RESUMEN
The clinical success of combined androgen deprivation therapy (ADT) and radiotherapy (RT) in prostate cancer created interest in understanding the mechanistic links between androgen receptor (AR) signaling and the DNA damage response (DDR). Convergent data have led to a model where AR both regulates, and is regulated by, the DDR. Integral to this model is that the AR regulates the transcription of DDR genes both at a steady state and in response to ionizing radiation (IR). In this study, we sought to determine which immediate transcriptional changes are induced by IR in an AR-dependent manner. Using PRO-seq to quantify changes in nascent RNA transcription in response to IR, the AR antagonist enzalutamide, or the combination of the two, we find that enzalutamide treatment significantly decreased expression of canonical AR target genes but had no effect on DDR gene sets in prostate cancer cells. Surprisingly, we also found that the AR is not a primary regulator of DDR genes either in response to IR or at a steady state in asynchronously growing prostate cancer cells. IMPLICATIONS: Our data indicate that the clinical benefit of combining ADT with RT is not due to direct AR regulation of DDR gene transcription, and that the field needs to consider alternative mechanisms for this clinical benefit.
Asunto(s)
Neoplasias de la Próstata Resistentes a la Castración , Neoplasias de la Próstata , Masculino , Humanos , Receptores Androgénicos/genética , Receptores Androgénicos/metabolismo , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/metabolismo , Antagonistas de Andrógenos/farmacología , Línea Celular Tumoral , Daño del ADN , Neoplasias de la Próstata Resistentes a la Castración/genéticaRESUMEN
As mobile and wearable devices continue to grow in popularity, there is strong yet unrealized potential to harness people's mobile sensing data to improve our understanding of their cellular and biologically-based diseases. Breakthrough technical innovations in tumor modeling, such as the three dimensional tumor microenvironment system (TMES), allow researchers to study the behavior of tumor cells in a controlled environment that closely mimics the human body. Although patients' health behaviors are known to impact their tumor growth through circulating hormones (cortisol, melatonin), capturing this process is a challenge to rendering realistic tumor models in the TMES or similar tumor modeling systems. The goal of this paper is to propose a conceptual framework that unifies researchers from digital health, data science, oncology, and cellular signaling, in a common cause to improve cancer patients' treatment outcomes through mobile sensing. In support of our framework, existing studies indicate that it is feasible to use people's mobile sensing data to approximate their underlying hormone levels. Further, it was found that when cortisol is cycled through the TMES based on actual patients' cortisol levels, there is a significant increase in pancreatic tumor cell growth compared to when cortisol levels are at normal healthy levels. Taken together, findings from these studies indicate that continuous monitoring of people's hormone levels through mobile sensing may improve experimentation in the TMES, by informing how hormones should be introduced. We hope our framework inspires digital health researchers in the psychosocial sciences to consider how their expertise can be applied to advancing outcomes across levels of inquiry, from behavioral to cellular.
RESUMEN
Prostate cancer (PCa) is a very complex disease that is a major cause of death in men worldwide. Currently, PCa dependence on the androgen receptor (AR) has resulted in use of AR antagonists and antiandrogen therapies that reduce endogenous steroid hormone production. However, within two to three years of receiving first-line androgen deprivation therapy, the majority of patients diagnosed with PCa progress to castration-resistant prostate cancer (CRPC). There is an urgent need for therapies that are more durable than antagonism of the AR axis. Studies of runt-related transcription factors (RUNX) and their heterodimerization partner, core-binding factor subunit b (CBFß), are revealing that the RUNX family are drivers of CRPC. In this review, we describe what is presently understood about RUNX members in PCa, including what regulates and is regulated by RUNX proteins, and the role of RUNX proteins in the tumor microenvironment and AR signaling. We discuss the implications for therapeutically targeting RUNX, the potential for RUNX as PCa biomarkers, and the current pressing questions in the field.
