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1.
J Pathol ; 236(3): 278-89, 2015 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-25693838

RESUMEN

Glutamine is conditionally essential in cancer cells, being utilized as a carbon and nitrogen source for macromolecule production, as well as for anaplerotic reactions fuelling the tricarboxylic acid (TCA) cycle. In this study, we demonstrated that the glutamine transporter ASCT2 (SLC1A5) is highly expressed in prostate cancer patient samples. Using LNCaP and PC-3 prostate cancer cell lines, we showed that chemical or shRNA-mediated inhibition of ASCT2 function in vitro decreases glutamine uptake, cell cycle progression through E2F transcription factors, mTORC1 pathway activation and cell growth. Chemical inhibition also reduces basal oxygen consumption and fatty acid synthesis, showing that downstream metabolic function is reliant on ASCT2-mediated glutamine uptake. Furthermore, shRNA knockdown of ASCT2 in PC-3 cell xenografts significantly inhibits tumour growth and metastasis in vivo, associated with the down-regulation of E2F cell cycle pathway proteins. In conclusion, ASCT2-mediated glutamine uptake is essential for multiple pathways regulating the cell cycle and cell growth, and is therefore a putative therapeutic target in prostate cancer.


Asunto(s)
Sistema de Transporte de Aminoácidos ASC/genética , Regulación Neoplásica de la Expresión Génica , Glutamina/metabolismo , Neoplasias de la Próstata/genética , Sistema de Transporte de Aminoácidos ASC/antagonistas & inhibidores , Sistema de Transporte de Aminoácidos ASC/metabolismo , Animales , Transporte Biológico , Ciclo Celular , Línea Celular Tumoral , Proliferación Celular , Regulación hacia Abajo , Ácidos Grasos/metabolismo , Técnicas de Silenciamiento del Gen , Xenoinjertos , Humanos , Masculino , Diana Mecanicista del Complejo 1 de la Rapamicina , Ratones , Ratones Desnudos , Antígenos de Histocompatibilidad Menor , Complejos Multiproteicos/genética , Complejos Multiproteicos/metabolismo , Metástasis de la Neoplasia , Oxígeno/metabolismo , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/patología , Neoplasias de la Próstata/prevención & control , ARN Interferente Pequeño , Serina-Treonina Quinasas TOR/genética , Serina-Treonina Quinasas TOR/metabolismo
2.
BMC Genomics ; 16: 145, 2015 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-25766521

RESUMEN

BACKGROUND: Strand specific RNAseq data is now more common in RNAseq projects. Visualizing RNAseq data has become an important matter in Analysis of sequencing data. The most widely used visualization tool is the UCSC genome browser that introduced the custom track concept that enabled researchers to simultaneously visualize gene expression at a particular locus from multiple experiments. Our objective of the software tool is to provide friendly interface for visualization of RNAseq datasets. RESULTS: This paper introduces a visualization tool (RNASeqBrowser) that incorporates and extends the functionality of the UCSC genome browser. For example, RNASeqBrowser simultaneously displays read coverage, SNPs, InDels and raw read tracks with other BED and wiggle tracks -- all being dynamically built from the BAM file. Paired reads are also connected in the browser to enable easier identification of novel exon/intron borders and chimaeric transcripts. Strand specific RNAseq data is also supported by RNASeqBrowser that displays reads above (positive strand transcript) or below (negative strand transcripts) a central line. Finally, RNASeqBrowser was designed for ease of use for users with few bioinformatic skills, and incorporates the features of many genome browsers into one platform. CONCLUSIONS: The features of RNASeqBrowser: (1) RNASeqBrowser integrates UCSC genome browser and NGS visualization tools such as IGV. It extends the functionality of the UCSC genome browser by adding several new types of tracks to show NGS data such as individual raw reads, SNPs and InDels. (2) RNASeqBrowser can dynamically generate RNA secondary structure. It is useful for identifying non-coding RNA such as miRNA. (3) Overlaying NGS wiggle data is helpful in displaying differential expression and is simple to implement in RNASeqBrowser. (4) NGS data accumulates a lot of raw reads. Thus, RNASeqBrowser collapses exact duplicate reads to reduce visualization space. Normal PC's can show many windows of NGS individual raw reads without much delay. (5) Multiple popup windows of individual raw reads provide users with more viewing space. This avoids existing approaches (such as IGV) which squeeze all raw reads into one window. This will be helpful for visualizing multiple datasets simultaneously. RNASeqBrowser and its manual are freely available at http://www.australianprostatecentre.org/research/software/rnaseqbrowser or http://sourceforge.net/projects/rnaseqbrowser/.


