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PDF - 260KB, Supplementary Figure 1. Validation of the Docetaxel-resistant prostate cancer cell line DU145-Rx. Supplementary Figure 1. Validation of the Docetaxel-resistant prostate cancer cell line DU145-Rx. Supplementary Figure 3. Co-treatment-induced cell death in Docetaxel-resistant DU145-Rx cells is associated with enhanced autophagy. Supplementary Table 1. SILAC ratios of up- or down-regulated tyrosine phosphorylation sites.

BackgroundThe interferon response can influence the primary and metastatic activity of breast cancers and can interact with checkpoint immunotherapy to modulate its effects. UsingN-ethyl-N-nitrosourea mutagenesis, we found a mouse with an activating mutation in oligoadenylate synthetase 2 (Oas2), a sensor of viral double stranded RNA, that resulted in an interferon response and prevented lactation in otherwise healthy mice.MethodsTo determine if sole activation ofOas2could alter the course of mammary cancer, we combined theOas2mutation with theMMTV-PyMToncogene model of breast cancer and examined disease progression and the effects of checkpoint immunotherapy using Kaplan–Meier survival analysis with immunohistochemistry and flow cytometry.ResultsOas2mutation prevented pregnancy from increasing metastases to lung. Checkpoint immunotherapy with antibodies against programmed death-ligand 1 was more effective when theOas2mutation was present.ConclusionsThese data establish OAS2 as a therapeutic target for agents designed to reduce metastases and increase the effectiveness of checkpoint immunotherapy.

Cancer cells invoke phenotypic plasticity programs to drive disease progression and evade chemotherapeutic insults, yet until now there have been no validated clinical therapies targeting this process. Here, we identify a phenotypic plasticity signature associated with poor survival in basal/triple-negative breast cancer, in which androgen signalling is prominent. We establish that anti-androgen therapies block cancer stem cell function and prevent chemotherapy-induced emergence of new cancer stem cells. In particular, the anti-androgen agent seviteronel synergizes with chemotherapy to improve chemotherapeutic inhibition of primary and metastatic tumour growth and prevent the emergence of chemotherapy-resistant disease. We validate cytoplasmic AR expression as a clinical phenotypic plasticity biomarker that predicts poor survival and poor response to chemotherapy, and positive response to seviteronel plus chemotherapy. This new targeted combination therapy validates modulating phenotypic plasticity as an effective strategy to prevent and treat chemotherapy-resistant cancers with transformative clinical potential.STATEMENT OF SIGNIFICANCEThere are currently no curative therapies for patients with chemotherapy-resistant cancer. We demonstrate that modulating phenotypic plasticity prevents the emergence of chemotherapy-resistant disease in triple-negative breast cancer. This represents the first known validated clinical therapy leveraging phenotypic plasticity. Moreover, we identify a highly effective anti-androgen drug and a biomarker to select and treat patients best-suited to this new therapy. A clinical trial is underway (NCT04947189).SUMMARY SENTENCEBlocking phenotypic plasticity is an effective targeted therapeutic strategy to treat cance

Pancreatic ductal adenocarcinoma (PDAC) remains one of the deadliest malignancies. It is phenotypically heterogeneous with a highly unstable genome and provides few common therapeutic targets. We found that MCL1, Cofilin1 (CFL1) and SRC mRNA were highly expressed by a wide range of these cancers, suggesting that a strategy of dual MCL-1 and SRC inhibition might be efficacious for many patients. Immunohistochemistry revealed that MCL-1 protein was present at high levels in 94.7% of patients in a cohort of PDACs from Australian Pancreatic Genome Initiative (APGI). High MCL1 and Cofilin1 mRNA expression was also strongly predictive of poor outcome in the TCGA dataset and in the APGI cohort. In culture, MCL-1 antagonism reduced the level of the cytoskeletal remodeling protein Cofilin1 and phosphorylated SRC on the active Y416 residue, suggestive of reduced invasive capacity. The MCL-1 antagonist S63845 synergized with the SRC kinase inhibitor dasatinib to reduce cell viability and invasiveness through 3D-organotypic matrices. In preclinical murine models, this combination reduced primary tumor growth and liver metastasis of pancreatic cancer xenografts. These data suggest that MCL-1 antagonism, while reducing cell viability, may have an additional benefit in increasing the antimetastatic efficacy of dasatinib for the treatment of PDAC.

