Nobuyuki Sekita

Epigenetic Regulation of Androgen Receptor Gene Expression in Human Prostate Cancers

Epigenetic mechanisms including DNA methylation and histone deacetylation are thought to play important roles in gene transcriptional inactivation. Heterogenous expression of androgen receptor (AR), which appears to be related to variable responses to endocrine therapy in prostate cancer (PCa) may also be due to epigenetic factors. The methylation status of the 5′ CpG island of the AR in 3 prostate cancer cell lines and 10 primary and 14 hormone-refractory PCa samples was determined using the bisulfite PCR methods. In DU145, CpG-rich regions of the AR were hypermethylated. By an immunohistochemical analysis, only one PCa sample had no AR expression, the others being heterogenous. Bisulfite sequencing and methylation-specific PCR analysis showed aberrant methylation of AR 5′-regulatory region in 20% of 10 primary and 28% of 14 hormone-refractory PCa samples. To clarify the effect of epigenetic regulation on AR expression, we treated three prostate cancer cell lines with a demethylating agent, 5-aza-2′-deoxycytidine (azaC), and a histone deacetylase inhibitor, Trichostatin A (TSA). In DU145, re-expression of AR mRNA was detected after treatment with azaC and/or TSA. Our results suggest that epigenetic regulations including CpG methylation and histone acetylation may play important roles in the regulation of the AR.

Are alpha-blockers involved in lower urinary tract dysfunction in multiple system atrophy? A comparison of prazosin and moxisylyte.

Lower urinary tract dysfunction is a major cause of morbidity in patients with multiple system atrophy (MSA). alpha1-Adrenergic receptors are present in the proximal urethra where impaired relaxation may be responsible for voiding difficulty and a large amount of residual urine. An open study was designed to evaluate whether the blockade of these receptors by prazosin (a nonselective alpha1 blocker) and moxisylyte (an alpha1A-selective blocker) would improve bladder emptying in patients with MSA. Post-micturition residual volumes and clinical symptoms of 49 patients with MSA were evaluated at trial entry and after 4 weeks (prazosin; n=21 and moxisylyte; n=28). The respective means for the prazosin and moxisylyte groups were 38.1% and 35.2% reductions in residual urine volume (P<0.05), and there was lessening of urinary symptoms. Side effects due to orthostatic hypotension were seen in 23.8% of the prazosin group but in only 10.7% of the moxisylyte group. These effects were common in patients with postural hypotension of more than -30 mmHg at trial entry (P<0.05). Modulation of alpha1-receptors may function in the management of lower urinary tract dysfunction in MSA.

Incidence of inguinal hernia after prostate surgery: Open radical retropubic prostatectomy versus open simple prostatectomy versus transurethral resection of the prostate

Our objective was to determine the incidence of inguinal hernia (IH) after surgery for prostatic diseases. Medical records of 395 patients who underwent radical retropubic prostatectomy (RRP; n = 155), open simple prostatectomy (OP; n = 35), or transurethral resection of the prostate (TURP; n = 205) at the Chibaken Saiseikai Narashino Hospital from April 2000 to March 2007 were retrospectively evaluated. The incidence of IH was 23.9% in the RRP group, 18.9% in the OP group, and 2% in the TURP group. Overall, 91.9% in the RRP and 83.3% in the OP group developed an IH within 2 years postoperatively. The laterality of IH after open surgery was mainly on the right side. Subclinical IH were seen in 25% of RRP cases. The existence of subclinical IH was the only significant risk factor for postoperative IH in this analysis. Furthermore, OP and RRP procedures significantly increased the risk of postoperative IH compared with TURP. The hernia‐free ratios were significantly lower after RRP and OP than after TURP (vs RRP: P < 0.001; vs OP: P < 0.001). Our findings confirm that a lower abdominal incision itself is associated with postoperative IH in patients undergoing prostate surgery. Attention must be paid to pre‐existing subclinical IH through careful preoperative assessment. Patients should be followed for more than 2 years due to the high incidence of postoperative IH.