Asunto(s)
Bases de Datos Genéticas , Genoma , Análisis de Secuencia de ARN/métodos , Programas Informáticos , Biología Computacional/métodos , Mutación INDEL/genética , Internet
3.
BMC Genomics ; 16: 1021, 2015 Dec 01.
Artículo en Inglés | MEDLINE | ID: mdl-26626734

RESUMEN

BACKGROUND: Fusion transcripts are found in many tissues and have the potential to create novel functional products. Here, we investigate the genomic sequences around fusion junctions to better understand the transcriptional mechanisms mediating fusion transcription/splicing. We analyzed data from prostate (cancer) cells as previous studies have shown extensively that these cells readily undergo fusion transcription. RESULTS: We used the FusionMap program to identify high-confidence fusion transcripts from RNAseq data. The RNAseq datasets were from our (N = 8) and other (N = 14) clinical prostate tumors with adjacent non-cancer cells, and from the LNCaP prostate cancer cell line that were mock-, androgen- (DHT), and anti-androgen- (bicalutamide, enzalutamide) treated. In total, 185 fusion transcripts were identified from all RNAseq datasets. The majority (76%) of these fusion transcripts were 'read-through chimeras' derived from adjacent genes in the genome. Characterization of sequences at fusion loci were carried out using a combination of the FusionMap program, custom Perl scripts, and the RNAfold program. Our computational analysis indicated that most fusion junctions (76%) use the consensus GT-AG intron donor-acceptor splice site, and most fusion transcripts (85%) maintained the open reading frame. We assessed whether parental genes of fusion transcripts have the potential to form complementary base pairing between parental genes which might bring them into physical proximity. Our computational analysis of sequences flanking fusion junctions at parental loci indicate that these loci have a similar propensity as non-fusion loci to hybridize. The abundance of repetitive sequences at fusion and non-fusion loci was also investigated given that SINE repeats are involved in aberrant gene transcription. We found few instances of repetitive sequences at both fusion and non-fusion junctions. Finally, RT-qPCR was performed on RNA from both clinical prostate tumors and adjacent non-cancer cells (N = 7), and LNCaP cells treated as above to validate the expression of seven fusion transcripts and their respective parental genes. We reveal that fusion transcript expression is similar to the expression of parental genes. CONCLUSIONS: Fusion transcripts maintain the open reading frame, and likely use the same transcriptional machinery as non-fusion transcripts as they share many genomic features at splice/fusion junctions.


Asunto(s)
Regulación Neoplásica de la Expresión Génica , Neoplasias de la Próstata/genética , Sitios de Carácter Cuantitativo , Empalme del ARN , Transcripción Genética , Andrógenos/farmacología , Antineoplásicos Hormonales/farmacología , Biología Computacional/métodos , Secuencia Conservada , Conjuntos de Datos como Asunto , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Masculino , Motivos de Nucleótidos , Sitios de Empalme de ARN , Secuencias Repetitivas de Ácidos Nucleicos
4.
Nucleic Acids Res ; 41(2): 727-37, 2013 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-23221645

RESUMEN

miRDeep and its varieties are widely used to quantify known and novel micro RNA (miRNA) from small RNA sequencing (RNAseq). This article describes miRDeep*, our integrated miRNA identification tool, which is modeled off miRDeep, but the precision of detecting novel miRNAs is improved by introducing new strategies to identify precursor miRNAs. miRDeep* has a user-friendly graphic interface and accepts raw data in FastQ and Sequence Alignment Map (SAM) or the binary equivalent (BAM) format. Known and novel miRNA expression levels, as measured by the number of reads, are displayed in an interface, which shows each RNAseq read relative to the pre-miRNA hairpin. The secondary pre-miRNA structure and read locations for each predicted miRNA are shown and kept in a separate figure file. Moreover, the target genes of known and novel miRNAs are predicted using the TargetScan algorithm, and the targets are ranked according to the confidence score. miRDeep* is an integrated standalone application where sequence alignment, pre-miRNA secondary structure calculation and graphical display are purely Java coded. This application tool can be executed using a normal personal computer with 1.5 GB of memory. Further, we show that miRDeep* outperformed existing miRNA prediction tools using our LNCaP and other small RNAseq datasets. miRDeep* is freely available online at http://www.australianprostatecentre.org/research/software/mirdeep-star.


