Testosterone Pellets Research Articles

Abstract 1800: Targeted therapy using saracatinib adjunctive to castration inhibits progression to castration-resistant prostate cancer in a murine model

Abstract Introduction: Prostate cancer treatment with androgen deprivation is typically efficacious for two years prior to the development of castration resistant disease (CRPC). The Src family kinases (SFK) are upregulated with androgen deprivation and have been implicated in the progression to CRPC. Saracatinib is a specific SFK inhibitor that inhibits castration resistant growth and metastases in an animal model using LNCaP cells overexpressing the neuropeptide gastrin-releasing peptide. We test the hypothesis that saracatinib given immediately after androgen deprivation inhibits castration resistant tumor recurrence using a CWR22 xenograft model. Materials and Methods: CWR22 prostate cancer xenografts were injected subcutaneously into nude mice primed with testosterone pellets (21-day release). Twenty-six mice were castrated after adequate tumor development (day 0). Mice in the treatment arm were administered saracatinib from day 7 to day 150. Mice were sacrificed at days 0, 3, 30 and 150. Tumors and sera were collected for analysis including tumor weight, serum testosterone, serum PSA and gene-profile studies. The data was analyzed using the t-test and analysis of variance. Results: CRPC recurrence was significantly inhibited with the administration of saracatinib. The mean weight of tumor recurrence was 0.18 grams with saracatinib compared to 3.20 grams in the control group (p=0.015). Saracatinib was associated with a reduction in serum PSA (1.6 versus 14.7 ng/dl) and testosterone (21.1 versus 24.1 pg/ml) compared to controls, though statistical significance was not reached (p=0.08 and p=0.63, respectively). Gene expression of signaling kinases and their downstream olecules including Src, Etk, FAK, JAK, Stat3, c-myc, β-catenin, NF-κB, Akt, and mTOR is being compared between time points. Conclusions: Saracatinib significantly inhibits castration resistant tumor recurrence when started with androgen deprivation. Additional investigation into the genetic and molecular changes induced by saracatinib with gene microarray and polymerase chain reaction is ongoing. Saracatinib may have a role in combination with androgen deprivation to delay the onset of castration resistant disease. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1800. doi:1538-7445.AM2012-1800

1570 TADALAFIL, THE PHOSPHODIESTERASE-5 INHIBITOR, ATTENUATES THE CELL GROWTH OF HUMAN BENIGN PROSTATIC HYPERPLASIA XENOGRAFTS IN SUPER-SCID MICE

You have accessJournal of UrologyBenign Prostatic Hyperplasia: Basic Research1 Apr 20121570 TADALAFIL, THE PHOSPHODIESTERASE-5 INHIBITOR, ATTENUATES THE CELL GROWTH OF HUMAN BENIGN PROSTATIC HYPERPLASIA XENOGRAFTS IN SUPER-SCID MICE Hiroshi Kiuchi, Akira Tsujimura, Hidenobu Okuda, Keisuke Yamamoto, Shinichirou Fukuhara, Iwao Yoshioka, Tetsuya Takao, Yasushi Miyagawa, Norio Nonomura, and Taisei Nomura Hiroshi KiuchiHiroshi Kiuchi Suita, Japan More articles by this author , Akira TsujimuraAkira Tsujimura Suita, Japan More articles by this author , Hidenobu OkudaHidenobu Okuda Suita, Japan More articles by this author , Keisuke YamamotoKeisuke Yamamoto Suita, Japan More articles by this author , Shinichirou FukuharaShinichirou Fukuhara Suita, Japan More articles by this author , Iwao YoshiokaIwao Yoshioka Suita, Japan More articles by this author , Tetsuya TakaoTetsuya Takao Suita, Japan More articles by this author , Yasushi MiyagawaYasushi Miyagawa Suita, Japan More articles by this author , Norio NonomuraNorio Nonomura Suita, Japan More articles by this author , and Taisei NomuraTaisei Nomura Ibaraki, Japan More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2012.02.1342AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Benign prostatic hyperplasia (BPH) is characterized by a pathologic proliferation of the prostate glandular and stromal tissues, and subsequent induction of lower urinary tract symptoms. Emerging evidences suggest that phosphodiesterase-5 (PDE-5) inhibitors improve urinary symptoms in patients with BPH. However, inhibition of prostate growth by PDE-5 inhibitors has not been fully elucidated. The objective of this study is to evaluate the effect of Tadalafil, the PDE-5 inhibitor, on suppression of prostate cell growth using a novel human BPH xenograft model. METHODS Human BPH tissue was obtained from a single patient who had undertaken retropubic prostatectomy for the treatment of BPH and was transplanted under the skin of super-SCID mice; SCID mice with macrophage dysfunction. BPH tissues were collected after 2 months of treatment with Tadalafil orally or nothing (control) with sustained release testosterone pellet. Immunohistochemical study was performed to examine the histology and expression of cell type-specific markers in xenografts, including PSA, androgen receptor (AR), p63 (basal cell marker), cytokeratin 8(K8, luminal cell marker) and human-specific CD31 (human vessel marker). We next assess the effect of Tadalafil on the prostate cell proliferation by Ki67 and apoptosis by cleaved caspase-3. Endothelial NOS (eNOS) and COX-2 expression were also examined. RESULTS The implanted tissue contained normal prostatic components; the gland showed two layers of epithelial cells. Inner layer expressed K8, AR and PSA, and outer layer was positive for p63. A considerable amount of the human originated vessels was apparent. After 2 months of Tadalafil treatment, a proliferation index (percentage of Ki-positive cells) was significantly reduced, compared with control in epithelial cells (1.4% vs. 3.9%, p<0.05) as well as in stromal tissues (51.7 vs.14.1 cells/mm2, p<0.05). Cleaved caspase-3 positive cells were significantly increased by Tadalafil. In addition, Tadalafil induced up-regulation of eNOS expression and down-regulation of COX-2 expression. CONCLUSIONS Our novel xenograft model has maintained appropriate histology and protein expression, and may serve as a useful human BPH model. Our study also indicates that Tadalafil attenuates the cell growth and induces apoptosis in benign prostate cells through NO signaling. These results provide a therapeutic potential of Tadalafil for reducing prostate volume in patients with BPH. © 2012 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 187Issue 4SApril 2012Page: e636 Advertisement Copyright & Permissions© 2012 by American Urological Association Education and Research, Inc.MetricsAuthor Information Hiroshi Kiuchi Suita, Japan More articles by this author Akira Tsujimura Suita, Japan More articles by this author Hidenobu Okuda Suita, Japan More articles by this author Keisuke Yamamoto Suita, Japan More articles by this author Shinichirou Fukuhara Suita, Japan More articles by this author Iwao Yoshioka Suita, Japan More articles by this author Tetsuya Takao Suita, Japan More articles by this author Yasushi Miyagawa Suita, Japan More articles by this author Norio Nonomura Suita, Japan More articles by this author Taisei Nomura Ibaraki, Japan More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

