Hormone Ablation Research Articles

C-Met/miR-130b axis as novel mechanism and biomarker for castration resistance state acquisition.

Although a significant subset of prostate tumors remain indolent during the entire life, the advanced forms are still one of the leading cause of cancer-related death. There are not reliable markers distinguishing indolent from aggressive forms. Here we highlighted a new molecular circuitry involving microRNA and coding genes promoting cancer progression and castration resistance. Our preclinical and clinical data demonstrated that c-Met activation increases miR-130b levels, inhibits androgen receptor expression, promotes cancer spreading and resistance to hormone ablation therapy. The relevance of these findings was confirmed on patients' samples and by in silico analysis on an independent patient cohort from Taylor's platform. Data suggest c-Met/miR-130b axis as a new prognostic marker for patients' risk assessment and as an indicator of therapy resistance. Our results propose new biomarkers for therapy decision-making in all phases of the pathology. Data may help identify high-risk patients to be treated with adjuvant therapy together with alternative cure for castration-resistant forms while facilitating the identification of possible patients candidates for anti-Met therapy. In addition, we demonstrated that it is possible to evaluate Met/miR-130b axis expression in exosomes isolated from peripheral blood of surgery candidates and advanced patients offering a new non-invasive tool for active surveillance and therapy monitoring.

Association of Cytokines and Chemokines in Pathogenesis of Breast Cancer.

Breast cancer touches women's life worldwide. Expected outcome is not achieved due to molecular heterogeneity and complex biology despite substantial advancement in diagnosis, prevention and treatment of breast cancer. Patients with estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2 (Her2) positive tumors receive hormone ablation and Her2 directed therapy. While patients diagnosed with triple-negative breast cancer receive chemotherapy in both the early and advanced stages. However, chemotherapeutic efficacies are not the same in every patient, which has fostered a major effort to identify new targets to treat breast cancer. Positive therapeutic outcome after immune checkpoint inhibitors emphasizes the significance of the host immune system in breast cancer. Cancer develops in immune competent host wherein cytokines, while shaping the immune system, also serve as growth signals for cancer cells. The dynamics of cross talk between immune system and cancer cells mediated by cytokines and chemokines changes during cancer initiation, progression, and therapeutic interventions. Hence, better understanding of molecular footprint of cancer cells, as well as crosstalk between cancer cells and host immune system is needed to develop patient specific treatment and management of breast cancer.

Progesterone induces progesterone receptor gene (PGR) expression via rapid activation of protein kinase pathways required for cooperative estrogen receptor alpha (ER) and progesterone receptor (PR) genomic action at ER/PR target genes

Progesterone Receptors (PRs) are critical effectors of estrogen receptor (ER) signaling required for mammary gland development and reproductive proficiency. In breast and reproductive tract malignancies, PR expression is a clinical prognostic marker of ER action. While estrogens primarily regulate PR expression, other factors likely contribute to a dynamic range of receptor expression across diverse tissues. In this study, we identified estrogen-independent but progestin (R5020)-dependent regulation of ER target genes including PGR in ER+/PR+ cancer cell lines. R5020 (10nM–10μM range) induced dose-dependent PR mRNA and protein expression in the absence of estrogen but required both PR and ERα. Antagonists of either PR (RU486, onapristone) or ERα (ICI 182,780) attenuated R5020 induction of TFF1, CTSD, and PGR. Chromatin immunoprecipitation (ChIP) assays performed on ER+/PR+ cells demonstrated that both ERα and PR were recruited to the same ERE/Sp1 site-containing region of the PGR proximal promoter in response to high dose progestin (10μM). Recruitment of ERα and PR to chromatin and subsequent PR mRNA induction were dependent upon rapid activation of MAPK/ERK and AKT; inhibition of these kinase pathways via U0126 or LY294002 blocked these events. Overall, we have identified a novel mechanism of ERα activation initiated by rapid PR-dependent kinase pathway activation and associated with phosphorylation of ERα Ser118 for estrogen-independent but progestin-dependent ER/PR cross talk. These studies may provide insight into mechanisms of persistent ER-target gene expression during periods of hormone (i.e. estrogen) ablation and suggest caution following prolonged treatment with aromatase or CYP17 inhibitors (i.e. contexts when progesterone levels may be abnormally elevated).

