RESUMO
There are diverse phenotypes of castration-resistant prostate cancer, including neuroendocrine disease, that vary in their sensitivity to drug treatment. The efficacy of BET and CBP/p300 inhibitors in prostate cancer is attributed, at least in part, to their ability to decrease androgen receptor (AR) signalling. However, the activity of BET and CBP/p300 inhibitors in prostate cancers that lack the AR is unclear. In this study, we showed that BRD4, CBP, and p300 were co-expressed in AR-positive and AR-null prostate cancer. A combined inhibitor of these three proteins, NEO2734, reduced the growth of both AR-positive and AR-null organoids, as measured by changes in viability, size, and composition. NEO2734 treatment caused consistent transcriptional downregulation of cell cycle pathways. In neuroendocrine models, NEO2734 treatment reduced ASCL1 levels and other neuroendocrine markers, and reduced tumour growth in vivo. Collectively, these results show that epigenome-targeted inhibitors cause decreased growth and phenotype-dependent disruption of lineage regulators in neuroendocrine prostate cancer, warranting further development of compounds with this activity in the clinic. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.
Assuntos
Proteína p300 Associada a E1A , Receptores Androgênicos , Transdução de Sinais , Masculino , Humanos , Receptores Androgênicos/metabolismo , Receptores Androgênicos/genética , Animais , Proteína p300 Associada a E1A/metabolismo , Proteína p300 Associada a E1A/genética , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias da Próstata/patologia , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Regulação Neoplásica da Expressão Gênica , Camundongos , Ensaios Antitumorais Modelo de Xenoenxerto , Proteínas que Contêm Bromodomínio , Proteína de Ligação a CREBRESUMO
BACKGROUND: There are relatively few widely used models of prostate cancer compared to other common malignancies. This impedes translational prostate cancer research because the range of models does not reflect the diversity of disease seen in clinical practice. In response to this challenge, research laboratories around the world have been developing new patient-derived models of prostate cancer, including xenografts, organoids, and tumor explants. METHODS: In May 2023, we held a workshop at the Monash University Prato Campus for researchers with expertise in establishing and using a variety of patient-derived models of prostate cancer. This review summarizes our collective ideas on how patient-derived models are currently being used, the common challenges, and future opportunities for maximizing their usefulness in prostate cancer research. RESULTS: An increasing number of patient-derived models for prostate cancer are being developed. Despite their individual limitations and varying success rates, these models are valuable resources for exploring new concepts in prostate cancer biology and for preclinical testing of potential treatments. Here we focus on the need for larger collections of models that represent the changing treatment landscape of prostate cancer, robust readouts for preclinical testing, improved in vitro culture conditions, and integration of the tumor microenvironment. Additional priorities include ensuring model reproducibility, standardization, and replication, and streamlining the exchange of models and data sets among research groups. CONCLUSIONS: There are several opportunities to maximize the impact of patient-derived models on prostate cancer research. We must develop large, diverse and accessible cohorts of models and more sophisticated methods for emulating the intricacy of patient tumors. In this way, we can use the samples that are generously donated by patients to advance the outcomes of patients in the future.
Assuntos
Neoplasias da Próstata , Masculino , Humanos , Reprodutibilidade dos Testes , Neoplasias da Próstata/terapia , Neoplasias da Próstata/patologia , Próstata/patologia , Organoides/patologia , Xenoenxertos , Microambiente TumoralRESUMO
BACKGROUND: Cyclin-dependent kinase 9 (CDK9) stimulates oncogenic transcriptional pathways in cancer and CDK9 inhibitors have emerged as promising therapeutic candidates. METHODS: The activity of an orally bioavailable CDK9 inhibitor, CDKI-73, was evaluated in prostate cancer cell lines, a xenograft mouse model, and patient-derived tumor explants and organoids. Expression of CDK9 was evaluated in clinical specimens by mining public datasets and immunohistochemistry. Effects of CDKI-73 on prostate cancer cells were determined by cell-based assays, molecular profiling and transcriptomic/epigenomic approaches. RESULTS: CDKI-73 inhibited proliferation and enhanced cell death in diverse in vitro and in vivo models of androgen receptor (AR)-driven and AR-independent models. Mechanistically, CDKI-73-mediated inhibition of RNA polymerase II serine 2 phosphorylation resulted in reduced expression of BCL-2 anti-apoptotic factors and transcriptional defects. Transcriptomic and epigenomic approaches revealed that CDKI-73 suppressed signaling pathways regulated by AR, MYC, and BRD4, key drivers of dysregulated transcription in prostate cancer, and reprogrammed cancer-associated super-enhancers. These latter findings prompted the evaluation of CDKI-73 with the BRD4 inhibitor AZD5153, a combination that was synergistic in patient-derived organoids and in vivo. CONCLUSION: Our work demonstrates that CDK9 inhibition disrupts multiple oncogenic pathways and positions CDKI-73 as a promising therapeutic agent for prostate cancer, particularly aggressive, therapy-resistant subtypes.
