Akira Sakurada

A prospective trial of systematic nodal dissection for lung cancer by video-assisted thoracic surgery: can it be perfect?

BACKGROUNDnThere have been no reports evaluating the completeness of systematic nodal dissection with video-assisted thoracic surgery (VATS). In order to elucidate the completeness of the dissection, we have conducted a prospective trial with patients having primary lung cancer.nnnMETHODSnPatients with clinical stage I lung cancer were the candidates for this study. Thoracotomy was performed with a small skin incision of 7 cm to 8 cm in length. Through these small wounds and two trocars, pulmonary resection was performed and then hilar and mediastinal lymph nodes were dissected. After that, a standard thoracotomy was carried out by another surgeon to complete systematic nodal dissection.nnnRESULTSnVideo-assisted thoracic surgery lobectomy with lymph node dissection was accomplished in 17 right lung cancer patients and 12 left lung cancer patients. On the right side, the average numbers of resected lymph nodes by VATS and remnant lymph nodes were 40.3 and 1.2, respectively. The average weights of dissected tissues by VATS and remnant tissues were 10.0 g and 0.2 g, respectively. On the left side, there were 37.1 and 1.2 lymph nodes and 8.3 g and 0.2 g of weight of dissected tissues. No nodal involvement was observed in the remnant lymph nodes.nnnCONCLUSIONSnThe lymph node dissection with VATS was technically feasible and the remnant (missed by VATS) lymph nodes and tissues were 2% to 3%, which seems acceptable for clinical stage I lung cancer.

Microarray analysis of promoter methylation in lung cancers

AbstractAberrant DNA methylation is an important event in carcinogenesis. Of the various regions of a gene that can be methylated in cancers, the promoter is the most important for the regulation of gene expression. Here, we describe a microarray analysis of DNA methylation in the promoter regions of genes using a newly developed promoter-associated methylated DNA amplification DNA chip (PMAD). For each sample, methylated Hpa II-resistant DNA fragments and Msp I-cleaved (unmethylated + methylated) DNA fragments were amplified and labeled with Cy3 and Cy5 respectively, then hybridized to a microarray containing the promoters of 288 cancer-related genes. Signals from Hpa II-resistant (methylated) DNA (Cy3) were normalized to signals from Msp I-cleaved (unmethylated + methylated) DNA fragments (Cy5). Normalized signals from lung cancer cell lines were compared to signals from normal lung cells. About 10.9% of the cancer-related genes were hypermethylated in lung cancer cell lines. Notably, HIC1, IRF7, ASC, RIPK3, RASSF1A, FABP3, PRKCDBP, and PAX3 genes were hypermethylated in most lung cancer cell lines examined. The expression profiles of these genes correlated to the methylation profiles of the genes, indicating that the microarray analysis of DNA methylation in the promoter region of the genes is convenient for epigenetic study. Further analysis of primary tumors indicated that the frequency of hypermethylation was high for ASC (82%) and PAX3 (86%) in all tumor types, and high for RIPK3 in small cell carcinoma (57%). This demonstrates that our PMAD method is effective at finding epigenetic changes during cancer.

A microarray-based method for detecting methylated loci

AbstractCpG island DNA methylation plays an important role in regulating gene expression in development and carcinogenesis. We developed a new microarray-based method called methylation amplification DNA chip (MAD) for detecting differences in methylation. In this method, only methylated CpG islands from the two samples that we wanted to compare were amplified and used for hybridization. The resource material for the microarray was derived from the methylated DNA library of the sample in which we wanted to detect hypermethylation. Choosing the methylated DNA library as the resource material of the microarray increased the percentage of DNA fragments derived from hypermethylated loci on the microarray.

hnRNP B1 protein may be a possible prognostic factor in squamous cell carcinoma of the lung.

Heterogeneous nuclear ribonucleoprotein (hnRNP) B1 is an RNA-binding protein that is required for the maturation of mRNA precursor. It was previously reported that hnRNP A2/B1 was overexpressed at the early clinical stage of lung cancer, and that hnRNP B1 protein, a splicing variant of hnRNP A2 mRNA, was elevated in lung cancer tissues. In this study, we applied the immunohistochemical method, using anti-hnRNP B1 antibody to analyze the usefulness of the hnRNP B1 antibody as a prognostic marker and also as a marker useful for early detection. A total of 206 specimens were examined. Histological examination revealed this protein to be positive in 79 (71.2%) of 111 squamous cell carcinomas and in 45 (64.3%) of 70 adenocarcinomas, respectively. This protein was also expressed in 24 (63.2%) of 38 roentgenographically occult carcinomas and in seven (63.6%) of 11 dysplastic lesions. These findings suggest the possible participation of this protein in early carcinogenesis. Furthermore, the survival curve of the squamous cell carcinoma patients with hnRNP B1 overexpresseion showed a better prognosis compared with that of the patients without hnRNP B1 expression (P=0.014), whereas in adenocarcinoma patients, there was no such a difference between them (P=0.889). These findings indicate that hnRNP B1 could be a useful marker for the early detection of bronchogenic squamous cell carcinoma and that it may be a prognostic factor in this cell type.

A prospective evaluation of transbronchial ultrasonography for assessment of depth of invasion in early bronchogenic squamous cell carcinoma.