Abstract 4340: DNA repair genes aberrations in germline DNA in metastatic castration-resistant prostate cancer patients

Abstract INTRODUCTION: DNA repair defects are found in mCRPC and are therapeutically actionable; germline BRCA mutation-associated (gBRCA) prostate cancer has a poor prognosis. We hypothesized that metastatic castration resistant prostate cancer (mCRPC) is enriched for germline DNA repair mutations and that these may be relevant to patient outcome. METHODS: Targeted-sequencing for DNA repair genes was conducted in germline DNA from patients consenting to 3 clinical trials between 2013-2015. Germline DNA was extracted from saliva or buccal swabs using the Oragene kit; libraries were constructed using a customized Qiagen panel and sequenced using the Illumina MiSeq. Family history and clinical data were prospectively collected. For time to event analyses unadjusted Cox regression models were used and comparisons were made using log-rank tests. RESULTS: Germline samples from 154 mCRPC patients were available. Median age at diagnosis was 61years (y), median time to castration-resistance was 14.5 months (m) and median overall survival (OS) from initial diagnosis of prostate cancer was 106.8m; 69% (91/131; 24 N/A) of patients were initially diagnosed with Gleason≥8 tumors. 130/154 (84.4%) and 131/154 (85.0%) received Docetaxel and Abiraterone respectively. Of 154 patients, 4 were previously known to be gBRCA2 mutation carriers and were removed from the prevalence analysis but included in the clinical analyses; 22/150 (14.7%, 95%CI 9.4-21.4%) harboured a truncating or frameshift mutation in a DNA repair gene (9 BRCA2, 6%; 4 ATM, 2.7%; 2 PALB2, 1.3%, 1 each for CHEK2, FANCI, MRE11A, NBN, RAD51C, RAD51D and MSH6). Overall, patients with any germline DNA repair aberrations had a worse median OS (75.8 vs 106.8 m; log-rank p = 0.04). Time to resistance to primary hormonal ablation was shorter specifically for gBRCA2 mutations carriers (11.0 vs 14.8 m; log-rank p = 0.01) but not for non-BRCA2 repair aberrations. Age at diagnosis was similar in patients with or without DNA repair germline mutations (median 61.3 vs 61.7y, Mann-Whitney p = 0.41) as well as frequency of Gleason≥8 tumors (16/21 [76%] vs 75/109 [68%]; Mann-Whitney p = 0.23) Response rates to Docetaxel (14/18 [77.8%] vs 64/94 [68.1%]; Fisher exact p = 0.67) and Abiraterone (10/21 [47.6%] vs 44/94 [46.8%]; Fisher exact p = 0.73) were similar among individuals with and without mutations. Family cancer history was collected in 125/154 cases (81%). While having cases of ovarian/prostate/breast/pancreas cancers in these patients’ families associated with a higher likelihood of finding a germline mutation (Odds Ratio 3.36, p = 0.03), 5 of 68 (7.4%) men with no cases of ovarian/prostate/breast/pancreas cancers registered in their families carried a germline mutation in a DNA repair gene. CONCLUSIONS: mCRPC is enriched for patients with germline mutations in DNA repair genes (15%), with 6% having gBRCA2 mutation. Germline DNA repair aberrations are associated with a worse prognosis from mCRPC. Citation Format: JOAQUIN MATEO, Suzanne Carreira, Helen Mossop, Pasquale Rescigno, Michael Kolinsky, Elena Castro, Ada Balasopoulou, Jo Hunt, Desamparados Roda, Claudia Bertan, Jane Goodall, Susana Miranda, Penny Flohr, Nuria Porta, Zsofia Kote-Jarai, David Olmos, Christopher J. Lord, Emma Hall, Ros Eeles, Johann S. de Bono. DNA repair genes aberrations in germline DNA in metastatic castration-resistant prostate cancer patients. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4340.

Stimulation of cellular senescent processes, including secretory phenotypes and anti-oxidant responses, after androgen deprivation therapy in human prostate cancer

Endocrine resistance is a major problem in prostate cancer. Recent studies suggest that cellular plasticity plays a key role in therapy resistance. Yet little is known about the cellular changes of human prostate cancer after androgen deprivation therapy (ADT). In this study, we investigated cellular senescence, senescence-associated secretory phenotypes (SASPs), and anti-oxidant responses. Hormone ablation upregulated senescence-associated (SA)-β-Gal activity in prostate glands, as well as the expressions of p27KIP1 and p53, in a mouse castration model. In line with this, the expressions of p21CIP1 and p27KIP1 were significantly more upregulated in human non-pathological prostatic glands after ADT than in untreated specimens. In a study of SASP markers, the expressions of IL6 and IL8 were also more upregulated in human non-pathological prostatic glands after ADT than in untreated specimens. IL6, IL8, and MMP2 were expressed more strongly in human prostate cancer specimens resected after ADT than in untreated tumors. Of note, treatment with the anti-oxidant reagent NAC significantly suppressed SA-β-Gal activity in androgen-sensitive human prostate cancer LNCaP cells. In immunohistochemical analyses on anti-oxidant response genes, NRF2 and NQO1 were more upregulated after hormone ablation in human prostate gland and carcinoma specimens after ADT than in untreated specimens or in murine prostate glands after castration. Taken together, these findings suggest that ADT induces cellular senescence processes accompanied by secretory phenotypes and anti-oxidant responses in prostate. These cellular changes may be attractive targets for preventing endocrine resistance in prostate cancer.

