RESUMO
Growth hormone-releasing hormone antagonists (GHRHAnt) have been associated with antitumor and antioxidative activities. The present study investigates for the first time the effects of those compounds towards pro-inflammatory cytokine expression in a murine model of cecal ligation and puncture (CLP) - induced sepsis. The results indicate that GHRHAnt JV-1-36 significantly suppressed IL-1α, IL-6, and pSTAT3 activation in septic lungs. Moreover, GHRHAnt treatment reduced bronchoalveolar lavage fluid (BALF) protein concentration, suggesting a protective effect of that compound in sepsis-induced lung edema. Based on those findings, it is suggested that GHRHAnt may represent an exciting new therapeutic possibility in sepsis-induced endotoxemia and lung injury.
Assuntos
Hormônio Liberador de Hormônio do Crescimento , Pulmão , Sepse , Animais , Sepse/tratamento farmacológico , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Hormônio Liberador de Hormônio do Crescimento/metabolismo , Camundongos , Pulmão/patologia , Pulmão/metabolismo , Pulmão/efeitos dos fármacos , Masculino , Fator de Transcrição STAT3/metabolismo , Líquido da Lavagem Broncoalveolar , Interleucina-6/metabolismo , Modelos Animais de DoençasRESUMO
COVID-19 is characterized by an excessive inflammatory response and macrophage hyperactivation, leading, in severe cases, to alveolar epithelial injury and acute respiratory distress syndrome. Recent studies have reported that SARS-CoV-2 spike (S) protein interacts with bacterial lipopolysaccharide (LPS) to boost inflammatory responses in vitro, in macrophages and peripheral blood mononuclear cells (PBMCs), and in vivo. The hypothalamic hormone growth hormone-releasing hormone (GHRH), in addition to promoting pituitary GH release, exerts many peripheral functions, acting as a growth factor in both malignant and non-malignant cells. GHRH antagonists, in turn, display potent antitumor effects and antinflammatory activities in different cell types, including lung and endothelial cells. However, to date, the antinflammatory role of GHRH antagonists in COVID-19 remains unexplored. Here, we examined the ability of GHRH antagonist MIA-602 to reduce inflammation in human THP-1-derived macrophages and PBMCs stimulated with S protein and LPS combination. Western blot and immunofluorescence analysis revealed the presence of GHRH receptor and its splice variant SV1 in both THP-1 cells and PBMCs. Exposure of THP-1 cells to S protein and LPS combination increased the mRNA levels and protein secretion of TNF-α and IL-1ß, as well as IL-8 and MCP-1 gene expression, an effect hampered by MIA-602. Similarly, MIA-602 hindered TNF-α and IL-1ß secretion in PBMCs and reduced MCP-1 mRNA levels. Mechanistically, MIA-602 blunted the S protein and LPS-induced activation of inflammatory pathways in THP-1 cells, such as NF-κB, STAT3, MAPK ERK1/2 and JNK. MIA-602 also attenuated oxidative stress in PBMCs, by decreasing ROS production, iNOS and COX-2 protein levels, and MMP9 activity. Finally, MIA-602 prevented the effect of S protein and LPS synergism on NF-кB nuclear translocation and activity. Overall, these findings demonstrate a novel antinflammatory role for GHRH antagonists of MIA class and suggest their potential development for the treatment of inflammatory diseases, such as COVID-19 and related comorbidities.
Assuntos
COVID-19 , Glicoproteína da Espícula de Coronavírus , Humanos , Células Endoteliais , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Inflamação/tratamento farmacológico , Leucócitos Mononucleares , Lipopolissacarídeos , SARS-CoV-2 , Fator de Necrose Tumoral alfaRESUMO
Endometrial stromal cells remodeling is critical during human pregnancy. Growth hormone-releasing hormone and its functional receptor have been shown to be expressed in gynecological cancer cells and eutopic endometrial stromal cells. Recent studies have demonstrated the potential clinical uses of antagonists of growth hormone-releasing hormone as effective antitumor agents because of its directly antagonistic effect on the locally produced growth hormone-releasing hormone in gynecological tumors. However, the impact of growth hormone-releasing hormone antagonists on normal endometrial stromal cell growth remained to be elucidated. The aim of this study was to investigate the effect of a growth hormone-releasing hormone antagonist (JMR-132) on cell proliferation and apoptosis of human decidual stromal cells and the underlying molecular mechanisms. Our results showed that growth hormone-releasing hormone and the splice variant 1 of growth hormone-releasing hormone receptor are expressed in human decidual stromal cells isolated from the decidual tissues of early pregnant women receiving surgical abortion. In addition, treatment of stroma cells with JMR-132 induced cell apoptosis with increasing cleaved caspase-3 and caspase-9 activities and decrease cell viability in a time- and dose-dependent manner. Using a dual inhibition approach (pharmacological inhibitors and siRNA-mediated knockdown), we showed that JMR-132-induced activation of apoptotic signals are mediated by the activation of ERK1/2 and JNK signaling pathways and the subsequent upregulation of GADD45alpha. Taken together, JMR-132 suppresses cell survival of decidual stromal cells by inducing apoptosis through the activation of ERK1/2- and JNK-mediated upregulation of GADD45alpha in human endometrial stromal cells. Our findings provide new insights into the potential impact of growth hormone-releasing hormone antagonist on the decidual programming in humans.
