RESUMEN
Organic anion-transporting polypeptide (OATP) 1B1/3-mediated drug-drug interaction (DDI) potential is evaluated in vivo with rosuvastatin (RST) as a probe substrate in clinical studies. We calibrated our assay with RST and estradiol 17-ß-D-glucuronide (E217ßG)/cholecystokinin-8 (CCK8) as in vitro probes for qualitative and quantitative prediction of OATP1B-mediated DDI potential for RST. In vitro OATP1B1/1B3 inhibition using E217ßG and CCK8 yielded higher area under the curve (AUC) ratio (AUCR) values numerically with the static model, but all probes performed similarly from a qualitative cutoff-based prediction, as described in regulatory guidances. However, the magnitudes of DDI were not captured satisfactorily. Considering that clearance of RST is also mediated by gut breast cancer resistance protein (BCRP), inhibition of BCRP was also incorporated in the DDI prediction if the gut inhibitor concentrations were 10 × IC50 for BCRP inhibition. This combined static model closely predicted the magnitude of RST DDI with root-mean-square error values of 0.767-0.812 and 1.24-1.31 with and without BCRP inhibition, respectively, for in vitro-in vivo correlation of DDI. Physiologically based pharmacokinetic (PBPK) modeling was also used to simulate DDI between RST and rifampicin, asunaprevir, and velpatasvir. Predicted AUCR for rifampicin and asunaprevir was within 1.5-fold of that observed, whereas that for velpatasvir showed a 2-fold underprediction. Overall, the combined static model incorporating both OATP1B and BCRP inhibition provides a quick and simple mathematical approach to quantitatively predict the magnitude of transporter-mediated DDI for RST for routine application. PBPK complements the static model and provides a framework for studying molecules when a dynamic model is needed. SIGNIFICANCE STATEMENT: Using 22 drugs, we show that a static model for organic anion-transporting polypeptide (OATP) 1B1/1B3 inhibition can qualitatively predict potential for drug-drug interaction (DDI) using a cutoff-based approach, as in regulatory guidances. However, consideration of both OATP1B1/3 and gut breast cancer resistance protein inhibition provided a better prediction of the magnitude of the transporter-mediated DDI of these inhibitors with rosuvastatin. Based on these results, we have proposed an empirical mechanistic-static approach for a more reliable prediction of transporter-mediated DDI liability with rosuvastatin that drug development teams can leverage.
Asunto(s)
Transportador 1 de Anión Orgánico Específico del Hígado/metabolismo , Modelos Biológicos , Rosuvastatina Cálcica/farmacocinética , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/metabolismo , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2/antagonistas & inhibidores , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2/metabolismo , Área Bajo la Curva , Colecistoquinina/farmacocinética , Evaluación Preclínica de Medicamentos , Interacciones Farmacológicas , Estradiol/análogos & derivados , Estradiol/farmacocinética , Células HEK293 , Humanos , Transportador 1 de Anión Orgánico Específico del Hígado/antagonistas & inhibidores , Proteínas de Neoplasias/antagonistas & inhibidores , Proteínas de Neoplasias/metabolismo , Fragmentos de Péptidos/farmacocinética , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/antagonistas & inhibidoresRESUMEN
New nanocarriers are obtained by assembling two amphiphilic monomers: one containing the bioactive peptide CCK8 spaced, by a polydisperse poly(ethylene glycol), from two hydrophobic tails ((C18)2PEG2000CCK8), and the other containing a chelating agent able to give stable radiolabeled indium-111 complexes linked to the same hydrophobic moiety ((C18)2DTPAGlu). The size and shape of the supramolecular aggregates were structurally characterized by dynamic light scattering, small-angle neutron scattering, and cryogenic transmission electronic microscopy. Under the experimental conditions we investigated (pH 7.4 and molar ratio between monomers 30:70), there is the presence of high polydisperse aggregates: rod-like micelles with a radius of approximately 40 A and length >700 A, open bilayer fragments with thickness approximately 65 A, and probably vesicles. The presence of the bioactive peptide well exposed on the external surface of the aggregate allows selective targeting of nanocarriers towards the cholecystokinin receptors overexpressed by the cancerous cells. In vitro binding assays and in vivo biodistribution studies by nuclear medicine experiments using indium-111 are reported. Moreover, preliminary data concerning the drug loading capability of the aggregates and their drug efficiency on the target cells is reported by using the cytotoxic drug doxorubicin. Incubation of receptor-positive and control cells with peptide-containing aggregates filled with doxorubicin shows significantly lower cell survival in receptor-expressing cells relative to the control, for samples incubated in the presence of doxorubicin.
