Motoshi Kawahara

Multigene Methylation Analysis for Detection and Staging of Prostate Cancer

Purpose: Aberrant gene promoter methylation profiles have been well-studied in human prostate cancer. Therefore, we rationalize that multigene methylation analysis could be useful as a diagnostic biomarker. We hypothesize that a new method of multigene methylation analysis could be a good diagnostic and staging biomarker for prostate cancer. Experimental Design: To test our hypothesis, prostate cancer samples (170) and benign prostatic hyperplasia samples (69) were examined by methylation-specific PCR for three genes: adenomatous polyposis coli (APC), glutathione S-transferase pi (GSTP1), and multidrug resistance 1 (MDR1). The methylation status of representative samples was confirmed by bisulfite DNA sequencing analysis. We further investigated whether methylation score (M score) can be used as a diagnostic and staging biomarker for prostate cancer. The M score of each sample was calculated as the sum of the corresponding log hazard ratio coefficients derived from multivariate logistic regression analysis of methylation status of various genes for benign prostatic hyperplasia and prostate cancer. The optimal sensitivity and specificity of the M score for diagnosis and for staging of prostate cancer was determined by receiver-operator characteristic (ROC) curve analysis. A pairwise comparison was employed to test for significance using the area under the ROC curve analysis. For each clinicopathologic finding, the association with prostate-specific antigen (PSA) failure-free probability was determined using Kaplan-Meier curves and a log-rank test was used to determine significance. The relationship between M score and clinicopathologic findings was analyzed by either the Mann-Whitney U test, Kruskal-Wallis test, or the Spearman rank correlation test. Results: The frequency of positive methylation-specific PCR bands for APC, GSTP1, and MDR1 genes in prostate cancer samples was 64.1%, 54.0%, and 55.3%, respectively. In benign prostatic hyperplasia samples, it was 8.7%, 5.8%, and 11.6%, respectively. There was a significant correlation of M score with high pT category (P < 0.001), high Gleason sum (P < 0.001), high preoperative PSA (P = 0.027), and advanced pathologic features. For all patients, the M score had a sensitivity of 75.9% and a specificity of 84.1% as a diagnostic biomarker using a cutoff value of 1.0. In patients with low or borderline PSA levels (<10.0 ng/mL), the M score was significantly higher in prostate cancers than in benign prostatic hyperplasias (2.635 ± 0.200 and 0.357 ± 0.121, respectively). ROC curve analysis revealed that the M score had a sensitivity of 65.4% and a specificity of 94.2% when 1.0 was used as a cutoff value. For all patients, M score can distinguish organ-confined (≤pT2) from locally advanced cancer (≥pT3) with a sensitivity of 72.1% and a specificity of 67.8%. Moreover, considering patients with PSA levels of <10 ng/mL, the M score has a sensitivity of 67.1% and a specificity of 85.7%. The ROC curve analysis showed a significant difference between M score and PSA (P = 0.010). Conclusions: This is the first report demonstrating that M score is a new method for multigene methylation analysis that can serve as a good diagnostic and staging biomarker for prostate cancer.

CpG Hypermethylation of MDR1 Gene Contributes to the Pathogenesis and Progression of Human Prostate Cancer

Multidrug resistance 1 (MDR1) gene encodes for P-glycoprotein (P-gp), a Mr 170,000 transmembrane calcium-dependent efflux pump that is inactivated in prostate cancer. We hypothesize that inactivation of the MDR1 gene through CpG methylation contributes to the pathogenesis and progression of prostate cancer. To test this hypothesis, CpG methylation status of the MDR1 promoter and its correlation with clinicopathological findings were evaluated in 177 prostate cancer samples and 69 benign prostate hypertrophy (BPH) samples. Cellular proliferation index and apoptotic index were determined by proliferating cell nuclear antigen (PCNA) and single-strand DNA immunostaining, respectively. After 5-aza-2′-deoxycytidine treatment, increased expression of MDR1 mRNA transcript was found in prostate cancer cell lines (DU145, DuPro, and ND1). MDR1 methylation frequency was significantly higher in prostate cancer samples compared with BPH samples (54.8 versus 11.6%, respectively, P < 0.001). Logistic regression analysis revealed that PC patients are 11.5 times more likely to have MDR1 methylation than BPH patients (95% confidence interval 4.87–27.0) and that MDR1 methylation is independent of the age. Significant correlation of MDR1 methylation was observed with high pT category (P < 0.001), high Gleason sum (P = 0.008), high preoperative prostate-specific antigen (P = 0.01), and advancing pathological features. In addition, PCNA-labeling index were significantly higher in methylation-specific PCR (MSP)-positive than in MSP-negative prostate cancer samples (P = 0.048). In contrast, no significant difference in apoptotic index was found between MSP-positive and -negative prostate cancer samples. These findings suggest that CpG hypermethylation of MDR1 promoter is a frequent event in prostate cancer and is related to disease progression via increased cell proliferation in prostate cancer cells.