Assuntos
Apoptose/efeitos dos fármacos , Decídua/citologia , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Células Estromais/efeitos dos fármacos , Proteínas de Ciclo Celular/genética , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Decídua/efeitos dos fármacos , Implantação do Embrião/efeitos dos fármacos , Feminino , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/fisiologia , Gravidez , Sermorelina/análogos & derivados , Sermorelina/farmacologia , Células Estromais/fisiologia , Regulação para Cima/efeitos dos fármacosRESUMO
Colorectal cancer (CRC) is an aggressive tumor in which new treatment options deliver negative results on cure rates and long-term survival. The anticancer effects of growth hormone-releasing hormone (GHRH) antagonists have been reported in various experimental tumors, but their activity in CRC is unknown. In the present study, we demonstrated that chronic treatment with GHRH antagonist of MIAMI class, MIA-690, promoted survival and gradually blunted tumor progression in experimentally induced colitis-associated cancer in mice, paralleled by reduced inflammation in colon tissue. In particular, MIA-690 improved disease activity index score, and reduced loss of weight and mortality, by improving the survival rates, compared with vehicle-treated group. MIA-690 was also found to reduce various inflammatory and oxidative markers, such as serotonin, prostaglandin (PG)E2 and 8-iso-PGF2α levels, as well as COX-2, iNOS, TNF-α, IL-6 and NF-kB gene expression. Moreover, MIA-690 inhibited the protein expression of c-Myc, P-AKT and Bcl-2 and upregulated p53 protein expression. In conclusion, we showed that MIA-690 suppresses CRC progression and growth by reducing inflammatory and oxidative markers and modulating apoptotic and oncogenic pathways. Further investigations are required for translating these findings into the clinics.
Assuntos
Neoplasias Colorretais , Hormônio Liberador de Hormônio do Crescimento , Animais , Masculino , Camundongos , Apoptose/efeitos dos fármacos , Carcinogênese/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Neoplasias Colorretais/metabolismo , Regulação para Baixo , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Mediadores da Inflamação/metabolismo , Camundongos Endogâmicos C57BL , Estresse Oxidativo/efeitos dos fármacos , Distribuição Aleatória , Regulação para CimaRESUMO
Growth hormone-releasing hormone (GHRH) is a hypothalamic hormone, which regulates the secretion of growth hormone (GH) from the anterior pituitary gland. The effects of GHRH extend beyond the GH-insulin-like growth factor I axis, and that neuropeptide has been involved in the potentiation of several malignancies and other inflammatory disorders. The development of GHRH antagonists (GHRHAnt) delivers an exciting possibility to counteract the pathogenesis of the GHRH-related effects in human pathophysiology, especially when considered that GHRHAnt support endothelial barrier integrity. Those GHRHAnt-mediated effects are exerted at least in part due to the suppression of major inflammatory pathways, and the modulation of major cytoskeletal components. In the present study, we measured the production of reactive oxygen species (ROS) in bovine pulmonary artery endothelial cells, human cerebral microvascular endothelial cells, and human lung microvascular endothelial cells exposed to GHRH or a commercially available GHRHAnt. Our findings reveal the antioxidative effects of GHRHAnt in all three cell lines, which express GHRH receptors. The redox status of NIH/3T3 cells, which do not produce GHRH receptors, was not significantly affected by GHRH or GHRHAnt. Hence, the application of GHRHAnt in pathologies related to increased ROS production should be further investigated.
