RESUMO
Nearly all prostate cancer deaths are from metastatic castration-resistant prostate cancer (mCRPC), but there have been few whole-genome sequencing (WGS) studies of this disease state. We performed linked-read WGS on 23 mCRPC biopsy specimens and analyzed cell-free DNA sequencing data from 86 patients with mCRPC. In addition to frequent rearrangements affecting known prostate cancer genes, we observed complex rearrangements of the AR locus in most cases. Unexpectedly, these rearrangements include highly recurrent tandem duplications involving an upstream enhancer of AR in 70%-87% of cases compared with <2% of primary prostate cancers. A subset of cases displayed AR or MYC enhancer duplication in the context of a genome-wide tandem duplicator phenotype associated with CDK12 inactivation. Our findings highlight the complex genomic structure of mCRPC, nominate alterations that may inform prostate cancer treatment, and suggest that additional recurrent events in the non-coding mCRPC genome remain to be discovered.
Assuntos
Neoplasias de Próstata Resistentes à Castração/patologia , Receptores Androgênicos/genética , Sequenciamento Completo do Genoma , Idoso , Anilidas/uso terapêutico , Quinases Ciclina-Dependentes/genética , Quinases Ciclina-Dependentes/metabolismo , Elementos Facilitadores Genéticos/genética , Duplicação Gênica , Rearranjo Gênico , Genes myc , Loci Gênicos , Haplótipos , Humanos , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , PTEN Fosfo-Hidrolase/genética , Fenótipo , Antígeno Prostático Específico/sangue , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/genética , Inibidores de Proteínas Quinases/uso terapêutico , Piridinas/uso terapêuticoRESUMO
Exitron splicing (EIS) creates a cryptic intron (called an exitron) within a protein-coding exon to increase proteome diversity. EIS is poorly characterized, but emerging evidence suggests a role for EIS in cancer. Through a systematic investigation of EIS across 33 cancers from 9,599 tumor transcriptomes, we discovered that EIS affected 63% of human coding genes and that 95% of those events were tumor specific. Notably, we observed a mutually exclusive pattern between EIS and somatic mutations in their affected genes. Functionally, we discovered that EIS altered known and novel cancer driver genes for causing gain- or loss-of-function, which promotes tumor progression. Importantly, we identified EIS-derived neoepitopes that bind to major histocompatibility complex (MHC) class I or II. Analysis of clinical data from a clear cell renal cell carcinoma cohort revealed an association between EIS-derived neoantigen load and checkpoint inhibitor response. Our findings establish the importance of considering EIS alterations when nominating cancer driver events and neoantigens.
Assuntos
Epitopos/genética , Éxons/genética , Perfilação da Expressão Gênica , Íntrons/genética , Neoplasias/genética , Oncogenes , Splicing de RNA/genética , Sequência de Aminoácidos , Linhagem Celular , Estudos de Coortes , Humanos , Mutação/genéticaRESUMO
Prostate cancer (PC) is the most frequently diagnosed malignancy and a leading cause of cancer deaths in US men. Many PC cases metastasize and develop resistance to systemic hormonal therapy, a stage known as castration-resistant prostate cancer (CRPC). Therefore, there is an urgent need to develop effective therapeutic strategies for CRPC. Traditional drug discovery pipelines require significant time and capital input, which highlights a need for novel methods to evaluate the repositioning potential of existing drugs. Here, we present a computational framework to predict drug sensitivities of clinical CRPC tumors to various existing compounds and identify treatment options with high potential for clinical impact. We applied this method to a CRPC patient cohort and nominated drugs to combat resistance to hormonal therapies including abiraterone and enzalutamide. The utility of this method was demonstrated by nomination of multiple drugs that are currently undergoing clinical trials for CRPC. Additionally, this method identified the tetracycline derivative COL-3, for which we validated higher efficacy in an isogenic cell line model of enzalutamide-resistant vs. enzalutamide-sensitive CRPC. In enzalutamide-resistant CRPC cells, COL-3 displayed higher activity for inhibiting cell growth and migration, and for inducing G1-phase cell cycle arrest and apoptosis. Collectively, these findings demonstrate the utility of a computational framework for independent validation of drugs being tested in CRPC clinical trials, and for nominating drugs with enhanced biological activity in models of enzalutamide-resistant CRPC. The efficiency of this method relative to traditional drug development approaches indicates a high potential for accelerating drug development for CRPC.
