Anterior gradient 2 and 3 – two prototype androgen‐responsive genes transcriptionally upregulated by androgens and by oestrogens in prostate cancer cells

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/96748/1/febs12118.pd

Dissecting molecular mechanisms of resistance to NOTCH1-targeted therapy in T-cell acute lymphoblastic leukemia xenografts

Despite substantial progress in treatment of T-cell acute lymphoblastic leukemia (T-ALL), mortality remains relatively high, mainly due to primary or acquired resistance to chemotherapy. Further improvements in survival demand better understanding of T-ALL biology and development of new therapeutic strategies. The Notch pathway has been involved in the pathogenesis of this disease and various therapeutic strategies are currently under development, including selective targeting of NOTCH receptors by inhibitory antibodies. We previously demonstrated that the NOTCH1-specific neutralizing antibody OMP52M51 prolongs survival in TALL patient-derived xenografts bearing NOTCH1/FBW7 mutations. However, acquired resistance to OMP52M51 eventually developed and we used patient-derived xenografts models to investigate this phenomenon. Multi-level molecular characterization of T-ALL cells resistant to NOTCH1 blockade and serial transplantation experiments uncovered heterogeneous types of resistance, not previously reported with other Notch inhibitors. In one model, resistance appeared after 156 days of treatment, it was stable and associated with loss of Notch inhibition, reduced mutational load and acquired NOTCH1 mutations potentially affecting the stability of the heterodimerization domain. Conversely, in another model resistance developed after only 43 days of treatment despite persistent down-regulation of Notch signaling and it was accompanied by modulation of lipid metabolism and reduced surface expression of NOTCH1. Our findings shed light on heterogeneous mechanisms adopted by the tumor to evade NOTCH1 blockade and support clinical implementation of antibody-based target therapy for Notch-addicted tumors

Identification and characterization of androgen-regulated genes involved in prostate carcinogenesis and prostate cancer progression