Prolactin, EGFR, vimentin and α-actin profiles in elderly rat prostate subjected to steroid hormonal imbalance.

The aim of this study was to characterize and relate the prolactin (PR), epidermal growth factor receptor (EGFR), α-actin and vimentin immunoreactivity in the prostate of elderly rats subjected to steroid hormonal imbalance. Senile and young rats were divided into the young group (YNG), the senile group (SE), the castrated group (CAS), the estrogen-deficient group (ED), the castrated+estrogen group (CASE), and the estrogen-deficient+androgen group (EDTEST). PR and EGFR increased in the estrogen and androgen ablation groups. In addition, EGFR influenced the immunolocalization by changing it from the prostatic stroma to the epithelium in elderly rats. Hormone ablation in elderly rats, not only related to androgen but also estrogen, led to increased stromal EGFR immunolocalization. The α-actin pattern decreased in the groups with estrogenic imbalance. Moreover, vimentin increased in the senile and estrogen deficient group. To conclude, we can suggest that EGFR contributed towards the proliferative process in the prostate, by means however, of different mechanisms, considering the androgenic and estrogenic pathways. Also, our results indicated that prolactin could be activated not only in an androgen-independent pathway but also in an estrogen independent pathway. Finally, PR and vimentin immunolocalization increase, in the prostatic stroma in the group showing estrogenic ablation, could be one of the factors which contribute to the reactive stroma formation.

Acknowledging unreported problems with active surveillance for prostate cancer: a prospective single-centre observational study

ObjectiveTo report outcomes of patients with localised prostate cancer (PCa) managed with active surveillance (AS) in a standard clinical setting.DesignSingle-centre, prospective, observational study.SettingNon-academic, average-size hospital in Switzerland.ParticipantsProspective, observational study at...

Medical Help-Seeking for Sexual Concerns in Prostate Cancer Survivors.

IntroductionAlthough sexual dysfunction is common after prostate cancer, men's decisions to seek help for sexual concerns are not well understood.AimDescribe predictors of actual prior help-seeking and intended future medical help-seeking for sexual dysfunction in prostate cancer survivors.MethodsA cross-sectional survey of 510 prostate cancer survivors assessed masculine beliefs, attitudes, support/approval from partner/peer networks (subjective norm), and perceived control as predictors of medical help-seeking for sexual concerns. A theory of planned behavior (TPB) perspective was used to examine actual prior and planned future behavior and contributing factors. Statistical analyses included multiple and logistic regressions.Main Outcome MeasuresIntention to see a doctor for sexual advice or help in the next 6 months was measured using the intention subscale adapted from the Attitudes to Seeking Help after Cancer Scale. Prior help-seeking was measured with a dichotomous yes/no scale created for the study.ResultsMen were Mage 71.69 years (SD = 7.71); 7.54 years (SD = 4.68) post-diagnosis; received treatment(s) (58.1% radical prostatectomy; 47.1% radiation therapy; 29.4% hormonal ablation); 81.4% reported severe ED (IIED 0–6) and 18.6% moderate–mild ED (IIED 7–24). Overall, 30% had sought sexual help in the past 6 months, and 24% intended to seek help in the following 6 months. Prior help-seeking was less frequent among men with severe ED. Sexual help-seeking intentions were associated with lower education, prior sexual help-seeking, sexual importance/ priority, emotional self-reliance, positive attitude, and subjective norm (R2 = 0.56).ConclusionThe TPB has utility as a theoretical framework to understand prostate cancer survivors' sexual help-seeking decisions and may inform development of more effective interventions. Masculine beliefs were highly salient. Men who were more emotionally self-reliant and attributed greater importance to sex formed stronger help-seeking intentions. Subjective norm contributed most strongly to help-seeking intentions suggesting that health professionals/partners/peers have a key role as support mechanisms and components of psycho-sexual interventions.

Effect of FSH supplementation on the GnRH antagonist suppressed growth of PC-3 cell xenografts in nude mice.

