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W D Tilley

Publications and source records attributed to W D Tilley.

At least 19 recordsLinked to original sources
PubMed2001

Versican accumulation in human prostatic fibroblast cultures is enhanced by prostate cancer cell-derived transforming growth factor beta1.

We have previously demonstrated that peritumoral stromal matrix derived from prostate cancer patients who relapse after radical surgery contains elevated levels of versican. The purpose of this study was to determine whether prostate cancer cells control stromal cell secretion of versican. Serum-free conditioned medium from three prostate adenocarcinoma cell lines, LNCaP, PC3, and DU145, was added to cultures of fibroblasts established from prostatic tissue of patients with benign prostatic hyperplasia, and the medium was harvested at 24, 48, and 72 h. Immunoblotting with an antiversican core protein antibody revealed that prostatic fibroblast medium harvested at 72 h contained increased levels of versican after treatment with either LNCaP-, PC3- or DU145-conditioned medium (2.5-, 4.5-, and 5-fold, respectively) compared with control cultures. This increase in versican in the culture medium was not observed after coincubation with transforming growth factor beta1-neutralizing antisera. The results of this study suggest that prostate tumor cells induce host stromal cells to secrete increased versican levels via a paracrine mechanism mediated by transforming growth factor beta1.

Adenocarcinoma↗
PubMed2001

Contribution of the androgen receptor to prostate cancer predisposition and progression.

Although prostate cancer is heterogeneous in its etiology and progression, androgen signaling through the androgen receptor (AR) appears to be involved in all aspects of the disease, from initiation to development of treatment resistance. Lifetime exposure to a constitutively more active AR, encoded by AR alleles as defined by two translated polymorphic microsatellites (CAG and GGC), results in a significant increase in prostate cancer risk. The AR gene is amplified or a target for somatic gain-of-function mutations in metastatic prostate cancer. Gain-of-function AR gene mutations may result in inappropriate activation of the AR, thereby contributing to the failure of conventional androgen-ablation treatments. In cases where no genetically altered receptors are observed, altered signaling through the AR, achieved by cross-talk with other signaling pathways (e.g. kinase-mediated pathways) and/or inappropriate expression of coregulatory proteins, may contribute to disease progression. Thus, the AR-signaling axis contributes to many aspects of prostate cancer, including initiation, progression and resistance to current forms of therapy. This recognition represents a paradigm shift in our understanding of the molecular mechanisms involved in progression of prostate cancer, and provides insight into novel AR-targeted therapies which ultimately may be more effective than current forms of androgen ablation.

Adenocarcinoma↗
PubMed2001

Mutations at the boundary of the hinge and ligand binding domain of the androgen receptor confer increased transactivation function.

The androgen receptor (AR), a member of the steroid receptor superfamily of nuclear transcription factors, mediates androgen signaling in diverse target tissues. Here we report AR gene mutations identified in human prostate cancer and the autochthonous transgenic adenocarcinoma of the mouse prostate model that colocate to residues (668)QPIF(671) at the boundary of the hinge and ligand-binding domain, resulting in receptors that exhibit 2- to 4-fold increased activity compared with wild-type AR in response to dihydrotestosterone, estradiol, progesterone, adrenal androgens, and the AR antagonist, hydroxyflutamide, without an apparent effect on receptor levels, ligand binding kinetics, or DNA binding. The expression of these or similar variants could explain the emergence of hormone refractory disease in a subset of patients. Homology modeling indicates that amino acid residues (668)QPIF(671) form a ridge bordering a potential protein-protein interaction surface. The naturally occurring AR gene mutations reported in this study result in decreased hydrophobicity of this surface, suggesting that altered receptor-protein interaction mediates the precocious activity of the AR variants.

Adenocarcinoma↗
PubMed2001

Collocation of androgen receptor gene mutations in prostate cancer.

