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Steven P Balk

Publications and source records attributed to Steven P Balk.

At least 19 recordsLinked to original sources
PubMed2007

Combining an mTOR antagonist and receptor tyrosine kinase inhibitors for the treatment of prostate cancer.

Inhibition of the mammalian target of rapamycin (mTOR) signaling pathway is a potentially useful therapeutic strategy in the treatment of advanced prostate cancer. However mTOR antagonists used as single agents are not likely to result in dramatic clinical responses, so that it is useful to identify prospective agents that might be useful in combination. We treated CWR22Rv1 and LNCaP prostate cancer cells with an mTOR inhibitor, rapamycin, alone, or in combination with either of two receptor protein kinase (RTK) inhibitors. We assessed the effects of these treatments on cell survival and activation of down-stream mTOR target proteins. Treatment with either PD16839, an EGFr antagonist, or imatinib mesylate (Gleevec), a PDGFr, c-kit and bcr/abl antagonist, enhanced the anti-proliferative effects of rapamycin. We therefore assessed the effects of treatment with the RTK antagonist alone and in combination with rapamycin on mTOR targeted proteins. RTK antagonists alone had no effect or paradoxically increased phosphorylation of the mTOR targeted proteins, p70 S6 kinase and ribosomal S6. In contrast, when these cells were treated with either RTK antagonist in the presence of rapamycin, there was a dramatic decrease in phosphorylation of these two mTOR-targeted proteins. These effects were not mediated through phospho-AKT. Since two separate RTK antagonists had additive antiproliferative effects in combination with an mTOR antagonist and were associated with a dramatic decrease in mTOR targeted proteins in cells with or without PTEN expression, the strategy deserves further evaluation for the treatment of prostate cancer.

Antineoplastic Agents↗
PubMed2006

Androgen receptor phosphorylation and stabilization in prostate cancer by cyclin-dependent kinase 1.

Androgen receptors (ARs) are phosphorylated at multiple sites in response to ligand binding, but the kinases mediating AR phosphorylation and the importance of these kinases in AR function have not been established. Here we show that cyclin-dependent kinase 1 (Cdk1) mediates AR phosphorylation at Ser-81 and increases AR protein expression, and that Cdk1 inhibitors decrease AR Ser-81 phosphorylation, protein expression, and transcriptional activity in prostate cancer (PCa) cells. The decline in AR protein expression mediated by the Cdk inhibitor roscovitine was prevented by proteosome inhibitors, indicating that Cdk1 stabilizes AR protein, although roscovitine also decreased AR message levels. Analysis of an S81A AR mutant demonstrated that this site is not required for transcriptional activity or Cdk1-mediated AR stabilization in transfected cells. The AR is active and seems to be stabilized by low levels of androgen in "androgen-independent" PCas that relapse subsequent to androgen-deprivation therapy. Significantly, the expression of cyclin B and Cdk1 was increased in these tumors, and treatment with roscovitine abrogated responses to low levels of androgen in the androgen-independent C4-2 PCa cell line. Taken together, these findings identify Cdk1 as a Ser-81 kinase and indicate that Cdk1 stabilizes AR protein by phosphorylation at a site(s) distinct from Ser-81. Moreover, these results indicate that increased Cdk1 activity is a mechanism for increasing AR expression and stability in response to low androgen levels in androgen-independent PCas, and that Cdk1 antagonists may enhance responses to androgen-deprivation therapy.

Androgens↗
PubMed2006

Androgens induce prostate cancer cell proliferation through mammalian target of rapamycin activation and post-transcriptional increases in cyclin D proteins.

Androgen receptor (AR) plays a central role in prostate cancer, with most tumors responding to androgen deprivation therapies, but the molecular basis for this androgen dependence has not been determined. Androgen [5alpha-dihydrotestosterone (DHT)] stimulation of LNCaP prostate cancer cells, which have constitutive phosphatidylinositol 3-kinase (PI3K)/Akt pathway activation due to PTEN loss, caused increased expression of cyclin D1, D2, and D3 proteins, retinoblastoma protein hyperphosphorylation, and cell cycle progression. However, cyclin D1 and D2 message levels were unchanged, indicating that the increases in cyclin D proteins were mediated by a post-transcriptional mechanism. This mechanism was identified as mammalian target of rapamycin (mTOR) activation. DHT treatment increased mTOR activity as assessed by phosphorylation of the downstream targets p70 S6 kinase and 4E-BP1, and mTOR inhibition with rapamycin blocked the DHT-stimulated increase in cyclin D proteins. Significantly, DHT stimulation of mTOR was not mediated through activation of the PI3K/Akt or mitogen-activated protein kinase/p90 ribosomal S6 kinase pathways and subsequent tuberous sclerosis complex 2/tuberin inactivation or by suppression of AMP-activated protein kinase. In contrast, mTOR activation by DHT was dependent on AR-stimulated mRNA synthesis. Oligonucleotide microarrays showed that DHT-stimulated rapid increases in multiple genes that regulate nutrient availability, including transporters for amino acids and other organic ions. These results indicate that a critical function of AR in PTEN-deficient prostate cancer cells is to support the pathologic activation of mTOR, possibly by increasing the expression of proteins that enhance nutrient availability and thereby prevent feedback inhibition of mTOR.

