Biomedical subjects

P S Rennie

Publications and source records attributed to P S Rennie.

At least 19 recordsLinked to original sources

PubMed2006

Targeting and killing of prostate cancer cells using lentiviral constructs containing a sequence recognized by translation factor eIF4E and a prostate-specific promoter.

To develop a gene therapy that would selectively kill prostate cancer cells while sparing normal cells, we have constructed lentiviral vectors that contain a therapeutic gene with a short DNA sequence in the 5'-untranslated region (UTR) that is recognized by the translation initiation factor, eIF4E, which is often overexpressed in malignant cells. Infection of cancer (LNCaP, PC-3M, DU145, and MCF-7 cells) and noncancer cell lines (BPH-1, 267-B1, Plat-E, and Huvec-c cells) with lentivirus having a CMV-promoter and EGFP reporter resulted in high levels of EGFP expression in all cells, whereas, inclusion of the eIF4E UTR recognition sequence restricted high expression to cancer cells and Plat-E cells, which also express substantial levels of eIF4E. Infection of the cells with lentiviral vectors having this UTR in front of the HSV thymidine kinase suicide gene resulted in differential sensitivity to the killing effects of ganciclovir, with at least 100-fold more drug required to kill noncancer cells than cancer cells. Furthermore, in experiments where the CMV promoter was replaced by the prostate-specific ARR(2)PB promoter, the killing effects of ganciclovir were restricted to prostate cancer cells and not seen in nonprostate cancer cells. Our results indicate that combined translational regulation, by incorporation of an eIF4E-UTR recognition sequence into a therapeutic gene, together with transcriptional regulation with a prostate-specific promoter, may provide a means to selectively destroy prostate cancer cells while sparing normal prostate cells.

Cell Death↗

PubMed2000

Acquisition of chemoresistant phenotype by overexpression of the antiapoptotic gene testosterone-repressed prostate message-2 in prostate cancer xenograft models.

Testosterone-repressed prostate message-2 (TRPM-2) expression is highly up-regulated in normal and malignant prostate cells after androgen withdrawal. Although recent studies have suggested a protective role of TRPM-2 expression against apoptosis in several experimental models, the functional role of TRPM-2 in chemotherapy-induced apoptosis remains undefined. Here, we demonstrated that overexpression of TRPM-2 in human androgen-dependent LNCaP prostate cancer cells by stable transfection rendered them highly resistant to paclitaxel treatment than control LNCaP cells, with a 20-fold higher IC50 through the inhibition of apoptotic cell death. In mice bearing TRPM-2-overexpressing LNCaP tumors, tumor volume and serum prostate-specific antigen increased two to three times faster after castration and paclitaxel treatment compared with mice bearing control tumors. We then tested the efficacy of combined treatment with antisense TRPM-2 oligodeoxynucleotide (ODN) and paclitaxel in the mouse androgen-dependent Shionogi tumor model. Antisense TRPM-2 ODN treatment significantly enhanced paclitaxel chemosensitivity of Shionogi tumor cells in a dose-dependent manner, reducing the IC50 by 75%. Combined treatment of Shionogi cells with 500 nM antisense TRPM-2 ODN and 10 nM paclitaxel-induced apoptosis, either agent alone did not. Adjuvant administration of antisense TRPM-2 ODN and polymeric micellar paclitaxel after castration resulted in reduced TRPM-2 levels in vivo and a significant delay of emergence of androgen-independent recurrent Shionogi tumors compared with administration of either agent alone. Furthermore, combined treatment of mice bearing androgen-independent recurrent Shionogi tumors with antisense TRPM-2 ODN and micellar paclitaxel inhibited tumor growth compared with treatment with either agent alone. Collectively, these findings demonstrate that TRPM-2 overexpression helps confer a chemoresistant phenotype through inhibition of apoptosis, and that antisense TRPM-2 ODN may be useful in enhancing the effects of cytotoxic chemotherapy in hormone-refractory prostate cancer.

Animals↗

PubMed2000

Testosterone-repressed prostate message-2 is an antiapoptotic gene involved in progression to androgen independence in prostate cancer.

