Toshi Menju

UCHL1 provides diagnostic and antimetastatic strategies due to its deubiquitinating effect on HIF-1α

Hypoxia-inducible factor 1 (HIF-1) plays a role in tumour metastases; however, the genes that activate HIF-1 and subsequently promote metastases have yet to be identified. Here we show that Ubiquitin C-terminal hydrolase-L1 (UCHL1) abrogates the von Hippel–Lindau-mediated ubiquitination of HIF-1α, the regulatory subunit of HIF-1, and consequently promotes metastasis. The aberrant overexpression of UCHL1 facilitates distant tumour metastases in a HIF-1-dependent manner in murine models of pulmonary metastasis. Meanwhile, blockade of the UCHL1–HIF-1 axis suppresses the formation of metastatic tumours. The expression levels of UCHL1 correlate with those of HIF-1α and are strongly associated with the poor prognosis of breast and lung cancer patients. These results indicate that UCHL1 promotes metastases as a deubiquitinating enzyme for HIF-1α, which justifies exploiting it as a prognostic marker and therapeutic target of cancers.

Mutations in Keap1 are a potential prognostic factor in resected non-small cell lung cancer.

Mutations in Kelch‐like ECH‐associated protein 1 (Keap1) have been reported to protect tumor cells from chemotherapeutic agents. However, their prognostic significance in nonsmall cell lung cancer (NSCLC) is still unclear. In this study, we examined the effect of Keap1 gene mutations on survival and disease‐free interval using resected primary NSCLC tissue.

GEP100-Arf6-AMAP1-Cortactin Pathway Frequently Used in Cancer Invasion Is Activated by VEGFR2 to Promote Angiogenesis

Angiogenesis and cancer invasiveness greatly contribute to cancer malignancy. Arf6 and its effector, AMAP1, are frequently overexpressed in breast cancer, and constitute a central pathway to induce the invasion and metastasis. In this pathway, Arf6 is activated by EGFR via GEP100. Arf6 is highly expressed also in human umbilical vein endothelial cells (HUVECs) and is implicated in angiogenesis. Here, we found that HUVECs also highly express AMAP1, and that vascular endothelial growth factor receptor-2 (VEGFR2) recruits GEP100 to activate Arf6. AMAP1 functions by binding to cortactin in cancer invasion and metastasis. We demonstrate that the same GEP100-Arf6-AMAP1-cortactin pathway is essential for angiogenesis activities, including cell migration and tubular formation, as well as for the enhancement of cell permeability and VE-cadherin endocytosis of VEGF-stimulated HUVECs. Components of this pathway are highly expressed in pathologic angiogenesis, and blocking of this pathway effectively inhibits VEGF- or tumor-induced angiogenesis and choroidal neovascularization. The GEP100-Arf6-AMAP1-cortactin pathway, activated by receptor tyrosine kinases, appears to be common in angiogenesis and cancer invasion and metastasis, and provides their new therapeutic targets.

Engagement of Overexpressed Her2 with GEP100 Induces Autonomous Invasive Activities and Provides a Biomarker for Metastases of Lung Adenocarcinoma

Overexpression of Her2/ErbB2/Neu in cancer is often correlated with recurrent distant metastasis, although the mechanism still remains largely elusive. We have previously shown that EGFR, when tyrosine-phosphorylated, binds to GEP100/BRAG2 to activate Arf6, which induces cancer invasion and metastasis. We now show that overexpressed Her2 in lung adenocarcinoma cells also employs GEP100. Like EGFR-GEP100 binding, this association is primarily mediated by the pleckstrin homology (PH) domain of GEP100 and Tyr1139/Tyr1196 of Her2. Tyr1139/Tyr1196 are autonomously phosphorylated, when Her2 is overexpressed. Accordingly, invasive activities mediated by the Her2-GEP100 pathway are not dependent on external factors. Blocking Her2-GEP100 binding, as well as its signaling pathway all inhibit cancer invasive activities. Moreover, our clinical study indicates that co-overexpression of Her2 with GEP100 in primary lung adenocarcinomas of patients is correlated with the presence of their node-metastasis with a statistical significance. Since the GEP100 PH domain interacts with both Her2 and EGFR, targeting this domain may provide novel cancer therapeutics.