The tumour stroma, and in particular the extracellular matrix (ECM), is a salient feature of solid tumours that plays a crucial role in shaping their progression. Many desmoplastic tumours including breast cancer involve the significant accumulation of type I collagen. However, recently it has become clear that the precise distribution and organisation of matrix molecules such as collagen I is equally as important in the tumour as their abundance. Cancer-associated fibroblasts (CAFs) coexist within breast cancer tissues and play both pro- and anti-tumourigenic roles through remodelling the ECM. Here, using temporal proteomic profiling of decellularized tumours, we interrogate the evolving matrisome during breast cancer progression. We identify 4 key matrisomal clusters, and pinpoint collagen type XII as a critical component that regulates collagen type I organisation. Through combining our proteomics with single-cell transcriptomics, and genetic manipulation models, we show how CAF-secreted collagen XII alters collagen I organisation to create a pro-invasive microenvironment supporting metastatic dissemination. Finally, we show in patient cohorts that collagen XII may represent an indicator of breast cancer patients at high risk of metastatic relapse. ispartof: NATURE COMMUNICATIONS vol:13 issue:1 ispartof: location:England status: published

Background Docetaxel, the standard chemotherapy for metastatic castration-resistant prostate cancer (CRPC) also enhances the survival of patients with metastatic castration-sensitive prostate cancer (CSPC) when combined with androgen-deprivation therapy. Focal Adhesion Kinase (FAK) activation is a mediator of docetaxel resistance in prostate cancer cells. The aim of this study was to investigate the effect of the second generation FAK inhibitor VS-6063 on docetaxel efficacy in pre-clinical CRPC and CSPC models. Methods Docetaxel-resistant CRPC cells, mice with PC3 xenografts, and ex vivo cultures of patient-derived primary prostate tumors were treated with VS-6063 and/or docetaxel, or vehicle control. Cell counting, immunoblotting, and immunohistochemistry techniques were used to evaluate the treatment effects. Results Docetaxel and VS-6063 co-treatment caused a greater decrease in the viability of docetaxel-resistant CRPC cells, and a greater inhibition in PC3 xenograft growth compared to either monotherapy. FAK expression in human primary prostate cancer was positively associated with advanced tumor stage. Patient-derived prostate tumor explants cultured with both docetaxel and VS-6063 displayed a higher percentage of apoptosis in cancer cells, than monotherapy treatment. Conclusions Our findings suggest that co-administration of the FAK inhibitor, VS-6063, with docetaxel represents a potential therapeutic strategy to overcome docetaxel resistance in prostate cancer.

Contains fulltext : 240100.pdf (Publisher’s version ) (Open Access) Basal breast cancer is associated with younger age, early relapse, and a high mortality rate. Here, we use unbiased droplet-based single-cell RNA sequencing (RNA-seq) to elucidate the cellular basis of tumor progression during the specification of the basal breast cancer subtype from the luminal progenitor population in the MMTV-PyMT (mouse mammary tumor virus-polyoma middle tumor-antigen) mammary tumor model. We find that basal-like cancer cells resemble the alveolar lineage that is specified upon pregnancy and encompass the acquisition of an aberrant post-lactation developmental program of involution that triggers remodeling of the tumor microenvironment and metastatic dissemination. This involution mimicry is characterized by a highly interactive multicellular network, with involution cancer-associated fibroblasts playing a pivotal role in extracellular matrix remodeling and immunosuppression. Our results may partially explain the increased risk and poor prognosis of breast cancer associated with childbirth.