975 ORTHOTOPIC TUMORGRAFTS DERIVED FROM MEN WITH LOCALIZED PROSTATE CANCER REFLECT INITIAL TUMOR PATHOLOGY – A NEW MODEL TO STUDY PROSTATE CANCER

INTRODUCTION AND OBJECTIVES: Animal models that authentically reproduce tumor growth are still sparse in prostate cancer research for several reasons: Whilst orthotopic implantation is technically difficult, most models favor subcutaneous tumor growth that provides little information about natural tumor growth behaviour and tumor stroma interaction. Established prostate cancer cell lines as in vivo xenografts are not able to reflect the variety of tumor specific growth patterns and growth behaviour. Primary cell cultures are difficult to handle and an induction of orthotopic tumors has not been successful yet. Therefore, a tumorgraft model using tumor tissue from prostatectomy specimens was recently developed. METHODS: Balb/c mice were used to graft fresh prostate tumor tissue by surgical orthotopic implantation. Tissue implantation was performed at four inoculation sides in each animal and testosterone propionate was supplemented. Animals were tracked after 12, 16 and 20 weeks by means of 30MHz ultrasound to monitor tumor engraftment. Autopsy, histology and PSA measurements were used to confirm orthotopic tumors after 13, 21 and 28 weeks. RESULTS: To date, tumor implantation was performed in 17 mice. 3 animals were excluded due to early dead or repetitive loss of the testosterone pellet. After successful testosterone supplementation PSA measurement was possible in 12 of 14 mice (86%, range: 0,009 80ng/ml). So far, orthotopic tumor growth was histologically verified in 6 of 14 mice. Ultrasound seems feasible to detect tumors. Of interest, tumorgrafts maintain essential features of the original tumors as tumor growth pattern, PSA expression, androgen receptor expression and high proliferation using human Ki67 immunostaining. CONCLUSIONS: The tumorgrafts resembled histological properties of organ confined prostate cancer. Measurements of serum PSA and three-dimensional ultrasound were useful for monitoring of tumor engraftment and growth. Further studies like comparative genomic hybridization to genetically characterize tumorgrafts are necessary. Besides their role as a bank of tumor samples for repetitive in vitro studies, tumorgrafts display a new approach for an optimized in vivo model of prostate cancer and will allow further investigations on specific components of tumor initiation and progression, including the interactions of prostate cancer cells with the surrounding stroma. Whether this model is also applicable for metastatic progression needs to be further investigated.

