Laparoscopic Radical Prostatectomy with a Remote Controlled Robot.

PURPOSE: Robotics in surgery is a recent innovation. This technology offers a number of attractive features in laparoscopy. It overcomes the difficulties with fixed port sites by restoring all 6 degrees of freedom at the instrument tips, provides new possibilities for miniaturization of surgical tasks and allows remote controlled surgery. We investigated the applicability of remote controlled robotic surgery to laparoscopic radical prostatectomy. MATERIALS AND METHODS: Our previous experience with laparoscopic prostatectomy served as a basis for adapting robotic surgery to this procedure. A surgeon at a different location who activated the tele-manipulators of the da Vinci∗ robotic system performed all steps of the intervention. A scrub nurse and second surgeon who stood at patient side had limited roles to port and instrument placement, exposure of the operative field, assistance in hemostasis and removal of the operative specimen. Our patient was a 63-year-old man presenting with a T1c tumor discovered on 1 positive sextant biopsy with a 3+3 Gleason score and 7 ng./ml. preoperative serum prostate specific antigen. RESULTS: The robot provided an ergonomic surgical environment and remarkable dexterity enhancement. Operating time was 420 minutes, and the hospital stay lasted 4 days. The bladder catheter was removed 3 days postoperatively, and 1 week later the patient was fully continent. Pathological examination showed a pT3a tumor with negative margins. CONCLUSIONS: Robotically assisted laparoscopic radical prostatectomy is feasible. This new technology enhances surgical dexterity. Further developments in this field may have new applications in laparoscopic tele-surgery.

Regional copy number-independent deregulation of transcription in cancer.

Genetic and epigenetic alterations have been identified that lead to transcriptional deregulation in cancers. Genetic mechanisms may affect single genes or regions containing several neighboring genes, as has been shown for DNA copy number changes. It was recently reported that epigenetic suppression of gene expression can also extend to a whole region; this is known as long-range epigenetic silencing. Various techniques are available for identifying regional genetic alterations, but no large-scale analysis has yet been carried out to obtain an overview of regional epigenetic alterations. We carried out an exhaustive search for regions susceptible to such mechanisms using a combination of transcriptome correlation map analysis and array CGH data for a series of bladder carcinomas. We validated one candidate region experimentally, demonstrating histone methylation leading to the loss of expression of neighboring genes without DNA methylation.

Occupational exposure to polycyclic aromatic hydrocarbons influenced neither the frequency nor the spectrum of FGFR3 mutations in bladder urothelial carcinoma.

Occupational exposure to polycyclic aromatic hydrocarbons (PAH) is associated with an increased risk of urothelial carcinoma (UC). FGFR3 is found mutated in about 70% of Ta tumors, which represent the major group at diagnosis. The influence of PAH on FGFR3 mutations and whether it is related to the emergence or shaping of these mutations is not yet known. We investigated the influence of occupational PAH on the frequency and spectrum of FGFR3 mutations. We included on 170 primary urothelial tumors from five hospitals from France. Patients (median age, 64 yr) were interviewed to gather data on occupational exposure to PAH, revealing 104 non- and possibly PAH exposed patients, 66 probably and definitely exposed patients. Tumors were classified as follows: 75 pTa, 52 pT1, and 43 > or =pT2. Tumor grades were as follows: 6 low malignant potential neoplasms (LMPN) and 41 low-grade and 123 high-grade carcinomas. The SnaPshot method was used to screen for the following FGFR3 mutations: R248C, S249C, G372C, Y375C, A393E, K652E, K652Q, K652M, and K652T. Occupational PAH exposure was not associated with a particular stage or grade of tumors. Thirty-nine percent of the tumors harbored FGFR3 mutations. After adjustment for smoking, occupational exposure to PAH did not influence the frequency [OR, 1.10; 95% CI, 0.78-1.52], or spectrum of FGFR3 mutations. Occupational exposure to PAH influenced neither the frequency nor the spectrum of FGFR3 mutations and there was no direct relationship between these mutations and this occupational hazard.

The expression of Twist has an impact on survival in human bladder cancer and is influenced by the smoking status.