Asunto(s)
Perfilación de la Expresión Génica , Secuenciación de Nucleótidos de Alto Rendimiento , MicroARNs/química , Análisis de Secuencia de ARN , Programas Informáticos , Línea Celular Tumoral , Humanos , Masculino , MicroARNs/metabolismo , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/metabolismo
5.
BMC Bioinformatics ; 15: 275, 2014 Aug 12.
Artículo en Inglés | MEDLINE | ID: mdl-25117656

RESUMEN

BACKGROUND: Small RNA sequencing is commonly used to identify novel miRNAs and to determine their expression levels in plants. There are several miRNA identification tools for animals such as miRDeep, miRDeep2 and miRDeep*. miRDeep-P was developed to identify plant miRNA using miRDeep's probabilistic model of miRNA biogenesis, but it depends on several third party tools and lacks a user-friendly interface. The objective of our miRPlant program is to predict novel plant miRNA, while providing a user-friendly interface with improved accuracy of prediction. RESULT: We have developed a user-friendly plant miRNA prediction tool called miRPlant. We show using 16 plant miRNA datasets from four different plant species that miRPlant has at least a 10% improvement in accuracy compared to miRDeep-P, which is the most popular plant miRNA prediction tool. Furthermore, miRPlant uses a Graphical User Interface for data input and output, and identified miRNA are shown with all RNAseq reads in a hairpin diagram. CONCLUSIONS: We have developed miRPlant which extends miRDeep* to various plant species by adopting suitable strategies to identify hairpin excision regions and hairpin structure filtering for plants. miRPlant does not require any third party tools such as mapping or RNA secondary structure prediction tools. miRPlant is also the first plant miRNA prediction tool that dynamically plots miRNA hairpin structure with small reads for identified novel miRNAs. This feature will enable biologists to visualize novel pre-miRNA structure and the location of small RNA reads relative to the hairpin. Moreover, miRPlant can be easily used by biologists with limited bioinformatics skills.miRPlant and its manual are freely available at http://www.australianprostatecentre.org/research/software/mirplant or http://sourceforge.net/projects/mirplant/.


Asunto(s)
Biología Computacional/métodos , MicroARNs/genética , Plantas/genética , ARN de Planta/genética , Análisis de Secuencia de ARN , Programas Informáticos , Secuencia de Bases
6.
Biol Chem ; 395(9): 1127-32, 2014 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-25153393

RESUMEN

We assessed whether alternative transcripts (using KLK2, KLK3 and KLK4 as models) are differentially regulated by androgens and anti-androgens as an indicator of prostate cancers as they acquire treatment resistance. Using RNAseq of LNCaP cells treated with dihydrotestosterone, bicalutamide and enzalutamide, we show that the expression of variant KLK transcripts is markedly different to other variant transcripts at those loci. We also reveal that KLK variants are also over 2-fold more highly expressed in prostate cancers compared to their corresponding normal prostate. We propose that androgens and anti-androgens can activate specific variant transcripts of critical prostate cancer genes during treatment resistance.


Asunto(s)
Empalme Alternativo/genética , Antagonistas de Andrógenos/farmacología , Andrógenos/farmacología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Calicreínas/genética , Neoplasias de la Próstata/genética , Empalme Alternativo/efectos de los fármacos , Línea Celular Tumoral , Humanos , Calicreínas/metabolismo , Masculino , Antígeno Prostático Específico/genética , Antígeno Prostático Específico/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo
7.
RNA ; 16(6): 1156-66, 2010 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-20406994

RESUMEN

In humans, more than 30,000 chimeric transcripts originating from 23,686 genes have been identified. The mechanisms and association of chimeric transcripts arising from chromosomal rearrangements with cancer are well established, but much remains unknown regarding the biogenesis and importance of other chimeric transcripts that arise from nongenomic alterations. Recently, a SLC45A3-ELK4 chimera has been shown to be androgen-regulated, and is overexpressed in metastatic or high-grade prostate tumors relative to local prostate cancers. Here, we characterize the expression of a KLK4 cis sense-antisense chimeric transcript, and show other examples in prostate cancer. Using non-protein-coding microarray analyses, we initially identified an androgen-regulated antisense transcript within the 3' untranslated region of the KLK4 gene in LNCaP cells. The KLK4 cis-NAT was validated by strand-specific linker-mediated RT-PCR and Northern blotting. Characterization of the KLK4 cis-NAT by 5' and 3' rapid amplification of cDNA ends (RACE) revealed that this transcript forms multiple fusions with the KLK4 sense transcript. Lack of KLK4 antisense promoter activity using reporter assays suggests that these transcripts are unlikely to arise from a trans-splicing mechanism. 5' RACE and analyses of deep sequencing data from LNCaP cells treated +/-androgens revealed six high-confidence sense-antisense chimeras of which three were supported by the cDNA databases. In this study, we have shown complex gene expression at the KLK4 locus that might be a hallmark of cis sense-antisense chimeric transcription.


