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TISSUE MICROARRAY ANALYSES OF MULTIPLE BIOMARKERS AS CANDIDATE PROGNOSTIC INDICATORS FOR PREDICTING RELAPSE FOLLOWING RADICAL PROSTATECTOMY
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TISSUE MICROARRAY ANALYSES OF MULTIPLE BIOMARKERS AS CANDIDATE PROGNOSTIC INDICATORS FOR PREDICTING RELAPSE FOLLOWING RADICAL PROSTATECTOMY
METHYLATION PROFILE OF N33, HIC, GSTP1 AND CDKN2 AT DIFFERENT STAGES OF PROSTATE CARCINOGENESIS
Nariculam J., Freeman A., Feneley M., Masters J. University College London, Institute of Urology, London, United Kingdom INTRODUCTION & OBJECTIVES: Tissue microarrays (TMA) provide a powerful tool to evaluate biomarker expression simultaneously in large numbers of tissue samples. This study used TMAs to evaluate expression of four biomarkers and their prognostic value independent of established clinical indicators, in a well-defined sample of patients. BCL-2, Ki67, p53 and E-cadherin expression were examined in relation to relapse status following radical prostatectomy (relapse or non-relapse) for patient pairs matched for preoperative PSA, pathological stage and Gleason Sum score. MATERIAL & METHODS: Using our radical prostatectomy database of over 1200 patients, 72 patients (36 patient pairs) were identified, characterised by distinct biochemical relapse status but otherwise matched for pT3 pathological stage, preoperative serum PSA level, and Gleason Sum score. The relapse group consisted of 36 men with PSA recurrence within two years of surgery (PSA level > 0.2ng/ml). The non-relapse group consisted of 36 men with undetectable PSA > 3 years after surgery. No patients had any adjuvant or neoadjuvant therapy. TMA blocks containing 210 cores were constructed from formalin fixed paraffin embedded tissue from radical prostatectomy specimens. Cores were taken from cancer and benign areas to produce 4 replicate TMA blocks. BCL-2, Ki67, p53 and E-cadherin expression were examined immunohistochemically. Extent of staining was assessed semi-quantitatively, except for Ki67, where a labelling index was calculated. Relative biomarker expression was related to relapse status. RESULTS: Comparing non-neoplastic and cancer tissue, p53 expression was higher in 11 relapsers (30%) and 7 non-relapsers (20%) and in 18 pairs (50%) expression was identical. Ki67 expression was higher in 12 relapsers (33%) and 8 non-relapsers (22%) and was identical in 16 pairs (44%). BCl-2 expression was higher in 7 relapsers (20%) and 5 non-relapsers (14%) and identical in 24 pairs (67%). E-cadherin expression was higher in 1 relapser and in the other 35 pairs expression was identical. CONCLUSIONS: p53, BCL-2, E-cadherin and Ki67 expression did not predict relapse status following radical prostatectomy in patients matched for pathological stage, Gleason grade and preoperative PSA. These markers are therefore unlikely to have prognostic value independent of these well-established clinical prognostic factors. TMA blocks constructed from samples matched for clinical prognostic variables provides a powerful tool for evaluating molecular markers as independent prognostic indicators, using relatively small numbers of cases.