Docetaxel and cabazitaxel are taxane chemotherapy treatments for metastatic castration-resistant prostate cancer (CRPC). However, therapeutic resistance remains a major issue. MicroRNAs are short non-coding RNAs that can silence multiple genes, regulating several signalling pathways simultaneously. Therefore, synthetic microRNAs may have therapeutic potential in CRPC by regulating genes involved in taxane response and minimise compensatory mechanisms that cause taxane resistance. To identify microRNAs that can improve the efficacy of taxanes in CRPC, we performed a genome-wide screen of 1280 microRNAs in the CRPC cell lines PC3 and DU145 in combination with docetaxel or cabazitaxel treatment. Mimics of miR-217 and miR-181b-5p enhanced apoptosis significantly in PC3 cells in the presence of these taxanes. These mimics downregulated at least a thousand different transcripts, which were enriched for genes with cell proliferation and focal adhesion functions. Individual knockdown of a selection of 46 genes representing these transcripts resulted in toxic or taxane sensitisation effects, indicating that these genes may be mediating the effects of the microRNA mimics. A range of these genes are expressed in CRPC metastases, suggesting that these microRNA mimics may be functional in CRPC. With further development, these microRNA mimics may have therapeutic potential to improve taxane response in CRPC patients.

Quantification of cellular antigens and their interactions via antibody-based detection methods are widely used in scientific research. Accurate high-throughput quantitation of these assays using general image analysis software can be time consuming and challenging, particularly when attempted by users with limited image processing and analysis knowledge. To overcome this, we have designed Andy’s Algorithms, a series of automated image analysis pipelines for FIJI, that permits rapid, accurate and reproducible batch-processing of 3,3′-diaminobenzidine (DAB) immunohistochemistry, proximity ligation assays (PLAs) and other common assays. Andy’s Algorithms incorporates a step-by-step tutorial and optimization pipeline to make batch image analysis simple for the untrained user and adaptable across laboratories. Andy’s algorithms provide a simpler, faster, standardized work flow compared to existing programs, while offering equivalent performance and additional features, in a free to use open-source application of FIJI. Andy’s Algorithms are available at GitHub, publicly accessed at https://github.com/andlaw1841/Andy-s-Algorithm.

We identified a non-synonymous mutation in Oas2 (I405N), a sensor of viral double-stranded RNA, from an ENU-mutagenesis screen designed to discover new genes involved in mammary development. The mutation caused post-partum failure of lactation in healthy mice with otherwise normally developed mammary glands, characterized by greatly reduced milk protein synthesis coupled with epithelial cell death, inhibition of proliferation and a robust interferon response. Expression of mutant but not wild type Oas2 in cultured HC-11 or T47D mammary cells recapitulated the phenotypic and transcriptional effects observed in the mouse. The mutation activates the OAS2 pathway, demonstrated by a 34-fold increase in RNase L activity, and its effects were dependent on expression of RNase L and IRF7, proximal and distal pathway members. This is the first report of a viral recognition pathway regulating lactation., Author summary Using ENU-mutagenesis in mice we discovered a pedigree with lactation failure. Mammary development through puberty and pregnancy appeared normal in mutant animals, but the activation of lactation failed in the immediate post partum period and no milk reached the pups. Failure of lactation was accompanied by greatly diminished milk protein synthesis, decreased epithelial cell proliferation, increased epithelial cell death and a robust interferon response. A non-synonymous mutation in Oas2 (I405N) in the viral sensor Oas2 was found and expression of mutant Oas2 in mammary cells recapitulated these phenotypes. RNase L, the most proximal effector of OAS2 action, was activated in the mammary glands of mutant mice and in mammary cells expressing mutant Oas2. Knockdown of RNase L, or the distal pathway member IRF7, prevented these effects, indicating that the mutation in OAS2 caused activation of the viral signaling pathway. These results show that viral detection in the mammary gland can prevent lactation.