Assuntos
Neoplasias de Próstata Resistentes à Castração , Masculino , Humanos , Neoplasias de Próstata Resistentes à Castração/patologia , Nitrilas/farmacologia , Descoberta de Drogas , Castração , Resistencia a Medicamentos Antineoplásicos , Receptores Androgênicos/metabolismoRESUMO
Prostate cancer is the second leading cause of male cancer death in the United States. The androgen receptor (AR) transcription factor is a master regulator of normal glandular homeostasis in the prostate, as well as growth and survival of prostate cancer cells. Therefore, AR-targeted therapies are effective for improving overall survival of patients with advanced prostate cancer that is incurable by surgery or radiation. However, prostate cancer will inevitably progress on AR-targeted therapies to a castration-resistant prostate cancer (CRPC) phenotype that accounts for virtually all prostate cancer-specific death. mRNA transcript variants of the AR gene are expressed in CRPC cells and can be translated to produce AR variant (AR-V) proteins that function as ligand-independent, constitutively active transcription factors. AR-Vs are able to support growth of CRPC cells by promoting expression of AR target genes that are normally suppressed by AR-targeted therapies. Knowledge of mechanisms that govern expression of AR-Vs is incomplete. Studies have shown genomic rearrangements of the AR gene underlie expression of diverse AR-Vs in certain CRPC tumors, but post-transcriptional processes represent a broader regulatory mechanism for expression of AR-Vs in CRPC. This review focuses on alternative splicing, 3' end processing, miRNA-mediated mRNA repression, of AR and AR-V expression and the potential these mechanisms hold as therapeutic targets for CRPC.
Assuntos
Processamento Alternativo , Antagonistas de Receptores de Andrógenos/uso terapêutico , Regulação Neoplásica da Expressão Gênica , Terapia de Alvo Molecular , Neoplasias da Próstata/tratamento farmacológico , RNA Mensageiro/genética , Receptores Androgênicos/química , Animais , Rearranjo Gênico , Humanos , Masculino , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Receptores Androgênicos/genéticaRESUMO
A hallmark of prostate cancer progression is dysregulation of lipid metabolism via overexpression of fatty acid synthase (FASN), a key enzyme in de novo fatty acid synthesis. Metastatic castration-resistant prostate cancer (mCRPC) develops resistance to inhibitors of androgen receptor (AR) signaling through a variety of mechanisms, including the emergence of the constitutively active AR variant V7 (AR-V7). Here, we developed an FASN inhibitor (IPI-9119) and demonstrated that selective FASN inhibition antagonizes CRPC growth through metabolic reprogramming and results in reduced protein expression and transcriptional activity of both full-length AR (AR-FL) and AR-V7. Activation of the reticulum endoplasmic stress response resulting in reduced protein synthesis was involved in IPI-9119-mediated inhibition of the AR pathway. In vivo, IPI-9119 reduced growth of AR-V7-driven CRPC xenografts and human mCRPC-derived organoids and enhanced the efficacy of enzalutamide in CRPC cells. In human mCRPC, both FASN and AR-FL were detected in 87% of metastases. AR-V7 was found in 39% of bone metastases and consistently coexpressed with FASN. In patients treated with enzalutamide and/or abiraterone FASN/AR-V7 double-positive metastases were found in 77% of cases. These findings provide a compelling rationale for the use of FASN inhibitors in mCRPCs, including those overexpressing AR-V7.