LAPAROSCOPIC RADICAL PROSTATECTOMY AND ROBOTIC SURGERY I Wednesday, 5 April, 12.45-14.15, Room Concorde 2 / Level 4 115 AN ANIMAL MODEL FOR HEAT DISTRIBUTION AROUND ENDOSHEARS COAGULATION DEVICES - APPLICATIONS FOR LAPAROSCOPIC NERVE-SPARING SURGERY
Alekseev B.1, Shegai P.1, Kekeeva T.2, Nemtsova M.3, Zaletaev D.3, Rusakov I.1 1
Moscow Hertzen Oncology Institute, Oncourology, Moscow, Russia, 2Moscow Medical Academy, Genetics, Moscow, Russia, 3Research Centre for Medical Genetics Rams, Medical Genetics, Moscow, Russia INTRODUCTION & OBJECTIVES: DNA methylation of CpG sites of tumour suppressor genes is a frequent epigenetic event in the cancer pathogenesis. We examined the methylation profile of N33, HIC, GSTP1 and CDKN2 genes in the spectrum of prostate cancer disease progression, from benign prostate hyperplasia (BPH) and prostatic intraepithelial neoplasia (PIN) to primary prostate cancer. MATERIAL & METHODS: Biopsy specimens were obtained from patients with BPH (n=24), high grade PIN (n=32) and adenocarcinoma (n=52), 28 prostate tissues were obtained from patients with prostate cancer after radical prostatectomy. Methylation status was determined using methylation-sensitive PCR. RESULTS: Methylation frequency of HIC1 in BPH samples was 37% and methylation frequencies of PIN and adenocarcinoma were significantly higher 66% and 65% accordingly (P=0,05). Promoter hypermethylation of p16 was detected in 25% of hyperplasia, 29% of neoplasia and 36% of carcinoma. The similar pattern of methylation was shown for GSTP1 gene (25%, 16% and 42%). It was interesting that methylation of N33 was absent in the preinvasive lesions but was frequent in adenocarcinoma (18%, P=0,02). Methylation percentages in prostatectomy tissues were: GSTP1, 32%; HIC1, 71%; p16, 18%; N33, 20%. There were no significant differences in the frequencies of methylation in biopsy and prostatectomy prostate adenocarcinomas specimens for individual genes. CONCLUSIONS: We can conclude that N33 methylation is related to late, tumour-associated events, while HIC1, GSTP1 and CDKN2 methylation appears with different frequencies at cancer precursor lesions. These early epigenetic DNA alterations are the accessory component of extensive tumour reorganizations and provide clinically useful markers to diagnose early prostate cancer lesions and assess disease prognosis.
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Katz R.1, Danai Y.2, Gofrit O.1, Peleg E.2, Adler D.2, Pode D.1 1 Hadassah Hebrew University Hospital, Urology, Jerusalem, Israel, 2Hadassah Hebrew University Hospital, Biomedical Engineering, Jerusalem, Israel
INTRODUCTION & OBJECTIVES: Advanced laparoscopic procedures such as laparosacopic radical prostatectomy or retroperitoneal lymph node dissection often include preservation of locoregional nerve fibers. The use of electrical coagulation in such procedures is limited due to tissue conductance of current and heat production. While ultrasonic shears coagulation devices (e.g. harmonic scalpel) do not use electricity, heat is still produced. Previous works showed that tissue temperatures of 48-57ºC may cause irreversible damage to peripheral nerve fibers. We designed an animal model to characterize the heat spread around ultrasonic devices and assess whether it is potentially neurodestructive. MATERIAL & METHODS: 10 rats were anesthetized; their skin was reflected, exposing the muscles. An ultrasonic probe was introduced into the tissue and using 4 thermocouples, tissue temperatures during coagulation were measured in distances of 4, 8, 12 and 16 millimetres from the probe and in a circumferential manner around it. RESULTS: In distances of 4-8 millimetres from the device, temperatures as high as 81ºc were recorded. The heat had a typical pattern of distribution. The area above the vertical vibrating blade was the hottest and the region below the horizontal fixed blade was significantly colder. CONCLUSIONS: Although ultrasonic coagulation is efficient, local heat production exists and reaches neurodestructive levels with a typical tissue distribution pattern. Further histological studies are required to assess the nerve damage following the application of ultrasonic coagulation.
