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1.
Ultrasound Obstet Gynecol ; 60(3): 425-427, 2022 09.
Article in English | MEDLINE | ID: mdl-35653222

ABSTRACT

Anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies have been found in breast milk following both natural SARS-CoV-2 infection and coronavirus disease 2019 (COVID-19) vaccination. This was a prospective study to evaluate the temporal changes in amount and neutralization capacity of anti-SARS-CoV-2 antibodies in breast milk stimulated by natural infection and by vaccination. Serial breast milk samples were collected from postnatal women who were recruited through convenience sampling. We found a rapid increase in neutralizing SARS-CoV-2-specific antibodies in breast milk from both study groups. Amongst the infection group, the median immunoglobulin A (IgA) level was 16.99 (range, 0-86.56) ng/mL and median binding capacity was 33.65% (range, 0-67.65%), while in the vaccination group these were 30.80 (range, 0-77.40) ng/mL and 23.80% (range, 0-42.80%), respectively. In both groups, both binding capacity and IgA levels decreased progressively over time after peaking. Neutralizing activity had become undetectable by about 150 days after the first dose of the vaccine, but a vaccine booster dose restored secretion of neutralizing IgA, albeit with different levels of response in different individuals. This highlights the importance of the vaccine booster dose in sustaining neutralizing antibody levels in breast milk, which may potentially provide protection for very young children, who cannot receive the COVID-19 vaccine. © 2022 International Society of Ultrasound in Obstetrics and Gynecology.


Subject(s)
COVID-19 Vaccines , COVID-19 , Antibodies, Viral , COVID-19/prevention & control , Child , Child, Preschool , Female , Humans , Immunoglobulin A , Milk, Human , Prospective Studies , SARS-CoV-2 , Vaccination
2.
Br J Cancer ; 125(11): 1462-1465, 2021 11.
Article in English | MEDLINE | ID: mdl-34316019

ABSTRACT

The National Cancer Imaging Translational Accelerator (NCITA) is creating a UK national coordinated infrastructure for accelerated translation of imaging biomarkers for clinical use. Through the development of standardised protocols, data integration tools and ongoing training programmes, NCITA provides a unique scalable infrastructure for imaging biomarker qualification using multicentre clinical studies.


Subject(s)
Biomarkers, Tumor/metabolism , Diagnostic Tests, Routine/methods , Neoplasms/diagnostic imaging , Humans , Research Design , United Kingdom
3.
Br J Cancer ; 111(3): 424-9, 2014 Jul 29.
Article in English | MEDLINE | ID: mdl-24946001

ABSTRACT

BACKGROUND: Salvage therapeutic options for biochemical failure after primary radiation-based therapy include radical prostatectomy, cryoablation, high-intensity focused ultrasound (HIFU), brachytherapy (for post-EBRT patients) and androgen deprivation therapy (ADT). ADT and salvage prostate cryoablation (SPC) are two commonly considered treatment options for RRPC. However, there is an urgent need for high-quality clinical studies to support evidence-based decisions on treatment choice. Our study aims to determine the feasibility of randomising men with RRPC for treatment with ADT and SPC. METHODS: The randomised controlled trial (CROP) was developed, which incorporated protocols to assess parameters relating to cryotherapy procedures and provide training workshops for optimising patient recruitment. Analysis of data from the recruitment phase and patient questionnaires was performed. RESULTS: Over a period of 18 months, 39 patients were screened for eligibility. Overall 28 patients were offered entry into the trial, but only 7 agreed to randomisation. The majority reason for declining entry into the trial was an unwillingness to be randomised into the study. 'Having the chance of getting cryotherapy' was the major reason for accepting the trial. Despite difficulty in retrieving cryotherapy temperature parameters from prior cases, 9 of 11 cryotherapy centres progressed through the Cryotherapists Qualification Process (CQP) and were approved for recruiting into the CROP study. CONCLUSIONS: Conveying equipoise between the two study arms for a salvage therapy was challenging. The use of delayed androgen therapy may have been seen as an inferior option. Future cohort studies into available salvage options (including prostate cryotherapy) for RRPC may be more acceptable to patients than randomisation within an RCT.


Subject(s)
Neoplasm Recurrence, Local/drug therapy , Prostatic Neoplasms/drug therapy , Androgen Antagonists/therapeutic use , Cryosurgery , Feasibility Studies , Humans , Male , Multicenter Studies as Topic , Neoplasm Recurrence, Local/surgery , Patient Acceptance of Health Care , Prostatic Neoplasms/radiotherapy , Prostatic Neoplasms/surgery , Randomized Controlled Trials as Topic , Salvage Therapy
4.
Br J Cancer ; 109(4): 950-6, 2013 Aug 20.
Article in English | MEDLINE | ID: mdl-23880823

ABSTRACT

BACKGROUND: Evidence increasingly supports that prostate cancer is initiated by the malignant transformation of stem cells (SCs). Furthermore, many SC-signalling pathways are shown to be shared in prostate cancer. Therefore, we planned transcriptome characterisation of adult prostate SCs as a strategy to consider new targets for cancer treatment. METHODS: Intuitive pathway analysis was used for putative target discovery in 12 matched selections of human prostate SCs, transiently amplifying cells and terminally differentiated cells. These were pooled into three groups according to the stage of differentiation for mRNA microarray analysis. Targets identified were validated using uncultured primary tissue (n=12), functional models of prostate cancer and a tissue microarray consisting of benign (n=42) and malignant prostate (n=223). RESULTS: A deficiency in class 1 UDP glucuronosyltransferase (UGT) enzymes (UGT1A) was identified in prostate SCs, which are involved in androgen catabolism. Class 1 UGT enzyme expression was also downregulated in cancer SCs and during progression to metastatic castration-resistant prostate cancer (CRPC). Reduction of UGT1A expression in vitro was seen to improve cell survival and increase androgen receptor (AR) activity, as shown by upregulation of prostate-specific antigen expression. INTERPRETATION: Inactivation of intracellular androgen catabolism represents a novel mechanism to maintain AR activity during CRPC.


