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1.
bioRxiv ; 2024 Jun 24.
Artigo em Inglês | MEDLINE | ID: mdl-38260576

RESUMO

Androgen receptor (AR) splice variants, of which ARv7 is the most common, are increased in prostate cancer (PC) that develops resistance to androgen signaling inhibitor drugs, but the extent to which these variants drive AR activity, and whether they have novel functions or dependencies, remain to be determined. We generated a subline of VCaP PC cells (VCaP16) that is resistant to the AR inhibitor enzalutamide (ENZ) and found that AR activity was independent of the full-length AR (ARfl), despite its continued high-level expression, and was instead driven by ARv7. The ARv7 cistrome and transcriptome in VCaP16 cells mirrored that of the ARfl in VCaP cells, although ARv7 chromatin binding was weaker, and strong ARv7 binding sites correlated with higher affinity ARfl binding sites across multiple models and clinical samples. Notably, although ARv7 expression in VCaP cells increased rapidly in response to ENZ, there was a long lag before it gained chromatin binding and transcriptional activity. This lag was associated with an increase in chromatin accessibility, with the AR and nuclear factor I (NFI) motifs being most enriched at these more accessible sites. Moreover, the transcriptional effects of combined NFIB and NFIX knockdown versus ARv7 knockdown were highly correlated. These findings indicate that ARv7 can drive the AR program, but that its activity is dependent on adaptations that increase chromatin accessibility to enhance its intrinsically weak chromatin binding.

2.
Cancer Res ; 83(7): 1016-1030, 2023 04 04.
Artigo em Inglês | MEDLINE | ID: mdl-36622276

RESUMO

Noncanonical Wnt signaling by WNT5a has oncogenic and tumor suppressive activities, but downstream pathways mediating these specific effects remain to be fully established. In a subset of prostate cancer organoid culture and xenograft models, inhibition of Wnt synthesis stimulated growth, whereas WNT5a or a WNT5a mimetic peptide (Foxy5) markedly suppressed tumor growth. WNT5a caused a ROR2-dependent decrease in YAP1 activity, which was associated with increased phosphorylation of MST1/2, LATS1, MOB1, and YAP1, indicating Hippo pathway activation. Deletion of MST1/2 abrogated the WNT5a response. WNT5a similarly activated Hippo in ROR2-expressing melanoma cells, whereas WNT5a in ROR2-negative cells suppressed Hippo. This suppression was associated with increased inhibitory phosphorylation of NF2/Merlin that was not observed in ROR2-expressing cells. WNT5a also increased mRNA encoding Hippo pathway components including MST1 and MST2 and was positively correlated with these components in prostate cancer clinical datasets. Conversely, ROR2 and WNT5a expression was stimulated by YAP1, and correlated with increased YAP1 activity in clinical datasets, revealing a WNT5a/ROR2 negative feedback loop to modulate YAP1 activity. Together these findings identify Hippo pathway activation as a mechanism that mediates the tumor suppressive effects of WNT5a and indicate that expression of ROR2 may be a predictive biomarker for responsiveness to WNT5a-mimetic drugs. SIGNIFICANCE: WNT5a signaling through ROR2 activates the Hippo pathway to downregulate YAP1/TAZ activity and suppress tumor growth, identifying ROR2 as a potential biomarker to identify patients that could benefit from WNT5a-related agents.


Assuntos
Via de Sinalização Hippo , Neoplasias da Próstata , Masculino , Humanos , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/genética , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Transdução de Sinais/fisiologia , Proteína Wnt-5a/metabolismo , Fosforilação
3.
Artigo em Inglês | MEDLINE | ID: mdl-34568716

RESUMO

Metastatic prostate cancer is initially sensitive to androgen receptor inhibition, but eventually becomes castration-resistant prostate cancer (mCRPC). Early use of more intensive therapies targeting androgen receptor and other oncogenic drivers in treatment-naïve primary prostate cancer (PC) may be more effective than that in advanced mCRPC. However, analysis of primary tumors may not reveal targetable metastatic drivers that are subclonal in the primary tumor or acquired at metastatic sites. METHODS: PC samples spanning one patient's clinical course: diagnostic biopsies, pre- or post-enzalutamide metastatic biopsies, and rapid autopsy samples including a patient-derived xenograft (PDX) were analyzed by targeted exome sequencing followed by phylogenetic analysis. RESULTS: Left- and right-lobe primary PC tumors appeared to diverge, with the right acquiring additional shared mutations and striking differences in copy number alterations that later appeared in metastatic samples during the treatment course and at autopsy, whereas the left base tumor maintained a quiet copy number alteration landscape and partitioned into a dead-end node. RB1 loss, a common finding in advanced castration-resistant disease, was identified throughout mCRPC samples, but not in the primary tumor. Significantly, a truncal EGFR-activating mutation (R108K) was identified in the primary tumor and was also found to be maintained in the mCRPC samples and in a PDX model. Furthermore, the PDX model remained sensitive to the EGFR inhibitor erlotinib, despite the presence of both RB1 and BRCA2 losses. CONCLUSION: These findings indicate that truncal alterations identified in primary PC can drive advanced mCRPC, even in the presence of additional strong oncogenic drivers (ie, RB1 and BRCA2 loss), and suggest that earlier detection and targeting of these truncal alterations may be effective at halting disease progression.