Abstract A3: A tumorgraft model of castration-resistant primary prostate cancer

Abstract Androgen receptor (AR) signaling plays a central role in all stages of prostate cancer (PCa). Androgen deprivation therapy (ADT) is used as adjuvant therapy for high-risk PCa as well as the first-line therapy for metastatic PCa. However, tumor remissions are temporary and patients almost inevitably progress to become castration resistant prostate cancer (CRPC). Most CRPC still expresses AR and depends on AR for growth, therefore, there is an urgent need to understand the regulation of AR expression and to find novel ways of inhibiting this pathway in CRPC. We developed a novel realistic model for primary CRPC by establishing tumorgrafts with thin, precision-cut tissue slices derived from high-grade primary PCa under the renal capsule of immunodeficient mice supplemented with subcutaneous testosterone pellets. One month after the implantation, the testosterones pellets were removed and the mice were castrated. Tumorgrafts from control and castrated mice were harvested one month after castration and fixed in formalin. Immunohistochemistry was performed to examine the expression of genes of interest in the grafts. Tumorgrafts derived from high-grade primary PCa resemble the original tumors in expression of all genes examined. Specifically, they are positive for AR, prostate specific antigen (PSA), cytokeratin 18, and AMCAR, but negative for p63. After castration, cancer cells remained in grafts derived from two of four tumors, whereas cancer cells in grafts derived from the other two tumors were almost completely eliminated, suggesting that some tumors respond to androgen deprivation therapy (ADT) better than the others. In the castration-resistant tumorgrafts (CRTs), PSA expression was diminished in cancer cells, but AR and cytokeratin 18 expression remained, similar to responses of human prostate to ADT. In addition, cancer cells in CRTs were less proliferative than in control grafts as determined by Ki67 staining consistent with the observation that ADT inhibit cell proliferation in human prostate. More importantly, CRTs expressed AR target genes that have been reported to be upregulated under androgen ablation in other experimental models. For example, the number of cyclin A-expressing cells was significantly higher in CRTs than in control grafts. Our results demonstrated that our novel CRT model has the appropriate characteristics to serve as a useful tool to model primary CRPC. Our results demonstrated that our novel CRT model has the appropriate characteristics to serve as a useful tool to model primary CRPC and may be used as a surrogate model for clinical trials to achieve rapid and less expressive screening of therapeutic agents for neoadjuvant or adjuvant therapy of PCa. Citation Format: Hongjuan Zhao, Alan Thong, Rosie Nolley, Stephen Reese, Jennifer Santos, Donna Peehl. A tumorgraft model of castrationresistant primary prostate cancer [abstract]. In: Proceedings of the AACR Special Conference on Advances in Prostate Cancer Research; 2012 Feb 6-9; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(4 Suppl):Abstract nr A3.

A Multi‐Institutional Observational Study of Testosterone Levels after Testosterone Pellet (Testopel®) Insertion

Implantable testosterone pellets were approved by the Food and Drug Administration in 1972 for the treatment of testosterone deficiency syndrome (TDS). Clinical use of this testosterone delivery modality has been limited until its recent reintroduction (Testopel(®) , Slate Pharmaceuticals, Durham, NC, USA). Six academic institutions collaborated and combined their databases to more fully characterize serum testosterone levels after the pellet implantations. To assess the time-dependent serum testosterone levels after subcutaneous testosterone pellets in clinical practice for the treatment of TDS. Data were retrospectively pooled and analyzed from data in six academic institutions. Variables included patient age, total testosterone concentrations before and after implantation, the number of testosterone pellets implanted, and the time from implantation to measurement of serum testosterone concentrations. Three hundred eighty men undergoing 702 insertions were included for analysis using JMP (version 4.0.4; SAS Institute, Cary, NC, USA). Main outcome measures were postimplantation total testosterone levels and investigator-reported adverse events. Testosterone levels as a function of the number of pellets implanted and time from implantation were assessed. Implantation of six to ≥10 testosterone pellets (450 to ≥750 mg) increased total testosterone into the therapeutic range at 1 month postimplantation and sustained therapeutic levels (>300) for 4-6 months. Higher pellet numbers (10-12 pellets) were associated with higher, more consistent, and longer maintenance of testosterone levels within the therapeutic range. Four extrusions and three hematomas were reported early in our experience; other investigator-reported adverse events were generally mild to moderate in nature and transient in duration. No subjects required analgesics. Testosterone pellets (Testopel(®) , Slate Pharmaceuticals) provide sustained levels of testosterone for at least 4 months and up to 6 months in men with TDS. Implantation of ≥8 pellets achieved optimal results with respect to peak mean testosterone level and duration of effect. Testosterone pellets were generally well tolerated.

Primary Xenografts of Human Prostate Tissue as a Model to Study Angiogenesis Induced by Reactive Stroma