Subject(s)
Adult Stem Cells/enzymology , Gene Expression Regulation, Neoplastic , Glucuronosyltransferase/genetics , Neoplastic Stem Cells/enzymology , Prostate/enzymology , Prostatic Neoplasms/enzymology , RNA, Messenger/analysis , Androgens/metabolism , Cell Line , Cell Line, Tumor , Cell Survival , Down-Regulation , Gene Expression Profiling , Glucuronosyltransferase/metabolism , Humans , Kallikreins/metabolism , Male , Prostate/cytology , Prostate-Specific Antigen/metabolism , Prostatic Neoplasms/genetics , Real-Time Polymerase Chain Reaction , Receptors, Androgen/metabolism , Transcriptome , Up-Regulation
5.
Br J Cancer ; 108(1): 139-48, 2013 Jan 15.
Article in English | MEDLINE | ID: mdl-23321516

ABSTRACT

BACKGROUND: Prostate cancer cell growth is dependent upon androgen receptor (AR) activation, which is regulated by specific kinases. The aim of the current study is to establish if AR phosphorylation by Cdk1 or ERK1/2 is of prognostic significance. METHODS: Scansite 2.0 was utilised to predict which AR sites are phosphorylated by Cdk1 and ERK1/2. Immunohistochemistry for these sites was then performed on 90 hormone-naive prostate cancer specimens. The interaction between Cdk1/ERK1/2 and AR phosphorylation was investigated in vitro using LNCaP cells. RESULTS: Phosphorylation of AR at serine 515 (pAR(S515)) and PSA at diagnosis were independently associated with decreased time to biochemical relapse. Cdk1 and pCdk1(161), but not ERK1/2, correlated with pAR(S515). High expression of pAR(S515) in patients with a PSA at diagnosis of ≤20 ng ml(-1) was associated with shorter time to biochemical relapse (P=0.019). This translated into a reduction in disease-specific survival (10-year survival, 38.1% vs 100%, P<0.001). In vitro studies demonstrated that treatment with Roscovitine (a Cdk inhibitor) caused a reduction in pCdk1(161) expression, pAR(S515)expression and cellular proliferation. CONCLUSION: In prostate cancer patients with PSA at diagnosis of ≤20 ng ml(-1), phosphorylation of AR at serine 515 by Cdk1 may be an independent prognostic marker.


Subject(s)
MAP Kinase Signaling System/physiology , Prostatic Neoplasms/metabolism , Purines/pharmacokinetics , Receptors, Androgen/metabolism , Aged , Biomarkers, Tumor/antagonists & inhibitors , CDC2 Protein Kinase/antagonists & inhibitors , CDC2 Protein Kinase/metabolism , Disease-Free Survival , Humans , Male , Phosphorylation , Prognosis , Prostate-Specific Antigen/metabolism , Recurrence , Roscovitine , Serine/metabolism
6.
Br J Cancer ; 108(1): 149-54, 2013 Jan 15.
Article in English | MEDLINE | ID: mdl-23321517

ABSTRACT

BACKGROUND: Aberrant mitogen/extracellular signal-regulated kinase 5 (MEK5)-extracellular signal-regulated protein kinase 5 (ERK5)-mediated signalling has been implicated in a number of tumour types including prostate cancer (CaP). The mechanism for ERK5 activation in CaP remains to be fully elucidated. Studies have recently implicated the role of microRNA (miRNA) mir143 expression in the regulation of ERK5 expression. METHODS: We utilised a tissue microarray (TMA) of 530 CaP cores from 168 individual patients and stained for both mir143 and ERK5. These TMAs were scored by a combination of observer and automated methods. RESULTS: We observed a strong inverse relation between ERK5 and mir143, which manifested itself most strongly in the subgroup of 417 cores with non-zero mir143 and ERK5 immunoreactivity, or with only one of mir143 or ERK5 being zero (cc=0.2558 and P<0.0001). Mir143 neither correlate with Gleason scores or prostate-specific antigen levels, nor was it a predictor of disease-specific survival on univariate analysis. CONCLUSION: Although the mechanism for ERK5 activation in CaP remains to be fully elucidated, we have further validated the potential role of mir143 in regulating ERK5 levels in the clinical context. In addition, we demonstrate that the automated counting method for nuclear ERK5 is a clinically useful alterative to observer counting method in patient stratification in the context of ERK5 targeting therapy.


