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Prostate cancer and immunotherapy: A preliminary study about frequency of telomerase-specific CD8+ lymphocytes
133 CHEMOSENSITISATION OF HUMAN PROSTATE CANCER USING ANTISENSE AGENTS TARGETING THE TYPE I INSULIN-LIKE GROWTH FACTOR RECEPTOR
134 ANANDAMIDE AND CAPSAICIN INHIBIT PROLIFERATION OF HUMAN PROSTATE CANCER CELLVIACANNABINOID RECEPTOR AND VANILLOID RECEPTOR. RESPECTIVELY
Hellawell G.‘. Ferguson D.‘. Brewster 8’. Macaulay V.’ Kamivama ‘institute of Molecular Medicine, Cancer Research UK Oncology Laboratories, Oxford, United Kingdom, ‘The John Radcliffe Hospttal, Nuflield Department of Pathology. Oxford, United Kingdom. ‘The Churchill Hospttal, Department of Urology. Oxford. United Kingdom INTRODUCTION & OBJECTIVES: The type I msulin-hkc growth factor receptor (IGFlR) is tmportant for tumour growth and apoptosis protection. We have recently shown that the IGFI R ts overexpressed by prostate cancer compared with benign prostatic epithelium, and IGFI R expression commonly persists in androgenmdependent metastatic disease at levels comparable to those in the primary (Cancer Res 62:2942-50, 2002). In vitro reports recently observed that the lGFlR is uprcgulated in prostate cancer cells during the development of androgenIndependence. The objective of this study was to determine whether antisense mediated IGFI R downregulation could enhance the chemosensttivity of androgentndependent prostate cancer cells in vitro MATERIAL & METHODS: We transfectcd human androgclr-itldcpendent DU I45 prostate cancer cells with lGFl R antisense oligonucleotidcs (ASOs) or antisense RNA. Transfected cultures vvere treated with cisplatm, mitoxantrone, paclitaxel or vehicle control, and survival was measured by clonogcnic assay. DlJ I45 prostate cancer cell clones expressing ICiFl R antisense RN4 were devclopcd to assess in viva growth in nude mtcc. RESULTS: IGFIR ASOs caused dose-dependent inhibition of IGFI R exprcsston to levels of 30.40% of those m control-treated cells. This was accompanied by marked reductton m DU 145 growth measured by MTS assay, and clonogenic survival was rcduccd by up to 80%. Growth of’ IGFIR antisense RNA expressing DUl45 cell clones was ahnost completely abolished in viva. Furthermore. IGFlR ASOs enhanced chemosensitivity to cisplatin, mitoxantrone and paclitaxel with 1.5 to 2-fold reduction in IC-50. Ultrastructural analysis of the DU145 clones treated with cisplatin confirmed that the AS-IGFI R cells wcrc more susceptible to apoptosis.
M.. Furuya Y.. Takihana Y.. Araki I.. Tanabe N., Takeda M
University of Yamanashi,
lirology,
Yamanashi,
Japan
INTRODUCTION & OBJECTIVES: It is known that both cannabinoid receptor subtype I (CBI) and vanilloid receptor (VRI) are expressed not only in nerve system but in peripheral non-neuronal tissues, and are related to controlling proliferation in various cells. The present study was designed to Investigate the expression of CBI and VRI in human prostate and the effect of C‘BI and VRI on the proliferation of human prostate cancer ceils. MATERIAL & METHODS: Immunohistochemistry and RT-PCR were performed to determine the distribution and expression of CBI and VRI in human prostate tissues and cell lines, three normal cells and three malignant cells. DUl45 cells were treated with anandamide (non-selective agonist for (‘Bl) or capsaicm (selectivse agonist for VRI), and then cell viability was measured by MTT assay. To further analyse whether the cell death was medtated via CBI and VRI DUl45 cells were treated with selective antagonists of these receptors. RESULTS: In prostate tissues, CBI was highly expressed in epithelial cells, while VRI was in stromal regions. And in prostate cell lines, the expression of CBI was considerable in DU145 cells, while VRl was expressed in all cell lines, regardless of normal or malignant. In DUl45, anandamide elicited cell death, which was significantly inhibited by AM251. a selective antagonist for CBI receptor. And similarly. capsaicin elicited cell death, which was inhibited by capsarepine, a selective antagonist for VR I receptor.
CONCLUSIONS: These results mdicatc that lGFlR downregulation can enhance chemosensitivity of androgcn-independent prostate cancer in part via enhanced suscepttbility to apoptosis. In a clmical srttmg. targeted suppression of the IGFI R ming ASG technology may cnhancc the effects of conventional chemotherapy and therefore otfcrs promtsc as novel therapy for patients with androgen-independent disease.