Assuntos
Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Bovinos , Linhagem Celular Transformada , Hormônio Liberador de Hormônio do Crescimento/metabolismo , Hormônio Liberador de Hormônio do Crescimento/farmacologia , Humanos , Peróxido de Hidrogênio/metabolismo , Camundongos , Células NIH 3T3 , Artéria Pulmonar/citologia , Receptores de Neuropeptídeos/metabolismo , Receptores de Hormônios Reguladores de Hormônio Hipofisário/metabolismoRESUMO
Optic neuropathies are leading causes of irreversible visual impairment and blindness, currently affecting more than 100 million people worldwide. Glaucoma is a group of optic neuropathies attributed to progressive degeneration of retinal ganglion cells (RGCs). We have previously demonstrated an increase in survival of RGCs by the activation of macrophages, whereas the inhibition of macrophages was involved in the alleviation on endotoxin-induced inflammation by antagonist of growth hormone-releasing hormone (GHRH). Herein, we hypothesized that GHRH receptor (GHRH-R) signaling could be involved in the survival of RGCs mediated by inflammation. We found the expression of GHRH-R in RGCs of adult rat retina. After optic nerve crush, subcutaneous application of GHRH agonist MR-409 or antagonist MIA-602 promoted the survival of RGCs. Both the GHRH agonist and antagonist increased the phosphorylation of Akt in the retina, but only agonist MR-409 promoted microglia activation in the retina. The antagonist MIA-602 reduced significantly the expression of inflammation-related genes Il1b, Il6, and Tnf Moreover, agonist MR-409 further enhanced the promotion of RGC survival by lens injury or zymosan-induced macrophage activation, whereas antagonist MIA-602 attenuated the enhancement in RGC survival. Our findings reveal the protective effect of agonistic analogs of GHRH on RGCs in rats after optic nerve injury and its additive effect to macrophage activation, indicating a therapeutic potential of GHRH agonists for the protection of RGCs against optic neuropathies especially in glaucoma.
Assuntos
Hormônio Liberador de Hormônio do Crescimento/agonistas , Macrófagos/patologia , Neuroproteção , Traumatismos do Nervo Óptico/patologia , Células Ganglionares da Retina/patologia , Animais , Sobrevivência Celular/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Hormônio do Crescimento/metabolismo , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Inflamação/genética , Inflamação/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Microglia/efeitos dos fármacos , Microglia/metabolismo , Microglia/patologia , Neuroproteção/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos Endogâmicos F344 , Receptores de Neuropeptídeos/metabolismo , Receptores de Hormônios Reguladores de Hormônio Hipofisário/metabolismo , Células Ganglionares da Retina/efeitos dos fármacos , Células Ganglionares da Retina/metabolismo , Fator de Transcrição STAT3/metabolismo , Sermorelina/análogos & derivados , Sermorelina/farmacologia , Transdução de Sinais/efeitos dos fármacos , Corpo Vítreo/efeitos dos fármacos , Corpo Vítreo/metabolismo , Zimosan/farmacologiaRESUMO
Growth hormone releasing hormone is a hypothalamic neuropeptide, which regulates the release of growth hormone from the anterior pituitary gland. Growth hormone releasing hormone antagonists are anticancer agents, associated with strong anti-inflammatory activities. In the present study, we investigated the effects of the GHRH antagonist MIA-602 in the integrity of the brain microvascular endothelium in vitro. Our observations suggest that MIA-602 protects against the H2O2-induced breakdown of the brain endothelium and enhances its integrity by inducing P53, deactivating cofilin, and suppressing the RhoA inflammatory pathway. Thus, GHRH antagonists may offer an exciting possibility for the treatment of pathologies related to the blood brain barrier dysfunction, including the Parkinson's and Alzheimer's diseases.
Assuntos
Encéfalo/irrigação sanguínea , Endotélio/patologia , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Peróxido de Hidrogênio/toxicidade , Microvasos/patologia , Fármacos Neuroprotetores/farmacologia , Actinas/metabolismo , Endotélio/efeitos dos fármacos , Células HeLa , Humanos , Sermorelina/análogos & derivados , Sermorelina/farmacologia , Proteína Supressora de Tumor p53/metabolismoRESUMO
Growth hormone-releasing hormone is a hypothalamic neuropeptide, which regulates the secretion of growth hormone by the anterior pituitary gland. Recent evidence suggest that it exerts growth factor activities in a diverse variety of in vivo and in vitro experimental malignancies, which are counteracted by growth hormone-releasing hormone antagonists. Those peptides support lung endothelial barrier integrity by suppressing major inflammatory pathways and by inducing the endothelial defender P53. The present effort provides information regarding the effects of growth hormone-releasing hormone in the regulation of P53 and the unfolded protein response. Furthermore, it suggests the possible application of growth hormone-releasing hormone antagonists towards the management of acute lung injury, including the lethal acute respiratory distress syndrome.