Androgen-Rezeptor (AR) Signalweg spielt eine essentielle Rolle beim Prostatakrebs (PCa). Sowohl die Krebsentwicklung als auch die Progression zu einem aggressiven kastrationsresitenten Tumor (CRPC) erfolgen unter seinem direkten Einfluss. Die Hemmung des AR ist aus diesem Grunde die Hauptstrategie bei der Prostatakrebstherapie. Leider ist die Wirksamkeit zeitlich begrenzt und Identifizierung und Charakterisierung von AR Effektoren, die die AR Wirkung vermitteln, stehen im Mittelpunkt der Bemühungen PCa Diagnose, Prognose und die medizinische Versorgung der betroffenen Patienten zu verbessern. In dieser Arbeit wurde das AR Cistrome untersucht und zwar mit einem besonderem Augenmerk auf den Zusammenhang zwischen AR Effekten und sogenannten nicht-kodierenden RNAs (miRNAs). miRNA wurden in den letzten Jahren zunehmend als wichtige Regulatoren der Genexpression im Zusammenhang mit Krebsentstehung und progression erkannt und untersucht. Im Detail hat sich in dieser Arbeit gezeigt, dass der AR nach Aktivierung durch Androgene in Prostatatumorzellen neben einigen anderen miRNAs miR-22 und miR-29a induziert. Interessanterweise wirken miR-22 und miR-29a als tumorhemmende miRNAs, indem sie zwei Krebs-assoziierte Transkripte, LAMC1 und MCL1, reduzieren. In der Arbeit wurden zwei Ebenen der Regulation für diese beiden miRNAs in PCa Tumoren charakterisiert, einerseits ein epigenetisches Silencing der genomischen Regionen und andererseits eine AR Regulation über entsprechende androgenresponsive Elemente in den kodierenden Genen. Ein weiterer Teil dieser Arbeit widmete sich einer eingehenden Untersuchung der beiden Gene AGR2 und AGR3 und deren Regulation in Prostatatumoren durch androgene und östrogene Hormone. Aufgrund ihrer beschrieben Rollen, gelten beide Gene als vielversprechende Biomarker für Prostatakrebs sowie für Brustkrebs. Beide AGR Gene wurden auf Grund der in den Genloci vorhandenen -Bindungsstellen und ihrer Induktion durch Androgene dem AR cistrome zugeordnet. Darüber hinaus verdeutlichte die beobachtete Induktion von AGR2 und AGR3 durch Östrogene, die in PCa Zellen beide Gene durch Aktivierung von Wildtyp-AR induzieren können, die Fähigkeit von Tumorzellen, sich an Veränderungen in ihrer Tumormikroumgebung anzupassen um damit einer Tumorbehandlung zu entkommen. Insgesamt tragen die Erkenntnisse dieser Arbeit dazu bei, mehr Licht auf das komplexe regulatorische Netzwerk, von AR, miRNAs und AR/miRNAs nachgeschalteten Zielgenen zu werfen. Dieses Netzwerk unterliegt einer permanenten Entwicklung während der Prostatakrebs fortschreitet und aggressiver wird. Durch diese Erkenntnisse werden Bemühungen unterstützt integrative Therapieansätze zu entwickeln und spezifischere Krebs-Biomarker zu etablieren, um so das Management von betroffenen Patienten zu verbessern.Androgen receptor (AR) signaling is required for prostate cancer (PCa) onset as well as development to the more aggressive stage named castration-resistant prostate cancer (CRPC). Targeting AR represents the mainstay strategy to treat PCa, however, since it has a time-limited efficacy identification of AR down-stream effectors with an essential role in mediating AR-signaling is necessary to improve PCa diagnosis, prognosis and medical care. In this work, the AR cistrome has been investigated with particular emphasis on the link between AR and the non-coding RNAs known as microRNAs (miRNAs) that recently have popped-out as powerful regulators of gene expression and being involved in carcinogenesis. In detail, it has been demonstrated that AR binds to miR-22 and miR-29a and induces their expression significantly in prostate cancer cells following androgen stimulation. Intriguingly, miR-22 and miR-29a act as tumor suppressive miRNAs modulating two cancer-associated transcripts, LAMC1 and MCL1, respectively. Indeed, their levels are reduced in primary prostate tumor samples via epigenetic silencing of the genomic regions harboring AR regulatory elements. One additional interest of this study has been an in-depth examination of AGR2 and AGR3 genes regulation by androgens and oestrogens in the context of PCa, because of their extensively described roles as promising biomarkers for prostate as well as breast cancer. Both AGR genes have been confirmed being members of the AR cistrome on the basis of their strong up-regulation in response to androgens and the presence of androgen receptor binding sites within their genomic locus. Moreover, the recognition of AGR2 and AGR3 induction by oestrogens via activation of wild-type AR validates the tumor ability to adapt to microenvironmental changes and develop mechanisms for treatment escape. Overall, these findings contributed to shed light on the intricate network including AR, miRNAs and AR/miRNAs down-stream target genes. This network evolves as prostate cancer progresses and strengthens the need to develop an integrative therapeutic approach as well as more specific cancer biomarkers to improve PCa management.by Lorenza PasqualiniAbweichender Titel laut Übersetzung der Verfasserin/des VerfassersEnth. u.a. 2 Veröff. d. Verf. aus den Jahren 2013 - 2014 . - Zsfassung in dt. SpracheInnsbruck, Med. Univ., Diss., 2015OeBB(VLID)76172

Identification and characterization of androgen-regulated genes involved in prostate carcinogenesis and prostate cancer progression

