RESUMEN
Apoptosis is characterized by membrane blebbing and apoptotic body formation. Caspase cleavage of ROCK1 generates an active fragment that promotes actin-myosin-mediated contraction and membrane blebbing during apoptosis. Expression of caspase-resistant non-cleavable ROCK1 (Rock1 NC) prolonged survival of mice that rapidly develop B cell lymphomas due to Eµ-Myc transgene expression. Eµ-Myc; Rock1 NC mice had significantly fewer bone marrow cells relative to those in Eµ-Myc mice expressing wild-type ROCK1 (Rock1 WT), which was associated with altered cell cycle profiles. Circulating macrophage numbers were lower in Eµ-Myc; Rock1 NC mice, but there were higher levels of bone marrow macrophages, consistent with spontaneous cell death in Eµ-Myc; Rock1 NC mouse bone marrows being more inflammatory. Rock1 WT recipient mice transplanted with pre-neoplastic Eµ-Myc; Rock1 NC bone marrow cells survived longer than mice transplanted with Eµ-Myc; Rock1 WT cells, indicating that the survival benefit was intrinsic to the Eµ-Myc; Rock1 NC bone marrow cells. The results suggest that the apoptotic death of Eµ-Myc; Rock1 NC cells generates a proliferation-suppressive microenvironment in bone marrows that reduces cell numbers and prolongs B cell lymphoma mouse survival.
Asunto(s)
Caspasas , Linfoma de Células B , Proteínas Proto-Oncogénicas c-myc , Quinasas Asociadas a rho , Animales , Ratones , Apoptosis , Células de la Médula Ósea/metabolismo , Caspasas/metabolismo , Ciclo Celular , Linfoma de Células B/patología , Linfoma de Células B/genética , Macrófagos/metabolismo , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas Proto-Oncogénicas c-myc/metabolismo , Quinasas Asociadas a rho/genética , Quinasas Asociadas a rho/metabolismo , Análisis de SupervivenciaRESUMEN
The actin and microtubule cytoskeletons are critically important for cancer cell proliferation, and drugs that target microtubules are widely-used cancer therapies. However, their utility is compromised by toxicities due to dose and exposure. To overcome these issues, we characterized how inhibition of the actin and microtubule cytoskeleton regulatory LIM kinases could be used in drug combinations to increase efficacy. A previously-described LIMK inhibitor (LIMKi) induced dose-dependent microtubule alterations that resulted in significant mitotic defects, and increased the cytotoxic potency of microtubule polymerization inhibitors. By combining LIMKi with 366 compounds from the GSK Published Kinase Inhibitor Set, effective combinations were identified with kinase inhibitors including EGFR, p38 and Raf. These findings encouraged a drug discovery effort that led to development of CRT0105446 and CRT0105950, which potently block LIMK1 and LIMK2 activity in vitro, and inhibit cofilin phosphorylation and increase αTubulin acetylation in cells. CRT0105446 and CRT0105950 were screened against 656 cancer cell lines, and rhabdomyosarcoma, neuroblastoma and kidney cancer cells were identified as significantly sensitive to both LIMK inhibitors. These large-scale screens have identified effective LIMK inhibitor drug combinations and sensitive cancer types. In addition, the LIMK inhibitory compounds CRT0105446 and CRT0105950 will enable further development of LIMK-targeted cancer therapy.