Assuntos
Quinase 9 Dependente de Ciclina , Epigênese Genética , Neoplasias da Próstata , Masculino , Quinase 9 Dependente de Ciclina/antagonistas & inibidores , Humanos , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Neoplasias da Próstata/metabolismo , Animais , Camundongos , Epigênese Genética/efeitos dos fármacos , Linhagem Celular Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto , Proliferação de Células/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Receptores Androgênicos/metabolismo , Receptores Androgênicos/genética , Transcrição Gênica/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacosRESUMO
BACKGROUND: Prostate cancer is dependent on androgen receptor (AR) signaling, and androgen deprivation therapy (ADT) has proven effective in targeting prostate cancer. However, castration-resistant prostate cancer (CRPC) eventually emerges. AR signaling inhibitors (ARSI) have been also used, but resistance to these agents develops due to genetic AR alterations and epigenetic dysregulation. METHODS: In this study, we investigated the role of OCT1, a member of the OCT family, in an AR-positive CRPC patient-derived xenograft established from a patient with resistance to ARSI and chemotherapy. We conducted a genome-wide analysis chromatin immunoprecipitation followed by sequencing and bioinformatic analyses using public database. RESULTS: Genome-wide analysis of OCT1 target genes in PDX 201.1 A revealed distinct OCT1 binding sites compared to treatment-naïve cells. Bioinformatic analyses revealed that OCT1-regulated genes were associated with cell migration and immune system regulation. In particular, C-terminal Binding Protein 2 (CTBP2), an OCT1/AR target gene, was correlated with poor prognosis and immunosuppressive effects in the tumor microenvironment. Metascape revealed that CTBP2 knockdown affects genes related to the immune response to bacteria. Furthermore, TISIDB analysis suggested the relationship between CTBP2 expression and immune cell infiltration in prostate cancer, suggesting that it may contribute to immune evasion in CRPC. CONCLUSIONS: Our findings shed light on the genome-wide network of OCT1 and AR in AR-positive CRPC and highlight the potential role of CTBP2 in immune response and tumor progression. Targeting CTBP2 may represent a promising therapeutic approach for aggressive AR-positive CRPC. Further validation will be required to explore novel therapeutic strategies for CRPC management.
Assuntos
Oxirredutases do Álcool , Proteínas Correpressoras , Regulação Neoplásica da Expressão Gênica , Fator 1 de Transcrição de Octâmero , Neoplasias de Próstata Resistentes à Castração , Receptores Androgênicos , Masculino , Humanos , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Neoplasias de Próstata Resistentes à Castração/metabolismo , Receptores Androgênicos/metabolismo , Receptores Androgênicos/genética , Camundongos , Animais , Fator 1 de Transcrição de Octâmero/metabolismo , Fator 1 de Transcrição de Octâmero/genética , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Regulação para Cima , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/genética , Microambiente Tumoral , Transdução de SinaisRESUMO
Estrogen receptor alpha (ERα) activity is associated with increased cancer cell proliferation. Studies aiming to understand the impact of ERα on cancer-associated phenotypes have largely been limited to its transcriptional activity. Herein, we demonstrate that ERα coordinates its transcriptional output with selective modulation of mRNA translation. Importantly, translational perturbations caused by depletion of ERα largely manifest as "translational offsetting" of the transcriptome, whereby amounts of translated mRNAs and corresponding protein levels are maintained constant despite changes in mRNA abundance. Transcripts whose levels, but not polysome association, are reduced following ERα depletion lack features which limit translation efficiency including structured 5'UTRs and miRNA target sites. In contrast, mRNAs induced upon ERα depletion whose polysome association remains unaltered are enriched in codons requiring U34-modified tRNAs for efficient decoding. Consistently, ERα regulates levels of U34-modifying enzymes and thereby controls levels of U34-modified tRNAs. These findings unravel a hitherto unprecedented mechanism of ERα-dependent orchestration of transcriptional and translational programs that may be a pervasive mechanism of proteome maintenance in hormone-dependent cancers.
