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Objectives: Muscarinic M3 (M3) receptors mediate cholinergic smooth muscle contraction of the bladder. Current drugs targeting bladder M3 receptors for micturition disorders have a risk of cholinergic side effects due to excessive receptor activation and insufficient selectivity. We investigated the effect of ASP8302, a novel positive allosteric modulator (PAM) of M3 receptors, on bladder function in rats. Methods: Modulation of carbachol‐induced increases in intracellular Ca2+ was assessed in cells expressing rat muscarinic receptors. Potentiation of bladder contractions was evaluated using isolated rat bladder strips and by measuring intravesical pressure in anesthetized rats. Conscious cystometry was performed to investigate the effects on residual urine volume and voiding efficiency in rat voiding dysfunction models induced by the α1‐adrenoceptor agonist midodrine and muscarinic receptor antagonist atropine, and bladder outlet obstruction. To assess potential side effects, the number of stools and tracheal insufflation pressure were measured in conscious and anesthetized rats, respectively. Results: ASP8302 demonstrated PAM effects on the rat M3 receptor in cell assays, and augmented cholinergic bladder contractions both in vivo and in vitro. ASP8302 improved voiding efficiency and reduced residual urine volume in two voiding dysfunction models as effectively as distigmine bromide, but unlike distigmine bromide did not affect the number of stools or tracheal insufflation pressure. Conclusions: Our results in rats indicate that ASP8302 improves voiding dysfunction by potentiating bladder contraction with fewer effects on cholinergic responses in other organs, and suggest a potential advantage over current cholinomimetic drugs for treating micturition disorders caused by insufficient bladder contraction. [ABSTRACT FROM AUTHOR]

Background: Mitochondrial dysfunction contributes to many types of human disorders and cancer progression. Inner membrane mitochondrial protein (IMMT) plays an important role in the maintenance of mitochondrial structure and function. The aims of this study were to examine IMMT expression in lung adenocarcinoma and evaluate its correlation with clinicopathological parameters and patient prognosis. Methods: IMMT expression was immunohistochemically studied in 176 consecutive lung adenocarcinoma resection tissues, and its correlations with clinicopathological parameters were evaluated. Kaplan‐Meier survival analysis and Cox‐proportional hazards models were used to estimate the effect of IMMT expression on survival. Results: High‐IMMT expression was detected in 84 of 176 (47.7%) lung adenocarcinomas. Levels were significantly correlated with advanced disease stage (stage II and III; P = 0.024), larger tumor size (>3 cm; P = 0.002), intratumoral vascular invasion (P < 0.001), and poorer adenocarcinoma patient prognosis (P = 0.002). Based on 176 patients with adenocarcinoma, multivariate analysis revealed that IMMT expression was an independent predictor of poorer survival (HR, 1.99; 95% confidence interval [CI], 1.06–3.74; P = 0.031). Further, treating A549 cells derived from lung adenocarcinoma, with IMMT siRNA resulted in significantly decreased proliferation. Conclusion: Here, we first demonstrated that high‐IMMT expression is related to some clinicopathological parameters, and that its expression is an independent prognostic predictor of poorer survival in patients with lung adenocarcinoma; further studies are required to clarify the biological function of IMMT in lung adenocarcinoma. However, results suggest that this protein could be a novel prognostic indicator and therapeutic target. [ABSTRACT FROM AUTHOR]

We conducted a large‐scale surveillance study as a post–marketing commitment to investigate the safety and effectiveness of alectinib in patients with ALK‐positive non–small‐cell lung cancer (NSCLC) in Japan. Patients receiving 300 mg twice‐daily alectinib (September 2014 to June 2015) were monitored until termination of alectinib or completion of 18 months of treatment at 519 Japanese study sites. The primary endpoint was the incidence of adverse drug reactions (ADR), which are important identified risks for alectinib in Japanese patients. Overall survival (OS), a key secondary endpoint, was assessed according to information on outcome. Overall, 1251 patients were enrolled. The median patient age was 62.0 years; 12.9% of patients were aged ≥75 years. At baseline, 63.0% of patients had received crizotinib and 40.6% had brain metastases. Altogether, 1512 ADR occurred in 654 patients (53.6%), with 164 grade ≥3 ADR in 123 patients (10.1%). Commonly occurring ADR were hepatic disorders (all grades, 19.8%; grade ≥3, 2.0%), decreased neutrophil and/or white blood cell count (all grades, 7.6%; grade ≥3, 1.1%), and interstitial lung disease (all grades, 3.8%; grade ≥3,.7%). Median OS was not estimable. The 18‐month cumulative OS rate was longer in patients with ECOG performance status ≤1 (vs 2 or ≥3; 83.7% vs 44.5% or 27.2%), without prior crizotinib (vs with; 81.1% vs 73.4%), receiving first‐line alectinib (vs second and third or later line; 83.0% vs 79.2% or 71.9%), without brain metastases (vs with; 79.5% vs 71.5%). These data confirm the favorable safety and effectiveness of alectinib in patients with ALK‐positive NSCLC in Japan. The highly selective ALK inhibitor alectinib is approved as the standard of care for advanced anaplastic lymphoma kinase (ALK)‐positive non–small‐cell lung cancer (NSCLC). This large‐scale surveillance study was implemented as part of a post–marketing commitment to investigate the safety and effectiveness of alectinib in patients with ALK‐positive NSCLC in Japan. Data from this real‐world study confirm the favorable safety and effectiveness profile of alectinib in Japanese patients with ALK‐positive NSCLC. [ABSTRACT FROM AUTHOR]