e630 Background: One of the fundamental differences between GnRH agonist and antagonist treatment is the permanent suppression of FSH by antagonist, while a rebound in FSH secretion has been documented upon agonist treatments. To assess the potential beneficial effects of permanent FSH suppression in the hormone ablation treatment of prostate cancer we studied the effect of the GnRH antagonist degarelix in the absence and presence of FSH supplementation on the growth of PC-3 human prostate cancer cell xenografts in nude mice. Methods: Intact (n = 20) and gonadectomized (n = 20) nude male mice were inoculated with 2 x 106PC-3 cells. Half of each group received degarelix treatment at 10 mg/kg body weight s.c. In a second experiment (n = 10/group), degarelix treatment was supplemented with recombinant human FSH at10 IU/kg/day using i.p. ALZET osmotic minipumps. Tumor growth in both experiments was monitored for 4 weeks by external inspection and caliper measurement, after which the mice were sacrificed. Results: The first experiment demonstrated that degarelix treatment significantly reduced tumor growth by 33% and 35%, respectively, in intact and gonadectomized mice as compared to non-treated controls (p<0.0001 for each). In the second experiment, FSH reversed the inhibitory effect of degarelix in intact mice. Conclusions: These results demonstrated that the suppression of PC-3 xenograft growth by GnRH antagonist treatment was completely inhibited by concomitant FSH treatment. The findings prove experimentally the principle that combined FSH and LH suppression by GnRH antagonist could offer an advantage over the isolated LH suppression by GnRH agonist upon prostate cancer treatment. [Table: see text]

Quantifying the potential of C-11 acetate PET scanning in avanced prostate cancer patients.

300 Background: Detection of advanced prostate cancer has historically relied on radionuclide bone scanning and cross-sectional imaging with varying degrees of sensitivity and specificity based on the clinical scenario. In the current era, use of NaF and Choline based whole body scans have led to increased detection, but access to these scan can be limited and expense can be prohibitive. Similar to choline based scanning, acetate is metabolically active in cancer, including prostate. Methods: Fifty-six men previously diagnosed with prostate cancer with PSA recurrences after some failed primary treatment, have undergone whole body C-11 Acetate PET scans at Indiana University since June of 2013. Patient demographics, prior treatments, PSA values, prior imaging, post-scan therapies, and therapeutic outcomes have been recorded prospectively and updated as follow-up information becomes available. Results: Analysis of these initial 56 men demonstrated 31/56 men had a C-11 Acetate PET scan that lead to further definitive loco-regional treatments, including salvage radiation therapy ( 12), salvage regional lymph node dissection (11) and salvage cryo-ablation (8), while 9/56 men had initiation of hormonal ablation, secondary to detection of systemic disease. The remainder of these men had negative scans and are currently under continued active surveillance without evidence of disease progression requiring either loco-regional or systemic therapy. Closer analysis of the 31 men who received loco-regional treatments revealed 10 men who benefited with clear PSA declines, 4 men who failed to demonstrate a benefit from the definitive loco-regional therapy and 17 of men are too early to determine at the time of analysis. Careful pathological analysis of the regional lymph dissections revealed the limit of detection of the C-11 Acetate PET scan of greater than or equal to 3mm. Correlations of SUV readings of detected prostate cancer lesions with PSA kinetics, suggest importance of both PSA doubling time and absolute PSA value cutoffs. Conclusions: The C-11 Acetate PET scan is a useful adjunct to current imagining techniques and provides additional clinical information to guide clinicians caring for prostate cancer patients.

Abstract 4057: Gender and the immune system in lung and liver colonization of murine melanoma in a B16 metastatic model

Abstract Introduction: Until age 45, incidence rates of melanoma are higher in woman than men, but by age 60, rates in men are more than double and by age 80, they are almost triple. With the low rate of survival for distant metastatic patients with melanoma, especially in men, studies aimed at elucidating the mechanisms underlying metastasis and the gender disparity are warranted. The purpose of this study is to identify the immune cells and cellular interactions that are crucial for initial colonization of metastatic melanoma in a gender dependent manner. Experimental Procedures: We used a B16 melanoma metastatic model in which 2×105 cells are injected via the tail vein of B6 mice and allowed to colonize for 14 days. Positive organs are analyzed by visual counting of the metastases, extraction of the melanin pigment and measurement by spectrophotometry, and by flow cytometry. To investigate the role of innate immunity, we injected ip either anti-Ly6G, 1A8 (neutrophil depletion), anti NK1.1, PK136 (natural killer [NK] cell depletion) or control IgG every 3 days, starting 1 day before tumor injection. For hormone ablation studies, an LHRH antagonist was injected. Data: Naïve mice injected with B16 cells revealed that females have higher lung tumor burden than males. Previous castration studies indicate that this may be dependent on androgens. Intriguingly, neutrophil depletion increases tumor burden only in male mice when compared to male IgG and does not significantly affect tumor burden in female mice, indicating a potential gender difference in neutrophils. Neutrophil depletion in male mice also reduced NK activation and IFN-γ production, indicating a stimulatory relationship between neutrophils and NK cells. A subsequent NK depletion study revealed significantly increased tumor burden in both male and female NK-depleted mice when compared to the gender-matched controls. However, the increased tumor burden normally seen in lungs of females compared to males was still apparent when comparing lungs of NK depleted male mice to NK depleted female mice. This study also revealed that NK cells are crucial for the prevention of liver metastasis as both genders of NK depleted mice revealed multiple melanoma tumor nodules on the surface of the liver. Conclusion: Our data indicates that both neutrophils, and to a greater extent, NK cells are important for the initial response against B16 melanoma colonization of the lung and may be necessary to prevent metastasis. For the liver, NK cells were absolutely crucial for the prevention of melanoma colonization in this organ. Neutrophil depletion, as well as our previous castration data, indicated that there is a gender specific differential response of these cells, which may contribute to the gender disparity seen in human incidence and survival. Further, the importance of NK cells in killing of tumor cells indicates a population of patients that may benefit from ex vivo NK expansion therapy currently in clinical trials. Citation Format: Janet L. Markman, Daiko Wakita, Timothy R. Crother, Moshe Arditi. Gender and the immune system in lung and liver colonization of murine melanoma in a B16 metastatic model. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4057. doi:10.1158/1538-7445.AM2015-4057