Assuntos
Neoplasias da Mama/genética , Receptor alfa de Estrogênio/genética , Regulação Neoplásica da Expressão Gênica , Polirribossomos/genética , Biossíntese de Proteínas , RNA Mensageiro/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Proliferação de Células , Receptor alfa de Estrogênio/metabolismo , Feminino , Humanos , Células MCF-7 , Polirribossomos/metabolismo , RNA Mensageiro/metabolismo , Transdução de Sinais , Ativação TranscricionalRESUMO
It is well known that hydrogen peroxide (H2O2) is a signaling molecule essential for vital physiological reactions in mammalian cells, such as cell survival, intercellular communication, and cancer metabolism. However, to fully understand the function of H2O2, it is critical to monitor its intracellular and/or extracellular concentrations. Current techniques implemented to address this need require large sample volumes, expensive instrumentation, and long sample preparation and analysis times, inapplicable to inline or online monitoring. In this paper, a new integrated microfluidic device capable of overcoming these limitations is demonstrated for the colorimetric detection of extracellular hydrogen peroxide H2O2. The device contains an optical waveguide to determine absorbance changes and micromixers to enable complete mixing of reagents using a passive approach. This novel H2O2-sensing device has allowed the detection of H2O2 in the range of 0.5-60 µM with a detection limit of 167 ± 5.8 nM and a sensitivity of 13.5 ± 0.1 AU/mM. Proof of concept of the device was demonstrated by quantifying H2O2 release from benign prostatic epithelial (BPH-1) cells upon stimulation with phorbol 12-myristate 13-acetate (PMA). Results show that this integrated device can be potentially utilized to continuously monitor cell-released metabolites autonomously without constant human supervision during the process. Furthermore, this can be achieved without interfering with the cell culture conditions, as only a very small volume of conditioned media (less than 0.4 µL), and not the cells, is required.
Assuntos
Colorimetria , Peróxido de Hidrogênio , Animais , Humanos , Peróxido de Hidrogênio/análise , Dispositivos Lab-On-A-Chip , Acetato de TetradecanoilforbolRESUMO
OBJECTIVES: To perform a systematic review and meta-analysis of the literature to understand the variation in the reporting of neuroendocrine staining and determine the influence of reporting neuroendocrine staining at diagnosis on patient outcomes. METHODS: Medical databases were searched to identify studies in which adenocarcinoma specimens were stained with any of the following four neuroendocrine markers: chromogranin A (CgA), neuron-specific enolase (NSE), synaptophysin and CD56. The prevalence of neuroendocrine staining and correlation of the prevalence of neuroendocrine staining to patient outcomes were analysed using a random-effects model. All statistical tests were two-sided. RESULTS: Sixty-two studies spanning 7616 patients were analysed. The pooled prevalence for the most common marker, CgA (41%), was similar to that of NSE (39%) and higher than that of synaptophysin (31%). The prevalence of CgA staining was significantly influenced by reporting criteria, where objective thresholds reduced the variation in prevalence to 26%. No correlation was found between CgA prevalence and tumour grade. Patients positive for CgA staining using objective criteria had more rapid biochemical progression (hazard ratio [HR] 1.98, 95% confidence interval [CI] 1.49 to 2.65) and poorer prostate cancer-specific survival (HR 7.03, 95% CI 2.55 to 19.39) compared to negative patients, even among those with low-risk cancers. CONCLUSION: Discrepancies in the reported prevalence of neuroendocrine cells in adenocarcinoma are driven by the inconsistent scoring criteria. This study unequivocally demonstrates that when neuroendocrine cell staining is assessed with objective criteria it identifies patients with poor clinical outcomes. Future studies are needed to determine the exact quantifiable thresholds for use in reporting neuroendocrine cell staining to identify patients at higher risk of progression.
Assuntos
Adenocarcinoma , Células Neuroendócrinas , Neoplasias da Próstata , Adenocarcinoma/patologia , Biomarcadores Tumorais/análise , Cromogranina A , Humanos , Masculino , Células Neuroendócrinas/química , Células Neuroendócrinas/patologia , Fosfopiruvato Hidratase , Neoplasias da Próstata/patologia , SinaptofisinaRESUMO
PURPOSE OF REVIEW: Many clinical trials are currently underway to target the epigenome of castration-resistant prostate cancer. In this review, we summarize the major epigenetic alterations that occur during prostate cancer progression, describe their biological consequences, and highlight potential of therapies that target epigenetic regulators for use in patients. RECENT FINDINGS: Epigenetic alterations frequently occur in tumour suppressor genes, DNA repair genes, and genes that regulate cell proliferation and differentiation. Unlike genetic alterations, epigenetic changes are reversible, making them promising targets for cancer therapy. Epigenetic regulators can be divided into three broad groups: writers, readers, and erasers , each with specific drug targets that are being assessed in phase I and II clinical trials for prostate cancer. CBP/p300, and BRD4 are coregulators of the androgen receptor and inhibit androgen signalling, making bromodomain extra-terminal inhibitors and CBP/p300 inhibitors attractive targets in prostate cancer. Enhancer of zeste homolog 2, a histone methyltransferase, is also a potential target in castrate-resistant prostate cancer. An emerging direction is to combine epigenetic inhibitors with other compounds to enhance their efficacy. SUMMARY: Preclinical studies indicate that the epigenome is a potential target in prostate cancer, and clinical trials are testing multiple agents that target the epigenome in different ways. However, the process of translating these therapies into the clinic is ongoing and none have yet been approved for castrate-resistant prostate cancer.