Ethnic group-related differences in CpG hypermethylation of the GSTP1 gene promoter among African-American, Caucasian and Asian patients with prostate cancer

The incidence and mortality of prostate cancer (PC) is approximately 2‐fold higher among African‐Americans as compared to Caucasians and very low in Asian. We hypothesize that inactivation of GSTP1 genes through CpG methylation plays a role in the pathogenesis of PC, and its ability to serve as a diagnostic marker that differs among ethnic groups. GSTP1 promoter hypermethylation and its correlation with clinico‐pathological findings were evaluated in 291 PC (Asian = 170; African‐American = 44; Caucasian = 77) and 172 benign prostate hypertrophy samples (BPH) (Asian = 96; African‐American = 38; Caucasian = 38) using methylation‐specific PCR. In PC cells, 5‐aza‐dC treatment increased expression of GSTP1 mRNA transcripts. The methylation of all CpG sites was found in 191 of 291 PC (65.6%), but only in 34 of 139 BPH (24.5%). The GSTP1 hypermethylation was significantly higher in PC as compared to BPH in each ethnic group (p < 0.0001). Logistic regression analysis (PC vs. BPH) showed that African‐Americans had a higher hazard ratio (HR) (13.361) compared to Caucasians (3.829) and Asian (8.603). Chi‐square analysis showed correlation of GSTP1 hypermethylation with pathological findings (pT categories and higher Gleason sum) in Asian PC (p < 0.0001) but not in African‐Americans and Caucasian PC. Our results suggest that GSTP1 hypermethylation is a sensitive biomarker in African‐Americans as compared to that in Caucasians or Asian, and that it strongly influences tumor progression in Asian PC. Ours is the first study investigating GSTP1 methylation differences in PC among African‐American, Caucasian and Asian.

Smoking influences aberrant CpG hypermethylation of multiple genes in human prostate carcinoma

Aberrant CpG methylation profiles of gene promoters and their correlation with advanced pathologic features have been well investigated in prostate carcinoma (PC). Several case–control and prospective studies have revealed a positive association between current smoking and PC. The authors hypothesized that smoking influences both progression and prognosis of PC through CpG hypermethylation of related genes.

Diagnosis and treatment of priapism: experience with 5 cases

OBJECTIVES Priapism is a rare disease, but needs early intervention and appropriate management. We present 5 cases, 2 of nonischemic high-flow priapism and 3 of ischemic low-flow priapism. METHODS Focusing on the differential diagnosis of priapism between the nonischemic high-flow type and the ischemic low-flow type, we reviewed the medical records of 5 patients. RESULTS Of the examinations carried out, cavernosography, blood gas analysis of cavernosal blood, color Doppler ultrasound, and internal pudendal arteriography were useful in differentiating the type of priapism. Complete detumescence of the penis in 2 cases of high-flow priapism and 3 cases of low-flow priapism was achieved by selective embolization with gelform and by glandular-cavernosal shunting, respectively. No recurrence was observed in any patient, and postoperative erectile function was preserved in 4 patients and is unknown in 1. CONCLUSIONS These results indicate that angiographic studies provide the most reliable information for the differentiation of the type of priapism. However, color flow Doppler ultrasound and cavernosal blood gas determination can obviate the need for angiographic studies and are noninvasive. Although conservative treatment or even expectant management may be feasible with high-flow priapism, aggressive treatment should be carried out for low-flow priapism immediately after initial treatment fails to achieve detumescence of the penis. Selective embolization of the internal pudendal artery may be the treatment of choice for patients with high-flow priapism.

An epidemiologic survey of methicillin-resistant Staphylococcus aureus by combined use of mec-HVR genotyping and toxin genotyping in a university hospital in Japan.