CONCLUSIONS: CB I and VK I were expressed in human prostate tissues. In DUl45 cells, cell death induced by anandamidc or capsaicin was elicited via C‘B I or VR I, respectively.
135
136
PROSTATE CANCER AND IMMUNOTHERAPY: A PRELIMINARY STUDY ABOUT FREQUENCY OF TELOMERASE-SPECIFIC CDS+ LYMPHOCYTES Traverao P.‘, Romagnolt Carmignani Ci.
A.‘. Filact G’. Fravrga
of Cicnoa. DIMI. (icnoa.
INTRODUCTION & OBJECTIVES: Telomerase is an interesting tumourassociated antigen (TAA) because it is expressed m majority of cancers and because it is possible to generate telomerase specific CD8+ T lymphocytes (CTL) in the peripheral blood of healthy subjects or affected by cancer. We try to find out frequency of telomerase specific CTL in patients with prostate carcinoma. MATERIAL & METHODS: We used the l-540 antigen, encompassing the 540. 548 pepttde sequence of tclomcrase, that is presented by HLA-A2 class I molecules, We enrolled 5 HLA-,421 pattents: 4 wtth clinically localired prostate cancer (which have been subjected to a Radical Retropubic Prostatectomy) and I wtth metastatic prostate. 4 HLA-A2 t pattents without prostate cancer and 2 HLAA2- patients with prostate cancer were the control group. We did subsequent analyses a) calculation of number of CDX+ T lymphocytes reacting to HLA-A2 by tmmunofluorescence after staining with FITC-labelled HLA-tetramer loaded with l-540 peptidc, b) calculation of frequency of l-540 specitic CTI, precursors using “ltmitmg dtlution” c) generation of I-540 specific CTL lines. d RESUIXS: Percentage of l-S40 specific C‘DX+ T lymphocytes was significantly higher in patients HLA-A2+ affected by prostate cancer (P
Supplements
IN BPH AND PROSTATE CANCER PATIENTS
Mado-cbnchcr S ‘. Ponholrcr A.‘. Schat/l C; Ntjhuts L.‘. AdlercreutL II:
(i\ur 4 ‘, Hatdmger (i ‘. dc Vrtcs (’ ‘1
M.‘. Naselli A ‘_ Indiveri F.‘.
‘Ilntvcrsity of Cienoa. Ut-ology. Cicnoa. Italy. llniversity Italy
European
PHYTO-OESTROGENS
2 (2003) No. 1, pp. 36
I)onauspttal. Dept. of Urology, Vienna. Austrta. Umverstty of Vtennn, Dept. of llrology. Vienna. Austria. ‘University of Vienna. Institute of (‘ancer Research, Vienna, Austria, ‘Clnivcrsity of Helsinki, Inst. for Prey. Med. Nutr. and Cancer, Helsinki. Finland INTRODUCTION & OBJECTIVES: Epidemtological differences (west vs. cast) in prostate cancer (PC) incidence have been in part attributed to the consumption of phyto-oestrogens, which is substantially higher in Asians. Aim of our study was to determine whether serum levels of phyto-oestrogcns differ in BPH and PC-patients in an Austrian population and to estimate their correlation to PSA and endocrine parameters. MATERIAL & METHODS: In this cast control study, men (Caucasian only) wtth newly diagnosed, untreated prostate cancer and, as a control population, agcmatched patients with untreated lower urtnary tract symptoms due to benign prostatic hyperplasta (BPH) wcrc entered. In BPII-patients. PC was excluded clinically (negative digito-rectal examination and PSA) or histologically. Serum levels of the phyto-oestrogens datd7ein. gcnistcin and enterolactone were analysed by time-resolved fluoroimnlunoaasays; in parallel a serum cndocrinc study was obtained. RESULTS: A total of 194 men, 97 BPH- and 97 PC-patiems entcrcd this case control study. Mean age was comparable in BPH- (hX+lOyrs: meanistandard deviation) and PC- (64+6yrs; ~~0.05) casts. Enterolactone (50+209 vs. 22+34nmol/L), Daidzein (17+44 vs. 10+35nmol/L) and Genistein (33187 vs. 19+45nmol/L) wcrc all higher m BPH-patients, yet these differences did not reach statistical significance. We subsequently correlated phyto-oestrogens to PSA and endocrine parameters in the entire study population (n=l94); these analyses were controlled for age. No correlations between phyto-oestrogens and PSA, LH, total testosterone and estradiol were observed. Daidzein (p=O.O 19). Genistein (p=O.O02) and the sum of all three phyto-estrogens (p=O.OOS), however, were all positively correlated to FSH. CONCI.USIONS: These data suggest that photo-oestrogcns arc involved m the pathogenesis of prostate cancer and underline their potential for prostate cancer chemoprevention. The close correlation between FSH and phyto-oestrogens directs towards a potenttal regulatory mechanism and warrants further atudtcc.