Characterization of the mechanism(s) of androgen-driven human angiogenesis could have significant implications for modeling new forms of anti-angiogenic therapies for CaP and for developing targeted adjuvant therapies to improve efficacy of androgen-deprivation therapy. However, models of angiogenesis by human endothelial cells localized within an intact human prostate tissue architecture are until now extremely limited. This report characterizes the burst of angiogenesis by endogenous human blood vessels in primary xenografts of fresh surgical specimens of benign prostate or prostate cancer (CaP) tissue that occurs between Days 6–14 after transplantation into SCID mice pre-implanted with testosterone pellets. The wave of human angiogenesis was preceded by androgen-mediated up-regulation of VEGF-A expression in the stromal compartment. The neo-vessel network anastomosed to the host mouse vascular system between Days 6–10 post-transplantation, the angiogenic response ceased by Day 15, and by Day 30 the vasculature had matured and stabilized, as indicated by a lack of leakage of serum components into the interstitial tissue space and by association of nascent endothelial cells with mural cells/pericytes. The angiogenic wave was concurrent with the appearance of a reactive stroma phenotype, as determined by staining for α-SMA, Vimentin, Tenascin, Calponin, Desmin and Masson's trichrome, but the reactive stroma phenotype appeared to be largely independent of androgen availability. Transplantation-induced angiogenesis by endogenous human endothelial cells present in primary xenografts of benign and malignant human prostate tissue was preceded by induction of androgen-driven expression of VEGF by the prostate stroma, and was concurrent with and the appearance of a reactive stroma phenotype. Androgen-modulated expression of VEGF-A appeared to be a causal regulator of angiogenesis, and possibly of stromal activation, in human prostate xenografts.

Deficits in Spermatogenesis but not Neurogenesis are Alleviated by Chronic Testosterone Therapy in R6/1 Huntington’s Disease Mice

Despite the well established central pathophysiology of Huntington's disease (HD), less is known about systemic impairments that are emerging as significant contributors to the morbidity of this neurodegenerative condition. Given the evidence of neuroendocrine dysfunction in HD patients and the pro-neural properties of sex-hormones, we explored the therapeutic potential of hormone therapy in the HD R6/1 mouse model (HD mice). HD mice over-express exon-1 of the defective human HD gene and replicate many of the clinical behavioural, biochemical and physiological impairments. Seven-week-old HD and wild-type littermate mice had either saline (control) or testosterone (treatment; 160μg/day over 90days) pellets implanted s.c. and were subsequently subjected to behavioural, molecular and cellular analysis. Separate mice were used to establish a decrease in serum testosterone concentrations in HD mice at 12weeks of age. Baseline serum testosterone was significantly reduced in control 19-week-old HD mice, whereas treatment significantly raised serum testosterone in both wild-type and HD mice. Testosterone treatment had a limited effect on the development of rotarod deficiencies in HD mice and no effect on progressive body weight loss or the development of central mutant huntingtin-containing aggregates. Testosterone treatment induced hypo-locomotion in both genotypes. Deficits in hippocampal-dependent cognition and neurogenesis were not rescued in testosterone-treated HD mice. By contrast, wild-type-treatment mice experienced significantly increased neuronal survival and differentiation. Testosterone treatment in HD mice did rescue androgen receptor levels in the hippocampus and testes, significantly improved severe testicular atrophy and restored spermatogenesis. We conclude that chronic testosterone provides systemic efficacy in treating spermatogenesis deficits and testicular atrophy but not central cellular and behavioural pathologies in R6/1 HD mice.

Outcomes with the “V” Implantation Technique vs. Standard Technique for Testosterone Pellet Therapy

Standard technique (ST) for implantation of testosterone pellets involves making a single linear track in the subcutaneous tissue of the buttock from the incision. After our initial experience with this modality, we modified this surgical technique to our current "V" technique (VT). This involves two tracks both caudally directed and emanating from the same skin incision but angulated approximately 10-15 degrees apart. While this allows additional pellets to be inserted more easily, and increased space to place those pellets further from the skin incision, it minimally increases the surgical procedure. We sought to examine the impact of this technical modification on therapeutic efficacy and surgical complication rates. Retrospective chart review of all patients treated with testosterone pellets at our institution. Complication rates for infection, extrusion, hematoma, and pain. One hundred sixty-eight patients underwent 281 implantation procedures (40 via ST and 241 via VT). The mode number of pellets used with ST was 8 (range 6-8) and with VT was 10 (range 10-13). Incidence of pellet extrusion was 7.5% with ST and 0.8% with VT. Infection complicated ST in 5% of cases but only 1.2% with VT. No cases of hematoma were seen with ST but 1.2% of VT cases. Pain prompting discontinuation of therapy was seen in 7.5% with ST and 1.7% with VT. Significant pain without discontinuation was seen in 5% with ST and 1.2% with VT. Only in 1 of the 3 cases of hematoma was the individual on blood thinners. Fifty-eight other insertions were performed on blood thinners without significant hematoma. None of the individuals who developed infection or bleeding required additional surgical therapy. VT allows successful placement of larger number of pellets, with low rates of complications, especially extrusion, even in men on anticoagulants.