Assuntos
Neoplasias de Próstata Resistentes à Castração , Biópsia , Carcinogênese , Humanos , Masculino , Filogenia , Neoplasias de Próstata Resistentes à Castração/genética
4.
Cancer Lett ; 519: 172-184, 2021 10 28.
Artigo em Inglês | MEDLINE | ID: mdl-34256096

RESUMO

One mechanism for reactivation of androgen receptor (AR) activity after androgen deprivation therapy in castration-resistant prostate cancer (CRPC) is expression of splice variants such as ARv7 that delete the ligand binding domain and have constitutive activity. Exogenous overexpressed ARv7 can function as a homodimer or heterodimer with full length AR (ARfl), which is highly expressed with ARv7 in CRPC. However, the extent to which endogenous ARv7 function is dependent on heterodimerization with ARfl remains to be determined. We used double-crosslinking to stabilize AR complexes on chromatin in a CRPC cell line expressing endogenous ARfl and ARv7 (LN95 cells), and established that only trace levels of ARfl were associated with ARv7 on chromatin. Consistent with this result, depletion of ARfl with an AR degrader targeting the AR ligand binding domain did not decrease ARv7 binding to chromatin or its association with HOXB13, but did decrease overall AR transcriptional activity. Comparable results were obtained in CWR22RV1 cells, another CRPC cell line expressing ARfl and ARv7. These results indicate that ARv7 function in CRPC is not dependent on ARfl, and that both contribute independently to overall AR activity.


Assuntos
Neoplasias da Próstata/genética , Receptores Androgênicos/genética , Linhagem Celular , Linhagem Celular Tumoral , Cromatina/genética , Células HEK293 , Proteínas de Homeodomínio/genética , Humanos , Ligantes , Masculino , Próstata/metabolismo , Domínios Proteicos/genética
5.
Clin Cancer Res ; 27(17): 4836-4847, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34168052

RESUMO

PURPOSE: A subset of primary prostate cancer expresses programmed death-ligand 1 (PD-L1), but whether they have a unique tumor immune microenvironment or genomic features is unclear. EXPERIMENTAL DESIGN: We selected PD-L1-positive high-grade and/or high-risk primary prostate cancer, characterized tumor-infiltrating lymphocytes with multiplex immunofluorescence, and identified genomic alterations in immunogenic and nonimmunogenic tumor foci. RESULTS: One quarter of aggressive localized prostate cancer cases (29/115) had tumor PD-L1 expression more than 5%. This correlated with increased density of CD8+ T cells, a large fraction coexpressing PD-1, versus absent PD-1 expression on sparse CD8 T cells in unselected cases. Most CD8+PD-1+ cells did not express terminal exhaustion markers (TIM3 or LAG3), while a subset expressed TCF1. Consistent with these CD8+PD-1+TCF1+ cells being progenitors, they were found in antigen-presenting cell niches in close proximity to MHC-II+ cells. CD8 T-cell density in immunogenic prostate cancer and renal cell carcinoma (RCC) was nearly identical. Shallow RB1 and BRCA2 losses, and deep deletions of CHD1, were prevalent, the latter being strongly associated with a dendritic cell gene set in The Cancer Genome Atlas. Tumor mutation burden was variable; neither high microsatellite instability nor CDK12 alterations were present. CONCLUSIONS: A subset of localized prostate cancer is immunogenic, manifested by PD-L1 expression and CD8+ T-cell content comparable with RCC. The CD8+ T cells include effector cells and exhausted progenitor cells, which may be expanded by immune checkpoint inhibitors (ICI). Genomic losses of RB1, BRCA2, and CHD1 may be drivers of this phenotype. These findings indicate that immunotherapies may be effective in biomarker-selected subpopulations of patients with localized prostate cancer.


Assuntos
Linfócitos T CD8-Positivos , Neoplasias da Próstata , Antígeno B7-H1/genética , Genes Supressores de Tumor , Humanos , Linfócitos do Interstício Tumoral , Masculino , Fenótipo , Neoplasias da Próstata/genética , Microambiente Tumoral/genética
6.
Mol Oncol ; 15(7): 1901-1920, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33932081

RESUMO

Androgen receptor (AR) is the principal molecule in prostate cancer (PCa) etiology and therapy. AR re-activation still remains a major challenge during treatment of castration-resistant prostate cancer (CRPC) tumors that relapse after castration therapies. Recent reports have indicated the enrichment of Ser81-phosphorylated AR (pS81) in the nucleus of CRPC cells, and CDK1 and CDK9 as the kinases phosphorylating AR at S81. In the current study we showed that pS81 is preferentially localized in the nucleus in both rapid biopsy metastatic CRPC samples and PCa xenografts, and nuclear pS81 localization is correlated with AR transactivation in tumor xenografts. Chromatin immunoprecipitation (ChIP) analysis demonstrated an alignment of S81 phosphorylation and AR-mediated transactivation with the chromatin locus openness. Moreover, pS81-specific ChIP-Seq showed a disproportional occupancy of pS81 on AR-activated promoters, while 3C-ChIP assays further indicated an enrichment of pS81 at the PSA enhancer-promoter loop, a known AR activating hub. In the latter, CDK9 was shown to modulate the transactivation of the AR and RNA Pol II. Indeed, ChIP and re-ChIP assays also confirmed that AR-dependent activation of the PSA enhancer and promoter mediated by pS81 was coupled with activation of Pol II and the pTEFb complex. Mechanistically, we determined that CDK1 and CDK9 sustained the pS81 AR modification in the soluble and chromatin-bound fractions of PCa cells, respectively. Finally, we demonstrated that CDK1 activity was maintained throughout the cell cycle, and that CDK1 inhibitors restored androgen sensitivity in CRPC tumor cells. Based on these findings, CDK1 and CDK9 could be targeted as pS81 kinases in patients with CRPC, either alone or in conjunction with direct AR antagonists.


