RESUMEN
ß-lactoglobulin is one of the nutrition allergens present in the milk of many mammals, with the exception of human. This protein belongs to the family of lipocalins, consisting of nine antiparallel ß-strands (ß-A to ß-I) and one α-helix. This structure allows it to serve as a nanotransporter of various nature ligands in a pH dependent manner, which allows us to confidently consider it as a reliable carrier of drugs directly into the intestine, bypassing the destructive acidic environment of the stomach. Based on the latest data, this review describes the currently known methods of reducing the allergenicity of beta-lactoglobulin, as well as the mechanisms and methods of forming complexes of this protein with ligands, which emphasizes its importance and versatility and explains the growing interest in studying its properties in recent decades, and also opens up prospects for its practical application in medicine and pharmaceuticals.
Asunto(s)
Alérgenos/metabolismo , Lactoglobulinas/metabolismo , Leche/química , Alérgenos/análisis , Alérgenos/química , Animales , Hipersensibilidad a los Alimentos , Humanos , Concentración de Iones de Hidrógeno , Lactoglobulinas/análisis , Lactoglobulinas/química , Lactoglobulinas/genética , Ligandos , Reacción de Maillard , Leche/metabolismo , Estado NutricionalRESUMEN
BACKGROUND: Ferritin is a globular intracellular protein that acts as the main reservoir for iron. Malignancies are associated with increased plasma ferritin concentrations. A number of studies show that tumor cells express high levels of transferrin receptors (TfR). Increased TfR expression was observed in prostate carcinoma. Apoferritin (APO) can be used as a protein nanotransporter into which a suitable medicinal substance can be encapsulated. Nanoparticles increase the permeability of tumor cells to nanotransporters and have a photothermal effect. The aim of this study was to encapsulate doxorubicin (DOX) into APO and to modify the resulting APO/DOX with gold (AuNPs) and silver nanoparticles prepared by green synthesis (AgNPsGS). METHODS: APO was characterized using 10% sodium dodecylsulphate polyacrylamide gel electrophoresis (SDS-PAGE) - 120 V, 60 min, 24 mM Tris, 0.2 M glycine, 3 mM SDS. DOX fluorescence (Ex 480 nm; Em 650 nm) was observed, with a typical absorption maximum at 560 nm. Electrochemical measurement was performed in Brdicka solution (three-electrode setup). AgNPsGS were prepared by green synthesis using clover (Trifolium pratense L.). RESULTS: An electrophoretic study of APO and APO/DOX (5-100 μg/mL) was performed and the behavior of APO and APO/DOX (10 μM) as a function of pH was monitored. In an acidic environment, APO forms subunits of about 20 kDa; in an alkaline medium, it forms a globular protein of about 450 kDa. A change in APO/DOX mobility (about by 10%) was observed. A film of gold nanoparticles was applied to the APO/DOX surface. APO/DOX-AuNPs were washed with ultra-pure water. pH-dependent release of DOX a was monitored. The amount of DOX analyzed was increased by up to 50%. Furthermore, an AgNPsGS-DOX complex (1 mg AgNPsGS/100 μM DOX) was generated and prepared. Subsequently, the AgNPsGS-DOX complex was encapsulated into APO. To further improve therapeutic efficacy, the APO/AgNPsGS-DOX complex was coated with an Au layer. APO/AgNPsGS-DOX/AuNPs were stable and DOX was released from the complex after physical parameters had changed. CONCLUSION: APO nanocomplexes were prepared and modified to increase therapeutic efficacy against tumors. Tumor cell targeting was achieved by binding to TfR and via increased tumor cell permeability and retention. Release of the drug was made possible due to a pH change and photothermal activation that will now be tested. This work was supported by COST European Cholangiocarcinoma Network CA18122 and International Collaboration Project of The European Technology Platform for Nanomedicine. The authors declare they have no potential conflicts of interest concerning drugs, products, or services used in the study. The Editorial Board declares that the manuscript met the ICMJE recommendation for biomedical papers. Submitted: 21. 3. 2019 Accepted: 14. 5. 2019.
