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Antibody-targeted lipid nanoparticles (Ab-LNPs) are rapidly gaining traction as multifaceted platforms in precision medicine, adept at delivering a diverse array of therapeutic agents, including nucleic acids and small molecules. This review provides an incisive overview of the latest developments in the field of Ab-LNP technology, with a special emphasis on pivotal design aspects such as antibody engineering, bioconjugation strategies, and advanced formulation techniques. Furthermore, it addresses critical chemistry, manufacturing, and controls (CMC) considerations and thoroughly examines the in vivo dynamics of Ab-LNPs, underscoring their promising potential for clinical application. By seamlessly blending scientific advancements with practical industrial perspectives, this review casts a spotlight on the burgeoning role of Ab-LNPs as an innovative and potent tool in the realm of targeted drug delivery.
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Lipossomos , Nanopartículas , Sistemas de Liberação de Medicamentos , Nanopartículas/química , Anticorpos , RNA Interferente PequenoRESUMO
1,2-Distigmasterylhemisuccinoyl-sn-glycero-3-phosphocholine (DSHemsPC) is a new lipid in which two molecules of stigmasterol (an inexpensive plant sterol) are covalently linked via a succinic acid to glycerophosphocholine. Our previous study revealed that liposome (Lip)-intercalated amphotericin B (AMB) prepared from DSHemsPC (DSHemsPC-AMB-Lip) possesses excellent colloidal properties and in vitro antifungal and antileishmanial activities similar to those of the liposomal AMB preparation AmBisome. The aim of this study was to determine the biodistribution and evaluate the antileishmanial effects of DSHemsPC-AMB-Lip in Leishmania major-infected BALB/c mice. The serum profile and tissue concentrations of AMB were similar in DSHemsPC-AMB-Lip- and AmBisome-treated mice after intravenous (i.v.) injection. Multiple i.v. doses of the micellar formulation of AMB (Fungizone; 1 mg/kg of body weight), DSHemsPC-AMB-Lip (5 mg/kg), and AmBisome (5 mg/kg) were used in L. major-infected BALB/c mouse models of early and established lesions. In a model of the early lesions of cutaneous leishmaniasis (CL), the results indicated that the level of footpad inflammation was significantly (P < 0.001) lower in mice treated with DSHemsPC-AMB-Lip and AmBisome than mice treated with empty liposomes or 5% dextrose. The splenic and footpad parasite load was also significantly (P < 0.001) lower in these groups of mice than in control mice that received 5% DW or free liposome. The in vivo activity of DSHemsPC-AMB-Lip was comparable to that of AmBisome, and both provided improved results compared to those achieved with Fungizone at the designated doses. The results suggest that systemic DSHemsPC-AMB-Lip administration may be useful for the treatment of leishmaniasis, and because it costs less to produce DSHemsPC-AMB-Lip than AmBisome, DSHemsPC-AMB-Lip merits further investigation.
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Anfotericina B/farmacocinética , Anfotericina B/uso terapêutico , Antiprotozoários/uso terapêutico , Leishmania major/efeitos dos fármacos , Leishmaniose Cutânea/tratamento farmacológico , Fosfatidilcolinas/farmacocinética , Fosfatidilcolinas/uso terapêutico , Estigmasterol/análogos & derivados , Animais , Antiprotozoários/farmacocinética , Modelos Animais de Doenças , Vias de Administração de Medicamentos , Portadores de Fármacos/uso terapêutico , Feminino , Leishmania major/patogenicidade , Leishmaniose Cutânea/parasitologia , Lipossomos/uso terapêutico , Camundongos , Camundongos Endogâmicos BALB C , Carga Parasitária , Baço/parasitologia , Estigmasterol/farmacocinética , Estigmasterol/uso terapêuticoRESUMO
We report here the first exploration of the nature of the hydrophobic region of bilayer membranes formed from sterol-modified phospholipids [Huang, Z.; Szoka, F. C., Sterol-Modified Phospholipids: Cholesterol and Phospholipid Chimeras with Improved Biomembrane Properties. J. Am. Chem. Soc. 2008, 130 (46), 15702-15712] & [Ding, J.; Starling, A. P.; East, J. M.; Lee, A. G., Binding Sites for Cholesterol on Ca(2+)-ATPase Studied by Using a Cholesterol-Containing Phospholipid. Biochemistry 1994, 33 (16), 4974-4979]. Using 2H NMR spectroscopy, we present our results for the phase behavior and acyl chain ordering of multilamellar vesicles (MLVs) of a sterol-modified phospholipid, 1-cholesterylhemisuccinoyl-2-palmitoyl(d31)-sn-glycero-3-phosphocholine (hereafter referred to as CholPPC-d31). We compared our results with the conformational order induced by cholesterol at various concentrations in 1-palmitoyl,2-palmitoyl(d31)-sn-glycero-3-phosphocholine (DPPC-d31)/cholesterol