AMP-Activated Protein Kinases↗
PubMed2006

Increased expression of genes converting adrenal androgens to testosterone in androgen-independent prostate cancer.

Androgen receptor (AR) plays a central role in prostate cancer, and most patients respond to androgen deprivation therapies, but they invariably relapse with a more aggressive prostate cancer that has been termed hormone refractory or androgen independent. To identify proteins that mediate this tumor progression, gene expression in 33 androgen-independent prostate cancer bone marrow metastases versus 22 laser capture-microdissected primary prostate cancers was compared using Affymetrix oligonucleotide microarrays. Multiple genes associated with aggressive behavior were increased in the androgen-independent metastatic tumors (MMP9, CKS2, LRRC15, WNT5A, EZH2, E2F3, SDC1, SKP2, and BIRC5), whereas a candidate tumor suppressor gene (KLF6) was decreased. Consistent with castrate androgen levels, androgen-regulated genes were reduced 2- to 3-fold in the androgen-independent tumors. Nonetheless, they were still major transcripts in these tumors, indicating that there was partial reactivation of AR transcriptional activity. This was associated with increased expression of AR (5.8-fold) and multiple genes mediating androgen metabolism (HSD3B2, AKR1C3, SRD5A1, AKR1C2, AKR1C1, and UGT2B15). The increase in aldo-keto reductase family 1, member C3 (AKR1C3), the prostatic enzyme that reduces adrenal androstenedione to testosterone, was confirmed by real-time reverse transcription-PCR and immunohistochemistry. These results indicate that enhanced intracellular conversion of adrenal androgens to testosterone and dihydrotestosterone is a mechanism by which prostate cancer cells adapt to androgen deprivation and suggest new therapeutic targets.

3-Hydroxysteroid Dehydrogenases↗
PubMed2006

CD1d mediates T-cell-dependent resistance to secondary infection with encephalomyocarditis virus (EMCV) in vitro and immune response to EMCV infection in vivo.

The innate and adaptive immune responses have evolved distinct strategies for controlling different viral pathogens. Encephalomyocarditis virus (EMCV) is a picornavirus that can cause paralysis, diabetes, and myocarditis within days of infection. The optimal innate immune response against EMCV in vivo requires CD1d. Interaction of antigen-presenting cell CD1d with distinct natural killer T-cell ("NKT") populations can induce rapid gamma interferon (IFN-gamma) production and NK-cell activation. The T-cell response of CD1d-deficient mice (lacking all NKT cells) against acute EMCV infection was further studied in vitro and in vivo. EMCV persisted at higher levels in CD1d-knockout (KO) splenocyte cultures infected in vitro. Furthermore, optimal resistance to repeat cycles of EMCV infection in vitro was also shown to depend on CD1d. However, this was not reflected in the relative levels of NK-cell activation but rather by the responses of both CD4(+) and CD8(+) T-cell populations. Repeated EMCV infection in vitro induced less IFN-gamma and alpha interferon (IFN-alpha) from CD1d-deficient splenocytes than with the wild type. Furthermore, the level of EMCV replication in wild-type splenocytes was markedly and specifically increased by addition of blocking anti-CD1d antibody. Depletion experiments demonstrated that dendritic cells contributed less than the combination of NK and NKT cells to anti-EMCV responses and that none of these cell types was the main source of IFN-alpha. Finally, EMCV infection in vivo produced higher levels of viremia in CD1d-KO mice than in wild-type animals, coupled with significantly less lymphocyte activation and IFN-alpha production. These results point to the existence of a previously unrecognized mechanism of rapid CD1d-dependent stimulation of the antiviral adaptive cellular immune response.

Acute Disease↗
PubMed2006

Activation of beta-catenin signaling in prostate cancer by peptidyl-prolyl isomerase Pin1-mediated abrogation of the androgen receptor-beta-catenin interaction.

