Katsuki Tsuchiyama

Expression of MicroRNAs associated with Gleason grading system in prostate cancer: miR-182-5p is a useful marker for high grade prostate cancer

Expression profiles of some microRNAs (miRNAs) were associated with clinicopathological findings in human prostate cancer (PC), but the relative expression of miRNAs among Gleason patterns (GPs) remains unclear. In this study, we investigated the expression of several known microRNAs in each GP of PC.

Regulation of microRNA expression by hepatocyte growth factor in human head and neck squamous cell carcinoma

Hepatocyte growth factor (HGF) is a multifunctional molecule that acts as mitogen, motogen, and/or morphogen in a variety of cells. MET, a specific receptor tyrosine kinase for HGF, is upregulated in various tumors including squamous cell carcinoma of the human head and neck (HNSCC), but how HGF affects the expression of downstream functional genes has not yet been elucidated in detail. In the present study, we examined the expression of microRNA (miRNA), non‐coding small RNA that regulate cell proliferation and functions by interfering with the translation of target mRNA, with or without HGF stimulation in HNSCC cell line HSC3. Among several miRNAs, in which the expression was altered after HGF stimulation, we focused on miR‐200c and miR‐27b, both of which were drastically downregulated after HGF stimulation. Expression of ZEB1, a target mRNA for miR‐200c, was upregulated 3 and 6 h after HGF stimulation, and that of E‐cadherin, a downstream molecule of ZEB1, was downregulated 12 h after HGF stimulation. Expression of ST14/matriptase, an enzyme for extracellular matrix (ECM) degradation and HGF activation and a target mRNA for miR‐27b, was drastically upregulated in the protein level after HGF stimulation, although it was not statistically altered in the mRNA level. These results suggest that miR‐200c and miR‐27b downregulated by HGF might play an important role in epithelial–mesenchymal transition mediated by ZEB1/E‐cadherin and ECM degradation and HGF autoactivation mediated by ST14/matriptase, respectively. Altered expression of miRNA directly regulated by HGF might contribute enhanced progressive and invasive characteristics of HNSCC by regulating the translation of HGF‐induced functional molecules. (Cancer Sci 2011; 102: 2164–2171)

IPSS is lower in hypertensive patients treated with angiotensin-II receptor blocker: Posthoc analyses of a lower urinary tract symptoms population†‡

The existence of Angiotensin‐II (Ang‐II) receptors in the bladder wall and the pronounced contractile effect of Ang‐II on the human detrusor muscle have been well established. Studies have presented the role of Ang‐II as a mediator in smooth muscle growth and collagen production in the bladder with outlet obstruction. We investigated the associations between male lower urinary tract symptoms (LUTS) and hypertension (HT), and examined whether the medications used for HT treatment, particularly Ang‐II receptor blockers (ARBs) influence LUTS.

Arterial Transit Time-corrected Renal Blood Flow Measurement with Pulsed Continuous Arterial Spin Labeling MR Imaging

Purpose: The importance of arterial transit time (ATT) correction for arterial spin labeling MRI has been well debated in neuroimaging, but it has not been well evaluated in renal imaging. The purpose of this study was to evaluate the feasibility of pulsed continuous arterial spin labeling (pcASL) MRI with multiple post-labeling delay (PLD) acquisition for measuring ATT-corrected renal blood flow (ATC-RBF). Materials and Methods: A total of 14 volunteers were categorized into younger (n = 8; mean age, 27.0 years) and older groups (n = 6; 64.8 years). Images of pcASL were obtained at three different PLDs (0.5, 1.0, and 1.5 s), and ATC-RBF and ATT were calculated using a single-compartment model. To validate ATC-RBF, a comparative study of effective renal plasma flow (ERPF) measured by 99mTc-MAG3 scintigraphy was performed. ATC-RBF was corrected by kidney volume (ATC-cRBF) for comparison with ERPF. Results: The younger group showed significantly higher ATC-RBF (157.68 ± 38.37 mL/min/100 g) and shorter ATT (961.33 ± 260.87 ms) than the older group (117.42 ± 24.03 mL/min/100 g and 1227.94 ± 226.51 ms, respectively; P < 0.05). A significant correlation was evident between ATC-cRBF and ERPF (P < 0.05, r = 0.47). With suboptimal single PLD (1.5 s) settings, there was no significant correlation between ERPF and kidney volume-corrected RBF calculated from single PLD data. Conclusion: Calculation of ATT and ATC-RBF by pcASL with multiple PLD was feasible in healthy volunteers, and differences in ATT and ATC-RBF were seen between the younger and older groups. Although ATT correction by multiple PLD acquisitions may not always be necessary for RBF quantification in the healthy subjects, the effect of ATT should be taken into account in renal ASL–MRI as debated in brain imaging.