Assuntos
Neoplasias de Próstata Resistentes à Castração , Neoplasias da Próstata , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/uso terapêutico , Proliferação de Células , Epigênese Genética , Humanos , Masculino , Proteínas Nucleares/genética , Proteínas Nucleares/uso terapêutico , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Fatores de Transcrição/genética , Fatores de Transcrição/uso terapêuticoRESUMO
Amplifications of the androgen receptor (AR) occur in up to 80% of men with castration-resistant prostate cancer (CRPC). Recent studies highlighted that these amplifications not only span the AR gene but usually encompass a distal enhancer. This represents a newly recognised, non-coding mechanism of resistance to AR-directed therapies, including enzalutamide. To study disease progression before and after AR amplification, we used tumour samples from a castrate-sensitive primary tumour and castrate-resistant metastasis of the same patient. For subsequent functional and genomic studies, we established serially transplantable patient-derived xenografts (PDXs). Whole genome sequencing showed that alterations associated with poor prognosis, such as TP53 and PTEN loss, existed before androgen deprivation therapy, followed by co-amplification of the AR gene and enhancer after the development of metastatic CRPC. The PDX of the primary tumour, without the AR amplification, was sensitive to AR-directed treatments, including castration, enzalutamide, and apalutamide. The PDX of the metastasis, with the AR amplification, had higher AR and AR-V7 expression in castrate conditions, and was resistant to castration, apalutamide, and enzalutamide in vivo. Treatment with a BET inhibitor outperformed the AR-directed therapies for the metastasis, resulting in tumour regression for some, but not all, grafts. Therefore, this study provides novel matched PDXs to test potential treatments that target the overabundance of AR in tumours with AR enhancer amplifications. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Assuntos
Antineoplásicos/farmacologia , Neoplasias de Próstata Resistentes à Castração/patologia , Neoplasias da Próstata/patologia , Receptores Androgênicos/genética , Antagonistas de Androgênios/farmacologia , Androgênios/metabolismo , Animais , Benzamidas/farmacologia , Modelos Animais de Doenças , Progressão da Doença , Xenoenxertos , Humanos , Masculino , Camundongos , Nitrilas/farmacologia , Orquiectomia , Feniltioidantoína/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Tioidantoínas/farmacologia , Sequenciamento Completo do GenomaRESUMO
The growth and progression of solid tumors involves dynamic cross-talk between cancer epithelium and the surrounding microenvironment. To date, molecular profiling has largely been restricted to the epithelial component of tumors; therefore, features underpinning the persistent protumorigenic phenotype of the tumor microenvironment are unknown. Using whole-genome bisulfite sequencing, we show for the first time that cancer-associated fibroblasts (CAFs) from localized prostate cancer display remarkably distinct and enduring genome-wide changes in DNA methylation, significantly at enhancers and promoters, compared to nonmalignant prostate fibroblasts (NPFs). Differentially methylated regions associated with changes in gene expression have cancer-related functions and accurately distinguish CAFs from NPFs. Remarkably, a subset of changes is shared with prostate cancer epithelial cells, revealing the new concept of tumor-specific epigenome modifications in the tumor and its microenvironment. The distinct methylome of CAFs provides a novel epigenetic hallmark of the cancer microenvironment and promises new biomarkers to improve interpretation of diagnostic samples.