Asunto(s)
Subunidades alfa del Factor de Unión al Sitio Principal/metabolismo , Neoplasias de la Próstata Resistentes a la Castración/metabolismo , Neoplasias de la Próstata/metabolismo , Factores de Transcripción/metabolismo , Antagonistas de Andrógenos/uso terapéutico , Humanos , Masculino , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata Resistentes a la Castración/tratamiento farmacológico , Receptores Androgénicos/metabolismoRESUMEN
Lung cancer rates are rising globally and non-small cell lung cancer (NSCLC) has a five year survival rate of only 24%. Unfortunately, the development of drugs to treat cancer is severely hampered by the inefficiency of translating pre-clinical studies into clinical benefit. Thus, we sought to apply a tumor microenvironment system (TMES) to NSCLC. Using microvascular endothelial cells, lung cancer derived fibroblasts, and NSCLC tumor cells in the presence of in vivo tumor-derived hemodynamic flow and transport, we demonstrate that the TMES generates an in-vivo like biological state and predicts drug response to EGFR inhibitors. Transcriptomic and proteomic profiling indicate that the TMES recapitulates the in vivo and patient molecular biological state providing a mechanistic rationale for the predictive nature of the TMES. This work further validates the TMES for modeling patient tumor biology and drug response indicating utility of the TMES as a predictive tool for drug discovery and development and potential for use as a system for patient avatars.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Células Endoteliales/metabolismo , Neoplasias Pulmonares/metabolismo , Modelos Biológicos , Microambiente Tumoral , Animales , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Células Endoteliales/patología , Humanos , Neoplasias Pulmonares/patología , Ratones , Ratones Desnudos , Ratones SCIDRESUMEN
We have previously demonstrated that checkpoint kinase 2 (CHK2) is a critical negative regulator of androgen receptor (AR) transcriptional activity, prostate cancer (PCa) cell growth, and androgen sensitivity. We have now uncovered that the AR directly interacts with CHK2 and ionizing radiation (IR) increases this interaction. This IR-induced increase in AR-CHK2 interactions requires AR phosphorylation and CHK2 kinase activity. PCa associated CHK2 mutants with impaired kinase activity reduced IR-induced AR-CHK2 interactions. The destabilization of AR - CHK2 interactions induced by CHK2 variants impairs CHK2 negative regulation of cell growth. CHK2 depletion increases transcription of DNAPK and RAD54, increases clonogenic survival, and increases resolution of DNA double strand breaks. The data support a model where CHK2 sequesters the AR through direct binding decreasing AR transcription and suppressing PCa cell growth. CHK2 mutation or loss of expression thereby leads to increased AR transcriptional activity and survival in response to DNA damage.
Asunto(s)
Quinasa de Punto de Control 2/metabolismo , Neoplasias de la Próstata/metabolismo , Receptores Androgénicos/metabolismo , Línea Celular Tumoral , Supervivencia Celular , Quinasa de Punto de Control 2/genética , Reparación del ADN , Humanos , Inmunoprecipitación , Masculino , Fosforilación , Unión Proteica , Radiación IonizanteRESUMEN
Herpes simplex virus (HSV) entry into cells is a multistep process that engages the host cell machinery. The proteasome is a large, ATP-dependent, multisubunit protease that plays a critical role in the maintenance of cell homeostasis. A battery of assays were used to demonstrate that proteasome inhibitors blocked an early step in HSV entry that occurred after capsid penetration into the cytosol but prior to capsid arrival at the nuclear periphery. Proteasome-dependent viral entry was not reliant on host or viral protein synthesis. MG132, a peptide aldehyde that competitively inhibits the degradative activity of the proteasome, had a reversible inhibitory effect on HSV entry. HSV can use endocytic or nonendocytic pathways to enter cells. These distinct entry routes were both dependent on proteasome-mediated proteolysis. In addition, HSV successfully entered cells in the absence of a functional host ubiquitin-activating enzyme, suggesting that viral entry is ubiquitin independent. We propose that proteasomal degradation of virion and/or host proteins is required for efficient delivery of incoming HSV capsids to the nucleus.
