The preparation of specifically iodine-125 (125I)-labeled peptides of high purity and specific activity represents a key tool for the detailed characterization of their binding properties in interaction with their binding partners. Early synthetic methods for the incorporation of iodine faced challenges such as harsh reaction conditions, the use of strong oxidants and low reproducibility. Herein, we review well-established radiolabeling strategies available to incorporate radionuclide into a protein of interest, and our long-term experience with a mild, simple and generally applicable technique of 125I late-stage-labeling of biomolecules using the Pierce iodination reagent for the direct solid-phase oxidation of radioactive iodide. General recommendations, tips, and details of optimized chromatographic conditions to isolate pure, specifically 125I-mono-labeled biomolecules are illustrated on a diverse series of (poly)peptides, ranging up to 7.6 kDa and 67 amino acids (aa). These series include peptides that contain at least one tyrosine or histidine residue, along with those featuring disulfide crosslinking or lipophilic derivatization. This mild and straightforward late-stage-labeling technique is easily applicable to longer and more sensitive proteins, as demonstrated in the cases of the insulin-like growth factor binding protein-3 (IGF-BP-3) (29 kDa and 264 aa) and the acid-labile subunit (ALS) (93 kDa and 578 aa).

• Klíčová slova
• 125I-labeling of peptides, High specific activity, Intramolecular effect of 125I decay, Late-stage peptide labeling, Radiochemical stability, Radiohalogenated prosthetic groups, Site-specific labeling,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

Since 1975, the incidence of obesity has increased to epidemic proportions, and the number of patients with obesity has quadrupled. Obesity is a major risk factor for developing other serious diseases, such as type 2 diabetes mellitus, hypertension, and cardiovascular diseases. Recent epidemiologic studies have defined obesity as a risk factor for the development of neurodegenerative diseases, such as Alzheimer's disease (AD) and other types of dementia. Despite all these serious comorbidities associated with obesity, there is still a lack of effective antiobesity treatment. Promising candidates for the treatment of obesity are anorexigenic neuropeptides, which are peptides produced by neurons in brain areas implicated in food intake regulation, such as the hypothalamus or the brainstem. These peptides efficiently reduce food intake and body weight. Moreover, because of the proven interconnection between obesity and the risk of developing AD, the potential neuroprotective effects of these two agents in animal models of neurodegeneration have been examined. The objective of this review was to explore anorexigenic neuropeptides produced and acting within the brain, emphasizing their potential not only for the treatment of obesity but also for the treatment of neurodegenerative disorders.

• Klíčová slova
• Alzheimer´s-like pathology, anorexigenic neuropeptides, antiobesity treatment, neuroprotection,
• MeSH
• Alzheimerova nemoc farmakoterapie   metabolismus   patologie   prevence a kontrola   MeSH
• hypothalamus účinky léků   metabolismus   patologie   MeSH
• látky proti obezitě * farmakologie   terapeutické užití   MeSH
• lidé MeSH
• mozek účinky léků   metabolismus   patologie   MeSH
• neurodegenerativní nemoci farmakoterapie   metabolismus   prevence a kontrola   MeSH
• neuropeptidy * metabolismus   farmakologie   terapeutické užití   MeSH
• neuroprotektivní látky * farmakologie   terapeutické užití   MeSH
• obezita * farmakoterapie   metabolismus   MeSH
• přijímání potravy účinky léků   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• látky proti obezitě * MeSH
• neuropeptidy * MeSH
• neuroprotektivní látky * MeSH