Consistent with both the development of the normal prostate gland and prostate tumorigenesis being dependent on testicular androgens, targeting the androgen-signaling axis (i.e., androgen ablation therapy) remains the predominant treatment regime for patients with metastatic prostate cancer. Although there is a very good initial response to androgen ablation, these treatments are essentially palliative. Recent evidence suggests that treatment failure may not result from a loss of androgen signaling but, rather, from the acquisition of genetic changes that lead to aberrant activation of the androgen-signaling axis. A consistent finding is that androgen receptor (AR) gene mutations, present in metastatic prostate cancer and in human prostate cancer cell lines as well as in xenograft and other animal models, result in decreased specificity of ligand-binding and inappropriate receptor activation by estrogens, progestins, adrenal androgens, glucocorticoids and/or AR antagonists. Because a significant proportion of missense mutations in the AR gene reported in prostate cancer collocate to the signature sequence and AF-2, two discrete regions of the ligand-binding domain critical for androgen signaling, we recently proposed that collocation of mutations identified in prostate cancer would identify additional regions of the AR important in receptor function. This approach led to the identification of a four-amino acid region at the boundary of the hinge and ligand-binding domains of the receptor that forms half of a potential protein-protein binding site. AR gene mutations have also been identified that collocate to areas in the DNA-binding domain, to the NH(2)-terminal transactivation domain, and to the hinge region in prostate tumors. In nearly every case, missense mutations in the AR gene identified in prostate cancer that collocate to discrete regions of the receptor contribute to altered androgen signaling and provide a potential mechanism to explain the reemergence of tumor growth during the course of hormone ablation therapies.

Androgens↗
PubMed2000

Hormone status selects for spontaneous somatic androgen receptor variants that demonstrate specific ligand and cofactor dependent activities in autochthonous prostate cancer.

We have used the autochthonous transgenic adenocarcinoma of mouse prostate (TRAMP) model to investigate the relationship between somatic mutation in the androgen receptor (AR) and the emergence of androgen-independent prostate cancer. Here we report the identification, isolation, and characterization of distinct classes of AR variants from spontaneous prostate tumors in the TRAMP model. Using cDNA cloning, single stranded conformation polymorphism and sequencing strategies, 15 unique somatic mutations in the AR were identified in prostate tumors obtained from eight TRAMP mice between 24 and 29 weeks of age. At least one mutation was isolated from each mouse. All mutations were single base substitutions, 10 were missense and 5 were silent. Nine mutations in the AR were identified in tumors of four mice that were castrated at 12 weeks of age. Interestingly, the majority of mutations (seven out of nine, 78%) identified in the androgen-independent tumors colocalized in the AR transactivation domain. The remaining mutations colocalized in the AR ligand binding domain. In general, the AR variants demonstrated promoter-, cell-, and cofactor-specific activities in response to various hormones. All AR variants isolated in this study maintained strong sensitivity for androgens, and four AR variants isolated from castrated mice demonstrated increased activities in the absence of ligand. The K638M and F677S variants demonstrated increased activities in response to androgen, and K638M also demonstrated increased response to estradiol. In the presence of AR coactivator ARA70 the E231G variant demonstrated increased activity in response to both androgen and estradiol. However, in the presence of AR coactivator ARA160 the E231G variant was selectively responsive to androgen. Collectively these analyses not only indicate that somatic mutations in the AR gene occur spontaneously in TRAMP tumors but also how changes in the hormonal environment may drive the selection of spontaneous somatic mutations that provide a growth advantage.

Animals↗
PubMed2000

Breast cancer susceptibility gene 1 (BRCAI) is a coactivator of the androgen receptor.

In the present study, the role of BRCA1 in ligand-dependent androgen receptor (AR) signaling was assessed. In transfected prostate and breast cancer cell lines, BRCA1 enhanced AR-dependent transactivation of a probasin-derived reporter gene. The effects of BRCA1 were mediated through the NH2-terminal activation function (AF-1) of the receptor. Cotransfection of p160 coactivators markedly potentiated BRCA1-mediated enhancement of AR signaling. In addition, BRCA1 was shown to interact physically with both the AR and the p160 coactivator, glucocorticoid receptor interacting protein 1. These findings suggest that BRCA1 may directly modulate AR signaling and, therefore, may have implications regarding the proliferation of normal and malignant androgen-regulated tissues.