Subject(s)
MicroRNAs/metabolism , Mitogen-Activated Protein Kinase 7/metabolism , Prostatic Neoplasms/genetics , Gene Expression Regulation , Humans , Male , Prostatic Neoplasms/metabolism
7.
Br J Cancer ; 107(9): 1554-63, 2012 Oct 23.
Article in English | MEDLINE | ID: mdl-23093296

ABSTRACT

BACKGROUND: Cell line models suggest that activation of NFκB is associated with progression of prostate cancer. This pathway may be a therapeutic target if these observations translate to clinical specimens. METHODS: Immunohistochemistry measured NFκBp65 (p65), NFκBp65 nuclear localisation signal (NLS), NFκBp65 phosphorylated at ser 276 (p65(ser276)), NFκBp65 phosphorylated at ser 536 (p65(ser536)), IκBα phosphorylated at ser 32/36 (pIκBα(ser32/36)) and MMP-9 protein expression in 61 matched hormone naive prostate cancer (HNPC) and castrate-resistant prostate cancer (CRPC) tumours. Animal and cell models were used to investigate the role of NFκB inhibition in prostate carcinogenesis. RESULTS: In HNPC tumours, NLS expression significantly associated with a shorter time to disease recurrence and disease-specific death. In CRPC tumours p65, pIκBα(ser32/36) and MMP-9 expression significantly associated with shorter time to death from disease recurrence and shorter disease-specific death. MMP-9 and pIκBα(ser32/36) expression significantly associated with metastases at recurrence and were independent of Gleason sum and prostate-specific antigen at recurrence. Expression of phosphorylated Akt was associated with increased p65 activation in mouse models and inhibition of NFκB in LNCaP cells significantly reduced cellular proliferation and induced apoptosis. CONCLUSION: These results provide further evidence that the NFκB pathway could be exploited as a target for CRPC.


Subject(s)
NF-kappa B/metabolism , Prostatic Neoplasms/metabolism , Prostatic Neoplasms/pathology , Aged , Animals , Biomarkers, Tumor/biosynthesis , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Cell Growth Processes/physiology , Cell Line, Tumor , Cohort Studies , Disease Models, Animal , Disease Progression , Humans , I-kappa B Proteins/metabolism , Immunohistochemistry , Kaplan-Meier Estimate , Male , Matrix Metalloproteinase 9/biosynthesis , Mice , NF-KappaB Inhibitor alpha , NF-kappa B/biosynthesis , NF-kappa B/genetics , Neoplasms, Hormone-Dependent/genetics , Neoplasms, Hormone-Dependent/metabolism , Neoplasms, Hormone-Dependent/pathology , Phosphorylation , Prostatic Neoplasms/genetics , Signal Transduction , Transcription Factor RelA/metabolism , Up-Regulation
8.
Br J Cancer ; 105(9): 1362-9, 2011 Oct 25.
Article in English | MEDLINE | ID: mdl-21952621

ABSTRACT

BACKGROUND: Prostate cancer (PC) represents a global health issue. Treatment for locally advanced and metastatic PC remains unsatisfactory. The androgen receptor (AR) has been validated in having a key role in both naïve and castrate-resistant PC (CRPC). However, the significance of other signalling pathways in CRPC is less well validated. METHODS: To gain a better insight into the molecular signalling cascades involved in clinical CRPC, we performed gene expression profiling using the Illumina DASL assay and studied matched hormone-naive (HN) and CR prostate tumours (n=10 pairs). Ingenuity Pathways Analysis (IPA) was used to identify potential networks involved, and further validation was performed in in vitro cell models and clinical tumours. RESULTS: Expression of 50 genes was significantly different between HN and CRPC. IPA revealed two networks of particular interest, including AR and FGFR1, respectively. FGFR1 expression was confirmed to be significantly upregulated in CRPC (P ≤ 0.005), and abnormal FGFR1 expression was associated with shorter time to biochemical relapse in HNPC (P=0.006) and less favourable disease-specific survival in CRPC (P=0.018). CONCLUSION: For the first time, our gene expression profiling experiment on archival tumour materials has identified upregulated FGFR1 expression to be associated with PC progression to the CR state.


Subject(s)
Androgen Antagonists/pharmacology , Castration , Gonadotropin-Releasing Hormone/pharmacology , Prostatic Neoplasms/genetics , Receptor, Fibroblast Growth Factor, Type 1/genetics , Aged , Aged, 80 and over , Cell Line, Tumor , Gene Expression Profiling , Humans , Male , Middle Aged , Neoplasms, Hormone-Dependent/genetics , Orchiectomy , Receptors, Androgen/genetics , Recurrence , Signal Transduction , Up-Regulation
9.
Cell Death Dis ; 2: e124, 2011 Mar 03.
Article in English | MEDLINE | ID: mdl-21368895

ABSTRACT

Mutations in the Ras family of proteins (predominantly in H-Ras) occur in approximately 40% of urothelial cell carcinoma (UCC). However, relatively little is known about subsequent mutations/pathway alterations that allow tumour progression. Indeed, expressing mutant H-Ras within the mouse bladder does not lead to tumour formation, unless this is expressed at high levels. The Wnt signalling pathway is deregulated in approximately 25% of UCC, so we examined if this correlated with the activation of MAPK signalling in human UCC and found a significant correlation. To test the functional significance of this association we examined the impact of combining Ras mutation (H-Ras(Q61L) or K-Ras(G12D)) with an activating ß-catenin mutation within the mouse bladder using Cre-LoxP technology. Although alone, neither Ras mutation nor ß-catenin activation led to UCC (within 12 months), mice carrying both mutations rapidly developed UCC. Mechanistically this was associated with reduced levels of p21 with dependence on the MAPK signalling pathway. Moreover, tumours from these mice were sensitive to MEK inhibition. Importantly, in human UCC there was a negative correlation between levels of p-ERK and p21 suggesting that p21 accumulation may block tumour progression following Ras mutation. Taken together these data definitively show Ras pathway activation strongly cooperates with Wnt signalling to drive UCC in vivo.