Reactive stroma in the prostate during late life: The role of microvasculature and antiangiogenic therapy influences.

Prostate cancer is associated to a reactive stroma microenvironment characterized by angiogenic processes that are favorable for tumor progression. Senescence has been identified as a predisposing factor for prostate malignancies. In turn, the relationships between aging, reactive stroma, and the mechanisms that induce this phenotype are largely unknown. Thus, we investigated the occurrence of reactive stroma in the mouse prostate during advanced age as well as the effects of antiangiogenic and androgen ablation therapies on reactive stroma recruitment. Male mice (52-week-old FVB) were treated with two classes of angiogenesis inhibitors: direct (TNP-470; 15 mg/kg; s.c.) and/or indirect (SU5416; 6 mg/kg; i.p.). Androgen ablation was carried out by finasteride administration (20 mg/kg; s.c.), alone or in association to both inhibitors. The Transgenic Adenocarcinoma of the Mouse Prostate (TRAMP) model was used as a paradigm of cancer-associated reactive stroma. The dorsolateral prostate was collected for α-actin (αSMA), vimentin (VIM), and transforming growth factor-beta (TGF-β) immunohistochemical and Western blotting analyses as well as for CD34/αSMA and CD34/VIM colocalization. Senescence was associated with increased αSMA, VIM, and TGF-β expression as well as with the recruitment of CD34/αSMA and CD34/VIM dual-positive fibroblasts. These observations were similar to those verified in TRAMP mice. Antiangiogenic treatment promoted the recovery of senescence-associated stromal changes. Hormonal ablation, despite having led to impaired CD34/αSMA and CD34/VIM dual-positive cell recruitment, did not result in decreased stimulus to reactive stroma development, due to enhanced TGF-β expression in relation to the aged controls. Reactive stroma develops in the prostate of non-transgenic mice as a result of aging. The periacinar microvasculature is a candidate source for the recruitment of reactive stroma-associated cells, which may be derived either from perivascular-resident mesenchymal stem cells (MSCs) or from an endothelial-to-mesenchymal transition (EndMT) process. Thus, antiangiogenic therapy is a promising approach for preventing age-associated prostate malignancies by means of its negative interference in the development of reactive stroma phenotype from the vascular wall.

Comparison of the effects of pretreatment with Veramix sponge (medroxyprogesterone acetate) or CIDR (natural progesterone) in combination with an injection of estradiol-17β on ovarian activity, endocrine profiles, and embryo yields in cyclic ewes superovulated in the multiple-dose Folltropin-V (porcine FSH) regimen

Follicular wave status at the beginning of exogenous FSH administration is an important contributor to variability in superovulatory responses in ruminants. Studies in ewes have shown a decrease in the number of ovulations when superovulation is initiated in the presence of ostensibly ovulatory-sized ovarian follicles. Hormonal ablation of large antral follicles with the progestin–estradiol (E2-17β) treatment significantly reduces this variability in superovulated anestrous ewes, but the effects of the treatment in cycling ewes have not yet been assessed. Sixteen Rideau Arcott × Polled Dorset ewes (November–December) received either medroxyprogesterone acetate (MAP)–releasing intravaginal sponges (60 mg) or controlled internal drug release (CIDR) devices (containing 300 mg of natural progesterone) for 14 days (Days 0–14), with a single intramuscular injection of 350 μg of E2-17β on Day 6. The superovulatory treatment consisted of six injections of porcine FSH (Folltropin-V) given twice daily, followed by a bolus GnRH injection (50 μg intramuscular) on Day 15. There were no differences (P < 0.05) in the ovulatory responses and embryo yields between the two groups of ewes. In both subsets of animals, the next follicular wave emerged ∼2.5 days after an E2-17β injection (P > 0.05). A decline in maximum follicle size after an E2-17β injection was more abrupt in CIDR- compared with MAP-treated animals, and the ewes pretreated with exogenous progesterone had significantly more 3-mm follicles at the start of the superovulatory treatment. The metabolic clearance rate of exogenous E2-17β appeared to be greater in MAP-treated ewes, but circulating concentrations of porcine FSH failed to increase significantly after each Folltropin-V injection in CIDR-treated animals. The CIDR-treated ewes exceeded (P < 0.05) their MAP-treated counterparts in serum E2-17β concentrations during superovulation. In spite of differences in antral follicle numbers and endocrine profiles between MAP- and CIDR-treated cyclic ewes receiving E2-17β before ovarian superstimulation, there were no differences in superovulatory responses.