Asunto(s)
Colecistoquinina/química , Portadores de Fármacos/química , Nanoestructuras/química , Fragmentos de Péptidos/química , Animales , Colecistoquinina/farmacocinética , Microscopía por Crioelectrón , Femenino , Radioisótopos de Indio , Ratones , Micelas , Fragmentos de Péptidos/farmacocinética , Dispersión de Radiación , Distribución TisularRESUMEN
INTRODUCTION: During therapy with radiolabeled peptides, the kidney is most often the critical organ. Newly developed peptides are evaluated preclinically in different animal models before their application in humans. In this study, the renal retention of several radiolabeled peptides was compared in male and female rats and mice. METHODS: After intravenous injection of radiolabeled peptides [somatostatin, cholecystokinin (CCK), minigastrin, bombesin and neurotensin analogues], renal uptake was determined in both male and female Lewis rats and C57Bl mice. In addition, ex vivo autoradiography of renal sections was performed to localize accumulated radioactivity. RESULTS: An equal distribution pattern of renal radioactivity was found for all peptides: high accumulation in the cortex, lower accumulation in the outer medulla and no radioactivity in the inner medulla of the kidneys. In both male rats and mice, an increasing renal uptake was found: [(111)In-DTPA]CCK8<[(111)In-DTPA-Pro(1),Tyr(4)]bombesin approximately [(111)In-DTPA]neurotensin<[(111)In-DTPA]octreotide<<[(111)In-DTPA]MG0. Renal uptake of [(111)In-DTPA]octreotide in rats showed no gender difference, and renal radioactivity was about constant over time. In mice, however, renal uptake in females was significantly higher than that in males and decreased rapidly over time in both genders. Moreover, renal radioactivity in female mice injected with [(111)In-DTPA]octreotide showed a different localization pattern. CONCLUSIONS: Regarding the renal uptake of different radiolabeled peptides, both species showed the same ranking order. Similar to findings in patients, rats showed comparable and constant renal retention of radioactivity in both genders, in contrast to mice. Therefore, rats appear to be the more favorable species for the study of the renal retention of radioactivity.
Asunto(s)
Bombesina/farmacocinética , Colecistoquinina/análogos & derivados , Colecistoquinina/farmacocinética , Gastrinas/farmacocinética , Antagonistas de Hormonas/farmacocinética , Riñón/diagnóstico por imagen , Riñón/metabolismo , Neurotensina/farmacocinética , Somatostatina/farmacocinética , Animales , Autorradiografía , Femenino , Inmunohistoquímica , Técnicas In Vitro , Radioisótopos de Indio , Marcaje Isotópico , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Desnudos , Ácido Pentético/farmacocinética , Cintigrafía , Ratas , Ratas Endogámicas Lew , Caracteres Sexuales , Especificidad de la Especie , Distribución TisularRESUMEN
BACKGROUND AND PURPOSE: Acute intraperitoneal (i.p.) administration of cholecystokinin (CCK) is known to induce a significant, but short-lasting, reduction in food intake, followed by recovery within hours. Therefore, we had covalently coupled CCK to a 10 kDa polyethylene glycol and showed that this conjugate, PEG-CCK(9), produced a significantly longer anorectic effect than unmodified CCK(9). The present study assessed the dose-dependency of this response and the effect of two selective CCK(1) receptor antagonists, with different abilities to cross the blood-brain barrier (BBB), on PEG-CCK(9)-induced anorexia. EXPERIMENTAL APPROACH: Food intake was measured, for up to 23 h, after i.p. administration of different doses (2, 4, 8, 16 and 32 microg kg(-1)) of CCK(9) or PEG-CCK(9) in male Wistar rats. Devazepide (100 microg kg(-1)), which penetrates the BBB or 2-NAP (3 mg kg(-1)), which does not cross the BBB, were coadministered i.p. with PEG-CCK(9) (6 microg kg(-1)) and food intake was monitored. KEY RESULTS: In PEG-CCK(9)-treated rats, a clear dose-dependency was seen for both the duration and initial intensity of the anorexia whereas, for CCK(9), only the initial intensity was dose-dependent. Intraperitoneal administration of devazepide or 2-NAP, injected immediately prior to PEG-CCK(9), completely abolished the anorectic effect of PEG-CCK(9). CONCLUSIONS AND IMPLICATIONS: The duration of the anorexia for PEG-CCK(9) was dose-dependent, suggesting that PEGylation of CCK(9) increases its circulation time. Both devazepide and 2-NAP completely abolished the anorectic effect of i.p. PEG-CCK(9) indicating that its anorectic effect was solely due to stimulation of peripheral CCK(1) receptors.
Asunto(s)
Anorexia/inducido químicamente , Colecistoquinina/farmacología , Ingestión de Alimentos/efectos de los fármacos , Fragmentos de Péptidos/farmacología , Receptor de Colecistoquinina A/efectos de los fármacos , Saciedad/efectos de los fármacos , Animales , Depresores del Apetito/administración & dosificación , Depresores del Apetito/química , Depresores del Apetito/farmacocinética , Depresores del Apetito/farmacología , Ácido Aspártico/análogos & derivados , Ácido Aspártico/farmacología , Barrera Hematoencefálica , Colecistoquinina/administración & dosificación , Colecistoquinina/química , Colecistoquinina/farmacocinética , Preparaciones de Acción Retardada , Devazepida/farmacología , Relación Dosis-Respuesta a Droga , Inyecciones Intraperitoneales , Masculino , Naftalenosulfonatos/farmacología , Fragmentos de Péptidos/administración & dosificación , Fragmentos de Péptidos/química , Fragmentos de Péptidos/farmacocinética , Polietilenglicoles/química , Ratas , Ratas Wistar , Receptor de Colecistoquinina A/metabolismoRESUMEN
The radiolabeling of the natural octapeptide CCK8, derivatized with a cysteine residue (Cys-Gly-CCK8), by using the metal fragment [99mTc(N)(PNP3)]2+ (PNP3 = N,N-bis(dimethoxypropylphosphinoethyl)methoxyethylamine) is reported. The [99mTc(N)(NS-Cys-Gly-CCK8)(PNP3)]+ complex was obtained according to two methods (one-step or two-step procedure) that gave the desired compound in high yield. The complex is stable in aqueous solution and in phosphate buffer. In vitro challenge experiments with an excess of cysteine and glutathione indicate that no transchelation reactions occur, confirming the high thermodynamic stability and kinetic inertness of this compound. Stability studies carried out in human and mouse serum, as well as in mouse liver homogenates, show that the radiolabeled compound remains intact for prolonged incubation at 37 degrees C. Binding properties give Kd (19.0 +/- 4.6 nmol/l) and Bmax (approximately 10(6) sites/cell) values in A431 cells overexpressing the CCK2-R. In vivo evaluation of the compound shows rapid and specific targeting to CCK2-R, a fourfold higher accumulation compared to nonreceptor expressing tumors.
