Aki Iwai

Glucocorticoids Suppress Tumor Angiogenesis and In vivo Growth of Prostate Cancer Cells

Purpose: Glucocorticoids, such as prednisone, hydrocortisone, and dexamethasone, are known to produce some clinical benefit for patients with hormone-refractory prostate cancer (HRPC). However, the underlying mechanisms by which glucocorticoids affect HRPC growth are not well established as yet. Here, we hypothesize that the therapeutic effect of glucocorticoids on HRPC can be attributed to a direct inhibition of angiogenesis through the glucocorticoid receptor by down-regulating two major angiogenic factors, vascular endothelial growth factor (VEGF) and interleukin-8 (IL-8). Experimental Design: The effects of dexamethasone on VEGF and IL-8 expression and cell proliferation were examined using DU145, which expresses glucocorticoid receptor. The effects of dexamethasone on DU145 xenografts were determined by analyzing VEGF and IL-8 gene expression, microvessel density, and tumor volume. Results: Dexamethasone significantly down-regulated VEGF and IL-8 gene expression by 50% (P < 0.001) and 89% (P < 0.001), respectively, and decreased VEGF and IL-8 protein production by 55% (P < 0.001) and 74% (P < 0.001), respectively, under normoxic condition. Similarly, hydrocortisone down-regulated VEGF and IL-8 gene expression. The effects of dexamethasone were completely reversed by the glucocorticoid receptor antagonist RU486. Even under hypoxia-like conditions, dexamethasone inhibited VEGF and IL-8 expression. In DU145 xenografts, dexamethasone significantly decreased tumor volume and microvessel density and down-regulated VEGF and IL-8 gene expression, whereas dexamethasone did not affect the in vitro proliferation of the cells. Conclusion: Glucocorticoids suppressed androgen-independent prostate cancer growth possibly due to the inhibition of tumor-associated angiogenesis by decreasing VEGF and IL-8 production directly through glucocorticoid receptor in vivo.

p51/p63 Controls Subunit α3 of the Major Epidermis Integrin Anchoring the Stem Cells to the Niche

p51/p63, a member of the tumor suppressor p53 gene family, is crucial for skin development. We describe here identification of ITGA3 encoding integrin α3 as a target of its trans-activating function, proposing that p51/p63 allows epidermal stem cells to express laminin receptor α3β1 for anchorage to the basement membrane. When activated by genotoxic stress or overexpressed ectopically in non-adherent cells, p51/p63 transduced a phenotype to attach to extracellular matrices, which was accompanied by expression of ITGA3. Motifs matching the p53-binding consensus sequence were located in a scattered form in intron 1 of human ITGA3, and served as p51/p63-responsive elements in reporter assays. In addition to the trans-activating ability of the TA isoform, we detected a positive effect of the ΔN isoform on ITGA3. The high level α3 production in human keratinocyte stem cells diminished upon elimination of p51/p63 by small interfering RNA or by Ca2+-induced differentiation. Furthermore, a chromatin immunoprecipitation experiment indicated a physical interaction of p51/p63 with intron 1 of ITGA3. This study provides a molecular basis for the standing hypothesis that p51/p63 is essential for epidermal-mesenchymal interactions.

Down-regulation of vascular endothelial growth factor in renal cell carcinoma cells by glucocorticoids.

Metastatic renal cell carcinomas (RCC) remain highly resistant to systemic therapy. RCCs are highly vascular tumors, which overproduce angiogenic peptides such as vascular endothelial growth factor (VEGF) even under normoxic conditions. A potential suggested role of antiangiogenic therapeutic strategies is the treatment of RCC by inhibiting VEGF production. The down-regulation of VEGF expression by glucocorticoids has recently been demonstrated in several cells. In this study, the direct effects of glucocorticoids on VEGF production by RCC cells were evaluated. Four RCC cell lines A498, RCC270, Caki1, and ACHN were treated with dexamethasone (DEX), hydrocortisone (HC), 5-alpha-dihydrotestosterone (DHT), or estradiol (E2). RU486 was used as a glucocorticoid receptor (GR) antagonist. Cell growth was studied with MTS assays. VEGF mRNA and protein were evaluated with quantitative real-time RT-PCR and ELISA, respectively, and GR expression was examined using RT-PCR and immunocytochemistry. All four RCC cell lines expressed GR. DEX at 100 nM down-regulated VEGF secretions by more than 50% in three lines (A498, RCC270, and Caki1) and had a weak inhibitory effect on ACHN cells. The effect of DEX on reducing VEGF mRNA levels in A498 cells was concentration-dependent and maximal at 100 nM (80% inhibition). HC had similar but weaker effects on VEGF production in the RCC cells, but E2 and DHT had no effect. RU486 reversed the effects of DEX. DEX at 1-1000 nM did not affect cell growth in any of the four RCC cell lines. This is the first study showing that glucocorticoids, at concentrations achievable in vivo by oral administration of low doses of DEX, have an inhibitory effect on VEGF mRNA expression and protein secretion of RCC cells possibly through the GR pathway. Furthermore, DEX might have a potential role in antiangiogenic therapeutic strategies by inhibiting VEGF production during metastatic RCC treatment.