Asunto(s)
ADN sin Sentido/genética , Variación Genética , Calicreínas/genética , Neoplasias de la Próstata/genética , Transcripción Genética , Antígenos de Neoplasias/genética , Quimera/genética , Mapeo Cromosómico , ADN de Neoplasias/genética , Exones , Reordenamiento Génico , Estudio de Asociación del Genoma Completo , Humanos , Masculino , Proteínas de Transporte de Membrana/genética , Neoplasias de la Próstata/patología , Proteína Elk-4 del Dominio ets/genética
9.
Prostate ; 70(10): 1134-45, 2010 Jul 01.
Artículo en Inglés | MEDLINE | ID: mdl-20503398

RESUMEN

BACKGROUND: Relaxin, a potent peptide hormone of the insulin-like family normally produced and secreted by the human prostate, is upregulated in castrate resistant prostate cancer progression. In various tissues, relaxin increases angiogenesis and cell motility through upregulation of vascular endothelial growth factor, matrix metalloproteases, and nitric oxide, and therefore maybe an attractive target for cancer therapeutics. METHODS: To examine the role of relaxin in prostate cancer progression, LNCaP cells stably transfected with relaxin (LNCaP(RLN)) were used to form xenograft tumors, and microarray expression analysis was subsequently performed to determine novel pathways regulated by relaxin. Prostate cancer tissue microarrays from patient samples were stained by immunohistochemistry for further validation and correlation of the findings. RESULTS: Expression analysis identified novel relaxin regulated pathways, including the ProtocadherinY (PCDHY)/Wnt pathway. PCDHY, which upregulates Wnt11, has previously been shown to stabilize beta-catenin, causing beta-catenin to translocate from the cytoplasmic membrane to the nucleus and initiate TCF-mediated signaling. LNCaP(RLN) xenografts exhibit increased PCDHY expression and increased cytoplasmic localization of beta-catenin, suggesting relaxin directs Wnt11 overexpression through PCDHY upregulation. Similarly, prostate cancer samples from patients who have undergone androgen ablation have increased Wnt11 expression, which is further upregulated in castrate resistant tissues. Like relaxin, Wnt11, and PCDHY are negatively regulated by androgens, and further analysis indicated that the overexpression of relaxin results in dysregulation of androgen-regulated genes. CONCLUSIONS: These data suggest that prostate cancer cell motility and altered androgen receptor activity attributed to relaxin may be mediated in part by Wnt11.


Asunto(s)
Cadherinas/metabolismo , Neoplasias Hormono-Dependientes/metabolismo , Neoplasias de la Próstata/metabolismo , Relaxina/metabolismo , Animales , Northern Blotting , Cadherinas/genética , Línea Celular Tumoral , Humanos , Inmunohistoquímica , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Neoplasias Hormono-Dependientes/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , Neoplasias de la Próstata/genética , ARN/química , ARN/genética , Receptores Acoplados a Proteínas G/biosíntesis , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Péptidos/biosíntesis , Receptores de Péptidos/genética , Receptores de Péptidos/metabolismo , Relaxina/biosíntesis , Relaxina/genética , Transducción de Señal , Estadísticas no Paramétricas , Transfección , Trasplante Heterólogo , Regulación hacia Arriba , Proteína Wnt1/genética , Proteína Wnt1/metabolismo , beta Catenina/metabolismo
10.
Prostate ; 70(3): 239-51, 2010 Feb 15.
Artículo en Inglés | MEDLINE | ID: mdl-19790237

RESUMEN

BACKGROUND: De novo androgen synthesis and subsequent androgen receptor (AR) activation has recently been shown to contribute to castration-resistant prostate cancer (CRPC) progression. Herein we provide evidence that fatty acids (FA) can trigger androgen synthesis within steroid starved prostate cancer (CaP) tumor cells. METHODS: Tumoral FA and steroid levels were assessed by GC-MS and LC-MS, respectively. Profiles of genes and proteins involved in FA activation of steroidogenesis were assessed by fluorescence microscopy, immunohistochemistry, microarray expression profiling and Western blot analysis. RESULTS: In human CaP tissues the levels of proteins responsible for FA activation of steroid synthesis were observed to be altered during progression to CRPC. Further investigating this mechanism in LNCaP cells, we demonstrate that specific FA, arachidonic acid, is synthesized in an androgen-dependent and AR-mediated manner. Arachidonic acid is known to induce steroidogenic acute regulatory protein (StAR) in steroidogenic cells. When bound to hormone sensitive lipase (HSL), StAR shuttles free cholesterol into the mitochondria for downstream conversion into androgens. We show that arachidonic acid induces androgen production in steroid starved LNCaP cells coincidently in the same conditions that HSL and StAR are predominantly localized in the mitochondria. Furthermore, their activities are verified by a functional increase in mitochondrial uptake of cholesterol in this steroid starved environment. CONCLUSIONS: We propose that this characterized arachidonic acid induced steroidogenesis mechanism significantly contributes to the activation of AR in CRPC progression and therefore recommend that fatty acid pathways be targeted therapeutically in progressing CaP.