Asunto(s)
Colecistoquinina/química , Colecistoquinina/metabolismo , Compuestos de Organotecnecio/química , Compuestos de Organotecnecio/farmacocinética , Fragmentos de Péptidos/química , Fragmentos de Péptidos/metabolismo , Receptor de Colecistoquinina B/análisis , Animales , Tampones (Química) , Colecistoquinina/farmacocinética , Humanos , Marcaje Isotópico/métodos , Ratones , Ratones Desnudos , Biología Molecular/métodos , Fragmentos de Péptidos/farmacocinética , Radiofármacos/química , Receptor de Colecistoquinina B/genética , Receptor de Colecistoquinina B/metabolismo , Distribución Tisular , Trasplante HeterólogoRESUMEN
Otsuka Long Evans Tokushima Fatty (OLETF) rats have a deletion in the gene encoding the cholecystokinin-1 (CCK1) receptor. This deletion prevents protein expression, making the OLETF rat a CCK1 receptor knockout model. Consistent with the absence of CCK1 receptors, OLETF rats do not reduce their food intake in response to exogenously administered CCK and consume larger than normal meals. This deficit in within-meal feedback signaling is evident in liquid as well as solid meals. Neonatal OLETF rats show similar differences in independent ingestion tests. Intake is higher and is reflected in greater licking behavior. Neonatal OLETF rats also have diminished latencies to consume and higher initial ingestion rats. Adult OLETF rats are hyperphagic and obese. Although arcuate nucleus peptide gene expression is apparently normal in OLETF rats, when obesity is prevented through pair-feeding to amounts consumed by control Long Evans Tokushima Otsuka (LETO) rats, dorsomedial hypothalamic NPY mRNA expression is significantly elevated in OLETF rats. NPY overexpression is also evident in preobese, juvenile OLETF rats suggesting a causal role for this overexpression in the hyperphagia and obesity. Running wheel exercise normalizes food intake and body weight in OLETF rats. When access to exercise is provided at a time when OLETF rats are obese, the effects are limited to the period of exercise. When running wheel access is available to younger, preobese OLETF rats, exercise results in long lasting reductions in food intake and body weight and improved glucose regulation. These lasting metabolic effects of exercise may be secondary to an exercise induced reduction in DMH NPY mRNA expression.
Asunto(s)
Colecistoquinina/administración & dosificación , Hiperfagia/genética , Obesidad/genética , Receptores de Colecistoquinina/genética , Animales , Colecistoquinina/farmacocinética , Ingestión de Energía , Conducta Alimentaria , Eliminación de Gen , Hiperfagia/metabolismo , Núcleo Talámico Mediodorsal/metabolismo , Obesidad/metabolismo , ARN Mensajero/genética , Ratas , Ratas Endogámicas OLETF , Receptores de Colecistoquinina/metabolismoRESUMEN
Regulatory peptide receptors are overexpressed in numerous human cancers. These receptors have been used as molecular targets by which radiolabeled peptides can localize cancers in vivo and, more recently, to treat cancers with peptide receptor radiation therapy (PRRT). This review describes the candidate tumors eligible for such radiotherapy on the basis of their peptide receptor content and discusses factors in PRRT eligibility. At the present time, PRRT is performed primarily with somatostatin receptor- and cholecystokinin-2 (CCK2)-receptor-expressing neuroendocrine tumors with radiolabeled octreotide analogs or with radiolabeled CCK2-selective analogs. In the future, PRRT may be extended to many other tumor types, including breast, prostate, gut, pancreas, and brain tumors, that have recently been shown to overexpress several other peptide receptors, such as gastrin-releasing peptide-, neurotensin-, substance P-, glucagon-like peptide 1-, neuropeptide Y-, or corticotropin-releasing factor-receptors. A wide range of radiolabeled peptides is being developed for clinical use. Improved somatostatin or CCK(2) analogs as well as newly designed bombesin, neurotensin, substance P, neuropeptide Y, and glucagon-like peptide-1 analogs offer promise for future PRRT.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Neoplasias/metabolismo , Neoplasias/radioterapia , Péptidos/farmacocinética , Péptidos/uso terapéutico , Radioisótopos/farmacocinética , Radioisótopos/uso terapéutico , Receptores de Péptidos/metabolismo , Animales , Bombesina/farmacocinética , Bombesina/uso terapéutico , Colecistoquinina/farmacocinética , Colecistoquinina/uso terapéutico , Ensayos Clínicos como Asunto , Sistemas de Liberación de Medicamentos/métodos , Evaluación Preclínica de Medicamentos/tendencias , Humanos , Guías de Práctica Clínica como Asunto , Pautas de la Práctica en Medicina/tendencias , Radiofármacos/farmacocinética , Radiofármacos/uso terapéutico , Somatostatina/análogos & derivados , Somatostatina/farmacocinética , Somatostatina/uso terapéutico , Sustancia P/farmacocinética , Sustancia P/uso terapéutico , Resultado del TratamientoRESUMEN
