RESUMEN
Bombesin mediates several biological activities in the gastrointestinal (GI) tract and central nervous system in mammals, including smooth muscle contraction, secretion of GI hormones and regulation of homeostatic mechanisms. Here, we report a novel bombesin-like peptide isolated from Boana raniceps. Its amino acid sequence, GGNQWAIGHFM-NH2, was identified and structurally confirmed by HPLC, MS/MS and 454-pyrosequencing; the peptide was named BR-bombesin. The effect of BR-bombesin on smooth muscle contraction was assessed in ileum and esophagus, and its anti-secretory activity was investigated in the stomach. BR-bombesin exerted significant contractile activity with a concentration-response curve similar to that of commercially available bombesin in ileum strips of Wistar rats. In esophageal strips, BR-bombesin acted as an agonist, as many other bombesin-related peptides act, although with different behavior compared to the muscarinic agonist carbachol. Moreover, BR-bombesin inhibited stomach secretion by approximately 50% compared to the untreated control group. This novel peptide has 80% and 70% similarity with the 10-residue C-terminal domain of human neuromedin B (NMB) and human gastrin releasing peptide (GRP10), respectively. Molecular docking analysis revealed that the GRP receptor had a binding energy equal to - 7.3 kcal.mol-1 and - 8.5 kcal.mol-1 when interacting with bombesin and BR-bombesin, respectively. Taken together, our data open an avenue to investigate BR-bombesin in disorders that involve gastrointestinal tract motility and acid gastric secretion.
Asunto(s)
Bombesina , Receptores de Bombesina , Animales , Anuros/metabolismo , Bombesina/metabolismo , Bombesina/farmacología , Mamíferos/metabolismo , Simulación del Acoplamiento Molecular , Péptidos/farmacología , Ratas , Ratas Wistar , Receptores de Bombesina/genética , Receptores de Bombesina/metabolismo , Estómago , Espectrometría de Masas en TándemRESUMEN
OBJECTIVES: Cancer has been investigated using various pre-targeting techniques or models focusing on radiobombesin analogues; however, both are not offered together. In this study, nano-bombesin labeling by a pre-targeting system was undertaken to develop an alternative approach for prostate tumor treatment. METHODS: A two-step pre-targeting system utilizing a combination of streptavidin (SA), biotinylated morpholino (B-MORF), biotinylated BBN (B-BBN) with two different spacers (b-Ala and PEG), and a radiolabeled cMORF was evaluated in vitro and in vivo. RESULTS: Final conjugation conditions consisted of a 1:1:2 ratio of SA:B-MORF:B-BBN, followed by addition of 99mTc-cMORF to compensate for free MORF. In vitro binding experiments with prostate cancer cells (PC-3) revealed that total binding was time-dependent for the Ala spacer but not for the PEG spacer. The highest accumulation (5.06 ± 1.98 %) was achieved with 1 hour of incubation, decreasing as time progressed. Specific binding fell to 1.05 ± 0.35 %. The pre-targeting biodistribution in healthy Swiss mice was measured at different time points, with the best responses observed for 7-h and 15-h incubations. The effector, 99mTc-MAG3-cMORF, was administered 2 h later. Strong kidney excretion was always documented. The greatest tumor uptake was 2.58 ± 0.59 %ID/g at 7 h for B-bAla-BBN, with a region of interest (ROI) value of 3.9 % during imaging. The tumor/blood ratio was low due to the slow blood clearance; however, the tumor/muscle ratio was 5.95. CONCLUSIONS: The pre-targeting approach with a peptide was a viable concept. Further evaluation with modified sequences of MORF, including less cytosine, and additional test intervals could be worthwhile.