Asunto(s)
Quinasas Lim/antagonistas & inhibidores , Mitosis/efectos de los fármacos , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/enzimología , Neoplasias de la Mama/patología , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/enzimología , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Femenino , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/enzimología , Neoplasias Pulmonares/patología , Células MCF-7 , Microtúbulos/metabolismo , Mitosis/fisiología , Neoplasias/enzimología , Neuroblastoma/tratamiento farmacológico , Neuroblastoma/enzimología , Neuroblastoma/patologíaRESUMEN
Prostate cancer affects a large proportion of the male population, and is primarily driven by androgen receptor (AR) activity. First-line treatment typically consists of reducing AR signaling by hormone depletion, but resistance inevitably develops over time. One way to overcome this issue is to block AR function via alternative means, preferably by inhibiting protein targets that are more active in tumors than in normal tissue. By staining prostate cancer tumor sections, elevated LIM kinase 1 (LIMK1) expression and increased phosphorylation of its substrate Cofilin were found to be associated with poor outcome and reduced survival in patients with nonmetastatic prostate cancer. A LIMK-selective small molecule inhibitor (LIMKi) was used to determine whether targeted LIMK inhibition was a potential prostate cancer therapy. LIMKi reduced prostate cancer cell motility, as well as inhibiting proliferation and increasing apoptosis in androgen-dependent prostate cancer cells more effectively than in androgen-independent prostate cancer cells. LIMK inhibition blocked ligand-induced AR nuclear translocation, reduced AR protein stability and transcriptional activity, consistent with its effects on proliferation and survival acting via inhibition of AR activity. Furthermore, inhibition of LIMK activity increased αTubulin acetylation and decreased AR interactions with αTubulin, indicating that the role of LIMK in regulating microtubule dynamics contributes to AR function. These results indicate that LIMK inhibitors could be beneficial for the treatment of prostate cancer both by reducing nuclear AR translocation, leading to reduced proliferation and survival, and by inhibiting prostate cancer cell dissemination.
Asunto(s)
Núcleo Celular/metabolismo , Quinasas Lim/metabolismo , Neoplasias de la Próstata/metabolismo , Receptores Androgénicos/metabolismo , Bibliotecas de Moléculas Pequeñas/farmacología , Factores Despolimerizantes de la Actina/metabolismo , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Quinasas Lim/antagonistas & inhibidores , Quinasas Lim/genética , Masculino , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/mortalidad , Receptores Androgénicos/genética , Análisis de Supervivencia , Tubulina (Proteína)/genética , Tubulina (Proteína)/metabolismoRESUMEN
Accumulating evidence from basic and clinical studies supports the concept that signaling pathways downstream of Rho GTPases play important roles in tumor development and progression. As a result, there has been considerable interest in the possibility that specific proteins in these signal transduction pathways could be potential targets for cancer therapy. A number of inhibitors targeting critical effector proteins, activators or the Rho GTPases themselves, have been developed. We will review the strategies currently being used to develop inhibitors of Rho GTPases and downstream signaling kinases and discuss candidate entities. Although molecularly targeted drugs that inhibit Rho GTPase signaling have not yet been widely adopted for clinical use, their potential value as cancer therapeutics continues to drive considerable pharmaceutical research and development.
Asunto(s)
Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Inhibidores Enzimáticos/farmacología , Inhibidores Enzimáticos/uso terapéutico , Neoplasias/tratamiento farmacológico , Transducción de Señal/efectos de los fármacos , Proteínas de Unión al GTP rho/antagonistas & inhibidores , Animales , Citoesqueleto/metabolismo , Activación Enzimática/efectos de los fármacos , Factores de Intercambio de Guanina Nucleótido/antagonistas & inhibidores , Humanos , Quinasas Lim/antagonistas & inhibidores , Terapia Molecular Dirigida , Neoplasias/enzimología , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Quinasas p21 Activadas/antagonistas & inhibidores , Proteínas de Unión al GTP rho/metabolismo , Quinasas Asociadas a rho/antagonistas & inhibidoresRESUMEN
Insulin receptor substrate-2 (IRS-2) belongs to the IRS family of adaptor proteins that function as signaling intermediates for growth factor, cytokine, and integrin receptors, many of which have been implicated in cancer. Although the IRS proteins share significant homology, distinct functions have been attributed to each family member in both normal and tumor cells. In cancer, IRS-2 is positively associated with aggressive tumor behavior. In the current study, we show that IRS-2 expression, but not IRS-1 expression, is positively regulated by hypoxia, which selects for tumor cells with increased metastatic potential. We identify IRS-2 as a novel hypoxia-responsive gene and establish that IRS-2 gene transcription increases in a hypoxia-inducible factor-dependent manner in hypoxic environments. IRS-2 is active to mediate insulin-like growth factor I-dependent signals in hypoxia, and enhanced activation of Akt in hypoxia is dependent on IRS-2 expression. Functionally, the elevated expression of IRS-2 facilitates breast carcinoma cell survival and invasion in hypoxia. Collectively, our results reveal a novel mechanism by which IRS-2 contributes to the aggressive behavior of hypoxic tumor cells.