Assuntos
Lipogênese , Proteínas de Neoplasias/metabolismo , Neoplasias de Próstata Resistentes à Castração/metabolismo , Receptores Androgênicos/metabolismo , Transdução de Sinais , Animais , Linhagem Celular Tumoral , Inibidores Enzimáticos/farmacologia , Ácido Graxo Sintase Tipo I/antagonistas & inibidores , Ácido Graxo Sintase Tipo I/genética , Ácido Graxo Sintase Tipo I/metabolismo , Humanos , Masculino , Camundongos , Metástase Neoplásica , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Receptores Androgênicos/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Nucleotide-based drugs, such as antisense oligonucleotides (ASOs), have unique advantages in treating human diseases as they provide virtually unlimited ability to target any gene. However, their clinical translation faces many challenges, one of which is poor delivery to the target tissue in vivo. This problem is particularly evident in solid tumors. Here, we functionalized liposomes with a tumor-homing and -penetrating peptide, iRGD, as a carrier of an ASO against androgen receptor (AR) for prostate cancer treatment. The iRGD-liposomes exhibited a high loading efficiency of AR-ASO, and an efficient knockdown of AR gene products was achieved in vitro, including AR splice variants. In vivo, iRGD-liposomes significantly increased AR-ASO accumulation in the tumor tissue and decreased AR expression relative to free ASOs in prostate tumors established as subcutaneous xenografts. Similar results were obtained with intra-tibial xenografts modeling metastasis to bones, the predominant site of metastasis for prostate cancer. In treatment studies, iRGD-liposomes markedly improved the AR-ASO efficacy in suppressing the growth of both subcutaneous xenografts and intra-tibial xenografts. The inhibitory effect on tumor growth was also significantly prolonged by the delivery of the AR-ASO in the iRGD-liposomes. Meanwhile, iRGD-liposomes did not increase ASO accumulation or toxicity in healthy organs. Overall, we provide here a delivery system that can significantly increase ASO accumulation and efficacy in solid tumors. These benefits are achieved without significant side effects, providing a way to increase the antitumor efficacy of ASOs.
RESUMO
Resistance to androgen receptor (AR)-targeted therapies in prostate cancer (PC) is a major clinical problem. A key mechanism of treatment resistance in advanced PC is the generation of alternatively spliced forms of the AR termed AR variants (AR-Vs) that are refractory to targeted agents and drive tumour progression. Our understanding of how AR-Vs function is limited due to difficulties in distinguishing their discriminate activities from full-length AR (FL-AR). Here we report the development of a novel CRISPR-derived cell line which is a derivative of CWR22Rv1 cells, called CWR22Rv1-AR-EK, that has lost expression of FL-AR, but retains all endogenous AR-Vs. From this, we show that AR-Vs act unhindered by loss of FL-AR to drive cell growth and expression of androgenic genes. Global transcriptomics demonstrate that AR-Vs drive expression of a cohort of DNA damage response genes and depletion of AR-Vs sensitises cells to ionising radiation. Moreover, we demonstrate that AR-Vs interact with PARP1 and PARP2 and are dependent upon their catalytic function for transcriptional activation. Importantly, PARP blockade compromises expression of AR-V-target genes and reduces growth of CRPC cell lines suggesting a synthetic lethality relationship between AR-Vs and PARP, advocating the use of PARP inhibitors in AR-V positive PC.
Assuntos
Sistemas CRISPR-Cas , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Receptores Androgênicos/genética , Algoritmos , Linhagem Celular Tumoral , Proliferação de Células , Dano ao DNA , Reparo do DNA , Ensaios de Seleção de Medicamentos Antitumorais , Técnicas Genéticas , Humanos , Lentivirus , Masculino , Receptores Androgênicos/biossíntese , Análise de Sequência de RNA , TranscriptomaRESUMO
Androgens and the androgen receptor (AR) play crucial roles in the biology of normal and diseased prostate tissue, including prostate cancer (PCa). This dependence is evidenced by the use of androgen depletion therapy (ADT) as the primary treatment for locally advanced, metastatic, or relapsed PCa. This dependence is further evidenced by the various mechanisms employed by PCa cells to re-activate the AR to circumvent the growth-inhibitory effects of ADT. Re-activation of the AR during ADT is central to the disease evolving into the lethal castration resistant PCa (CRPC) phenotype, which is responsible for nearly all PCa mortality. Thus, understanding the regulation of AR and AR signaling is important for understanding the development and progression of PCa. This understanding provides the foundation for development of newer approaches for targeting CRPC therapeutically.