Androgen receptor (AR) interacts with beta-catenin and can suppress its coactivation of T cell factor 4 (Tcf4) in prostate cancer (PCa) cells. Pin1 is a peptidyl-prolyl cis/trans isomerase that stabilizes beta-catenin by inhibiting its binding to the adenomatous polyposis coli gene product and subsequent glycogen synthase kinase 3beta (GSK-3beta)-dependent degradation. Higher Pin1 expression in primary PCa is correlated with disease recurrence, and this study found that Pin1 expression was markedly increased in metastatic PCa. Consistent with this result, increased expression of Pin1 in transfected LNCaP PCa cells strongly accelerated tumor growth in vivo in immunodeficient mice. Pin1 expression in LNCaP cells enhanced beta-catenin/Tcf4 transcriptional activity, as assessed using Tcf4-regulated reporter genes, and increased expression of endogenous Tcf4 and c-myc. However, in contrast to results in cells with intact PTEN and active GSK-3beta, Pin1 expression in LNCaP PCa cells, which are PTEN deficient, did not increase beta-catenin. Instead, Pin1 expression markedly inhibited the beta-catenin interaction with AR, and Pin1 abrogated the ability of AR to antagonize beta-catenin/Tcf4 binding and transcriptional activity. These findings demonstrate that AR can suppress beta-catenin signaling, that the AR-beta-catenin interaction can be regulated by Pin1, and that abrogation of this interaction can enhance beta-catenin/Tcf4 signaling and contribute to aggressive biological behavior in PCa.

Androgen Receptor Antagonists↗
PubMed2006

Androgen receptor remains critical for cell-cycle progression in androgen-independent CWR22 prostate cancer cells.

The majority of prostate cancers (PCa) that relapse after androgen deprivation therapy (androgen-independent PCa) continue to express androgen receptor (AR). To study the functional importance of AR in these tumors, we derived androgen-independent CWR22 PCa xenografts in castrated mice and generated a cell line from one of these xenografts (CWR22R3). Similarly to androgen-independent PCa in patients, the relapsed xenografts and cell line expressed AR and were resistant to treatment with bicalutamide. However, expression of the AR-regulated PSA gene in the CWR22R3 cell line was markedly decreased compared to the relapsed xenografts in vivo. Transfections with androgen-regulated reporter genes further indicated that the cells lacked androgen-independent AR transcriptional activity and were not hypersensitive to low androgen concentrations despite constitutive activation of the Erk/MAP kinases. Nonetheless, AR remained essential for androgen-independent growth because retroviral shRNA-mediated AR down-regulation resulted in marked long-term growth suppression. This was associated with increased levels of p27(kip1) and hypophosphorylation of retinoblastoma protein but not with decreases in D-type cyclin levels or MAP kinase activation. These results reveal a potentially critical function of AR in androgen-independent PCa that is distinct from its previously described transcriptional or nontranscriptional functions.

Androgens↗
PubMed2005

CD1d ligation on human monocytes directly signals rapid NF-kappaB activation and production of bioactive IL-12.

Natural killer T cells (NKT cells) expressing a semi-invariant CD1d-reactive T cell receptor (invariant NKT, iNKT) can be rapidly activated by monocytes or immature dendritic cells (iDCs) bearing a CD1d-presented glycolipid antigen and can in turn stimulate these myeloid cells to mature and produce IL-12. Previous studies have shown that iNKT-produced IFNgamma and CD40 ligand contribute to this dendritic cell maturation. This study demonstrates that CD1d ligation alone, in the absence of iNKT, could rapidly (within 24 h) stimulate production of bioactive IL-12p70 by CD1d+ human peripheral blood monocytes as well as iDCs. IFNgamma alone had no effect, but it markedly enhanced CD1d-stimulated IL-12 production. Monocyte differentiation, as assessed by CD40 and CD1a up-regulation, was also accelerated by CD1d stimulation, consistent with this representing a physiological response. CD1d ligation on the human monocytic cell line THP-1 similarly specifically stimulated IL-12 production. Biochemical studies showed that IL-12 release correlated with rapid phosphorylation of IkappaB, a critical step in NF-kappaB activation. Selective NF-kappaB inhibition blocked this CD1d-stimulated IL-12 production. Finally, CD1d ligation could also enhance IL-12 production in the presence of suboptimal LPS or CD40 stimulation. These findings demonstrate an innate immune signaling function for CD1d and provide a mechanism for the rapid activation of monocytes and iDCs by CD1d-reactive T cells.