Urethral Sensations are Related to the Development of Detrusor Overactivity

Urgency is the core symptom of the overactive bladder symptom complex, but the underlying mechanisms are not fully understood. Clinical findings have led to the assumption that bladder outlet obstruction (BOO) caused by benign prostatic enlargement (BPE) induces storage symptoms and detrusor overactivity. Presumably, BOO by BPE accounts for urgency; however, urgency is not always caused by BOO. Sensory nerves in the wall of the urethra fire in response to urethral fluid flow, and this activity initiates bladder contractions in the quiescent bladder and augments ongoing contractions in the active bladder. In humans, prostatic urethral anesthesia results in significant increases in bladder capacity among BPH patients without neurological diseases, therefore sensory stimuli from an anatomically altered prostatic urethra has the possibility to induce urgency and detrusor overactivity. Studies in animals demonstrate the basis for an excitatory urethra to bladder reflex. Urethral stimulation by prostaglandin E2 induces an excitatory effect on micturition reflex by activation of C‐fiber afferent nerves. α1A‐adrenoceptor blocker has an inhibitory effect on the micturition reflex, suggesting excitatory urethra to bladder reflex is mediated by α1A‐adrenoceptor. Even if there is no obstruction, increase in urethral sensory due to BPE may induce the development of the detrusor overactivity.

MP29-12 PRECLINICAL ASSESSMENT OF EARLY THERAPEUTIC EFFECT OF SUNITINIB ON RENAL CELL CARCINOMA USING 18F-FLUOROTHYMIDINE POSITRON EMISSION TOMOGRAPHY IN A XENOGRAFT MODEL

vaccines. Clinical and immunologic results vary. We have recently shown that genomic profiling of peripheral blood mononuclear cells (PBMC) may predict response. Transcription factors play a key role in the myelopoesis of monocytes and DC. We hypothesized that the profile of key transcription factors plays a key role in the efficacy of DC vaccines. METHODS: Monocytes from RCC patients and healthy donors were isolated via cold agglutination or ELUTRA. For some experiments, DC were matured with GM-CSF/ IL-4 and a variety of maturation protocols (TNF-alpha, cytokine cocktail, Flt-3).RNA was extracted using QIAGEN RNeasy Mini Kit. It’s concentration and quality was assessed using Nanodrop (Thermo Scientific) and Bioanalyzer (Agilent Technologies). Only high quality RNA (RIN > 7.2) was used for reverse transcription (Bio-Rad iScript cDNA synthesis kit) and real time qPCR (in triplicates using ABI SYBR-Select Master Mix with target specific primers and the ABI 7900HT Thermocycler). The expression of key transcription factors IRF-4, IRF-8, IL-10, IL-12 were analyzed using previously reported primer pairs. To determine relative concentrations, cycle threshold (Ct) values were compared to the reference gene GAP-DH. The statistical analysis was conducted in JMP 11 from SAS. RESULTS: Healthy donors demonstrated a significant variation of expression levels of all transcription factors examined. When DC were matured with a variety of maturation protocols, these resulted in significant differences in expression of all transcription factors. The greatest variations were observed in the expression levels of IRF-4, IL10 and IL-12. The greatest induction of IRF-4 and IL-12 and lowest induction of IL-10 were observed with the cytokine cocktail. In an IFNgamma release assay, these cells also demonstrated the greatest T cell stimulatory capacity. RCC patient monocytes showed showed significantly lower expression of IRF-4 and IRF-8 than healthy age-adjusted donor monocytes (p1⁄40.004 and 0.0214, respectively). Immunotherapy resulted in upregulation of IL-10 when compared to pretreatment mRNA levels (p1⁄40.007). CONCLUSIONS: In summary, this is the first study to assess the profile of key transcription factors as predictors for DC vaccine quality. We demonstrate significant inter-individual variability in monocytes prior to DC vaccine and throughout the generation of DC vaccines. These data may lead to improved cellular anti-cancer vaccines.

Tumour multiplicity as a risk factor for the development of bladder tumours after primary upper urinary tract cancer

Objective: To determine the independent risk factors for intravesical tumour recurrence in patients with primary urothelial cancer of the upper urinary tract. Patients and methods: Of the 60 patients who underwent nephroureterectomy for urothelial cancer of the upper urinary tract, the data from 49 patients were retrospectively reviewed. Patients with a previous history or concomitance of bladder cancer were excluded from the study. Multivariate analysis by Cox’s proportional hazards model was used to determine independent risk factors for intravesical tumour recurrence. Results: Of the 49 patients reviewed, 22 (44.9%) experienced subsequent intravesical tumour recurrence during a mean follow-up period of 26 months (range 3–103). On multivariate analysis, tumour multiplicity had a statistically significant impact on the risk of intravesical tumour recurrence (P < 0.01). Conclusion: Neither the pathology of the upper urinary tract cancers nor the method of treatment was associated with recurrent bladder cancers. Only tumour multiplicity had a significant impact on the incidence of intravesical tumour recurrence.