BRCA1 Protein↗
PubMed1999

Androgen receptor agonist activity of the synthetic progestin, medroxyprogesterone acetate, in human breast cancer cells.

Medroxyprogesterone acetate (MPA), which is frequently used as second line hormonal therapy for the treatment of metastatic breast cancer, binds with high affinity to the progesterone receptor (PR). However, the androgenic side-effects of MPA suggest that it may also activate androgen receptor (AR) regulated pathways. Treatment of the human breast cancer cell lines MDA-MB-453, ZR-75-1 and T47-D with high dose (100 nM) MPA resulted in 26-30% inhibition of cell growth, which was partially reversed by co-treatment with a 10-fold excess of the synthetic antiandrogen, anandron. Scatchard analysis demonstrated specific, high affinity (non-PR) binding of [3H]MPA to cytosols prepared from the PR-/AR+ MDA-MB-453 and PR+/AR+ ZR-75-1, but not the PR-/AR- BT-20 breast cancer cell lines. Competition of [3H]MPA binding to MDA-MB-453 cytosols by equimolar concentrations of androgens (5alpha-dihydrotestosterone (DHT), R1881) and the antiandrogen, anandron was consistent with binding of MPA to the AR. In ZR-75-1 cell cytosol fractions, DHT, R1881 and anandron only partially competed out [3H]MPA binding, suggesting that androgens displace [3H]MPA binding to AR but not to PR. Induction by MPA of AR transactivation was demonstrated in MDA-MB-453 and ZR-75-1 cells, and in the CV-1 cell line transfected with a full-length AR. In these cell lines the increased activity of the androgen responsive reporter gene (MMTV-CAT) by 1 nM MPA was fully (MDA-MB-453, CV-1) or partially (ZR-75-1) inhibited by co-culture with 1 microM anandron. These findings indicate that MPA is an AR agonist and suggest that the in vivo effects of MPA in breast cancer patients may in part be mediated by the AR.

Androgen Antagonists↗
PubMed1999

Elevated levels of peritumoral chondroitin sulfate are predictive of poor prognosis in patients treated by radical prostatectomy for early-stage prostate cancer.

The disease course of localized prostate cancer is highly variable, and patients potentially curable by aggressive management are not readily identified by current clinical practice. Chondroitin sulfate (CS) glycosaminoglycan is a candidate biomarker as elevated levels of CS in peritumoral stroma of prostate cancer have been associated with prostate-specific antigen (PSA) failure. Immunoreactive CS was measured using image analysis of archived radical prostatectomy tissues, obtained from 157 men with a median of 47 months (range, 16-111 months) clinical follow-up. CS level, Gleason score, and preoperative serum PSA levels were independent predictors of PSA failure by Cox's multivariate analysis. Patients with low CS levels had significantly fewer PSA failures after radical prostatectomy than patients with high levels of CS (Kaplan-Meier plot; 32% PSA failures at 5 years for CS mean integrated absorbance cut point < 7.0 versus 50% for CS > or = 7.0, P = 0.0001). In the subgroup of patients with preoperative serum PSA levels < 10 ng/ml, CS was particularly useful in discriminating retrospectively those patients most suited for surgery (Kaplan-Meier plot; 14% PSA failures at 5 years for CS mean integrated absorbance cut point < 7.0 versus 47% for CS > or = 7.0, P = 0.0001). We conclude that measurements of CS level can assist in predicting patient outcome after surgery. Additionally, our data suggest that the combination of CS and PSA measurements may improve outcome prediction for patients with intermediate Gleason scores.

Adenocarcinoma↗
PubMed1999

Molecular detection of prostate cells in ejaculate and urethral washings in men with suspected prostate cancer.