Antigens, CD1↗
PubMed2005

Identification of hypoxia-inducible factor-1alpha (HIF-1alpha) polymorphism as a mutation in prostate cancer that prevents normoxia-induced degradation.

BACKGROUND: Hypoxia-inducible factor-1alpha (HIF-1alpha) regulates cellular responses to hypoxia and is rapidly degraded under normoxia through von Hippel-Lindau (VHL) mediated ubiquitination. Although HIF-1alpha stabilization appears to be the molecular basis for VHL-associated cancers, stabilizing mutations in HIF-1alpha have not been reported. METHODS: A series of 15 metastatic androgen independent prostate cancers were examined for mutations in the oxygen-dependent domain (ODD) of HIF-1alpha by PCR amplification and DNA sequencing. RESULTS: A somatic proline to serine mutation in codon 582 (P582S) was identified in one sample. Transfection studies with a HIF-1alpha regulated reporter gene showed increased transcriptional activity that correlated with higher mutant HIF-1alpha protein expression. Increased expression of the P582S mutant induced by iron chelation, which blocks proline hydroxylation of wild-type HIF-1alpha, was markedly attenuated. The mutant also showed increased stability under normoxic versus hypoxic conditions. CONCLUSION: The P582S HIF-1alpha is a stable variant and HIF-1alpha mutation is a mechanism for enhancing HIF-1alpha activity in human cancer. The recent identification of the identical P582S HIF-1alpha as a polymorphism suggests that this variant may increase tumor susceptibility or cause more aggressive biological behavior.

Biopsy↗
PubMed2005

CD1d and CD1d-restricted iNKT-cells play a pivotal role in contact hypersensitivity.

CD1d-restricted T-cells are activated by glycolipids presented by the major histocompatibility complex class-Ib molecule CD1d, found on the surface of antigen-presenting cells (APC). This interaction between APC, most notably dendritic cells (DC), and CD1d-restricted T-cells is an important regulatory step in the initiation of adaptive immune responses. It is well known that DC play a crucial role in the induction of contact hypersensitivity (CHS), a frequently studied form of in vivo T-cell-mediated immunity. In this study, we show that CD1d-restricted T-cells are also necessary for CHS, because both wild-type mice treated systemically or topically with CD1d glycolipid antagonists and CD1d-restricted T-cell-null mice have markedly diminished CHS responses. Thus, pharmacologic antagonists of CD1d can be used as effective inhibitors of CHS, a prototype for a variety of delayed-type tissue hypersensitivity responses.

Administration, Topical↗
PubMed2005

The analysis of systemic tolerance elicited by antigen inoculation into the vitreous cavity: vitreous cavity-associated immune deviation.

The immune privilege that exists in the eye is maintained by various mechanisms. One of the best studied is a form of systemic tolerance termed anterior chamber-associated immune deviation (ACAID). We have investigated the mechanisms by which ocular inflammation associated with the vitreous cavity (VC) is reduced, by injecting either ovalbumin (OVA) or allogeneic splenocytes into the VCs of mice, and assessed the effect of this on delayed type hypersensitivity (DTH) responses. After antigen inoculation into the VC, antigen-specific DTH responses were significantly impaired and we named this phenomenon 'vitreous cavity-associated immune deviation' (VCAID). VCAID could also be induced by inoculating antigen-pulsed macrophages into the VC. However, VCAID did not develop either in mice with inflamed eyes, whether as a result of experimental autoimmune uveitis or coadministration of interleukin (IL)-6 in the VC, or in knockout mice deficient for natural killer T (NKT) cells. Finally, we found that so-called 'hyalocytes' are the only cells present in the VCs of normal mice, uniformly distributed on the retinal surface. Interestingly, they express F4/80, suggesting that hyalocytes are candidate antigen-presenting cells (APCs) responsible for mediating VCAID. As for the anterior chamber model, systemic tolerance can be induced in the VC in non-inflamed eyes and in the presence of invariant NKT cells.

Animals↗
PubMed2004

Recruitment of the androgen receptor via serum response factor facilitates expression of a myogenic gene.