A broad spectrum of radiolabeled peptides with high affinity for receptors expressed on tumor cells is currently under preclinical and clinical investigation for scintigraphic imaging and radionuclide therapy. The present paper evaluates two (99m)Tc-labeled forms of the C-terminal octapeptide of cholecystokinin (CCK8): sulfated (s)CCK8, with high affinity for CCK1 and CCK2 receptors, and nonsulfated (ns)CCK8, with high affinity for CCK2 receptors but low affinity for CCK1 receptors. Peptides were conjugated with the bifunctional chelator N-hydroxysuccinimidyl hydrazino niconitate (s-HYNIC). (99m)Tc-labeling, performed in the presence of nicotinic acid and tricine, was highly efficient (approximately 95%) and yielded products with a high specific activity (approximately 700 Ci/mmol) and good stability (approximately 5% release of radiolabel during 16 h incubation in phosphate buffered saline at 37 degrees C). Chinese hamster ovary cells stably expressing the CCK1 receptor (CHO-CCK1 cells) internalized approximately 3% of added (99m)Tc-sCCK8 per confluent well during 2 h at 37 degrees C. Internalization was effectively blocked by excess unlabeled sCCK8. CHO-CCK1 cells did not internalize (99m)Tc-nsCCK8. Displacement of (99m)Tc-sCCK8 and -nsCCK8 by unlabeled CCK-8 (performed at 0 degrees C to prevent internalization) revealed 50% inhibitory concentrations (IC(50)) of 8 nM and >1 microM, respectively. CHO-CCK2 cells internalized approximately 25% and approximately 5% of added (99m)Tc-sCCK8 and -nsCCK8, respectively. In both cases internalization was blocked by excess unlabeled peptide. IC(50) values for the displacement of (99m)Tc-sCCK8 and -nsCCK8 were 3 nM and 10 nM, respectively. CHO-CCK1 cell-derived tumors present in one flank of athymic mice accumulated 2.0% of injected (99m)Tc-sCCK8 per gram tissue at 1 h postinjection. This value decreased to 0.6% following coinjection with excess unlabeled peptide. Uptake of (99m)Tc-nsCCK8 was low (0.2%) and not did change by excess unlabeled peptide (0.3%). Accumulation of (99m)Tc-sCCK8 and -nsCCK8 by CHO-CCK2 cell-derived tumors (present in the other flank) amounted to 4.2% and 0.6%, respectively. In both cases uptake was significantly reduced by excess unlabeled peptide to 1.0% and 0.4% for sCCK8 and nsCCK8, respectively. Accumulation of (99m)Tc-sCCK8 was also high in pancreas (11.7%), stomach (2.0%), and kidney (2.1%), whereas uptake of (99m)Tc-nsCCK8 was high in stomach (0.7%) and kidney (1.4%). Both radiolabeled peptides showed a rapid blood clearance. In conclusion, these data show that CCK8 analogues can be efficiently labeled with (99m)Tc using s-HYNIC as chelator and nicotinic acid/tricine as coligand system without compromising receptor binding. Furthermore, the present study demonstrates that CCK1 tumors hardly accumulate (99m)Tc-nsCCK8, CCK2 tumors accumulate 2 times more (99m)Tc-sCCK8 than CCK1 tumors, and CCK2 tumors accumulate 15 times more (99m)Tc-sCCK8 than (99m)Tc-nsCCK8. Although accumulation in some nontarget organs was also higher with (99m)Tc-sCCK8, this may not reflect the human situation due to a different receptor expression pattern in humans as compared to mice. Therefore, further studies are warranted to investigate the possible use of (99m)Tc-sCCK8 for scintigraphic imaging of CCK receptor-positive tumors in humans.
Asunto(s)
Colecistoquinina/química , Neoplasias/diagnóstico por imagen , Fragmentos de Péptidos/química , Péptidos/química , Cintigrafía/métodos , Receptores de Colecistoquinina/metabolismo , Tecnecio/química , Animales , Células CHO , Colecistoquinina/farmacocinética , Cricetinae , Humanos , Marcaje Isotópico , Ratones , Estructura Molecular , Neoplasias/metabolismo , Fragmentos de Péptidos/farmacocinética , Péptidos/farmacocinética , Receptores de Colecistoquinina/efectos de los fármacos , Distribución TisularRESUMEN
AIM: To evaluate the effect of early intrajejunal nutrition (EIN) on the natural course, entero-hormone secretion and its efficacy on dogs with acute pancreatitis. METHODS: An acute pancreatitis model was induced by injecting 1 ml/kg of combined solution (2.5% sodium taurocholate and 8,000-10,000 BAEE units trypsin/ml) into the pancreas via pancreatic duct. Fifteen dogs were divided into parenteral nutrition (PN) group and EIN group. Two groups were isonitrogenous and isocaloric. EIN was used at postoperative 24 h. Serum glucose, calcium, amylase and lysosomal enzymes were determined before and 1, 4, 7 d after acute pancreatitis was induced. All the dogs were injected 50 uCi 125I-BSA 4 h before sacrificed on the 7th day. The 125I -BSA index of the pancreas/muscle, pancreas/blood, and pancreas pathology score (PPS) were determined. The peripheral plasma cholecystokinin (CCK), secretin (SEC) and gastrin were measured by ELISA and RIA, and was quantitative analysis of pancreatic juice and amylase, pancreatolipase and HCO3-, Cl-, Na+ and K+ performed by an autochemical analyzer at 30, 60, 120 and 180 min after beginning PN or EIN on the first day. RESULTS: There was no difference between two groups in the contents of serum calcium, amylase and lysosomal enzymes, 125I-BSA index of pancreas/muscle and pancreas/blood and PPS. The contents of CCK and gastrin in EIN were higher than those in PN group at 60 and 120 min (P<0.05). The content of SEC post-infusion of nutrition solution was higher than that of pre-infusion of nutrition solution in both groups, and only at 60 min SEC in EIN group was higher than that in PN group. The content of gastrin in EIN was higher than that in PN group at 120 and 180 min (P<0.05). The changes of pancreatic juice, amylase, pancreatolipase and HCO3-, Cl-, Na+ and K+ between two groups did not reach significantly statistical difference (P>0.05). CONCLUSION: EIN does not stimulate entero-hormone and pancreatic juice secretion, and enzyme-protein synthesis and release. EIN has no effect on the natural course of acute pancreatitis.