Glucocorticoids Suppress Tumor Lymphangiogenesis of Prostate Cancer Cells

Purpose: Glucocorticoids such as prednisone, hydrocortisone, and dexamethasone are known to provide some clinical benefit for patients with hormone-refractory prostate cancer. However, the underlying mechanisms by which glucocorticoids affect hormone-refractory prostate cancer progression are not well established as yet. Our previous study has shown that glucocorticoids inhibit tumor angiogenesis possibly by down-regulation of vascular endothelial growth factor (VEGF) and interleukin 8. Here, we hypothesized that the therapeutic effect of dexamethasone on hormone-refractory prostate cancer can be partly attributed to a direct inhibition of lymphangiogenesis through the glucocorticoid receptor by down-regulating a major lymphangiogenic factor, VEGF-C. Experimental Design: The effects of dexamethasone on the expression of VEGF-C and its receptor, VEGF receptor-3 (VEGFR-3), were examined using an androgen-independent human prostate cancer cell line, DU145, which expresses glucocorticoid receptor. The effects of dexamethasone on tumor-associated lymphangiogenesis in DU145 xenografts were determined by analyzing VEGF-C gene expression, lymphatic vessel density, and relative lymphatic vessel area. Results: Dexamethasone significantly down-regulated VEGF-C gene expression and protein production by 48% (P = 0.003) and 44% (P = 0.002), respectively, under normoxic condition. Similarly, hydrocortisone down-regulated VEGF-C gene expression. The effects of dexamethasone were completely reversed by the glucocorticoid receptor antagonist RU486. Even under hypoxia-like conditions, dexamethasone inhibited VEGF-C gene expression. In DU145 xenografts, dexamethasone significantly down-regulated VEGF-C gene expression and decreased lymphangiogenesis. Dexamethasone did not affect VEGFR-3 gene expression in vitro and in vivo. Conclusion: Glucocorticoids suppressed tumor-associated lymphangiogenesis by down-regulating VEGF-C through glucocorticoid receptor in androgen-independent prostate cancer cells in vivo.

Suppression of VEGF transcription in renal cell carcinoma cells by pyrrole-imidazole hairpin polyamides targeting the hypoxia responsive element

Hypoxia inducible factor (HIF), a master regulator of critical genes for cell survival under hypoxic conditions, is known to be related to tumorigenesis and progression of renal cell carcinoma. N-methylpyrrole (Py)-N-methylimidazole (Im) hairpin polyamides are synthetic organic compounds that recognize and bind to the minor grooves of specific DNA sequences. We synthesized three Py-Im hairpin polyamides targeting the flanking sequences of hypoxia responsive element (HRE; a binding site of HIF) in the promoter region of the vascular endothelial growth factor (VEGF) gene. The effects of the polyamides on HIF-induced transcription were evaluated by a luciferase assay using a reporter plasmid containing a VEGF promoter. Real time reverse-transcriptase polymerase chain reaction and enzyme-linked immunosorbent assay were performed to examine the effects of the polyamides on the transcription and secretion of VEGF in A498 renal cell carcinoma cells, which have a frame-shift mutation in the von Hippel-Lindau gene. A combination of three Py-Im hairpin polyamides suppressed HIF-induced transcription in reporter assays using 293 cells and successfully suppressed transcription and translation of the VEGF gene in A498 cells. Inhibition of the HIF-HRE interaction was confirmed by an electrophoresis mobility shift assay. An approach using Py-Im hairpin polyamides may be a new strategy for the treatment of renal cell carcinoma.

Simple prophylactic procedure of inguinal hernia after radical retropubic prostatectomy: isolation of the spermatic cord.