Assuntos
Lesão Pulmonar Aguda/metabolismo , Hormônio Liberador de Hormônio do Crescimento/metabolismo , Pulmão/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Lesão Pulmonar Aguda/tratamento farmacológico , Lesão Pulmonar Aguda/patologia , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Antagonistas de Hormônios/uso terapêutico , Humanos , Pulmão/efeitos dos fármacos , Pulmão/patologia , Sermorelina/análogos & derivados , Sermorelina/uso terapêutico , Transdução de Sinais , Resposta a Proteínas não DobradasRESUMO
Growth hormone-releasing hormone (GHRH) antagonist MIA-690 and GHRH agonist MR-409, previously synthesized and developed by us have demonstrated potent antitumor effects. However, little is known about the effects of these analogs on brain functions. We investigated the potential antinflammatory and antioxidant effects of GHRH antagonist MIA-690 and GHRH agonist MR-409, on isolated mouse prefrontal cortex specimens treated with lipopolysaccharide (LPS). Additionally, we studied their effects on emotional behavior after chronic in vivo treatment. Ex vivo, MIA-690 and MR-409 inhibited LPS-induced inflammatory and pro-oxidative markers. In vivo, both MIA-690 and MR-409 induced anxiolytic and antidepressant-like effects, increased norepinephrine and serotonin levels and decreased nuclear factor-kB, tumor necrosis factor-α and interleukin-6 gene expression in prefrontal cortex. Increased nuclear factor erythroid 2-related factor 2 expression was also found in mice treated with MIA-690 and MR-409. MIA-690 showed higher efficacy in inhibiting all tested inflammatory and oxidative markers. In addition, MR-409 induced a down regulation of the gene and protein expression of pituitary-type GHRH-receptor in prefrontal cortex of mice after 4 weeks of treatment at 5 µg/day. In conclusion, our results demonstrate anxiolytic and antidepressant-like effects of GHRH analogs that could involve modulatory effects on monoaminergic signaling, inflammatory and oxidative status.
Assuntos
Anti-Inflamatórios , Antioxidantes , Comportamento Animal/efeitos dos fármacos , Emoções/efeitos dos fármacos , Hormônio Liberador de Hormônio do Crescimento/agonistas , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Sermorelina/análogos & derivados , Animais , Ansiolíticos , Antidepressivos , Expressão Gênica/efeitos dos fármacos , Interleucina-6/genética , Interleucina-6/metabolismo , Camundongos , NF-kappa B/genética , NF-kappa B/metabolismo , Norepinefrina/metabolismo , Córtex Pré-Frontal/metabolismo , Receptores de Neuropeptídeos/genética , Receptores de Neuropeptídeos/metabolismo , Receptores de Hormônios Reguladores de Hormônio Hipofisário/genética , Receptores de Hormônios Reguladores de Hormônio Hipofisário/metabolismo , Sermorelina/farmacologia , Serotonina/metabolismo , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Prostate cancer is the second leading cause of cancer-related deaths among men in developed countries. Neuroendocrine prostate cancer, in particular, is associated with an aggressive phenotype and a poor prognosis. Neuroendocrine cells produce and secrete peptide hormones and growth factors in a paracrine/autocrine manner which promote the progression of the disease. Recent studies have demonstrated that extracellular vesicles or exosomes are released by prostate cancer cells, supporting the spread of prostate cancer. Hence, the aim of this study was to investigate the effect of growth hormone-releasing hormone (GHRH) on neuroendocrine differentiation (NED) in the androgen-dependent prostate cancer cell line LNCaP and the molecular mechanisms underlying these effects. GHRH induced an increase in the percentage of neurite-bearing cells and in the protein levels of Neuron-Specific Enolase. Both effects were blocked by the GHRH receptor antagonist MIA-690. In addition, pretreatment of these cells with the calcium chelator BAPTA, the EGFR inhibitor AG-1478 or the HER2 inhibitor AG-825 reduced the effect of GHRH, suggesting that the GHRH-induced stimulation of NED involves calcium channel activation and EGFR/HER2 transactivation. Finally, PC3-derived exosomes led to an increase in NED, cell proliferation and cell adhesion. Altogether, these findings suggest that GHRH antagonists should be considered for in the management of neuroendocrine prostate cancer.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Células Neuroendócrinas/efeitos dos fármacos , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/metabolismo , Androgênios/farmacologia , Adesão Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Humanos , Masculino , Células Neuroendócrinas/metabolismo , Células PC-3 , Quinazolinas/farmacologia , Receptor ErbB-2/metabolismo , Tirfostinas/farmacologiaRESUMO
Growth Hormone-Releasing Hormone (GHRH) regulates the release of growth hormone from the anterior pituitary gland. GHRH also acts as a growth and inflammatory factor in a variety of experimental models in oncology. In the current study, we used bovine pulmonary arterial cells in order to investigate the effects of GHRH and its antagonistic and agonistic analogs in key intracellular pathways that regulate endothelial permeability. GHRH antagonists suppressed the activation of MLC2, ERK1/2, JAK2/STAT3 pathway and increased the intracellular P53 and pAMPK levels. In contrast, both GHRH and GHRH agonist MR409 exerted the opposite effects. Furthermore, GHRH antagonists supported the integrity of endothelial barrier, while GHRH and GHRH agonists had the contrary effects, as reflected in measurements of transendothelial resistance. Our observations support the evidence for the anti - inflammatory role of GHRH antagonists in the vasculature. Moreover, our results suggest that GHRH antagonists should be considered as promising therapeutic agents for treating severe respiratory abnormalities, such as the lethal Acute Respiratory Distress Syndrome (ARDS).