Asunto(s)
Complejo de la Endopetidasa Proteasomal/metabolismo , Simplexvirus/fisiología , Internalización del Virus , Animales , Línea Celular , Chlorocebus aethiops , Cricetinae , Inhibidores de Cisteína Proteinasa/farmacología , Genes Reporteros , Humanos , Leupeptinas/farmacología , Ratones , Inhibidores de Proteasoma , Enzimas Activadoras de Ubiquitina/metabolismo , Ensayo de Placa Viral , beta-Galactosidasa/genética , beta-Galactosidasa/metabolismoRESUMEN
The development of drugs to treat cancer is hampered by the inefficiency of translating pre-clinical in vitro monoculture and mouse studies into clinical benefit. There is a critical need to improve the accuracy of evaluating pre-clinical drug efficacy through the development of more physiologically relevant models. In this study, a human triculture 3D in vitro tumor microenvironment system (TMES) was engineered to accurately mimic the tumor microenvironment. The TMES recapitulates tumor hemodynamics and biological transport with co-cultured human microvascular endothelial cells, pancreatic ductal adenocarcinoma, and pancreatic stellate cells. We demonstrate that significant tumor cell transcriptomic changes occur in the TMES that correlate with the in vivo xenograft and patient transcriptome. Treatment with therapeutically relevant doses of chemotherapeutics yields responses paralleling the patients' clinical responses. Thus, this model provides a unique platform to rigorously evaluate novel therapies and is amenable to using patient tumor material directly, with applicability for patient avatars.
Asunto(s)
Biomimética/métodos , Carcinoma Ductal Pancreático/patología , Microambiente Tumoral , Proliferación Celular/efectos de los fármacos , Humanos , Microambiente Tumoral/efectos de los fármacosRESUMEN
Over half of BRAFV600E melanomas display intrinsic resistance to BRAF inhibitors, in part due to adaptive signaling responses. In this communication we ask whether BRAFV600E melanomas share common adaptive responses to BRAF inhibition that can provide clinically relevant targets for drug combinations. We screened a panel of 12 treatment-naïve BRAFV600E melanoma cell lines with MAP Kinase pathway inhibitors in pairwise combination with 58 signaling inhibitors, assaying for synergistic cytotoxicity. We found enormous diversity in the drug combinations that showed synergy, with no two cell lines having an identical profile. Although the 6 lines most resistant to BRAF inhibition showed synergistic benefit from combination with lapatinib, the signaling mechanisms by which this combination generated synergistic cytotoxicity differed between the cell lines. We conclude that adaptive responses to inhibition of the primary oncogenic driver (BRAFV600E) are determined not only by the primary oncogenic driver but also by diverse secondary genetic and epigenetic changes ("back-seat drivers") and hence optimal drug combinations will be variable. Because upregulation of receptor tyrosine kinases is a major source of drug resistance arising from diverse adaptive responses, we propose that inhibitors of these receptors may have substantial clinical utility in combination with inhibitors of the MAP Kinase pathway.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Evaluación Preclínica de Medicamentos , Resistencia a Antineoplásicos/genética , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Melanoma/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas B-raf/antagonistas & inhibidores , Animales , Línea Celular Tumoral , Humanos , Indoles/farmacología , Lapatinib , Melanoma/genética , Ratones , Ratones Desnudos , Ratones SCID , Proteínas Proto-Oncogénicas B-raf/genética , Quinazolinas/farmacología , Sulfonamidas/farmacologíaRESUMEN
Recent data show that extracellular signals are transmitted through a network of proteins rather than hierarchical signaling pathways, suggesting that the inhibition of a single component of a canonical pathway is insufficient for the treatment of cancer. The biologic outcome of signaling through a network is inherently more robust and resistant to inhibition of a single network component. In this study, we conducted a functional chemical genetic screen to identify novel interactions between signaling inhibitors that would not be predicted on the basis of our current understanding of signaling networks. We screened over 300 drug combinations in nine melanoma cell lines and have identified pairs of compounds that show synergistic cytotoxicity. The synergistic cytotoxicities identified did not correlate with the known RAS and BRAF mutational status of the melanoma cell lines. Among the most robust results was synergy between sorafenib, a multikinase inhibitor with activity against RAF, and diclofenac, a nonsteroidal anti-inflammatory drug (NSAID). Drug substitution experiments using the NSAIDs celecoxib and ibuprofen or the MAP-ERK kinase inhibitor PD325901 and the RAF inhibitor RAF265 suggest that inhibition of COX and mitogen-activated protein kinase signaling are targets for the synergistic cytotoxicity of sorafenib and diclofenac. Cotreatment with sorafenib and diclofenac interrupts a positive feedback signaling loop involving extracellular signal-regulated kinase, cellular phospholipase A2, and COX. Genome-wide expression profiling shows synergy-specific downregulation of survival-related genes. This study has uncovered novel functional drug combinations and suggests that the underlying signaling networks that control responses to targeted agents can vary substantially, depending on unexplored components of the cell genotype.