Asunto(s)
Antibióticos Antineoplásicos/química , Apoferritinas/química , Doxorrubicina/análogos & derivados , Nanopartículas del Metal/química , Antibióticos Antineoplásicos/farmacología , Línea Celular Tumoral , Doxorrubicina/química , Doxorrubicina/farmacología , Liberación de Fármacos , Oro/química , Humanos , Concentración de Iones de Hidrógeno , Receptores de Transferrina/metabolismo , Plata/químicaRESUMEN
Protein supplementation therapy using in vitro-transcribed (IVT) mRNA for genetic diseases contains huge potential as a new class of therapy. From the early ages of synthetic mRNA discovery, a great number of studies showed the versatile use of IVT mRNA as a novel approach to supplement faulty or absent protein and also as a vaccine. Many modifications have been made to produce high expressions of mRNA causing less immunogenicity and more stability. Recent advancements in the in vivo lung delivery of mRNA complexed with various carriers encouraged the whole mRNA community to tackle various genetic lung diseases. This review gives a comprehensive overview of cells associated with various lung diseases and recent advancements in mRNA-based protein replacement therapy. This review also covers a brief summary of developments in mRNA modifications and nanocarriers toward clinical translation.
Asunto(s)
Terapia de Reemplazo Enzimático/métodos , Enfermedades Pulmonares/tratamiento farmacológico , ARN Mensajero/administración & dosificación , ARN Mensajero/genética , Transcripción Genética , Animales , Sistemas de Liberación de Medicamentos/métodos , Humanos , Lípidos/química , Pulmón/metabolismo , Enfermedades Pulmonares/metabolismo , Enfermedades Pulmonares/patología , Ratones , Nanopartículas/química , Polímeros/química , Procesamiento Postranscripcional del ARNRESUMEN
INTRODUCTION: Modular nanotransporters (MNTs) are vehicles designed to transport drugs from the cell surface via receptor-mediated endocytosis and endosomal escape to nucleus. Hence their conjugation to Auger electron emitters, can cause severe cell killing, by nuclear localization. Herein we evaluate the use of MNT as a platform for targeted radiotherapy with (67)Ga. METHODS: EGF was the targeting ligand on the MNT, and NOTA was selected for its radiolabeling with (67)Ga. In the radiolabeling study we dealt with the precipitation of MNT (pI 5.7) at the labeling pH (4.5-5.5) of (67)Ga. Cellular and nuclei uptake of (67)Ga-NOTA-MNT by the A431 cell line was determined. Its specific cytotoxicity was compared to that of (67)Ga-EDTA, (67)Ga-NOTA-BSA and (67)Ga-NOTA-hEGF, in A431 and U87MGWTT, cell lines, by clonogenic assay. Dosimetry studies were also performed. RESULTS: (67)Ga-NOTA-MNT was produced with 90% yield and specific activity of 25.6mCi/mg. The in vitro kinetics revealed an increased uptake over 24h. 55% of the internalized radioactivity was detected in the nuclei at 1h. The cytotoxicity of (67)Ga-NOTA-MNT on A431 cell line was 17 and 385-fold higher when compared to non-specific (67)Ga-NOTA-BSA and (67)Ga-EDTA. While its cytotoxic potency was 13 and 72-fold higher when compared to (67)Ga-NOTA-hEGF in the A431 and the U87MGWTT cell lines, respectively, validating its nuclear localization. The absorbed dose, for 63% cell killing, was 8Gy, confirming the high specific index of (67)Ga. CONCLUSION: These results demonstrate the feasibility of using MNT as a platform for single cell kill targeted radiotherapy by Auger electron emitters.