Assuntos
Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Hormônio do Crescimento/metabolismo , Inflamação/tratamento farmacológico , Pulmão/efeitos dos fármacos , Animais , Bovinos , HumanosRESUMO
Malignant pleural mesothelioma (MPM) is an aggressive malignancy associated with exposure to asbestos, with poor prognosis and no effective therapies. The strong inhibitory activities of growth hormone-releasing hormone (GHRH) antagonists have been demonstrated in different experimental human cancers, including lung cancer; however, their role in MPM remains unknown. We assessed the effects of the GHRH antagonists MIA-602 and MIA-690 in vitro in MPM cell lines and in primary MPM cells, and in vivo in MPM xenografts. GHRH, GHRH receptor, and its main splice variant SV1 were found in all the MPM cell types examined. In vitro, MIA-602 and MIA-690 reduced survival and proliferation in both MPM cell lines and primary cells and showed synergistic inhibitory activity with the chemotherapy drug pemetrexed. In MPM cells, GHRH antagonists also regulated activity and expression of apoptotic molecules, inhibited cell migration, and reduced the expression of matrix metalloproteinases. These effects were accompanied by impairment of mitochondrial activity and increased production of reactive oxygen species. In vivo, s.c. administration of MIA-602 and MIA-690 at the dose of 5 µg/d for 4 wk strongly inhibited the growth of MPM xenografts in mice, along with reduction of tumor insulin-like growth factor-I and vascular endothelial growth factor. Overall, these results suggest that treatment with GHRH antagonists, alone or in association with chemotherapy, may offer an approach for the treatment of MPM.
Assuntos
Antineoplásicos/farmacologia , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Mesotelioma/metabolismo , Mesotelioma/patologia , Neoplasias Pleurais/metabolismo , Neoplasias Pleurais/patologia , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Modelos Animais de Doenças , Expressão Gênica , Hormônio Liberador de Hormônio do Crescimento/genética , Hormônio Liberador de Hormônio do Crescimento/metabolismo , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Mesotelioma/tratamento farmacológico , Mesotelioma Maligno , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Neoplasias Pleurais/tratamento farmacológico , Receptores de Neuropeptídeos/genética , Receptores de Neuropeptídeos/metabolismo , Receptores de Hormônios Reguladores de Hormônio Hipofisário/genética , Receptores de Hormônios Reguladores de Hormônio Hipofisário/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
P53 is a transcription factor very often mutated in malignancies. It functions towards the regulation of important cellular activities, such as cell cycle, senescence and apoptosis. Since inflammation and cancer are strongly associated through common pathways, P53 can suppress inflammation in a plethora of human tissues. Growth Hormone - Releasing Hormone is a hypothalamic peptide with a great capacity to affect the complex networks of cellular regulation via GHRH - specific receptors. GHRH antagonistic and agonistic analogs have been developed for clinical applications, including treatment of benign prostatic hyperplasia, breast, prostate and lung cancers, diabetes and neurodegenerative diseases. The epicenter of the current manuscript is the protective role of P53 against inflammation and cancer and emphasizes the p53 - mediated beneficial effects of GHRH antagonists in various human diseases.
Assuntos
Hormônio Liberador de Hormônio do Crescimento/metabolismo , Neoplasias/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Animais , Hormônio Liberador de Hormônio do Crescimento/agonistas , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Humanos , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Inflamação/patologia , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Proteína Supressora de Tumor p53/agonistas , Proteína Supressora de Tumor p53/antagonistas & inibidoresRESUMO
BACKGROUND: Inflammation plays a key role in the etiology of benign prostatic hyperplasia (BPH) through multiple pathways involving the stimulation of proliferation by cytokines and growth factors as well as the induction of the focal occurrence of epithelial-to-mesenchymal transition (EMT). We have previously reported that GHRH acts as a prostatic growth factor in experimental BPH and in autoimmune prostatitis models and its blockade with GHRH antagonists offer therapeutic approaches for these conditions. Our current study was aimed at the investigation of the beneficial effects of GHRH antagonists in λ-carrageenan-induced chronic prostatitis and at probing the downstream molecular pathways that are implicated in GHRH signaling. METHODS: To demonstrate the complications triggered by recurrent/chronic prostatic inflammation in Sprague-Dawley rats, 50 µL 3% carrageenan was injected into both ventral prostate lobes two times, 3 weeks apart. GHRH antagonist, MIA-690, was administered 5 days after the second intraprostatic injection at 20 µg daily dose for 4 weeks. GHRH-induced signaling events were identified in BPH-1 and in primary prostate epithelial (PrEp) cells at 5, 15, 30, and 60 min with Western blot. RESULTS: Inflammation induced prostatic enlargement and increased the area of the stromal compartment whereas treatment with the GHRH antagonist significantly reduced these effects. This beneficial activity was consistent with a decrease in prostatic GHRH, inflammatory marker COX-2, growth factor IGF-1 and inflammatory and EMT marker TGF-ß1 protein levels and the expression of multiple genes related to EMT. In vitro, GHRH stimulated multiple pathways involved in inflammation and growth in both BPH-1 and PrEp cells including NFκB p65, AKT, ERK1/2, EGFR, STAT3 and increased the levels of TGF-ß1 and Snail/Slug. Most interestingly, GHRH also stimulated the transactivation of the IGF receptor. CONCLUSIONS: The study demonstrates that GHRH antagonists could be beneficial for the treatment of prostatic inflammation and BPH in part by inhibiting the growth-promoting and inflammatory effects of locally produced GHRH.