OBJECTIVES: Twist is considered as transcription factor that regulates epithelial mesenchymal transition (EMT) by at least inhibition of E-cadherin expression. EMT is a key event in the tumor invasion process. The purpose of this study is to investigate the expression of Twist but also those of E- and N-cadherin in human primary bladder tumor and to evaluate its prognostic value. As smoking cigarettes is a strong bladder cancer risk factor, we tried to evaluate the impact of the tobacco status on these molecular abnormalities. MATERIALS AND METHODS: To delineate on the oncogenic role for Twist in human bladder cancer, we evaluated the E- and N-cadherin but also Twist expression (n = 70) by immunohistochemistry. We evaluated the prognostic value of these expressions. Moreover, we tried to correlate these protein expressions to the smoking status of the patients. Overall survival (OS) and progression-free survival (PFS) were evaluated using the Kaplan-Meier method, and multivariate analysis was performed using the Cox proportional hazard analysis. RESULTS: Of the 70 bladder tumors, 28 (40%) cases were positive for Twist expression, 16 (23%) cases were negative for E-cadherin expression, and 12 (17%) were positive for N-cadherin expression. When categorized into negative vs. positive expression, Twist was associated with the stage (P = 0.001), the grade (P < 0.001), the progression (P = 0.02), and the E-cadherin expression (P = 0.01). Moreover, positive Twist expression clearly predicted poorer PFS (P = 0.02). In the multivariate analysis, both positive Twist expression and loss of E-cadherin expression were independent prognostic factors for PFS (P = 0.046 and P = 0.001, respectively) and only loss of E-cadherin expression for the OS (P < 0.001). We also demonstrated that almost 60% (16/28) of patients with Twist-positive expression were current smokers at the time of the diagnosis, corroborating the fact that smoking modulates the expression of EMT markers including Twist. CONCLUSION: Positive Twist expression may be a useful prognostic marker for patients with bladder cancer. Its expression seems to be correlated to the tobacco status of the patients.

Gefitinib inhibits the growth and invasion of urothelial carcinoma cell lines in which Akt and MAPK activation is dependent on constitutive epidermal growth factor receptor activation.

PURPOSE: Abnormally high levels of epidermal growth factor receptor (EGFR) protein are associated with advanced tumor stage/grade. The objective of this study was to evaluate the effects of the specific EGFR tyrosine kinase inhibitor gefitinib on activation of the Akt and mitogen-activated protein kinase (MAPK) pathways in human urothelial cell carcinoma (UCC) cell lines and to identify potential markers of gefitinib responsiveness in biopsy samples of UCC. EXPERIMENTAL DESIGN: Changes in markers of UCC growth and invasion after exposure to gefitinib were studied in six human UCC cell lines expressing various levels of EGFR. The findings were related to activation of Akt and MAPK. We studied the influence of gefitinib on intraepithelial expansion of the responsive 1207 cell line. EGFR, Akt, and MAPK activation was studied by Western blot analysis of a panel of 57 human UCC. RESULTS: Gefitinib had a growth-inhibitory and anti-invasive effect in two of six UCC cell lines (i.e., 647V and 1207). Gefitinib was also able to block the expansion of 1207 at the expense of normal urothelial cells. These effects did not depend on the level of expression of EGFR but they were associated with the down-regulation of MAPK and Akt activity; in 1207 cells, gefitinib activity was associated with p27 up-regulation and p21 and matrix metalloproteinase-9 down-regulation. Similarly, the Akt and MAPK pathways were found to be strongly phosphorylated in association with EGFR activation in a subset of human UCC specimens. CONCLUSIONS: Activation of EGFR, Akt, and MAPK defines a subset of UCC which might provide information for the identification of gefitinib responders.

A human- and male-specific protocadherin that acts through the wnt signaling pathway to induce neuroendocrine transdifferentiation of prostate cancer cells.

