Hiroshi Eto

Overexpression of Bcl-2 in bladder cancer cells inhibits apoptosis induced by cisplatin and adenoviral-mediated p53 gene transfer

To investigate the effects of the expression of Bcl-2 protein in bladder cancer on the apoptosis induced by cisplatin or adenoviral-mediated p53 gene (Ad5CMV-p53) transfer, we transfected the bcl-2 gene into KoTCC-1, a human bladder cancer cell line that does not express the Bcl-2 protein. The Bcl-2-transfected KoTCC-1 (KoTCC-1/B) exhibited significantly higher resistance to both cisplatin and Ad5CMV-p53 transfer than did either the parental KoTCC-1 (KoTCC-1/P) or the vector-only transfected cell line (KoTCC-1/C). The flow cytometric analysis of the propidium iodide-stained nuclei and DNA fragmentation analysis after cisplatin or Ad5CMV-p53 treatment revealed DNA degradation in both KoTCC-1/P and KoTCC-1/C, whereas KoTCC1/B showed a marked inhibition of DNA degradation. Following the treatment with cisplatin or Ad5CMV-p53, the accumulation of p53 protein was highly detectable for a long period in KoTCC-1/B compared to that in KoTTC-1/P and KoTCC-1/C. Furthermore, the cisplatin and Ad5CMV-p53 treatments each reduced the volume of the subcutaneous tumors established in nude mice formed by KoTCC-1/P or KoTCC-1/C; in contrast, their reductive effects on the tumors formed by KoTCC-1/B were significantly suppressed. The intraperitoneal tumor cell implantation model revealed that the prognoses of mice injected with KoTCC-1/B were significantly inferior to those of the mice injected with either KoTCC-1/P or KoTCC-1/C after treatment with cisplatin or Ad5CMV-p53. These findings suggest that the expression of Bcl-2 in bladder cancer cells interferes with the therapeutic effects of cisplatin and Ad5CMV-p53 through the inhibition of the apoptotic pathway.

Basic fibroblast growth factor regulates matrix metalloproteinases production and in vitro invasiveness in human bladder cancer cell lines.

PURPOSE This study was designed to investigate the effect of endogenous basic fibroblast growth factor (FGF-2) on matrix metalloproteinases (MMPs) production and in vitro invasive potential of human bladder cancer cell lines. MATERIALS AND METHODS The human bladder cancer cell lines, HT1376 and KoTCC-1, were used in this study. The mRNA for FGF receptor has been shown to be expressed in both cell lines; the mRNA for FGF-2 is expressed in only KoTCC-1. The effects of FGF-2 expression on HT1376 by gene transfection and those of FGF-2 antisense oligonucleotides treatment on KoTCC-1 were analyzed by zymography and in vitro tumor cell invasion assay. RESULTS The introduction of human FGF-2 gene into HT1376 cells markedly enhanced both the MMP-2 and MMP-9 production, and the in vitro invasive potential was also increased. In contrast, the exposure of KoTCC-1 cells to FGF-2 specific antisense oligonucleotides decreased the MMP-2 production and in vitro invasive potential, but the exposure to FGF-2 sense oligonucleotides did not. CONCLUSIONS These findings suggest that FGF-2 plays an important role in the invasive process of human bladder cancer in part through the regulation of MMPs production.

Detection of Micrometastases in Pelvic Lymph Nodes in Patients Undergoing Radical Cystectomy for Focally Invasive Bladder Cancer by Real-time Reverse Transcriptase-PCR for Cytokeratin 19 and Uroplakin II