We established the KU‐Lu‐8 monoclonal antibody (MoAb) using a lung cancer cell line as an antigen and a random immunization method. The KU‐Lu‐8 MoAb recognizes basigin (BSG), which is a transmembrane‐type glycoprotein that is strongly expressed on the cell membranes of lung cancer cells. This study aimed to clarify the relationships between BSG expression and clinicopathological parameters and determine the prognostic significance of BSG expression in pulmonary adenocarcinoma (AC) patients. To evaluate the significance of BSG expression in lung cancer, we immunohistochemically analyzed 113 surgically resected pulmonary adenocarcinomas, and the associations between BSG expression and various clinicopathological parameters were evaluated. Kaplan‐Meier survival analysis and Cox proportional hazards models were used to investigate the effects of BSG expression on survival. Clinicopathologically, BSG expression was significantly associated with tumor differentiation, vascular invasion, lymphatic invasion, and a poor prognosis. In particular, BSG expression was significantly correlated with poorer survival in patients with stage I AC. The high BSG expression group (compared with the low BSG expression group) exhibited adjusted hazard ratios for mortality of 4.694. BSG expression is indicative of a poor prognosis in AC patients, particularly in those with stage I disease. [ABSTRACT FROM AUTHOR]

Abstract: Objectives: To clarify the role of bradykinin in urogenital pain, we investigated bradykinin involvement in rat models of testicular pain. Methods: Bradykinin (0.1, 0.3, 1, 3 and 10 mmol/L) or distilled water was injected into the testes of male Wistar rats, and induced pain behaviors in conscious rats were evaluated. The effect of pretreatment with bradykinin B2 receptor antagonist FK3657 on bradykinin‐induced pain behavior was then examined. We also evaluated the analgesic effect of FK3657 in a rat acetic acid‐induced testicular pain as well as changes in the intratesticular bradykinin concentration after testicular injection of acetic acid. Results: An injection of bradykinin into the testes of conscious rats induced pain behaviors that were dose‐proportionally reduced by prior administration of FK3657. In addition, FK3657 dose‐dependently inhibited the pain responses induced by testicular injection of 1% acetic acid. An increase in intratesticular bradykinin concentration was detected after the testicular injection of 1% acetic acid. Conclusions: Here, we found that intratesticular bradykinin evokes pain behavior via stimulation of bradykinin B2 receptors and that intratesticular acetic acid injection induces intratesticular bradykinin synthesis, consequently leading to pain behavior. These findings suggest that the potential utility of bradykinin B2 receptor antagonists as a novel target for treating urogenital pain. [ABSTRACT FROM AUTHOR]

Objective We investigated the role of bradykinin in urethral function by examining contractile responses in urethral smooth muscle strips isolated from humans and the intraurethral pressure in rats and dogs. Methods The contractile responses of human urethral tissue for bradykinin (0.01-10 µmol/L) were examined, and changes in intraurethral pressure induced by bradykinin (0.003-10 µg/kg) in anesthetized rats or dogs were measured. In addition, the effects of pretreatment with the bradykinin B2 receptor antagonist FK3657 were also examined. Results In smooth muscle strips obtained from human urethra, bradykinin induced contraction, which was inhibited by FK3657 in a concentration-dependent manner. In anesthetized rats and dogs, intravenously administered bradykinin dose-dependently increased intraurethral pressure. FK3657 shifted the intraurethral pressure dose-response curve for bradykinin to the right in rats. The bradykinin-induced elevation of intraurethral pressure was also dose-dependently inhibited by FK3657 in dogs. Conclusions The present study provides evidence that bradykinin elicits urethral smooth muscle contraction via the bradykinin B2 receptor, suggesting the potential utility of this receptor as a novel target for the treatment of voiding dysfunction. [ABSTRACT FROM AUTHOR]