Protocadherin-PC (PCDH-PC) is a gene on the human Y chromosome that is selectively expressed in apoptosis- and hormone-resistant human prostate cancer cells. The protein encoded by PCDH-PC is cytoplasmically localized and has a small serine-rich domain in its COOH terminus that is homologous to the beta-catenin binding site of classical cadherins. Variants of prostate cancer cells that express PCDH-PC have high levels of nuclear beta-catenin protein and increased wnt-signaling. In this study, we show that transfection of human prostate cancer cells (LNCaP) with PCDH-PC or culture of these cells in androgen-free medium (a condition that up-regulates PCDH-PC expression) activates wnt signaling as assessed by nuclear accumulation of beta-catenin, increased expression of luciferase from a reporter vector promoted by Tcf binding elements and increased expression of wnt target genes. Moreover, LNCaP cells transfected with PCDH-PC or grown in androgen-free medium transdifferentiate to neuroendocrine-like cells marked by elevated expression of neuron-specific enolase and chromogranin-A. Neuroendocrine transdifferentiation was also observed when LNCaP cells were transfected by stabilized beta-catenin. Increased wnt signaling and neuroendocrine transdifferentiation of LNCaP cells induced by culture in androgen-free medium was suppressed by short interfering RNAs that target PCDH-PC as well as by dominant-negative Tcf or short interfering RNA against beta-catenin, supporting the hypothesis that increased expression of PCDH-PC is driving neuroendocrine transdifferentiation by activating wnt signaling. These findings have significant implications for the process through which prostate cancers progress to hormone resistance in humans.

Prognostic value of histologic subtypes in renal cell carcinoma: a multicenter experience.

PURPOSE: To analyze to what extent histologic subtype is of prognostic importance in renal cell carcinoma based on a large, international, multicenter experience. PATIENTS AND METHODS: Four thousand sixty-three patients from eight international centers were included in this retrospective study. Histologic subtype (1997 International Union Against Cancer [UICC] criteria of tumor response), age, sex, TNM stage, Fuhrman grade, tumor size, Eastern Cooperative Oncology Goup performance status (ECOG PS), and overall survival were determined in all cases. The prognostic values of clear cell, papillary, and chromophobe histologic features were assessed by uni- and multivariate analysis using the Kaplan-Meier method and Cox model, respectively. RESULTS: Clear cell, papillary, and chromophobe carcinomas accounted for 3,564 (87.7%), 396 (9.7%) and 103 (2.5%) cases, respectively. In univariate analysis, a trend toward a better survival was observed when clear cell, papillary, and chromophobe histologies were considered prognostic categories (log-rank P = .0007). However, in multivariate analysis, TNM stage, Fuhrman grade and ECOG PS, but not histology, were retained as independent prognostic variables (P < .001). CONCLUSION: The stratification in three main renal cell carcinoma histologic subtypes as defined by the 1997 UICC-American Joint Committee on Cancer consensus should not be considered a major prognostic variable comparable to TNM stage, Fuhrman grade and ECOG PS.

FGFR3 and TP53 gene mutations define two distinct pathways in urothelial cell carcinoma of the bladder.

FGFR3 and TP53 mutations are frequent in superficial papillary and invasive disease, respectively. We used denaturing high-performance liquid chromatography and sequencing to screen for FGFR3 and TP53 mutations in 81 newly diagnosed urothelial cell carcinomas. Tumors were classified as follows: 31 pTa, 1 carcinoma in situ, 30 pT1, and 19 pT2-T4. Tumor grades were as follows: 10 G1, 29 G2, and 42 G3. FGFR3 mutations were associated with low-stage (P < 0.0001), low-grade (P < 0.008) tumors, whereas TP53 mutations were associated with high-stage (P < 0.003), high-grade (P < 0.02) tumors. Mutations in these two genes were almost mutually exclusive. Our results suggest that FGFR3 and TP53 mutations define separate pathways at initial diagnosis of urothelial cell carcinoma.

[Complications of retroperitoneal laparoscopy based on a series of 500 cases]. / Complications de la laparoscopie rétropéritonéale: experience aprés 500 cas.