PURPOSE: To determine whether prostatic cells were normally present in ejaculate and if the sensitivity and specificity of the detection of malignant prostate cells in ejaculate and urethral washings from men with suspected prostate cancer could be improved using the more sensitive molecular technique of reverse transcriptase-polymerase chain reaction (RT-PCR). MATERIALS AND METHODS: RT-PCR for prostate-specific antigen (PSA), prostate-specific membrane antigen (PSM) and Apoliprotein D (3 putative prostate-specific and/or cancer-specific markers) was performed on RNA extracts of ejaculate (80) and urethral washings (52) from 77 men with suspected prostate cancer and 12 young controls (<30 years of age) and urines from 5 men who had radical prostatectomies and 10 women. RESULTS: PSA, PSM and Apolipoprotein D expression was detected in ejaculates and urethral washings from both patient and control groups. No differences were observed in the results obtained for 58 men with suspected or 19 men with confirmed prostate cancer or the 18 vasectomized men within the patient group. Urines from the 5 men who had radical prostatectomies and 10 women were all negative for PSA, but PSM was detected in 2 female urines and in 3 radical prostatectomy samples. As few as 10 LNCaP prostate tumor cells could be detected by PSA RT-PCR when added to female urine. CONCLUSION: We have established a sensitive method of detecting prostatic cells in ejaculate and urethral washings and shown that PSA RT-PCR is a reliable indicator of prostate cells in these samples. However, RT-PCR for PSA, PSM and Apoliprotein D were not useful for discriminating malignant from non-malignant prostate cells.

Adult↗
PubMed1998

Immunolocalization of apolipoprotein D, androgen receptor and prostate specific antigen in early stage prostate cancers.

PURPOSE: To determine the cellular distribution and levels of immunohistochemical staining for apolipoprotein D (Apo-D), prostate specific antigen (PSA) and androgen receptor (AR) in early stage prostate cancers. MATERIALS AND METHODS: Cellular distribution of Apo-D, PSA and AR in 30 stage A/B prostate cancers and in non-malignant glandular tissue contained in the same sections was detected immunohistochemically, and staining was evaluated by computerized video image analysis. RESULTS: Staining for Apo-D (percentage positive cellular area) was significantly increased in tumor cells of early stage prostate cancers compared with non-malignant glandular tissue. PSA and AR were present at high levels in both early stage prostate tumors and non-malignant prostate. CONCLUSIONS: Malignant transformation in the prostate is associated with increased cellular levels of Apo-D.

Apolipoproteins↗
PubMed1998

Role of the androgen receptor in human breast cancer.

Although the androgen receptor (AR)3 is often co-expressed with the estrogen receptor (ER) and progesterone receptor (PR) in human breast tumors, its role in breast cancer is poorly understood. Specific growth stimulatory and inhibitory actions of androgens have been described in human breast cancer cell lines. The mechanisms by which androgens exert these contrasting growth effects are unknown. A commonly utilized second line therapy for the treatment of advanced breast cancer is high dose medroxyprogesterone acetate (MPA). Although MPA, a synthetic progestin, was thought to act exclusively through the PR, the androgenic side-effects observed in women taking MPA suggest that its action may also be mediated in part by the AR. In support of this hypothesis, the level of AR measured by radioligand binding in primary breast tumors was correlated with the duration of response to MPA treatment following failure of tamoxifen therapy. Recent data suggest that the presence of structurally altered AR in breast cancers may account for unresponsiveness to MPA in some of these cases. Further studies are warranted to determine the role of AR mediated pathways in regulating breast tumor growth. In particular, identification of androgen-regulated genes may lead to new possibilities for the hormonal treatment of breast cancer.

Breast Neoplasms↗
PubMed1998

Prostate-specific antigen and gross cystic disease fluid protein-15 are co-expressed in androgen receptor-positive breast tumours.