Although initially reported as an androgen-repressed gene in the rat prostate, the functional role of testosterone-repressed prostate message-2 (TRPM-2) in apoptosis remains undefined. Inhibition of castration-induced apoptosis by calcium channel blocker treatment in androgen-dependent Shionogi tumors resulted in the prevention of TRPM-2 gene up-regulation, suggesting that TRPM-2 is not directly androgen-repressed, but is regulated by apoptotic stimuli. The overexpression of the TRPM-2 gene in human androgen-dependent LNCaP prostate cancer cells by stable transfection rendered them highly resistant to androgen ablation in vivo. We then tested the efficacy of antisense TRPM-2 oligodeoxynucleotide (ODN) therapy in the Shionogi tumor model and demonstrated that the systemic administration of antisense TRPM-2 ODNs in mice bearing Shionogi tumors after castration resulted in a more rapid onset of apoptosis and time to complete regression, as well as a significant delay of emergence of androgen-independent recurrent tumors compared to control ODN treatment. Collectively, these findings illustrate that TRPM-2 is an antiapoptotic rather than an androgen-repressed gene that confers resistance to androgen ablation and thereby helps accelerate the progression to androgen independence.

Animals↗

PubMed2000

Characterization of an androgen-specific response region within the 5' flanking region of the murine epididymal retinoic acid binding protein gene.

The epididymis provides the optimal milieu for sperm maturation and storage. Epididymal secretory proteins are believed to be involved in that process. Androgens are the major endocrine and paracrine regulatory signals that regulate gene expression in the epididymis. We have previously identified an androgen-dependent retinoic acid-binding protein (mE-RABP) that is secreted into the luminal fluid from the mouse mid/distal caput epididymidis. The mE-RABP protein belongs to the lipocalin superfamily and may be involved in the trafficking of retinoic acid within the epididymis. We have recently demonstrated that 5 kilobases of the 5' flanking region of the mE-RABP gene contained all the information for the hormonal regulation and the tissue-, region-, and cell-specific expression of the mE-RABP gene. In this study, we have identified a complex androgen-specific response region (ARR) within the first 600 base pairs of the mE-RABP gene promoter. Androgen (DHT) but not glucocorticoid (DEX) activates the ARR in HeLa and PC-3 cells. Two androgen receptor binding sites have been located at positions -445/-459 and -102/-88 and were named ARBS-1 and ARBS-0, respectively. Point mutations of ARBS-0 resulted in a slight decrease of the androgen response. However, mutations of ARBS-1 led to a total loss of the androgen responsiveness, suggesting that it was a major cis-acting element. When ARBS-1 is isolated from its promoter context, it serves as a weak androgen-responsive element that was activated by both androgens and glucocorticoids. Also, the -543/-88 DNA promoter fragment behaved as a poor androgen-responsive region, suggesting that regulatory elements located within the proximal mE-RABP promoter were required for a full androgen response. In conclusion, the mE-RABP ARR is a good model for the study of molecular mechanisms that lead to an androgen-specific responsiveness in vivo.

Androgens↗

PubMed2000

Overexpression of insulin-like growth factor binding protein-5 helps accelerate progression to androgen-independence in the human prostate LNCaP tumor model through activation of phosphatidylinositol 3'-kinase pathway.

Although insulin-like growth factor (IGF) binding protein-5 (IGFBP-5) is highly up-regulated in normal and malignant prostate tissues after androgen withdrawal, its functional role in castration-induced apoptosis and androgen-independent progression remains undefined. To analyze the functional significance of IGFBP-5 overexpression in IGF-I-mediated mitogenesis and progression to androgen-independence, IGFBP-5-overexpressing human androgen-dependent LNCaP prostate cancer cells were generated by stable transfection. The growth rates of IGFBP-5-transfected LNCaP cells were significantly faster, compared with either the parental or vector-only transfected LNCaP cells in both the presence and absence ofdihydrotestosterone. IGFBP-5-induced increases in LNCaP cell proliferation occurs through both IGF-I-dependent and -independent pathways, with corresponding increases in the cyclin D1 messenger RNA expression and the fraction of cells in S + G2/M phases of the cell cycle. Changes in Akt/protein kinase B, a downstream component of phosphatidylinositol 3'-kinase (PI3K) pathway, in the LNCaP sublines also paralleled changes in their growth rates. Although treatment with a PI3K inhibitor induced apoptosis in both control and IGFBP-5-overexpressing LNCaP cells, this PI3K inhibitor-induced apoptosis was prevented by exogenous IGF-I treatment only in IGFBP-5 transfectants, suggesting that IGFBP-5 overexpression can potentiate the antiapoptotic effects of IGF-I. Furthermore, tumor growth and serum prostate-specific antigen levels increased several fold faster in mice bearing IGFBP-5-transfected LNCaP tumors after castration, despite having similar tumor incidence and tumor growth rates with controls when grown in intact mice before castration. Collectively, these data suggest that IGFBP-5 overexpression in prostate cancer cells after castration is an adaptive cell survival mechanism that helps potentiate the antiapoptotic and mitogenic effects of IGF-I, thereby accelerating progression to androgen independence through activation of the PI3K-Akt/ protein kinase B signaling pathway.