Androgen suppresses proliferation of castration‐resistant LNCaP 104‐R2 prostate cancer cells through androgen receptor, Skp2, and c‐Myc

Androgen ablation therapy is the primary treatment for metastatic prostate cancer. However, this therapy is associated with several undesired side-effects, including increased risk of cardiovascular diseases. To study if termination of long-term androgen ablation and restoration of testosterone levels could suppress the growth of relapsed hormone-refractory prostate tumors, we implanted testosterone pellets in castrated nude mice carrying androgen receptor (AR)-positive LNCaP 104-R2 cells, which relapsed from androgen-dependent LNCaP 104-S cells after long-term androgen deprivation. 104-R2 tumor xenografts regressed after testosterone pellets were implanted. Of 33 tumors, 24 adapted to elevation of testosterone level and relapsed as androgen-insensitive tumors. Relapsed tumors (R2Ad) expressed less AR and prostate-specific antigen. We then studied the molecular mechanism underlying the androgenic regulation of prostate cancer cell proliferation. Androgen suppresses proliferation of 104-R2 by inducing G(1) cell cycle arrest through reduction of S-phase kinase-associated protein 2 (Skp2) and c-Myc, and induction of p27(Kip1). 104-R2 cells adapted to androgen treatment and the adapted cells, R2Ad, were androgen-insensitive cells with a slower growth rate and low protein level of AR, high levels of c-Myc and Skp2, and low levels of p27(Kip1). Nuclear AR and prostate-specific antigen expression is present in 104-R2 cells but not R2Ad cells when androgen is absent. Overexpression of AR in R2Ad cells regenerated an androgen-repressed phenotype; knockdown of AR in 104-R2 cells generated an androgen-insensitive phenotype. Overexpression of Skp2 and c-Myc in 104-R2 cells blocked the growth inhibition caused by androgens. We concluded that androgens cause growth inhibition in LNCaP 104-R2 prostate cancer cells through AR, Skp2, and c-Myc.

A Phase IV Prospective Evaluation of the Safety and Efficacy of Extended Release Testosterone Pellets for the Treatment of Male Hypogonadism

Men with hypogonadism exhibit decreased serum testosterone levels and may experience a constellation of clinical symptoms, including decrease in muscle mass, loss of sexual desire, impotence, and infertility. While previous studies have shown that implantation of extended release testosterone pellets can provide therapeutic levels of testosterone over several months, additional data are needed to establish this approach as the standard of care for male hypogonadism. To evaluate the safety and efficacy of testosterone pellets over 6 months as a treatment for male hypogonadism in a clinical practice setting. A phase IV, single center, open-label study designed to assess the safety and efficacy of subcutaneous insertion of 8 to 12 testosterone 75 mg pellets (450 mg to 900 mg), during a single implantation procedure in hypogonadal men. Subjects who successfully completed the protocol were allowed to enroll in an extension study that included another implantation and 6 months of follow-up. Safety was determined by investigator-reported adverse events, changes in vital signs, physical exam findings, and laboratory tests. Efficacy was based on serum laboratory tests, physical exams, implantation site evaluations, and vital signs. Secondary objectives were to assess patient preference for testosterone pellets and to maintain optimal total testosterone. Mean testosterone significantly increased and luteinizing hormone (LH) levels significantly decreased from pre-implantation values at weeks 1, 4, and 12, and had returned to pre-implantation levels by week 24. Prostate-specific antigen levels remained unchanged for the duration of the study. Improvements in several symptoms of hypogonadism were determined with multiple questionnaires. Implanted testosterone pellets were generally well tolerated. Implanted testosterone pellets can normalize testosterone and LH levels and improve symptoms for at least 3 months and up to 6 months in men with hypogonadism, and should be considered as a therapeutic option for hypogonadal men.

Abstract LB-203: Aromatase expression promotes breast cancer tumorigenesis and bone metastasis

Abstract Aromatase P450 (CYP19) is the rate-limiting enzyme catalyzing the final step in estrogen biosynthesis, and an important target in breast cancer therapy. In post-menopausal women, ovaries cease to make estrogen, yet the incidence of breast cancer increases with age as aromatase present in breast adipose stroma and epithelium catalyzes local estrogen formation. The circulating estrogen level in postmenopausal women is very low and it reflects the sum of local estrogen biosynthesis in various sites. Although breast cancer cells have been shown to express aromatase prompting the hypothesis that intracrine action of estrogen in breast cancer cells may drive tumor progression, there has been little evidence in addressing the hypothesis. We stably transfected the ER positive (ER+) breast cancer ZR-75-1 cells with an human aromatase expression vector and obtained a few clones expressing a moderately higher level of aromatase in comparison with the control vector-transfected cells. One of the clones called ZR-75-1/Aro/Clone10 (Cl10) was selected for further study due to its increased intracellular estradiol (E2) concentration compared to other clones. The Cl10 cells formed tumors after being xenografted into the inguinal mammary fat pad of female athymic mice while the control cells were incapable of forming tumors without estrogen supplementation. Implantation of testosterone pellet further stimulated the growth of the tumor and a cell line, ZR-75-1/Aro/Clone10/TT1 (TT1) was established from the tumor. Both Cl10 and TT1 cells expressed similar levels of aromatase. Interestingly, the aromatase expression increased when the cells were cultured in suspension suggesting circulating ER+ breast cancer cells may up-regulate intracrine estrogen activity for survival after leaving the estrogen-rich adipose stroma at primary site. To investigate the importance of intracrine estrogen in promoting tumorigenesis and metastasis, we implanted the Cl10 and TT1 cells orthotopically and intracardiacally (I.C.) into female athymic mice. Notably, both cell lines generated orthotopic tumors with no estrogen supplementation after three weeks of inoculation. More interestingly, the mice with I.C. inoculated aromatase-expressing cells presented distant metastasis to bones in mandible and legs after two weeks of inoculation detected by whole mouse fluorescence and bioluminescence imaging as both cells were stably transfected with a luciferase and GFP expression vector. We will determine whether growth of the orthotopic tumors and bone metastases can be inhibited with systemic administration of an aromatase inhibitor and whether bone metastasis is osteoclastic or osteoblastic. Our studies show that a moderate expression of aromatase in breast cancer cells is sufficient to generate intracrine estrogenic action and promote tumorigenesis and skeletal metastasis in ER+ breast cancer cells. These aromatase-expressing models should be useful for future investigation on how ER+ breast cancer cells up-regulate aromatase expression in suspension and induces bone remodeling. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr LB-203.