Subject(s)
Mutation , Urinary Bladder Neoplasms/genetics , Urinary Bladder Neoplasms/metabolism , Urinary Bladder Neoplasms/pathology , beta Catenin/metabolism , ras Proteins/genetics , Animals , Female , Humans , Male , Mice , Mice, Transgenic , Neoplastic Processes , Signal Transduction , Wnt Proteins/genetics , Wnt Proteins/metabolism , beta Catenin/genetics , ras Proteins/metabolism
10.
Br J Cancer ; 104(4): 664-72, 2011 Feb 15.
Article in English | MEDLINE | ID: mdl-21266977

ABSTRACT

BACKGROUND: Aberrant mitogen/extracellular signal-regulated kinase 5 (MEK5)-extracellular signal-regulated protein kinase 5 (ERK5)-mediated signalling has been implicated in a number of tumour types including prostate cancer (PCa). The molecular basis of ERK5-driven carcinogenesis and its clinical relevance remain to be fully characterised. METHODS: Modulation of ERK5 expression or function in human PCa PC3 and PC3-ERK5 (stably transfected with ERK5) cells was performed using siRNA-mediated knockdown or the MEK inhibitor PD18435 respectively. In vitro significance of ERK5 signalling was assessed by assays for proliferation, motility, invasion and invadopodia. Expression of matrix metalloproteinases/tissue inhibitors of metalloproteases was determined by Q-RT-PCR. Extracellular signal-regulated protein kinase 5 expression in primary and metastatic PCa was examined using immunohistochemistry. RESULTS: Reduction of ERK5 expression or signalling significantly inhibited the motility and invasive capability of PC3 cells. Extracellular signal-regulated protein kinase 5-mediated signalling significantly promoted formation of in vivo metastasis in an orthotopic PCa model (P<0.05). Invadopodia formation was also enhanced by forced ERK5 expression in PC3 cells. Furthermore, in metastatic PCa, nuclear ERK5 immunoreactivity was significantly upregulated when compared with benign prostatic hyperplasia and primary PCa (P=0.013 and P<0.0001, respectively). CONCLUSION: Our in vitro, in vivo and clinical data support an important role for the MEK5-ERK5 signalling pathway in invasive PCa, which represents a potential target for therapy in primary and metastatic PCa.


Subject(s)
Mitogen-Activated Protein Kinase 7/physiology , Prostatic Hyperplasia/pathology , Prostatic Neoplasms/pathology , Animals , Benzamides/pharmacology , Cell Movement/drug effects , Cell Movement/genetics , Cell Proliferation/drug effects , Cells, Cultured , Drug Evaluation, Preclinical , Gene Expression Regulation, Neoplastic/drug effects , Gene Expression Regulation, Neoplastic/genetics , Humans , MAP Kinase Kinase 5/genetics , MAP Kinase Kinase 5/metabolism , MAP Kinase Kinase 5/physiology , MAP Kinase Signaling System/drug effects , MAP Kinase Signaling System/genetics , MAP Kinase Signaling System/physiology , Male , Mice , Mice, Nude , Mitogen-Activated Protein Kinase 7/genetics , Mitogen-Activated Protein Kinase 7/metabolism , Neoplasm Invasiveness , Phenotype , Prostatic Hyperplasia/genetics , Prostatic Hyperplasia/metabolism , Prostatic Neoplasms/genetics , Prostatic Neoplasms/metabolism , Protein Kinase Inhibitors/pharmacology , RNA, Small Interfering/pharmacology , Transfection , Transplantation, Heterologous
11.
Oncogene ; 30(2): 178-89, 2011 Jan 13.
Article in English | MEDLINE | ID: mdl-20818428

ABSTRACT

Although deregulation of the Wnt signalling pathway has been implicated in urothelial cell carcinoma (UCC), the functional significance is unknown. To test its importance, we have targeted expression of an activated form of ß-catenin to the urothelium of transgenic mice using Cre-Lox technology (UroIICRE(+) ß-catenin(exon3/+)). Expression of this activated form of ß-catenin led to the formation of localized hyperproliferative lesions by 3 months, which did not progress to malignancy. These lesions were characterized by a marked increase of the phosphatase and tensin homologue (PTEN) tumour suppressor protein. This appears to be a direct consequence of activating Wnt signalling in the bladder as conditional deletion of the adenomatous polyposis coli (Apc) gene within the adult bladder led rapidly to coincident ß-catenin and PTEN expression. This PTEN expression blocked proliferation. Next, we combined PTEN deficiency with ß-catenin activation and found that this caused papillary UCC. These tumours had increased pAKT signalling and were dependent on mammalian target of rapamycin (mTOR). Importantly, in human UCC, there was a significant correlation between high levels of ß-catenin and pAKT (and low levels of PTEN). Taken together these data show that deregulated Wnt signalling has a critical role in promoting UCC, and suggests that human UCC that have high levels of Wnt and PI3 kinase signalling may be responsive to mTOR inhibition.