Genes suppressed by DNA methylation in non-small cell lung cancer reveal the epigenetics of epithelial-mesenchymal transition

BackgroundDNA methylation is associated with aberrant gene expression in cancer, and has been shown to correlate with therapeutic response and disease prognosis in some types of cancer. We sought to investigate the biological significance of DNA methylation in lung cancer.ResultsWe integrated the gene expression profiles and data of gene promoter methylation for a large panel of non-small cell lung cancer cell lines, and identified 578 candidate genes with expression levels that were inversely correlated to the degree of DNA methylation. We found these candidate genes to be differentially methylated in normal lung tissue versus non-small cell lung cancer tumors, and segregated by histologic and tumor subtypes. We used gene set enrichment analysis of the genes ranked by the degree of correlation between gene expression and DNA methylation to identify gene sets involved in cellular migration and metastasis. Our unsupervised hierarchical clustering of the candidate genes segregated cell lines according to the epithelial-to-mesenchymal transition phenotype. Genes related to the epithelial-to-mesenchymal transition, such as AXL, ESRP1, HoxB4, and SPINT1/2, were among the nearly 20% of the candidate genes that were differentially methylated between epithelial and mesenchymal cells. Greater numbers of genes were methylated in the mesenchymal cells and their expressions were upregulated by 5-azacytidine treatment. Methylation of the candidate genes was associated with erlotinib resistance in wild-type EGFR cell lines. The expression profiles of the candidate genes were associated with 8-week disease control in patients with wild-type EGFR who had unresectable non-small cell lung cancer treated with erlotinib, but not in patients treated with sorafenib.ConclusionsOur results demonstrate that the underlying biology of genes regulated by DNA methylation may have predictive value in lung cancer that can be exploited therapeutically.

Virtual-assisted lung mapping: outcome of 100 consecutive cases in a single institute

OBJECTIVES We developed virtual-assisted lung mapping (VAL-MAP), a bronchoscopic multispot dye-marking technique using three dimensional (3D) virtual imaging, for precise thoracoscopic sublobar lung resection with safe surgical margins. We herein review the results of 100 consecutive cases of VAL-MAP in our institute to identify types of tumours or resections that benefit from VAL-MAP. METHODS Markings were bronchoscopically made within 2 days preoperatively using virtual 3D images. Post-VAL-MAP computer tomography (CT) scans localizing the actual markings were reconstructed into 3D images for intraoperative navigation. All data on patients, markings and outcomes were prospectively collected, and the contribution of VAL-MAP to the operation was graded by the surgeon. RESULTS Resections of 156 lung lesions in 100 consecutive patients were planned from July 2012 to March 2014. The lesion diameter was 8.3 ± 4.9 (range, 2-24) mm. The total number of actually conducted markings was 380 (3.83 ± 1.07 markings/patient). Eighty-four lesions were resected by 71 wedge resections using 158 markings (2.1 ± 0.1/resection; range, 1-3). Seventy lesions were resected by 63 segmentectomies using 224 markings (3.6 ± 0.1/resection; range, 2-6). Markings were identifiable on post-VAL-MAP CT mostly as ground-glass opacities (87.7%) and/or bronchial dilatation (56.1%). During the operation, 357 of 380 markings (93.9%) were visible on the pleural surface and significantly associated with marking visibility on CT. Multiple markings that were complementary to one another appeared to have contributed to the high rate of successful resection (99.3%) with satisfactory resection margins. The contribution of VAL-MAP to the operation as graded by surgeons demonstrated that VAL-MAP is most effective during wedge resection or complex segmentectomy for hardly palpable, small tumours, while VAL-MAP still plays an important role in simple segmentectomy or resection of palpable tumours by providing higher confidence levels to surgeons during the operation. Minor pneumothoraces were found on post-VAL-MAP CT images in 4 patients without symptoms or a need for treatment. CONCLUSIONS The present study further demonstrated the efficacy and safety of VAL-MAP. VAL-MAP is likely to benefit a broader range of patients than are conventional marking techniques by assisting with both accurate tumour identification and precise determination of resection lines.

Spontaneous rupture of mediastinal cystic teratoma into the pleural cavity : Report of two cases and review of the literature

The authors report on two female patients aged 12 and 14 years, who spontaneously developed a rupture of benign mediastinal cystic teratoma into the right pleural cavity. They presented with acute onset of severe chest pain and respiratory distress. The tumors were completely resected by thoracotomy. The serum and pleural fluid levels of carcinoembryonic antigens, CA-125 and CA19-9 were invariably elevated, then decreased to normal range after the surgical resection. Rapid diagnosis of this extremely rare complication is important because it may progress to a life-threatening condition.