OBJECTIVE: To evaluate the complications of retroperitoneal laparoscopy for upper urinary tract surgery. METHODS: From 1994 to 2003, 500 retroperitoneal laparoscopy procedures were performed: 143 radical nephrectomies, 104 simple nephrectomies, 95 adrenalectomies, 47 ureteropelvic junction plasties, 44 partial nephrectomies, 22 nephroureterectomies, 20 cyst resections, 9 diverticulectomies, 8 lymphadenectomies, 4 pyelotomies and 4 ureteric procedures. The standardized technique uses 5 trocars. RESULTS: There were 23 conversions (4.60%): 5 for retroperitoneal adhesions, 11 for intraoperative bleeding, 7 for technical impossibility. 14 patients (2.8%) required surgical revision: 5 urinomas and 2 urocutaneous fistulas treated by ureteric drainage, 2 deep abscesses, 2 cases of secondary bleeding, 2 colostomies for gastrointestinal fistula after colonic injury, 1 incisional hernia on a trocar orifice. There were postoperative 2 deaths (0.4%) due to septic shock and haemorrhagic shock. 21 patients (4.2%) presented medical postoperative complications: haematoma, hyperthermia, phlebitis and pulmonary infections. 15 patients (3%) were transfused. The most frequent complications occurred after partial nephrectomies and the most serious complications occurred after radical nephrectomies. CONCLUSION: The complication rate is low. Retroperitoneal laparoscopy allows reproducible and effective upper urinary tract surgery, but it is not recommended in patients with a history of retroperitoneal surgery.

The emergence of protocadherin-PC expression during the acquisition of apoptosis-resistance by prostate cancer cells.

In order to identify gene products associated with the development of acquired therapeutic resistance by prostate cancer cells, we created two novel apoptosis-resistant prostate cancer cell lines, LNCaP-TR (phorbol-ester [TPA]-Resistant) and LNCaP-SSR (Serum Starvation-Resistant) by repeated transient exposure of cultured human LNCaP cells to apoptotic stimuli followed by expansion of surviving cell populations. These cell lines were found to be cross-resistant to the alternative selective agent and also hormone-resistant when xenografted into castrated male immunodeficient mice. RNA from the LNCaP-TR line was comparatively screened using a subtractive hybridization-PCR procedure. This allowed us to identify a 249 bp cDNA fragment that hybridized to a 4.8 kb mRNA preferentially expressed by the apoptosis-resistant cells. Using RACE procedures, we cloned and sequenced the complete 4.8 kb cDNA. It is an unusual member of the protocadherin gene family containing two large overlapping open reading frames encoding homologous polypeptides, one having a signal sequence and the other lacking a signal sequence and we refer to it as protocadherin-PC. LNCaP cells directly transformed with protocadherin-PC cDNA were comparatively resistant to phorbol-ester induced apoptosis. Antibody recognition studies demonstrating the cytoplasmic nature of the protcadherin-PC translation product and its propensity to bind beta-catenin suggest that it might influence the apoptotic sensitivity of prostate cancer cells through a unique mechanism.

Molecular grading of urothelial cell carcinoma with fibroblast growth factor receptor 3 and MIB-1 is superior to pathologic grade for the prediction of clinical outcome.

PURPOSE: Fibroblast growth factor receptor 3 (FGFR3) mutations were recently found at a high frequency in well-differentiated urothelial cell carcinoma (UCC). We investigated the relationship between FGFR3 status and three molecular markers (MIB-1, P53, and P27kip1) associated with worse prognosis and determined the reproducibility of pathologic grade and molecular variables. PATIENTS AND METHODS: In this multicenter study, we included 286 patients with primary (first diagnosis) UCC. The histologic slides were reviewed. FGFR3 status was examined by polymerase chain reaction-single strand conformation polymorphism and sequencing. Expression levels of MIB-1, P53, and P27kip1 were determined by immunohistochemistry. Mean follow-up was 5.5 years (range, 0.4 to 18.4 years). RESULTS: FGFR3 mutations were detected in 172 (60%) of 286 UCCs. Grade 1 tumors had an FGFR3 mutation in 88% of patient samples and grade 3 tumors in 16% of patient samples. Conversely, aberrant expression patterns of MIB-1, P53, and P27kip1 were seen in 5%, 2%, and 3% of grade 1 tumors and in 85%, 60%, and 56% of grade 3 tumors, respectively. In multivariate analysis with recurrence rate, progression, and disease-specific survival as end points, the combination of FGFR3 and MIB-1 proved independently significant in all three cases. By using these two molecular markers, three molecular grades (mGs) could be identified: mG1 (mutation; normal expression), favorable prognosis; mG2 (two remaining combinations), intermediate prognosis; and mG3 (no mutation; high expression), poor prognosis. The molecular variables were more reproducible than pathologic grade (85% to 100% v 47% to 61%). CONCLUSION: The FGFR3 mutation represents the favorable molecular pathway of UCC. Molecular grading provides a new, simple, and highly reproducible tool for clinical decision making in UCC patients.

