RESUMEN
The gastrin-releasing peptide receptor (GRPR) is overexpressed in many solid malignancies, particularly in prostate and breast cancers, among others. We synthesized ProBOMB2, a novel bombesin derivative radiolabeled with 68Ga and 177Lu, and evaluated its ability to target GRPR in a preclinical model of human prostate cancer. Methods: ProBOMB2 was synthesized in solid phase using fluorenylmethoxycarbonyl chemistry. The chelator 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid was coupled to the N terminus and separated from the GRPR-targeting sequence by a cationic 4-amino-(1-carboxymethyl)-piperidine spacer. Binding affinity for both human and murine GRPR was determined using a cell-based competition assay, whereas a calcium efflux assay was used to measure the agonist and antagonist properties of the derivatives. ProBOMB2 was radiolabeled with 177Lu and 68Ga. SPECT and PET imaging and biodistribution studies were conducted using male immunocompromised mice bearing GRPR-positive PC-3 human prostate cancer xenografts. Results: Ga-ProBOMB2 and Lu-ProBOMB2 bound to PC-3 cells with an inhibition constant of 4.58 ± 0.67 and 7.29 ± 1.73 nM, respectively. 68Ga-ProBOMB2 and 177Lu-ProBOMB2 were radiolabeled with a radiochemical purity greater than 95%. Both radiotracers were excreted primarily via the renal pathway. PET images of PC-3 tumor xenografts were visualized with excellent contrast at 1 and 2 h after injection with 68Ga-ProBOMB2, and there was very low off-target organ accumulation. 177Lu-ProBOMB2 enabled clear visualization of PC-3 tumor xenografts by SPECT imaging at 1, 4, and 24 h after injection 177Lu-ProBOMB2 displayed higher tumor uptake than 68Ga-ProBOMB2 at 1 h after injection. 177Lu-ProBOMB2 tumor uptake at 1, 4, and 24 h after injection was 14.9 ± 3.1, 4.8 ± 2.1, and 1.7 ± 0.3 percentage injected dose per gram of tissue, respectively. Conclusion: 68Ga-ProBOMB2 and 177Lu-ProBOMB2 are promising radiotracers with limited pancreas uptake, good tumor uptake, and favorable pharmacokinetics for imaging and therapy of GRPR-expressing tumors.
Asunto(s)
Neoplasias de la Próstata , Receptores de Bombesina , Animales , Bombesina/farmacocinética , Línea Celular Tumoral , Radioisótopos de Galio/química , Humanos , Masculino , Ratones , Imagen Molecular , Neoplasias de la Próstata/metabolismo , Receptores de Bombesina/metabolismo , Distribución TisularRESUMEN
Isolated beef heart mitochondria have been exposed to tert-butyl hydroperoxide (tBHP) and peroxynitrite (PeN) in order to model the effects of reactive oxygen and nitrogen species on mitochondria in vivo. The formation of malondialdehyde (MDA), protein carbonyls, lipofuscin-like pigments (LFP), and nitrotyrosine was studied during incubations with various concentrations of oxidants for up to 24 h. The oxidants differed in their ability to oxidize particular substrates. Fatty acids were more sensitive to the low concentrations of tBHP, whereas higher concentrations of PeN consumed MDA. Oxidation of proteins producing carbonyls had different kinetics and also a probable mechanism with tBHP or PeN. Diverse proteins were affected by tBHP or PeN. In both cases, prolonged incubation led to the appearance of proteins with molecular weights lower than 29 kDa bearing carbonyl groups that might have been caused by protein fragmentation. PeN induced nitration of protein tyrosines that was more intensive in the soluble proteins than in the insoluble ones. LFP, the end products of lipid peroxidation, were formed more readily by PeN. On the other hand, fluorometric and chromatographic techniques have confirmed destruction of LFP by higher PeN concentrations. This is a unique feature that has not been described so far for any oxidant.
Asunto(s)
Mitocondrias Cardíacas/metabolismo , Especies de Nitrógeno Reactivo/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Animales , Bovinos , Ácido Peroxinitroso , terc-ButilhidroperóxidoRESUMEN
Postnatal heart development is characterized by critical periods of heart remodeling. In order to characterize the changes in the lipophilic fraction induced by free radicals, fatty acids and their oxidized products, lipofuscin-like pigments (LFP), were investigated. Fatty acids were analyzed by gas chromatography and LFP were studied by fluorescence techniques. A fluorophore characterized by spectral methods was further resolved by HPLC. Major changes in the composition of fatty acids occurred immediately after birth and then during maturation. Fluorescence of LFP changed markedly on postnatal days 1, 4, 8, and 14, and differed from the adult animals. LFP comprise several fluorophores that were present since fetal state till adulthood. No new major fluorophores were formed during development, just the abundances of individual fluorophores have been modulated which produced changes in the shape of the spectral arrays. HPLC resolved the fluorophore with excitation maximum at 360 nm and emission maximum at 410 nm. New chromatographically distinct species appeared immediately on postnatal day 1, and then on days 30 and 60. Consumption of polyunsaturated fatty acids immediately after birth and subsequent formation of LFP suggests that oxidative stress is involved in normal heart development.
Asunto(s)
Ácidos Grasos/metabolismo , Corazón/crecimiento & desarrollo , Lipofuscina/metabolismo , Animales , Animales Recién Nacidos , Femenino , Peroxidación de Lípido/fisiología , Masculino , Estrés Oxidativo/fisiología , Pigmentos Biológicos/metabolismo , Embarazo , Ratas , Ratas WistarRESUMEN
Oxidative stress, which is present in Alzheimer's disease (AD), results in the formation of various end-products of free radical reactions with proteins and lipids. At present there are no reliable diagnostic biomarkers of AD in the blood. Therefore, specific products of lipid peroxidation in the blood of AD patients were investigated. Lipophilic extracts of erythrocytes in the group of patients with AD (n = 44) and age-matched controls (n = 16) were studied. The end-products of lipid peroxidation, so called lipofuscin-like pigments (LFP), were analysed by fluorescence spectroscopy. It was found that the level of these products is significantly increased in erythrocytes of AD patients compared to controls. LFP were further separated by means of HPLC into individual fractions to study their composition in AD and controls. The specific fraction of LFP in AD patients, which was isolated, might represent a disease-specific product in the blood.