Asunto(s)
Antineoplásicos/análisis , Antineoplásicos/farmacología , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Melanoma/tratamiento farmacológico , Transducción de Señal/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/análisis , Bibliotecas de Moléculas Pequeñas/farmacología , Antineoplásicos/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Muerte Celular/efectos de los fármacos , Línea Celular Tumoral , Diclofenaco/farmacología , Diclofenaco/uso terapéutico , Sinergismo Farmacológico , Quinasas MAP Reguladas por Señal Extracelular/antagonistas & inhibidores , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Genoma Humano/genética , Humanos , Melanoma/enzimología , Melanoma/genética , Melanoma/patología , Niacinamida/análogos & derivados , Niacinamida/farmacología , Niacinamida/uso terapéutico , Compuestos de Fenilurea/farmacología , Compuestos de Fenilurea/uso terapéutico , Fosfolipasas A2 Citosólicas/metabolismo , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/enzimología , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/patología , Bibliotecas de Moléculas Pequeñas/uso terapéutico , SorafenibRESUMEN
Herpesviruses commandeer distinct cellular pathways to enter target cells. The mechanism by which herpes simplex virus (HSV) selects a pH-dependent, endocytic route or a pH-independent route remains to be elucidated. We investigated the role of the non-glycosylated viral envelope protein UL45 in HSV entry via endocytosis. UL45 plays a role in mediating cell-cell fusion and has been proposed to functionally interact with gB to regulate membrane fusion. Thus, we also probed the impact of UL45 on the structure and function of gB present in virions. A UL45 deletion virus successfully entered cells via low pH, endocytic pathway with wild type kinetics. In the absence or presence of UL45, the antigenic conformation of virion gB appeared unaltered. Antibodies to gB neutralized infection of the UL45-deletion virus and wild type virus to a similar extent, regardless of whether the target cells supported low pH endocytic or non-endocytic entry routes. Lastly, HSV virions were inactivated by low pH regardless of the presence of UL45. The results, together with previous studies, suggest that UL45 plays distinct roles in cell-cell fusion and virus-cell fusion during acid-dependent entry.
Asunto(s)
Endocitosis , Simplexvirus/fisiología , Proteínas del Envoltorio Viral/fisiología , Proteínas Virales/fisiología , Internalización del Virus , Animales , Células CHO , Cricetinae , Cricetulus , Eliminación de Gen , Concentración de Iones de Hidrógeno , Conformación Proteica , Simplexvirus/genética , Proteínas del Envoltorio Viral/química , Proteínas Virales/genéticaRESUMEN
Fusion-from-without (FFWO) is the rapid induction of cell fusion by virions in the absence of viral protein synthesis. The combination of two amino acid mutations in envelope glycoprotein B (gB), one in the ectodomain and one in the cytoplasmic tail, can confer FFWO activity to wild type herpes simplex virus (HSV). In this report, we analyzed the entry and cell fusion phenotypes of HSV that contains FFWO gB, with emphasis on the cellular receptors for HSV, nectin-1, nectin-2 and HVEM. The ability of an HSV strain with FFWO gB to efficiently mediate FFWO via a specific gD-receptor correlated with its ability to mediate viral entry by that receptor. A FFWO form of gB was not sufficient to switch the entry of HSV from a pH-dependent, endocytic pathway to a direct fusion, pH-independent pathway. The conformation of gB with FFWO activity was not globally altered relative to wild type. However, distinct monoclonal antibodies had reduced reactivity with FFWO gB, suggesting an altered antigenic structure relative to wild type. FFWO was blocked by preincubation of virions with neutralizing antibodies to gB or gD. Together with previous studies, the results indicate that the roles of gB in FFWO and in virus-cell fusion during entry are related but not identical. This study also suggests that the FFWO function of gB is not a specific determinant for the selection of HSV entry pathway and that antigenic differences in FFWO gB may reflect its enhanced fusion activity.