Androgens regulate breast cancer cell proliferation via androgen receptor (AR)-mediated mechanisms. To investigate further the androgen-responsiveness of human breast tumours, we examined the immunohistochemical expression of the AR and two androgen-regulated proteins, prostate-specific antigen (PSA) and gross cystic disease fluid protein-15 (GCDFP-15), in 72 primary breast tumours. AR immunoreactivity was present in the nuclei of breast tumour cells and was correlated with oestrogen receptor (ER; P < 0.05) and progesterone receptor (PR; P < 0.01) status. PSA and GCDFP-15 immunoreactivity was present in the cytoplasm of tumour cells but not the adjacent stromal cells. AR-positive cells were present in 85% (61/72) of breast tumours, and 98% (43/44) of PSA-positive and 92% (44/48) of GCDFP-15-positive tumours were also positive for AR. Positive immunoreactivity for both PSA and GCDFP-15 in breast tumours was highly dependent on AR status (odds ratios of 24.0 and 4.5 respectively), but unrelated to age, ER and PR status and axillary lymph node involvement. PSA immunoreactivity was more frequently observed in moderate and well-differentiated tumours and was significantly (P < 0.001) associated with GCDFP-15 immunoreactivity. In conclusion, PSA and GCDFP-15 immunoreactivity was dependent on the presence of AR, but not ER or PR in primary breast tumours.

Adult↗
PubMed1997

Vascular endothelial growth factor (VEGF) expression in prostate cancer and benign prostatic hyperplasia.

PURPOSE: Vascular endothelial growth factor (VEGF) is a potent inducer of endothelial cell growth and is expressed at elevated levels in several tumor types. In this study immunohistochemical localization and distribution of isoforms of VEGF were examined in malignant and non-malignant human prostatic tissues. MATERIALS AND METHODS: Immunohistochemical localization of VEGF was performed on thirty well, moderately and poorly differentiated stage D2 prostate cancer specimens and twenty benign prostatic hyperplasia (BPH) specimens. VEGF mRNA was determined by polymerase chain reaction and VEGF protein isoforms were detected by Western blotting of prostate cancer and BPH specimens. RESULTS: Cytoplasmic immunoreactivity for VEGF was detected in tumor cells and peritumoral stromal cells of prostate cancer specimens and in non-malignant glandular epithelial cells and interglandular stromal cells in BPH specimens. Staining was focal with areas of strongly to weakly stained cells adjacent to negatively staining areas. mRNA's for VEGF121, VEGF165 and VEGF189 were present in all benign and malignant prostate specimens. VEGF protein isoforms of molecular sizes corresponding to VEGF165 and VEGF189 were detected in cytosolic extracts of prostate cancers and BPH specimens by Western blotting. In addition, two novel higher molecular weight immunoreactive bands were detected in the prostate specimens. CONCLUSIONS: Widespread distribution of VEGF in prostate cancers and BPH specimens suggest that the VEGF165, VEGF189 isoforms and novel 90 and 110 kD forms detected may contribute to the establishment or progression of these conditions.

Blotting, Western↗
PubMed1997

Elevated stromal chondroitin sulfate glycosaminoglycan predicts progression in early-stage prostate cancer.

Curative therapies for clinically localized prostate cancer have significant morbidity, and those patients who might be cured by aggressive management are not easily identified using current clinical information. Better biomarkers of tumor behavior need to be identified to improve clinical outcome. Chondroitin sulfate (CS), a glycosaminoglycan, may be a potentially useful biomarker as it is known to influence cell growth and differentiation and might influence malignant progression. In this study, CS was immuno-localized to the periacinar and peritumoral fibromuscular stromal tissue of nonmalignant and malignant prostates. The CS concentration was increased in the prostate tissue of men with early-stage prostate cancer compared with tissue from men without cancer (P < 0.0001). Using Cox's univariate analysis, CS concentration, tumor grade, preoperative serum prostate-specific antigen (PSA), extracapsular extension of disease, positive surgical margins, and patient age were associated with an increased risk of PSA failure. The CS concentration was compared with the other features in two-variable regression analyses. CS and preoperative serum PSA concentrations were independent predictors of PSA failure. CS was a stronger prognostic feature than tumor grade and could predict outcome for patients with moderately differentiated tumors. Patients with a low CS concentration had significantly better progression-free survival following radical prostatectomy than patients with high levels of CS (Kaplan-Meier plot, 91% versus 49% PSA progression free at 5 years, respectively, P = 0.0038). Only postoperative pathological indices (extracapsular extension, surgical margins) were stronger predictors than CS. We conclude that measurement of prostatic CS concentrations at diagnosis may allow stratification of patients with early-stage prostate cancer for adjunctive or alternate therapies.