Assuntos
Neoplasias de Próstata Resistentes à Castração , Neoplasias da Próstata , Proteína Quinase CDC2/genética , Proteína Quinase CDC2/metabolismo , Linhagem Celular Tumoral , Quinase 9 Dependente de Ciclina/genética , Quinase 9 Dependente de Ciclina/metabolismo , Humanos , Masculino , Fosforilação , Neoplasias da Próstata/patologia , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/metabolismo , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Ativação Transcricional/genética
7.
J Clin Invest ; 130(6): 3287-3298, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-32478682

RESUMO

Despite widespread use of taxanes, mechanisms of action and resistance in vivo remain to be established, and there is no way of predicting who will respond to therapy. This study examined prostate cancer (PCa) xenografts and patient samples to identify in vivo mechanisms of taxane action and resistance. Docetaxel drug-target engagement was assessed by confocal anti-tubulin immunofluorescence to quantify microtubule bundling in interphase cells and aberrant mitoses. Tumor biopsies from metastatic PCa patients obtained 2 to 5 days after their first dose of docetaxel or cabazitaxel were processed to assess microtubule bundling, which correlated with clinical response. Microtubule bundling was evident in PCa xenografts 2 to 3 days after docetaxel treatment but was decreased or lost with acquired resistance. Biopsies after treatment with leuprolide plus docetaxel showed extensive microtubule bundling as did biopsies obtained 2 to 3 days after initiation of docetaxel or cabazitaxel in 2 patients with castration-resistant PCa with clinical responses. In contrast, microtubule bundling in biopsies 2 to 3 days after the first dose of docetaxel was markedly lower in 4 nonresponding patients. These findings indicate that taxanes target both mitotic and interphase cells in vivo and that resistance is through mechanisms that impair drug-target engagement. Moreover, the findings suggest that microtubule bundling after initial taxane treatment may be a predictive biomarker for clinical response.


Assuntos
Hidrocarbonetos Aromáticos com Pontes , Resistencia a Medicamentos Antineoplásicos , Microtúbulos/metabolismo , Neoplasias da Próstata/metabolismo , Taxoides , Animais , Hidrocarbonetos Aromáticos com Pontes/farmacocinética , Hidrocarbonetos Aromáticos com Pontes/farmacologia , Linhagem Celular Tumoral , Docetaxel/farmacocinética , Docetaxel/farmacologia , Humanos , Masculino , Camundongos , Camundongos Nus , Microtúbulos/patologia , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Taxoides/farmacocinética , Taxoides/farmacologia
8.
Artigo em Inglês | MEDLINE | ID: mdl-31528835

RESUMO

PURPOSE: Despite decreased screening-based detection of clinically insignificant tumors, most diagnosed prostate cancers are still indolent, indicating a need for better strategies for detection of clinically significant disease before treatment. We hypothesized that patients with detectable circulating tumor DNA (ctDNA) were more likely to harbor aggressive disease. METHODS: We applied ultra-low-pass whole-genome sequencing to profile cell-free DNA from 112 patients diagnosed with localized prostate cancer and performed targeted resequencing of plasma DNA for somatic mutations previously identified in matched solid tumor in nine cases. We also performed similar analyses of data from patients with metastatic prostate cancer. RESULTS: In all cases of localized prostate cancer, even in clinically high-risk patients who subsequently had recurrent disease, ultra-low-pass whole-genome sequencing and targeted resequencing did not detect ctDNA in plasma acquired before surgery or before recurrence. In contrast, using both approaches, ctDNA was detected in patients with metastatic prostate cancer. CONCLUSION: Our findings demonstrate clear differences between localized and advanced prostate cancer with respect to the dissemination and detectability of ctDNA. Because allele-specific alterations in ctDNA are below the threshold for detection in localized prostate cancer, other approaches to identify cell-free nucleic acids of tumor origin may demonstrate better specificity for aggressive disease.