In order to determine the appropriate treatment modality for roentgenographically occult bronchogenic squamous cell carcinoma (ROSCC), it is essential to evaluate the depth of invasion, because ROSCC invading beyond the cartilaginous layer cannot be effectively treated by photodynamic therapy (PDT) due to spread of disease. Transtracheal endoscopic ultrasonography (TUS) was useful for predicting the depth of invasion in some ROSCCs. In order to assess the actual significance of TUS as a diagnostic tool for predicting the depth of carcinoma invasion, we have conducted a prospective trial with 22 lesions of ROSCCs. We ultrasonographically classified the degree of the depth of invasion into two groups; A: invasion does not reach cartilaginous layer and B: invasion involves cartilaginous layer. Then the patients were treated by irradiation, PDT, or surgical resection. Pathological findings were also classified into A or B. In order to calculate the sensitivity for evaluating the depth of invasion by TUS, the cases without any tumor and/or malignant cells after PDT were regarded as pathological A. In the evaluation of the depth of carcinoma invasion staying inside the cartilaginous layer, the sensitivity and the positive predictive value were 85.7%, the specificity was 66.7%, and the accuracy was 80.0%. With TUS, preoperative evaluation of the depth of invasion would be more accurate, and the decision of treatment modality would be more appropriate, compared with the conventional bronchoscopic observation alone.

Chromosome 12, frequently deleted in human pancreatic cancer, may encode a tumor-suppressor gene that suppresses angiogenesis.

Several lines of evidence have suggested that the long arm of chromosome 12 may carry a tumor-suppressor gene(s) that plays a role in pancreatic ductal carcinogenesis. We have previously found a significant association between loss of heterozygosity of the 12q arm and a poor prognosis in pancreatic cancer patients. In this study, we introduced a normal copy of chromosome 12 into some pancreatic ductal carcinoma cells. Both anchorage-dependent and -independent proliferations as well as invasiveness were similar throughout the hybrid clones when compared with their corresponding parental cells. In sharp contrast, significant suppression of tumorigenesis was observed after inoculation of the hybrid clones into nude mice. Measurements made up to 1 month later showed that there was a significant delay in the growth of tumors into which the introduced normal copy of chromosome 12 had been restored. More significantly, using our dorsal skin chamber and an intravital microscopy system experiment in SCID mice, we demonstrated and visualized directly that implantation of the hybrids failed to promote the angiogenic phenotype encountered in the parental cells. Gene expression profiling using the complementary DNA microarray system identified a set of 24 genes differentially expressed between the hybrids and parental cells. An additional set of 18 genes was also identified that were differentially expressed between the hybrid clone that lost its growth-suppression activity and one that retained such activity. Another set of 25 genes mapped on 12q was detected that showed high expression levels in the hybrid clones retaining growth-suppressive activity. In summary, this study provides the first functional evidence of the existence of an additional tumor-suppressor gene(s) on chromosome 12, whose absence is responsible for the pathogenesis in pancreatic ductal carcinogenesis.

Endobronchial carcinoid tumor combined with pulmonary non-tuberculous mycobacterial infection: report of two cases

We report here two cases of endobronchial carcinoid tumor complicated with pulmonary infection with non-tuberculous mycobacteria (NTM). Case 1 was an 81-year-old woman with the left lower lobe atelectasis. Bronchoscopy showed complete obstruction of the left basal bronchus by a tumor and a sleeve lower lobectomy with mediastinal lymph node dissection was performed. Pathological examination showed typical carcinoid located in the left basal bronchus and many caseous granulomas containing mycobacteria in the lung parenchyma distal to the bronchus. Bacterial examinations of sputum and gastric juice after the operation showed a growth of Mycobacterium kansasii. Case 2 was a 50-year-old woman with the atelectasis of the left upper division. Bronchoscopy showed complete obstruction of the left upper division bronchus by a tumor and a left upper lobectomy with mediastinal lymph node dissection was performed. Pathological examination showed typical carcinoid located in the left upper division bronchus and many caseous granulomas in the lung parenchyma distal to the bronchus. The Ziehl-Neelsen stain showed many mycobacteria in these granulomas and they were identified as Mycobacterium avium by PCR analysis. Although NTM are not well recognized as possible pathogens of pulmonary infection related to bronchial obstruction by endobronchial carcinoma, our experiences rouse a caution to consider NTM as potential pathogens. We also discuss the possible mechanisms responsible for the specific relationship between carcinoid tumor and TNM.

Roentgenographically occult bronchogenic squamous cell carcinoma involving mediastinal lymph nodes after removal of initial lesion by the diagnostic examination

A 69-year-old male was suspected of having lung cancer by sputum cytology and diagnosed as roentgenographically occult squamous cell carcinoma (ROSCC) at the spur of left B(1+2)/B(3). However, after the first bronchoscopy, no suspicious lesion was detected by any examinations. Therefore, we considered that cancer cells had been removed completely by the initial examination, and the patient was followed up by sputum cytology, chest roentgenogram, and bronchoscopy. Sixteen months later from the initial examination, bronchoscopy was performed for follow-up. The bronchoscopic findings showed the elevation of the surface of left B(1+2) a+b, but the cytologic specimen by brushing toward B(1+2) a+b showed negative findings. However, the lesion had developed to polypoid-shaped tumor and obstructed B(1+2) a+b after the next 6 months. The tumor was diagnosed as squamous cell carcinoma, and hilar and mediastinal nodal involvement was suspected on chest computed tomography. The standard thoracotomy was performed and the pathological results showed positive for nodal involvement on hilus and mediastinum. The tumor is considered to arise from the residual cancer cells of initially detected ROSCC. In conclusion, although some ROSCCs regress by the diagnostic examinations, it is important to detect the recurrence of residual cancer cells as early as possible by intensive follow-up.