Androgen-Rezeptor (AR) Signalweg spielt eine essentielle Rolle beim Prostatakrebs (PCa). Sowohl die Krebsentwicklung als auch die Progression zu einem aggressiven kastrationsresitenten Tumor (CRPC) erfolgen unter seinem direkten Einfluss. Die Hemmung des AR ist aus diesem Grunde die Hauptstrategie bei der Prostatakrebstherapie. Leider ist die Wirksamkeit zeitlich begrenzt und Identifizierung und Charakterisierung von AR Effektoren, die die AR Wirkung vermitteln, stehen im Mittelpunkt der Bemühungen PCa Diagnose, Prognose und die medizinische Versorgung der betroffenen Patienten zu verbessern. In dieser Arbeit wurde das AR Cistrome untersucht und zwar mit einem besonderem Augenmerk auf den Zusammenhang zwischen AR Effekten und sogenannten nicht-kodierenden RNAs (miRNAs). miRNA wurden in den letzten Jahren zunehmend als wichtige Regulatoren der Genexpression im Zusammenhang mit Krebsentstehung und progression erkannt und untersucht. Im Detail hat sich in dieser Arbeit gezeigt, dass der AR nach Aktivierung durch Androgene in Prostatatumorzellen neben einigen anderen miRNAs miR-22 und miR-29a induziert. Interessanterweise wirken miR-22 und miR-29a als tumorhemmende miRNAs, indem sie zwei Krebs-assoziierte Transkripte, LAMC1 und MCL1, reduzieren. In der Arbeit wurden zwei Ebenen der Regulation für diese beiden miRNAs in PCa Tumoren charakterisiert, einerseits ein epigenetisches Silencing der genomischen Regionen und andererseits eine AR Regulation über entsprechende androgenresponsive Elemente in den kodierenden Genen. Ein weiterer Teil dieser Arbeit widmete sich einer eingehenden Untersuchung der beiden Gene AGR2 und AGR3 und deren Regulation in Prostatatumoren durch androgene und östrogene Hormone. Aufgrund ihrer beschrieben Rollen, gelten beide Gene als vielversprechende Biomarker für Prostatakrebs sowie für Brustkrebs. Beide AGR Gene wurden auf Grund der in den Genloci vorhandenen -Bindungsstellen und ihrer Induktion durch Androgene dem AR cistrome zugeordnet. Darüber hinaus verdeutlichte die beobachtete Induktion von AGR2 und AGR3 durch Östrogene, die in PCa Zellen beide Gene durch Aktivierung von Wildtyp-AR induzieren können, die Fähigkeit von Tumorzellen, sich an Veränderungen in ihrer Tumormikroumgebung anzupassen um damit einer Tumorbehandlung zu entkommen. Insgesamt tragen die Erkenntnisse dieser Arbeit dazu bei, mehr Licht auf das komplexe regulatorische Netzwerk, von AR, miRNAs und AR/miRNAs nachgeschalteten Zielgenen zu werfen. Dieses Netzwerk unterliegt einer permanenten Entwicklung während der Prostatakrebs fortschreitet und aggressiver wird. Durch diese Erkenntnisse werden Bemühungen unterstützt integrative Therapieansätze zu entwickeln und spezifischere Krebs-Biomarker zu etablieren, um so das Management von betroffenen Patienten zu verbessern.Androgen receptor (AR) signaling is required for prostate cancer (PCa) onset as well as development to the more aggressive stage named castration-resistant prostate cancer (CRPC). Targeting AR represents the mainstay strategy to treat PCa, however, since it has a time-limited efficacy identification of AR down-stream effectors with an essential role in mediating AR-signaling is necessary to improve PCa diagnosis, prognosis and medical care. In this work, the AR cistrome has been investigated with particular emphasis on the link between AR and the non-coding RNAs known as microRNAs (miRNAs) that recently have popped-out as powerful regulators of gene expression and being involved in carcinogenesis. In detail, it has been demonstrated that AR binds to miR-22 and miR-29a and induces their expression significantly in prostate cancer cells following androgen stimulation. Intriguingly, miR-22 and miR-29a act as tumor suppressive miRNAs modulating two cancer-associated transcripts, LAMC1 and MCL1, respectively. Indeed, their levels are reduced in primary prostate tumor samples via epigenetic silencing of the genomic regions harboring AR regulatory elements. One additional interest of this study has been an in-depth examination of AGR2 and AGR3 genes regulation by androgens and oestrogens in the context of PCa, because of their extensively described roles as promising biomarkers for prostate as well as breast cancer. Both AGR genes have been confirmed being members of the AR cistrome on the basis of their strong up-regulation in response to androgens and the presence of androgen receptor binding sites within their genomic locus. Moreover, the recognition of AGR2 and AGR3 induction by oestrogens via activation of wild-type AR validates the tumor ability to adapt to microenvironmental changes and develop mechanisms for treatment escape. Overall, these findings contributed to shed light on the intricate network including AR, miRNAs and AR/miRNAs down-stream target genes. This network evolves as prostate cancer progresses and strengthens the need to develop an integrative therapeutic approach as well as more specific cancer biomarkers to improve PCa management.by Lorenza PasqualiniAbweichender Titel laut Übersetzung der Verfasserin/des VerfassersEnth. u.a. 2 Veröff. d. Verf. aus den Jahren 2013 - 2014 . - Zsfassung in dt. SpracheInnsbruck, Med. Univ., Diss., 2015OeBB(VLID)76172