Background: Glutathione S-transferase 1 (GSTP1) inactivation is associated with CpG island promoter hypermethylation in the majority of prostate cancers (PCs). This study assessed whether the level of circulating methylated GSTP1 (mGSTP1) in plasma DNA is associated with chemotherapy response and overall survival (OS). Methods: Plasma samples were collected prospectively from a Phase I exploratory cohort of 75 men with castrate-resistant PC (CRPC) and a Phase II independent validation cohort (n=51). mGSTP1 levels in free DNA were measured using a sensitive methylation-specific PCR assay. Results: The Phase I cohort identified that detectable baseline mGSTP1 DNA was associated with poorer OS (HR, 4.2 95% CI 2.1–8.2; P

Background: Docetaxel is the first-line chemotherapy for castration-resistant prostate cancer (CRPC). However, response rates are ∼50% and determined quite late in the treatment schedule, thus non-responders are subjected to unnecessary toxicity. The potential of circulating microRNAs as early biomarkers of docetaxel response in CRPC patients was investigated in this study. Methods: Global microRNA profiling was performed on docetaxel-resistant and sensitive cell lines to identify candidate circulating microRNA biomarkers. Custom Taqman Array MicroRNA cards were used to measure the levels of 46 candidate microRNAs in plasma/serum samples, collected before and after docetaxel treatment, from 97 CRPC patients. Results: Fourteen microRNAs were associated with serum prostate-specific antigen (PSA) response or overall survival, according to Mann–Whitney U or log-rank tests. Non-responders to docetaxel and patients with shorter survival generally had high pre-docetaxel levels of miR-200 family members or decreased/unchanged post-docetaxel levels of miR-17 family members. Multivariate Cox regression with bootstrapping validation showed that pre-docetaxel miR-200b levels, post-docetaxel change in miR-20a levels, pre-docetaxel haemoglobin levels and visceral metastasis were independent predictors of overall survival when modelled together. Conclusions: Our study suggests that circulating microRNAs are potential early predictors of docetaxel chemotherapy outcome, and warrant further investigation in clinical trials.

Docetaxel remains the standard-of-care for men diagnosed with metastatic castrate-resistant prostate cancer (CRPC). However, only approximately 50% of patients benefit from treatment and all develop docetaxel-resistant disease. Here, we characterize global perturbations in tyrosine kinase signaling associated with docetaxel resistance and thereby develop a potential therapeutic strategy to reverse this phenotype. Using quantitative mass spectrometry–based phosphoproteomics, we identified that metastatic docetaxel-resistant prostate cancer cell lines (DU145-Rx and PC3-Rx) exhibit increased phosphorylation of focal adhesion kinase (FAK) on Y397 and Y576, in comparison with parental controls (DU145 and PC3, respectively). Bioinformatic analyses identified perturbations in pathways regulating focal adhesions and the actin cytoskeleton and in protein–protein interaction networks related to these pathways in docetaxel-resistant cells. Treatment with the FAK tyrosine kinase inhibitor (TKI) PF-00562271 reduced FAK phosphorylation in the resistant cells, but did not affect cell viability or Akt phosphorylation. Docetaxel administration reduced FAK and Akt phosphorylation, whereas cotreatment with PF-00562271 and docetaxel resulted in an additive attenuation of FAK and Akt phosphorylation and overcame the chemoresistant phenotype. The enhanced efficacy of cotreatment was due to increased autophagic cell death, rather than apoptosis. These data strongly support that enhanced FAK activation mediates chemoresistance in CRPC, and identify a potential clinical niche for FAK TKIs, where coadministration with docetaxel may be used in patients with CRPC to overcome chemoresistance. Mol Cancer Ther; 13(1); 190–201. ©2013 AACR.