Asunto(s)
Antagonistas de Andrógenos/uso terapéutico , Antineoplásicos Hormonales/uso terapéutico , Ácido Araquidónico/metabolismo , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/patología , Receptores Androgénicos/metabolismo , Esteroides/biosíntesis , Andrógenos/deficiencia , Andrógenos/metabolismo , Animales , Ácido Araquidónico/farmacología , Línea Celular Tumoral , Colesterol/farmacocinética , Progresión de la Enfermedad , Resistencia a Medicamentos , Humanos , Masculino , Ratones , Ratones Desnudos , Mitocondrias/metabolismo , Proteínas de Neoplasias/metabolismo , Trasplante de Neoplasias , Fosfoproteínas/metabolismo , Neoplasias de la Próstata/tratamiento farmacológico , Esterol Esterasa/metabolismo , Distribución Tisular , Trasplante Heterólogo , Regulación hacia Arriba
11.
Cancer Metab ; 8: 11, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32577235

RESUMEN

BACKGROUND: Metabolic reprograming, non-mutational epigenetic changes, increased cell plasticity, and multidrug tolerance are early hallmarks of therapy resistance in cancer. In this temporary, therapy-tolerant state, cancer cells are highly sensitive to ferroptosis, a form of regulated cell death that is caused by oxidative stress through excess levels of iron-dependent peroxidation of polyunsaturated fatty acids (PUFA). However, mechanisms underpinning therapy-induced ferroptosis hypersensitivity remain to be elucidated. METHODS: We used quantitative single-cell imaging of fluorescent metabolic probes, transcriptomics, proteomics, and lipidomics to perform a longitudinal analysis of the adaptive response to androgen receptor-targeted therapies (androgen deprivation and enzalutamide) in prostate cancer (PCa). RESULTS: We discovered that cessation of cell proliferation and a robust reduction in bioenergetic processes were associated with multidrug tolerance and a strong accumulation of lipids. The gain in lipid biomass was fueled by enhanced lipid uptake through cargo non-selective (macropinocytosis, tunneling nanotubes) and cargo-selective mechanisms (lipid transporters), whereas de novo lipid synthesis was strongly reduced. Enzalutamide induced extensive lipid remodeling of all major phospholipid classes at the expense of storage lipids, leading to increased desaturation and acyl chain length of membrane lipids. The rise in membrane PUFA levels enhanced membrane fluidity and lipid peroxidation, causing hypersensitivity to glutathione peroxidase (GPX4) inhibition and ferroptosis. Combination treatments against AR and fatty acid desaturation, lipase activities, or growth medium supplementation with antioxidants or PUFAs altered GPX4 dependence. CONCLUSIONS: Our work provides mechanistic insight into processes of lipid metabolism that underpin the acquisition of therapy-induced GPX4 dependence and ferroptosis hypersensitivity to standard of care therapies in PCa. It demonstrates novel strategies to suppress the therapy-tolerant state that may have potential to delay and combat resistance to androgen receptor-targeted therapies, a currently unmet clinical challenge of advanced PCa. Since enhanced GPX4 dependence is an adaptive phenotype shared by several types of cancer in response to different therapies, our work might have universal implications for our understanding of metabolic events that underpin resistance to cancer therapies.

12.
Mol Oncol ; 14(1): 105-128, 2020 01.
Artículo en Inglés | MEDLINE | ID: mdl-31630475

RESUMEN

Kallikrein-related peptidase 14 (KLK14) is one of the several secreted KLK serine proteases involved in prostate cancer (PCa) pathogenesis. While relatively understudied, recent reports have identified KLK14 as overexpressed during PCa development. However, the modulation of KLK14 expression during PCa progression and the molecular and biological functions of this protease in the prostate tumor microenvironment remain unknown. To determine the modulation of KLK14 expression during PCa progression, we analyzed the expression levels of KLK14 in patient samples using publicly available databases and immunohistochemistry. In order to delineate the molecular mechanisms involving KLK14 in PCa progression, we integrated proteomic, transcriptomic, and in vitro assays with the goal to identify substrates, related-signaling pathways, and functional roles of this protease. We showed that KLK14 expression is elevated in advanced PCa, and particularly in metastasis. Additionally, KLK14 levels were found to be decreased in PCa tissues from patients responsive to neoadjuvant therapy compared to untreated patients. Furthermore, we also identified that KLK14 expression reoccurred in patients who developed castrate-resistant PCa. The combination of proteomic and transcriptomic analysis as well as functional assays revealed several new KLK14 substrates (agrin, desmoglein 2, vitronectin, laminins) and KLK14-regulated genes (Interleukin 32, midkine, SRY-Box 9), particularly an involvement of the mitogen-activated protein kinase 1 and interleukin 1 receptor pathways, and an involvement of KLK14 in the regulation of cellular migration, supporting its involvement in aggressive features of PCa progression. In conclusion, our work showed that KLK14 expression is associated with the development of aggressive PCa suggesting that targeting this protease could offer a novel route to limit the progression of prostate tumors. Additional work is necessary to determine the benefits and implications of targeting/cotargeting KLK14 in PCa as well as to determine the potential use of KLK14 expression as a predictor of PCa aggressiveness or response to treatment.