OBJECTIVE To evaluate the usefulness of an assay using two polymerase chain reaction-based genotyping methods in the practical surveillance of methicillin-resistant Staphylococcus aureus (MRSA). METHODS Nosocomial infection and colonization were surveyed monthly in a university hospital in Japan for 20 months. Genotyping with mec-HVR is based on the size of the mec-associated hypervariable region amplified by polymerase chain reaction. Toxin genotyping uses a multiplex polymerase chain reaction method to amplify eight staphylococcal toxin genes. RESULTS Eight hundred nine MRSA isolates were classified into 49 genotypes. We observed differing prevalences of genotypes for different hospital wards, and could rapidly demonstrate the similarity of genotype for outbreak isolates. The incidence of genotype D: SEC/TSST1 was significantly higher in isolates causing nosocomial infections (49.5%; 48 of 97) than in nasal isolates (31.4%; 54 of 172) (P = .004), suggesting that this genotype may represent the nosocomial strains. CONCLUSION The combined use of these two genotyping methods resulted in improved discriminatory ability and should be further investigated.

Endoscopic treatment of a long fibroepithelial ureteral polyp

Abstract A case is reported of a 30‐year‐old woman with a long fibroepithelial polyp in the middle ureter treated with the Ho‐YAG laser endoscopically. She presented with an intermittent macroscopic hematuria and lower abdominal pain lasting for 1 year. The filling defect on urography occupying one‐third of the ureter was migratory depending on the patient position. Transurethral flexible ureterorenoscopy showed a large pedunculated tumor with a small base at the middle ureter. About 1 month after the endoscopic irradiation of the Ho‐YAG laser to the base of tumor, the tumor was spontaneously discharged and pathologic examination revealed it to be a fibroepithelial polyp without malignant component. Postoperatively, the patient remained asymptomatic and follow‐up excretory urographs showed no abnormal findings.

Endoscopic removal of intravesical thermometer using a rigid nephroscope and forceps

Intravesical thermometers are occasionally encountered in urologic practice. In the present paper, we describe the removal of a thermometer from the bladder of a woman who presented with irritable bladder symptoms. The thermometer was removed intact transurethrally using a rigid nephroscope and forceps, even though both ends of the thermometer were embedded in the bladder wall. Our retrieval technique may be of general use in such cases, and it should be attempted before resorting to open surgery.

Renal silica calculi in an infant

Abstract  We report on a rare case of urinary silica calculi in a 10‐month‐old boy. The boy showed acute pyelonephritis with left hydronephrosis. Ultrasonography and computed tomography revealed a calculus at the left ureteropelvic junction and three additional calculi in the left renal pelvis. Because his acute pyelonephritis was refractory to conventional chemotherapy, the patient underwent successful left percutaneous nephrostomy followed by percutaneous nephrolithotripsy for the renal calculi. All stones disappeared and his postoperative course was uneventful. On infrared spectrophotometry, the wavelength pattern of the stones exhibited two peaks at 1100 and 1650 cm−1, consistent with the determination that the calculi consisted of a mixture of silicate (78%) and calcium oxalate (22%). We consider that the etiology of the calculi in this child can be ascribed to the silicate‐rich water used to dilute milk. In Japan, 46 adult patients with urinary silicate calculi have been reported in the literature; however, there is no report of the disease in an infant in Japan.

Intravesical instillation of adriamycin in the presence or absence of verapamil for the treatment of superficial bladder cancer : preliminary report of a collaborative study

SummaryA case-controlled collaborative study on the intravesical administration of Adriamycin in the presence or absence of verapamil, a calcium-channel blocker, as chemotherapy of superficial bladder cancer was carried out at two universities, Okayama and Kagoshima, and their affiliated hospitals. Although little is known about the expression of P-glycoprotein in superficial bladder cancer, it may be a cause of multidrug resistance (MDR). Verapamil was used as an inhibitor of P-glycoprotein. Arm A consisted of Adriamycin given at 50 mg/50 ml saline, and arm B constituted Adriamycin given at 50 mg/40 ml saline plus 5 ampules (10 ml) of injectable verapamil. The drugs were instilled into the bladder for 3 consecutive days in each of 3 consecutive weeks for a total of 9 instillations. No significant difference in antitumor effects was observed between arm A and arm B. Recurrent tumors responded better than did primary tumors to both arm-A and arm-B treatments (P=0.012). In both treatment arms, significant differences (P=0.031) in the response rate were found between tumors with, diameters of <1 cm and those measuring 1–3 cm in diameter. Although the number of evaluable patients was limited, recurrent subjects who had previously received Adriamycin instillations responded in both treatment arms.