Chronic opiate usage, whether prescribed or illicit, has been associated with changes in bone mass and is a recognized risk factor for the development of osteoporosis; however, the mechanism behind this effect is unknown. Here we show that lack of dynorphin, an endogenous opioid, in mice (Dyn−/−), resulted in a significantly elevated cancellous bone volume associated with greater mineral apposition rate and increased resorption indices. A similar anabolic phenotype was evident in bone of mice lacking dynorphin’s cognate receptor, the kappa opioid receptor. Lack of opioid receptor expression in primary osteoblastic cultures and no change in bone cell function after dynorphin agonist treatment in vitro indicates an indirect mode of action. Consistent with a hypothalamic action, central dynorphin signaling induces extracellular signal-regulated kinase (ERK) phosphorylation and c-fos activation of neurons in the arcuate nucleus of the hypothalamus (Arc). Importantly, this signaling also leads to an increase in Arc NPY mRNA expression, a change known to decrease bone formation. Further implicating NPY in the skeletal effects of dynorphin, Dyn−/−/NPY−/− double mutant mice showed comparable increases in bone formation to single mutant mice, suggesting that dynorphin acts upstream of NPY signaling to control bone formation. Thus the dynorphin system, acting via NPY, may represent a pathway by which higher processes including stress, reward/addiction and depression influence skeletal metabolism. Moreover, understanding of these unique interactions may enable modulation of the adverse effects of exogenous opioid treatment without directly affecting analgesic responses.

Background Metastatic disease is largely resistant to therapy and accounts for almost all cancer deaths. Myeloid cell leukemia-1 (MCL-1) is an important regulator of cell survival and chemo-resistance in a wide range of malignancies, and thus its inhibition may prove to be therapeutically useful. Methods To examine whether targeting MCL-1 may provide an effective treatment for breast cancer, we constructed inducible models of BIMs2A expression (a specific MCL-1 inhibitor) in MDA-MB-468 (MDA-MB-468-2A) and MDA-MB-231 (MDA-MB-231-2A) cells. Results MCL-1 inhibition caused apoptosis of basal-like MDA-MB-468-2A cells grown as monolayers, and sensitized them to the BCL-2/BCL-XL inhibitor ABT-263, demonstrating that MCL-1 regulated cell survival. In MDA-MB-231-2A cells, grown in an organotypic model, induction of BIMs2A produced an almost complete suppression of invasion. Apoptosis was induced in such a small proportion of these cells that it could not account for the large decrease in invasion, suggesting that MCL-1 was operating via a previously undetected mechanism. MCL-1 antagonism also suppressed local invasion and distant metastasis to the lung in mouse mammary intraductal xenografts. Kinomic profiling revealed that MCL-1 antagonism modulated Src family kinases and their targets, which suggested that MCL-1 might act as an upstream modulator of invasion via this pathway. Inhibition of MCL-1 in combination with dasatinib suppressed invasion in 3D models of invasion and inhibited the establishment of tumors in vivo. Conclusion These data provide the first evidence that MCL-1 drives breast cancer cell invasion and suggests that MCL-1 antagonists could be used alone or in combination with drugs targeting Src kinases such as dasatinib to suppress metastasis. Electronic supplementary material The online version of this article (doi:10.1186/s13058-016-0781-6) contains supplementary material, which is available to authorized users.

Background: Docetaxel improves symptoms and survival in metastatic castration-resistant prostate cancer (CRPC). However, ∼50% of patients are chemoresistant. This study examined whether changes in cytokine levels predict for docetaxel resistance in vitro and in a clinical cohort. Methods: PC3 cells or their docetaxel-resistant subline (PC3Rx) were co-cultured with U937 monocytes, with and without docetaxel treatment, and cytokine levels were measured. The circulating levels of 28 cytokines were measured pre-/post cycle 1 of docetaxel from 55 men with CRPC, and compared with prostate-specific antigen (PSA) response. Results: PC3Rx-U937 co-culture expressed more cytokines, chiefly markers of alternative macrophage differentiation, compared with PC3-U937 co-culture. Docetaxel treatment enhanced cytokine production by PC3Rx-U937 co-culture, while reducing cytokine levels in PC3-U937. In patients, changes in the levels of seven circulating cytokines (macrophage inhibitory cytokine 1 (MIC1), interleukin (IL)-1ra, IL-1β, IL-4, IL-6, IL-12 and IFNγ) after cycle 1 of docetaxel were associated with progressive disease (all P