116 ANATOMICAL RETROGRADE LAPAROSCOPIC PROSTATECTOMY IMPROVES POSTOPERATIVE ERECTIONS WITHOUT INCREASING OF SURGICAL MARGINS: A COMPARATIVE STUDY Colombel M., Mege Lechevallier F., Marechal J.M., Poissonnier L., Murat F.J., Pricaz E., Gelet A., Xavier M. Hôpital Edouard Herriot, Urology, Lyons, France INTRODUCTION & OBJECTIVES: This analysis was designed to assess the ability of the retrograde laparoscopic protatectomy (that reproduce every steps of the anatomical prostatectomy as described by Walsh), to improve long term spontaneous erectile function without increasing positive margins rates. MATERIAL & METHODS: Men with functional preoperative erections who underwent radical prostatectomy for localised prostate cancer (low and intermediate risk) were stratified according to operative technique: open anatomical retrograde prostatectomy (group 1); laparoscopic anatomical retrograde prostatectomy (group 2); laparoscopic antegrade prostatectomy (group 3). Post operatively patients received oral Sildenafil and non responders were switched to intracavernosal injection therapy. Patients who completed the international index of erectile function (IIEF) and who had been followed at least 18 months were included. RESULTS: There were 68 patients in the open prostatectomy group, 72 in the laparoscopic anatomical retrograde prostatectomy group, and 70 in the laparoscopic antegrade prostatectomy group. There were no differences in age, comorbidity profile, intra operative nerve sparing status, or prostate cancer risk profile between groups. At 18 months post RP, there were statistically significant differences between open RP and laparoscopic prostatectomy in the percentage of patients who were capable of having medication unassisted intercourse (group 1: 35% vs. group 2-3: 48%) mean IIEF erectile function (group 1: 18±5 vs. group 2-3: 27±3), the percentage of patients with normal EF domain scores (group 1: 21% vs. group 2-3: 31%); the percentage of patients responding to sildenafil (group 1: 31% vs. group 2-3: 39%) and the percentage of patients responding to low doses (2,5μg) of PGE1 at 3 months post operatively (group 1: 68% vs. group 2-3: 88%). However, there were statistically significant differences in the rate of positive surgical margins between retrograde anatomical laparoscopic or open and laparoscopic antegrade PR (group 1: 12%, group 2: 11%, group 3: 19%). CONCLUSIONS: In our hand, laparoscopic prostatectomy allowed us to improve our results on post operative erectile function. However it is only by reproducing the anatomical retrograde prostatectomy as described by Walsh that we were able to keep our positive margins at an acceptable rate.
Eur Urol Suppl 2006;5(2):51
114
TISSUE MICROARRAY ANALYSES OF MULTIPLE BIOMARKERS AS CANDIDATE PROGNOSTIC INDICATORS FOR PREDICTING RELAPSE FOLLOWING RADICAL PROSTATECTOMY
METHYLATION PROFILE OF N33, HIC, GSTP1 AND CDKN2 AT DIFFERENT STAGES OF PROSTATE CARCINOGENESIS
Nariculam J., Freeman A., Feneley M., Masters J. University College London, Institute of Urology, London, United Kingdom INTRODUCTION & OBJECTIVES: Tissue microarrays (TMA) provide a powerful tool to evaluate biomarker expression simultaneously in large numbers of tissue samples. This study used TMAs to evaluate expression of four biomarkers and their prognostic value independent of established clinical indicators, in a well-defined sample of patients. BCL-2, Ki67, p53 and E-cadherin expression were examined in relation to relapse status following radical prostatectomy (relapse or non-relapse) for patient pairs matched for preoperative PSA, pathological stage and Gleason Sum score. MATERIAL & METHODS: Using our radical prostatectomy database of over 1200 patients, 72 patients (36 patient pairs) were identified, characterised by distinct biochemical relapse status but otherwise matched for pT3 pathological stage, preoperative serum PSA level, and Gleason Sum score. The relapse group consisted of 36 men with PSA recurrence within two years of surgery (PSA level > 0.2ng/ml). The non-relapse group consisted of 36 men with undetectable PSA > 3 years after surgery. No patients had any adjuvant or neoadjuvant therapy. TMA blocks containing 210 cores were constructed from formalin fixed paraffin embedded tissue from radical prostatectomy specimens. Cores were taken from cancer and benign areas to produce 4 replicate TMA blocks. BCL-2, Ki67, p53 and E-cadherin expression were examined immunohistochemically. Extent of staining was assessed semi-quantitatively, except for Ki67, where a labelling index was calculated. Relative biomarker expression was related to relapse status. RESULTS: Comparing non-neoplastic and cancer tissue, p53 expression was higher in 11 relapsers (30%) and 7 non-relapsers (20%) and in 18 pairs (50%) expression was identical. Ki67 expression was higher in 12 relapsers (33%) and 8 non-relapsers (22%) and was identical in 16 pairs (44%). BCl-2 expression was higher in 7 relapsers (20%) and 5 non-relapsers (14%) and identical in 24 pairs (67%). E-cadherin expression was higher in 1 relapser and in the other 35 pairs expression was identical. CONCLUSIONS: p53, BCL-2, E-cadherin and Ki67 expression did not predict relapse status following radical prostatectomy in patients matched for pathological stage, Gleason grade and preoperative PSA. These markers are therefore unlikely to have prognostic value independent of these well-established clinical prognostic factors. TMA blocks constructed from samples matched for clinical prognostic variables provides a powerful tool for evaluating molecular markers as independent prognostic indicators, using relatively small numbers of cases.
