Cellular stresses induce the nuclear accumulation of importin alpha and cause a conventional nuclear import block.

We report here that importin alpha accumulates reversibly in the nucleus in response to cellular stresses including UV irradiation, oxidative stress, and heat shock. The nuclear accumulation of importin alpha appears to be triggered by a collapse in the Ran gradient, resulting in the suppression of the nuclear export of importin alpha. In addition, nuclear retention and the importin beta/Ran-independent import of importin alpha also facilitate its rapid nuclear accumulation. The findings herein show that the classical nuclear import pathway is down-regulated via the removal of importin alpha from the cytoplasm in response to stress. Moreover, whereas the nuclear accumulation of heat shock cognate 70 is more sensitive to heat shock than the other stresses, importin alpha is able to accumulate in the nucleus at all the stress conditions tested. These findings suggest that the stress-induced nuclear accumulation of importin alpha can be involved in a common physiological response to various stress conditions.

Foreign bodies in the external auditory canal: Influence of age on incidence and outcomes in a Japanese population.

AIM: Otolaryngologists occasionally observe foreign bodies (FB) in the external auditory canal (EAC), although relatively few studies have focused on the role of age in this condition. METHODS: We retrospectively compared the incidences, outcomes and complications of FB in the EAC in different age groups. RESULTS: The patients at our center included 24 children (19%), 46 adults (37%) and 56 older adults (44%). Compared with adults, older adults were significantly more likely to have FB (peak age 75-79 years), be women (18/46 vs 34/56, P = 0.0461) and be unaware of their FB (18/46 vs 34/56, P = 0.0461). We observed that all EAC FB were more common during the summer, and biotic FB were not observed during the winter. Complications were more common in cases of biotic FB, compared with abiotic FB (5/27 vs 6/99, P = 0.0421). CONCLUSION: Our findings show that older adults are particularly susceptible to FB, are frequently unaware of their FB and can develop complications. These characteristics should be considered before treating FB in the EAC. Geriatr Gerontol Int 2017; 17: 2131-2135.

Neutrophil elastase and cancer.

This mini-review summarizes our recent experimental and clinical studies on neutrophil elastase (NE) and cancer based on our original view point. Neoplasms metastasize as a result of a complex series of events. This process requires various degradative enzymes including proteases. NE has broad substrate specificity under physiological conditions, and excessive NE results in digestion of not only elastin, but also other extracellular matrix proteins. Several cell lines from human breast cancer and human lung cancer produce immunoreactive NE. The amount of immunoreactive NE in tumor tissue is an independent prognostic indicator of patients with breast cancer and lung cancer. Furthermore, a specific NE inhibitor completely suppressed growth of cancer cells transplanted into severe combined immunodeficiency mice. The use of NE inhibitor would seem to be a promising way to prevent the invasion and metastasis of cancer.

Circulating tumor cells detected by reverse transcription-polymerase chain reaction for carcinoembryonic antigen mRNA: distinguishing follicular thyroid carcinoma from adenoma.

BACKGROUND: We prospectively tested whether circulating tumor cells could be detected in peripheral blood of patients with thyroid tumors by a reverse transcription-polymerase chain reaction (RT-PCR) to detect carcinoembryonic antigen (CEA) messenger RNA (mRNA). METHODS: We assayed for CEA mRNA by RT-PCR in peripheral blood sampled before and 2 to 3 weeks after curative surgery for thyroid tumors in 121 patients. Blood samples from 7 patients with chronic thyroiditis and 7 healthy subjects served as controls. RESULTS: No control samples were positive for CEA mRNA by RT-PCR. Of 121 preoperative samples from patients with thyroid tumor, 6 were positive (5.0%). Preoperative frequencies of CEA mRNA positivity in benign tumor, papillary carcinoma, follicular variant papillary carcinoma, minimally invasive follicular carcinoma, and widely invasive follicular carcinoma were 0%, 0%, 0%, 44.4% (4/9), and 50.0% (2/4), respectively. Among positive patients only one, who had widely invasive follicular carcinoma, remained positive after surgery. CONCLUSIONS: RT-PCR detection of tumor cells in preoperative blood often can distinguish malignant from benign follicular thyroid tumors.