Y2 receptors, particularly those in the brain, have been implicated in neuropeptide Y (NPY)-mediated effects on energy homeostasis and bone mass. Recent evidence also indicates a role for Y2 receptors in peripheral tissues in this process by promoting adipose tissue accretion; however their effects on energy balance remain unclear. Here, we show that adult-onset conditional knockdown of Y2 receptors predominantly in peripheral tissues results in protection against diet-induced obesity accompanied by significantly reduced weight gain, marked reduction in adiposity and improvements in glucose tolerance without any adverse effect on lean mass or bone. These changes occur in association with significant increases in energy expenditure, respiratory exchange ratio, and physical activity and despite concurrent hyperphagia. On a chow diet, knockdown of peripheral Y2 receptors results in increased respiratory exchange ratio and physical activity with no effect on lean or bone mass, but decreases energy expenditure without effecting body weight or food intake. These results suggest that peripheral Y2 receptor signaling is critical in the regulation of oxidative fuel selection and physical activity and protects against the diet-induced obesity. The lack of effects on bone mass seen in this model further indicates that bone mass is primarily controlled by non-peripheral Y2 receptors. This study provides evidence that novel drugs that target peripheral rather than central Y2 receptors could provide benefits for the treatment of obesity and glucose intolerance without adverse effects on lean and bone mass, with the additional benefit of avoiding side effects often associated with pharmaceuticals that act on the central nervous system.

BACKGROUND: Y2 receptor signalling is known to be important in neuropeptide Y (NPY)-mediated effects on energy homeostasis and bone physiology. Y2 receptors are located post-synaptically as well as acting as auto receptors on NPY-expressing neurons, and the different roles of these two populations of Y2 receptors in the regulation of energy homeostasis and body composition are unclear. METHODOLOGY/PRINCIPAL FINDINGS: We thus generated two conditional knockout mouse models, Y2(lox/lox) and NPYCre/+;Y2(lox/lox), in which Y2 receptors can be selectively ablated either in the hypothalamus or specifically in hypothalamic NPY-producing neurons of adult mice. Specific deletion of hypothalamic Y2 receptors increases food intake and body weight compared to controls. Importantly, specific ablation of hypothalamic Y2 receptors on NPY-containing neurons results in a significantly greater adiposity in female but not male mice, accompanied by increased hepatic triglyceride levels, decreased expression of liver carnitine palmitoyltransferase (CPT1) and increased expression of muscle phosphorylated acetyl-CoA carboxylase (ACC). While food intake, body weight, femur length, bone mineral content, density and cortical bone volume and thickness are not significantly altered, trabecular bone volume and number were significantly increased by hypothalamic Y2 deletion on NPY-expressing neurons. Interestingly, in situ hybridisation reveals increased NPY and decreased proopiomelanocortin (POMC) mRNA expression in the arcuate nucleus of mice with hypothalamus-specific deletion of Y2 receptors in NPY neurons, consistent with a negative feedback mechanism between NPY expression and Y2 receptors on NPY-ergic neurons. CONCLUSIONS/SIGNIFICANCE: Taken together these data demonstrate the anti-obesogenic role of Y2 receptors in the brain, notably on NPY-ergic neurons, possibly via inhibition of NPY neurons and concomitant stimulation of POMC-expressing neurons in the arcuate nucleus of the hypothalamus, reducing lipogenic pathways in liver and/or skeletal muscle in females. These data also reveal as an anti-osteogenic effect of Y2 receptors on hypothalamic NPY-expressing neurons on trabecular but not on cortical bone.