Androgen receptor (AR) induced precocious myogenesis in culture and myogenic specified gene activity. Increased levels of AR expression in replicating C2C12 myoblasts stimulated fusion into post-differentiated multinucleated myotubes and the appearance of skeletal alpha-actin transcripts, even in the absence of ligand. Furthermore, AR activated the skeletal alpha-actin promoter, which lacks GRE sites, in co-transfected C2C12 cells. AR co-activation of the skeletal alpha-actin promoter required co-expressed full-length serum response factor (SRF). In vitro, AR associated with SRF and was recruited by SRF to a alpha-actin promoter SRF binding site. Our data suggest that AR is capable of activating myogenic genes devoid of consensus AR binding sites via its recruitment by the myogenic enriched transcription factor, SRF.

Actins↗
PubMed2004

The androgen receptor recruits nuclear receptor CoRepressor (N-CoR) in the presence of mifepristone via its N and C termini revealing a novel molecular mechanism for androgen receptor antagonists.

The androgen receptor (AR) activates target gene expression in the presence of agonist ligands via the recruitment of transcriptional coactivators, but recent work shows that overexpression of the nuclear corepressors NCoR and SMRT attenuates this agonist-mediated AR activation. Here we demonstrate using NCoR siRNA and chromatin immunoprecipitation that endogenous NCoR is recruited to and represses the dihydrotestosterone (DHT)-liganded AR. Furthermore this study shows that NCoR and coactivators compete for AR in the presence of DHT. AR antagonists such as bicalutamide that are currently in use for prostate cancer treatment can also mediate NCoR recruitment, but mifepristone (RU486) at nanomolar concentrations is unique in its ability to markedly enhance the AR-NCoR interaction. The RU486-liganded AR interacted with a C-terminal fragment of NCoR, and this interaction was mediated by the two most C-terminal nuclear receptor interacting domains (RIDs) present in NCoR. Significantly, in addition to the AR ligand binding domain, the AR N terminus was also required for this interaction. Mutagenesis studies demonstrate that the N-terminal surface of the AR-mediating NCoR recruitment was distinct from tau5 and from the FXXLF motif that mediates agonist-induced N-C-terminal interaction. Taken together these data demonstrate that NCoR is a physiological regulator of the AR and reveal a new mechanism for AR antagonism that may be exploited for the development of more potent AR antagonists.

Androgen Receptor Antagonists↗
PubMed2004

Activation of p21-activated kinase 6 by MAP kinase kinase 6 and p38 MAP kinase.

The p21-activated kinases (PAKs) contain an N-terminal Cdc42/Rac interactive binding domain, which in the group 1 PAKs (PAK1, 2, and 3) regulates the activity of an adjacent conserved autoinhibitory domain. In contrast, the group 2 PAKs (PAK4, 5, and 6) lack this autoinhibitory domain and are not activated by Cdc42/Rac binding, and the mechanisms that regulate their kinase activity have been unclear. This study found that basal PAK6 kinase activity was repressed by a p38 mitogen-activated protein (MAP) kinase antagonist and could be strongly stimulated by constitutively active MAP kinase kinase 6 (MKK6), an upstream activator of p38 MAP kinases. Mutation of a consensus p38 MAP kinase target site at serine 165 decreased PAK6 kinase activity. Moreover, PAK6 was directly activated by MKK6, and mutation of tyrosine 566 in a consensus MKK6 site (threonine-proline-tyrosine, TPY) in the activation loop of the PAK6 kinase domain prevented activation by MKK6. PAK6 activation by MKK6 was also blocked by mutation of an autophosphorylated serine (serine 560) in the PAK6 activation loop, indicating that phosphorylation of this site is necessary for MKK6-mediated activation. PAK4 and PAK5 were similarly activated by MKK6, consistent with a conserved TPY motif in their activation domains. The activation of PAK6 by both p38 MAP kinase and MKK6 suggests that PAK6 plays a role in the cellular response to stress-related signals.

Cell Line↗
PubMed2004

Recruitment of beta-catenin by wild-type or mutant androgen receptors correlates with ligand-stimulated growth of prostate cancer cells.