Asunto(s)
Movimiento Celular/genética , Regulación Neoplásica de la Expresión Génica/genética , Calicreínas/metabolismo , Metástasis de la Neoplasia/genética , Neoplasias de la Próstata/metabolismo , Línea Celular Tumoral , Proliferación Celular/genética , Cromatografía Líquida de Alta Presión , Bases de Datos Genéticas , Regulación hacia Abajo , Humanos , Inmunohistoquímica , Calicreínas/genética , Masculino , Terapia Neoadyuvante , Neoplasias de la Próstata/enzimología , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/patología , Proteómica , Transducción de Señal/genética , Espectrometría de Masas en Tándem , Transcriptoma , Microambiente Tumoral/genética , Regulación hacia Arriba
13.
Mol Cancer Res ; 17(5): 1166-1179, 2019 05.
Artículo en Inglés | MEDLINE | ID: mdl-30808729

RESUMEN

De novo lipogenesis is a well-described androgen receptor (AR)-regulated metabolic pathway that supports prostate cancer tumor growth by providing fuel, membrane material, and steroid hormone precursor. In contrast, our current understanding of lipid supply from uptake of exogenous lipids and its regulation by AR is limited, and exogenous lipids may play a much more significant role in prostate cancer and disease progression than previously thought. By applying advanced automated quantitative fluorescence microscopy, we provide the most comprehensive functional analysis of lipid uptake in cancer cells to date and demonstrate that treatment of AR-positive prostate cancer cell lines with androgens results in significantly increased cellular uptake of fatty acids, cholesterol, and low-density lipoprotein particles. Consistent with a direct, regulatory role of AR in this process, androgen-enhanced lipid uptake can be blocked by the AR-antagonist enzalutamide, but is independent of proliferation and cell-cycle progression. This work for the first time comprehensively delineates the lipid transporter landscape in prostate cancer cell lines and patient samples by analysis of transcriptomics and proteomics data, including the plasma membrane proteome. We show that androgen exposure or deprivation regulates the expression of multiple lipid transporters in prostate cancer cell lines and tumor xenografts and that mRNA and protein expression of lipid transporters is enhanced in bone metastatic disease when compared with primary, localized prostate cancer. Our findings provide a strong rationale to investigate lipid uptake as a therapeutic cotarget in the fight against advanced prostate cancer in combination with inhibitors of lipogenesis to delay disease progression and metastasis. IMPLICATIONS: Prostate cancer exhibits metabolic plasticity in acquiring lipids from uptake and lipogenesis at different disease stages, indicating potential therapeutic benefit by cotargeting lipid supply.


Asunto(s)
Andrógenos/farmacología , Neoplasias Óseas/metabolismo , Neoplasias Óseas/secundario , Metabolismo de los Lípidos/efectos de los fármacos , Neoplasias de la Próstata/metabolismo , Neoplasias Óseas/genética , Línea Celular Tumoral , Colesterol/metabolismo , Progresión de la Enfermedad , Ácidos Grasos/metabolismo , Regulación Neoplásica de la Expresión Génica , Redes Reguladoras de Genes/efectos de los fármacos , Humanos , Lipoproteínas LDL/metabolismo , Masculino , Microscopía Fluorescente , Neoplasias de la Próstata/genética , Receptores Androgénicos/metabolismo , Transducción de Señal
14.
Artículo en Inglés | MEDLINE | ID: mdl-31379747

RESUMEN

Androgen deprivation therapy (ADT) is the standard treatment for advanced prostate cancer (PCa), yet many patients relapse with lethal metastatic disease. With this loss of androgens, increased cell plasticity has been observed as an adaptive response to ADT. This includes gain of invasive and migratory capabilities, which may contribute to PCa metastasis. Hyperinsulinemia, which develops as a side-effect of ADT, has been associated with increased tumor aggressiveness and faster treatment failure. We investigated the direct effects of insulin in PCa cells that may contribute to this progression. We measured cell migration and invasion induced by insulin using wound healing and transwell assays in a range of PCa cell lines of variable androgen dependency (LNCaP, 22RV1, DuCaP, and DU145 cell lines). To determine the molecular events driving insulin-induced invasion we used transcriptomics, quantitative real time-PCR, and immunoblotting in three PCa cell lines. Insulin increased invasiveness of PCa cells, upregulating Forkhead Box Protein C2 (FOXC2), and activating key PCa cell plasticity mechanisms including gene changes consistent with epithelial-to-mesenchymal transition (EMT) and a neuroendocrine phenotype. Additionally, analysis of publicly available clinical PCa tumor data showed metastatic prostate tumors demonstrate a positive correlation between insulin receptor expression and the EMT transcription factor FOXC2. The insulin receptor is not suitable to target clinically however, our data shows that actions of insulin in PCa cells may be suppressed by inhibiting downstream signaling molecules, PI3K and ERK1/2. This study identifies for the first time, a mechanism for insulin-driven cancer cell motility and supports the concept that targeting insulin signaling at the level of the PCa tumor may extend the therapeutic efficacy of ADT.