Asunto(s)
Colecistoquinina/farmacocinética , Yeyuno , Pancreatitis/dietoterapia , Pancreatitis/metabolismo , Nutrición Parenteral , Enfermedad Aguda , Amilasas/sangre , Animales , Glucemia/metabolismo , Calcio/sangre , Perros , Gastrinas/sangre , Radioisótopos de Yodo , Lisosomas/enzimología , Jugo Pancreático/metabolismo , Secretina/sangre , Albúmina Sérica Bovina/farmacocinéticaRESUMEN
PKC-delta is important in cell growth, apoptosis, and secretion. Recent studies show its stability is regulated by tyrosine phosphorylation (TYR-P), which can be stimulated by a number of agents. Many of these stimuli also activate phospholipase C (PLC) cascades and little is known about the relationship between these cascades and PKC-delta TYR-P. Cholecystokinin (CCK) stimulates PKCs but it is unknown if it causes PKC-delta TYR-P and if so, the relationship between these cascades is unknown. In rat pancreatic acini, CCK-8 stimulated rapid PKC-delta TYR-P by activation of the low affinity CCK(A) receptor state. TPA had a similar effect. BAPTA did not decrease CCK-stimulated PKC-delta TYR-P but instead, increased it. A23187 did not stimulate PKC-delta TYR-P. Wortmannin and LY 294002 did not alter CCK-stimulated PKC-delta TYR-P. GF 109203X, at low concentrations, increased PKC-delta TYR-P stimulated by CCK or TPA and at higher concentrations, inhibited it. The cPKC inhibitors, Gö 6976 and safingol, caused a similar increase in TPA- and CCK-stimulated PKC-delta TYR-P. These results demonstrate that CCK(A) receptor activation causes PKC-delta TYR-P through activation of only one of its two receptor affinity states. This PKC-delta TYR-P is not directly influenced by changes in [Ca(2+)](i); however, the resultant activation of PKC-alpha has an inhibitory effect. Therefore, CCK activates both stimulatory and inhibitory PKC cascades regulating PKC-delta TYR-P and, hence, likely plays an important role in regulating PKC-delta degradation and cellular abundance.
Asunto(s)
Colecistoquinina/metabolismo , Isoenzimas/metabolismo , Páncreas/enzimología , Proteína Quinasa C/metabolismo , Animales , Calcio/metabolismo , Colecistoquinina/farmacocinética , Técnicas de Cultivo , Isoenzimas/efectos de los fármacos , Masculino , Fragmentos de Péptidos/farmacocinética , Fosforilación/efectos de los fármacos , Fosfotransferasas/efectos de los fármacos , Fosfotransferasas/metabolismo , Proteína Quinasa C/efectos de los fármacos , Proteína Quinasa C-delta , Ratas , Ratas Wistar , Receptor de Colecistoquinina A , Receptores de Colecistoquinina/metabolismo , Regulación hacia Arriba/fisiologíaRESUMEN
The aim of this study is to label CCK-8 with Tc-99m and to investigate its radiopharmaceutical potential. CCK-8 was labeled with Tc-99m using GH and DTPA as bifunctional chelating agents. Labeling efficiency was higher than 99%. Complex was stable more than 5 hours at room temperature. 37 MBq Tc-99m-GH-CCK-8 or Tc-99m-DTPA-CCK-8 was administered intravenously to rabbits for biodistribution experiments. Dynamic and static images were obtained from anterior projection using a Camstar XC/T gamma camera. For quantitative evaluation, regions of interest were drawn on organs and time-activity curves were generated. The highest accumulation occurred in brain within 10 and 30 minutes after injection. Renal and hepatobiliary excretion were observed. Brain distribution studies in rats showed the highest activity was in hypothalamus. Results demonstrated that Tc-99m-GH-CCK-8 and Tc-99m-DTPA-CCK-8 analogs may be a useful new class of receptor-binding peptides for diagnosis and therapy of brain diseases related with CCK-B receptor-expressing tumors.
Asunto(s)
Colecistoquinina/farmacocinética , Hormona del Crecimiento/farmacocinética , Ácido Pentético/farmacocinética , Radiofármacos/farmacocinética , Receptores de Colecistoquinina/metabolismo , Compuestos de Tecnecio/farmacocinética , Animales , Encéfalo/diagnóstico por imagen , Colecistoquinina/análogos & derivados , Hormona del Crecimiento/análogos & derivados , Ácido Pentético/análogos & derivados , Conejos , Cintigrafía , Ratas , Ratas Wistar , Receptor de Colecistoquinina B , Distribución TisularRESUMEN
The gut and brain peptide cholecystokinin (CCK) exerts a number of central nervous effects. Among them are effects on attention and stimulus processing as revealed by modulations of event-related potentials (ERPs). In the present study the time course of central nervous effects after an intranasal administration of CCK-8 was investigated by means of ERPs. ERPs were recorded in an oddball paradigm 15, 30, 60, 90, 120, and 240 min after administration. Following the double-blind intranasal administration of CCK-8 and placebo, the late positive complex (LPC) of the ERP was significantly increased following CCK-8 compared to placebo. This effect was more pronounced in women than in men. The enhancement of the LPC by intranasal CCK-8 was not restricted to a specific recording time but reached its maximum 120 min after administration in men and women. Moreover, results tentatively indicate that 30 min after administration of CCK-8 the LPC increased only in women but not in men. The early effect of intranasal CCK-8 on LPC in women is unlikely to be caused by changes in plasma CCK-8 levels and suggests a direct nose-brain pathway.