Objectives We investigated the effect of the selective prostaglandin E2 EP2 receptor agonist CP-533,536 on voiding efficiency in rats with midodrine-induced functional urethral obstruction. Methods The effect of CP-533,536 (0.03-0.3 mg/kg, intravenous [i.v.]) on urethral perfusion pressure ( UPP) was investigated in anesthetized rats pre-treated with midodrine (1 mg/kg, i.v.), which forms an active metabolite that acts as an α1-adrenoceptor agonist. The effect of CP-533,536 (0.03-0.3 mg/kg, i.v.) on cystometric parameters was also investigated in anesthetized rats. In addition, the effect of CP-533,536 (0.03-0.3 mg/kg, i.v.) on residual urine volume ( RV) and voiding efficiency ( VE) was investigated in conscious rats treated with midodrine (1 mg/kg, i.v.). Results CP-533,536 dose-dependently decreased UPP elevated by midodrine in anesthetized rats. In contrast, CP-533,536 did not affect maximum voiding pressure, intercontraction interval, or intravesical threshold pressure. In conscious rats, midodrine (1 mg/kg, i.v.) markedly increased RV and reduced VE. CP-533,536 dose-dependently ameliorated increases in RV and decreases in VE induced by midodrine. Conclusions These results suggest that a selective EP2 receptor agonist could ameliorate the elevation of RV and improve the reduction of VE in rats with functional urethral obstruction caused by stimulation of α1-adrenoceptors. The mechanism of action might be not potentiation of bladder contraction but rather preferential relief of urethral constriction. [ABSTRACT FROM AUTHOR]

Objectives We investigated the relaxant effect of stimulation of prostaglandin E2 ( PGE2) receptor subtype EP2 as well as the involvement of a cyclic AMP ( cAMP)-dependent pathway related to stimulation of EP2 receptors in urethral function in rats by evaluating effects of PGE2 and selective EP2 receptor agonist CP-533,536. Methods Effects of PGE2 and CP-533,536 on cAMP accumulation were assessed in Chinese hamster ovary ( CHO)- K1 cells expressing rat EP2 or EP4 receptors. Relaxant responses to PGE2 and CP-533,536 (0.01-10 µmol/L) in rat urethral tissue pre-contracted with 10 µmol/L phenylephrine were evaluated, and cAMP levels in isolated rat urethral tissue treated with these compounds were determined as well. The effects of PGE2 and CP-533,536 (0.003-0.3 mg/kg intravenously) on urethral perfusion pressure ( UPP) in anesthetized rats were also evaluated. Results PGE2 concentration-dependently increased the accumulation of cAMP in cells expressing rat EP2 ( EC50 value = 1.3 nmol/L) and EP4 receptors ( EC50 value = 17 nmol/L). While CP-533,536 similarly increased the accumulation of cAMP in cells expressing rat EP2 receptors ( EC50 value = 3.0 nmol/L), no such effects were noted in cells expressing rat EP4 receptors up to 10 µmol/L. Both PGE2 and CP-533,536 produced relaxation and increased cAMP levels in urethral tissues in a concentration-dependent manner. PGE2 and CP-533,536 both dose-dependently decreased UPP in anesthetized rats. Conclusions Taken together, these results suggest that stimulation of EP2 receptors induces relaxation likely via activation of cAMP-dependent mechanisms in rat urethral tissue, leading to a reduction of UPP. [ABSTRACT FROM AUTHOR]

Objectives: To clarify the effect of cAMP on the Ca2+ -sensitized smooth muscle contraction in human detrusor, as well as the role of novel exchange protein directly activated by cAMP (Epac) in cAMP-mediated relaxation. Materials and Methods: All experimental protocols to record isometric tension force were performed using a-toxin-permeabilized human detrusor smooth muscle strips. The mechanisms of cAMP-mediated suppression of Ca2+ sensitization activated by 10 lM carbachol (CCh) and 100 lM GTP were studied using a selective rho kinase (ROK) inhibitor, Y-27632, and a selective protein kinase C (PKC) inhibitor, GF-109203X. The relaxation mechanisms were further probed using a selective protein kinase A (PKA) activator, 6-Bnz-cAMP and a selective Epac activator, 8-pCPT-20-O-Me-cAMP. Results: We observed that CCh-induced Ca2+ sensitization was inhibited by cAMP in a concentration-dependent manner. GF-109203X (10 μM) but not Y-27632 (10 μM) significantly enhanced the relaxation effect induced by cAMP (100 μM). 6-Bnz-cAMP (100 μM) predominantly decreased the tension force in comparison with 8-pCPT-20-O-Me-cAMP (100 μM). Conclusions: We showed that cAMP predominantly inhibited the ROK pathway but not the PKC pathway. The PKA-dependent pathway is dominant, while Epac plays a minor role in human detrusor smooth muscle Ca2+ sensitization. [ABSTRACT FROM AUTHOR]