9.
Am J Hematol ; 94(1): 62-73, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30295334

RESUMO

Myeloproliferative neoplasms (MPNs) driver mutations are usually found in JAK2, MPL, and CALR genes; however, 10%-15% of cases are triple negative (TN). A previous study showed lower rate of JAK2 V617F in primary myelofibrosis patients exposed to low doses of ionizing radiation (IR) from Chernobyl accident. To examine distinct driver mutations, we enrolled 281 Ukrainian IR-exposed and unexposed MPN patients. Genomic DNA was obtained from peripheral blood leukocytes. JAK2 V617F, MPL W515, types 1- and 2-like CALR mutations were identified by Sanger Sequencing and real time polymerase chain reaction. Chromosomal alterations were assessed by oligo-SNP microarray platform. Additional genetic variants were identified by whole exome and targeted sequencing. Statistical significance was evaluated by Fisher's exact test and Wilcoxon's rank sum test (R, version 3.4.2). IR-exposed MPN patients exhibited a different genetic profile vs unexposed: lower rate of JAK2 V617F (58.4% vs 75.4%, P = .0077), higher rate of type 1-like CALR mutation (12.2% vs 3.1%, P = .0056), higher rate of TN cases (27.8% vs 16.2%, P = .0366), higher rate of potentially pathogenic sequence variants (mean numbers: 4.8 vs 3.1, P = .0242). Furthermore, we identified several potential drivers specific to IR-exposed TN MPN patients: ATM p.S1691R with copy-neutral loss of heterozygosity at 11q; EZH2 p.D659G at 7q and SUZ12 p.V71 M at 17q with copy number loss. Thus, IR-exposed MPN patients represent a group with distinct genomic characteristics worthy of further study.


Assuntos
Acidente Nuclear de Chernobyl , Transtornos Mieloproliferativos/etiologia , Neoplasias Induzidas por Radiação/etiologia , Poluentes Radioativos/efeitos adversos , Adulto , Idoso , Calreticulina/genética , Aberrações Cromossômicas , DNA/genética , Feminino , Dosagem de Genes , Humanos , Janus Quinase 2/genética , Perda de Heterozigosidade , Masculino , Pessoa de Meia-Idade , Mutação de Sentido Incorreto , Transtornos Mieloproliferativos/epidemiologia , Transtornos Mieloproliferativos/genética , Neoplasias Induzidas por Radiação/epidemiologia , Neoplasias Induzidas por Radiação/genética , Receptores de Trombopoetina/genética , Ucrânia/epidemiologia , Sequenciamento do Exoma , Adulto Jovem
10.
Cancer Res ; 78(22): 6354-6362, 2018 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-30242112

RESUMO

The standard treatment for metastatic prostate cancer, androgen deprivation therapy (ADT), is designed to suppress androgen receptor (AR) activity. However, men invariably progress to castration-resistant prostate cancer (CRPC), and AR reactivation contributes to progression in most cases. To identify mechanisms that may drive CRPC, we examined a VCaP prostate cancer xenograft model as tumors progressed from initial androgen sensitivity prior to castration to castration resistance and then on to relapse after combined therapy with further AR-targeted drugs (abiraterone plus enzalutamide). AR activity persisted in castration-resistant and abiraterone/enzalutamide-resistant xenografts and was associated with increased expression of the AR gene and the AR-V7 splice variant. We then assessed expression of individual AR-regulated genes to identify those that persisted, thereby contributing to tumor growth, versus those that decreased and may therefore exhibit tumor suppressor activities. The most significantly decreased AR target gene was dipeptidyl peptidase 4 (DPP4), which encodes a membrane-anchored protein that cleaves dipeptides from multiple growth factors, resulting in their increased degradation. DPP4 mRNA and protein were also decreased in clinical CRPC cases, and inhibition of DPP4 with sitagliptin enhanced the growth of prostate cancer xenografts following castration. Significantly, DPP4 inhibitors are frequently used to treat type 2 diabetes as they increase insulin secretion. Together, these results implicate DPP4 as an AR-regulated tumor suppressor gene whose loss enhances growth factor activity and suggest that treatment with DPP4 inhibitors may accelerate emergence of resistance to ADT.Significance: These findings identify DPP4 as an AR-stimulated tumor suppressor gene that is downregulated during progression to castration-resistant prostate cancer, warning that treatment with DPP4 inhibitors, commonly used to treat type 2 diabetes, may accelerate prostate cancer progression following androgen deprivation therapy. Cancer Res; 78(22); 6354-62. ©2018 AACR.


Assuntos
Dipeptidil Peptidase 4/metabolismo , Progressão da Doença , Regulação para Baixo , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/metabolismo , Processamento Alternativo , Antagonistas de Androgênios/farmacologia , Androstenos/farmacologia , Animais , Benzamidas , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Epigênese Genética , Perfilação da Expressão Gênica , Humanos , Masculino , Camundongos , Camundongos SCID , Recidiva Local de Neoplasia , Transplante de Neoplasias , Nitrilas , Feniltioidantoína/análogos & derivados , Feniltioidantoína/farmacologia , RNA Interferente Pequeno/metabolismo , Receptores Androgênicos/metabolismo , Transdução de Sinais , Fosfato de Sitagliptina/farmacologia
11.
Cancer Lett ; 438: 97-104, 2018 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-30217568

RESUMO

Phosphorylation of serine 81 (pS81) in the N-terminal transactivation domain of the androgen receptor (AR) has been linked to its transcriptional activation in prostate cancer (PCa) cell lines, but in vivo studies have been limited. Moreover, the role of pS81 in the reactivation of AR when tumors relapse after androgen deprivation therapy (castration-resistant prostate cancer, CRPC) has not been determined. In this study we validate a pS81 antibody for immunohistochemistry (IHC) and show it yields strong nuclear staining in primary PCa clinical samples and in the VCaP PCa xenograft model. Moreover, this staining was decreased at 7 days post-castration in VCaP xenografts, coinciding with markedly decreased AR transcriptional activity. Staining with the pS81 antibody then was restored when the VCaP xenografts relapsed, which was associated with restoration of AR transcriptional activity. Significantly, analysis of CRPC clinical samples, including tumors that had progressed during treatment with abiraterone, showed strong nuclear staining with the pS81 antibody. Together these findings indicate that AR reactivation in CRPC is associated with S81 phosphorylation, and suggest that IHC for pS81 may be useful as a biomarker of AR activity in CRPC.