Postoperative pulmonary function and complications in living-donor lobectomy

BACKGROUND Successful living-donor lobar lung transplantation largely depends on the donors outcome. Because surgical skills and peri-operative management have evolved over time, this study evaluated the recent outcomes of donor lobectomies. METHODS Between 2008 and 2014, 48 consecutive living-donor lobar lung transplantations with 85 donor lobectomies were performed at Kyoto University. All donors were prospectively followed up regularly until 1 year after surgery. RESULTS Right and left lower lobectomies were performed in 49 and 36 donors, respectively. Pulmonary arterial branches were sacrificed at equal frequency in both lobectomies, whereas pulmonary arterioplasty was only performed in left lower lobectomy (n = 9). All donors were discharged after the lobectomies, and none died during follow-up. Post-operative complications occurred in 24 donors (28%) overall, without a significant difference between donor sides. Intraoperative complications were found in 2 donors. Early and late post-operative complications were noted in 17 and 6 donors, respectively. Pneumothorax, pleuritis, and pleural effusion were the most frequent. Post-operative pulmonary function sequentially recovered more than expected and was not significantly affected by the sacrifice of pulmonary arterial branches during lobectomy. By contrast, pulmonary function at 1 year after donor lobectomy in the donors who had peri-operative complications was significantly lower than that in the donors who did not, although even post-operative pulmonary function in the donors with peri-operative complications still recovered more than expected. CONCLUSIONS Living-donor lobectomies have been safely performed in recent decades with low morbidities and without mortality.

Predictive factors of myasthenic crisis after extended thymectomy for patients with myasthenia gravis

OBJECTIVES Postoperative myasthenic crisis (POMC) is one of the serious complications after extended thymectomy for patients with myasthenia gravis (MG). This study aims to clarify the risk factors of POMC occurrence. METHODS The clinical data of 55 MG patients (25 male, 30 female; median age, 51 years) who underwent extended thymectomy at Kyoto University from 2000 to 2013 were retrospectively reviewed. Surgical outcomes and pre- and perioperative predictive factors of POMC were analysed. RESULTS The preoperative Myasthenia Gravis Foundation of America stage was I, II, III and IV in 24, 22, 8 and 1 patients, respectively. Ten patients (18.2%) developed POMC; 6 required prolonged intubation over 24 h and 4 required reventilatory support. All patients were weaned after 5.6 (2-26) days of ventilator support, and were discharged. Univariate analysis revealed a correlation with a high preoperative anti-acetylcholine receptor antibody titre (P = 0.009), history of myasthenic crisis (MC) (P = 0.0004) and unstable MG after preoperative medical therapy (P = 0.003). Multivariate logistic regression analysis showed history of MC (odds ratio, 11.84; 95% confidential interval, 1.05-372; P = 0.045) and unstable MG (odds ratio, 29.45; 95% confidential interval, 2.00-1063; P = 0.013) independently predicted POMC. The surgical response rate was not significantly different between the two groups (66.7% with POMC, 85.4% without POMC; P = 0.334). CONCLUSIONS POMC occurred more frequently in unstable MG before surgery or in patients with a history of MC. Adequate preoperative medical therapy and perioperative care should be provided to these patients.

Hypoxia-inducible factor 1 promotes chemoresistance of lung cancer by inducing carbonic anhydrase IX expression

Lung cancer treatment is difficult owing to chemoresistance. Hypoxia‐inducible factor 1 (HIF‐1) and HIF‐1‐induced glycolysis are correlated with chemoresistance; however, this is not evident in lung cancer. We investigated the effect of HIF‐1α and carbonic anhydrase IX (CAIX), a transmembrane protein neutralizing intracellular acidosis, on chemoresistance and prognosis of lung cancer patients after induction chemoradiotherapy. Associations of HIF‐1α, glucose transporter 1 (GLUT1), and CAIX with chemoresistance of lung cancer were investigated using A549 lung cancer cells under normoxia or hypoxia in vitro. HIF‐1α‐induced reprogramming of glucose metabolic pathway in A549 cells and the effects of HIF‐1 and CAIX on the cytotoxicity of vinorelbine were investigated. Immunohistochemical analyses were performed to determine HIF‐1α, GLUT1, and CAIX expression levels in cancer specimens from lung cancer patients after induction chemoradiotherapy. Hypoxia induced HIF‐1α expression in A549 cells. Moreover, hypoxia induced GLUT1 and CAIX expression in A549 cells in a HIF‐1‐dependent manner. Glucose metabolic pathway was shifted from oxidative phosphorylation to glycolysis by inducing HIF‐1α in A549 cells. HIF‐1 and CAIX induced chemoresistance under hypoxia, and their inhibition restored the chemosensitivity of A549 cells. The expression levels of HIF‐1α, GLUT1, and CAIX were associated with poor overall survival of lung cancer patients after induction chemoradiotherapy. HIF‐1 and CAIX affected the chemosensitivity of A549 cells and prognosis of lung cancer patients. Therefore, inhibition of HIF‐1 and CAIX might improve prognosis of lung cancer patients after induction chemoradiotherapy. Further analysis might be helpful in developing therapies for lung cancer.