Assuntos
Neoplasias da Próstata/metabolismo , Receptores Androgênicos/metabolismo , Androgênios/metabolismo , Progressão da Doença , Humanos , Masculino , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Transdução de SinaisRESUMO
BACKGROUND: Insertions and deletions (indels) are a major class of genomic variation associated with human disease. Indels are primarily detected from DNA sequencing (DNA-seq) data but their transcriptional consequences remain unexplored due to challenges in discriminating medium-sized and large indels from splicing events in RNA-seq data. RESULTS: Here, we developed transIndel, a splice-aware algorithm that parses the chimeric alignments predicted by a short read aligner and reconstructs the mid-sized insertions and large deletions based on the linear alignments of split reads from DNA-seq or RNA-seq data. TransIndel exhibits competitive or superior performance over eight state-of-the-art indel detection tools on benchmarks using both synthetic and real DNA-seq data. Additionally, we applied transIndel to DNA-seq and RNA-seq datasets from 333 primary prostate cancer patients from The Cancer Genome Atlas (TCGA) and 59 metastatic prostate cancer patients from AACR-PCF Stand-Up- To-Cancer (SU2C) studies. TransIndel enhanced the taxonomy of DNA- and RNA-level alterations in prostate cancer by identifying recurrent FOXA1 indels as well as exitron splicing in genes implicated in disease progression. CONCLUSIONS: Our study demonstrates that transIndel is a robust tool for elucidation of medium- and large-sized indels from DNA-seq and RNA-seq data. Including RNA-seq in indel discovery efforts leads to significant improvements in sensitivity for identification of med-sized and large indels missed by DNA-seq, and reveals non-canonical RNA-splicing events in genes associated with disease pathology.
Assuntos
Análise Mutacional de DNA , Mutação INDEL , Análise de Sequência de RNA , Éxons/genética , Perfilação da Expressão Gênica , Humanos , Masculino , Metástase Neoplásica , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Splicing de RNA/genéticaRESUMO
BACKGROUND: Mutations or truncation of the ligand-binding domain (LBD) of androgen receptor (AR) underlie treatment resistance for prostate cancer (PCa). Thus, targeting the AR N-terminal domain (NTD) could overcome such resistance. METHODS: Luciferase reporter assays after transient transfection of various DNA constructs were used to assess effects of E1A proteins on AR-mediated transcription. Immunofluorescence microscopy and subcellular fractionation were applied to assess intracellular protein localization. Immunoprecipitation and mammalian two-hybrid assays were used to detect protein-protein interactions. qRT-PCR was employed to determine RNA levels. Western blotting was used to detect protein expression in cells. Effects of adenoviruses on prostate cancer cell survival were evaluated with CellTiter-Glo assays. RESULTS: Adenovirus 12 E1A (E1A12) binds specifically to the AR. Interestingly, the full-length E1A12 (266 aa) preferentially binds to full-length AR, while the small E1A12 variant (235 aa) interacts more strongly with AR-V7. E1A12 promotes AR nuclear translocation, likely through mediating intramolecular AR NTD-LBD interactions. In the nucleus, AR and E1A12 co-expression in AR-null PCa cells results in E1A12 redistribution from nuclear foci containing CBX4 (also known as Pc2), suggesting a preferential AR-E1A12 interaction over other E1A12 interactors. E1A12 represses AR-mediated transcription in reporter gene assays and endogenous AR target genes such as ATAD2 and MYC in AR-expressing PCa cells. AR-expressing PCa cells are more sensitive to death induced by a recombinant adenovirus expressing E1A12 (Ad-E1A12) than AR-deficient PCa cells, which could be attributed to the increased viral replication promoted by androgen stimulation. Targeting the AR by E1A12 promotes apoptosis in PCa cells that express the full-length AR or C-terminally truncated AR variants. Importantly, inhibition of mTOR signaling that blocks the expression of anti-apoptotic proteins markedly augments Ad-E1A12-induced apoptosis of AR-expressing cells. Mechanistically, Ad-E1A12 infection triggers apoptotic response while activating the PI3K-AKT-mTOR signaling axis; thus, mTOR inhibition enhances apoptosis in AR-expressing PCa cells infected by Ad-E1A12. CONCLUSION: Ad12 E1A inhibits AR-mediated transcription and suppresses PCa cell survival, suggesting that targeting the AR by E1A12 might have therapeutic potential for treating advanced PCa with heightened AR signaling.