Assuntos
Neoplasias de Próstata Resistentes à Castração/metabolismo , Receptores Androgênicos/metabolismo , Serina/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto , Androstenos/farmacologia , Animais , Benzamidas , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Humanos , Imuno-Histoquímica , Masculino , Camundongos Endogâmicos ICR , Camundongos SCID , Nitrilas , Feniltioidantoína/análogos & derivados , Feniltioidantoína/farmacologia , Fosforilação , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/genética , Receptores Androgênicos/genética , Serina/genética
12.
Cancer Res ; 78(16): 4716-4730, 2018 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-29921690

RESUMO

Primary prostate cancer can have extensive microheterogeneity, but its contribution to the later emergence of metastatic castration-resistant prostate cancer (mCRPC) remains unclear. In this study, we microdissected residual prostate cancer foci in radical prostatectomies from 18 men treated with neoadjuvant-intensive androgen deprivation therapy (leuprolide, abiraterone acetate, and prednisone) and analyzed them for resistance mechanisms. Transcriptome profiling showed reduced but persistent androgen receptor (AR) activity in residual tumors, with no increase in neuroendocrine differentiation. Proliferation correlated negatively with AR activity but positively with decreased RB1 expression, and whole-exome sequencing (WES) further showed enrichment for RB1 genomic loss. In 15 cases where 2 or 3 tumor foci were microdissected, WES confirmed a common clonal origin but identified multiple oncogenic alterations unique to each focus. These findings show that subclones with oncogenic alterations found in mCRPC are present in primary prostate cancer and are selected for by neoadjuvant-intense androgen deprivation therapy. In particular, this study indicates that subclonal RB1 loss may be more common than previously appreciated in intermediate- to high-risk primary prostate cancer and may be an early event, independent of neuroendocrine differentiation, in the development of mCRPC. Comprehensive molecular analyses of primary prostate cancer may detect aggressive subclones and possibly inform adjuvant strategies to prevent recurrence.Significance: Neoadjuvant androgen deprivation therapy for prostate cancer selects for tumor foci with subclonal genomic alterations, which may comprise the origin of metastatic castration-resistant prostate cancer. Cancer Res; 78(16); 4716-30. ©2018 AACR.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Resistencia a Medicamentos Antineoplásicos/genética , Recidiva Local de Neoplasia/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Acetato de Abiraterona/administração & dosagem , Idoso , Antagonistas de Androgênios/administração & dosagem , Hidrocarboneto de Aril Hidroxilases/genética , Biomarcadores Tumorais , Carcinogênese , Evolução Clonal , Família 2 do Citocromo P450/genética , Humanos , Masculino , Pessoa de Meia-Idade , Terapia Neoadjuvante , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/patologia , Prostatectomia , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Receptores Androgênicos/genética , Esteroide 16-alfa-Hidroxilase/genética
13.
Urol Oncol ; 36(3): 93.e13-93.e21, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29079132

RESUMO

PURPOSE: SATB1, a global genome organizer, has been shown to play a role in the development and progression of some solid tumors, but its role in bladder cancer is undetermined. Moreover, there is conflicting data about the role of SATB1 in other tumors. This study was initiated to assess a potential role for SATB1 with the hypothesis that SATB1 acts as a tumor promoter in bladder cancer. MATERIALS AND METHODS: We evaluated SATB1 expression in bladder cancer cell lines (HTB-5, HTB-9) and compared them to a benign urothelial cell line (UROtsa). Short-hairpin RNA was used to silence SATB1 in multiple cell lines, and cell death and cell proliferation were assessed using multiple assays. RESULTS: SATB1 expression was increased significantly in all cancer cell lines compared to benign urothelial cells. SATB1 expression was knocked down by short-hairpin RNA and functional outcomes, including cell number, cell-cycle arrest, cell viability, and apoptosis after cisplatin treatment, were measured. Surprisingly, knockdown of SATB1 in 2 high-grade cancer cell lines showed opposing functional roles. Compared to the non-silencing control, HTB-5 cells, showed decreased cellular proliferation and increased sensitivity to cisplatin, whereas HTB-9 cells, showed increased cell numbers and increased resistance to cisplatin. CONCLUSION: We conclude that our results in bladder cancer are consistent with the conflicting data reported in other cancers, and that SATB1 might have different roles in cancer dependent on genetic background and stage of the cancer.