Use of the University of California Los Angeles integrated staging system to predict survival in renal cell carcinoma: an international multicenter study.

PURPOSE: To evaluate ability of the University of California Los Angeles Integrated Staging System (UISS) to stratify patients with localized and metastatic renal cell carcinoma (RCC) into risk groups in an international multicenter study. PATIENTS AND METHODS: 4,202 patients from eight international academic centers were classified according to the UISS, which combines TNM stage, Fuhrman grade, and Eastern Cooperative Oncology Group performance status. Distribution of the UISS categories was assessed in the overall population and in each center. RESULTS: The UISS stratified both localized and metastatic RCC into three different risk groups (P <.001). For localized RCC, the 5-year survival rates were 92%, 67%, and 44% for low-, intermediate-, and high-risk groups, respectively. A trend toward a higher risk of death was observed in all centers for increasing UISS risk category. For metastatic RCC, the 3-year survival rates were 37%, 23%, and 12% for low-, intermediate-, and high-risk groups, respectively; in 6 of 8 centers, a trend toward a higher risk of death was observed for increasing UISS risk category. A greater variability in survival rates among centers was observed for high-risk patients. CONCLUSION: This study defines the general applicability of the UISS for predicting survival in patients with RCC. The UISS is an accurate predictor of survival for patients with localized RCC applicable to external databases. Although the UISS may be useful for patients with metastatic RCC, it may be less accurate in this subset of patients due to the heterogeneity of patients and treatments.

Combined microsatellite and FGFR3 mutation analysis enables a highly sensitive detection of urothelial cell carcinoma in voided urine.

PURPOSE: Fibroblast growth factor receptor 3 (FGFR3) mutations were reported recently at a high frequency in low-grade urothelial cell carcinoma (UCC). We investigated the feasibility of combining microsatellite analysis (MA) and the FGFR3 status for the detection of UCC in voided urine. EXPERIMENTAL DESIGN: In a prospective setting, 59 UCC tissues and matched urine samples were obtained, and subjected to MA (23 markers) and FGFR3 mutation analysis (exons 7, 10, and 15). In each case, a clinical record with tumor and urine features was provided. Fifteen patients with a negative cystoscopy during follow-up served as controls. RESULTS: A mutation in the FGFR3 gene was found in 26 (44%) UCCs of which 22 concerned solitary pTaG1/2 lesions. These mutations were absent in the 15 G3 tumors. For the 6 cases with leukocyturia, 46 microsatellite alterations were found in the tumor. Only 1 of these was also detected in the urine. This was 125 of 357 for the 53 cases without leukocyte contamination. The sensitivity of MA on voided urine was lower for FGFR3-positive UCC (15 of 21; 71%) as compared with FGFR3 wild-type UCC (29 of 32; 91%). By including the FGFR3 mutation, the sensitivity of molecular cytology increased to 89% and was superior to the sensitivity of morphological cytology (25%) for every clinical subdivision. The specificity was 14 of 15 (93%) for the two (molecular and morphological) cytological approaches. CONCLUSIONS: Molecular urine cytology by MA and FGFR3 mutation analysis enables a highly sensitive and specific detection of UCC. The similarity of molecular profiles in tumor and urine corroborate their clonal relation.

Beta-catenin-related anomalies in apoptosis-resistant and hormone-refractory prostate cancer cells.