Assuntos
Proteínas E1A de Adenovirus/metabolismo , Neoplasias da Próstata/terapia , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Proteínas E1A de Adenovirus/genética , Adenovírus Humanos/genética , Adenovírus Humanos/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/fisiologia , Células HEK293 , Humanos , Masculino , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/virologia , Domínios Proteicos , Ativação Transcricional , TransfecçãoRESUMO
BACKGROUND: With almost 30,000 deaths per year, prostate cancer is the second-leading cause of cancer-related death in men. Androgen Deprivation Therapy (ADT) has been the corner stone of prostate cancer treatment for decades. However, despite an initial response of prostate cancer to ADT, this eventually fails and the tumors recur, resulting in Castration Resistant Prostate Cancer (CRPC). Triptolide, a diterpene triepoxide, has been tested for its anti-tumor properties in a number of cancers for over a decade. Owing to its poor solubility in aqueous medium, its clinical application had been limited. To circumvent this problem, we have synthesized a water-soluble pro-drug of triptolide, Minnelide, that is currently being evaluated in a Phase 1 clinical trial against gastrointestinal tumors. In the current study, we assessed the therapeutic potential of Minnelide and its active compound triptolide against androgen dependent prostate cancer both in vitro as well as in vivo. METHODS: Cell viability was measured by a MTT based assay after treating prostate cancer cells with multiple doses of triptolide. Apoptotic cell death was measured using a caspase 3/7 activity. Androgen Receptor (AR) promoter-binding activity was evaluated by using luciferase reporter assay. For evaluating the effect in vivo, 22Rv1 cells were implanted subcutaneously in animals, following which, treatment was started with 0.21 mg/kg Minnelide. RESULTS: Our study showed that treatment with triptolide induced apoptotic cell death in CRPC cells. Triptolide treatment inhibited AR transcriptional activity and decreased the expression of AR and its splice variants both at the mRNA and the protein level. Our studies show that triptolide inhibits nuclear translocation of Sp1, resulting in its decreased transcriptional activity leading to downregulation of AR and its splice variants in prostate cancer cells. In vivo, Minnelide (0.21 mg/kg) regressed subcutaneous tumors derived from CRPC 22RV1 at our study endpoint. Our animal studies further confirmed that Minnelide was more efficacious than the standard of care therapies, Docetaxel and Enzalutamide. CONCLUSION: Our study indicates that Minnelide is very effective as a therapeutic option against CRPC at a dose that is currently tolerated by patients in the ongoing clinical trials. Prostate 77: 584-596, 2017. © 2017 Wiley Periodicals, Inc.
Assuntos
Regulação Neoplásica da Expressão Gênica , Organofosfatos/farmacologia , Fenantrenos/farmacologia , Neoplasias de Próstata Resistentes à Castração/metabolismo , Isoformas de Proteínas/biossíntese , Receptores Androgênicos/biossíntese , Animais , Linhagem Celular Tumoral , Diterpenos , Relação Dose-Resposta a Droga , Compostos de Epóxi , Humanos , Masculino , Camundongos , Camundongos Nus , Organofosfatos/uso terapêutico , Fenantrenos/uso terapêutico , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/genética , Distribuição Aleatória , Receptores Androgênicos/genética , Carga Tumoral/efeitos dos fármacos , Carga Tumoral/fisiologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Androgen receptor (AR) variants (AR-Vs) expressed in prostate cancer (PCa) lack the AR ligand binding domain (LBD) and function as constitutively active transcription factors. AR-V expression in patient tissues or circulating tumor cells is associated with resistance to AR-targeting endocrine therapies and poor outcomes. Here, we investigated the mechanisms governing chromatin binding of AR-Vs with the goal of identifying therapeutic vulnerabilities. By chromatin immunoprecipitation and sequencing (ChIP-seq) and complementary biochemical experiments, we show that AR-Vs display a binding preference for the same canonical high-affinity androgen response elements (AREs) that are preferentially engaged by AR, albeit with lower affinity. Dimerization was an absolute requirement for constitutive AR-V DNA binding and transcriptional activation. Treatment with the bromodomain and extraterminal (BET) inhibitor JQ1 resulted in inhibition of AR-V chromatin binding and impaired AR-V driven PCa cell growth in vitro and in vivo. Importantly, this was associated with a novel JQ1 action of down-regulating AR-V transcript and protein expression. Overall, this study demonstrates that AR-Vs broadly restore AR chromatin binding events that are otherwise suppressed during endocrine therapy, and provides pre-clinical rationale for BET inhibition as a strategy for inhibiting expression and chromatin binding of AR-Vs in PCa.