Subject(s)
Carcinoma, Papillary/metabolism , Cell Transformation, Neoplastic/metabolism , PTEN Phosphohydrolase/metabolism , Urinary Bladder Neoplasms/chemically induced , beta Catenin/metabolism , Adenomatous Polyposis Coli Protein/genetics , Animals , Cell Proliferation , Cohort Studies , Female , Humans , Hyperplasia/chemically induced , Male , Mice , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/analysis , Proto-Oncogene Proteins c-akt/metabolism , Sequence Deletion , TOR Serine-Threonine Kinases/metabolism , Urinary Bladder/metabolism , Urothelium/metabolism , Wnt Proteins/metabolism
12.
Br J Cancer ; 103(5): 701-7, 2010 Aug 24.
Article in English | MEDLINE | ID: mdl-20648010

ABSTRACT

BACKGROUND: The accuracy of prostate-specific antigen (PSA) testing in prostate cancer detection is constrained by low sensitivity and specificity. Dysregulated expression of minichromosome maintenance (Mcm) 2-7 proteins is an early event in epithelial multistep carcinogenesis and thus MCM proteins represent powerful cancer diagnostic markers. In this study we investigate Mcm5 as a urinary biomarker for prostate cancer detection. METHODS: Urine was obtained from 88 men with prostate cancer and from two control groups negative for malignancy. A strictly normal cohort included 28 men with complete, normal investigations, no urinary calculi and serum PSA <2 ng ml(-1). An expanded control cohort comprised 331 men with a benign final diagnosis, regardless of PSA level. Urine was collected before and after prostate massage in the cancer patient cohort. An immunofluorometric assay was used to measure Mcm5 levels in urine sediments. RESULTS: The Mcm5 test detected prostate cancer with 82% sensitivity (confidence interval (CI)= 72-89%) and with a specificity ranging from 73 (CI=68-78%) to 93% (CI=76-99%). Prostate massage led to increased Mcm5 signals compared with pre-massage samples (median 3440 (interquartile range (IQR) 2280 to 5220) vs 2360 (IQR <1800 to 4360); P=0.009), and was associated with significantly increased diagnostic sensitivity (82 vs 60%; P=0.012). CONCLUSIONS: Urinary Mcm5 detection seems to be a simple, accurate and noninvasive method for identifying patients with prostate cancer. Large-scale prospective trials are now required to evaluate this test in diagnosis and screening.


Subject(s)
Cell Cycle Proteins/urine , Prostatic Neoplasms/urine , Aged , Fluoroimmunoassay , Humans , Male , Massage , Pilot Projects , Prostate , Prostate-Specific Antigen/blood , Prostatic Neoplasms/diagnosis , Sensitivity and Specificity
13.
Cell Death Dis ; 1: e72, 2010 Sep 09.
Article in English | MEDLINE | ID: mdl-21364676

ABSTRACT

Pro-survival signalling mediated by the androgen receptor (AR) is implicated as a key contributor to prostate carcinogenesis. As prostate tumours are characterized by nutrient-poor, hypoxic and acidified microenvironments, one mechanism whereby AR signalling may contribute to survival is by promoting adaptation to cellular stress. Here we have identified a novel role for AR in the inhibition of autophagy induced by serum withdrawal. This blockade is attributed to AR-mediated upregulation of the endoplasmic reticulum (ER) chaperone glucose-regulated protein 78/BiP (Grp78/BiP), and occurs independently of ER stress response pathway activation. Interestingly, AR activation did not affect serum starvation-induced mammalian target of rapamycin inhibition, illustrating that the adaptive role for androgens lies not in the ability to modulate nutrient sensing, but in the promotion of ER stability. Finally, we show that the adaptive advantage conferred by AR-mediated Grp78/BiP upregulation is temporary, as upon chronic serum starvation, AR activation delayed but did not suppress the onset of autophagy and cell death. This study reveals a novel mechanism whereby maintained AR signalling promotes temporary adaptation to cellular stress and in turn may contribute to the evasion of prostate tumour cell death.


Subject(s)
Androgens/pharmacology , Apoptosis , Autophagy , Heat-Shock Proteins/metabolism , Prostatic Neoplasms/metabolism , Cell Line, Tumor , Endoplasmic Reticulum Chaperone BiP , Heat-Shock Proteins/genetics , Humans , Male , RNA Interference , RNA, Small Interfering/metabolism , Receptors, Androgen/metabolism , Signal Transduction , TOR Serine-Threonine Kinases/antagonists & inhibitors , TOR Serine-Threonine Kinases/metabolism , Up-Regulation
14.
Br J Cancer ; 101(11): 1891-9, 2009 Dec 01.
Article in English | MEDLINE | ID: mdl-19888221

ABSTRACT

BACKGROUND: The fibroblast growth factor (FGF) axis is an important mitogenic stimulus in prostate carcinogenesis. We have previously reported that transcript level of human similar expression to FGF (hSef), a key regulator of this pathway, is downregulated in clinical prostate cancer. In this study we further analysed the role of hSef in prostate cancer. METHODS: hSef function was studied in in vitro and in vivo prostate cancer models using stable over-expression clones. Protein expression of hSef was studied in a comprehensive tissue microarray. RESULTS: Stable over-expression of hSef resulted in reduced in vitro cancer cell proliferation, migration and invasive potential. In an in vivo xenograft model, the expression of hSef significantly retarded prostate tumour growth as compared with empty vector (P=0.03) and non-transfected (P=0.0001) controls. Histological examination further showed a less invasive tumour phenotype and reduced numbers of proliferating cells (P=0.0002). In signalling studies, hSef inhibited FGF-induced ERK phosphorylation, migration to the nucleus and activation of a reporter gene. Constitutively active Ras, however, was able to reverse these effects, suggesting that hSef exerts an effect either above or at the level of Ras in prostate cancer cells. In a large tissue microarray, we observed a significant loss of hSef protein in high-grade (P<0.0001) and metastatic (P<0.0001) prostate cancer. CONCLUSIONS: Considered together, the role of hSef in attenuating FGF signalling and evidence of downregulation in advanced tumours argue strongly for a tumour suppressor function in human prostate cancer.