Acute stress enhances the expression of neuroprotection- and neurogenesis-associated genes in the hippocampus of a mouse restraint model

Stress arises from an external demand placed on an organism that triggers physiological, cognitive and behavioural responses in order to cope with that request. It is thus an adaptive response useful for the survival of an organism. The objective of this study was to identify and characterize global changes in gene expression in the hippocampus in response to acute stress stimuli, by employing a mouse model of short-term restraint stress. In our experimental design mice were subjected to a one time exposure of restraint stress and the regulation of gene expression in the hippocampus was examined 3, 12 and 24 hours thereafter. Microarray analysis revealed that mice which had undergone acute restraint stress differed from non-stressed controls in global hippocampal transcriptional responses. An up-regulation of transcripts contributing directly or indirectly to neurogenesis and neuronal protection including, Ttr, Rab6, Gh, Prl, Ndufb9 and Ndufa6, was observed. Systems level analyses revealed a significant enrichment for neurogenesis, neuron morphogenesis- and cognitive functions-related biological process terms and pathways. This work further supports the hypothesis that acute stress mediates a positive action on the hippocampus favouring the formation and the preservation of neurons, which will be discussed in the context of current data from the literature

Putative Prostate Cancer Risk SNP in an Androgen Receptor-Binding Site of the Melanophilin Gene Illustrates Enrichment of Risk SNPs in Androgen Receptor Target Sites

Genome-wide association studies have identified genomic loci, whose single-nucleotide polymorphisms (SNPs) predispose to prostate cancer (PCa). However, the mechanisms of most of these variants are largely unknown. We integrated chromatin-immunoprecipitation-coupled sequencing and microarray expression profiling in TMPRSS2-ERG gene rearrangement positive DUCaP cells with the GWAS PCa risk SNPs catalog to identify disease susceptibility SNPs localized within functional androgen receptor-binding sites (ARBSs). Among the 48 GWAS index risk SNPs and 3,917 linked SNPs, 80 were found located in ARBSs. Of these, rs11891426:T>G in an intron of the melanophilin gene (MLPH) was within a novel putative auxiliary AR-binding motif, which is enriched in the neighborhood of canonical androgen-responsive elements. TG exchange attenuated the transcriptional activity of the ARBS in an AR reporter gene assay. The expression of MLPH in primary prostate tumors was significantly lower in those with the G compared with the T allele and correlated significantly with AR protein. Higher melanophilin level in prostate tissue of patients with a favorable PCa risk profile points out a tumor-suppressive effect. These results unravel a hidden link between AR and a functional putative PCa risk SNP, whose allele alteration affects androgen regulation of its host gene MLPH

miR-22 and miR-29a Are Members of the Androgen Receptor Cistrome Modulating LAMC1 and Mcl-1 in Prostate Cancer

The normal prostate as well as early stages and advanced prostate cancer (PCa) require a functional androgen receptor (AR) for growth and survival. The recent discovery of microRNAs (miRNAs) as novel effector molecules of AR disclosed the existence of an intricate network between AR, miRNAs and downstream target genes. In this study DUCaP cells, characterized by high content of wild-type AR and robust AR transcriptional activity, were chosen as the main experimental model. By integrative analysis of chromatin immunoprecipitation-sequencing (ChIP-seq) and microarray expression profiling data, miRNAs putatively bound and significantly regulated by AR were identified. A direct AR regulation of miR-22, miR-29a, and miR-17-92 cluster along with their host genes was confirmed. Interestingly, endogenous levels of miR-22 and miR-29a were found to be reduced in PCa cells expressing AR. In primary tumor samples, miR-22 and miR-29a were less abundant in the cancerous tissue compared with the benign counterpart. This specific expression pattern was associated with a differential DNA methylation of the genomic AR binding sites. The identification of laminin gamma 1 (LAMC1) and myeloid cell leukemia 1 (MCL1) as direct targets of miR-22 and miR-29a, respectively, suggested a tumor-suppressive role of these miRNAs. Indeed, transfection of miRNA mimics in PCa cells induced apoptosis and diminished cell migration and viability. Collectively, these data provide additional information regarding the complex regulatory machinery that guides miRNAs activity in PCa, highlighting an important contribution of miRNAs in the AR signaling