Androgens↗

PubMed1999

Determinants of DNA sequence specificity of the androgen, progesterone, and glucocorticoid receptors: evidence for differential steroid receptor response elements.

While androgen, progesterone, and glucocorticoid receptors perform distinct physiological functions by regulating unique sets of genes, in vitro they can transactivate a common high-affinity DNA-binding target. Naturally occurring steroid response elements display nucleotide divergence that lowers binding affinity in comparison to the optimal binding element, but enhances receptor-type specificity. We investigated the role of nucleotide deviations within the DNA-binding site for contribution to steroid receptor specificity. We hypothesized that receptor specificity drives the evolution of binding site sequence, rather than strictly receptor-binding affinity. Receptor-selective targets can evolve by some nucleotides selected on the basis of additional bond energy, and others may be selected by differential tolerance to discourage binding from inappropriate receptors. To identify receptor-specific binding sites, we mimicked these dual selection pressures in a receptor-competitive environment in which DNA binding sites for the androgen or progesterone receptors were selected in the presence of the glucocorticoid receptor. These analyses also demonstrated that steroid receptors strongly select nucleotides in the spacer and flanking regions of the half-site and do so in an asymmetric fashion, indicating that steroid receptors interact with DNA in an allosteric manner that affects the transcriptional activation potential.

Amino Acid Sequence↗

PubMed1999

Selective activation of the probasin androgen-responsive region by steroid hormones.

Glucocorticoid and androgen receptors have been shown to function through the same palindromic glucocorticoid response element (GRE) and yet have differential effects on gene transcription. In this study, we examined the functional and structural relationship of the androgen and glucocorticoid receptors with the androgen responsive region (ARR) of the probasin (PB) gene containing two androgen receptor binding sites, ARBS-1 and ARBS-2. Transfection studies indicated that one copy of each cis-acting DNA element was essential for maximal androgen-induced chloramphenicol acetyltransferase (CAT) activity and that androgen selectivity was maintained when multiple copies of the minimal wild type (wt) androgen responsive region containing both ARBS-1 and ARBS-2 (-244 to -96) were subcloned in front of the thymidine kinase promoter. Furthermore, replacing the androgen response region with 1, 2 or 3 copies of either ARBS-1 or ARBS-2 restored less than 4% of the biological activity seen with the wt PB ARR. Multiple copies of either ARBS-1 or ARBS-2 did not result in glucocorticoid-induced CAT gene activity. By comparison, 1 or 2 copies of the tyrosine aminotransferase (TAT) GRE, as well as the mouse mammary tumour virus GRE, were strong inducers of CAT activity in response to both androgen and glucocorticoid treatment. In addition, band shift assays demonstrated that although the synthetic glucocorticoid receptor, GR-DNA binding domain (GR-DBD), and the synthetic androgen receptor, AR2, could interact with the TAT GRE (dissociation constants Kd of 63.9 and 14.1 respectively), only AR2 but not GR-DBD binding could be detected on ARBS-1 and ARBS-2. Our findings provide further evidence that androgen-induced regulation of gene transcription can occur through androgen-specific DNA binding sites that are distinct from the common GRE.

Androgen-Binding Protein↗

PubMed1998

Differential transactivation by the androgen receptor in prostate cancer cells.