Prostate cancers respond to treatments that suppress androgen receptor (AR) function, with bicalutamide, flutamide, and cyproterone acetate (CPA) being AR antagonists in clinical use. As CPA has substantial agonist activity, it was examined to identify AR coactivator/corepressor interactions that may mediate androgen-stimulated prostate cancer growth. The CPA-liganded AR was coactivated by steroid receptor coactivator-1 (SRC-1) but did not mediate N-C terminal interactions or recruit beta-catenin, indicating a nonagonist conformation. Nonetheless, CPA did not enhance AR interaction with nuclear receptor corepressor, whereas the AR antagonist RU486 (mifepristone) strongly stimulated AR-nuclear receptor corepressor binding. The role of coactivators was further assessed with a T877A AR mutation, found in LNCaP prostate cancer cells, which converts hydroxyflutamide (HF, the active flutamide metabolite) into an agonist that stimulates LNCaP cell growth. The HF and CPA-liganded T877A ARs were coactivated by SRC-1, but only the HF-liganded T877A AR was coactivated by beta-catenin. L-39, a novel AR antagonist that transcriptionally activates the T877A AR, but still inhibits LNCaP growth, similarly mediated recruitment of SRC-1 and not beta-catenin. In contrast, beta-catenin coactivated a bicalutamide-responsive mutant AR (W741C) isolated from a bicalutamide-stimulated LNCaP subline, further implicating beta-catenin recruitment in AR-stimulated growth. Androgen-stimulated prostate-specific antigen gene expression in LNCaP cells could be modulated by beta-catenin, and endogenous c-myc expression was repressed by dihydrotestosterone, but not CPA. These results indicate that interactions between AR and beta-catenin contribute to prostate cell growth in vivo, although specific growth promoting genes positively regulated by AR recruitment of beta-catenin remain to be identified.

Cell Cycle↗
PubMed2004

Androgen receptor: a key molecule in the progression of prostate cancer to hormone independence.

Despite earlier detection and recent advances in surgery and radiation, prostate cancer is second only to lung cancer in male cancer deaths in the United States. Hormone therapy in the form of medical or surgical castration remains the mainstay of systemic treatment in prostate cancer. Over the last 15 years with the clinical use of prostate specific antigen (PSA), there has been a shift to using hormone therapy earlier in the disease course and for longer duration. Despite initial favorable response to hormone therapy, over a period of time these tumors will develop androgen-independence that results in death. The androgen receptor (AR) is central to the initiation and growth of prostate cancer and to its response to hormone therapy. Analyses have shown that AR continues to be expressed in androgen-independent tumors and AR signaling remains intact as demonstrated by the expression of the AR regulated gene, PSA. Androgen-independent prostate cancers have demonstrated a variety of AR alterations that are either not found in hormone naïve tumors or found at lower frequency. These changes include AR amplification, AR point mutation, and changes in expression of AR co-regulatory proteins. These AR changes result in a "super AR" that can respond to lower concentrations of androgens or to a wider variety of agonistic ligands. There is also mounting evidence that AR can be activated in a ligand independent fashion by compounds such as growth factors or cytokines working independently or in combination. These growth factors working through receptor tyrosine kinase pathways may promote AR activation and growth in low androgen environments. The clinical significance of these AR alterations in the development and progression of androgen-independent prostate cancer remains to be determined. Understanding the changes in AR signaling in the evolution of androgen-independent prostate cancer will be key to the development of more effective hormone therapy.

Disease Progression↗
PubMed2004

Androgen receptor down-regulation in prostate cancer with phosphorodiamidate morpholino antisense oligomers.

PURPOSE: Androgen receptor (AR) has a pivotal role in the growth and proliferation of prostate cancer (PCa). Even in advanced stages of PCa AR continues to be expressed and appears to be functional. Since the mechanisms of AR activation in androgen independent PCa have yet to be clearly defined, the decrease in AR protein by antisense compounds is an attractive therapeutic option. In this study we evaluated a novel antisense phosphorodiamidate morpholino oligomer (PMO) targeting the translational start site of AR mRNA in vitro and in vivo in a PCa xenograft and murine prostate. MATERIALS AND METHODS: AR antisense PMOs targeting the AR initiation AUG were tested in vitro and in LNCaP cells, and in vivo in LAPC-4 xenografts and normal mouse prostate. Effects on AR protein and PSA expression were assessed. RESULTS: AR antisense PMOs specifically down-regulated AR protein levels in a plasmid based screening system and also decreased endogenous AR levels in androgen responsive LNCaP cells in culture compared to control nonspecific PMOs. Pretreatment and posttreatment biopsies in the LAPC-4 xenograft model demonstrated that the antisense AR PMO administered intraperitoneally specifically decreased AR protein levels and serum PSA. Analysis of tissue distribution of the AR PMO by high performance liquid chromatography based methodology showed significant PMO levels in tumor tissue and mouse prostate, and there was a dose dependent decrease in AR protein levels in murine AR antisense PMO treated mouse prostates. CONCLUSIONS: An AR antisense PMO with unique chemical properties administered once daily can decrease AR protein levels and PSA in vivo. The reduction of AR protein with an antisense PMO may be an effective method of interfering with AR mediated growth in advanced human PCa.

Animals↗