15.
Oncogene ; 38(13): 2436, 2019 03.
Artículo en Inglés | MEDLINE | ID: mdl-30510231

RESUMEN

Following the publication of the above article, the authors noted an error in Figure 4, panel B. The colours of the localized and mCRPC samples were accidentally switched. The authors have corrected the colour scheme and added a key to the figure. They have also updated the colour scheme of panel C, both bars are now red instead of one red and one blue. The authors wish to apologize for any inconvenience caused.

16.
Oncogene ; 38(7): 913-934, 2019 02.
Artículo en Inglés | MEDLINE | ID: mdl-30194451

RESUMEN

The propensity of cancer cells to transition between epithelial and mesenchymal phenotypic states via the epithelial-mesenchymal transition (EMT) program can regulate metastatic processes, cancer progression, and treatment resistance. Transcriptional investigations using reversible models of EMT, revealed the mesenchymal-to-epithelial reverting transition (MErT) to be enriched in clinical samples of metastatic castrate resistant prostate cancer (mCRPC). From this enrichment, a metastasis-derived gene signature was identified that predicted more rapid cancer relapse and reduced survival across multiple human carcinoma types. Additionally, the transcriptional profile of MErT is not a simple mirror image of EMT as tumour cells retain a transcriptional "memory" following a reversible EMT. This memory was also enriched in mCRPC samples. Cumulatively, our studies reveal the transcriptional profile of epithelial-mesenchymal plasticity and highlight the unique transcriptional properties of MErT. Furthermore, our findings provide evidence to support the association of epithelial plasticity with poor clinical outcomes in multiple human carcinoma types.


Asunto(s)
Transición Epitelial-Mesenquimal , Neoplasias de la Próstata Resistentes a la Castración/metabolismo , Neoplasias de la Próstata Resistentes a la Castración/mortalidad , Línea Celular Tumoral , Supervivencia sin Enfermedad , Humanos , Masculino , Metástasis de la Neoplasia , Neoplasias de la Próstata Resistentes a la Castración/clasificación , Neoplasias de la Próstata Resistentes a la Castración/patología , Tasa de Supervivencia
17.
Mol Cell Endocrinol ; 478: 84-96, 2018 12 15.
Artículo en Inglés | MEDLINE | ID: mdl-30053582

RESUMEN

Mammalian sex determination hinges on sexually dimorphic transcriptional programs in developing fetal gonads. A comprehensive view of these programs is crucial for understanding the normal development of fetal testes and ovaries and the etiology of human disorders of sex development (DSDs), many of which remain unexplained. Using strand-specific RNA-sequencing, we characterized the mouse fetal gonadal transcriptome from 10.5 to 13.5 days post coitum, a key time window in sex determination and gonad development. Our dataset benefits from a greater sensitivity, accuracy and dynamic range compared to microarray studies, allows global dynamics and sex-specificity of gene expression to be assessed, and provides a window to non-transcriptional events such as alternative splicing. Spliceomic analysis uncovered female-specific regulation of Lef1 splicing, which may contribute to the enhanced WNT signaling activity in XX gonads. We provide a user-friendly visualization tool for the complete transcriptomic and spliceomic dataset as a resource for the field.


Asunto(s)
Empalme Alternativo/genética , Perfilación de la Expresión Génica , ARN Mensajero/genética , Procesos de Determinación del Sexo/genética , Animales , Femenino , Feto/metabolismo , Regulación del Desarrollo de la Expresión Génica , Redes Reguladoras de Genes , Gónadas/embriología , Gónadas/metabolismo , Factor de Unión 1 al Potenciador Linfoide/genética , Factor de Unión 1 al Potenciador Linfoide/metabolismo , Masculino , Ratones , ARN Mensajero/metabolismo , Reproducibilidad de los Resultados , Análisis de Secuencia de ARN , Caracteres Sexuales , Factores de Tiempo , Activación Transcripcional/genética
18.
Sci Rep ; 7(1): 16862, 2017 12 04.
Artículo en Inglés | MEDLINE | ID: mdl-29203868