Assuntos
Cromatina/metabolismo , Resistencia a Medicamentos Antineoplásicos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Neoplasias da Próstata/genética , Receptores Androgênicos/metabolismo , Ativação Transcricional/efeitos dos fármacos , Animais , Antineoplásicos/uso terapêutico , Azepinas/farmacologia , Benzamidas , Linhagem Celular , Linhagem Celular Tumoral , Dimerização , Masculino , Camundongos Nus , Nitrilas , Feniltioidantoína/análogos & derivados , Feniltioidantoína/uso terapêutico , Neoplasias da Próstata/metabolismo , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Receptores Androgênicos/genética , Elementos de Resposta , Triazóis/farmacologiaRESUMO
Androgen receptor (AR) target genes direct development and survival of the prostate epithelial lineage, including prostate cancer (PCa). Thus, endocrine therapies that inhibit the AR ligand-binding domain (LBD) are effective in treating PCa. AR transcriptional reactivation is central to resistance, as evidenced by the efficacy of AR retargeting in castration-resistant PCa (CRPC) with next-generation endocrine therapies abiraterone and enzalutamide. However, resistance to abiraterone and enzalutamide limits this efficacy in most men, and PCa remains the second-leading cause of male cancer deaths. Here we show that AR gene rearrangements in CRPC tissues underlie a completely androgen-independent, yet AR-dependent, resistance mechanism. We discovered intragenic AR gene rearrangements in CRPC tissues, which we modeled using transcription activator-like effector nuclease (TALEN)-mediated genome engineering. This modeling revealed that these AR gene rearrangements blocked full-length AR synthesis, but promoted expression of truncated AR variant proteins lacking the AR ligand-binding domain. Furthermore, these AR variant proteins maintained the constitutive activity of the AR transcriptional program and a CRPC growth phenotype independent of full-length AR or androgens. These findings demonstrate that AR gene rearrangements are a unique resistance mechanism by which AR transcriptional activity can be uncoupled from endocrine regulation in CRPC.
Assuntos
Rearranjo Gênico , Neoplasias da Próstata/genética , Engenharia de Proteínas/métodos , Receptores Androgênicos/genética , Sequência de Aminoácidos , Androstenos , Androstenóis/uso terapêutico , Animais , Sequência de Bases , Benzamidas , Western Blotting , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica , Redes Reguladoras de Genes , Humanos , Hibridização in Situ Fluorescente , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Dados de Sequência Molecular , Nitrilas , Análise de Sequência com Séries de Oligonucleotídeos , Orquiectomia , Feniltioidantoína/análogos & derivados , Feniltioidantoína/uso terapêutico , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Receptores Androgênicos/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
The androgen receptor (AR) is a transcription factor that has a pivotal role in the occurrence and progression of prostate cancer. The AR is activated by androgens that bind to its ligand-binding domain (LBD), causing the transcription factor to enter the nucleus and interact with genes via its conserved DNA-binding domain (DBD). Treatment for prostate cancer involves reducing androgen production or using anti-androgen drugs to block the interaction of hormones with the AR-LBD. Eventually the disease changes into a castration-resistant form of PCa where LBD mutations render anti-androgens ineffective or where constitutively active AR splice variants, lacking the LBD, become overexpressed. Recently, we identified a surfaced exposed pocket on the AR-DBD as an alternative drug-target site for AR inhibition. Here, we demonstrate that small molecules designed to selectively bind the pocket effectively block transcriptional activity of full-length and splice variant AR forms at low to sub-micromolar concentrations. The inhibition is lost when residues involved in drug interactions are mutated. Furthermore, the compounds did not impede nuclear localization of the AR and blocked interactions with chromatin, indicating the interference of DNA binding with the nuclear form of the transcription factor. Finally, we demonstrate the inhibition of gene expression and tumor volume in mouse xenografts. Our results indicate that the AR-DBD has a surface site that can be targeted to inhibit all forms of the AR, including enzalutamide-resistant and constitutively active splice variants and thus may serve as a potential avenue for the treatment of recurrent and metastatic prostate cancer.