The anorexigenic neuropeptide prolactin-releasing peptide (PrRP) is involved in the regulation of food intake and energy expenditure. Lipidization of PrRP stabilizes the peptide, facilitates central effect after peripheral administration and increases its affinity for its receptor, GPR10, and for the neuropeptide FF (NPFF) receptor NPFF-R2. The two most potent palmitoylated analogs with anorectic effects in mice, palm11-PrRP31 and palm-PrRP31, were studied in vitro to determine their agonist/antagonist properties and mechanism of action on GPR10, NPFF-R2 and other potential off-target receptors related to energy homeostasis. Palmitoylation of both PrRP31 analogs increased the binding properties of PrRP31 to anorexigenic receptors GPR10 and NPFF-R2 and resulted in a high affinity for another NPFF receptor, NPFF-R1. Moreover, in CHO-K1 cells expressing GPR10, NPFF-R2 or NPFF-R1, palm11-PrRP and palm-PrRP significantly increased the phosphorylation of extracellular signal-regulated kinase (ERK), protein kinase B (Akt) and cAMP-responsive element-binding protein (CREB). Palm11-PrRP31, unlike palm-PrRP31, did not activate either c-Jun N-terminal kinase (JNK), p38, c-Jun, c-Fos or CREB pathways in cells expressing NPFF-1R. Palm-PrRP31 also has higher binding affinities for off-target receptors, namely, the ghrelin, opioid (KOR, MOR, DOR and OPR-L1) and neuropeptide Y (Y1, Y2 and Y5) receptors. Palm11-PrRP31 exhibited fewer off-target activities; therefore, it has a higher potential to be used as an anti-obesity drug with anorectic effects.

• Klíčová slova
• GPR10, NPFF-R1, NPFF-R2, binding properties, neuropeptide FF, prolactin-releasing peptide, signaling pathways,
• MeSH
• CHO buňky MeSH
• Cricetulus MeSH
• hormon uvolňující prolaktin chemie   genetika   metabolismus   MeSH
• křečci praví MeSH
• lidé MeSH
• lipoylace * MeSH
• receptory neuropeptidů genetika   metabolismus   MeSH
• receptory spřažené s G-proteiny genetika   metabolismus   MeSH
• techniky in vitro MeSH
• vápník metabolismus   MeSH
• zvířata MeSH
• Check Tag
• křečci praví MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• hormon uvolňující prolaktin MeSH
• neuropeptide FF receptor MeSH Prohlížeč
• PRLH protein, human MeSH Prohlížeč
• PRLHR protein, human MeSH Prohlížeč
• receptory neuropeptidů MeSH
• receptory spřažené s G-proteiny MeSH
• vápník MeSH

Prolactin-releasing peptide (PrRP), a natural ligand for the GPR10 receptor, is a neuropeptide with anorexigenic and antidiabetic properties. Due to its role in the regulation of food intake, PrRP is a potential drug for obesity treatment and associated type 2 diabetes mellitus (T2DM). Recently, the neuroprotective effects of lipidized PrRP analogs have been proven. In this study, we focused on the molecular mechanisms of action of natural PrRP31 and its lipidized analog palm11-PrRP31 in the human neuroblastoma cell line SH-SY5Y to describe their cellular signaling and possible anti-apoptotic properties. PrRP31 significantly upregulated the phosphoinositide-3 kinase-protein kinase B/Akt (PI3K-PKB/Akt) and extracellular signal-regulated kinase/cAMP response element-binding protein (ERK-CREB) signaling pathways that promote metabolic cell survival and growth. In addition, we proved via protein kinase inhibitors that activation of signaling pathways is mediated specifically by PrRP31 and its palmitoylated analog. Furthermore, the potential neuroprotective properties were studied through activation of anti-apoptotic pathways of PrRP31 and palm11-PrRP31 using the SH-SY5Y cell line and rat primary neuronal culture stressed with toxic methylglyoxal (MG). The results indicate increased viability of the cells treated with PrRP and palm11-PrRP31 and a reduced degree of apoptosis induced by MG, suggesting their potential use in the treatment of neurological disorders.

• Klíčová slova
• SH-SY5Y, cellular signaling, inhibitors, methylglyoxal, neuroprotection, primary neuronal culture, prolactin-releasing peptide,
• MeSH
• apoptóza * MeSH
• hormon uvolňující prolaktin chemie   farmakologie   MeSH
• lidé MeSH
• nádorové buňky kultivované MeSH
• neuroblastom farmakoterapie   metabolismus   patologie   MeSH
• neuropeptidy chemie   farmakologie   MeSH
• neuroprotektivní látky chemie   farmakologie   MeSH
• proteiny regulující apoptózu metabolismus   MeSH
• signální transdukce MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• hormon uvolňující prolaktin MeSH
• neuropeptidy MeSH
• neuroprotektivní látky MeSH
• proteiny regulující apoptózu MeSH

