The mitotic apparatus plays a pivotal role in dividing cells to ensure each daughter cell receives a full set of chromosomes and complement of cytoplasm during mitosis. A human homologue of the Drosophila warts tumor suppressor, h-warts/LATS1, is an evolutionarily conserved serine/threonine kinase and a dynamic component of the mitotic apparatus. We have identified an interaction of h-warts/LATS1 with zyxin, a regulator of actin filament assembly. Zyxin is a component of focal adhesion, however, during mitosis a fraction of cytoplasmic-dispersed zyxin becomes associated with h-warts/LATS1 on the mitotic apparatus. We found that zyxin is phosphorylated specifically during mitosis, most likely by Cdc2 kinase, and that the phosphorylation regulates association with h-warts/LATS1. Furthermore, microinjection of truncated h-warts/LATS1 protein, including the zyxin-binding portion, interfered with localization of zyxin to mitotic apparatus, and the duration of mitosis of these injected cells was significantly longer than that of control cells. These findings suggest that h-warts/LATS1 and zyxin play a crucial role in controlling mitosis progression by forming a regulatory complex on mitotic apparatus.

We identified a human homolog of Drosophila warts tumor suppressor gene, termed h-warts, which was mapped at chromosome 6q24-25.1. The h-warts protein has a serine/ threonine kinase domain and is localized to centrosomes in interphase cells. However, it becomes localized to the mitotic apparatus, including spindle pole bodies, mitotic spindle, and midbody, in a highly dynamic manner during mitosis. Furthermore, h-warts is specifically phosphorylated in cells at mitotic phase, most likely by Cdc2 kinase. These findings suggest that hwarts functions as a component of the mitotic apparatus and is involved in proper progression of mitosis.z 1999 Federation of European Biochemical Societies.

We have cloned a cDNA for a novel human homolog of the Drosophila discs large (dlg) tumor suppressor protein, termed NE-dlg (neuronal and endocrine dlg). Northern blot analysis revealed that the gene is highly expressed in neuronal and endocrine tissues. Fluorescence in situ hybridization (FISH) and radiation hybrid mapping studies localized the NE-dlg gene to chromosome Xq13. We also found that the NE-dlg gene encoded a 100 kDa protein. Immunolocalization studies using an NE-dlg antibody showed that the protein tended to be expressed in non-proliferating cells, such as neurons, cells in Langerhans islets of the pancreas, myocytes of the heart muscles, and the prickle and functional layer cells of the esophageal epithelium. Proliferative cells, including various cultured cancer cell lines and basal cells in the esophageal epithelium, showed little expression of the NE-dlg protein. In addition, yeast two-hybrid screening and in vitro binding assays revealed that the NE-dlg interacted with the carboxyl-terminal region of the APC tumor suppressor protein. These data suggest that NE-dlg negatively regulates cell proliferation through its interaction with the APC protein.

The Ki-67 index is an important biomarker for indicating the proliferation of cancer cells and is considered to be an effective prognostic factor for breast cancer. However, a standard cut-off point for the Ki-67 index has not yet been established. Therefore, the aim of this retrospective study was to determine an optimal cut-off point in order to establish it as a more accurate prognostic factor. Immunohistochemical analysis of the Ki-67 index was performed on 4329 patients with primary breast cancer from August 1987 to March 2012. Out of this sample, there were 3186 consecutive cases from September 1997 with simultaneous evaluations of ER, PgR and HER2 status. Cox's proportional hazard model was used to perform univariate and multivariate analyses of the factors related to OS. The hazard ratios (HR) and the p values were then compared to determine the optimal cut-off point for the Ki-67 index. The median Ki-67 index value was 20.5% (mean value 26.2%). The univariate analysis revealed that there was a statistically significant negative correlation with DFS and OS and the multivariate analysis revealed that the Ki-67 index value was a significant factor for DFS and OS. The top seven cut-off points were then carefully chosen based on the results of the univariate analysis using the lowest p-values and the highest HR as the main selection criteria. The multivariate analysis of the factors for OS showed that the cut-off point of 20% had the highest HR in all of the cases. However, the cutoff point of 20% was only a significant factor for OS in the Luminal/HER2- subtype. There was no correlation between the Ki-67 index value and OS in any of the other subtypes. These data indicate that the optimal cut-off point of 20% is the most effective prognostic factor for Luminal/HER2- breast cancer.

An iron chelate, ferric nitrilotriacetate (Fe‐NTA), induces renal proximal tubular damage, a consequence of iron‐catalyzed free radical reactions, that finally leads to a high incidence of renal cell carcinoma (RCC) in rodents. Previous studies have identified, within 24 h after administration of Fe‐NTA, lipid peroxidation products, aldehyde‐modified proteins and a variety of modified DNA bases such as 8‐hydroxyguanine that may be mutagenic in vivo. In the present study, pathological features of the RCCs were studied, and, in an effort to correlate them with carcinogen‐specific molecular events in Fe‐NTA‐induced carcinogenesis, the H‐, K‐ and N‐ras oncogenes and the p53 tumor suppressor gene were investigated for the presence of mutations. Fe‐NTA‐induced RCCs showed similarity to human RCCs in that they are often invasive, metastatic and fatal. None (0 of 12) of the tumors had mutation in codons 12, 13 and 61 of the H‐, K‐ and N‐ras genes by direct sequencing. Only one (1 of 12) tumor with high grade histology revealed a CGC‐to‐CTC (Arg to Leu) transversion in codon 246 of the p53 gene by the use of single strand conformation polymorphism (SSCP) analysis and direct sequencing. High expression of mutant p53 protein was confirmed by Western blotting and immunohistochemistry. Study of three peritoneal mesotheliomas induced by Fe‐NTA revealed no mutation in ras and p53 genes. These results suggest that the ras and p53 genes are not the major targets of mutation in Fe‐NTA‐induced carcinogenesis of kidney and mesothelium. Instead, p53 mutation may work for potentiation of malignant character in Fe‐NTA‐induced renal carcinogenesis.