BACKGROUND: The purpose of this study was to determine the contribution of different transactivating regions of the androgen receptor (AR) to the induction of androgen-regulated promoters in poorly (PC3 cells) and well-differentiated (LNCaP cells) prostate cancer cell lines. METHODS: PC3 and LNCaP cells were co-transfected with plasmids expressing full-length AR or deletion mutants together with luciferase reporters linked to the probasin (PB) and PSA promoters; as well as to ARR3tk, a PB-derived recombinant promoter. RESULTS: Androgen induction of the ARR3tk promoter in the presence of AR was 8- to 10-fold higher than that seen with the PB promoter. Activation of ARR3tk was greatest with an androgen-independent construct in which the first 231 amino acids and the ligand binding domain had been removed, indicating that this promoter is more responsive to activating functions in the N-terminal domain than in the ligand binding domain. By comparison, induction of the PB promoter was greatest with the full-length AR, which suggests that the ligand binding domain also makes a major contribution to the activation of this promoter. In similar analyses with the PSA promoter, AR regions required for promoter induction was dependent on the host cell type. In PC3 cells, the predominant AR transactivation function was androgen-independent and resided in the N-terminal domain, whereas in LNCaP cells, the highest level of induction was androgen dependent and also required participation of the ligand binding domain. CONCLUSIONS: Our results indicate that the relative utilization of transactivating functions in N-terminal and ligand binding domains of the AR is promoter and cell specific.

Androgen-Binding Protein↗

PubMed1997

Androgenic induction of prostate-specific antigen gene is repressed by protein-protein interaction between the androgen receptor and AP-1/c-Jun in the human prostate cancer cell line LNCaP.

In exploring the possible mechanisms of androgen independence of prostate-specific antigen (PSA) gene expression, we investigated the effect of elevating AP-1 by both 12-O-tetradecanoylphorbol 13-acetate (TPA) treatment and transfection of the c-Jun expression vector in LNCaP cells. Transcription of PSA is initiated when ligand-activated androgen receptor (AR) binds to a region in the PSA promoter that contains an androgen-responsive element (ARE). It was found that TPA inhibited androgen-induced PSA gene expression by a mechanism that did not alter nuclear levels of AR protein. Overexpression of AP-1 (jun and fos proteins) also inhibited androgen-induced PSA promoter activity. These observations were apparently related to the disruption of AR.ARE complexes as demonstrated by the results of electrophoretic mobility shift assays. Specifically, c-Jun inhibited the formation of AR.ARE complexes and conversely that AR-glutathione S-transferase proteins inhibited the formation of c-Jun.TPA-responsive element (TRE) complexes. Consistent with the inhibitory effect of both proteins, anti-c-Jun antibody blocked the inhibition of AR.ARE complex formation by c-Jun. A similar, but less marked, effect was obtained when anti-AR antibody was used to prevent AR inhibition of c-Jun.TRE complex formation. These findings together with results obtained from co-immunoprecipitation experiments strongly suggest that mutual repression of DNA binding activity is due to direct interaction between the two proteins and that the degree of repression may be determined by the ratio of AR to c-Jun. The mechanism of repression studied in mutant analysis experiments yielded evidence of an interaction between the DNA- and ligand-binding domains of AR and the leucine zipper region of c-Jun. Thus, the AR is similar to other nuclear receptors in its ability to interact with AP-1. This association provides a link between AP-1 and AR signal transduction pathways and may play a role in the regulation of the androgen-responsive PSA gene.

Androgens↗

PubMed1997

A metastatic and androgen-sensitive human prostate cancer model using intraprostatic inoculation of LNCaP cells in SCID mice.

Several metastasizing murine and human animal models for prostate cancer are available. However, these models are androgen-independent and lack differentiated features such as androgen receptor and androgen-regulated gene expression like prostate-specific antigen (PSA). The objective of this study was to develop a metastasizing prostate cancer model with differentiated features using the human LNCaP cell line. Athymic and SCID mice were injected either s.c. or intraprostatically with 1 x 10(6) LNCaP cells. Changes in serum and tumor PSA mRNA levels were determined before and after castration to assess time to androgen-independent progression. Local tumor and metastatic growth was assessed at sacrifice after 12 weeks. Reverse transcription-PCR (RT-PCR) was used to detect circulating LNCaP cells. LNCaP tumor incidence after s.c. injection was 100% (65 of 65) in SCID mice and 80% in athymic mice. No lymph node or distant metastases were observed with s.c. tumors, and RT-PCR for PSA transcripts was negative. Primary tumor incidence after intraprostatic injection was 89% (39 of 44) in SCID mice and 60% in athymic mice. In 10 SCID mice with primary tumors followed for 12 weeks, retroperitoneal or mediastinal lymph node metastases were found in 100%, and microscopic pulmonary metastases were identified in 40%. RT-PCR for PSA transcripts was positive in 3 of 10 mice tested. Serum PSA levels in mice with s.c. and intraprostatic tumors decreased by 65% to nadir levels at 7 and 4 days after castration, respectively. Serum PSA and LNCaP tumor PSA mRNA levels increased to precastration levels earlier in SCID mice with intraprostatic tumors compared to those with s.c. tumors. Intraprostatic injection of LNCaP cells in SCID mice provides a useful animal model to investigate mechanisms of metastasis and to evaluate therapies targeted toward inhibiting the metastatic cascade.