Assuntos
Proteínas de Ligação à Região de Interação com a Matriz/metabolismo , Neoplasias da Bexiga Urinária/patologia , Apoptose/efeitos dos fármacos , Apoptose/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Cisplatino/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Perfilação da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Proteínas de Ligação à Região de Interação com a Matriz/genética , RNA Interferente Pequeno/metabolismo , Regulação para Cima , Neoplasias da Bexiga Urinária/genética
14.
Breast J ; 23(4): 461-464, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28139868

RESUMO

Next-generation sequencing promotes identification of mutations in non-BRCA1/2 genes in hereditary cancer families. The contribution of mutations in moderate penetrance genes to hereditary cancer risk is not well established. Here, we report a family with early onset breast and fallopian tube cancer that was identified as carrying germline mutations in BARD1 and ATM genes. Loss of heterozygosity studies suggest a causative role of the BARD1 mutation in the development of primary peritoneal cancer, but fail to confirm an association between germline ATM mutations and breast cancer development in this family. Complexities in interpreting implications of mutations in moderate-risk cancer susceptibility genes are discussed.


Assuntos
Proteínas Mutadas de Ataxia Telangiectasia/genética , Neoplasias da Mama/genética , Predisposição Genética para Doença/genética , Neoplasias Ovarianas/genética , Proteínas Supressoras de Tumor/genética , Ubiquitina-Proteína Ligases/genética , Adulto , Idoso , Diagnóstico Diferencial , Feminino , Humanos , Mutação , Linhagem
15.
Clin Cancer Res ; 23(4): 935-945, 2017 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-27683182

RESUMO

Purpose: Despite the efficacy of abiraterone, a CYP17A1 inhibitor, in metastatic castration-resistant prostate cancer (CRPC), nearly all patients develop resistance. The purpose of this phase II study was to evaluate mechanisms of resistance to more complete androgen synthesis inhibition with abiraterone and dutasteride.Experimental Design: Eligible patients with metastatic CRPC underwent a baseline metastasis biopsy. Patients received abiraterone and prednisone for two 4-week cycles. After this time, high-dose dutasteride (3.5 mg daily) was added. Patients continued therapy until study withdrawal or radiographic progression. Repeat metastasis biopsy was obtained at progression. The primary endpoint was to assess mechanisms of resistance. Serum hormone and abiraterone levels were assessed. Tissue was assessed for androgen receptor (AR) and AR splice variant-7 (ARV7) expression.Results: Forty patients were enrolled. Sixty percent (n = 24) achieved a ≥50% reduction in prostate-specific antigen (PSA). The median time to radiographic progression was 11 months. Nearly all baseline (n = 29 of 31) and posttreatment (n = 16 of 16) tumors tested for AR nuclear expression were positive. Of those tested, ARV7 expression was present in 48% (n = 10 of 21) of baseline and 42% (n = 5 of 12) of treatment discontinuation specimens. Compared with patients with higher serum abiraterone levels at treatment discontinuation, patients with lower levels had higher circulating androgens.Conclusions: Despite increased androgen synthesis inhibition, we demonstrate that tumor AR axis remains important in disease progression. We highlight that abiraterone metabolism and pharmacokinetics may play a role in resistance. The noncomparative design limits conclusions on the efficacy of dual therapy with abiraterone and dutasteride, but the results support development of further multifaceted approaches toward AR inhibition. Clin Cancer Res; 23(4); 935-45. ©2016 AACR.


Assuntos
Androstenos/administração & dosagem , Dutasterida/administração & dosagem , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Receptores Androgênicos/genética , Idoso , Androgênios/biossíntese , Androgênios/genética , Androstenos/sangue , Progressão da Doença , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , Neoplasias de Próstata Resistentes à Castração/diagnóstico por imagem , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/patologia , Radiografia
16.
Clin Cancer Res ; 22(14): 3672-82, 2016 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-26936914

RESUMO

PURPOSE: ErbB2 signaling appears to be increased and may enhance androgen receptor (AR) activity in a subset of patients with castration-resistant prostate cancer (CRPC), but agents targeting ErbB2 have not been effective. This study was undertaken to assess ErbB2 activity in abiraterone-resistant prostate cancer and to determine whether it may contribute to AR signaling in these tumors. EXPERIMENTAL DESIGN: AR activity and ErbB2 signaling were examined in the radical prostatectomy specimens from a neoadjuvant clinical trial of leuprolide plus abiraterone and in the specimens from abiraterone-resistant CRPC xenograft models. The effect of ErbB2 signaling on AR activity was determined in two CRPC cell lines. Moreover, the effect of combination treatment with abiraterone and an ErbB2 inhibitor was assessed in a CRPC xenograft model. RESULTS: We found that ErbB2 signaling was elevated in residual tumor following abiraterone treatment in a subset of patients and was associated with higher nuclear AR expression. In xenograft models, we similarly demonstrated that ErbB2 signaling was increased and associated with AR reactivation in abiraterone-resistant tumors. Mechanistically, we show that ErbB2 signaling and subsequent activation of the PI3K/AKT signaling stabilizes AR protein. Furthermore, concomitantly treating CRPC cells with abiraterone and an ErbB2 inhibitor, lapatinib, blocked AR reactivation and suppressed tumor progression. CONCLUSIONS: ErbB2 signaling is elevated in a subset of patients with abiraterone-resistant prostate cancer and stabilizes AR protein. Combination therapy with abiraterone and ErbB2 antagonists may be effective for treating the subset of CRPC with elevated ErbB2 activity. Clin Cancer Res; 22(14); 3672-82. ©2016 AACR.