Objectives: To evaluate the impact of pulmonary emphysema (PE) on the incidence and severity of radiation pneumonitis (RP) in patients with lung and mediastinal tumours. Methods: 92 patients were enrolled. Involved-field radiation therapy (non-small cell carcinoma or mediastinal tumours in 69 patients; median 70 Gy) and accelerated hyperfractionation (limited disease small cell carcinoma in 23 patients; median 45 Gy) were performed. Common Terminology Criteria for Adverse Events v.3.0 was used to evaluate RP and the relationship with the percentage of pulmonary volume irradiated to .20 Gy (V20) and PE. PE was diagnosed by the presence of low-attenuation areas (LAAs) on CT scans and was classified into Grades 0-4 according to the extent of the LAAs. Results: The median follow-up time was 16 months. The 6-month cumulative incidence of RP at Grade 3 or greater was 7.7% and 34.1% in patients with a V20 of ,25% and >25%, respectively (p50.017). In patients with PE Grades 0, 1, 2 and 3 or greater, the incidence of RP was 16.5%, 9.1%, 8.6% and 54.0%, respectively. As the PE Grade increased, the incidence of RP also increased significantly. Conclusion: The incidence and severity of RP are significantly higher in patients with a high V20 value as well as in those with severe PE.

Abstract. The Ki-67 index value is a prognostic factor in primary breast cancer and is a proliferation marker that also distinguishes between luminal type A and type B breast cancer. Moreover, a change in Ki-67 index values due to treatment and recurrence is considered to be important in treating breast cancer. In this study, we investigated whether the baseline Ki-67 value in the primary tumor is useful as a prognostic factor following disease recurrence. Immunohistochemical analysis of the Ki-67 index was performed on 4,701 patients with primary breast cancer from 1987 until March, 2013. Among these patients, there were 666 consecutive cases exhibiting recurrence after primary surgery. The fraction of proliferating cells was based on a count of at least 500 tumor cells in the area including the hot spot. The Ki-67 values were divided into 3 groups, namely <20, ≥20 and ≥50%. The investigated items included estrogen receptor (ER), progesterone receptor (PgR), human epidermal growth factor receptor 2 (HER2), tumor size, nodal status for the primary tumor, recurrence site (soft tissue, bone and viscera) and disease-free interval (DFI). The Cox's proportional hazard model was used to perform univariate and multivariate analyses of the factors associated with overall survival (OS) following recurrence. The median follow-up period was 65.9 months in the surviving group. The median Ki-67 value at baseline was 20% in all the cases and 27% in the recurrent cases. The Ki-67 values were low (24%) in patients with bone metastasis and significantly higher in patients with liver or brain metastasis (38 and 55%, respectively). Moreover, DFI was found to be inversely correlated with the Ki-67 values. Univariate analysis was performed to identify the prognostic factors for OS after recurrence. The significant factors included tumor size, lymph node status, ER, PgR, DFI, recurrence site and Ki-67 index value. Among these factors, a multivariate analysis identified the Ki-67 index value in the primary tumor as an independent significant factor, particularly in luminal type tumors. The Ki-67 index value in the primary tumor was a significant prognostic factor for OS after disease recurrence. It is, therefore, important to take the Ki-67 index value into consi deration for the treatment and follow-up of breast cancer patients.

Background: Breast cancer is associated with a relatively good prognosis. Prognostic factors examined to date are related to early recurrence while those related to late recurrence and their countermeasures remain unclear. Therefore, we examined the factors related to late recurrence. Patients and Methods: From January 1980 to August 2012, 4,774 patients who underwent primary treatment and estrogen (ER) and progesterone receptor (PgR) assessment were enrolled in this study. The patients were divided into two groups, those with a follow-up period <10 years and those without any recurrence at 10 years but who continued follow-up examinations. Recurrence occurred in 711 patients followed up for <10 years and in 51 patients for ≥10 years. Results: The overall 10-year cumulative disease-free survival rate was 79.5%, and the recurrence rate at ≥10 years was 5.8%. A multivariate analysis revealed that the factors related to late recurrence were PgR positivity and positive nodes. This result differed from that for early recurrence in terms of ER/PgR, Ki-67 index and p53 overexpression. Conclusion: PgR positivity and lymph node metastases significantly correlated with late recurrence. Therefore, it is important to evaluate appropriate measures such as treatment period and treatment regimen for hormone-sensitive patients.