Asunto(s)
Bombesina/metabolismo , Imagen Molecular/métodos , Morfolinas/farmacocinética , Nanopartículas , Neoplasias de la Próstata/metabolismo , Radioisótopos , Estreptavidina/farmacocinética , Animales , Bombesina/análogos & derivados , Línea Celular Tumoral , Cromatografía Líquida de Alta Presión/métodos , Modelos Animales de Enfermedad , Masculino , Ratones , Ratones Desnudos , Compuestos de Organotecnecio , Neoplasias de la Próstata/diagnóstico por imagen , Radioisótopos/química , Cintigrafía , Distribución Aleatoria , Factores de TiempoRESUMEN
OBJECTIVES: Cancer has been investigated using various pre-targeting techniques or models focusing on radiobombesin analogues; however, both are not offered together. In this study, nano-bombesin labeling by a pre-targeting system was undertaken to develop an alternative approach for prostate tumor treatment. METHODS: A two-step pre-targeting system utilizing a combination of streptavidin (SA), biotinylated morpholino (B-MORF), biotinylated BBN (B-BBN) with two different spacers (b-Ala and PEG), and a radiolabeled cMORF was evaluated in vitro and in vivo. RESULTS: Final conjugation conditions consisted of a 1:1:2 ratio of SA:B-MORF:B-BBN, followed by addition of 99mTc-cMORF to compensate for free MORF. In vitro binding experiments with prostate cancer cells (PC-3) revealed that total binding was time-dependent for the Ala spacer but not for the PEG spacer. The highest accumulation (5.06 ± 1.98 percent) was achieved with 1 hour of incubation, decreasing as time progressed. Specific binding fell to 1.05 ± 0.35 percent. The pre-targeting biodistribution in healthy Swiss mice was measured at different time points, with the best responses observed for 7-h and 15-h incubations. The effector, 99mTc-MAG3-cMORF, was administered 2 h later. Strong kidney excretion was always documented. The greatest tumor uptake was 2.58 ± 0.59 percentID/g at 7 h for B-bAla-BBN, with a region of interest (ROI) value of 3.9 percent during imaging. The tumor/blood ratio was low due to the slow blood clearance; however, the tumor/muscle ratio was 5.95. CONCLUSIONS: The pre-targeting approach with a peptide was a viable concept. Further evaluation with modified sequences of MORF, including less cytosine, and additional test intervals could be worthwhile.
Asunto(s)
Animales , Masculino , Ratones , Bombesina/metabolismo , Imagen Molecular/métodos , Morfolinas/farmacocinética , Nanopartículas , Neoplasias de la Próstata/metabolismo , Radioisótopos , Estreptavidina/farmacocinética , Bombesina/análogos & derivados , Bombesina , Línea Celular Tumoral , Cromatografía Líquida de Alta Presión/métodos , Modelos Animales de Enfermedad , Ratones Desnudos , Compuestos de Organotecnecio , Neoplasias de la Próstata , Distribución Aleatoria , Radioisótopos/química , Factores de TiempoRESUMEN
Medulloblastoma is the most common brain tumor of childhood. Emerging molecular targets in medulloblastoma include neurotrophin and neuropeptide receptors. In the present study, we have examined the influence of brain-derived neurotrophic factor (BDNF)/TrkB receptor- and gastrin-releasing peptide receptor (GRPR)-mediated signaling on the viability of human medulloblastoma cells. The expression of TrkB and GRPR was confirmed by immunohistochemistry and mRNA for both BDNF and GRPR was detected by reverse transcriptase polymerase chain reaction in Daoy, D283, and ONS76 cells. Treatment with BDNF significantly inhibited the viability of Daoy and D283, but not ONS76 cells, measured with the MTT assay. Neither the GRPR agonists GRP and bombesin nor the GRPR antagonist RC-3095 affected cell viability. Because previous findings have indicated that the viability of glioma cells might be enhanced by GRP when combined with the cAMP phosphodiesterase-4 (PDE4) inhibitor rolipram, we also examined the effects of rolipram alone or combined with GRP on cell viability. Rolipram significantly reduced the viability of all three cell lines, and the inhibitory effect of rolipram in Daoy cells was not modified by cotreatment with GRP. The results suggest that BDNF/TrkB and PDE4, but not the GRPR, regulate the viability of medulloblastoma cells.
Asunto(s)
Factor Neurotrófico Derivado del Encéfalo/metabolismo , Supervivencia Celular/fisiología , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/metabolismo , Meduloblastoma/metabolismo , Receptores de Bombesina/metabolismo , Animales , Antineoplásicos/metabolismo , Bombesina/análogos & derivados , Bombesina/genética , Bombesina/metabolismo , Factor Neurotrófico Derivado del Encéfalo/genética , Niño , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/genética , Péptido Liberador de Gastrina/genética , Péptido Liberador de Gastrina/metabolismo , Fármacos Gastrointestinales/metabolismo , Humanos , Neurotransmisores/metabolismo , Fragmentos de Péptidos/metabolismo , Inhibidores de Fosfodiesterasa/metabolismo , Receptor trkB/metabolismo , Receptores de Bombesina/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Rolipram/metabolismoRESUMEN
Increasing evidence indicates that the gastrin-releasing peptide receptor (GRPR) is implicated in regulating synaptic plasticity and memory formation in the hippocampus and other brain areas. However, the molecular mechanisms underlying the memory-impairing effects of GRPR antagonism have remained unclear. Here we report that basic fibroblast growth factor (bFGF/FGF-2) rescues the memory impairment induced by GRPR antagonism in the rat dorsal hippocampus. The GRPR antagonist [D-Tpi(6), Leu(13) psi(CH(2)NH)-Leu(14)] bombesin (6-14) (RC-3095) at 1.0 microg impaired, whereas bFGF at 0.25 microg enhanced, 24 h retention of inhibitory avoidance (IA) when infused immediately after training into the CA1 hippocampal area in male rats. Coinfusion with an otherwise ineffective dose of bFGF blocked the memory-impairing effect of RC-3095. These findings suggest that the memory-impairing effects of GRPR antagonists might be partially mediated by an inhibition in the function and/or expression of neuronal bFGF or diminished activation of intracellular protein kinase pathways associated with bFGF signaling.