Efficacy and safety of pilocarpine mouthwash in elderly patients with xerostomia.

OBJECTIVE: Xerostomia is a common condition and can impair quality of life. Pilocarpine tablet ingestion can relieve dry mouth symptoms, but produces numerous adverse effects. We investigated the safety and efficacy of pilocarpine mouthwash. METHODS: Forty elderly patients were randomly divided into a pilocarpine mouthwash or water rinse (control) group. Outcomes were assessed by visual analog scale (VAS) scores and stimulated salivary flow rate before and 1 month after treatment. Patients recorded all adverse effects. RESULTS: In the pilocarpine group, we evaluated safety in 24 patients and efficacy in 19 patients. In the water rinse group, we evaluated safety and efficacy in 14 patients. VAS scores were significantly reduced after pilocarpine mouthwash treatment (70 ± 12.9 to 47.9 ± 13.1, p < 0.05). Overall improvement was observed in 47% of the pilocarpine group compared to 14% of the controls (p < 0.05). Stimulated salivary flow rate significantly increased (0.71 ± 0.14 to 0.83 ± 0.12 mL/minute, p < 0.05) after pilocarpine mouthwash treatment. Five of 24 patients reported side effects after pilocarpine mouthwash use, predominantly limited to oral discomfort. CONCLUSIONS: Pilocarpine mouthwash relieved dry mouth symptoms and improved saliva production with minor side effects.

Preoperative evidence of circulating tumor cells by means of reverse transcriptase-polymerase chain reaction for carcinoembryonic antigen messenger RNA is an independent predictor of survival in non-small cell lung cancer: a prospective study.

OBJECTIVE: We conducted a prospective study of 103 consecutive patients with non-small cell lung cancer who underwent a curative lobectomy to test whether circulating tumor cells detected in the peripheral blood by means of reverse transcriptase-polymerase chain reaction of carcinoembryonic antigen messenger RNA is a prognostic indicator independent of tumor stage in patients with non-small cell lung cancer. METHODS: We assayed for carcinoembryonic antigen messenger RNA by means of reverse transcriptase-polymerase chain reaction in peripheral blood taken at the time of diagnosis before an operation and again 2 to 3 weeks after an operation from patients with non-small cell lung cancer who underwent a curative lobectomy between March 1996 and April 1998. We analyzed the prognostic value of carcinoembryonic antigen messenger RNA expression in the patients with non-small cell lung cancer in a univariate and multivariate manner. RESULTS: Patients with carcinoembryonic antigen messenger RNA in the preoperative blood samples had a poor survival when compared with those without carcinoembryonic antigen messenger RNA. Of these patients, the worst survival was seen in those with carcinoembryonic antigen messenger RNA in the postoperative blood samples. The multivariate stepwise analysis selected the preoperative carcinoembryonic antigen messenger RNA expression (P =.0004; relative risk, 0.21) and the pathologic stage of disease (P =.0002; relative risk, 1.43) as the independent prognostic factors for survival. CONCLUSIONS: The molecular detection of carcinoembryonic antigen messenger RNA in the preoperative peripheral blood is an independent prognostic factor in patients with non-small cell lung cancer who undergo a curative operation.

A pulmonary metastatic model of human non-small cell lung carcinoma cells that produce a neutrophil elastase-like molecule in severe combined immunodeficiency mice.