Subject(s)
Fibroblast Growth Factors/antagonists & inhibitors , Prostatic Neoplasms/metabolism , Receptors, Interleukin/biosynthesis , Animals , Cell Growth Processes/physiology , Cell Line, Tumor , Down-Regulation , Fibroblast Growth Factors/metabolism , Gene Expression Regulation, Neoplastic , Humans , Male , Mice , Mice, Nude , Prostatic Neoplasms/genetics , Prostatic Neoplasms/pathology , Receptor Protein-Tyrosine Kinases/metabolism , Receptors, Interleukin/genetics , Receptors, Interleukin/metabolism , Signal Transduction , Transfection
15.
J Pathol ; 215(1): 67-77, 2008 May.
Article in English | MEDLINE | ID: mdl-18273831

ABSTRACT

The RNA-binding protein Sam68 has been reported to be up-regulated in clinical cases of prostate cancer (PCa), where it is thought to contribute to cell proliferation and survival. Consistent with this, we observed over-expression of Sam68 in a panel of clinical prostate tumours as compared with benign controls. Since Sam68 is implicated in a number of signalling pathways, we reasoned that its role in PCa may involve modulation of the androgen receptor (AR) signalling cascade, which drives the onset and progression of PCa. We found that Sam68 interacts with the AR in vivo in LNCaP cells, and is dynamically recruited to androgen response elements within the promoter region of the prostate-specific antigen (PSA) gene. Based on its known functions and nuclear location, Sam68 might either: (a) co-regulate AR-dependent transcription positively or negatively; or (b) modulate AR-dependent alternative splicing by enhancing incorporation of a Sam68-responsive exon transcribed under the control of an androgen-responsive promoter. We tested these possibilities using functional assays. Both wild-type Sam68 protein and the Sam68(V229F) mutant, which is impaired in RNA binding, functioned as a ligand-dependent AR co-activator on an androgen-regulated reporter gene. In contrast, splicing of a Sam68-responsive variable exon, transcribed under control of an androgen-responsive promoter, was strongly repressed in the presence of AR and androgens. This splicing inhibition was reversed by ectopic expression of Sam68 but enhanced by Sam68(V229F). These results demonstrate that Sam68 has separable effects on AR-regulated transcriptional activity and alternative splicing, both of which may affect PCa phenotypes.


Subject(s)
Adaptor Proteins, Signal Transducing/metabolism , Alternative Splicing , DNA-Binding Proteins/metabolism , Gene Expression Regulation, Neoplastic , Prostatic Neoplasms/metabolism , RNA-Binding Proteins/metabolism , Receptors, Androgen/metabolism , Transcription, Genetic/physiology , Androgens/metabolism , Case-Control Studies , Gene Expression , Gene Expression Profiling , Humans , Hyaluronan Receptors/genetics , Immunohistochemistry , Male , Mutation , Oligonucleotide Array Sequence Analysis , Prostate-Specific Antigen/metabolism
16.
Oncogene ; 27(21): 2978-88, 2008 May 08.
Article in English | MEDLINE | ID: mdl-18071319

ABSTRACT

Abnormal intracellular signaling contributes to carcinogenesis and may represent novel therapeutic targets. mitogen/extracellular signal-regulated kinase kinase-5 (MEK5) overexpression is associated with aggressive prostate cancer. In this study, we examined the role of extracellular signal-regulated kinase (ERK5, an MAPK and specific substrate for MEK5) in prostate cancer. ERK5 immunoreactivity was significantly upregulated in high-grade prostate cancer when compared to benign prostatic hyperplasia (P<0.0001). Increased ERK5 cytoplasmic signals correlated closely with Gleason sum score (P<0.0001), bony metastases (P=0.0044) and locally advanced disease at diagnosis (P=0.0023), with a weak association with shorter disease-specific survival (P=0.036). A subgroup of patients showed strong nuclear ERK5 localization, which correlated with poor disease-specific survival and, on multivariant analysis, was an independent prognostic factor (P<0.0001). Analysis of ERK5 expression in matched tumor pairs (before and after hormone relapse, n=26) revealed ERK5 nuclear expression was significantly associated with hormone-insensitive disease (P=0.0078). Similarly, ERK5 protein expression was increased in an androgen-independent LNCaP subline. We obtained the following in vitro and in vivo evidence to support the above expression data: (1) cotransfection of ERK5wt and MEK5D constructs in PC3 cells results in predominant ERK5 nuclear localization, similar to that observed in aggressive clinical disease; (2) ERK5-overexpressing PC3 cells have enhanced proliferative, migrative and invasive capabilities in vitro (P<0.0001), and were dramatically more efficient in forming tumors, with a shorter mean time for tumors to reach a critical volume of 1000 mm(3), in vivo (P<0.0001); (3) the MEK1 inhibitor, PD184352, blocking ERK1/2 activation at low dose, did not suppress proliferation but did significantly decrease proliferation at a higher dose required to inhibit ERK5 activation. Taken together, our results establish the potential importance of ERK5 in aggressive prostate cancer.