Asunto(s)
Sistema Nervioso Central/efectos de los fármacos , Colecistoquinina/farmacología , Colecistoquinina/farmacocinética , Fragmentos de Péptidos/farmacología , Fragmentos de Péptidos/farmacocinética , Administración Intranasal , Adulto , Área Bajo la Curva , Colecistoquinina/administración & dosificación , Estudios Cruzados , Electroencefalografía/efectos de los fármacos , Electrooculografía/efectos de los fármacos , Potenciales Evocados/efectos de los fármacos , Femenino , Humanos , Masculino , Fragmentos de Péptidos/administración & dosificaciónRESUMEN
Sphincter of Oddi dysfunction is an underdiagnosed but important clinical condition. It should be considered in the differential diagnosis of biliary pain when the gallbladder sonogram shows no evidence of gallbladder disease. Hepatobiliary scanning (Tc-99m dimethyl iminodiacetic acid) may provide valuable information in the evaluation of these patients and may be helpful in monitoring response to treatment.
Asunto(s)
Enfermedades del Conducto Colédoco/diagnóstico por imagen , Esfínter de la Ampolla Hepatopancreática/diagnóstico por imagen , Adulto , Colangiopancreatografia Retrógrada Endoscópica , Colecistoquinina/farmacocinética , Enfermedades del Conducto Colédoco/cirugía , Femenino , Humanos , Persona de Mediana Edad , Cintigrafía , Radiofármacos , Esfínter de la Ampolla Hepatopancreática/cirugía , Esfinterotomía Transduodenal , Ácido Dietil-Iminodiacético de Tecnecio Tc 99mRESUMEN
Expression of the long form of the leptin receptor, the isoform that is considered to have full signaling capability, has been reported in the central nervous system and several peripheral cell types. However, only a few cell lines have been shown to express the long form of the receptor. AR42J, a cell line derived from azaserine-treated rat pancreas, is a common model for pancreatic acinar cell secretion. In this study, the presence of leptin-receptor variants and leptin action was evaluated in this cell line. Messenger RNAs for both the long and a short form of the leptin receptor were detected by reverse transcription-polymerase chain reaction (RT-PCR) in AR42J cells, and authenticity of the receptor was confirmed by DNA sequencing. Competitive binding studies demonstrated that binding of radiolabeled leptin was specific and did not cross-react with cholecystokinin (CCK). Biologic effects of leptin on amylase release and intracellular calcium mobilization were further assessed in the presence and the absence of CCK, a known pancreatic secretagogue. Although leptin alone (< or =200 ng/ml) did not affect basal amylase release, it inhibited amylase release stimulated by 1 nM CCK by 48%. Leptin alone had no significant effect on calcium mobilization. However, pretreatment of leptin (10 and 100 ng/ml) enhanced calcium responses stimulated by CCK. These data demonstrate that the rat pancreatic tumor cell line AR42J expresses a functional form of the leptin receptor that modulates the action of CCK in calcium mobilization and amylase release.
Asunto(s)
Amilasas/metabolismo , Colecistoquinina/farmacología , Leptina/farmacología , Páncreas/efectos de los fármacos , Páncreas/metabolismo , Receptores de Superficie Celular , Amilasas/antagonistas & inhibidores , Animales , Unión Competitiva/efectos de los fármacos , Calcio/metabolismo , Proteínas Portadoras/genética , Línea Celular , Colecistoquinina/farmacocinética , Relación Dosis-Respuesta a Droga , Fármacos Gastrointestinales/farmacología , Líquido Intracelular/metabolismo , Leptina/farmacocinética , Isoformas de Proteínas/genética , ARN Mensajero/metabolismo , Ratas , Receptores de Leptina , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Análisis de Secuencia de ADN , Sincalida/farmacologíaRESUMEN
UNLABELLED: The presence of cholecystokinin (CCK)-B (gastrin) receptors has been shown in more than 90% of medullary thyroid cancers (MTCs) and in a high percentage of small cell lung cancers, stromal ovarium cancers and several other tumor types. METHODS: The aim of this study was to evaluate in vitro and in vivo whether 111In-labeled CCK-B receptor-specific CCK8 analog [D-Asp26,Nle28,31]CCK26-33 (D-Asp-Tyr-Nle-Gly-Trp-Nle-Asp-Phe-NH2) is suitable for CCK-B receptor scintigraphy based on the finding that unlabeled nonsulfated diethylenetriamine pentaacidic acid [DTPA0]CCK8 and tetraazacyclododecanetetraacetic acid [DOTA0]CCK8 analogs show high and specific binding for CCK-B receptors in human tumors. Fifty percent inhibitory concentrations were in the low nanomolar range. RESULTS: In vitro, [111In-DOTA0]CCK8 showed specific internalization in CCK-B receptor-positive rat pancreatic tumor cells AR42J. Internalization of the analog appeared to be time and temperature dependent and receptor specific. From the data obtained with [111In-DOTA0]CCK8 and (125I)I-gastrin, the latter being a specific ligand for the CCK-B receptor, the rat pancreatic cell line CA20948 also appeared to be CCK-B receptor positive. This provides an in vitro and in vivo rat tumor model because this cell line can be grown to solid tumors in Lewis rats. In vivo biodistribution experiments in CA20948 tumor-bearing Lewis rats showed rapid clearance of [111In-DOTA0]CCK8, and specific uptake was found in the CCK-B receptor-expressing stomach and tumor. Furthermore, comparing [111In-DOTA0]CCK8 with the radioiodinated nonsulfated CCK10 analog (D-Tyr-Gly-Asp-Tyr-Nle-Gly-Trp-Nle-Asp-Phe-NH2), both ligands having high affinity for the CCK-B receptor, tumor-to-blood ratios were significantly higher for [111In-DOTA0]CCK8 than for 125I-CCK10, analogous to the findings with radioiodinated and 111In-labeled octreotide. The study in humans with [111In-DTPA0]CCK8 showed receptor-specific uptake in the CCK-B receptor-positive stomach and in metastases in the neck region up to 48 h after injection. CONCLUSION: [111In-DOTA0]CCK8 is most promising for scintigraphy and, after coupling to therapeutic radionuclides, for radionuclide therapy of human CCK-B receptor-positive tumors such as MTC and small cell lung cancer.