44 Background: Despite a range of new treatments, docetaxel (DTX) remains the first-line chemotherapy for castration-resistant prostate cancer (CRPC). However, only 50% of patients respond to docetaxel at the cost of potentially significant toxicity. Therefore, there is a need for new biomarkers to identify early response to therapy. This study aims to determine if circulating microRNAs are associated with DTX chemotherapy outcome in CRPC. Methods: Global microRNA profiling was performed on DTX-resistant and sensitive cell lines to identify candidate circulating microRNA biomarkers. Custom Taqman Array MicroRNA cards measured the levels of 46 candidate microRNAs in plasma/serum from 97 CRPC patients, collected pre- and three weeks post-cycle one of DTX. Responses were defined by the PCWG1 serum prostate-specific antigen (PSA) response criteria; partial response (PR), stable disease (SD), progressive disease (PD). Multiple T-test, Mann-Whitney U, Kaplan-Meier, Receiver Operating Characteristic (ROC), and Cox regression analyses were used to assess the associations between microRNA levels and clinical outcomes. Results: Eighteen microRNAs were associated with PSA response or overall survival (p

11 Background: GSTP1 is inactivated and displays aberrant CpG island promoter methylation in the majority of prostate cancers (PCs). The aim of this study was to assess whether changes in methylated glutathione s-transferase 1 (mGSTP1) levels are predictive of response to chemotherapy and overall survival (OS). Methods: Plasma samples were collected prospectively from a phase I exploratory cohort of 75 men with castration-resistant prostate cancer (CRPC) (40 treated with chemotherapy) and a phase II independent validation cohort (n=51). Plasma samples were collected at presentation with CRPC and pre- and post-cycle one of chemotherapy (90% Docetaxel). mGSTP1 levels were measured using a methylation-specific head-loop PCR assay from DNA isolated from either (1) immunomagnetic separation of circulating tumor cells (CTCs) or (2) free plasma DNA. The associations between plasma mGSTP1 levels, PSA response (defined by PCWG1 criteria), and OS were tested using non-parametric tests and survival analyses. Results: The ability to detect mGSTP1 was significantly higher for the plasma free DNA assay compared to DNA from CTCs (p

SummaryNeuropepetide Y (NPY) is best known for its powerful stimulation of food intake and its effects on reducing energy expenditure. However, the pathways involved and the regulatory mechanisms behind this are not well understood. Here we demonstrate that NPY derived from the arcuate nucleus (Arc) is critical for the control of sympathetic outflow and brown adipose tissue (BAT) function. Mechanistically, a key change induced by Arc NPY signaling is a marked Y1 receptor-mediated reduction in tyrosine hydroxylase (TH) expression in the hypothalamic paraventricular nucleus (PVN), which is also associated with a reduction in TH expression in the locus coeruleus (LC) and other regions in the brainstem. Consistent with this, Arc NPY signaling decreased sympathetically innervated BAT thermogenesis, involving the downregulation of uncoupling protein 1 (UCP1) expression in BAT. Taken together, these data reveal a powerful Arc-NPY-regulated neuronal circuit that controls BAT thermogenesis and sympathetic output via TH neurons.

e22175 Background: DTX improves symptoms and survival in advanced CRPC, however ~ 50% of patients have chemoresistant disease. Given the toxicity of cytotoxic treatment in this population, there is an urgent need for early chemoresistance markers, a greater understanding of chemoresistance mechanisms and the development of new therapies. This study examined whether early changes in cytokine levels predict chemoresistance and explored the role of macrophages in cytokine generation and chemoresistance in vitro. Methods: 28 cytokines were measured pre/post cycle 1 of chemotherapy from 59 men with metastatic CRPC. Cytokine levels were correlated with PSA response and OS. DTX sensitive PC3 and DTX resistant PC3Rx cells were cultured alone and in coculture with U937 monocytes +/- DTX treatment. Conditioned media (CM) was analysed by a 36 cytokine array panel and by ELISA for MIC1. CM from U937 cells exposed to DTX was used to treat PC3 cells combined with escalating DTX doses. Results: Change in the levels of 10 cytokines over 1 cycle of chemotherapy (MIC-1 p=0.003; IL1ra p=0.004; IL1b p=0.01; IL4 p