Asunto(s)
Neoplasias de la Mama/metabolismo , Proteínas Sustrato del Receptor de Insulina/biosíntesis , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Hipoxia de la Célula/fisiología , Línea Celular Tumoral , Supervivencia Celular/fisiología , Femenino , Humanos , Factor 1 Inducible por Hipoxia/metabolismo , Proteínas Sustrato del Receptor de Insulina/genética , Neoplasias Mamarias Experimentales/genética , Neoplasias Mamarias Experimentales/metabolismo , Neoplasias Mamarias Experimentales/patología , Ratones , Invasividad Neoplásica , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Transcripción GenéticaRESUMEN
The Insulin Receptor Substrate (IRS) proteins are cytoplasmic adaptor proteins that function as essential signaling intermediates downstream of activated cell surface receptors, many of which have been implicated in cancer. The IRS proteins do not contain any intrinsic kinase activity, but rather serve as scaffolds to organize signaling complexes and initiate intracellular signaling pathways. As common intermediates of multiple receptors that can influence tumor progression, the IRS proteins are positioned to play a pivotal role in regulating the response of tumor cells to many different microenvironmental stimuli. Limited studies on IRS expression in human tumors and studies on IRS function in human tumor cell lines and in mouse models have provided clues to the potential function of these adaptor proteins in human cancer. A general theme arises from these studies; IRS-1 and IRS-4 are most often associated with tumor growth and proliferation and IRS-2 is most often associated with tumor motility and invasion. In this review, we discuss the mechanisms by which IRS expression and function are regulated and how the IRS proteins contribute to tumor initiation and progression.
RESUMEN
Neonatal exposure of CD-1 mice to diethylstilbestrol (DES) or genistein (GEN) induces uterine adenocarcinoma in aging animals. Uterine carcinogenesis in this model is ovarian dependent because its evolution is blocked by prepubertal ovariectomy. This study seeks to discover novel uterine genes whose expression is altered by such early endocrine disruption via an epigenetic mechanism. Neonatal mice were treated with 1 or 1000 microg/kg DES, 50 mg/kg GEN, or oil (control) on d 1-5. One group of treated mice was killed before puberty on d 19. Others were ovariectomized or left intact, and killed at 6 and 18 months of age. Methylation-sensitive restriction fingerprinting was performed to identify differentially methylated sequences associated with neonatal exposure to DES/GEN. Among 14 candidates, nucleosomal binding protein 1 (Nsbp1), the gene for a nucleosome-core-particle binding protein, was selected for further study because of its central role in chromatin remodeling. In uteri of immature control mice, Nsbp1 promoter CpG island (CGI) was minimally methylated. Once control mice reached puberty, the Nsbp1 CGI became hypermethylated, and gene expression declined further. In contrast, in neonatal DES/GEN-treated mice, the Nsbp1 CGI stayed anomalously hypomethylated, and the gene exhibited persistent overexpression throughout life. However, if neonatal DES/GEN-treated mice were ovariectomized before puberty, the CGI remained minimally to moderately methylated, and gene expression was subdued except in the group treated with 1000 microg/kg DES. Thus, the life reprogramming of uterine Nsbp1 expression by neonatal DES/GEN exposure appears to be mediated by an epigenetic mechanism that interacts with ovarian hormones in adulthood.