134 ANANDAMIDE AND CAPSAICIN INHIBIT PROLIFERATION OF HUMAN PROSTATE CANCER CELLVIACANNABINOID RECEPTOR AND VANILLOID RECEPTOR. RESPECTIVELY
Hellawell G.‘. Ferguson D.‘. Brewster 8’. Macaulay V.’ Kamivama ‘institute of Molecular Medicine, Cancer Research UK Oncology Laboratories, Oxford, United Kingdom, ‘The John Radcliffe Hospttal, Nuflield Department of Pathology. Oxford, United Kingdom. ‘The Churchill Hospttal, Department of Urology. Oxford. United Kingdom INTRODUCTION & OBJECTIVES: The type I msulin-hkc growth factor receptor (IGFlR) is tmportant for tumour growth and apoptosis protection. We have recently shown that the IGFI R ts overexpressed by prostate cancer compared with benign prostatic epithelium, and IGFI R expression commonly persists in androgenmdependent metastatic disease at levels comparable to those in the primary (Cancer Res 62:2942-50, 2002). In vitro reports recently observed that the lGFlR is uprcgulated in prostate cancer cells during the development of androgenIndependence. The objective of this study was to determine whether antisense mediated IGFI R downregulation could enhance the chemosensttivity of androgentndependent prostate cancer cells in vitro MATERIAL & METHODS: We transfectcd human androgclr-itldcpendent DU I45 prostate cancer cells with lGFl R antisense oligonucleotidcs (ASOs) or antisense RNA. Transfected cultures vvere treated with cisplatm, mitoxantrone, paclitaxel or vehicle control, and survival was measured by clonogcnic assay. DlJ I45 prostate cancer cell clones expressing ICiFl R antisense RN4 were devclopcd to assess in viva growth in nude mtcc. RESULTS: IGFIR ASOs caused dose-dependent inhibition of IGFI R exprcsston to levels of 30.40% of those m control-treated cells. This was accompanied by marked reductton m DU 145 growth measured by MTS assay, and clonogenic survival was rcduccd by up to 80%. Growth of’ IGFIR antisense RNA expressing DUl45 cell clones was ahnost completely abolished in viva. Furthermore. IGFlR ASOs enhanced chemosensitivity to cisplatin, mitoxantrone and paclitaxel with 1.5 to 2-fold reduction in IC-50. Ultrastructural analysis of the DU145 clones treated with cisplatin confirmed that the AS-IGFI R cells wcrc more susceptible to apoptosis.
M.. Furuya Y.. Takihana Y.. Araki I.. Tanabe N., Takeda M
University of Yamanashi,
lirology,
Yamanashi,
Japan
INTRODUCTION & OBJECTIVES: It is known that both cannabinoid receptor subtype I (CBI) and vanilloid receptor (VRI) are expressed not only in nerve system but in peripheral non-neuronal tissues, and are related to controlling proliferation in various cells. The present study was designed to Investigate the expression of CBI and VRI in human prostate and the effect of C‘BI and VRI on the proliferation of human prostate cancer ceils. MATERIAL & METHODS: Immunohistochemistry and RT-PCR were performed to determine the distribution and expression of CBI and VRI in human prostate tissues and cell lines, three normal cells and three malignant cells. DUl45 cells were treated with anandamide (non-selective agonist for (‘Bl) or capsaicm (selectivse agonist for VRI), and then cell viability was measured by MTT assay. To further analyse whether the cell death was medtated via CBI and VRI DUl45 cells were treated with selective antagonists of these receptors. RESULTS: In prostate tissues, CBI was highly expressed in epithelial cells, while VRI was in stromal regions. And in prostate cell lines, the expression of CBI was considerable in DU145 cells, while VRl was expressed in all cell lines, regardless of normal or malignant. In DUl45, anandamide elicited cell death, which was significantly inhibited by AM251. a selective antagonist for CBI receptor. And similarly. capsaicin elicited cell death, which was inhibited by capsarepine, a selective antagonist for VR I receptor.
CONCLUSIONS: These results mdicatc that lGFlR downregulation can enhance chemosensitivity of androgcn-independent prostate cancer in part via enhanced suscepttbility to apoptosis. In a clmical srttmg. targeted suppression of the IGFI R ming ASG technology may cnhancc the effects of conventional chemotherapy and therefore otfcrs promtsc as novel therapy for patients with androgen-independent disease.
CONCLUSIONS: CB I and VK I were expressed in human prostate tissues. In DUl45 cells, cell death induced by anandamidc or capsaicin was elicited via C‘B I or VR I, respectively.