Subject(s)
MAP Kinase Kinase 5/genetics , Prostatic Neoplasms/enzymology , Aged , Aged, 80 and over , Cell Proliferation , Disease-Free Survival , Humans , Immunohistochemistry , Male , Middle Aged , Prognosis , Prostatic Neoplasms/pathology
17.
Phytother Res ; 22(1): 131-3, 2008 Jan.
Article in English | MEDLINE | ID: mdl-17685390

ABSTRACT

The effects of VI-28 (a Yang-invigorating Chinese herbal formula) treatment on the renal mitochondrial antioxidant system and susceptibility to gentamicin-induced nephrotoxicity were investigated in rats. VI-28 treatment (80 or 240 mg/kg/day x 12) enhanced the renal mitochondrial antioxidant system, as indicated by dose-dependent increases in the level/activities of reduced glutathione, Mn-superoxide dismutase, Se-glutathione peroxidase and glutathione S-transferases. VI-28 treatment protected against nephrotoxicity induced by gentamicin administration (100 mg/kg/day x 8) and the nephroprotection was associated with an enhancement in the renal mitochondrial antioxidant system. In conclusion, VI-28 treatment enhanced the renal mitochondrial antioxidant system, thereby protecting against gentamicin nephrotoxicity.


Subject(s)
Drugs, Chinese Herbal/therapeutic use , Kidney Diseases/drug therapy , Kidney/drug effects , Yang Deficiency/drug therapy , Animals , Antioxidants/metabolism , Drugs, Chinese Herbal/chemistry , Gentamicins , Glutathione/metabolism , Glutathione Transferase/metabolism , Kidney/metabolism , Kidney/pathology , Kidney Diseases/chemically induced , Kidney Diseases/metabolism , Male , Medicine, Chinese Traditional , Mitochondria/drug effects , Mitochondria/metabolism , Phytotherapy , Rats , Rats, Sprague-Dawley , Superoxide Dismutase/metabolism
18.
Endocr Relat Cancer ; 14(3): 613-24, 2007 Sep.
Article in English | MEDLINE | ID: mdl-17914092

ABSTRACT

GnRH II has important functional effects in steroid hormone-dependent tumours. Here we investigated the expression and regulation of GnRH II in prostate cancer. GnRH II protein was equally expressed in benign (73%) and malignant (78%) biopsies studied in a prostate tissue microarray (P = 0.779). There was no relationship between expression and clinical parameters in the cancer cohort. GnRH II was, however, significantly reduced in tumour biopsies following hormone ablation. This was further investigated in a prostate xenograft model where androgens increased GnRH II levels, while their withdrawal reduced it. In cell lines, we confirmed high levels of GnRH II in androgen receptor (AR)-positive LNCaP cells but low levels in AR-negative PC3 cells. In LNCaP cells, GnRH II induction by androgens was blocked by the AR inhibitor casodex, but not by cycloheximide treatment. Sequence analysis subsequently revealed a putative androgen response element in the upstream region of the GnRH II gene and direct interaction with the AR was confirmed in chromatin immunoprecipitation experiments. Finally, to test whether the effects of GnRH II were dependent on AR expression, LNCaP and PC3 cells were exposed to exogenous peptide. In both cell lines, GnRH II inhibited cell proliferation and migration, suggesting that its function is independent of AR status. These results provide evidence that GnRH II is widely expressed in prostate cancer and is an AR-regulated gene. Further studies are warranted to characterise the effects of GnRH II on prostate cancer cells and investigate its potential value as a novel therapy.


Subject(s)
Gene Expression Regulation, Neoplastic , Gonadotropin-Releasing Hormone/analogs & derivatives , Prostatic Neoplasms/genetics , Receptors, Androgen/physiology , Androgens/pharmacology , Animals , Base Sequence , Binding Sites , Cell Proliferation/drug effects , Gene Expression Regulation, Neoplastic/drug effects , Gonadotropin-Releasing Hormone/genetics , Gonadotropin-Releasing Hormone/metabolism , Humans , Male , Mice , Mice, Nude , Molecular Sequence Data , Neoplasm Invasiveness , Neoplasms, Hormone-Dependent/genetics , Neoplasms, Hormone-Dependent/metabolism , Neoplasms, Hormone-Dependent/pathology , Prostatic Neoplasms/metabolism , Prostatic Neoplasms/pathology , Receptors, Androgen/metabolism , Response Elements , Transplantation, Heterologous , Tumor Cells, Cultured
19.
J Pathol ; 213(1): 82-90, 2007 Sep.
Article in English | MEDLINE | ID: mdl-17607666

ABSTRACT

Fibroblast growth factor receptors (FGFRs) mediate the tumourigenic effects of FGFs in prostate cancer. These receptors are therefore potential therapeutic targets in the development of inhibitors to this pathway. To identify the most relevant targets, we simultaneously investigated FGFR1-4 expression using a prostate cancer tissue microarray (TMA) and in laser capture microdissected (LCM) prostate epithelial cells. In malignant prostates (n = 138) we observed significant FGFR1 and FGFR4 protein over-expression in comparison with benign prostates (n = 58; p < 0.0001). FGFR1 was expressed at high levels in the majority of tumours (69% of grade 3 or less, 74% of grade 4 and 70% of grade 5), while FGFR4 was strongly expressed in 83% of grade 5 cancers but in only 25% of grade 1-3 cancers (p < 0.0001). At the transcript level we observed a similar pattern, with FGFR1 and FGFR4 mRNA over-expressed in malignant epithelial cells compared to benign cells (p < 0.0005 and p < 0.05, respectively). While total FGFR2 was increased in some cancers, there was no association between expression and tumour grade or stage. Transcript analysis, however, revealed a switch in the predominant isoform expressed from FGFR2IIIb to FGFR2IIIc among malignant epithelial cells. In contrast, protein and transcript expression of FGFR3 was very similar between benign and cancer biopsies. The functional effect of targeting FGFR4 in prostate cancer cells has not previously been investigated. In in vitro experiments, suppression of FGFR4 by RNA interference effectively blocked prostate cancer cell proliferation (p < 0.0001) and invasion (p < 0.001) in response to exogenous stimulation. This effect was evident regardless of whether the cells expressed the FGFR4 Arg388 or Gly388 allele. In parallel experiments, FGFR3 suppression had no discernible effect on cancer cell behaviour. These results suggest evidence of selective over-expression of FGFR1 and FGFR4 in clinical prostate cancer and support the notion of targeted inhibition of these receptors to disrupt FGF signalling.