Null activity of selenium and vitamin e as cancer chemopreventive agents in the rat prostate.

To evaluate the potential efficacy of selenium and vitamin E as inhibitors of prostate carcinogenesis, four chemoprevention studies using a common protocol were done in a rat model of androgen-dependent prostate cancer. After stimulation of prostate epithelial cell proliferation by a sequential regimen of cyproterone acetate followed by testosterone propionate, male Wistar-Unilever rats received a single i.v. injection of N-methyl-N-nitrosourea (MNU) followed by chronic androgen stimulation via subcutaneous implantation of testosterone pellets. At 1 week post-MNU, groups of carcinogen-treated rats (39-44/group) were fed either a basal diet or a basal diet supplemented with l-selenomethionine (3 or 1.5 mg/kg diet; study 1), dl-alpha-tocopherol (vitamin E, 4,000 or 2,000 mg/kg diet; study 2), l-selenomethionine + vitamin E (3 + 2,000 mg/kg diet or 3 + 500 mg/kg diet; study 3), or selenized yeast (target selenium levels of 9 or 3 mg/kg diet; study 4). Each chemoprevention study was terminated at 13 months post-MNU, and prostate cancer incidence was determined by histopathologic evaluation. No statistically significant reductions in prostate cancer incidence were identified in any group receiving dietary supplementation with selenium and/or vitamin E. These data do not support the hypotheses that selenium and vitamin E are potent cancer chemopreventive agents in the prostate, and when considered with the recent clinical data reported in the Selenium and Vitamin E Cancer Prevention Trial (SELECT), show the predictive nature of this animal model for human prostate cancer chemoprevention.

Trough serum testosterone predicts the development of polycythemia in hypogonadal men treated for up to 21 years with subcutaneous testosterone pellets

Testosterone formulations that have more steady-state pharmacokinetics, such as subcutaneously implanted testosterone pellets, may cause less erythrocytosis than i.m. injections of shorter acting androgen esters. We, therefore, sought to define the prevalence, predictors, and proximate basis (role of erythropoietin) for polycythemia (hematocrit >0.50) in hypogonadal men receiving testosterone implants long term. A cross-sectional study was conducted in an academic andrology center with a longitudinal subgroup analysis. A total of 158 hypogonadal men aged 14-84 years (mean age 46.7 years) treated on average for 8 years (range 0-21 years). Trough blood testosterone and hematocrit. Serial serum erythropoietin concentrations were measured in 16 volunteers. Positive univariate associations between polycythemia (hematocrit >0.50) and log(testosterone) (odds ratio (OR) 24.7, 95% confidence interval (CI): 4.3, 141.2, P<0.01) and age (OR 1.1, 95% CI: 1.0, 1.1, P=0.03) and a borderline relationship with current smoking (OR 4.2, 95% CI: 0.9, 20.0, P=0.08) were unveiled. A sensitivity analysis using alternate definitions of polycythemia was performed to capture all potential covariants. Multivariate regression analysis incorporating all potential covariants disclosed the independent OR of developing polycythemia (after adjusting for smoking and age) for log(testosterone) to be 15.0 (95% CI: 2.5, 90.8). Duration of testosterone therapy did not alter the risk of polycythemia. A direct relationship between testosterone and erythropoietin was observed (P=0.05). Higher trough serum testosterone concentrations but not duration of treatment predict the development of polycythemia in men receiving long-acting depot testosterone treatment.