135
136
PROSTATE CANCER AND IMMUNOTHERAPY: A PRELIMINARY STUDY ABOUT FREQUENCY OF TELOMERASE-SPECIFIC CDS+ LYMPHOCYTES Traverao P.‘, Romagnolt Carmignani Ci.
A.‘. Filact G’. Fravrga
of Cicnoa. DIMI. (icnoa.
INTRODUCTION & OBJECTIVES: Telomerase is an interesting tumourassociated antigen (TAA) because it is expressed m majority of cancers and because it is possible to generate telomerase specific CD8+ T lymphocytes (CTL) in the peripheral blood of healthy subjects or affected by cancer. We try to find out frequency of telomerase specific CTL in patients with prostate carcinoma. MATERIAL & METHODS: We used the l-540 antigen, encompassing the 540. 548 pepttde sequence of tclomcrase, that is presented by HLA-A2 class I molecules, We enrolled 5 HLA-,421 pattents: 4 wtth clinically localired prostate cancer (which have been subjected to a Radical Retropubic Prostatectomy) and I wtth metastatic prostate. 4 HLA-A2 t pattents without prostate cancer and 2 HLAA2- patients with prostate cancer were the control group. We did subsequent analyses a) calculation of number of CDX+ T lymphocytes reacting to HLA-A2 by tmmunofluorescence after staining with FITC-labelled HLA-tetramer loaded with l-540 peptidc, b) calculation of frequency of l-540 specitic CTI, precursors using “ltmitmg dtlution” c) generation of I-540 specific CTL lines. d RESUIXS: Percentage of l-S40 specific C‘DX+ T lymphocytes was significantly higher in patients HLA-A2+ affected by prostate cancer (P
Supplements
IN BPH AND PROSTATE CANCER PATIENTS
Mado-cbnchcr S ‘. Ponholrcr A.‘. Schat/l C; Ntjhuts L.‘. AdlercreutL II:
(i\ur 4 ‘, Hatdmger (i ‘. dc Vrtcs (’ ‘1
M.‘. Naselli A ‘_ Indiveri F.‘.
‘Ilntvcrsity of Cienoa. Ut-ology. Cicnoa. Italy. llniversity Italy
European
PHYTO-OESTROGENS
2 (2003) No. 1, pp. 36
I)onauspttal. Dept. of Urology, Vienna. Austrta. Umverstty of Vtennn, Dept. of llrology. Vienna. Austria. ‘University of Vienna. Institute of (‘ancer Research, Vienna, Austria, ‘Clnivcrsity of Helsinki, Inst. for Prey. Med. Nutr. and Cancer, Helsinki. Finland INTRODUCTION & OBJECTIVES: Epidemtological differences (west vs. cast) in prostate cancer (PC) incidence have been in part attributed to the consumption of phyto-oestrogens, which is substantially higher in Asians. Aim of our study was to determine whether serum levels of phyto-oestrogcns differ in BPH and PC-patients in an Austrian population and to estimate their correlation to PSA and endocrine parameters. MATERIAL & METHODS: In this cast control study, men (Caucasian only) wtth newly diagnosed, untreated prostate cancer and, as a control population, agcmatched patients with untreated lower urtnary tract symptoms due to benign prostatic hyperplasta (BPH) wcrc entered. In BPII-patients. PC was excluded clinically (negative digito-rectal examination and PSA) or histologically. Serum levels of the phyto-oestrogens datd7ein. gcnistcin and enterolactone were analysed by time-resolved fluoroimnlunoaasays; in parallel a serum cndocrinc study was obtained. RESULTS: A total of 194 men, 97 BPH- and 97 PC-patiems entcrcd this case control study. Mean age was comparable in BPH- (hX+lOyrs: meanistandard deviation) and PC- (64+6yrs; ~~0.05) casts. Enterolactone (50+209 vs. 22+34nmol/L), Daidzein (17+44 vs. 10+35nmol/L) and Genistein (33187 vs. 19+45nmol/L) wcrc all higher m BPH-patients, yet these differences did not reach statistical significance. We subsequently correlated phyto-oestrogens to PSA and endocrine parameters in the entire study population (n=l94); these analyses were controlled for age. No correlations between phyto-oestrogens and PSA, LH, total testosterone and estradiol were observed. Daidzein (p=O.O 19). Genistein (p=O.O02) and the sum of all three phyto-estrogens (p=O.OOS), however, were all positively correlated to FSH. CONCI.USIONS: These data suggest that photo-oestrogcns arc involved m the pathogenesis of prostate cancer and underline their potential for prostate cancer chemoprevention. The close correlation between FSH and phyto-oestrogens directs towards a potenttal regulatory mechanism and warrants further atudtcc.