PURPOSE: beta-Catenin is a critical end component of the wnt signaling pathway that regulates cell growth, apoptosis, and migratory behavior in response to intercellular adhesion molecules. The aim of this study was to evaluate abnormalities of beta-catenin protein expression, subcellular localization, and activity in an in vitro model of acquired apoptosis-resistance in cultured PC cells and in primary human prostate cancers (PrCa). EXPERIMENTAL DESIGN: Apoptosis-resistant human prostate cancer cell line variants were derived from parental LNCaP cells by repeated brief exposure to apoptotic stimuli. The derivative and parental cells were analyzed for beta-catenin expression and intracellular localization using cell fractionation and Western blotting procedures. Endogenous transcriptional activity from the TCF/LEF-1 response element was also studied in these variants after transfection with a beta-catenin sensitive reporter plasmid. Finally, beta-catenin protein expression and intracellular localization were evaluated on 212 patients [122 localized PrCa and 90 hormone-refractory (HRPC) PrCa specimens by immunohistochemistry]. RESULTS: Western blot analysis showed that the intracellular partitioning of beta-catenin was shifted from the membrane fraction in parental cells to the cytoplasmic/nuclear fractions of the apoptosis-resistant cell lines. Coordinately, transcriptional activity from a TCF/LEF-promoted reporter plasmid was increased significantly in the apoptosis-resistant lines. In the primary prostate tumors analyzed, cytoplasmic and/or nuclear beta-catenin expression was correlated statistically with the HRPC status and Gleason score. In the group of localized PrCa, abnormal beta-catenin expression tended to be associated with a higher Gleason score and with pT3 disease. No mutation was found in patients with HRPC and abnormal beta-catenin expression. CONCLUSION: These data suggest that anomalies of beta-catenin expression occur in PrCa and that these anomalies are associated with disease progression, especially to the therapeutic-resistant state.

Antiproliferative effect of nitrosulindac (NCX 1102), a new nitric oxide-donating non-steroidal anti-inflammatory drug, on human bladder carcinoma cell lines.

Non-steroidal anti-inflammatory drugs (NSAIDs) are potent antitumoral agents but their side effects limit their clinical use. A novel class of drugs, nitric oxide-donating NSAIDs (NO-NSAIDs), was found to be safer and more active than classical NSAIDs. This study explored the effect of the NO-donating sulindac derivative, NCX 1102, on three human urothelial epithelial carcinoma cell lines (T24, 647V, and 1207) and primary cultures of normal urothelial cells. Cytotoxicity, antiproliferative effect, cell cycle alterations, morphological changes, and apoptosis were investigated after treatment with NCX 1102 in comparison with the native molecule. After treatment, there was a cytotoxic effect (with IC(50) at 48 h of 23.1 micro M on 647V, 19.4 micro M on T24, and 14.5 micro M on 1207) and an antiproliferative effect on all three cell lines with NCX 1102 but not with sulindac. No effect was detected on normal urothelial cells. Flow cytometric analysis showed a differential NCX 1102-induced accumulation of cells in various phases of the cell cycle, depending on cell line and concentration. NCX 1102 induced an occurrence of multinucleated cells in all cell lines and mitotic arrest in 647V and 1207. NCX 1102-treated T24 and 647V cell lines showed a significant difference of apoptotic cell amount when compared to controls. Our results demonstrated a greater antiproliferative potency of NCX 1102 compared to its parent molecule sulindac, and suggested that this new NO-NSAID may have therapeutic impact in the management of bladder cancer.

Long-term kidney graft survival across a positive historic but negative current sensitized cross-match.

BACKGROUND: The sensitive cross-match (XM) techniques that have been introduced for clinical transplantation can detect anti-donor immune reactivity despite a negative standard National Institute of Health (NIH) cross-match. One of them uses anti-kappa human light chain globulins (AHG). But there is some discussion about the clinical consequences of a positive AHG-XM in the historical sera that became negative in the sera collected just before the transplantation (pretransplant sera). This study was intended to assess the risk of kidney graft failure associated with a positive historic but negative pretransplant AHG-XM in allosensitized patients having a negative historic NIH-XM. METHODS: This retrospective study includes 90 consecutive renal transplants in immunized patients performed at one center between 1985 and 1991. All of the patients had negative historical and pretransplant standard NIH lymphocytotoxic cross-matches and received the same immunosuppressive regimen. The AHG-XMs were done retrospectively using peak historic and sera collected on the day of the transplantation. RESULTS: The AHG cross-match (AHG-XM) was positive in 17 patients, although the standard NIH cross-match was negative. Fourteen of them had a positive historical but negative pretransplant AHG-XM. The actuarial graft survival in this group of 14 patients was 100% at 1 year and 78% at 9 years compared with 90 and 67%, respectively, in patients with negative historical AHG-XM. In addition, the number of rejection episodes per patient as well as renal function at 1, 2, and 5 years were similar in the two groups. IgG anti-donor HLA class I accounted for the XM positivity in 12 of the 14 patients; most rapidly lost all antibody reactivity by NIH technique in an average time of 8 months before the transplantation. In conclusion, this study suggests that transplant patients having a negative historic NIH-XM but a positive historic AHG-XM may not be at high risk of graft failure especially if there is a well-documented sera history showing a marked decrease in PRA level before transplantation and a negative pretransplant AHG-XM.