STUDY OBJECTIVES: To establish a clinically relevant animal model of pulmonary metastases of human non-small cell lung carcinoma (NSCLC) cells in severe combined immunodeficiency (SCID) mice, which can be used for repetitive investigations, so as to improve our understanding and management of the cellular and molecular mechanisms of human lung cancer metastases. METHODS AND RESULTS: SCID mice subcutaneously injected in the flank with 1 x 10(6) EBC-1 cells derived from human lung squamous cell carcinoma were killed weekly for examination until 12 weeks after tumor inoculation. The biological characteristics of implanted tumors and their metastatic foci were investigated by hematoxylin-eosin staining and immunostaining for neutrophil elastase (NE). Three weeks after ectopic implantation, EBC-1 cell lines formed a tumor at the inoculation site and grew steadily to show a plateau at 10 weeks. EBC-1 cells formed multiple metastases in the lung 7 weeks after tumor inoculation; their numbers increased steadily until 12 weeks in all mice. Immunoreactivity for NE was intense in the metastatic tumor cells. Then, to establish the primary tumor amputation/pulmonary metastasis model and to evaluate how primary tumor amputation influences the development of pulmonary metastases at the cellular and molecular level, excision was performed before (3 weeks and 5 weeks after inoculation) and after (7 weeks and 9 weeks after inoculation) formation of lung metastases. When the primary tumor was excised 3 weeks after tumor inoculation, all mice had pulmonary metastasis at 12 weeks after inoculation. Blood samples obtained at 3 weeks after tumor inoculation contained human beta-actin messenger RNA, which represents circulating tumor cells. CONCLUSION: Our NSCLC EBC-1 pulmonary metastasis model is reliable, technically simple, and predictably results in pulmonary metastasis from early hematogenous spread. This model may be useful for elucidating the mechanism of pulmonary metastasis in human lung cancer, and testing anti-metastatic efficacy of therapeutic agents in vivo.

Persistent evidence of circulating tumor cells detected by means of RT-PCR for CEA mRNA predicts early relapse: a prospective study in node-negative breast cancer.

BACKGROUND: In 100 consecutive patients with node-negative breast cancer who underwent curative surgery, we prospectively tested whether detection of circulating tumor cells in peripheral blood by means of reverse transcription-polymerase chain reaction for carcinoembryonic antigen (CEA) messenger RNA (mRNA) could predict patient outcomes. METHODS: We performed reverse transcription-polymerase chain reaction in blood samples taken before surgery and in repeat samples taken 2 to 3 weeks after surgery. Univariate and multivariate analyses of relapse-free survival were performed. RESULTS: Patients with CEA mRNA in preoperative samples had poorer survival rates than those who had no detectable CEA mRNA. The worst survival rate was seen in those with CEA mRNA in both pre- and postoperative samples. Stepwise multivariate analysis selected CEA mRNA expression pattern (P=.001; relative risk=0.69) and histologic tumor grade (P=.002; relative risk=1.35) as independent prognostic factors for disease-free survival. CONCLUSIONS: Molecular detection of CEA mRNA in both pre- and postoperative blood samples is an independent, negative prognostic factor in patients with node-negative breast cancer undergoing curative surgery.

Immunohistochemical analysis of estrone sulfatase and aromatase in human breast cancer tissues.

We examined, by immunohistochemical analysis, the expression of aromatase and estrone sulfatase (E1-STS) which are the two major enzymes involved in in situ estrogen synthesis in breast cancer tissue. In the 83 cases examined, E1-STS, which hydrolyses estrone sulfate (E1S) to estrone (E1), was dominantly detected in tumor cells in 43 (59.0%) cases. Aromatase, which converts androgens to estrogens, was dominantly detected in stromal cells of the tumor. It was detected in 39 (47.0%) of the 83 cases. There was no significant correlation between the expression of these two enzymes and clinicopathological factors. We found a tendency for a correlation between aromatase expression and E1-STS expression in breast tumor (p=0.075). In terms of the possible use of these enzymes as prognostic indicators, patients who had aromatase in their tumor showed longer relapse-free survival than those lacking aromatase (p=0.045). Significant correlations between the expression of aromatase and the angiogenesis regulators, vascular endothelial growth factor and thymidine phosphorylase, were found (p=0.047 and p=0.046, respectively), though the presence of aromatase did not correlate with intratumoral microvessel density. This may indicate that aromatase is involved in vascular permeability and recruitment of endothelial cells rather than neovascularization. It may be useful to study the expression of aromatase and E1-STS using immunocytochemical analysis for understanding the tumor characteristics in breast cancer.