RESUMEN

Short tandem repeats (STRs) are repetitive sequences of a polymorphic stretch of two to six nucleotides. We hypothesized that STRs are associated with prostate cancer development and/or progression. We undertook RNA sequencing analysis of prostate tumors and adjacent non-malignant cells to identify polymorphic STRs that are readily expressed in these cells. Most of the expressed STRs in the clinical samples mapped to intronic and intergenic DNA. Our analysis indicated that three of these STRs (TAAA-ACTG2, TTTTG-TRIB1, and TG-PCA3) are polymorphic and differentially expressed in prostate tumors compared to adjacent non-malignant cells. TG-PCA3 STR expression was repressed by the anti-androgen drug enzalutamide in prostate cancer cells. Genetic analysis of prostate cancer patients and healthy controls (N > 2,000) showed a significant association of the most common 11 repeat allele of TG-PCA3 STR with prostate cancer risk (OR = 1.49; 95% CI 1.11-1.99; P = 0.008). A significant association was also observed with aggressive disease (OR = 2.00; 95% CI 1.06-3.76; P = 0.031) and high mortality rates (HR = 3.0; 95% CI 1.03-8.77; P = 0.045). We propose that TG-PCA3 STR has both diagnostic and prognostic potential for prostate cancer. We provided a proof of concept to be applied to other RNA sequencing datasets to identify disease-associated STRs for future clinical exploratory studies.


Asunto(s)
Antígenos de Neoplasias/genética , Repeticiones de Microsatélite/genética , Neoplasias de la Próstata/patología , ARN Largo no Codificante/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Alelos , Secuencia de Bases , Estudios de Casos y Controles , Genotipo , Humanos , Estimación de Kaplan-Meier , Masculino , Persona de Mediana Edad , Oportunidad Relativa , Pronóstico , Modelos de Riesgos Proporcionales , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/mortalidad , Factores de Riesgo
19.
Mol Cell Endocrinol ; 420: 159-68, 2016 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-26499396

RESUMEN

Simultaneous expression of highly homologous RLN1 and RLN2 genes in prostate impairs their accurate delineation. We used PacBio SMRT sequencing and RNA-Seq in LNCaP cells in order to dissect the expression of RLN1 and RLN2 variants. We identified a novel fusion transcript comprising the RLN1 and RLN2 genes and found evidence of its expression in the normal and prostate cancer tissues. The RLN1-RLN2 fusion putatively encodes RLN2 isoform with the deleted secretory signal peptide. The identification of the fusion transcript provided information to determine unique RLN1-RLN2 fusion and RLN1 regions. The RLN1-RLN2 fusion was co-expressed with RLN1 in LNCaP cells, but the two gene products were inversely regulated by androgens. We showed that RLN1 is underrepresented in common PCa cell lines in comparison to normal and PCa tissue. The current study brings a highly relevant update to the relaxin field, and will encourage further studies of RLN1 and RLN2 in PCa and broader.


Asunto(s)
Proteínas de Fusión Oncogénica/genética , Neoplasias de la Próstata/genética , Relaxina/genética , Andrógenos/farmacología , Línea Celular Tumoral , Exones/genética , Humanos , Masculino , Proteínas de Fusión Oncogénica/metabolismo , Sistemas de Lectura Abierta/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Relaxina/metabolismo , Análisis de Secuencia de ARN
20.
Oncotarget ; 6(41): 43944-63, 2015 Dec 22.
Artículo en Inglés | MEDLINE | ID: mdl-26733491

RESUMEN

As part of an anti-cancer natural product drug discovery program, we recently identified eusynstyelamide B (EB), which displayed cytotoxicity against MDA-MB-231 breast cancer cells (IC50 = 5 µM) and induced apoptosis. Here, we investigated the mechanism of action of EB in cancer cell lines of the prostate (LNCaP) and breast (MDA-MB-231). EB inhibited cell growth (IC50 = 5 µM) and induced a G2 cell cycle arrest, as shown by a significant increase in the G2/M cell population in the absence of elevated levels of the mitotic marker phospho-histone H3. In contrast to MDA-MB-231 cells, EB did not induce cell death in LNCaP cells when treated for up to 10 days. Transcript profiling and Ingenuity Pathway Analysis suggested that EB activated DNA damage pathways in LNCaP cells. Consistent with this, CHK2 phosphorylation was increased, p21CIP1/WAF1 was up-regulated and CDC2 expression strongly reduced by EB. Importantly, EB caused DNA double-strand breaks, yet did not directly interact with DNA. Analysis of topoisomerase II-mediated decatenation discovered that EB is a novel topoisomerase II poison.


Asunto(s)
Antineoplásicos/farmacología , Neoplasias de la Mama/patología , ADN-Topoisomerasas de Tipo II/efectos de los fármacos , Indoles/farmacología , Neoplasias de la Próstata/patología , Animales , Western Blotting , Puntos de Control del Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Daño del ADN/efectos de los fármacos , Femenino , Humanos , Masculino , Análisis de Secuencia por Matrices de Oligonucleótidos , Reacción en Cadena en Tiempo Real de la Polimerasa , Urocordados
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