Assuntos
Androstenos/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Neoplasias de Próstata Resistentes à Castração/genética , Receptor ErbB-2/genética , Receptores Androgênicos/genética , Transdução de Sinais/genética , Androgênios/genética , Animais , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Leuprolida/farmacologia , Masculino , Camundongos , Camundongos SCID , Fosfatidilinositol 3-Quinases/genética , Próstata/efeitos dos fármacos , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
17.
Urol Oncol ; 33(9): 387.e17-27, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-25618296

RESUMO

INTRODUCTION: L-Selectin (CD62L) is a vascular adhesion molecule constitutively expressed on leukocytes with a primary function of directing leukocyte migration and homing of lymphocytes to lymph nodes. In a gene expression microarray study comparing laser-captured microdissected high-grade muscle-invasive bladder cancer (MIBC) without prior treatment and low-grade bladder cancer (LGBC) human samples, we found CD62L to be the highest differentially expressed gene. We sought to examine the differential expression of CD62L in MIBCs and its clinical relevance. METHODS: Unfixed fresh and formalin-fixed paraffin-embedded human bladder cancer specimens and serum samples were obtained from the University of Connecticut Health Center tumor bank. Tumor cells were isolated from frozen tumor tissue sections by laser-captured microdissected followed by RNA isolation. Quantitative polymerase chain reaction was used to validate the level of CD62L transcripts. Immunohistochemistry and enzyme-linked immunosorbent assay were performed to evaluate the CD62L protein localization and expression level. Flow cytometry was used to identify the relative number of cells expressing CD62L in fresh tumor tissue. In silico studies were performed using the Oncomine database. RESULTS: Immunostaining showed a uniformly higher expression of CD62L in MIBC specimens vs. LGBCs specimens. Further, CD62L localization was seen in foci of metastatic tumor cells in lymph node specimens from patients with high-grade MIBC and known nodal involvement. Up-regulated expression of CD62L was also observed by flow cytometric analysis of freshly isolated tumor cells from biopsies of high-grade cancers vs. LGBC specimens. Circulating CD62L levels were also found to be higher in serum samples from patients with high-grade metastatic vs. high-grade nonmetastatic MIBC. In addition, in silico analysis of Oncomine Microarray Database showed a significant correlation between CD62L expression and tumor aggressiveness and clinical outcomes. CONCLUSION: These data confirm the expression of CD62L on urothelial carcinoma cells and suggest that CD62L may serve as biomarker to predict the presence of or risk for developing metastatic disease in patients with bladder cancer.


Assuntos
Biomarcadores Tumorais/análise , Carcinoma de Células de Transição/patologia , Selectina L/biossíntese , Neoplasias da Bexiga Urinária/patologia , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Humanos , Imuno-Histoquímica , Selectina L/análise , Microdissecção e Captura a Laser , Masculino , Gradação de Tumores , Metástase Neoplásica , Reação em Cadeia da Polimerase , Transcriptoma , Regulação para Cima
18.
Clin Cancer Res ; 21(6): 1273-80, 2015 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-25320358

RESUMO

PURPOSE: The CYP17A1 inhibitor abiraterone markedly reduces androgen precursors and is thereby effective in castration-resistant prostate cancer (CRPC). However, abiraterone increases progesterone, which can activate certain mutant androgen receptors (AR) identified previously in flutamide-resistant tumors. Therefore, we sought to determine if CYP17A1 inhibitor treatment selects for progesterone-activated mutant ARs. EXPERIMENTAL DESIGN: AR was examined by targeted sequencing in metastatic tumor biopsies from 18 patients with CRPC who were progressing on a CYP17A1 inhibitor (17 on abiraterone, 1 on ketoconazole), alone or in combination with dutasteride, and by whole-exome sequencing in residual tumor in one patient treated with neoadjuvant leuprolide plus abiraterone. RESULTS: The progesterone-activated T878A-mutant AR was present at high allele frequency in 3 of the 18 CRPC cases. It was also present in one focus of resistant tumor in the neoadjuvant-treated patient, but not in a second clonally related resistant focus that instead had lost one copy of PTEN and both copies of CHD1. The T878A mutation appeared to be less common in the subset of patients with CRPC treated with abiraterone plus dutasteride, and transfection studies showed that dutasteride was a more potent direct antagonist of the T878A versus the wild-type AR. CONCLUSIONS: These findings indicate that selection for tumor cells expressing progesterone-activated mutant ARs is a mechanism of resistance to CYP17A1 inhibition.


Assuntos
Androstenos/uso terapêutico , Progesterona/metabolismo , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Receptores Androgênicos/genética , Esteroide 17-alfa-Hidroxilase/antagonistas & inibidores , Idoso , Idoso de 80 Anos ou mais , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Linhagem Celular Tumoral , DNA Helicases/genética , Proteínas de Ligação a DNA/genética , Resistencia a Medicamentos Antineoplásicos/genética , Dutasterida/uso terapêutico , Humanos , Cetoconazol/uso terapêutico , Leuprolida/uso terapêutico , Masculino , Pessoa de Meia-Idade , Mutação/genética , PTEN Fosfo-Hidrolase/genética , Neoplasias de Próstata Resistentes à Castração/genética , Receptores Androgênicos/metabolismo
19.
Clin Cancer Res ; 20(6): 1590-600, 2014 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-24449822