Purpose: The objective of this study was to clarify the significance of micrometastases in pelvic lymph nodes in patients who underwent radical cystectomy for bladder cancer. Experimental Design: We included 40 patients with locally invasive bladder cancer who underwent radical cystectomy and pelvic lymphadenectomy. Expression of cytokeratin 19 (CK19), uroplakin II (UP II), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in 760 lymph nodes were assessed by a fully quantitative real-time reverse transcription-PCR (RT-PCR) assay. The quantification value of CK19 or UP II mRNA was described as each value relative to GAPDH mRNA. In this study, we regarded specimen in which either CK19 or UP II mRNA was positive as “presence of micrometastasis.” Results: Routine pathologic examinations detected tumor cells in 29 lymph nodes from six patients. Real-time RT-PCR identified positive expression of CK19 and UP II mRNAs in 49 lymph nodes from 10 patients and 98 lymph nodes from 16 patients, respectively. Of 633 lymph nodes from 34 patients with no pathologic evidence of nodal involvement, 13 nodes from five patients and 58 nodes from 10 patients were diagnosed as positive for CK19 and UP II mRNAs expression, respectively, by real-time RT-PCR. Presence of micrometastases was significantly associated with other conventional prognostic variables, including pathologic stage and microvascular invasion. Disease recurrence was occurred in eight patients, among whom four patients were negative for lymph node metastasis by routine pathologic examination and diagnosed as having micrometastasis by real-time RT-PCR assay. Furthermore, cause-specific survival rate in patients without micrometastasis was significantly higher than that in those with micrometastasis, irrespective of the presence of pathologic-positive nodes. Conclusions: Approximately 30% of locally invasive bladder cancer shed cancer cells to pelvic lymph nodes, and disease recurrence after radical cystectomy could be explained, at least in part, by micrometastases in pelvic lymph nodes.

MULTIFOCAL RENAL CELL CARCINOMA IN JAPANESE PATIENTS WITH TUMORS WITH MAXIMAL DIAMETERS OF 50 MM. OR LESS

PURPOSE We determined the risk of local recurrence in 64 Japanese patients a median of 69 years old with renal cell carcinoma who were possible candidates for nephron sparing surgery and who underwent radical nephrectomy. MATERIALS AND METHODS A total of 64 kidneys in which tumors 50 mm. or less were resected were prospectively examined pathologically in 3 mm. sections. The incidence of satellite tumors and the relationship between the pathological findings of the primary and satellite tumors were evaluated. RESULTS Satellite tumors were identified in 10 of the 64 kidneys (15.6%), a rate similar to that reported in the United States. The correlation of histological findings between primary and satellite tumors was 70% for tumor grade. Satellite tumor grade was less than that of the primary lesion in 3 cases. In 60% of the specimens with multifocal renal cancer satellite tumors were within 10 mm. of the margin of the primary tumor. At this distance, if partial nephrectomy had been performed, the satellite lesions would have been missed in 4 of these 10 patients (40%). Of the 10 kidneys with satellite renal tumors 8 (80%) had vascular invasion of the primary tumor. Multiple logistic regression analysis demonstrated that vascular invasion was a significant predictor of multifocality of renal cell carcinoma. CONCLUSIONS Our results suggest that vascular invasion is a risk factor for multifocality in Japanese patients with renal cell carcinoma. Therefore, careful and long-term followup is necessary in patients with renal cell carcinoma who have undergone nephron sparing surgery, especially those with vascular invasion of the primary tumor.

Quantitative Detection of Micrometastases in Pelvic Lymph Nodes in Patients with Clinically Localized Prostate Cancer by Real-time Reverse Transcriptase-PCR

Purpose: Routine pathologic examination can miss micrometastatic tumor foci in the lymph nodes of patients with prostate cancer, resulting in confusion during tumor staging and clinical decision-making. The objective of this study was to clarify the significance of micrometastases in pelvic lymph nodes in patients who underwent radical prostatectomy for prostate cancer. Experimental Design: The expression of prostate-specific antigen (PSA) and prostate-specific membrane antigen (PSMA) in 2,215 lymph nodes isolated from 120 patients with clinically localized prostate cancer was assessed by a fully quantitative real-time reverse transcriptase-PCR. We regarded specimens in which either PSA or PSMA mRNAs were positive as proof of the “presence of micrometastasis.” Immunohistochemical staining of lymph node specimens with an antibody against PSA was also done. Results: Pathologic examinations detected tumor cells in 29 lymph nodes from 11 patients, and real-time reverse transcriptase-PCR further identified micrometastasis in 143 lymph nodes from 32 patients with no pathologic evidence of lymph node involvement. The presence of micrometastatic cancer cells was confirmed by immunohistochemical staining in 61 lymph nodes from 17 patients with pathologically negative lymph nodes. The presence of micrometastases was significantly associated with other conventional prognostic variables, including serum PSA value, pathologic stage, Gleason score, and tumor volume. Biochemical recurrence was detected in 32 patients, 17 of whom were negative for lymph node metastasis by pathologic examination (including 4 patients with pathologically organ-confined disease), but were diagnosed as having micrometastasis. Biochemical recurrence–free survival rate in patients without micrometastasis was significantly higher than in those with micrometastasis irrespective of the presence of pathologically positive nodes. Furthermore, only the presence of micrometastasis was independently associated with biochemical recurrence regardless of other factors examined. Conclusions: These findings suggest that ∼30% of clinically localized prostate cancers shed cancer cells to the pelvic lymph nodes, and that biochemical recurrence after radical prostatectomy could be explained, at least in part, by micrometastases in pelvic lymph nodes.