Asunto(s)
Factor 2 de Crecimiento de Fibroblastos/metabolismo , Hipocampo/metabolismo , Trastornos de la Memoria/metabolismo , Fármacos Neuroprotectores/metabolismo , Receptores de Bombesina/antagonistas & inhibidores , Animales , Conducta Animal/fisiología , Bombesina/análogos & derivados , Bombesina/metabolismo , Hipocampo/citología , Masculino , Memoria/fisiología , Fragmentos de Péptidos/metabolismo , Ratas , Ratas Wistar , Receptores de Bombesina/metabolismoRESUMEN
Sepsis is a complex, multifactorial syndrome that can develop into conditions of different severity, described as septic shock or severe sepsis. In spite of the great progress in understanding the mechanisms involved in the pathogenesis and management of sepsis, only a few therapeutic strategies were able to show a decrease in the mortality from septic shock. Although sepsis consists on a systemic inflammatory response, the anti-inflammatory therapies have failed to improve the outcome of critically ill patients. Here the role of gastrin-releasing peptide in immune processes is reviewed and the data that have prompted the recent patent for GRP receptor antagonists RC-3095 as a therapeutic agent on inflammatory conditions are described.
Asunto(s)
Receptores de Bombesina/antagonistas & inhibidores , Sepsis/tratamiento farmacológico , Animales , Bombesina/metabolismo , Péptido Liberador de Gastrina/fisiología , Humanos , Sistema Inmunológico/efectos de los fármacos , Sistema Inmunológico/fisiología , Enfermedades Pulmonares/tratamiento farmacológico , Enfermedades Pulmonares/etiología , Patentes como Asunto , Sepsis/complicacionesRESUMEN
The mammalian bombesin (BB)-like peptide gastrin-releasing peptide (GRP) stimulates cell proliferation, displays a range of neuroendocrine activities, and acts as a growth factor in the pathogenesis of several types of human cancer. Several lines of evidence have indicated that GRP and its receptor (GRPR) might also be involved in the neurochemical alterations associated with psychiatric and neurological disorders. GRP and GRPR are distributed throughout the mammalian central nervous system (CNS). Altered levels of BB-like peptides have been found in the CNS of patients with schizophrenia and Parkinson's disease. Dysfunctions in GRPR-induced cellular calcium signaling have been reported in fibroblasts from patients with Alzheimer's disease. A translocation in the GRPR gene has been associated with autism. Pharmacological and genetic studies in rodents have shown that GRPRs in brain areas such as the dorsal hippocampus and amygdala are importantly involved in regulating synaptic plasticity and aspects of behavior that might be altered in disorders such as anxiety, schizophrenia, depression, autism and dementia. Behaviors modulated by the GRPR in rodents include grooming, food intake, stereotypy, social behavior, and emotionally-motivated learning and memory. Together, these findings support the view that the GRPR should be considered a therapeutic target for a subset of CNS diseases.