Assuntos
Antagonistas de Receptores de Andrógenos/farmacologia , Imidazóis/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Receptores Androgênicos/fisiologia , Tiazóis/farmacologia , Transporte Ativo do Núcleo Celular , Sequência de Aminoácidos , Animais , Sítios de Ligação , Núcleo Celular/metabolismo , Cromatina/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Células MCF-7 , Masculino , Camundongos Nus , Dados de Sequência Molecular , Terapia de Alvo Molecular , Neoplasias da Próstata/patologia , Ligação Proteica , Isoformas de Proteínas/química , Isoformas de Proteínas/fisiologia , Receptores Androgênicos/química , Transcrição Gênica , Ativação Transcricional , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Personal genome assembly is a critical process when studying tumor genomes and other highly divergent sequences. The accuracy of downstream analyses, such as RNA-seq and ChIP-seq, can be greatly enhanced by using personal genomic sequences rather than standard references. Unfortunately, reads sequenced from these types of samples often have a heterogeneous mix of various subpopulations with different variants, making assembly extremely difficult using existing assembly tools. To address these challenges, we developed SHEAR (Sample Heterogeneity Estimation and Assembly by Reference; http://vk.cs.umn.edu/SHEAR), a tool that predicts SVs, accounts for heterogeneous variants by estimating their representative percentages, and generates personal genomic sequences to be used for downstream analysis. RESULTS: By making use of structural variant detection algorithms, SHEAR offers improved performance in the form of a stronger ability to handle difficult structural variant types and better computational efficiency. We compare against the lead competing approach using a variety of simulated scenarios as well as real tumor cell line data with known heterogeneous variants. SHEAR is shown to successfully estimate heterogeneity percentages in both cases, and demonstrates an improved efficiency and better ability to handle tandem duplications. CONCLUSION: SHEAR allows for accurate and efficient SV detection and personal genomic sequence generation. It is also able to account for heterogeneous sequencing samples, such as from tumor tissue, by estimating the subpopulation percentage for each heterogeneous variant.
Assuntos
Software , Algoritmos , Genômica , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Internet , Análise de Sequência de RNA/normas , Interface Usuário-ComputadorRESUMO
The androgen receptor (AR) and estrogen receptor alpha (ERα) are steroid receptor transcription factors with critical roles in the development and progression of prostate and breast cancers. Advances in the understanding of mechanisms underlying the ligand-dependent activation of these transcription factors have contributed to the development of small molecule inhibitors that block AR and ERα actions. These inhibitors include competitive antagonists and degraders that directly bind the ligand binding domains of these receptors, luteinizing hormone releasing hormone (LHRH) analogs that suppress gonadal synthesis of testosterone or estrogen, and drugs that block specific enzymes required for biosynthesis of testosterone or estrogen. However, resistance to these therapies is frequent, and is often driven by selection for tumor cells with alterations in the AR or ESR1 genes and/or alternatively spliced AR or ESR1 mRNAs that encode variant forms AR or ERα. While most investigations involving AR have been within the context of prostate cancer, and the majority of investigations involving ERα have been within the context of breast cancer, important roles for AR have been elucidated in breast cancer, and important roles for ERα have been elucidated in prostate cancer. Here, we will discuss the roles of AR and ERα in breast and prostate cancers, outline the effects of gene- and mRNA-level alterations in AR and ESR1 on progression of these diseases, and identify strategies that are being developed to target these alterations therapeutically.
Assuntos
Neoplasias da Mama , Receptor alfa de Estrogênio , Neoplasias da Próstata , Receptores Androgênicos , Humanos , Receptores Androgênicos/metabolismo , Receptores Androgênicos/genética , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Masculino , Receptor alfa de Estrogênio/metabolismo , Receptor alfa de Estrogênio/genética , Feminino , Animais , Processamento AlternativoRESUMO
The timing and fitness effect of somatic copy number alterations (SCNA) in cancer evolution remains poorly understood. Here we present a framework to determine the timing of a clonal SCNA that encompasses multiple gains. This involves calculating the proportion of time from its last gain to the onset of population expansion (lead time) as well as the proportion of time prior to its first gain (initiation time). Our method capitalizes on the observation that a genomic segment, while in a specific copy number (CN) state, accumulates point mutations proportionally to its CN. Analyzing 184 whole genome sequenced samples from 75 patients across five tumor types, we commonly observe late gains following early initiating events, occurring just before the clonal expansion relevant to the sampling. These include gains acquired after genome doubling in more than 60% of cases. Notably, mathematical modeling suggests that late clonal gains may contain final-expansion drivers. Lastly, SCNAs bolster mutational diversification between subpopulations, exacerbating the circle of proliferation and increasing heterogeneity.