Assuntos
Metilação de DNA , Epigenômica/métodos , Neoplasias da Próstata/genética , Microambiente Tumoral/genética , Fibroblastos Associados a Câncer/metabolismo , Células Cultivadas , Fibroblastos/metabolismo , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Genoma Humano/genética , Humanos , Masculino , Regiões Promotoras Genéticas/genética , Neoplasias da Próstata/patologia , Sequenciamento Completo do Genoma/métodosRESUMO
In prostate cancer, cancer-associated fibroblasts (CAF) exhibit contrasting biological properties to non-malignant prostate fibroblasts (NPF) and promote tumorigenesis. Resolving intercellular signaling pathways between CAF and prostate tumor epithelium may offer novel opportunities for research translation. To this end, the proteome and phosphoproteome of four pairs of patient-matched CAF and NPF were characterized to identify discriminating proteomic signatures. Samples were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) with a hyper reaction monitoring data-independent acquisition (HRM-DIA) workflow. Proteins that exhibited a significant increase in CAF versus NPF were enriched for the functional categories "cell adhesion" and the "extracellular matrix." The CAF phosphoproteome exhibited enhanced phosphorylation of proteins associated with the "spliceosome" and "actin binding." STRING analysis of the CAF proteome revealed a prominent interaction hub associated with collagen synthesis, modification, and signaling. It contained multiple collagens, including the fibrillar types COL1A1/2 and COL5A1; the receptor tyrosine kinase discoidin domain-containing receptor 2 (DDR2), a receptor for fibrillar collagens; and lysyl oxidase-like 2 (LOXL2), an enzyme that promotes collagen crosslinking. Increased activity and/or expression of LOXL2 and DDR2 in CAF were confirmed by enzymatic assays and Western blotting analyses. Pharmacological inhibition of CAF-derived LOXL2 perturbed extracellular matrix (ECM) organization and decreased CAF migration in a wound healing assay. Further, it significantly impaired the motility of co-cultured RWPE-2 prostate tumor epithelial cells. These results indicate that CAF-derived LOXL2 is an important mediator of intercellular communication within the prostate tumor microenvironment and is a potential therapeutic target.
Assuntos
Aminoácido Oxirredutases/metabolismo , Fibroblastos Associados a Câncer/metabolismo , Neoplasias da Próstata/metabolismo , Proteômica , Microambiente Tumoral , Comunicação Autócrina , Linhagem Celular Tumoral , Movimento Celular , Células Epiteliais/patologia , Matriz Extracelular/metabolismo , Humanos , Masculino , Proteínas de Neoplasias/metabolismo , Comunicação Parácrina , Fosfoproteínas/metabolismo , Fosforilação , Próstata/metabolismo , Próstata/patologia , Proteoma/metabolismo , Reprodutibilidade dos Testes , Transdução de SinaisRESUMO
BACKGROUND: Breast cancer (BCa) mortality is decreasing with early detection and improvement in therapies. The incidence of BCa, however, continues to increase, particularly estrogen-receptor-positive (ER +) subtypes. One of the greatest modifiers of ER + BCa risk is childbearing (parity), with BCa risk halved in young multiparous mothers. Despite convincing epidemiological data, the biology that underpins this protection remains unclear. Parity-induced protection has been postulated to be due to a decrease in mammary stem cells (MaSCs); however, reports to date have provided conflicting data. METHODS: We have completed rigorous functional testing of repopulating activity in parous mice using unfractionated and MaSC (CD24midCD49fhi)-enriched populations. We also developed a novel serial transplant method to enable us to assess self-renewal of MaSC following pregnancy. Lastly, as each pregnancy confers additional BCa protection, we subjected mice to multiple rounds of pregnancy to assess whether additional pregnancies impact MaSC activity. RESULTS: Here, we report that while repopulating activity in the mammary gland is reduced by parity in the unfractionated gland, it is not due to a loss in the classically defined MaSC (CD24+CD49fhi) numbers or function. Self-renewal was unaffected by parity and additional rounds of pregnancy also did not lead to a decrease in MaSC activity. CONCLUSIONS: Our data show instead that parity impacts on the stem-like activity of cells outside the MaSC population.
Assuntos
Glândulas Mamárias Animais , Células-Tronco , Animais , Feminino , Integrina beta1 , Camundongos , Paridade , GravidezRESUMO
Hypospadias, a developmental defect of the penis, is one of the most common congenital malformations in humans. Its incidence has rapidly increased over recent decades, and this has been largely attributed to our increased exposure to endocrine-disrupting chemicals. Penis development is primarily an androgen-driven process; however, estrogen and xenoestrogens are known to affect penis development in both humans and mice. Here, we investigated the role of estrogen in the developing penis. Using a novel penis culture system, we showed that exogenous estrogen directly targets the developing penis in utero to cause hypospadias. In addition, we also uncovered an unexpected endogenous role for estrogen in normal postnatal penis development and showed that a loss of estrogen signaling results in a mild hypospadias phenotype, the most common manifestation of this disease in humans. Our findings demonstrated that both androgen and estrogen signaling are intrinsically required for normal urethral closure. These findings confirmed that penis development is not an entirely androgen-driven process but one in which endogenous estrogen signaling also plays a critical role.-Govers, L. C., Phillips, T. R., Mattiske, D. M., Rashoo, N., Black, J. R., Sinclair, A., Baskin, L. S., Risbridger, G. P., Pask, A. J. A critical role for estrogen signaling in penis development.
