pathologically confirmed lobar lymph node–negative disease by frozen-section examination during surgery. Of the 34 patients who underwent this limited resection, 22 were treated with complete hilar and mediastinal lymph node dissection (intentional group), whereas 12 patients who were deemed to be high risk in their toleration for lobectomy underwent VATS segmentectomy with incomplete hilar and mediastinal lymph node dissection (compromised group). The surgical and clinical parameters were evaluated and compared with those of segmentectomy under standard thoracotomy to evaluate the technical feasibility of VATS segmentectomy.ResultsWe found that VATS segmentectomy could be performed safely with a nil mortality rate and acceptably low morbidity. The mean period of observation was relatively short at 656.7 ± 572.1 and 783.4 ± 535.8 days in the intentional and compromised groups, respectively. At the time of writing, all intentional patients remain alive and free of recurrence. There were two cases of non–cancer-related death in the compromised group. Clinical data indicated that VATS segmentectomy caused the same number or fewer surgical insults compared with segmentectomy under standard thoracotomy.ConclusionsThe present results are intermediate only; the rate of long-term survival and the advantages of the less invasive procedure still need further investigation. Nevertheless, we believe that VATS segmentectomy with complete lymph node dissection is a reasonable treatment option for selected patients with small peripheral NSCLC.