Asunto(s)
Colecistoquinina , Radioisótopos de Indio , Neoplasias Pancreáticas/diagnóstico por imagen , Fragmentos de Péptidos , Receptores de Colecistoquinina/análisis , Neoplasias de la Tiroides/diagnóstico por imagen , Animales , Colecistoquinina/farmacocinética , Colecistoquinina/uso terapéutico , Colecistoquinina/toxicidad , Evaluación de Medicamentos , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Persona de Mediana Edad , Neoplasias Pancreáticas/química , Fragmentos de Péptidos/farmacocinética , Fragmentos de Péptidos/uso terapéutico , Fragmentos de Péptidos/toxicidad , Cintigrafía , Ratas , Ratas Endogámicas Lew , Ratas Wistar , Receptor de Colecistoquinina B , Neoplasias de la Tiroides/química , Distribución Tisular , Células Tumorales CultivadasRESUMEN
The present study explores the effect of chemical penetration enhancers and iontophoresis on the in vitro permeability of cholecystokinin-8 (CCK-8) through porcine epidermis and on the ultrastructural changes in stratum corneum as observed by transmission electron microscopy (TEM). Enhancer [i.e., ethanol (EtOH), and 10% oleic acid in combination with ethanol (OA/EtOH)] pretreatment significantly increased (p < 0.01) the permeability coefficient of CCK-8 in comparison with the control (pretreated epidermis without enhancer). Iontophoresis further increased the permeability of CCK-8 (p < 0.01) through the enhancer-pretreated epidermis in comparison with the control. These results showed the synergistic effect of iontophoresis and enhancers such as OA/EtOH that provides an additional driving force to maintain and control the target flux of CCK-8. The ultrastructure of stratum corneum treated with ethanol demonstrated a loss of structural components in the superficial stratum corneum cell layers. OA/EtOH transformed the highly compact cells of stratum corneum into a looser network of filaments, creating an increased free volume and greater intracellular surface area. Treatment of stratum corneum with OA/EtOH followed by iontophoresis resulted in further swelling of stratum corneum cell layers. In conclusion, OA/EtOH in combination with iontophoresis increased the permeability of CCK-8 by loosening and swelling of stratum corneum cell layers.
Asunto(s)
Colecistoquinina/farmacocinética , Absorción Cutánea/efectos de los fármacos , Piel/ultraestructura , Animales , Técnicas In Vitro , Iontoforesis , Microscopía Electrónica , Piel/efectos de los fármacos , PorcinosRESUMEN
The effects of the selective CCK-B agonists, BC 264 and BC 197, and the nonselective CCK agonist BDNL were investigated in the elevated plus-maze in rats. BDNL and BC 197 induced anxiogeniclike effects, in contrast to BC 264, which had no effect. The behavioral responses induced by BDNL were not significantly blocked by L-365,260, but were suppressed by CI-988, another selective CCK-B antagonist, and by high doses of L-364,718, a selective CCK-A antagonist. BC 197-induced effects were also blocked by CI-988. Competition experiments performed with [3H]pBC 264 using brain membranes of guinea pig, mouse, and rat were significantly better fitted when analyzed by a two site model than by a one site model with BC 197 but not with BC 264. Moreover, BC 264 produced anxiogeniclike effects when administered with increasing doses of L-365,260 and opposing effects with increasing doses of CI-988. Together these results give pharmacological and behavioral evidence for the existence of CCK-B receptor subtypes.