Epigenetic Regulation of the KAI1 Metastasis Suppressor Gene in Human Prostate Cancer Cell Lines

Expression of the KAI1 gene, a metastasis‐suppressor for prostate cancer, is reduced in all foci of prostatic metastasis. The altered regulatory mechanism is not strongly related to mutations or allelic losses of the KAI1 gene in prostate tumors. Since transcriptional silencing of genes has been found to be caused by epigenetic mechanisms, we have investigated the involvement of this epigenetic regulation of KAI1 expression in prostate cancers. The methylation status of the KAI1 promoter region was examined by restriction‐enzyme digestion and sequencing, after amplifying a 331‐bp fragment in the GC‐rich promoter region from 4 human prostate cancer cell lines treated with bisulfite. The same 4 cell lines were also exposed to various concentrations of the demethylating agent, 5‐aza‐2‐deoxycytidine (5‐AzaC) and/or the histone deacetylase inhibitor, trichostatin A (TSA). To clarify the influence of epigenetic modification on reduced KAI1 mRNA expression in the tumor cells, RT‐PCR and northern‐blot analyses were performed. Bisulfite‐sequencing data showed a few methylated CpG islands in the promoter. RT‐PCR analysis of 5‐AzaC and/or TSA‐treated cells indicated reversal of suppression of KAI1 transcription in two cell lines (PC‐3 and DU‐145), although the expression could not be detected by northern blots. From these results, it is suggested that epigenetic change is not the main mechanism of KAI1 down‐regulation, though there remains a possibility that methylation in a more upstream region might be associated with this regulation.

Implications of Serum Bone Turnover Markers in Prostate Cancer Patients With Bone Metastasis

OBJECTIVES To assess the diagnostic accuracy of serum bone turnover markers for detection of bone metastasis in patients with prostate cancer (PCa) and to assess the usefulness of these markers as predictors of mortality from PCa. METHODS Serum total alkaline phosphatase, bone-specific alkaline phosphatase, carboxy-terminal pyridinoline cross-linked telopeptide parts of type-I collagen (1CTP), tartrate-resistant acid phosphatase type 5 b, and prostate-specific antigen (PSA) levels were measured in 222 patients (58 with bone metastasis, 57 with T2M0 PCa, 55 with T3M0 PCa, and 52 without PCa). Multivariate stepwise logistic regression analysis was used to identify independent predictors of bone metastasis. Correlation of serum marker levels with bone metastasis was assessed using receiver operating characteristics analysis. Multivariate Cox proportional hazards analysis was used to predict cause-specific survival in PCa patients with bone metastasis. RESULTS Serum total alkaline phosphatase, bone-specific alkaline phosphatase, 1CTP, tartrate-resistant acid phosphatase type 5 b, and PSA levels were significantly elevated in patients with bone metastasis, and correlated significantly with the extent of disease on bone scintigraphy. Multivariate stepwise logistic regression analysis demonstrated that serum PSA and 1CTP were significant predictors of bone metastasis. Receiver operating characteristics analyses showed that serum 1CTP level was the most reliable predictor of bone metastasis (area under the curve = 0.85). Multivariate Cox proportional hazards analysis revealed that only serum 1CTP was an independent prognostic factor for PCa-related death. CONCLUSIONS Serum 1CTP level was a more reliable marker than the others to detect bone metastatic spread and to predict survival probability in PCa patients with bone metastasis.

External validation and head‐to‐head comparison of Japanese and Western prostate biopsy nomograms using Japanese data sets

The objective of this study was to perform external validation of a previously developed prostate biopsy nomogram (the CHIBA nomogram) and to compare it with previously published nomograms developed in Japanese and overseas populations. Two different cohorts of patients were used: one from the Chiba Cancer Center (n = 392) in which transperineal 16‐core biopsy was performed, and another from Chibaken Saiseikai Narashino Hospital (n = 269) in which transrectal 16‐core biopsy was carried out. All patients were Japanese men with serum prostate‐specific antigen levels less than 10 ng/mL. The predictive accuracy of our CHIBA nomogram and of four other published nomograms (Finnes sextant biopsy‐based logistic regression model, Karakiewiczs sextant biopsy‐based nomogram, Chuns 10‐core biopsy‐based nomogram and Kawakamis three‐dimensional biopsy‐based nomogram) was quantified based on area under the curve derived from receiver operating characteristic curves. Head‐to‐head comparison of area under the curve values demonstrated that our nomogram was significantly more accurate than all other models except Chuns (P = 0.012 vs Finnes, P = 0.000 vs Karakiewiczs, and P = 0.003 vs Kawakamis). Our nomogram appears to be more useful for the Japanese population than Western models. Moreover, external validation demonstrates that its predictive accuracy does not vary according to biopsy approach. This is the first report to demonstrate that the predictive accuracy of a nomogram is independent from the biopsy method.