Assuntos
Receptor alfa de Estrogênio/fisiologia , Estrogênios/farmacologia , Hipospadia/etiologia , Pênis/efeitos dos fármacos , Pênis/crescimento & desenvolvimento , Animais , Disruptores Endócrinos/farmacologia , Feminino , Humanos , Hipospadia/metabolismo , Hipospadia/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
Hypospadias is the abnormal opening of the urethra on the underside of the penis and occurs in approximately 1/125 live male births worldwide. The incidence rate of hypospadias has dramatically increased over the past few decades. This is now attributed, at least in part, to our exposure to endocrine-disrupting chemicals (EDCs) which alter the hormonal signals required for development of the penis. In humans androgens are the main drivers of fusion of the urethral folds to form the urethra within the shaft of the penis, a process required for termination of the urethra in its normal location at the tip of the penis. However, recent research has suggested that estrogen also plays a role in this process. To better understand how EDCs impact urethral development it is essential that we understand the normal function of hormones during development of the penis. To define the role of estrogen in urethral development we examined development of the penis in the aromatase (Cyp19a1) Knockout (ArKO) mouse strain in which endogenous estrogen production is completely ablated. We found that the ArKO penis had a mild hypospadias phenotype. The developing ArKO postnatal penis displayed an early disruption in preputial development, which likely causes the mild hypospadias observed in adults. Using qPCR, we found altered expression of keratin genes and key urethral patterning genes in response to the disrupted estrogen signaling. The hypospadias phenotype was almost identical to that reported for the estrogen receptor α (ERα) knockout confirming that ERα is the predominant receptor for mediating estrogen action during development of the mouse penis. Our results show that estrogen is required for normal prepucial development and placement of the mature urethral opening at the distal aspect of the penis. We also identified several genes which are potential downstream targets of estrogen during normal urethral closure. With this knowledge, we can now better understand how anti-estrogenic as well as estrogenic EDCs disrupt urethral closure to cause mild hypospadias in both mice and humans.
Assuntos
Aromatase/fisiologia , Estrogênios/metabolismo , Hipospadia/etiologia , Organogênese , Pênis/anormalidades , Receptores de Estrogênio/metabolismo , Animais , Hipospadia/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pênis/enzimologia , Transdução de SinaisRESUMO
BACKGROUND: Serially transplantable patient-derived xenografts (PDXs) are invaluable preclinical models for studying tumor biology and evaluating therapeutic agents. As these models are challenging to establish from prostate cancer specimens, the ability to preserve them through cryopreservation has several advantages for ongoing research. Despite this, there is still uncertainty about the ability to cryopreserve PDXs of prostate cancer. This study compared three different cryopreservation protocols to identify a method that can be used to reproducibly cryopreserve a diverse cohort of prostate cancer PDX models. METHODS: One serially transplantable prostate cancer PDX from the Melbourne Urological Research Alliance cohort was used to compare three cryopreservation protocols: slow freezing in fetal calf serum (FCS) with 10% dimethyl sulfoxide (DMSO), FCS with 10% DMSO supplemented with the Rho-associated kinase (ROCK) inhibitor Y-27632 and vitrification. The efficiency of the slow freezing protocols was then assessed in 17 additional prostate cancer PDXs. Following cryopreservation, PDXs were re-established in host mice that were either intact and supplemented with testosterone or castrated. Graft take rate, tumor growth, histological features, and transcriptome profiles before and after cryopreservation were compared. RESULTS: Slow freezing maintained the viability and histological features of prostate cancer PDXs, and the addition of a ROCK inhibitor increased their growth following cryopreservation. Using the slow freezing method, we re-established 100% of PDXs grown in either testosterone-supplemented or castrated host mice. Importantly, the long-term tumor growth rate and transcriptome profile were maintained following cryopreservation. CONCLUSION: This study has identified a protocol to reliably cryopreserve and re-establish a diverse cohort of serially transplantable PDXs of prostate cancer. This study has the potential to significantly improve the practicality of maintaining PDX models. Cryopreservation may also increase the accessibility of these important resources and provide new opportunities for preclinical studies on a broader spectrum of prostate tumors.