Asunto(s)
Ansiedad/metabolismo , Compuestos de Fenilurea , Receptores de Colecistoquinina/metabolismo , Secuencia de Aminoácidos , Animales , Ansiedad/psicología , Conducta Animal/efectos de los fármacos , Benzodiazepinonas/farmacocinética , Benzodiazepinonas/farmacología , Colecistoquinina/análogos & derivados , Colecistoquinina/farmacocinética , Colecistoquinina/farmacología , Devazepida , Cobayas , Técnicas In Vitro , Indoles/farmacocinética , Indoles/farmacología , Masculino , Meglumina/análogos & derivados , Meglumina/farmacocinética , Meglumina/farmacología , Ratones , Datos de Secuencia Molecular , Páncreas/efectos de los fármacos , Páncreas/metabolismo , Fragmentos de Péptidos/farmacocinética , Fragmentos de Péptidos/farmacología , Ratas , Ratas Sprague-Dawley , Ratas Wistar , Receptores de Colecistoquinina/agonistas , Receptores de Colecistoquinina/antagonistas & inhibidoresRESUMEN
BACKGROUND: Cholecystokinin (CCK) 58 is the predominant molecular form of CCK in canine and human intestine and circulating blood. There is no report on the metabolism and clearance rate of CCK-58. The aim of this study was to compare the in vivo half-life and metabolism of CCK-58 with that of synthetic CCK-8. METHODS: CCK-58 was purified from canine intestine by consecutive high-performance liquid chromatographic (HPLC) and fast protein liquid chromatographic steps. The peptides were given to 12 dogs as an intravenous (IV) bolus injection to determine the half-life of circulating CCK. Six dogs were given CCK-58 or CCK-8 as a constant IV infusion to determine plasma clearance rates and stability in circulating blood. Circulating molecular forms of CCK were determined by radioimmunoassay after extraction of CCK from plasma and characterization by HPLC. RESULTS: The half-life of CCK-58 was 4.4 +/- 0.6 minutes compared with 1.3 +/- 0.1 minutes for CCK-8. Less than 5% of CCK-58 could be detected as smaller forms during constant IV infusion. CONCLUSIONS: The longer half-life of CCK-58 compared with CCK-8 and the minimal conversion into smaller forms during constant IV infusion are consistent with the finding that CCK-58 is not only the major stored form but also the circulating form of CCK after endogenous stimulation in dogs.
Asunto(s)
Colecistoquinina/farmacocinética , Sincalida/farmacocinética , Animales , Colecistoquinina/administración & dosificación , Colecistoquinina/aislamiento & purificación , Perros , Semivida , Infusiones Intravenosas , Inyecciones Intravenosas , Mucosa Intestinal/química , Tasa de Depuración Metabólica , Sincalida/administración & dosificación , Sincalida/síntesis químicaRESUMEN
A new synthetic analogue of cholecystokinin, Thr28Nle31CCK25-33(CCK9) is compared with caerulein with regards to plasma bioactivity, degradation rate, side effects, and stimulation of pancreatic secretion. 24 healthy male volunteers were intubated with a double lumen Lagerlöf-tube. 30 min after correct positioning of the tube subjects received a continuous intravenous infusion of synthetic secretin (1 U/kg) together with either ceruletide (61.5 pM/kg) or CCK9 (30 pM/kg) both for 45 min. 30 pM/kg of CCK9 have been shown by others to cause maximal enzyme secretion (1). 61.5 pM/kg (= 100 ng) of caerulein are probably supramaximal but used by many centers for direct pancreatic function tests. Plasma CCK was measured by bioassay which compares amylase release of isolated rat pancreatic acini stimulated by plasma extracts with known standards of CCK8. Lipase, amylase, trypsin, chymotrypsin, and bicarbonate were measured in 15 min fractions after the onset of CCK infusion. Both drugs caused a similar stimulation of pancreatic enzyme secretion during infusion of the respective analogue which declined after termination. After the onset of CCK9 infusion plasma bioactivity reached a plateau around 20 pM at 15 min. Values declined after 30 min. After termination of infusion bioactivity rapidly declined within 3 min but still remained slightly elevated after further 15 min as compared to basal values (3.9 vs 0.6 pM). Plasma kinetics of caerulein were quite similar. However, despite a dose which was only twice as high as compared to CCK9, plasma bioactivity was six times higher with plateau values of about 120 pM.(ABSTRACT TRUNCATED AT 250 WORDS)
Asunto(s)
Ceruletida/farmacocinética , Colecistoquinina/farmacocinética , Páncreas/efectos de los fármacos , Pruebas de Función Pancreática , Fragmentos de Péptidos/farmacocinética , Adulto , Amilasas/sangre , Animales , Bicarbonatos/sangre , Disponibilidad Biológica , Ceruletida/farmacología , Colecistoquinina/farmacología , Quimotripsina/sangre , Técnicas de Cultivo , Humanos , Infusiones Intravenosas , Secreciones Intestinales/efectos de los fármacos , Lipasa/sangre , Masculino , Fragmentos de Péptidos/farmacología , Ratas , Ratas Endogámicas , Tripsina/sangreRESUMEN
The aim of the present study was to analyze the effect of the specific cholecystokinin (CCK) receptor antagonist loxiglumide on hepatic and pancreatic processing of CCK-8 and the CCK analogue cerulein. Rat liver perfusion was performed in a non-recirculating system. CCK concentrations were measured by radioimmunoassay in perfusates from the inflow cannula (portal vein) and the outflow cannula (hepatic vein). In rat pancreatic acini, the effect of loxiglumide on internalization and surface-binding of radiolabelled CCK-8 was determined. Cerulein (20 nM, 2 nM) was extracted in a single pass through the liver by 29.7 and 25.4%, respectively. The hepatic uptake of CCK-8 (50 pM, 2 nM) was more than 90 and 89.9%, respectively. Loxiglumide drastically inhibited hepatic extraction of both peptides and reduced internalization of 125I-CCK-8 in pancreatic acini dose dependently by 39-93%. These results demonstrate that the potent CCK receptor antagonist loxiglumide significantly decreased CCK uptake by the liver and pancreas.