Assuntos
Variações do Número de Cópias de DNA , Mutação Puntual , Humanos , Variações do Número de Cópias de DNA/genética , Mutação , Cognição , Exercício FísicoRESUMO
Androgen receptor (AR) drives prostate cancer (PC) growth and progression, and targeting AR signaling is the mainstay of pharmacological therapies for PC. Resistance develops relatively fast as a result of refueled AR activity. A major gap in the field is the lack of understanding of targetable mechanisms that induce persistent AR expression in castrate-resistant PC (CRPC). This study uncovers an unexpected function of active Stat5 signaling, a known promoter of PC growth and clinical progression, as a potent inducer of AR gene transcription. Stat5 suppression inhibited AR gene transcription in preclinical PC models and reduced the levels of wild-type, mutated, and truncated AR proteins. Pharmacological Stat5 inhibition by a specific small-molecule Stat5 inhibitor down-regulated Stat5-inducible genes as well as AR and AR-regulated genes and suppressed PC growth. This work introduces the concept of Stat5 as an inducer of AR gene transcription in PC. Pharmacological Stat5 inhibitors may represent a new strategy for suppressing AR and CRPC growth.
Assuntos
Neoplasias de Próstata Resistentes à Castração , Receptores Androgênicos , Masculino , Humanos , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/metabolismo , Transdução de Sinais , Transcrição Gênica , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão GênicaRESUMO
B7-H3 (CD276) is a transmembrane glycoprotein of the B7 immune checkpoint superfamily that has emerged as a promising therapeutic target. To better understand the applicability of B7-H3-directed therapies, we analyzed 156,791 samples comprising 50 cancer types to interrogate the clinical, genomic, transcriptomic, and immunologic correlates of B7-H3 mRNA expression. DNA (592-gene/whole-exome) and RNA (whole-transcriptome) sequencing was performed from samples submitted to Caris Life Sciences. B7-H3 high versus low expression was based on top and bottom quartiles for each cancer type. Patients' overall survival was determined from insurance claims data. Pathway analysis was performed using gene set enrichment analyses. Immune cell fractions were inferred using quanTIseq. B7-H3 is expressed across several human malignancies including prostate, pancreatic, ovarian, and lung cancers. High B7-H3 expression is associated with differences in overall survival, possibly indicating a prognostic role of B7-H3 for some cancers. When examining molecular features across all cancer types, we did not identify recurrent associations between B7-H3 expression and genetic alterations in TP53, RB1, and KRAS. However, we find consistent enrichment of epithelial-to-mesenchymal transition, Wnt, TGFß, and Notch signaling pathways. In addition, tumors with high B7-H3 expression are associated with greater proportions of M1 macrophages, but lower fractions of CD8+ T cells. We have begun to define the genomic, transcriptomic, clinical, and immunologic features associated with B7-H3 expression in 50 cancer types. We report novel clinical and molecular features of B7-H3-high tumors which may inform how current B7-H3 therapeutics should be deployed and prioritized. SIGNIFICANCE: B7-H3-targeting therapeutics have shown promising results in initial clinical trials. In this pan-cancer analysis of B7-H3 mRNA expression, we found that B7-H3 exhibits robust expression in many common cancer types. These results may inform further development of B7-H3-targeting therapeutics and may guide clinical decisions for patients with limited treatment options.
Assuntos
Antígenos B7 , Neoplasias , Humanos , Antígenos B7/genética , Antígenos B7/metabolismo , Neoplasias/genética , Neoplasias/imunologia , Neoplasias/mortalidade , Neoplasias/terapia , Neoplasias/metabolismo , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Regulação Neoplásica da Expressão Gênica , Prognóstico , Masculino , FemininoRESUMO
The impact of variations in the three-dimensional structure of the genome has been recognized, but solid cancer tissue studies are limited. Here, we performed integrated deep Hi-C sequencing with matched whole-genome sequencing, whole-genome bisulfite sequencing, 5-hydroxymethylcytosine (5hmC) sequencing and RNA sequencing across a cohort of 80 biopsy samples from patients with metastatic castration-resistant prostate cancer. Dramatic differences were present in gene expression, 5-methylcytosine/5hmC methylation and in structural variation versus mutation rate between A and B (open and closed) chromatin compartments. A subset of tumors exhibited depleted regional chromatin contacts at the AR locus, linked to extrachromosomal circular DNA (ecDNA) and worse response to AR signaling inhibitors. We also identified topological subtypes associated with stark differences in methylation structure, gene expression and prognosis. Our data suggested that DNA interactions may predispose to structural variant formation, exemplified by the recurrent TMPRSS2-ERG fusion. This comprehensive integrated sequencing effort represents a unique clinical tumor resource.