Assuntos
Criopreservação/métodos , Xenoenxertos , Transplante de Neoplasias/métodos , Neoplasias da Próstata/patologia , Animais , Modelos Animais de Doenças , Humanos , Masculino , Camundongos , Transplante de Neoplasias/patologiaRESUMO
Prostate carcinoma and benign prostate hyperplasia (BPH) with associated lower urinary tract symptoms (LUTS) are among the most prevalent and clinically relevant diseases in men. BPH is characterized by an enlargement of prostate tissue associated with increased tone of smooth muscle cells (SMCs) which surround the single glands composing the prostate. Secretions of the glands leave the prostate through local excretory ducts during the emission phase of ejaculation. Pharmacological treatment of BPH suggests different local drug targets based on reduction of prostate smooth muscle tone as the main effect and disturbed ejaculation as a common side effect. This highlights the need for detailed investigation of single prostate glands and ducts. We combined structural and functional imaging techniques-notably, clear lipid-exchanged, acrylamide-hybridized rigid imaging/immunostaining/ in situ hybridization-compatible tissue-hydrogel (CLARITY) and time-lapse imaging-and defined glands and ducts as distinct SMC compartments in human and rat prostate tissue. The single glands of the prostate (comprising the secretory part) are characterized by spontaneous contractions mediated by the surrounding SMCs, whereas the ducts (excretory part) are quiescent. In both SMC compartments, phosphodiesterase (PDE)-5 is expressed. PDE5 inhibitors have recently emerged as alternative treatment options for BPH. We directly visualized that the PDE5 inhibitors sildenafil and tadalafil act by reducing spontaneous contractility of the glands, thereby reducing the muscle tone of the organ. In contrast, the ductal (excretory) system and thus the prostate's contribution to ejaculation is unaffected by PDE5 inhibitors. Our differentiated imaging approach reveals new details about prostate function and local drug actions and thus may support clinical management of BPH.-Kügler, R., Mietens, A., Seidensticker, M., Tasch, S., Wagenlehner, F. M., Kaschtanow, A., Tjahjono, Y., Tomczyk, C. U., Beyer, D., Risbridger, G. P., Exintaris, B., Ellem, S. J., Middendorff, R. Novel imaging of the prostate reveals spontaneous gland contraction and excretory duct quiescence together with different drug effects.
Assuntos
Ejaculação/efeitos dos fármacos , Contração Muscular/efeitos dos fármacos , Músculo Liso/patologia , Inibidores da Fosfodiesterase 5/farmacologia , Próstata/patologia , Hiperplasia Prostática/patologia , Neoplasias da Próstata/patologia , Idoso , Animais , Humanos , Masculino , Pessoa de Meia-Idade , Músculo Liso/diagnóstico por imagem , Músculo Liso/efeitos dos fármacos , Próstata/diagnóstico por imagem , Próstata/efeitos dos fármacos , Hiperplasia Prostática/diagnóstico por imagem , Hiperplasia Prostática/tratamento farmacológico , Neoplasias da Próstata/diagnóstico por imagem , Neoplasias da Próstata/tratamento farmacológico , Ratos , Ratos Wistar , Imagem com Lapso de TempoRESUMO
BACKGROUND: While it has been challenging to establish prostate cancer patient-derived xenografts (PDXs), with a take rate of 10-40% and long latency time, multiple groups throughout the world have developed methods for the successful establishment of serially transplantable human prostate cancer PDXs using a variety of immune deficient mice. In 2014, the Movember Foundation launched a Global Action Plan 1 (GAP1) project to support an international collaborative prostate cancer PDX program involving eleven groups. Between these Movember consortium members, a total of 98 authenticated human prostate cancer PDXs were available for characterization. Eighty three of these were derived directly from patient material, and 15 were derived as variants of patient-derived material via serial passage in androgen deprived hosts. A major goal of the Movember GAP1 PDX project was to provide the prostate cancer research community with a summary of both the basic characteristics of the 98 available authenticated serially transplantable human prostate cancer PDX models and the appropriate contact information for collaborations. Herein, we report a summary of these PDX models. METHODS: PDX models were established in immunocompromised mice via subcutaneous or subrenal-capsule implantation. Dual-label species (ie, human vs mouse) specific centromere and telomere Fluorescence In Situ Hybridization (FISH) and immuno-histochemical (IHC) staining of tissue microarrays (TMAs) containing replicates of the PDX models were used for characterization of expression of a number of phenotypic markers important for prostate cancer including AR (assessed by IHC and FISH), Ki67, vimentin, RB1, P-Akt, chromogranin A (CgA), p53, ERG, PTEN, PSMA, and epithelial cytokeratins. RESULTS: Within this series of PDX models, the full spectrum of clinical disease stages is represented, including androgen-sensitive and castration-resistant primary and metastatic prostate adenocarcinomas as well as prostate carcinomas with neuroendocrine differentiation. The annotated clinical characteristics of these PDXs were correlated with their marker expression profile. CONCLUSION: Our results demonstrate the clinical relevance of this series of PDXs as a platform for both basic science studies and therapeutic discovery/drug development. The present report provides the prostate cancer community with a summary of the basic characteristics and a contact information for collaborations using these models.