Introducing a prognostic score for pretherapeutic assessment of seminal vesicle invasion in patients with clinically localized prostate cancer.

PURPOSE: To identify prostate cancer patients who will have the most likely benefit from sparing the seminal vesicles during 3D conformal radiation therapy. METHODS AND MATERIALS: From 1988 to 2001, 532 patients underwent radical prostatectomy for clinically localized prostate cancer. Primary endpoint was the pathological evidence of seminal vesicle invasion. Variables for univariate and multivariate analyses were age, prostate weight, clinical stage, PSA level, Gleason score, number and site of positive prostate sextant biopsies. Multivariate logistic regression with backward stepwise variable selection was used to identify a set of independent predictors of seminal vesicle invasion, and the variable selection procedure was validated by non-parametric bootstrap. RESULTS: Seminal vesicle invasion was reported in 14% of the cases. In univariate analysis, all variables except age and prostate weight were predictors of seminal vesicle invasion. In multivariate analysis, only the number of positive biopsies (P<0.0001), Gleason score (P<0.007) and PSA (P<0.0001) were predictors for seminal vesicles invasion. Based on the multivariate model, we were able to develop a prognostic score for seminal vesicle invasion, which allowed us to discriminate two patient groups: A group with low risk of seminal vesicles invasion (5.7%), and the second with a higher risk of seminal vesicles invasion (32.7%). CONCLUSIONS: Using the number of positive biopsies, Gleason score and PSA, it is possible to identify patients with low risk of seminal vesicles invasion. In this population, seminal vesicles might be excluded as a target volume in radiation therapy of prostate cancer.

Profiles of the 2 INK4a gene products, p16 and p14ARF, in human reference urothelium and bladder carcinomas, according to pRb and p53 protein status.

The INK4a/ARF locus encodes 2 cell cycle regulatory proteins: p16 and p14(ARF). P16 inhibits the activities of cdks, which maintain the retinoblastoma protein (pRb) in its active hypophosphorylated state. P14(ARF) blocks MDM2-induced p53 degradation and transactivational silencing. In this study, we investigated the expression of p16 and p14(ARF) in reference human urothelium and in 51 urothelial carcinomas (UCs) of all stages and grades, by reverse transcription-polymerase chain reaction (RT-PCR). Patterns of p14(ARF) and p16 expression were compared with each other and then with patterns of p53 and pRb protein expression, respectively, as determined by immunohistochemistry. P14(ARF) and p16 mRNAs were present at low levels or were undetectable in reference urothelia and in most superficial tumors, whereas they were present at high levels in a subset of tumors of advanced stage and high grade. The expression profiles of these 2 mRNAs were correlated in all but 4 cases, indicating that the 2 INK4a products may have nonredundant functions. Forty-six of the 51 tumors (90%) presented changes to or a lack of activation of the p14(ARF)-p53 pathway and were p53 positive (n = 10), p14(ARF) negative (n = 23), or both p53 positive and p14(ARF) negative (n = 13), suggesting that these 2 components of the pathway may be altered or nonactivated. Markedly high levels of p16 mRNA (n = 5) were associated with the absence of pRb expression, with the exception of 1 case in which the p16 gene contained a deletion mutation. A lack of p16 mRNA or low levels of this mRNA were associated with pRb detection in all but 1 case. In invasive UCs, the p16-pRb pathway, the p14(ARF)-p53 pathway, or in many cases both pathways were altered or not activated, demonstrating the involvement of these pathways in invasive bladder tumorigenesis.