Assuntos
Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Hiperplasia Prostática/tratamento farmacológico , Prostatite/tratamento farmacológico , Animais , Carragenina , Linhagem Celular , Citocinas/metabolismo , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Masculino , Hiperplasia Prostática/induzido quimicamente , Hiperplasia Prostática/metabolismo , Prostatite/induzido quimicamente , Prostatite/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/fisiologiaRESUMO
We investigated the effects of novel antagonists of growth hormone releasing hormone (GHRH)-MIA602 and MIA690-on three human small cell lung cancer (SCLC) lines (H446, DMS53 and H69) and two non-SCLC (NSCLC) lines (HCC827 and H460). In vitro exposure of cancer cells to these GHRH antagonists significantly inhibited cell viability, increased cell apoptosis, decrease cellular levels of cAMP and reduced cell migration. In vivo, the antagonists strongly inhibited tumor growth in xenografted nude mice models. Subcutaneous administration of MIA602 at the dose of 5 µg/day for 4-8 weeks reduced the growth of HCC827, H460 and H446 tumors by 69.9%, 68.3% and 53.4%, respectively, while MIA690 caused a reduction of 76.8%, 58.3% and 54.9%, respectively. Western blot and qRT-PCR analyses demonstrated a downregulation of expression of the pituitary-type GHRH-R and its splice-variant, cyclinD1/2, cyclin-dependent kinase4/6, p21-activated kinase-1, phosphorylation of activator of transcription 3 and cAMP response element binding protein; and an upregulation of expression of E-cadherin, ß-catenin and P27kip1 in cancer cells and in xenografted tumor tissues. The study demonstrates the involvement of GHRH antagonists in multiple signaling pathways in lung cancers. Our findings suggest the merit of further investigation with these GHRH antagonists on the management of both SCLC and NSCLC.
Assuntos
Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Sermorelina/análogos & derivados , Carcinoma de Pequenas Células do Pulmão/metabolismo , Animais , Apoptose/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , AMP Cíclico/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Modelos Animais de Doenças , Feminino , Expressão Gênica , Hormônio Liberador de Hormônio do Crescimento/genética , Hormônio Liberador de Hormônio do Crescimento/metabolismo , Humanos , Camundongos , Camundongos Nus , Fator de Transcrição STAT3/metabolismo , Sermorelina/farmacologia , Transdução de Sinais/efeitos dos fármacos , Carcinoma de Pequenas Células do Pulmão/tratamento farmacológico , Carcinoma de Pequenas Células do Pulmão/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The potential therapeutic effects of agonistic analogs of growth hormone-releasing hormone (GHRH) and their mechanism of action were investigated in diabetic retinopathy (DR). Streptozotocin-induced diabetic rats (STZ-rats) were treated with 15 µg/kg GHRH agonist, MR-409, or GHRH antagonist, MIA-602. At the end of treatment, morphological and biochemical analyses assessed the effects of these compounds on retinal neurovascular injury induced by hyperglycemia. The expression levels of GHRH and its receptor (GHRH-R) measured by qPCR and Western blotting were significantly down-regulated in retinas of STZ-rats and in human diabetic retinas (postmortem) compared with their respective controls. Treatment of STZ-rats with the GHRH agonist, MR-409, prevented retinal morphological alteration induced by hyperglycemia, particularly preserving survival of retinal ganglion cells. The reverse, using the GHRH antagonist, MIA-602, resulted in worsening of retinal morphology and a significant alteration of the outer retinal layer. Explaining these results, we have found that MR-409 exerted antioxidant and anti-inflammatory effects in retinas of the treated rats, as shown by up-regulation of NRF-2-dependent gene expression and down-regulation of proinflammatory cytokines and adhesion molecules. MR-409 also significantly down-regulated the expression of vascular endothelial growth factor while increasing that of pigment epithelium-derived factor in diabetic retinas. These effects correlated with decreased vascular permeability. In summary, our findings suggest a neurovascular protective effect of GHRH analogs during the early stage of diabetic retinopathy through their antioxidant and anti-inflammatory properties.