Prostate adenocarcinoma producing syndrome of inappropriate secretion of antidiuretic hormone

Abstract The syndrome of inappropriate secretion of antidiuretic hormone (ADH) was recognized in a 68‐year‐old man with a poorly differentiated metastatic adenocarcinoma of the prostate. Elevated levels of ADH were found in the tissues of the primary tumor and lymph node metastasis. The patients clinical course is detailed and the pathophysiology of this syndrome is discussed. To our knowledge, this case is the ninth reported case of syndrome of inappropriate secretion of ADH with adenocarcinoma of the prostate. Antidiuretic hormone activity was proven in only three cases including this case.

Overexpression of TIAP/m-survivin in thymocytes enhances cell proliferation

TIAP/m-survivin, a member of the inhibitor of apoptosis (IAP) protein family, is expressed in a cell cycle dependent manner. It is strongly expressed in various subsets of thymocytes. To investigate a role of TIAP/m-survivin in thymocytes, mice carrying the lck-TIAP transgene were established. Two out of six transgenic mice expressed large amounts of TIAP mRNA and protein in thymocytes. Although T cell development and apoptosis of thymocytes were largely unaffected in lck-TIAP mice, transgenic thymocytes displayed hyperproliferation in response to PMA and ionomycin but not to anti-CD3 antibody. Thus, overexpression of TIAP/m-survivin augments cell proliferation of thymocytes to a certain stimulation.

Increased in situ gelatinolytic activity in renal cell tumor tissues correlates with tumor size, grade and vessel invasion.

Degradation of collagen, or gelatinolysis, by tumor cells is one of the most important events in tumorigenesis. We investigate the possible relationship between the in situ gelatinolytic activities exerted by matrix metalloproteinases (MMPs) and clinico‐pathological factors in renal cell tumor (RCT) patients. Using the film in situ zymography (FIZ) method, we determined in situ localization of MMP‐like gelatinolytic activities in cancerous and normal tissues in the kidney (n = 51). To clarify the MMP(s) responsible for the gelatinolytic activity in RCTs, we examined the expressions of MMP‐2 and MMP‐9 in the kidney tissues by means of gelatin zymography (GZG). MMP expression was also detected by RT‐PCR and Western blotting analysis. We then investigated the associations of MMP expression, as detected by GZG, with the intensity of gelatinolytic activity, as determined by FIZ. We analyzed the possible relationship of FIZ findings to several clinico‐pathological factors such as tumor size, grade, vessel invasion, histologic type, stage and metastasis. FIZ demonstrated that all tumor and normal kidney tissues showed in situ gelatinase activities, and that gelatinolytic activities in RCTs were much stronger than those of normal kidney tissues. There was a statistically significant correlation between the intensity of MMP‐like gelatinolytic activity and tumor size, tumor grade and vessel invasion (p < 0.05), but not between it and histological type, tumor stage or metastatic status. FIZ showed that tumor tissues in 5 of the 6 patients with fatal outcome exhibited the intense gelatinolytic pattern. Stronger in situ gelatinolytic patterns were documented in cases with higher MMP‐2 expression. The molecular species of MMPs detected by GZG were confirmed by RT‐PCR and Western blotting analysis. The FIZ technique enables a direct assessment of in situ gelatinolytic activity in RCT tissues. The intensity of the activity seems to affect the biology of RCT tissues. Our results also indicate a major role for MMP‐2 in in situ gelatinolysis in RCT tissues.

External validation and comparison of two nomograms predicting the probability of Gleason sum upgrading between biopsy and radical prostatectomy pathology in two patient populations: a retrospective cohort study

The aim of this study is to validate and compare the predictive accuracy of two nomograms predicting the probability of Gleason sum upgrading between biopsy and radical prostatectomy pathology among representative patients with prostate cancer. We previously developed a nomogram, as did Chun et al. In this validation study, patients originated from two centers: Toho University Sakura Medical Center (n = 214) and Chibaken Saiseikai Narashino Hospital (n = 216). We assessed predictive accuracy using area under the curve values and constructed calibration plots to grasp the tendency for each institution. Both nomograms showed a high predictive accuracy in each institution, although the constructed calibration plots of the two nomograms underestimated the actual probability in Toho University Sakura Medical Center. Clinicians need to use calibration plots for each institution to correctly understand the tendency of each nomogram for their patients, even if each nomogram has a good predictive accuracy.