Asunto(s)
Encefalopatías/metabolismo , Encéfalo/metabolismo , Péptido Liberador de Gastrina/metabolismo , Trastornos Mentales/metabolismo , Receptores de Bombesina/metabolismo , Animales , Conducta Animal/fisiología , Bombesina/metabolismo , Encéfalo/efectos de los fármacos , Encéfalo/fisiopatología , Química Encefálica/genética , Encefalopatías/tratamiento farmacológico , Encefalopatías/fisiopatología , Predisposición Genética a la Enfermedad/genética , Humanos , Trastornos Mentales/tratamiento farmacológico , Trastornos Mentales/fisiopatología , Plasticidad Neuronal/genética , Receptores de Bombesina/efectos de los fármacos , Receptores de Bombesina/genéticaRESUMEN
Accumulating evidence shows the involvement of neuropeptides in cardiovascular control in mammals as well as non-mammalian species. Our own immunohistochemical studies indicate a sparse innervation only in cyclostomes, holostean fish and lungfish, a more extensive variation and distribution in elasmobranchs and teleosts, and a rich and varied innervation of the cardiovascular system in crocodiles and lizards. Vasoactive intestinal polypeptide (VIP), neuropeptide Y (NPY), gastrin releasing peptide (GRP) and tachykinins are present in most vertebrate groups. VIP is vasodilatory in the Atlantic cod (Gadus morhua) as in most mammalian species, but increases gut vascular resistance in the spiny dogfish (Squalus acanthias). NPY potentiates the effect of noradrenaline on skate (Raja rhina) coronary vessels, suggesting an interaction between adrenergic mechanisms and NPY early in evolution, but studies in the spiny dogfish and the crocodile also demonstrate different mechanisms for the action of NPY and adrenaline in some species. Bombesin/GRP increases flow to the gut in the spiny dogfish by an increase in somatic vascular resistance, while visceral resistance remains unchanged. In the caiman (Caiman crocodylus crocodylus) bombesin causes a shunting of blood from the lung to the gut. Substance P and other tachykinins in general increase flow to the gut, and on some occasions also increase somatic blood flow. Flow in the anastomosis of the crocodile (Crocodylus porosus) gut is increased by substance P. The results presented here are a review of several published and unpublished studies.
Asunto(s)
Animales , Sistema Cardiovascular/fisiología , Neuropéptidos/fisiología , Bombesina/metabolismo , Bombesina/fisiología , Sistema Cardiovascular/metabolismo , Peces/fisiología , Caimanes y Cocodrilos/fisiología , Neuropéptido Y/metabolismo , Neuropéptido Y/fisiología , Neuropéptidos/metabolismo , Sustancia P/fisiología , Sustancia P/metabolismo , Taquicininas/metabolismo , Taquicininas/fisiología , Péptido Intestinal Vasoactivo/metabolismo , Péptido Intestinal Vasoactivo/fisiologíaRESUMEN
Accumulating evidence shows the involvement of neuropeptides in cardiovascular control in mammals as well as non-mammalian species. Our own immunohistochemical studies indicate a sparse innervation only in cyclostomes, holostean fish and lungfish, a more extensive variation and distribution in elasmobranchs and teleosts, and a rich and varied innervation of the cardiovascular system in crocodiles and lizards. Vasoactive intestinal polypeptide (VIP), neuropeptie Y (NPY), gastrin releasing peptide (GRP) and tachykinins are present in most vertebrate groups. VIP is vasodilatory in the Atlantic cod (Gadus morhua) as in most mammalian species, but increases gut vascular resistance in the spiny dogfish (Squalus acanthias). NPY potentiates the effect of noradrenaline on skate (Raja rhina) coronary vessels, suggesting an interaction between adrenergic mechanisms and NPY early in evolution, but studies in the spiny dogfish and the crocodile also demonstrate different mechanisms for the action of NPY and adrenaline in some species. Bombesin/GRP increases flow to the gut in the spiny dogfish by an increase in somatic vascular resistance, while visceral resistance remains unchanged. In the caiman (Caiman crocodylus crocodylus) bombesin causes a shunting of blood from the lung to the gut. Substance P and other tachykinins in general increase flow to the gut, and on some occasions also increase somatic blood flow. Flow in the anastomosis of the crocodile (Crocodylus porosus) gut is increased by substance P. The results presented here are a review of several published and unpublished studies.
Asunto(s)
Fenómenos Fisiológicos Cardiovasculares , Neuropéptidos/fisiología , Caimanes y Cocodrilos/fisiología , Animales , Bombesina/metabolismo , Bombesina/fisiología , Sistema Cardiovascular/metabolismo , Peces/fisiología , Neuropéptido Y/metabolismo , Neuropéptido Y/fisiología , Neuropéptidos/metabolismo , Sustancia P/metabolismo , Sustancia P/fisiología , Taquicininas/metabolismo , Taquicininas/fisiología , Péptido Intestinal Vasoactivo/metabolismo , Péptido Intestinal Vasoactivo/fisiologíaRESUMEN
A 31-year-old patient with a clinical picture of obstructive jaundice had surgical treatment, and a primary carcinoid of the ampulla of Vater (VA) was found. The tumor was studied with light microscopy, immunohistochemistry, and electron microscopy. The neoplasm had histopathologic and cytopathologic features similar to those encountered in typical neuroendocrine neoplasms. It is interesting that immunohistochemical techniques disclosed the presence of vasointestinal polypeptide, cholecystokinin, and bombesin; however, unlike most neuroendocrine neoplasms arising in VA, no somatostatin-immunoreactive cells were found.