Assuntos
Anti-Inflamatórios/farmacologia , Retinopatia Diabética/tratamento farmacológico , Hormônio Liberador de Hormônio do Crescimento/agonistas , Sermorelina/análogos & derivados , Animais , Anti-Inflamatórios/uso terapêutico , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Citocinas/genética , Citocinas/metabolismo , Retinopatia Diabética/metabolismo , Fator de Transcrição de Proteínas de Ligação GA/genética , Fator de Transcrição de Proteínas de Ligação GA/metabolismo , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Humanos , Masculino , Ratos , Ratos Sprague-Dawley , Retina/efeitos dos fármacos , Retina/metabolismo , Sermorelina/farmacologia , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Growth hormone-releasing hormone (GHRH) is a peptide hormone secreted by the hypothalamus that regulates the synthesis and secretion of growth hormone (GH) in the pituitary. The extra-hypothalamic GHRH and its cognate receptors (GHRHR and splice variants) play a mitogenic role by stimulating cell proliferation and preventing apoptotic cell death. It is well established that GHRH antagonists inhibit the growth, tumorigenicity, and metastasis of various human malignancies. In this work, we studied the effect of two new GHRH antagonists, MIA602 and MIA690, on thyroid cancer. We studied the effect of MIA602 and MIA690 on thyroid cancer in vitro, using human thyroid cancer cell lines, and in vivo, using chicken embryo chorioallantoic membrane (CAM) assays. We found that mRNA for GHRH and GHRH receptor is expressed in thyroid cell lines and in samples of thyroid tumors. Immunohistochemistry confirmed the expression of GHRHR protein in specimens of thyroid tumor. We observed that GHRH antagonists inhibited the growth and increased apoptosis of thyroid cancer cells. In vivo, the antagonists inhibited growth and angiogenesis of engrafted thyroid tumors. Our results suggest that GHRH expression may play a role in growth of thyroid cancer and that GHRH antagonists can be a therapeutic option for thyroid cancer patients.
Assuntos
Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Neoplasias da Glândula Tireoide/metabolismo , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Embrião de Galinha , Expressão Gênica , Hormônio Liberador de Hormônio do Crescimento/genética , Hormônio Liberador de Hormônio do Crescimento/metabolismo , Humanos , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , RNA Mensageiro/genética , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/patologiaRESUMO
The etiology of benign prostatic hyperplasia (BPH) is multifactorial, and chronic inflammation plays a pivotal role in its pathogenesis. Growth hormone-releasing hormone (GHRH) is a hypothalamic neuropeptide that has been shown to act as paracrine/autocrine factor in various malignancies including prostate cancer. GHRH and its receptors are expressed in experimental models of BPH, in which antagonists of GHRH suppressed the levels of proinflammatory cytokines and altered the expression of genes related to epithelial-to-mesenchymal transition (EMT). We investigated the effects of GHRH antagonist on prostatic enlargement induced by inflammation. Autoimmune prostatitis in Balb/C mice was induced by a homogenate of reproductive tissues of male rats. During the 8-wk induction of chronic prostatitis, we detected a progressive increase in prostatic volume reaching 92% at week 8 compared with control (P < 0.001). Daily treatment for 1 mo with GHRH antagonist MIA-690 caused a 30% reduction in prostate volume (P < 0.05). Conditioned medium derived from macrophages increased the average volume of spheres by 82.7% (P < 0.001) and elevated the expression of mRNA for N-cadherin, Snail, and GHRH GHRH antagonist reduced the average volume of spheres stimulated by inflammation by 75.5% (P < 0.05), and TGF-ß2 by 91.8% (P < 0.01). The proliferation of primary epithelial cells stimulated by IL-17A or TGF-ß2 was also inhibited by 124.1% and 69.9%, respectively. GHRH stimulated the growth of BPH-1 and primary prostate spheres. This study provides evidence that GHRH plays important roles in prostatic inflammation and EMT and suggests the merit of further investigation to elucidate the effects of GHRH antagonists in prostatitis and BPH.