Abstract B15: Bone marrow stromal cell-secreted inflammatory cytokines promote treatment resistance and survival of prostate cancer cells

Abstract It is well established that prostate cancer (PCa) cells hone to, adapt, and thrive in the bone. Importantly, bone metastases are found in over 80% of PCa deaths. Therefore, it is imperative to elucidate and characterize the molecular mechanisms that enable PCa cells to thrive in the bone and resist conventional PCa therapies, such as hormone ablation or anti-androgens. We found that the HS-5 bone marrow stromal cell (BMSC) line, a secretory BMSC line that can support hematopoietic stem cell growth, secretes paracrine factors that induce apoptosis in PCa cells. However a subpopulation of the PCa cells can survive the BMSC-induced death. The surviving PCa subpopulation differentiates into a neuronal morphology and looses androgen receptor (AR) expression, reminiscent of treatment-resistant neuroendocrine PCa. Ideally, the loss of AR would induce PCa cell death; however we observe a concomitant increase in the autophagy-related cell survival protein, p62/SQSTM1, and induction of cytoprotective autophagy. Furthermore, we find that interleukin-1 beta (IL-1β) and IL-6, inflammatory cytokine secreted by HS-5 BMSCs, are sufficient to promote PCa neuronal morphology, reduce AR accumulation, upregulate p62/SQSTM1 levels, and induce autophagy. In addition, genetic silencing of p62/SQSTM1 causes PCa cell death and pharmacological inhibition of cytokine or autophagy signaling attenuates PCa neuroendocrine differentiation. Thus, our PCa-bone marrow stromal cell model suggests that PCa cells can co-opt the immune system in the bone microenvironment leading to PCa androgen independence, treatment resistance, and survival in the bone. Furthermore, our data suggests that inhibiting inflammatory cytokine signaling and/or the autophagy process are rational strategies to combat PCa bone metastasis. Citation Format: Megan Chang, Micaela Morgado, Viral Patel, Michael Gwede, Mary Cindy Farach-Carson, Nikki Delk. Bone marrow stromal cell-secreted inflammatory cytokines promote treatment resistance and survival of prostate cancer cells. [abstract]. In: Abstracts: AACR Special Conference on Cellular Heterogeneity in the Tumor Microenvironment; 2014 Feb 26-Mar 1; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(1 Suppl):Abstract nr B15. doi:10.1158/1538-7445.CHTME14-B15

Zoledronic acid induces an immune response through increased central memory and effector memory gamma/delta T cells in early and metastatic breast cancer patients

Introduction: Zoledronic acid (ZA) in combination with endocrine therapy (ET) and ovarian ablation yielded a disease-free survival and overall survival advantage in women with estrogen receptor+ early stage breast cancer (EBC). Evidence from preclinical studies suggests that ZA increases gamma/delta T cells (GDT). Materials and Methods: We examined immune responses following ZA in 24 BC patients by collecting peripheral blood mononuclear cells at day 0, 1, 7, and 28. GDT population and cytokine responses were assayed using flow cytometry and multi-analyte profiling beads (Luminex), respectively, and relative changes from baseline were analyzed using the Wilcoxon signed-rank test. Results: In total, 18 (75%) patients had metastatic breast cancer (MBC), 6 (25%) had EBC, 18 (75%) received ET, 4 (17%) chemotherapy (C), and 2 (8%) no concurrent therapy. Following ZA, an increase in both effector (CD3+/Vdelta2+/CD45RA−/CD27−) ( P = 0.0005) and central memory GDT (CD3+/Vdelta2+/CD45RA−/CD27+) ( P = 0.006), as well as a decrease in naive GDT (CD3+/Vdelta2+/CD45RA+/CD27+) ( P = 0.003) were observed at day 7. Cytokines, including interleukin-1 (IL-1) receptor antagonist ( P P P P P Conclusion: In both EBC and MBC patients, ZA appears to induce a significant change in GDTs and cytokines associated with cell-mediated immunity offering a possible biologic mechanism for ZA's anticancer activity. Further studies are necessary to determine which BC patients might achieve clinically meaningful anticancer benefits from ZA.