membranes. On the basis of the existing literature [Foglia, F.; Barlow, D. J.; Szoka, F. C.; Huang, Z.; Rogers, S. E.; Lawrence, M. J., Structural Studies of the Monolayers and Bilayers Formed by a Novel Cholesterol-Phospholipid Chimera. Langmuir 2011, 27 (13), 8275-8281], we expected to find that the deuterated palmitoyl chain in CholPPC-d31 membranes had an order parameter profile similar to the deuterated palmitoyl chain of sn-2 labeled DPPC-d31 in MLVs of a mixture of DPPC-d31 with 40 mol % unconstrained cholesterol. Our data indicate that the ordering ability of cholesterol in CholPPC is significantly reduced compared to free cholesterol in DPPC. This result emphasizes that cholesterol molecules must be free to move in the bilayers to reach their maximum ordering ability. In other words, when compared to unconstrained cholesterol, the constrained cholesterol moiety in CholPPC causes nonoptimal chain packing.
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Colesterol/química , Deutério , Bicamadas Lipídicas , Espectroscopia de Ressonância Magnética , FosfolipídeosRESUMO
This study reported oleogel-based emulsions (OGEs, W/O) stabilized by carnauba wax. The effects of different external factors (heating temperature, crystallization temperature, and shear application during crystallization) on the microstructure and linear/nonlinear rheological properties of OGEs were investigated. Microstructural observation suggested that the OGEs had a uniform droplet distribution, and the carnauba wax crystals trapped oil in the continuous phase. The gelatinized oil phase allowed the OGEs to have a solid appearance and typical yielding behavior. The small amplitude oscillation shear analysis showed that lower heating temperature, higher crystallization temperature, and suitable shear application resulted in a stronger, more stable, and tighter packed network structure and better resistance to deformation of the OGEs. For nonlinear behavior, the elastic dominant behavior of OGEs transformed into viscous dominant behavior at large strain amplitudes, accompanied by more energy dissipation, strain stiffening, and a transition from shear thickening to shear thinning.
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Temperatura Alta , Compostos Orgânicos , Emulsões/química , Compostos Orgânicos/química , Temperatura , ReologiaRESUMO
Oleogel-structured emulsion (OGE) is an emulsion in which the presence of oleogelator in the oil phase causes the oil phase to change from a liquid to a solid state. OGEs are increasingly being explored for their better stability, delivery efficiency, and adjustable rheological properties compared with conventional emulsions. However, many oleogelators are relatively unstable in the presence of water, and a good understanding of the relationship between emulsification and oleogelator as well as the synergistic or antagonistic effects on emulsion formation, structure, and functionality is required. Hence, the question arises whether different kinds of oleogelators are feasible in the preparation of OGEs. This review aims to assess the relationship between the oleogelator and emulsifiers or aqueous phases. Moreover, the fabrication methods, physicochemical properties, and applications are reviewed. It can be concluded that the food industry may have a great revolution with the development of OGEs.
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Emulsificantes , Água , Emulsões , ReologiaRESUMO
This study aimed to explore the impacts of gallic acid (GA)/protocatechuic acid (PA) on the structural and functional characteristics of whey proteins (WP) through covalent binding. To this purpose, the covalent complexes of WP-PA and WP-GA at different concentration gradients were prepared by the alkaline method. SDS-PAGE indicated that PA/GA was cross-linked by covalent bonds. The decreased contents of free amino and sulfhydryl groups suggested that WP formed covalent bonds with PA/GA by amino and sulfhydryl groups, and the structure of WP became slightly looser after covalent modification by PA/GA. When the concentration of GA was added up to 10 mM, the structure of WP was slightly loosened with a reduction of α-helix content by 2.3% and an increase in random coil content by 3.0%. The emulsion stability index of WP increased by 14.9 min after interaction with GA. Moreover, the binding of WP and 2-10 mM PA/GA increased the denaturation temperature by 1.95 to 19.87 °C, indicating the improved thermal stability of the PA/GA-WP covalent complex. Additionally, the antioxidant capacity of WP was increased with increasing GA/PA concentration. This work may offer worthful information for enhancing the functional properties of WP and the application of the PA/GA-WP covalent complexes in food emulsifiers.