Analogs of anorexigenic neuropeptides, such as prolactin-releasing peptide (PrRP), have a potential as new anti-obesity drugs. In our previous study, palmitic acid attached to the N-terminus of PrRP enabled its central anorexigenic effects after peripheral administration. In this study, two linkers, γ-glutamic acid at Lys11 and a short, modified polyethylene glycol at the N-terminal Ser and/or Lys11, were applied for the palmitoylation of PrRP31 to improve its bioavailability. These analogs had a high affinity and activation ability to the PrRP receptor GPR10 and the neuropeptide FF2 receptor, as well as short-term anorexigenic effect similar to PrRP palmitoylated at the N-terminus. Two-week treatment with analogs that were palmitoylated through linkers to Lys11 (analogs 1 and 2), but not with analog modified both at the N-terminus and Lys11 (analog 3) decreased body and liver weights, insulin, leptin, triglyceride, cholesterol and free fatty acid plasma levels in a mouse model of diet-induced obesity. Moreover, the expression of uncoupling protein-1 was increased in brown fat suggesting an increase in energy expenditure. In addition, treatment with analogs 1 and 2 but not analog 3 significantly decreased urinary concentrations of 1-methylnicotinamide and its oxidation products N-methyl-2-pyridone-5-carboxamide and N-methyl-4-pyridone-3-carboxamide, as shown by NMR-based metabolomics. This observation confirmed the previously reported increase in nicotinamide derivatives in obesity and type 2 diabetes mellitus and the effectiveness of analogs 1 and 2 in the treatment of these disorders.

• MeSH
• beta-laktamasy metabolismus   MeSH
• CHO buňky MeSH
• Cricetulus MeSH
• dieta * MeSH
• hormon uvolňující prolaktin chemie   metabolismus   MeSH
• kompetitivní vazba MeSH
• křečci praví MeSH
• metabolomika MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• nukleární magnetická rezonance biomolekulární MeSH
• obezita etiologie   metabolismus   MeSH
• peptidy chemie   farmakologie   MeSH
• sekvence aminokyselin MeSH
• zvířata MeSH
• Check Tag
• křečci praví MeSH
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• beta-laktamasy MeSH
• hormon uvolňující prolaktin MeSH
• peptidy MeSH

OBJECTIVES: Obesity is a frequent metabolic disorder but an effective therapy is still scarce. Anorexigenic neuropeptides produced and acting in the brain have the potential to decrease food intake and ameliorate obesity but are ineffective after peripheral application. We have designed lipidized analogs of prolactin-releasing peptide (PrRP), which is involved in energy balance regulation as demonstrated by obesity phenotypes of both PrRP- and PrRP-receptor-knockout mice. RESULTS: Lipidized PrRP analogs showed binding affinity and signaling in PrRP receptor-expressing cells similar to natural PrRP. Moreover, these analogs showed high binding affinity also to anorexigenic neuropeptide FF-2 receptor. Peripheral administration of myristoylated and palmitoylated PrRP analogs to fasted mice induced strong and long-lasting anorexigenic effects and neuronal activation in the brain areas involved in food intake regulation. Two-week-long subcutaneous administration of palmitoylated PrRP31 and myristoylated PrRP20 lowered food intake, body weight and improved metabolic parameters, and attenuated lipogenesis in mice with diet-induced obesity. CONCLUSIONS: Our data suggest that the lipidization of PrRP enhances stability and mediates its effect in central nervous system. Strong anorexigenic and body-weight-reducing effects make lipidized PrRP an attractive candidate for anti-obesity treatment.

• MeSH
• energetický metabolismus MeSH
• hormon uvolňující prolaktin analogy a deriváty   farmakologie   MeSH
• látky proti obezitě farmakologie   MeSH
• lipidy chemie   MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• obezita prevence a kontrola   MeSH
• poločas MeSH
• přijímání potravy MeSH
• regulace chuti k jídlu MeSH
• signální transdukce MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• hormon uvolňující prolaktin MeSH
• látky proti obezitě MeSH
• lipidy MeSH