Human hormone-refractory prostate cancers can harbor mutations in the O(2)-dependent degradation domain of hypoxia inducible factor-1alpha (HIF-1alpha).

PURPOSE: Androgen ablation, the preferred therapy for advanced prostate cancer, reduces blood flow and induces hypoxia in androgen-dependent tissues. Given the transient effectiveness of this therapy, we must consider whether a hypoxia-resistance mechanism might be involved in the development of therapeutic resistance by prostate cancer cells. The transcription factor protein, hypoxia-inducible factor 1alpha (HIF-1alpha), helps increase the expression of gene products that enable cells to survive conditions of hypoxic stress. Enhanced HIF-1alpha expression during hypoxia results from a drastic reduction of its degradation rate within a critical region of the protein referred to as the "oxygen-dependent degradation (ODD) domain". We sequenced HIF-1alpha cDNAs amplified from human prostate cancer cell lines and from hormone resistant prostate cancer specimens to determine whether prostate cancer cells might harbor mutations within the HIF-1alpha ODD domain. METHODS: HIF-1alpha cDNAs were RT-PCR amplified from three prostate cancer cell lines (LNCaP, PC-3, and DU145), from five different human hormone-resistant prostate cancers and one normal prostate, all microdissected, and were sequenced to determine whether the HIF-1alpha gene products were wildtype or mutant. One specimen containing a hormone-resistant prostate tumor that expressed a mutated HIF-1alpha cDNA was further microdissected into benign and tumorous regions and DNAs extracted from these regions were directly amplified by PCR and sequenced to determine whether the HIF-1alpha mutation was specific to the tumor. RESULTS: Although the HIF-1alpha cDNAs of all cell lines, the normal prostate, and three of the tumors were found to have a wildtype sequence, HIF-1alpha cDNAs amplified from two hormone-resistant tumors had nucleic acid substitutions that resulted in significant amino acid changes within the ODD domain of the HIF-1alpha protein. Analysis of the DNA extracted from a benign or tumorous region of one of these specimens showed that only the wildtype (nonmutated) form of the HIF-1alpha gene was amplified from the normal DNA whereas only the mutated form of the HIF-1alpha gene was amplified from the tumor. CONCLUSIONS: Some human hormone-refractory prostate cancers have mutations in a critical regulatory domain of the HIF-1alpha protein. We believe that these mutations might enable expression of this protein under inappropriate conditions and contribute to the development of therapeutic resistance by the cancer cells. This hypothesis is currently being tested.

Prognostic value of EGF receptor and tumor cell proliferation in bladder cancer: therapeutic implications.

Changes in growth factor receptor expression may confer a growth advantage on tumour cells. Epidermal growth factor-receptor (EGF-R) has been associated with the genesis of bladder tumours. We sought a link between EGF-R expression and MIB-1 cell proliferation and examined their prognostic value in the progression of bladder cancer. Fresh frozen samples from 113 transitional cell carcinomas (TCC) of the bladder and 10 healthy bladders were studied by immunohistochemistry, using monoclonal antibodies for EGF-R expression and MIB-1 for cell proliferation. Qualitative and quantitative immunostaining were analyzed in relation to time to progression and compared with clinical and pathologic parameters for prognostic significance in univariate and multivariate analysis (stepwise logistic regression). EGF-R stained more intensively in invasive tumours. Median nuclear over-expression of MIB-1 was 28%. Progression free survival rate estimates (log rank test) were significantly lower in patients EGF-R positive and with MIB-1 score above 28% (P < 0.0001, P < 0.0001, respectively). Multivariate analysis indicated that MIB-1 immunostaining was the most significant independent variable and EGF-R expression had no additional prognostic value over clinical stage and grade and cell proliferation. The MIB-1 proliferation index is a stronger predictor of bladder tumour progression than is EGF-R over-expression. This marker yield significant prognostic information in addition to stage and grade and may be of value for the clinical management of superficial and invasive bladder carcinomas. The pattern of EGF-R immunostaining and its association with tumour progression makes it a candidate for antigrowth factor therapy.