Animals↗

PubMed1996

Intermittent androgen suppression delays progression to androgen-independent regulation of prostate-specific antigen gene in the LNCaP prostate tumour model.

In most patients with prostate cancer, continuous androgen suppression (CAS) therapy causes tumour regression and an accompanying decrease in serum prostate specific antigen (PSA). However, with tumour progression, regulation of both tumour growth and PSA gene expression becomes androgen-independent. Because androgen resistance develops, in part, from adaptive cell survival mechanisms activated by androgen withdrawal, we hypothesize that intermittent re-exposure to androgens may prolong time to androgen-independent progression. The objective of this study was to determine whether intermittent androgen suppression (IAS) could delay the onset of androgen-independent PSA gene regulation in LNCaP prostate tumour model when compared to CAS. Five or six cycles of IAS were possible before progression developed. IAS prolonged time to androgen-independent PSA gene regulation from an average of 26 days in CAS to 77 days in IAS. Serum PSA increased above pre-castrate levels in all mice treated with CAS by 28 days post-castration, but remained below pre-castrate levels in 75% of IAS-treated mice by 60 days post-castration. By 15 weeks post-castration, serum PSA levels increased 7-fold above pre-castrate levels in CAS-treated mice compared to 1.9-fold increase in IAS-treated mice. PSA mRNA expression levels highly correlated with serum PSA levels in both groups. Maintenance of androgen dependency through IAS may be due to androgen-induced differentiation and/or down-regulation of androgen-suppressed gene expression.

Androgens↗

PubMed1996

Induction of cell-free, in vitro transcription by recombinant androgen receptor peptides.

An in vitro, cell-free transcription system, based on prostate-derived transcriptional machinery and very powerful androgen response elements (AREs), has been developed. Multiple (p(ARR3)LovTATA) AREs from the androgen-regulated probasin gene were linked to G-free cassettes and used in nuclear extracts prepared from prostate carcinoma cell lines (PC3 and LNCaP cells) to test specific induction of transcription by full-length AR and by glutathione-S-transferase (GST)-fusion peptides in which the androgen receptor (AR) DNA-binding domain alone (AR524-649), or together with the ligand-binding domain (AR524-902), or a portion of the NH2-terminal domain (AR232-649) were incorporated. In the presence of AR, nuclear extracts from PC3 cells had greater activity in supporting transcription than those from LNCaP cells; and lower background activity than those from HeLa cells. All of the AR forms correctly initiated in vitro transcription of ARE-templates in an androgen-independent manner. The amount of specific, inducible transcript was dependent on the concentration of AR peptide present. AR524-902 was the most potent transactivator tested, with the maximal level of specific transcript over 900-fold higher than the minimal level. At all concentrations this peptide was three to four times more active than either AR524-649 or AR232-649. In conclusion, we have developed a very specific and sensitive cell-free transcription system for delineating trans-activational regions of the AR.

Androgen-Binding Protein↗

PubMed1996

Characterization of 5alpha-reductase gene expression in stroma and epithelium of human prostate.