Could Denosumab Support Bone Integrity in Sickle Cell Disease

Sickle cell anaemia is a congenital disease characterized by painful vaso-occlusive crises. It is associated with vitamin D deficiency, osteoporosis and recurrent attacks of micro-infarctions lead to bone osteonecrosis. The most common sites of osteonecrosis in sickle cell disease are the femoral and humeral heads which could lead to severe pain and extensive physical disability. Denosumab is a new human monoclonal antibody product; it is a receptor activator of nuclear factor kappa B ligand RANKL. That decreases osteoclastic bone restoration. It is Food and Drug Administration FDA approved for osteoporosis for women with high risk of fracture; and it is also being used for the treatment and prevention of bone loss in patients undergoing hormone ablation therapy for breast and prostate cancer. Denosumab is not restricted for sickle cell disease patients within its approved indications. It has less nephrotoxicity effect than the bisphosphonates which is a big advantage for sickle cell patients. It also used as an adjuvant agent to reduce metastatic bone pain in solid tumors. The extension of Denosumab use for sickle cell patients’ osteoporosis and osteonecrosis deserve a randomized multi central trail aiming to improve bone integrity and possible reduction of the devastating pain in sickle cell disease patients.

Targeting bone metabolism in patients with advanced prostate cancer: current options and controversies.

Maintaining bone health remains a clinical challenge in patients with prostate cancer (PC) who are at risk of developing metastatic bone disease and increased bone loss due to hormone ablation therapy. In patients with cancer-treatment induced bone loss (CTIBL), antiresorptive agents have been shown to improve bone mineral density (BMD) and to reduce the risk of fractures. For patients with bone metastases, both zoledronic acid and denosumab delay skeletal related events (SREs) in the castration resistant stage of disease. Novel agents targeting the Wnt inhibitors dickkopf-1 and sclerostin are currently under investigation for the treatment of osteoporosis and malignant bone disease. New antineoplastic drugs such as abiraterone, enzalutamide, and Radium-223 are capable of further delaying SREs in patients with advanced PC. The benefit of antiresorptive treatment for patients with castration sensitive PC appears to be limited. Recent trials on the use of zoledronic acid for the prevention of bone metastases failed to be successful, whereas denosumab delayed the occurrence of bone metastases by a median of 4.1 months. Currently, the use of antiresorptive drugs to prevent bone metastases still remains a field of controversies and further trials are needed to identify patient subgroups that may profit from early therapy.

Salvage focal prostate cryoablation for locally recurrent prostate cancer after radiotherapy: Initial results from the cryo on-line data registry

Several investigators have tried to apply salvage focal prostate cryoablation to small numbers of patients with biopsy-proven unilateral recurrent prostate cancer (PCa) after radiotherapy with the aim of decreasing complications of salvage cryoablation. We report contemporary outcomes of salvage focal cryoablation for locally recurrent PCa after radiotherapy within the Cryo On-Line Data (COLD) Registry. We queried the COLD Registry to identify patients diagnosed as locally recurrent PCa after radiotherapy and treated with salvage focal cryoablation. Patients with hormone ablation after cryotherapy were excluded. The biochemical disease-free survival and morbidities were analyzed. Biochemical failure was defined using the Phoenix definition. From 2002 to 2012, 91 patients with biopsy-proven radio-recurrent PCa underwent salvage focal cryoablation with curative intent. The biochemical disease-free survival rates were 95.3%, 72.4%, and 46.5% at 1, 3, and 5 years, respectively. Positive biopsies after salvage focal cryoablation were observed in four of 14 patients who underwent biopsy (28.6%). Rectourethral fistula was observed in three cases (3.3%). Urinary retention was observed in six cases (6.6%). Incontinence (requiring pad use) was reported in five cases (5.5%). Intercourse was reported in 10 of 20 patients (50%) who reported potency before salvage focal cryoablation. The outcomes from this observational study indicate that salvage focal cryoablation can be an effective treatment with encouraging potency preservation for patients with locally recurrent PCa after radiotherapy. However, other morbidity including rectourethral fistula and incontinence are not clearly lower than for patients treated with salvage whole gland cryoablation. Studies with longer follow-up, more patients, and direct comparison to salvage whole gland cryoablation are needed before recommending salvage focal cryoablation as a standard treatment option for these patients.