Abstract:  To reduce the incidence of inguinal hernia (IH) after radical retropubic prostatectomy (RRP), a simple prophylactic procedure was carried out during RRP. A consecutive 82 patients who had undergone RRP for clinically localized prostate cancer between July 2002 and October 2006 at Toride Kyodo General Hospital were enrolled. From July 2002 to November 2003, 20 patients underwent conventional RRP. Thereafter, 62 patients underwent conventional RRP with blunt dissection of the peritoneum at the internal inguinal ring and isolation of the spermatic cord from the peritoneum as a prophylactic procedure for IH. There was no significant difference in patient characteristics between the two groups. In the conventional RRP group, IH occurred in 10 patients during a median range follow‐up period of 41 (1 to 73) months. In contrast, in the RRP plus prophylactic procedure group, IH occurred in one patient (1.6%) during a median range follow‐up period of 41 (25 to 59) months. The incidence of IH after RRP plus the prophylactic procedure was significantly lower than that after conventional RRP, indicating the efficacy of the presented procedure.

Perioperative Complications of Radical Cystectomy After Induction Chemoradiotherapy in Bladder-sparing Protocol Against Muscle-invasive Bladder Cancer: A Single Institutional Retrospective Comparative Study with Primary Radical Cystectomy

OBJECTIVE To compare rates of early morbidity after radical cystectomy in patients treated with or without induction chemoradiotherapy (CRT) using a standardized reporting methodology. METHODS All 193 consecutive patients undergoing radical cystectomy for bladder cancer between 1989 and 2010 were retrospectively reviewed. Induction chemoradiotherapy consists of radiation at 40 Gy to the small pelvis and two cycles of concurrent cisplatin at 20 mg/day for 5 days. Deaths within 90 days after radical cystectomy and complications arising within 30 days were recorded and graded according to the Clavien-Dindo classification. Grades 1-2 were considered minor; Grades 3-5 were considered major. RESULTS Eighty-seven patients underwent radical cystectomy following chemoradiotherapy (chemoradiotherapy group) while the remaining 106 primarily underwent radical cystectomy (no chemoradiotherapy group). No Grade 4-5 complication was observed. Overall, 118 patients (61%) experienced 36 major and 122 minor complications. There was no significant difference in the incidence of overall complications between the chemoradiotherapy and no chemoradiotherapy groups (67 vs. 57%). Overall urinary anastomosis-related complications and major gastrointestinal complications, most of which were Grade 3 ileus, were more frequent in the chemoradiotherapy group than the no chemoradiotherapy group (11 vs. 2%, P = 0.007; and 14 vs. 4%, P = 0.02; respectively). Multivariate analysis identified induction chemoradiotherapy as an independent risk factor for overall urinary anastomosis-related complications (relative risk 6.0, P = 0.01) but not for major gastrointestinal complications. CONCLUSIONS Induction chemoradiotherapy at 40 Gy in bladder-sparing protocols against MIBC is unlikely to increase the rate of severe complications of radical cystectomy.

A Novel Repeat Biopsy Nomogram Based on Three-dimensional Extended Biopsy

OBJECTIVES To develop a nomogram based on a cohort examined with 3-dimensional (3D) protocol for diagnosis of prostate cancer on repeat biopsy. METHODS Of 4074 consecutive men undergoing prostate biopsy at our institutions between 2000 and 2009, 775 men with at least 1 previous negative biopsy underwent repeat biopsy with a 3D protocol. Men with previous atypical glands or atypical small acinar proliferation and/or without available prostate-specific antigen (PSA) kinetics information were excluded. The remaining 515 men constituted the study cohort. We developed a logistic regression-based nomogram with 70% of the cohort selected randomly; we validated it with the remaining 30%. Predictive accuracy and performance characteristics were assessed using the area under the receiver operating characteristic curve (AUC) and calibration plots, respectively. The threshold probability was evaluated with decision curve analysis. RESULTS We developed a novel repeat biopsy nomogram incorporating age, free to total PSA ratio, prostate volume, history of previous extended biopsy, and PSA doubling time. Validation confirmed predictive accuracy with an AUC value of 0.791. Calibration plots showed good agreement. The decision curve of the nomogram was superior to the decision curve of biopsying all men in a range of threshold probability over 0.15. At the threshold of 0.2, the number of unnecessary biopsies could be reduced by 10 per 100, without missing prostate cancer. CONCLUSIONS We developed a novel repeat biopsy nomogram based on a cohort examined with 3D protocol. This repeat biopsy nomogram is clinically beneficial, preventing a substantial number of unnecessary biopsies.