OVER EXPRESSION OF CD44V8-10 IN URINARY EXFOLIATED CELLS AS AN INDEPENDENT PROGNOSTIC PREDICTOR IN PATIENTS WITH UROTHELIAL CANCER

PURPOSE CD44 is a widely expressed cell surface adhesion molecule, of which various isoforms arise from alternative RNA splicing mechanisms. Over expression of specific CD44 splice variants, namely CD44v8-10, is evident in various malignant tumors and is considered to be associated with disease progression. In this study, we investigated whether the transcriptional level of CD44v8-10 relative to that of the standard CD44 isoform would predict the extent and prognosis of urothelial cancer. MATERIALS AND METHODS The CD44v8-10- to -standard CD44 ratio was measured in the tissue (40 urothelial cancer specimens and corresponding normal urinary tissue) and spontaneously voided urine samples of 150 patients with urothelial cancer and 50 with benign urological disease by reverse transcriptase-polymerase chain reaction using the set of primers capable of amplifying all CD44 splice variant isoforms. RESULTS Initially any CD44 variant isoforms were barely detectable in normal urinary tissues, whereas CD44v8-10 was predominantly expressed in most urothelial cancer specimens. Furthermore, the CD44v8-10- to -standard CD44 ratio in urothelial cancer was closely associated with tumor progression. We then compared the ratio in urothelial cancer tissue and urinary exfoliated cells, and noted a linear and significant correlation of these 2 values in the same patients. Therefore, we investigated whether the CD44v8-10- to -standard CD44 ratio in urinary exfoliated cells would predict the prognosis and disease progression. The mean ratio in the urinary exfoliated cells of patients with invasive urothelial cancer was significantly higher than in those with superficial urothelial cancer. Of the patients with superficial bladder cancer disease-free survival rate of those with an elevated versus a normal ratio was significantly lower. Moreover, of the patients with advanced urothelial carcinoma who underwent complete resection disease-free survival of those with an elevated CD44v8-10- to -standard CD44 ratio was significantly lower than that of patients with a normal ratio. CONCLUSIONS These results indicate that CD44v8-10 is strongly expressed in tumor tissue and evident at high levels in urinary exfoliated cells of patients with invasive versus superficial urothelial cancer. An elevated CD44v8-10- to -standard CD44 ratio in urinary exfoliated cells may serve as a novel prognostic predictor and indicator of disease extent in patients with urothelial cancer.

Clinical outcome of maximum androgen blockade using flutamide as second‐line hormonal therapy for hormone‐refractory prostate cancer

To investigate the efficacy of maximum androgen blockade (MAB) using flutamide as second‐line hormonal therapy for advanced hormone‐refractory prostate cancer (HRPC).

A comparison of the biological features between prostate cancers arising in the transition and peripheral zones

To investigate differences in the biological features of prostate cancer according to the zonal origin.

Predictors of prostate cancer on repeat transrectal ultrasound-guided systematic prostate biopsy.

Abstract Background: We analyzed the outcome of repeated transrectal ultrasound (TRUS)‐guided systematic prostate biopsy in Japanese men whose clinical findings were suspected of prostate cancer after previous negative biopsies.

Microscopic venous invasion in renal cell carcinoma as a predictor of recurrence after radical surgery

Background: The objective of the present study was to investigate the significance of microscopic venous invasion (MVI) as a prognostic factor for patients with renal cell carcinoma (RCC) who underwent radical surgery.