Assuntos
Proliferação de Células/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Hormônio Liberador de Hormônio do Crescimento/análogos & derivados , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Animais , Doenças Autoimunes/genética , Doenças Autoimunes/metabolismo , Doenças Autoimunes/patologia , Proliferação de Células/genética , Células Cultivadas , Células Epiteliais/metabolismo , Expressão Gênica/efeitos dos fármacos , Hormônio Liberador de Hormônio do Crescimento/genética , Hormônio Liberador de Hormônio do Crescimento/metabolismo , Hormônio Liberador de Hormônio do Crescimento/farmacologia , Humanos , Masculino , Camundongos Endogâmicos BALB C , Próstata/efeitos dos fármacos , Próstata/metabolismo , Próstata/patologia , Hiperplasia Prostática/genética , Hiperplasia Prostática/metabolismo , Hiperplasia Prostática/patologia , Prostatite/genética , Prostatite/metabolismo , Prostatite/patologia , Ratos , Fator de Crescimento Transformador beta2/farmacologiaRESUMO
The syntheses and biological evaluations of new GHRH analogs of Miami (MIA) series with greatly increased anticancer activity are described. In the design and synthesis of these analogs, the following previous substitutions were conserved: D-Arg2, Har9, Abu15, and Nle27. Most new analogs had Ala at position 8. Since replacements of both Lys12 and Lys21 with Orn increased resistance against enzymatic degradation, these modifications were kept. The substitutions of Arg at both positions 11 and 20 by His were also conserved. We kept D-Arg28, Har29 -NH2 at the C-terminus or inserted Agm or 12-amino dodecanoic acid amide at position 30. We incorporated pentafluoro-Phe (Fpa5), instead of Cpa, at position 6 and Tyr(Me) at position 10 and ω-amino acids at N-terminus of some analogs. These GHRH analogs were prepared by solid-phase methodology and purified by HPLC. The evaluation of the activity of the analogs on GH release was carried out in vitro on rat pituitaries and in vivo in male rats. Receptor binding affinities were measured in vitro by the competitive binding analysis. The inhibitory activity of the analogs on tumor proliferation in vitro was tested in several human cancer cell lines such as HEC-1A endometrial adenocarcinoma, HCT-15 colorectal adenocarcinoma, and LNCaP prostatic carcinoma. For in vivo tests, various cell lines including PC-3 prostate cancer, HEC-1A endometrial adenocarcinoma, HT diffuse mixed ß cell lymphoma, and ACHN renal cell carcinoma cell lines were xenografted into nude mice and treated subcutaneously with GHRH antagonists at doses of 1-5µg/day. Analogs MIA-602, MIA-604, MIA-610, and MIA-640 showed the highest binding affinities, 30, 58, 48, and 73 times higher respectively, than GHRH (1-29) NH2. Treatment of LNCaP and HCT-15 cells with 5µM MIA-602 or MIA-690 decreased proliferation by 40%-80%. In accord with previous tests in various human cancer lines, analog MIA-602 showed high inhibitory activity in vivo on growth of PC-3 prostate cancer, HT-mixed ß cell lymphoma, HEC-1A endometrial adenocarcinoma and ACHN renal cell carcinoma. Thus, GHRH analogs of the Miami series powerfully suppress tumor growth, but have only a weak endocrine GH inhibitory activity. The suppression of tumor growth could be induced in part by the downregulation of GHRH receptors levels.
Assuntos
Proliferação de Células/efeitos dos fármacos , Hormônio Liberador de Hormônio do Crescimento/administração & dosagem , Hormônio do Crescimento/biossíntese , Neoplasias/tratamento farmacológico , Animais , Linhagem Celular Tumoral , Hormônio do Crescimento/metabolismo , Hormônio Liberador de Hormônio do Crescimento/análogos & derivados , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Hormônio Liberador de Hormônio do Crescimento/síntese química , Humanos , Camundongos , Neoplasias/patologia , Ratos , Relação Estrutura-AtividadeRESUMO
More than 25% of patients diagnosed with endometrial carcinoma have invasive primary cancer accompanied by metastases. Growth hormone-releasing hormone (GHRH) plays an important role in reproduction. Here, we examined the effect of a GHRH antagonist on the motility of endometrial cancer cells and the mechanisms of action of the antagonist in endometrial cancer. Western blotting and immunohistochemistry (IHC) were used to determine the expression of the GHRH receptor protein. The activity of Twist and N-cadherin was determined by Western blotting. Cell motility was assessed by an invasion and migration assay. GHRH receptor siRNA was applied to knockdown the GHRH receptor in endometrial cancer cells. The GHRH antagonist inhibited cell motility in a dose-dependent manner. The GHRH antagonist inhibited cell motility and suppressed the expression of Twist and N-cadherin, and the suppression was abolished by GHRH receptor siRNA pretreatment. Moreover, the inhibition of Twist and N-cadherin with Twist siRNA and N-cadherin siRNA, respectively, suppressed cell motility. Our study indicates that the GHRH antagonist inhibited the cell motility of endometrial cancer cells through the GHRH receptor via the suppression of Twist and N-cadherin. Our findings represent a new concept in the mechanism of GHRH antagonist-suppressed cell motility in endometrial cancer cells and suggest the possibility of exploring GHRH antagonists as potential therapeutics for the treatment of human endometrial cancer.