Subject(s)
Gene Expression Regulation, Neoplastic , Prostatic Neoplasms/genetics , Receptors, Fibroblast Growth Factor/genetics , Case-Control Studies , Cell Line, Tumor , Cell Proliferation , Gene Expression Profiling , Humans , Immunohistochemistry , Male , Microdissection , Microscopy, Confocal , Oligonucleotide Array Sequence Analysis , Polymorphism, Single Nucleotide , Prostatic Neoplasms/metabolism , Protein Isoforms/genetics , RNA Interference , RNA, Small Interfering/pharmacology , Receptor, Fibroblast Growth Factor, Type 1/analysis , Receptor, Fibroblast Growth Factor, Type 1/genetics , Receptor, Fibroblast Growth Factor, Type 1/metabolism , Receptor, Fibroblast Growth Factor, Type 2/analysis , Receptor, Fibroblast Growth Factor, Type 2/genetics , Receptor, Fibroblast Growth Factor, Type 2/metabolism , Receptor, Fibroblast Growth Factor, Type 3/analysis , Receptor, Fibroblast Growth Factor, Type 3/genetics , Receptor, Fibroblast Growth Factor, Type 3/metabolism , Receptor, Fibroblast Growth Factor, Type 4/analysis , Receptor, Fibroblast Growth Factor, Type 4/genetics , Receptor, Fibroblast Growth Factor, Type 4/metabolism , Receptors, Fibroblast Growth Factor/analysis , Receptors, Fibroblast Growth Factor/metabolism , Transcription, Genetic
20.
J Cell Physiol ; 212(3): 572-8, 2007 Sep.
Article in English | MEDLINE | ID: mdl-17541959

ABSTRACT

Abnormal differentiation in epithelial stem cells or their immediate proliferative progeny, the transiently amplifying population (TAP), may explain malignant pathogenesis in the human prostate. These models are of particular importance as differing sensitivities to androgen among epithelial cell subpopulations during differentiation are recognised and may account for progression to androgen independent prostate cancer. Androgens are crucial in driving terminal differentiation and their indirect effects via growth factors from adjacent androgen responsive stroma are becoming better characterised. However, direct effects of androgen on immature cells in the context of a prostate stem cell model have not been investigated in detail and are studied in this work. In alpha2beta1hi stem cell enriched basal cells, androgen analogue R1881 directly promoted differentiation by the induction of differentiation-specific markers CK18, androgen receptor (AR), PSA and PAP. Furthermore, treatment with androgen down-regulated alpha2beta1 integrin expression, which is implicated in the maintenance of the immature basal cell phenotype. The alpha2beta1hi cells were previously demonstrated to lack AR expression and the direct effects of androgen were confirmed by inhibition using the anti-androgen bicalutamide. AR protein expression in alpha2beta1hi cells became detectable when its degradation was repressed by the proteosomal inhibitor MG132. Stratifying the alpha2beta1hi cells into stem (CD133(+)) and transient amplifying population (TAP) (CD133(-)) subpopulations, AR mRNA expression was found to be restricted to the CD133(-) (TAP) cells. The presence of a functional AR in the TAP, an androgen independent subpopulation for survival, may have particular clinical significance in hormone resistant prostate cancer, where both the selection of immature cells and functioning AR regulated pathways are involved.


Subject(s)
Cell Differentiation/drug effects , Cell Proliferation/drug effects , Epithelial Cells/drug effects , Metribolone/pharmacology , Neoplastic Stem Cells/drug effects , Prostatic Neoplasms/metabolism , Receptors, Androgen/drug effects , Testosterone Congeners/pharmacology , AC133 Antigen , Acid Phosphatase , Aged , Aged, 80 and over , Androgen Antagonists/pharmacology , Anilides/pharmacology , Antigens, CD/analysis , Cell Transformation, Neoplastic/drug effects , Cell Transformation, Neoplastic/metabolism , Cells, Cultured , Cysteine Proteinase Inhibitors/pharmacology , Dose-Response Relationship, Drug , Epithelial Cells/metabolism , Epithelial Cells/pathology , Fibroblast Growth Factor 7/metabolism , Glycoproteins/analysis , Humans , Integrin alpha2beta1/metabolism , Keratin-18/biosynthesis , Leupeptins/pharmacology , Male , Middle Aged , Neoplastic Stem Cells/metabolism , Neoplastic Stem Cells/pathology , Nitriles/pharmacology , Peptides/analysis , Phenotype , Prostate-Specific Antigen/biosynthesis , Prostatic Neoplasms/genetics , Prostatic Neoplasms/pathology , Proteasome Endopeptidase Complex/metabolism , Proteasome Inhibitors , Protein Tyrosine Phosphatases/biosynthesis , RNA, Messenger/biosynthesis , Receptors, Androgen/biosynthesis , Receptors, Androgen/genetics , Signal Transduction/drug effects , Tosyl Compounds/pharmacology
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