Abstract 5033: Relationship between microRNA expression and Gleason grading system in prostate cancer

Background: Recent studies have shown that some microRNA (miRNA) expression profiles are associated with clinicopathological findings, including “Gleason score (GS)” in human prostate cancer (PC). However, the direct relationship between “Gleason pattern (GP)” and miRNA expression remains unclear. In this study, we investigated the miRNA expression profile in each GP (GP 3, GP 4, and GP 5). Methods: Formalin-fixed, paraffin-embedded (FFPE) tissue samples obtained from radical prostatectomy (patient set 1, n = 43, including [GP 3] n = 22, [GP 4] n = 35, and [GP 5] n = 12) and prostate needle biopsy (patient set 2, n = 9, [GP 4] n = 9) were used. Cancer tissues and adjacent normal counterparts were collected separately using laser-captured microdissection with subsequent isolation of total RNA. Real time reverse transcription polymerase chain reaction (RT-PCR) was performed to determine the relative expression of miRNAs, including miR-31, -34c, -96, -182, -183, -205, -221, and miR-222, which are currently reported to be involved in the progression of PC. Results: In the radical prostatectomy samples (patient set 1), relative expression (mean ± SE) of miR-31, miR-34c, and miR-205 in every GP was significantly decreased (miR-31: [GP 3] 0.028 ± 0.007, [GP 4] 0.047 ± 0.017, and [GP 5] 0.071 ± 0.022; miR-34c: 0.208 ± 0.066, 0.419 ± 0.219, and 0.210 ± 0.054; and miR-205: 0.021 ± 0.008, 0.015 ± 0.003, and 0.050 ± 0.023, all p-values Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5033. doi:1538-7445.AM2012-5033

Abstract 3972: Regulation of microRNA expression by hepatocyte growth factorin human head and neck sqaumous cell carcinoma (HNSCC)

Introduction: Hepatocyte growth factor (HGF) is a multifunctional growth factor that acts as mitogen, motogen, and/or morphogen in a variety of cells. MET, a specific receptor tyrosine kinase for HGF, is up-regulated in various tumors including human head and neck squamous cell carcinoma (HNSCC) and its signal transduction induces tumor progression and invasion. However, how HGF affects the expression of downstream functional gene expression has not yet been elucidated in detail. MicroRNAs (miRNAs) are non-coding small RNAs that regulate posttranscriptional gene expression by interfering the translation of target mRNAs. Aberrant expression of miRNA is known to induce various human tumors including HNSCC, but the regulation of miRNA expression is still unknown. In this study, we examined the expression of miRNAs with or without HGF stimulation in human HNSCC cell line HSC3 and the expression of their putative target genes was evaluated. Methods: Human HNSCC cell line HSC3 was cultured with or without HGF for designated periods. Alteration of miRNA expression was examined by miRNA microarray analysis. Validation study of miRNAs showing altered expression was performed by quantitative RT-PCR (qRT-PCR). The expression of putative target mRNAs and their translated proteins were examined by qRT-PCR and western blot analysis, respectively. Results: The expression of several miRNAs including let-7a, miR-16, and miR-205, which are already known as tumor-suppressive miRNAs, was altered after HGF stimulation by miRNA microarray. Among them, we focused on miR-200c and miR-27b which were drastically down-regulated after HGF stimulation, because of their unique target mRNAs involving HGF-mediated functional molecules. The expression of ZEB1, a target mRNA for 200c, was up-regulated 3hr after HGF stimulation and that of E-cadherin, a downstream molecule of ZEB1, was up-regulated 12hr after HGF stimulation. The expression of ST14/matriptase, an activation enzyme of HGF and a target mRNA for miR-27b, was not statistically altered after HGF stimulation, but the protein level was drastically up-regulated 12hr after HGF stimulation. These changes were attenuated in part by the addition of oligo pre-mi27b or pre-miR200c into the culture medium. Conclusion: Our present results suggest that miR-200c down-regulated by HGF might play an important role for epithelial-mesenchymal transition (EMT), through up-regulation of ZEB1 followed by down-regulation of E-cadherin. In addition, it was also suggested that miR-27b down-regulated by HGF might induce auto-activation of HGF in HNSCC. These alterations of miRNA expression by HGF stimulation could be induce the enhanced progressive and invasive characteristics of HNSCC. This is a first report that HGF may directly regulate some miRNAs that affect the expression of HGF-mediated downstream functional molecules. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3972. doi:10.1158/1538-7445.AM2011-3972