Subcutaneous Testosterone Pellet Implant (Testopel®) Therapy for Men with Testosterone Deficiency Syndrome: A Single‐Site Retrospective Safety Analysis

Long-acting subcutaneous testosterone pellets provide sustained and steady testosterone levels for 3 to 6 months. Testopel subcutaneous crystalline testosterone pellets are U.S.-approved for the treatment of men with testosterone deficiency syndrome. Published experience with testosterone pellets manufactured by Organon has noted relatively high rates of pellet extrusion and infection. To report safety and limited efficacy data from our patients treated for testosterone deficiency syndrome with Testopel subcutaneous testosterone pellets. Infection with or without pellet extrusion, as determined by longitudinal follow-up. Single-site, retrospective analysis of medical records from December 2003 through April 2008. A total of 80 men met inclusion and exclusion criteria. In the 292 implant procedures performed, four adverse events were reported including one implantation site infection. No spontaneous pellet extrusions were reported. Total and free testosterone concentrations were significantly higher at follow-up than at baseline for all patients. Eighty-six percent of patients were satisfied with this treatment modality based on symptom improvement or having subsequent implant procedures. Testosterone replacement with long-acting Testopel pellets had a lower rate of infection (0.3%, 1/292 procedures) as compared with historical data from the Organon testosterone pellet (1.4-6.8%). Additionally, the rate of pellet extrusion was substantially lower (0.3%, 1/292 procedures) as compared with historical data (8.5-12%). None of the patients who complied with post-implant procedure instructions experienced infection or pellet extrusion. Patient satisfaction was high and serum hormone values were improved. The low infection and extrusion rates observed may have been the result of the manufacturing process, which results in small, smooth-surfaced pellets; the absence of foreign material within the pellet packaging; and/or differences in the surgical implantation technique used. Though Testopel pellets have been used in the United States for several decades, more research is needed to document their safety and efficacy.

Effect of testosterone on development of the lumen in seminiferous tubules of the rat.

Although systemically administered testosterone can effect the postnatal maturation of elements in the seminiferous tubule in hypophysectomized rats and mice, it does not elicit the same degree of development which occurs in normal control animals. In view of reports of precocious spermatogenesis in androgen secreting Leydig cell tumors, the present study was designed to determine if high local levels of testosterone accelerate development of the seminiferous tubules. Testosterone pellets were inserted under the tunica albuginea of the right testis of 7 day old rats. At 17, 23 and 28 days of age development of the seminiferous tubules, as judged by the formation of tubule lumens, was more extensive in the treated testes than in contralateral and sham operated controls. Tubule diameters were not necessarily correlated with lumen formation. This study demonstrates that high local levels of testosterone accelerate seminiferous tubule development in the rat and indicates that tubule diameter may not be a valid basis for estimating development of the testis. It is suggested that testosterone exerts this effect through its actions on the Sertoli cells.

SUBCUTANEOUS TESTOSTERONE PELLET IMPLANT THERAPY IN THE UNITED STATES FOR MEN WITH TESTOSTERONE DEFICIENCY SYNDROME: A SINGLE-SITE RETROSPECTIVE SAFETY AND EFFICACY STUDY

INTRODUCTION AND OBJECTIVES: Long-acting subcutaneous testosterone pellets provide sustained and steady testosterone levels 3-6 months. Testopel® subcutaneous crystalline testosterone pellets (Slate) are US Food and Drug Administration (FDA) approved for the treatment of men with testosterone deficiency syndrome. Previous experience with other subcutaneous testosterone pellets (Organon, not approved in the US) has noted a relatively high rate of pellet extrusion and infection (8.5%). Our objective was to report safety and efficacy data from US patients with testosterone deficiency syndrome treated with subcutaneous testosterone pellets. METHODS: A US, single-site, retrospective analysis of patients seen in our clinic between December 2003 and April 2008. RESULTS: A total of 166 men presented with testosterone deficiency syndrome and 80 (48%) met inclusion and exclusion criteria in this 4.5 year period. Eighty-six percent were satisfied and went on to have a second or more implant procedure(s). The mean time between implant procedures was 174 days (5.7 months; range, 45-689 days). Of the 292 subcutaneous implant procedures available for analysis, there were no reports of infections and extrusions. Total and free testosterone concentrations were significantly higher at follow-up (mean, 58 days; range, 13 120 days) than at baseline for all patients. CONCLUSIONS: In this single site retrospective study, longacting subcutaneous testosterone pellets have a significantly lower rate of extrusion and infection compared with historical data (8.5%) from the only other subcutaneous testosterone pellet implant known to be approved by a major regulatory agency. This may be due to the unique manufacturing process for the US-approved pellets resulting in a smaller, more regular pellet surface that is not packaged in wool or cotton compared to the Organon product. Patient satisfaction was high and blood tests confirmed improved hormone values. More research is needed concerning the safety and efficacy of long-acting subcutaneous testosterone pellets.

Testosterone treatment in elderly men

Testosterone has been used in testicular and hypothalamo-pituitary diseases since the 1940s. There is growing interest in the use of testosterone in aging men, and this has stimulated research into the benefits of male hormone replacement. Testosterone treatment of men with hypogonadism might have beneficial effects on body composition, muscle strength, sexual function, and cognition. There are several modes of administration of the male hormone, with injectable testosterone esters and implanted testosterone pellets being the mainstay of treatment until recently. These preparations are increasingly being replaced by transdermal patches, gels, and long-acting parenteral preparations. Testosterone patches and gels are ideally for elderly men. Treatment with the male hormone is relatively safe, if patients are selected appropriately and monitored carefully. The most important adverse effects are on the prostate. In this review, we briefly discuss the indications, contraindications, and benefits of testosterone treatment. Further, we list the adverse effects, advantages, and disadvantages of various testosterone preparations in elderly men.