Immunohistochemical expression of SKALP/elafin in squamous cell carcinoma of human lung.

The immunohistochemical expression of skin-derived anti-leukoproteinase (SKALP)/elafin in lung squamous cell carcinoma (SqCC) and in uninvolved bronchial epithelium was studied. The results were compared with the immunohistochemical staining of proliferating cell nuclear antigen (PCNA) and the TDT-mediated dUTP-digoxigenin nick end labeling (TUNEL) assay. SKALP/elafin was expressed in SqCC with high incidence (82.6%). By contrast, the expression was not observed in uninvolved bronchial epithelium. SKALP/elafin expression was located in the cell layers just underneath the cornified envelope of each cell nest, and DNA fragmentation was observed in the same cell layers. PCNA was expressed in the basal layer of each cornified envelope, and both expressions were clearly separated. Immunoreactive score of SKALP/elafin expression was significantly correlated with the differentiation and it tended to increase in accordance with the degree of differentiation. These results suggested that SKALP/elafin was possibly involved in the cell differentiation program, finally leading to keratinization in SqCC of human lung. SKALP/elafin may be a beneficial molecular target for detection or even the development of a new therapeutic method against cancer.

Prognostic significance of immunoreactive neutrophil elastase in human breast cancer: long-term follow-up results in 313 patients.

OBJECTIVE: We have measured the concentration of immunoreactive neutrophil elastase (ir-NE) in the tumor extracts of 313 primary human breast cancers. Sufficient time has elapsed, and we are now ready to analyze its prognostic value in human breast cancer. METHODS: ir-NE concentration in tumor extracts was determined with an enzyme-linked immunosorbent assay that enables a rapid measurement of both free-form ir-NE and the A1-protease inhibitor-complexed form of ir-NE. We analyzed the prognostic value of this enzyme in human breast cancer in univariate and multivariate analyses. RESULTS: Patients with breast cancer tissue containing a high concentration of ir-NE had poor survival compared to those with a low concentration of ir-NE at the cutoff point of 9.0 microg/100 mg protein (P = .0012), which had been previously determined in another group of 49 patients. Multivariate stepwise analysis selected lymph node status (P = .0004; relative risk = 1.46) and ir-NE concentration (P = .0013; relative risk = 1.43) as independent prognostic factors for recurrence. CONCLUSIONS: Tumor ir-NE concentration is an independent prognostic factor in patients with breast cancer who undergo curative surgery. This enzyme may play an active role in tumor progression that leads to metastasis in human breast cancer.

Comparison of video-assisted minithoracotomy and standard open thoracotomy for the treatment of non-small cell lung cancer.

This study represents a retrospective comparison of video-assisted thoracic surgery (VATS) lobectomy with standard open lobectomy for non-small cell lung cancer (NSCLC). The endpoints of this study include surgical stress as measured by interleukin 6 concentration and patient survival. A retrospective review was performed of 240 consecutive patients with clinical stage IA or IB NSCLC who underwent either VATS lobectomy (n=67) or conventional open lobectomy (n=173). The amount of blood loss was significantly less in the VATS group (110+/-75 ml) as compared to 165+/-90 ml for the open lobectomy group (P<0.05). A significantly lower incidence of post-thoracotomy pain occurred in the VATS group (6.2+/-4.1 times/3 days) than in the open lobectomy group (13.5+/-5.8 times/3 days, P<0.0001). The postoperative interleukin (IL)-6 serum concentration of was significantly lower in the VATS group (112+/-43 pg/ml) than that in the open lobectomy group (351+/-133 pg/ml, P<0.001). There was no statistically significant difference in survival between the VATS and open lobectomy groups. The median follow-up was 42 months in both groups. VATS lobectomy for NSCLC is a reasonable treatment option for selected patients with stage I NSCLC.