RESUMO

PURPOSE: Mechanisms mediating androgen receptor (AR) reactivation in prostate cancer that progresses after castration (castration-resistant prostate cancer; CRPC) and subsequent treatment with abiraterone (CYP17A1 inhibitor that further suppresses androgen synthesis) remain unclear. EXPERIMENTAL DESIGN: Prostate cancer xenografts were examined to identify mechanism of progression after castration and abiraterone. RESULTS: AR reactivation in abiraterone-resistant VCaP xenografts was not associated with restoration of intratumoral androgens or alterations in AR coregulators. In contrast, mRNA encoding full-length AR (AR-FL) and a constitutively active splice variant (AR-V7) were increased compared with xenografts before castration, with an increase in AR-V7 relative to AR-FL. This shift toward AR-V7 was due to a feedback mechanism whereby the androgen-liganded AR stimulates expression of proteins that suppress generation of AR-V7 relative to AR-FL transcripts. However, despite the increases in AR-V7 mRNA, it remained a minor transcript (<1%) relative to AR-FL in resistant VCaP xenografts and CRPC clinical samples. AR-V7 protein expression was similarly low relative to AR-FL in castration-resistant VCaP xenografts and androgen-deprived VCaP cells, but the weak basal AR activity in these latter cells was further repressed by AR-V7 siRNA. CONCLUSIONS: AR-V7 at these low levels is not adequate to restore AR activity, but its rapid induction after androgen deprivation allows tumors to retain basal AR activity that may be needed for survival until more potent mechanisms emerge to activate AR. Agents targeting AR splice variants may be most effective when used very early in conjunction with therapies targeting the AR ligand-binding domain.


Assuntos
Processamento Alternativo , Antagonistas de Androgênios/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Neoplasias da Próstata/genética , Receptores Androgênicos/genética , Androstenos , Androstenóis , Animais , Castração , Humanos , Immunoblotting , Masculino , Camundongos , Neoplasias da Próstata/metabolismo , Isoformas de Proteínas , RNA Interferente Pequeno , Receptores Androgênicos/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transfecção , Ensaios Antitumorais Modelo de Xenoenxerto
20.
Bone ; 56(1): 31-41, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23639875

RESUMO

Intermittent PTH is the major anabolic therapy for osteoporosis while continuous PTH causes bone loss. PTH acts on the osteoblast (OB) lineage to regulate bone resorption and formation. PTH also induces cyclooxygenase-2 (COX-2), producing prostaglandin E2 (PGE(2)) that can act on both OBs and osteoclasts (OCs). Because intermittent PTH is more anabolic in Cox-2 knockout (KO) than wild type (WT) mice, we hypothesized COX-2 might contribute to the effects of continuous PTH by suppressing PTH-stimulated differentiation of mesenchymal stem cells into OBs. We compared effects of continuous PTH on bone marrow stromal cells (BMSCs) and primary OBs (POBs) from Cox-2 KO mice, mice with deletion of PGE(2) receptors (Ptger(4) and Ptger(2) KO mice), and WT controls. PTH increased OB differentiation in BMSCs only in the absence of COX-2 expression or activity. In the absence of COX-2, PTH stimulated differentiation if added during the first week of culture. In Cox-2 KO BMSCs, PTH-stimulated differentiation was prevented by adding PGE(2) to cultures. Co-culture of POBs with M-CSF-expanded bone marrow macrophages (BMMs) showed that the inhibition of PTH-stimulated OB differentiation required not only COX-2 or PGE(2) but also BMMs. Sufficient PGE(2) to mediate the inhibitory effect was made by either WT POBs or WT BMMs. The inhibitory effect mediated by COX-2/PGE(2) was transferred by conditioned media from RANKL-treated BMMs and could be blocked by osteoprotegerin, which interferes with RANKL binding to its receptor on OC lineage cells. Deletion of Ptger(4), but not Ptger(2), in BMMs prevented the inhibition of PTH-stimulated OB differentiation. As expected, PGE(2) also stimulated OB differentiation, but when given in combination with PTH, the stimulatory effects of both were abrogated. These data suggest that PGE(2), acting via EP4R on BMMs committed to the OC lineage, stimulated secretion of a factor or factors that acted to suppress PTH-stimulated OB differentiation. This suppression of OB differentiation could contribute to the bone loss seen with continuous PTH in vivo.


Assuntos
Células da Medula Óssea/citologia , Diferenciação Celular/efeitos dos fármacos , Dinoprostona/farmacologia , Macrófagos/metabolismo , Osteoblastos/citologia , Hormônio Paratireóideo/farmacologia , Adipogenia/efeitos dos fármacos , Adipogenia/genética , Animais , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Bovinos , Diferenciação Celular/genética , Células Cultivadas , Meios de Cultivo Condicionados/farmacologia , Ciclo-Oxigenase 2/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Sistema Hematopoético/citologia , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Camundongos , Camundongos Knockout , Osteoblastos/efeitos dos fármacos , Osteoblastos/enzimologia , Osteocalcina/genética , Osteocalcina/metabolismo , Ligante RANK/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Prostaglandina E Subtipo EP2/deficiência , Receptores de Prostaglandina E Subtipo EP2/metabolismo , Receptores de Prostaglandina E Subtipo EP4/deficiência , Receptores de Prostaglandina E Subtipo EP4/metabolismo , Células Estromais/citologia , Células Estromais/efeitos dos fármacos , Células Estromais/enzimologia
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