The expression of 5alpha-reductase type 1 and type 2 isoenzymes in hyperplastic human prostate tissue and several human prostate cell lines was investigated by Northern blot analyses, reverse transcription-polymerase chain reaction (RT-PCR), and enzyme activity. Separation of stroma and epithelium was confirmed histologically and only preparations with no apparent contamination were employed in the subsequent studies. Poly(A)+ RNA was isolated from stromal and epithelial fractions and analysed by Northern blot and RT-PCR. Inhibition of epithelial and stromal 5alpha-reductase activities by 17beta-N,N-diethylcarbamoyl-4-methyl-4-aza-5alpha-androstan- 3-one (4MA) was assessed using a range of concentrations between 10(-13) and 10(-5) M. Results from Northern blot analyses and RT-PCR showed that the prostate stroma expressed 5alpha-reductase type 1 and type 2 isoenzymes, whereas the prostate epithelium only expressed 5alpha-reductase type 1. This was consistent with biphasic inhibition of 5alpha-reductase activity by 4MA in stroma and monophasic inhibition in epithelium. Cultured epithelial cells derived from human prostate only expressed 5alpha-reductase type 1 and had Vmax and Km values that approximated the lower end of the range reported for surgically removed prostate epithelium. The foregoing data explains the disparate activities of 5alpha-reductase, previously reported, in stroma and epithelium. The differential localization of these isoenzymes in the prostate suggests that future therapy of androgen-sensitive disease may be more successful through the use of selective inhibitors of the different 5alpha-reductase isoenzymes.

Aged↗

PubMed1996

Effects of intermittent androgen suppression on the stem cell composition and the expression of the TRPM-2 (clusterin) gene in the Shionogi carcinoma.

The proportion of tumorigenic stem cells and the expression of the apoptosis-related gene, TRPM-2 (clusterin), were studied in populations of Shionogi carcinoma cells subjected to multiple cycles of androgen withdrawal and replacement (intermittent androgen suppression). The parent androgen-dependent cell line was initially transplanted into a male mouse which was castrated when the estimated weight of the resultant tumour became approximately 3 g. After the tumour had regressed to 40% or less of the original weight, it was transplanted into the next non-castrated male. This was repeated for four cycles of transplantation and castration-induced apoptosis before the tumour progressed to an androgen-independent state. The proportion of total stem cells in the tumour, as determined by in vivo limiting dilution assays in male mice, was constant during the first three cycles but increased 15-fold between the third and fourth cycles. In the parent androgen-dependent tumour before androgen ablation, the androgen-independent stem cell population formed 0.8% of the total stem cell compartment. After the fourth cycle this population increased to 47%; a population of similar size (33%, P = 0.8) was found in the androgen-independent recurrent form of the tumour induced by one-time castration. Whether androgen withdrawal therapy was intermittent or continuous, conversion to androgen independence thus occurred when one-third to one-half of the total stem cell compartment was populated by androgen-independent stem cells. The androgen-repressed TRPM-2 (clusterin) gene was actively expressed in regressing tumours after androgen ablation, and also became constitutively expressed in non-regressing tumours after the first and subsequent cycles of androgen withdrawal. Staining of cytoplasm and nuclei with anti-clusterin antibody was observed in androgen-dependent tumour cells after each cycle of intermittent androgen suppression; the nuclear staining was more intense in recurrent androgen-independent cells. The anomalous nuclear localization of clusterin, an anti-cytolytic TRPM-2 encoded protein, may serve to inhibit early events in the apoptotic process and thereby foster the generation and outgrowth of androgen-independent stem cells in an androgen-depleted environment.

Androgens↗

PubMed1996

Control of tumor progression by maintenance of apoptosis.

The ability to induce multiple apoptotic regressions of an androgen-dependent tumor cell population by repeated cycles of androgen withdrawal and replacement may be advantageous in therapeutic strategies aimed at delaying or preventing tumor progression. With greater insight into factors that either initiate or limit apoptosis, more efficient application of intermittent therapy might be achieved, especially if methods could be devised to increase the length or number of treatment cycles. Both calreticulin and clusterin represent proteins with a potential role in the regulation of apoptosis. Calreticulin may inhibit target gene transcription by interacting with steroid hormone receptors, thereby masking their DNA-binding sites and triggering the onset of the apoptotic process. Clusterin, on the other hand, is a membrane-stabilizing protein that appears to be involved in limiting the autophagic lysis of epithelial cells during apoptosis. Also, the increasing tendency for nuclear localization of clusterin after androgen withdrawal may preserve the nuclear environment, limiting the lethal effect of treatment. Thus, tumor progression, characterized by the loss of apoptotic potential, appears to be linked in part to the inappropriate activation of TRPM-2 gene, which accounts for the constitutive expression of clusterin.