Developmental and extracellular matrix-remodeling processes in rosiglitazone-exposed neonatal rat cardiomyocytes

Objective: The objective of this study was to investigate the effects of rosiglitazone (Avandia®) on gene expression in neonatal rat ventricular myocytes. Materials &amp; methods: Myocytes were exposed to rosiglitazone ex vivo. The two factors examined in the experiment were drug exposure (rosiglitazone and dimethyl sulfoxide vs dimethyl sulfoxide), and length of exposure to drug (½ h, 1 h, 2 h, 4 h, 6 h, 8 h, 12 h, 18 h, 24 h, 36 h and 48 h). Results: Transcripts that were consistently expressed in response to the drug were identified. Cardiovascular system development, extracellular matrix and immune response are represented prominently among the significantly modified gene ontology terms. Conclusion: Hmgcs2, Angptl4, Cpt1a, Cyp1b1, Ech1 and Nqo1 mRNAs were strongly upregulated in cells exposed to rosiglitazone. Enrichment of transcripts involved in cardiac muscle cell differentiation and the extracellular matrix provides a panel of biomarkers for further analysis in the context of adverse cardiac outcomes in humans. Original submitted 15 November 2013; Revision submitted 14 February 2014 </jats:p

Early assessment of KRAS mutation in cfDNA correlates with risk of progression and death in advanced non-small-cell lung cancer

Background Liquid biopsy has the potential to monitor biological effects of treatment. KRAS represents the most commonly mutated oncogene in Caucasian non-small-cell lung cancer (NSCLC). The aim of this study was to explore association of dynamic plasma KRAS genotyping with outcome in advanced NSCLC patients. Methods Advanced NSCLC patients were prospectively enrolled. Plasma samples were collected at baseline (T1), after 3 or 4 weeks, according to treatment schedule (T2) and at first radiological restaging (T3). Patients carrying KRAS mutation in tissue were analysed in plasma with droplet digital PCR. Semi-quantitative index of fractional abundance of mutated allele (MAFA) was used. Results KRAS-mutated cohort included 58 patients, and overall 73 treatments (N = 39 chemotherapy and N = 34 immune checkpoint inhibitors) were followed with longitudinal liquid biopsy. Sensitivity of KRAS detection in plasma at baseline was 48.3% (95% confidence interval (CI): 35.0-61.8). KRAS mutation at T2 was associated with increased probability of experiencing progressive disease as best radiological response (adjusted odds ratio: 7.3; 95% CI: 2.1-25.0, p = 0.0016). Increased MAFA (T1-T2) predicted shorter progression-free survival (adjusted hazard ratio (HR): 2.1; 95% CI: 1.2-3.8, p = 0.0142) and overall survival (adjusted HR: 3.2; 95% CI: 1.2-8.4, p = 0.0168). Conclusions Longitudinal analysis of plasma KRAS mutations correlated with outcome: its early assessment during treatment has great potentialities for monitoring treatment outcome in NSCLC patients

Biomarkers of hippocampal gene expression in a mouse restraint chronic stress model

Objective: Acute stress provides many beneficial effects whereas chronic stress contributes to a variety of human health issues including anxiety, depression, gastrointestinal problems, cardiac disease, sleep disorders and obesity. The goal of this work was to identify, using a rodent model, hippocampal gene signatures associated with prolonged chronic stress representing candidate biomarkers and therapeutic targets for early diagnosis and pharmacological intervention for stress induced disease. Materials & methods: Mice underwent 'restraint stress' over 7 consecutive days and hippocampal gene-expression changes were analyzed at 3, 12 and 24 h following the final restraint treatment. Results: Data indicated that mice exposed to chronic restraint stress exhibit a differential gene-expression profile compared with non-stressed controls. The greatest differences were observed 12 and 24 h following the final stress test. Conclusion: Our study indicated that Gpr88, Ttr, Gh and Tac1 mRNAs were modulated in mice exposed to chronic restraint stress. These transcripts represent a panel of biomarkers and druggable targets for further analysis in the context of chronic stress associated disease in humans