Time course of changes in Gambel's white-crowned sparrow song behavior following transitions in breeding condition

Seasonal changes in behavior and in its underlying neural substrate are common across animal taxa. These changes are often triggered by steroid sex hormones. Song in seasonally breeding songbirds provides an excellent example of this phenomenon. In these species, dramatic seasonal changes mediated by testosterone and its metabolites occur in adult song behavior and in the neural circuitry controlling song. While song rate can quickly change in response to seasonal breeding cues, it is unknown how quickly other aspects of song change, particularly the stereotypy of song phonology and syntax. In this study we determined whether and how quickly song rate, phonology, and syntax change in response to breeding and non-breeding physiological cues. We asked these questions using Gambel's white-crowned sparrows ( Zonotrichia leucophrys gambelii), a closed-ended learner with well-characterized changes in the neural circuitry controlling song behavior. We exposed ten photosensitive sparrows to long-day photoperiod and implanted them with subcutaneous testosterone pellets (day 0) to simulate breeding conditions. We continuously recorded song and found that song rate increased quickly, reaching maximum around day 6. The stereotypy of song phonology changed more slowly, reaching maximum by day 10 or later. Song syntax changed minimally after day 6, the earliest time point examined. After 21 days, we transitioned five birds from breeding to non-breeding condition. Song rate declined precipitously. These results suggest that while song rate changes quickly, song phonology changes more slowly, generally following or in parallel with previously investigated changes in the neural substrate.

Differential Regulation of Aortic Growth in Male and Female Rodents Is Associated With AAA Development

The objective was to examine effects of gonadal hormone manipulation on aortic diameter and macrophage infiltration in rodents during abdominal aortic aneurysm (AAA) formation. Experiment 1: 17-beta estradiol and testosterone pellets were implanted in male (ME) and female (FT) rats. No pellet was implanted in shams (MES, FTS). Experiment 2: Testes and ovaries were removed from males (MO) and females (FO), respectively. No organs were removed from shams (MOS, FOS). Experiment 3: Male and female rats were orchiectomized and oophorectomized, respectively. Four weeks post-castration, testosterone (MOT) and 17-beta estradiol (FOE) pellets were implanted. Shams underwent castration, but no pellet was implanted (MOTS, FOES). All rats underwent infrarenal aortic infusion with elastase postimplantation/postcastration. Diameters were measured on postoperative d 14. Tissue was stained for macrophages by immunohistochemistry. Diameter (P = 0.046) and macrophage counts (P = 0.014) decreased in ME compared with shams, but not in females treated with testosterone (FT). Diameter (P = 0.019) and macrophage infiltration (P = 0.024) decreased in MO compared with shams, but not in FO. Diameter increased in MOT compared with MOTS (P = 0.033), but decreased in FOE compared with FOES (P = 0.002). Macrophages decreased in FOE compared with FOES (P = 0.002). This study documents a decrease in AAA diameter in males treated with estrogen or undergoing orchiectomy, but no changes in females treated with testosterone or undergoing oophorectomy; and an increase in diameter in MOT and a decrease in FOE. These data suggest that gonadal hormones differentially regulate AAA growth in association with changes in macrophages.

Dietary n-3 Polyunsaturated Fatty Acids Enhance Hormone Ablation Therapy in Androgen-Dependent Prostate Cancer

Hormone ablation therapy typically causes regression of prostate cancer and represents an important means of treating this disease, particularly after metastasis. However, hormone therapy inevitably loses its effectiveness as tumors become androgen-independent, and this conversion often leads to death because of subsequent poor responses to other forms of treatment. Because environmental factors, such as diet, have been strongly linked to prostate cancer, we examined the affects of dietary polyunsaturated fatty acids (PUFAs; at 1.5 wt%) on growth of androgen-dependent (CWR22) and androgen-independent (CWR22R) human prostatic cancer xenografts, the acute response of CWR22 tumors to ablation therapy, and their progression to androgen independence. Significant diet-induced changes in tumor n-3 or n-6 PUFA content had no affect on CWR22 or CWR22R tumors growing with or without androgen support, respectively. However, dietary changes that increased tumor eicosapentaenoic acid and linoleic acid content enhanced responses to ablation therapy, measured by cancer cell apoptosis and mitosis. In addition, relapse to androgen-independent growth (measured by renewed increases in tumor volume and serum prostate-specific antigen after ablation) positively correlated with tumor arachidonic acid content. There was no correlation between expression of 15-lipoxygenase isozymes or their products and tumor growth or responses to ablation. In conclusion, dietary n-3 PUFA may enhance the response of prostate cancer to ablation therapy and retard progression to androgen-independent growth by altering tumor PUFA content.