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Ácido Gálico , Hidroxibenzoatos , Proteínas do Soro do Leite , EmulsõesRESUMO
Therapeutics at or close to the nanoscale, such as liposomal irinotecan, offer significant promise for the treatment of solid tumors. Their potential advantage over the unencapsulated or free form of the drug is due in part to their altered biodistribution. For slow and sustained release, significant optimization of formulation is needed to achieve the required level of stability and allow long-term storage of the drug product. Gradient-based liposomal formulation of camptothecins such as irinotecan poses unique challenges owing to the camptothecin- and acid-catalyzed hydrolysis of phospholipid esters in the inner monolayer of the liposomal membrane. We demonstrated that a narrow set of conditions related to the external pH, temperature, intraliposomal concentration, identity of the drug-trapping agent, physical form of the drug inside the liposomes, and final drug load have a marked impact on the stability of the liposome phospholipid membrane. The physical form of the drug inside the liposome was shown to be an insoluble gel with an irinotecan-to-sulfate ratio approximating 1:1, reducing the potential for irinotecan-catalyzed phospholipid hydrolysis in the internal phospholipid monolayer. As a result of this work, a stable and active liposome formulation has been developed that maintains phospholipid chemical stability following long-term storage at 2-8°C.
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Lipossomos , Fosfolipídeos , Irinotecano , Estabilidade de Medicamentos , Distribuição Tecidual , Camptotecina , CatáliseRESUMO
The natural active components derived from plants have attracted widespread attention due to their abundant species and source advantages [...].
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The utilization of starch in the food and medical industry can be facilitated by using new non-chemical methods to make starch the only wall material to encapsulate microcapsules. In this study, soluble small molecular fraction obtained from corn starch by gelatinization and centrifugation methods and commercial soluble starch were used independently to encapsulate oil under the condition that wall material and core material were 2:1. Molecular weight of these starch fractions was measured firstly. The peak molecular weight of the soluble small molecular fraction of corn starch and commercial soluble starch was 3.537 × 105 Da and 2.720 × 104 Da, respectively. Basic physicochemical characteristics and application characteristics of the microcapsules were then characterized and compared. The soluble small molecular fraction of corn starch encapsulated microcapsule and the commercial soluble starch encapsulated microcapsule had high encapsulation efficiency (higher than 88%), high boiling water solubility (higher than 74%), high rehydration stability (higher than 2 h). Most importantly, the encapsulated oil of these microcapsules could be slowly released under the action of α-amylase and amyloglucosidase. Overall, both the soluble small molecular fraction of corn starch and commercial soluble starch could be used as microcapsule wall materials and might have great application potential in food and medicine.
Assuntos
CápsulasRESUMO
W/O/W emulsions were easily prepared by oleogelation of the oil phase using rice bran wax (RBX) and their microstructure, stability, rheology and protection of proanthocyanidins and ß-carotene were investigated. Formation of the W/O/W emulsion was confirmed using confocal laser scanning microscopy and staining of the inner aqueous phase by tartrazine. The average particle size and viscosity of the emulsion increased as the RBX concentration increased. Moreover, RBX increased the stability of the emulsion and the emulsion was the most stable when the RBX concentration was 8.0% or 10.0%. On the other hand, the W/O/W emulsions were used to simultaneously encapsulate proanthocyanidins and ß-carotene. Specifically, proanthocyanidins and ß-carotene in RBX-containing emulsions were more stable and had higher bioaccessibility than in the emulsion without RBX. Besides, both their chemical stability and bioaccessibility reached the maximum value when the RBX concentration was 8.0% or 10.0%. In summary, the optimal RBX concentration was 8.0%.