Abstract 3423: The genomic evolution of prostate cancer under the selective pressure of anti-androgen therapy

Abstract Background The implementation of novel technologies such as array comparative genomic hybridization (aCGH) and next-generation sequencing has led to a deeper understanding of the genomic nature of cancer. However, these analyses have classically been done without respecting intra-tumor heterogeneity. Here, we applied a methodology that allows us analyzing the genomic profile of distinct tumor populations from individual tumors and their clonal evolution during the progression to castration-resistant disease, thus enabling the discovery of novel genetic alterations in the course of the development of therapy resistance. Methods Matched pre- and post- hormone treated fresh frozen and/or formalin fixed prostate cancer samples were selected from our biobank. Clonal tumor populations were flow-sorted according to their nuclear DNA content. Sorted tumor populations were subjected to whole genome CGH and to full exome sequencing analyses by the use of Agilent SurePrint 2x400k microarrays and the SureSelect All Exon Kit, respectively. Target aberrations were evaluated using a prostate cancer tissue micro array (TMA) and fluorescence in situ hybridization as well as immunohistochemistry. Results The de-convolution of the genomic evolution in the investigated tumors shows that genetic aberrations and mutations are newly occurring under the selective pressure of anti-androgen therapy. More importantly, judging on the temporal dynamics of changes in allelic frequencies, we cannot only observe events that become more abundant, but also cancer associated mutations that are negatively selected for. We are currently following up several of these targets, of which at least one shows significant association with overall survival and differences in its expression pattern between hormone naïve and castration resistant cancer. Conclusions Genomic profiling of distinct clonal tumor populations during prostate cancer progression allows for analysis of intra-tumoral heterogeneity and the underlying clonal evolution. Importantly, this approach identifies genomic aberrations that were selected for under the pressure of hormone ablation therapy. Citation Format: Joël Roman Gsponer, Tanja Dietsche, Alexander Rufle, Thomas Lorber, Darius Juskevicius, Valeria Perrina, Elisabeth Lenkiewicz, Tobias Zellweger, Alexander Bachmann, Michael T. Barrett, Cyrill A. Rentsch, Christian Ruiz, Lukas Bubendorf. The genomic evolution of prostate cancer under the selective pressure of anti-androgen therapy. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3423. doi:10.1158/1538-7445.AM2014-3423

Abstract 3021: Roles of EMT on stem cell properties of prostate stem and cancer cells during castration-resistant prostate cancer progression

Abstract Androgen ablation remains the mainstay of treatment for men with advanced and metastatic prostate cancer. However, despite the introduction of new generation anti-androgens, a majority of men succumb to castration resistant prostate cancer (CRPC). The molecular mechanisms governing the emergence of treatment resistance in CRPC patients are not well understood. Recent experience suggests that tumor regeneration from castration-resistant stem-like cells induce resistance to hormonal therapy. Therefore, elucidating novel targets essential for driving stem-like activity is critical to prevent and defeat CRPC. It has been shown that normal and cancer stem cells exploit normal development process of epithelial-mesenchymal transition (EMT) to survive and metastasize, and that EMT confer stem cell properties to more differentiated cancer cell progeny in breast and other cancers. However, it is unclear if EMT is linked with stem cells in normal and malignant prostate. Our lab has reported that N-cadherin, a marker of EMT, is upregulated after neoadjuvant hormone ablation and in CRPC and is sufficient to cause metastasis and CRPC. Therapeutic targeting of N-cadherin by novel N-cadherin antibody inhibited metastatic and CRPC progression. The cell population displaying N-cadherin co-expressed a number of stem cell-associated genes in CRPC models. Here, we verified EMT linked to stem cells in both normal prostate and CRPC. We found in LAPC-9 CRPC tumors, the cell population expressing N-cadherin behaved like stem cells with enhanced sphere-forming ability, which could be specifically inhibited by novel N-cadherin antibody 2A9. We isolated stem-like CD49fhi/Trop2hi cells from prostatectomy specimens and found that forced N-cadherin expression promoted sphere formation of those cells. Our evaluation of gene expression in N-cadherin-positive prostate cancer cell lines and CRPC tumors demonstrated that N-cadherin expression activated common EMT transcriptional regulators including Zeb1. We asked if Zeb1 regulated stem cell properties in normal and malignant prostate. We found that forced Zeb1 expression induced EMT with enhanced cell invasiveness in LNCaP human prostate cancer cells. However, Zeb1 overexpression inhibited cell proliferation and CRPC tumor growth of LNCaP. Zeb1 overexpression also surprisingly inhibited sphere formation of normal stem/progenitor cells from prostatectomy specimens. Our data suggest that Zeb1-regulated EMT promotes both quiescence and invasiveness in normal and malignant prostate in which the quiescent cells may survive and play a role in treatment resistance, while N-cadherin mediates stem cell proliferation and self-renewal. Our research will likely provide useful information of EMT-related biomarkers for preventing and developing efficient therapeutics to combat the treatment resistance to new generation anti-androgens. Citation Format: Shu Lin, Evelyn Kono, Joyce Yamashiro, Sean Lee, Owen Witte, Andrew Goldstein, Robert E. Reiter. Roles of EMT on stem cell properties of prostate stem and cancer cells during castration-resistant prostate cancer progression. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3021. doi:10.1158/1538-7445.AM2014-3021