Adolescent↗
PubMed1996

Expression of the androgen receptor and an androgen-responsive protein, apolipoprotein D, in human breast cancer.

Little is known regarding the activity and function of the androgen receptor (AR) in human breast cancer. In the present study AR was evaluated in untreated primary breast cancers using antisera to the amino- and carboxy-termini of the receptor and quantitated using colour video image analysis. A strong correlation between tissue concentration and percentage AR-positive cells was observed for each antiserum. However, comparison of percentage positive cells using the amino- and carboxy-terminal AR antisera in individual breast cancer specimens revealed a subset of tumours with discordantly increased staining for the carboxy terminus. These findings suggest the presence of amino-terminal-truncated AR in a proportion of breast cancer cells or presence of AR mutations or associated protein alterations that affect binding of the amino-terminal AR antiserum. Immunohistochemical expression of the androgen-regulated glycoprotein, apolipoprotein D (apo-D), was also evaluated in the breast cancer specimens. Focal positivity of apo-D staining, which did not always co-localise with AR-positive cells, was observed within breast tumours. Furthermore, no correlation was evident between percentage positive cells stained for AR and apo-D in breast cancer specimens. These findings indicate that, although apo-D expression is androgen regulated in human breast cancer cell lines in vitro, its expression in primary breast cancers may be regulated by other factors. The expression of AR in primary breast cancers also suggests that the receptor may be involved in tumour responsiveness or in abnormal responses to endocrine therapies.

Adult↗
PubMed1996

Partial androgen insensitivity caused by an androgen receptor mutation at amino acid 907 (Gly-->Arg) that results in decreased ligand binding affinity and reduced androgen receptor messenger ribonucleic acid levels.

Androgen insensitivity is an X-linked disorder of sexual differentiation resulting from mutations in the androgen receptor (AR) gene. In this paper, we report the clinical phenotype and molecular analysis of two siblings with severe partial androgen insensitivity due to a novel mutation in the ligand-binding domain of the AR gene. Binding studies using cultured genital skin fibroblasts demonstrated reduced AR affinity and binding capacity. Nucleotide sequence analysis of the AR gene of both siblings revealed a point mutation causing a glycine to arginine amino acid substitution at position 907 within a conserved region of the ligand-binding domain. A silent guanine to adenine substitution was also identified in the protein-coding region of exon 1. Using an expression vector in which the identified mutation was recreated by site-directed mutagenesis, the mutant receptor was found to have a reduced binding affinity (Kd = 3.06 nmol/L) for mibolerone compared with that of normal AR (Kd = 1.71 nmol/L) when expressed in COS-7 cells. In cotransfection experiments using CV-1 cells and a mouse mammary tumor virus-chloramphenicol acetyltransferase reporter system, the concentration of dihydrotestosterone required to induce half-maximal chloramphenicol acetyltransferase gene expression was 50-fold higher in cells transfected with the mutant AR complementary DNA than in cells transfected with normal AR complementary DNA. AR messenger ribonucleic acid levels in genital skin fibroblasts determined by both competitive PCR amplification and ribonuclease protection assay were decreased compared with normal values. Our studies demonstrate the importance of this region of the AR gene in normal AR function and AR gene expression.

Adult↗