Assuntos
Xenoenxertos , Transplante de Neoplasias/métodos , Neoplasias da Próstata/patologia , Ensaios Antitumorais Modelo de Xenoenxerto , Animais , Biomarcadores Tumorais/metabolismo , Modelos Animais de Doenças , Humanos , Masculino , Camundongos , Neoplasias da Próstata/metabolismoRESUMO
OBJECTIVE: To determine the relevance of intraductal carcinoma of the prostate (IDC-P) in advanced prostate cancer by first examining whether IDC-P was originally present in patients who later developed advanced prostate cancer and then using patient-derived xenografts (PDXs) to investigate the response of IDC-P to androgen deprivation therapy (ADT). MATERIALS AND METHODS: We conducted a retrospective pathology review of IDC-P in primary prostate biopsy or surgery specimens from 38 men who subsequently developed advanced prostate cancer. Overall survival was calculated using the Kaplan-Meier method. To demonstrate the response of IDC-P to ADT, we established PDXs from seven patients with familial and/or high-risk sporadic prostate cancer. After castration and testosterone restoration of host mice, we measured the volume and proliferation of IDC-P within PDX grafts. RESULTS: We found that IDC-P was a prominent feature in the primary prostate specimens, present in 63% of specimens and often co-existing with poorly differentiated adenocarcinoma. Overall survival was similar in patients with or without IDC-P. In the PDXs from all seven patients, IDC-P was identified and present at a similar volume to adenocarcinoma. Residual IDC-P lesions persisted after host castration and, similar to castrate-tolerant adenocarcinoma, testosterone restoration led to tumour regeneration. CONCLUSION: The study showed that IDC-P is prevalent in aggressive prostate cancer and contains cells that can withstand androgen deprivation. Thus, IDC-P appears functionally relevant in advanced prostate cancer. The presence of IDC-P may be a trigger to develop innovative clinical management plans.
Assuntos
Antagonistas de Androgênios/uso terapêutico , Carcinoma Ductal/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Adenocarcinoma/patologia , Idoso , Idoso de 80 Anos ou mais , Animais , Carcinoma Ductal/patologia , Xenoenxertos/patologia , Humanos , Estimativa de Kaplan-Meier , Masculino , Camundongos Endogâmicos NOD , Camundongos SCID , Pessoa de Meia-Idade , Neoplasias de Próstata Resistentes à Castração/patologia , Estudos Retrospectivos , Transplante Heterólogo , Ensaios Antitumorais Modelo de Xenoenxerto/métodosRESUMO
Castration-resistant prostate cancer is a lethal disease. The cell type(s) that survive androgen deprivation remain poorly described, despite global efforts to understand the various mechanisms of therapy resistance. We recently identified in wild-type (WT) mouse prostates a rare population of luminal progenitor cells that we called LSCmed according to their FACS profile (Lin- /Sca-1+ /CD49fmed ). Here, we investigated the prevalence and castration resistance of LSCmed in various mouse models of prostate tumourigenesis (Pb-PRL, Ptenpc-/- , and Hi-Myc mice). LSCmed prevalence is low (â¼8%, similar to WT) in Hi-Myc mice, where prostatic androgen receptor signalling is unaltered, but is significantly higher in the two other models, where androgen receptor signalling is decreased, rising up to more than 80% in Ptenpc-/- prostates. LSCmed tolerate androgen deprivation and persist or are enriched 2-3 weeks after castration. The tumour-initiating properties of LSCmed from Ptenpc-/- mice were demonstrated by regeneration of tumours in vivo. Transcriptomic analysis revealed that LSCmed represent a unique cell entity as their gene expression profile is different from luminal and basal/stem cells, but shares markers of each. Their intrinsic androgen signalling is markedly decreased, explaining why LSCmed tolerate androgen deprivation. This also illuminates why Ptenpc-/- tumours are castration-resistant since LSCmed represent the most prevalent cell type in this model. We validated CK4 as a specific marker for LSCmed on sorted cells and prostate tissues by immunostaining, allowing for the detection of LSCmed in various mouse prostate specimens. In castrated Ptenpc-/- prostates, there was significant proliferation of CK4+ cells, further demonstrating their key role in castration-resistant prostate cancer progression. Taken together, this study identifies LSCmed as a probable source of prostate cancer relapse after androgen deprivation and as a new therapeutic target for the prevention of castrate-resistant prostate cancer. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.