Objectives To clarify the pathophysiological factor underlying neural crosstalk among pelvic organs, we investigated the possible role of nerve growth factor ( NGF) in the neural crosstalk between the testes and urinary bladder. Methods Nerve growth factor (10, 30, and 100 µg/mL) or saline was injected into the testes of male Wistar rats. The change in bladder capacity via cystometry and duration of spontaneous scratching behavior induced by NGF in conscious rats was measured. The effects of pretreatment with capsaicin on NGF-induced changes in bladder capacity and behavior were examined. Further, we evaluated the effect of analgesics, indomethacin and morphine, and pretreatment with compound 48/80 on NGF-induced scratching behavior to elucidate the mechanism of the behavior. Results Injection of saline into the testes had no effect on bladder capacity or behavior. However, an injection of NGF (30 and 100 µg/mL) reduced bladder capacity, which was regarded as bladder overactivity, and evoked scratching behavior in a dose-dependent manner. Pretreatment with capsaicin inhibited NGF-induced bladder overactivity and scratching behavior. Neither indomethacin nor pretreatment with compound 48/80 affected the scratching behavior, but morphine inhibited the behavior. Conclusions The present study provides evidence of a possible new role of NGF in the testes regarding the activation of testicular primary afferent neurons mediated by capsaicin-sensitive C-fibers, which evokes bladder overactivity via neural crosstalk between the testes and the urinary bladder as well as testicular pain. [ABSTRACT FROM AUTHOR]

The aim of the present study was to clarify protein profiling in small cell lung carcinoma (SCLC) and pulmonary large cell neuroendocrine carcinoma (LCNEC). The proteomic approach was used, and involved cell lysate from two cell lines (N231 derived from SCLC and LCN1 derived from LCNEC), with 2-D gel electrophoresis (2-DE). In the present study, 25 protein spots with greater than twofold quantitative differences between LCN1 and N231 cells on 2-DE gels were confirmed. Within the 25 identified proteins, cytokeratins (CK) 7, 8, 18 and 19 were upregulated in LCN1 cells compared with N231 cells. The expression of CK7, 8, 18, and 19 was further studied on immunohistochemistry with 81 formalin-fixed and paraffin-embedded pulmonary carcinomas, which included 27 SCLC, 30 LCNEC, 14 adenocarcinomas, and 10 squamous cell carcinomas. Although the expression of CK7, 8, 18, and 19 was observed in all histological types, the mean immunostaining scores of CK7, 8, 18, and 19 were significantly higher in LCNEC than in SCLC ( P < 0.001, P < 0.001, P < 0.01 and P < 0.001, respectively). These data suggest that the biological characteristics of LCNEC and SCLC may be different and the expression of CK may serve as differential diagnostic markers. [ABSTRACT FROM AUTHOR]

Tamibarotene (TM411) is a synthetic retinoic acid receptor-α/-β selective retinoid that is chemically more stable than all- trans retinoic acid. This study was designed to evaluate the activity of TM411 in multiple myeloma (MM) and the effects of TM411 combined with a glucocorticoid (GC). In vitro, five human myeloma cells were treated with TM411 alone, GC alone, or TM411 + GC. Cell survival was analyzed by the tetrazolium dye assay and the Hoechst 33342/propidium iodide double-staining method. The effect of TM411 + GC was assessed by the isobologram method. In vivo, the growth-inhibitory effects of the drugs on RPMI-8226 cell xenografts established in SCID mice were examined. The effects of the agents on IL-6-mediated signaling pathways were also analyzed by Western blotting. TM411 was 2- to 10-fold more potent, in terms of its growth-inhibitory effect, than all- trans retinoic acid. The combination of TM411 and GC was found to show a markedly synergistic interaction. While increased expressions of the IL-6 receptor, phosphorylated MAPK, and Akt were observed after exposure to GC, TM411 attenuated this increase in the expressions, suggesting that such modification of the effect of GC by TM411 might be the possible mechanism underlying the synergistic interaction. Furthermore, TM411 + GC showed a supra-additive inhibitory effect in a xenograft model as compared with TM411 or GC alone. These results imply that the combination of TM411 + GC might be highly effective against MM, and suggest the need for clinical evaluation of TM411 + GC for the treatment of MM. ( Cancer Sci 2009; 100: 1137–1143) [ABSTRACT FROM AUTHOR]