LAPAROSCOPIC RADICAL PROSTATECTOMY AND ROBOTIC SURGERY I Wednesday, 5 April, 12.45-14.15, Room Concorde 2 / Level 4 115 AN ANIMAL MODEL FOR HEAT DISTRIBUTION AROUND ENDOSHEARS COAGULATION DEVICES - APPLICATIONS FOR LAPAROSCOPIC NERVE-SPARING SURGERY
Alekseev B.1, Shegai P.1, Kekeeva T.2, Nemtsova M.3, Zaletaev D.3, Rusakov I.1 1
Moscow Hertzen Oncology Institute, Oncourology, Moscow, Russia, 2Moscow Medical Academy, Genetics, Moscow, Russia, 3Research Centre for Medical Genetics Rams, Medical Genetics, Moscow, Russia INTRODUCTION & OBJECTIVES: DNA methylation of CpG sites of tumour suppressor genes is a frequent epigenetic event in the cancer pathogenesis. We examined the methylation profile of N33, HIC, GSTP1 and CDKN2 genes in the spectrum of prostate cancer disease progression, from benign prostate hyperplasia (BPH) and prostatic intraepithelial neoplasia (PIN) to primary prostate cancer. MATERIAL & METHODS: Biopsy specimens were obtained from patients with BPH (n=24), high grade PIN (n=32) and adenocarcinoma (n=52), 28 prostate tissues were obtained from patients with prostate cancer after radical prostatectomy. Methylation status was determined using methylation-sensitive PCR. RESULTS: Methylation frequency of HIC1 in BPH samples was 37% and methylation frequencies of PIN and adenocarcinoma were significantly higher 66% and 65% accordingly (P=0,05). Promoter hypermethylation of p16 was detected in 25% of hyperplasia, 29% of neoplasia and 36% of carcinoma. The similar pattern of methylation was shown for GSTP1 gene (25%, 16% and 42%). It was interesting that methylation of N33 was absent in the preinvasive lesions but was frequent in adenocarcinoma (18%, P=0,02). Methylation percentages in prostatectomy tissues were: GSTP1, 32%; HIC1, 71%; p16, 18%; N33, 20%. There were no significant differences in the frequencies of methylation in biopsy and prostatectomy prostate adenocarcinomas specimens for individual genes. CONCLUSIONS: We can conclude that N33 methylation is related to late, tumour-associated events, while HIC1, GSTP1 and CDKN2 methylation appears with different frequencies at cancer precursor lesions. These early epigenetic DNA alterations are the accessory component of extensive tumour reorganizations and provide clinically useful markers to diagnose early prostate cancer lesions and assess disease prognosis.