Does megestrol acetate down-regulate interleukin-6 in patients with cancer-associated anorexia and weight loss? A North Central Cancer Treatment Group investigation.

Megestrol acetate improves appetite and abrogates weight loss in some patients with advanced cancer. Moreover, preliminary studies suggest that progestational agents down-regulate interleukin-6 (IL-6), an inflammatory cytokine widely implicated in cancer-associated anorexia and weight loss. The present investigation examined the effects of megestrol acetate on IL-6 in an attempt to confirm these earlier, preliminary studies. The translational component of a large multi-institutional trial, this investigation examined 85 patients with advanced cancer and weight loss. Patients had been randomly assigned to receive megestrol acetate liquid suspension 800 mg/day + placebo tablets, or oral dronabinol tablets 2.5 mg b.i.d. + liquid placebo, or both agents. Other testing included serial physician-reported weight and patient-reported appetite and global quality of life. We found no significant differences in 1-month changes in serum IL-6 according to whether patients had been treated with megestrol acetate, dronabinol, or the combination: the mean differences +/- standard deviation were -1.52+/-4.7 pg/ml, -0.62+/-3.5 pg/ml, and -0.2+/-3.1 pg/ml, respectively (P=0.40, by one-way ANOVA). Among the patients who noted alterations in their appetite over 1 month, we observed no significant changes in IL-6. Finally, changes in serum IL-6 were not associated with shifts in weight or global quality of life. Our investigation provides no evidence that megestrol acetate down-regulates IL-6 in patients with cancer-associated anorexia and weight loss.

Synthesis and evaluation of 6-methylene-bridged uracil derivatives. Part 1: discovery of novel orally active inhibitors of human thymidine phosphorylase.

A series of novel 6-methylene-bridged uracil derivatives have been prepared as inhibitors of human thymidine phosphorylase (TP). To enhance the in vivo antitumor activity of fluorinated pyrimidine 2'-deoxyribonucleosides such as 2'-deoxy-5-(trifluoromethyl)uridine (F(3)dThd), a potent TP inhibitor preventing their degradation to an inactive compound, has become a target of medicinal chemistry. We present here the synthesis and evaluation of novel human TP inhibitors. Introduction of an N-substituted aminomethyl side chain at the 6-position of 5-chlorouracil has improved water solubility and enhanced inhibitory activity compared with the known TP inhibitor, 6-amino-5-chlorouracil. Compound 42 was reasonably well absorbed in mice after oral administration. When combined with F(3)dThd, compound 42 exerted its TP inhibitory potency by increasing the maximum plasma concentrations of the former as evidenced in experiments with monkeys. Positive changes in pharmacokinetic profile were accompanied by the enhanced in vivo antitumor activity of this combination when compared to F(3)dThd alone, in mice bearing human tumor xenografts. Both biochemical and pharmacological effects appeared to fit the concept as anticipated.

Synthesis and evaluation of 6-methylene-bridged uracil derivatives. Part 2: optimization of inhibitors of human thymidine phosphorylase and their selectivity with uridine phosphorylase.

A series of novel 6-methylene-bridged uracil derivatives have been optimized for clinical use as the inhibitors of human thymidine phosphorylase (TP). We describe their synthesis and evaluation. Introduction of a guanidino or an amidino group enhanced the in vitro inhibitory activity of TP comparing with formerly reported inhibitor 1. Their selectivity for TP based on uridine phosphorylase inhibitory activity was also evaluated. Compound 2 (TPI) has been selected for clinical evaluation based on its strong TP inhibition and excellent modulation of 2'-deoxy-5-(trifluoromethyl)uridine (F(3)dThd) pharmacokinetics. As a result, TAS-102 (a combination of F(3)dThd and TPI) is currently in phase 1 clinical studies.