Aims The role of Ih channels at the spinal level in the control of bladder function was examined in urethane anesthetized female rats. Methods Bladder activity was recorded via a transurethral catheter under isovolumetric conditions or via an intravesical catheter during continuous infusion cystometry. ZD7288, a selective Ih channel inhibitor, was administered intrathecally at L6-S1 segmental levels of the spinal cord. Results Under isovolumetric conditions, intrathecal 0.3 µg of ZD7288 was inactive, whereas 1 and 3 µg abolished bladder activity for 5.6 ± 1.8 and 20.7 ± 3.9 min, respectively. Three micrograms also tended to decrease the amplitude of the contractions when they reappeared. During continuous cystometry, an intrathecal injection of ZD7288 at 0.3-3 µg dose-dependently increased the intercontraction intervals by 7.5%, 33.1%, and 57.5% from pre-drug values. A higher dose of ZD7288 (3 µg) tended to increase the pressure threshold for inducing micturition by 16.6% and the maximum voiding pressure by 11.1%. On the other hand, cardiovascular parameters such as heart rate and mean blood pressure were not affected by 0.3-3 µg of intrathecal ZD7288. Conclusions These results indicate that Ih channels play an important role in the control of micturition. Because Ih channel inhibition in the lumbosacral spinal cord reduced the frequency of the micturition reflex without significantly affecting the amplitude of reflex bladder contraction, Ih channels might preferentially be involved in afferent processing in the spinal cord to control the micturition reflex. Neurourol. Urodynam. 27:838-844, 2008, © 2008 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

In lung cancer, somatic mutations of epidermal growth factor receptor ( EGFR) are concentrated in exons 18–21, especially in adenocarcinoma (Ad), but these mutations have rarely been reported in small cell lung carcinoma (SCLC). Combined SCLC is rare, and the EGFR mutation status and its relationship to the clinicopathological features of this tumor type have not yet been elucidated. We retrospectively studied six patients with combined SCLC with Ad components among 64 consecutive patients who underwent resection of SCLC. The clinicopathological features of each patient were reviewed, especially for the distribution pattern of the Ad component and lymph node metastases. EGFR mutations were screened by high-resolution melting analysis in each case, and were confirmed by sequencing of each mutation in the microdissected SCLC or Ad components. Regarding EGFR, no specific mutation was detected in five of the six patients, whereas one female patient who had never smoked had a missense mutation. In this case, both the SCLC and Ad components shared the same mutation in exon 21 (L858R). We identified a patient with combined SCLC with Ad sharing an identical EGFR mutation in both the SCLC and Ad components. In addition to the clinicopathological characteristics of this rare histological type of lung cancer, these findings provide useful information for better understanding the biology, natural history and clinical management of SCLC. ( Cancer Sci 2007; 98: 1714–1719) [ABSTRACT FROM AUTHOR]

Background and Aims: Animal models of heart failure (HF) are useful to clarify the mechanism and to develop therapeutic interventions. To produce an easy ischemic HF model, we induced myocardial infarction (MI) in pigs by placing a tube in the coronary artery. Methods: Twelve pigs underwent echocardiography and were randomly allocated to the myocardial infarction group (MI group) and the control group. In the MI pigs, a 4.2 F nylon tube was placed via the carotid artery in the left circumflex coronary (LCx) artery to induce MI. Three months thereafter, thoracotomy was performed in the both groups and left ventricular (LV) pressure–volume relation was evaluated. Results: Body weight, LV dimension and function did not differ in the baseline state between the two groups. Three months after the tube placement, LV diameter was larger (47±3 vs. 42±2 mm) and its fractional shortening was lower in the MI group than the control group. In addition, aortic flow was decreased, LV ejection fraction was decreased (25±5 vs. 52±6%) and LV diastolic pressure was elevated (14±3 vs. 8±2 mmHg) in the MI group compared with the control group. The extent of MI was 26±5% of the LV in the MI pigs. Conclusion: The method of placing a tube in the coronary artery does not need thoracotomy or an additional procedure and enables the production of an ischemic HF model of pigs. [Copyright &y& Elsevier]