P8
Katz R.1, Danai Y.2, Gofrit O.1, Peleg E.2, Adler D.2, Pode D.1 1 Hadassah Hebrew University Hospital, Urology, Jerusalem, Israel, 2Hadassah Hebrew University Hospital, Biomedical Engineering, Jerusalem, Israel
INTRODUCTION & OBJECTIVES: Advanced laparoscopic procedures such as laparosacopic radical prostatectomy or retroperitoneal lymph node dissection often include preservation of locoregional nerve fibers. The use of electrical coagulation in such procedures is limited due to tissue conductance of current and heat production. While ultrasonic shears coagulation devices (e.g. harmonic scalpel) do not use electricity, heat is still produced. Previous works showed that tissue temperatures of 48-57ºC may cause irreversible damage to peripheral nerve fibers. We designed an animal model to characterize the heat spread around ultrasonic devices and assess whether it is potentially neurodestructive. MATERIAL & METHODS: 10 rats were anesthetized; their skin was reflected, exposing the muscles. An ultrasonic probe was introduced into the tissue and using 4 thermocouples, tissue temperatures during coagulation were measured in distances of 4, 8, 12 and 16 millimetres from the probe and in a circumferential manner around it. RESULTS: In distances of 4-8 millimetres from the device, temperatures as high as 81ºc were recorded. The heat had a typical pattern of distribution. The area above the vertical vibrating blade was the hottest and the region below the horizontal fixed blade was significantly colder. CONCLUSIONS: Although ultrasonic coagulation is efficient, local heat production exists and reaches neurodestructive levels with a typical tissue distribution pattern. Further histological studies are required to assess the nerve damage following the application of ultrasonic coagulation.
116 ANATOMICAL RETROGRADE LAPAROSCOPIC PROSTATECTOMY IMPROVES POSTOPERATIVE ERECTIONS WITHOUT INCREASING OF SURGICAL MARGINS: A COMPARATIVE STUDY Colombel M., Mege Lechevallier F., Marechal J.M., Poissonnier L., Murat F.J., Pricaz E., Gelet A., Xavier M. Hôpital Edouard Herriot, Urology, Lyons, France INTRODUCTION & OBJECTIVES: This analysis was designed to assess the ability of the retrograde laparoscopic protatectomy (that reproduce every steps of the anatomical prostatectomy as described by Walsh), to improve long term spontaneous erectile function without increasing positive margins rates. MATERIAL & METHODS: Men with functional preoperative erections who underwent radical prostatectomy for localised prostate cancer (low and intermediate risk) were stratified according to operative technique: open anatomical retrograde prostatectomy (group 1); laparoscopic anatomical retrograde prostatectomy (group 2); laparoscopic antegrade prostatectomy (group 3). Post operatively patients received oral Sildenafil and non responders were switched to intracavernosal injection therapy. Patients who completed the international index of erectile function (IIEF) and who had been followed at least 18 months were included. RESULTS: There were 68 patients in the open prostatectomy group, 72 in the laparoscopic anatomical retrograde prostatectomy group, and 70 in the laparoscopic antegrade prostatectomy group. There were no differences in age, comorbidity profile, intra operative nerve sparing status, or prostate cancer risk profile between groups. At 18 months post RP, there were statistically significant differences between open RP and laparoscopic prostatectomy in the percentage of patients who were capable of having medication unassisted intercourse (group 1: 35% vs. group 2-3: 48%) mean IIEF erectile function (group 1: 18±5 vs. group 2-3: 27±3), the percentage of patients with normal EF domain scores (group 1: 21% vs. group 2-3: 31%); the percentage of patients responding to sildenafil (group 1: 31% vs. group 2-3: 39%) and the percentage of patients responding to low doses (2,5μg) of PGE1 at 3 months post operatively (group 1: 68% vs. group 2-3: 88%). However, there were statistically significant differences in the rate of positive surgical margins between retrograde anatomical laparoscopic or open and laparoscopic antegrade PR (group 1: 12%, group 2: 11%, group 3: 19%). CONCLUSIONS: In our hand, laparoscopic prostatectomy allowed us to improve our results on post operative erectile function. However it is only by reproducing the anatomical retrograde prostatectomy as described by Walsh that we were able to keep our positive margins at an acceptable rate.
Eur Urol Suppl 2006;5(2):51