Amino Acid Sequence↗

PubMed1996

[Hormone release and intermittent hormonal therapy in the LN CaP model of human prostate cancer].

Androgen-independent progression invariably occurs following castration of patients with prostate cancer. Animal model data suggest that androgen resistance may result from adaptive cell survival mechanisms activated by androgen withdrawal. If this is true, then re-exposure to, or low levels of, androgens may suppress or downregulate these mechanisms. The objective of this study is to determine whether intermittent androgen suppression (IAS) delays the onset of nonandrogen-regulated PSA production in the LNCaP prostate tumour model, when compared to continuous androgen suppression (CAS). Serum PSA levels correlate highly with LNCaP tumour volume and decrease rapidly following castration; beginning 3-4 weeks post-castration LN CaP tumours become androgen-independent with respect to PSA gene expression and produce PSA in amounts similar to precastrate state. In this study, IAS-treated mice were implanted with testosterone pellets beginning two weeks post-castration; cycles of testosterone replacement for 1 week and withdrawal for 2 weeks were repeated until serum PSA levels no longer returned to baseline. IAS therapy prolonged time to androgen-independent PSA production 3-fold, from an average of 26 days in the CAS group to 77 days in the IAS group. Serum and LNCaP tumour mRNA PSA levels remained below precastrate levels by 60 days post-castration in 75% of the IAS group, while serum PSA in all mice in the CAS group exceeded precastrate PSA by 28 days post-castration. Following castration, serum PSA levels increased 9-fold faster with CAS compared to IAS therapy. Observations using IAS in the LNCaP tumour model suggest that the onset of androgen-independent PSA gene regulation is prolonged 3-fold, perhaps due to androgen-induced differentiation and/or downregulation of androgen-suppressed gene expression.

Androgens↗

PubMed1995

[Theoretical considerations and initial clinical results of intermittent hormone treatment of patients with advanced prostatic carcinoma].

Androgen suppression is the routine approach to the treatment of advanced prostate cancer. Using intermittent androgen suppression by taking the advantage of the reversible action of medical castration results in the maintenance of apoptotic potential. The experiments in the androgen-dependent androgen-dependent Shionogi carcinoma tumor model as well as clinical experience in a group of men with prostate malignancy are presented in this report. These consecutive cycles of androgen withdrawal and replacement afford an improved quality of life when the patient is off therapy. It is possible to reduce toxicity, cost of treatment and to delay tumor progression. Whether survival is affected in a beneficial or adverse way still remains to be studied.

Androgen Antagonists↗

PubMed1994

Cooperative binding of androgen receptors to two DNA sequences is required for androgen induction of the probasin gene.

The functional and structural interactions of two androgen receptor-binding sites in the 5'-flanking DNA of the rat probasin gene were determined. Deletion mapping and DNase I footprinting analysis had previously identified two androgen receptor-binding sites (ARBS) necessary for androgen induction of the probasin gene: ARBS-1, which resembled a glucocorticoid-responsive element, and ARBS-2, which had a unique sequence. In this study, maximal androgen induction in transient transfection studies only occurred when both sites were present. Neither binding site functioned independently, and deletion of the DNA sequence between the sites resulted in a 60% loss of androgen inducibility. Moreover, point mutations in either ARBS-1 or ARBS-2 led to > 90% loss in activity. Scatchard analysis indicated that ARBS-1 and ARBS-2 bound a synthetic androgen receptor, AR2, with Kd values of 20.0 and 6.7 nM, respectively. Consistent with the higher affinity, ARBS-2 bound AR2 at half the threshold concentration (200 ng) of that required in reciprocal DNase I footprinting experiments with ARBS-1. By comparison, protection occurred at a much lower threshold concentration of AR2 (60 ng) and to the same extent over each site when both sites were present, suggesting a cooperative interaction between the two sites. The cooperative effect was further substantiated when a point mutation in ARBS-1 blocked AR2 binding not only to ARBS-1, but also to ARBS-2. Similarly, a point mutation in ARBS-2 also prevented receptor binding to both sites. Androgen-specific regulation of probasin gene transcription therefore required an androgen-responsive region (positions -286 and +28) containing two androgen receptor-binding sites, where the binding of the androgen receptor to both sites occurred in a cooperative, mutually dependent manner.

Androgen-Binding Protein↗