Asunto(s)
Electrones , Receptores ErbB/metabolismo , Compuestos Heterocíclicos/uso terapéutico , Isotiocianatos/uso terapéutico , Péptidos/química , Péptidos/metabolismo , Radioterapia/métodos , Transporte Biológico , Línea Celular Tumoral , Núcleo Celular/metabolismo , Radioisótopos de Galio/uso terapéutico , Regulación Neoplásica de la Expresión Génica , Compuestos Heterocíclicos/química , Humanos , Isotiocianatos/química , Marcaje Isotópico , Transporte de Proteínas , RadiometríaRESUMEN
MicroRNAs (miRNAs) are non-coding endogenous RNAs that direct post-transcriptional regulation of gene expression by several mechanisms. Activity is primarily through binding to the 3' untranslated regions (UTRs) of messenger RNAs (mRNA) resulting in degradation and translation repression. Unlike other small-RNAs, miRNAs do not require perfect base pairing, and thus, can regulate a network of broad, yet specific, genes. Although we have only just begun to gain insights into the full range of biologic functions of miRNA, their involvement in the onset and progression of disease has generated significant interest for therapeutic development. Mounting evidence suggests that miRNA-based therapies, either restoring or repressing miRNAs expression and activity, hold great promise. However, despite the early promise and exciting potential, critical hurdles often involving delivery of miRNA-targeting agents remain to be overcome before transition to clinical applications. Limitations that may be overcome by delivery include, but are not limited to, poor in vivo stability, inappropriate biodistribution, disruption and saturation of endogenous RNA machinery, and untoward side effects. Both viral vectors and nonviral delivery systems can be developed to circumvent these challenges. Viral vectors are efficient delivery agents but toxicity and immunogenicity limit their clinical usage. Herein, we review the recent advances in the mechanisms and strategies of nonviral miRNA delivery systems and provide a perspective on the future of miRNA-based therapeutics.
Asunto(s)
Sistemas de Liberación de Medicamentos , Técnicas de Transferencia de Gen , MicroARNs/administración & dosificación , MicroARNs/uso terapéutico , Animales , Dendrímeros/química , Sistemas de Liberación de Medicamentos/métodos , Regulación de la Expresión Génica , Humanos , Ácido Láctico/química , Lípidos/química , MicroARNs/genética , MicroARNs/farmacocinética , Polietileneimina/química , Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico-Ácido Poliglicólico , Distribución TisularRESUMEN
BACKGROUND: Modular nanotransporters (MNT) are recombinant multifunctional polypeptides created to exploit a cascade of cellular processes, initiated with membrane receptor recognition to deliver selective short-range and highly cytotoxic therapeutics to the cell nucleus. This research was designed for in vivo concept testing for this drug delivery platform using two modular nanotransporters, one targeted to the α-melanocyte-stimulating hormone (αMSH) receptor overexpressed on melanoma cells and the other to the epidermal growth factor (EGF) receptor overexpressed on several cancers, including glioblastoma, and head-and-neck and breast carcinoma cells. METHODS: In vivo targeting of the modular nanotransporter was determined by immuno-fluorescence confocal laser scanning microscopy and by accumulation of (125)I-labeled modular nanotransporters. The in vivo therapeutic effects of the modular nanotransporters were assessed by photodynamic therapy studies, given that the cytotoxicity of photosensitizers is critically dependent on their delivery to the cell nucleus. RESULTS: Immunohistochemical analyses of tumor and neighboring normal tissues of mice injected with multifunctional nanotransporters demonstrated preferential uptake in tumor tissue, particularly in cell nuclei. With (125)I-labeled MNT{αMSH}, optimal tumor:muscle and tumor:skin ratios of 8:1 and 9.8:1, respectively, were observed 3 hours after injection in B16-F1 melanoma-bearing mice. Treatment with bacteriochlorin p-MNT{αMSH} yielded 89%-98% tumor growth inhibition and a two-fold increase in survival for mice with B16-F1 and Cloudman S91 melanomas. Likewise, treatment of A431 human epidermoid carcinoma-bearing mice with chlorin e(6)- MNT{EGF} resulted in 94% tumor growth inhibition compared with free chlorin e(6), with 75% of animals surviving at 3 months compared with 0% and 20% for untreated and free chlorin e(6)-treated groups, respectively. CONCLUSION: The multifunctional nanotransporter approach provides a new in vivo functional platform for drug development that could, in principle, be applicable to any combination of cell surface receptor and agent (photosensitizers, oligonucleotides, radionuclides) requiring nuclear delivery to achieve maximum effectiveness.