Assuntos
Emulsões/química , Proantocianidinas/química , Óleo de Farelo de Arroz/química , beta Caroteno/química , Óleo de Milho/química , Armazenamento de Alimentos , Tamanho da Partícula , Proantocianidinas/farmacocinética , Reologia , Viscosidade , Água/química , beta Caroteno/farmacocinéticaRESUMO
Current therapeutic treatments improving the impaired transportation of oxygen in acute respiratory distress syndrome (ARDS) have been found to be relevant and beneficial for the therapeutic treatment of COVID-19 patients suffering from severe respiratory complications. Hence, we report the preclinical and the preliminary results of the Phase I/II clinical trial of LEAF-4L6715, a liposomal nanocarrier encapsulating the kosmotropic agent trans-crocetin (TC), which, once injected, enhance the oxygenation of vascular tissue and therefore has the potential to improve the clinical outcomes of ARDS and COVID-19 in severely impacted patients. We demonstrated that the liposomal formulation enabled to increase from 30 min to 48 h the reoxygenation properties of free TCs in vitro in endothelial cells, but also to improve the half-life of TC by 6-fold in healthy mice. Furthermore, we identified 25 mg/kg as the maximum tolerated dose in mice. This determined concentration led to the validation of the therapeutic efficacy of LEAF-4 L6715 in a sepsis mouse model. Finally, we report the preliminary outcomes of an open-label multicenter Phase I/II clinical trial (EudraCT 2020-001393-30; NCT04378920), which was aimed to define the appropriate schedule and dosage of LEAF-4L6715 and to confirm its tolerability profile and preliminary clinical activity in COVID-19 patients treated in intensive care unit.
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COVID-19 , Síndrome do Desconforto Respiratório , Animais , Carotenoides , Células Endoteliais , Humanos , Camundongos , Respiração Artificial , Síndrome do Desconforto Respiratório/tratamento farmacológico , SARS-CoV-2 , Vitamina A/análogos & derivadosRESUMO
Metal chelation-ligand interactions, such as occur between nitrilotriacetic acid (NTA)-nickel and multihistidines, enable the noncovalent attachment of histidine-modified proteins to liposomes and other particles. We compared three lipids: a mono-NTA lipid (ca. 10 microM affinity) and two tris-NTA lipid derivatives (ca. 3 nM and 0.2 nM affinity) in their ability to retain two different his(6)-containing proteins on NTA-liposomes in the presence of serum or plasma and after intravenous injection in mice. At nanomolar affinities, the off-rate of a his(6)-ligand is sufficiently long so that his(6)-proteins attached to particle surfaces will remain with the particle for hours; thus, we hypothesized that the increased his(6) affinity of multivalent NTA-modified liposomes would retain his(6)-proteins longer both in vitro and in vivo. For each of the three lipids, we found a robust association and complete activity retention of two his(6)-modified proteins: a far red-fluorescent protein, monomeric Katushka (mKate), and a prodrug-converting enzyme, yeast cytosine deaminase (yCD). Proteins associated more tightly in vitro with tris-NTA liposomes than with mono-NTA liposomes in the presence of refiltered fetal calf serum and mouse plasma. Free yCD exchanged with previously associated mKate for tris-NTA binding sites on the liposome surface. This exchange was due to the exchange of the proteins for NTA occupancy and not due to the exchange of tris-NTA lipid out of the liposome. The amount of yCD on the surface was similar if the proteins were co-associated or if mKate was pre-associated. This exchange confirms that NTA associated proteins are in a dynamic state and can exchange with multihistidine proteins in the biological milieu. There was no difference in circulation time of the protein when it was intravenously administered by itself or attached to any of the NTA-modified liposomes because in vivo the protein was rapidly released from the NTA liposomes. Upon recovery from blood, liposomes containing tris-NTA accumulated a different plasma protein profile than control liposomes, suggesting that Ni-NTA specifically interacts with some plasma proteins. The reason for the rapid protein dissociation from the liposome in vivo is not clear; it could be due to displacement by endogenous histidine-containing proteins or to natural chelators that remove nickel from the NTA. Regardless of the cause, improvements in chelator or ligand design are needed before metal chelation will be capable of retaining histidine-modified proteins on NTA liposomes after in vivo administration.
Assuntos
Histidina/química , Lipídeos/química , Lipossomos/química , Ácido Nitrilotriacético/análogos & derivados , Compostos Organometálicos/química , Proteínas/metabolismo , Animais , Meia-Vida , Histidina/metabolismo , Ligantes , Lipossomos/administração & dosagem , Lipossomos/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Ácido Nitrilotriacético/química , Ácido Nitrilotriacético/metabolismo , Compostos Organometálicos/metabolismo , Ligação Proteica , Proteínas/químicaRESUMO
Waxy rice starch was hydrolyzed by ß-amylase and then modified by octenyl succinic anhydride (OSA) to prepare octenyl succinate ß-limit dextrin (OSA ß-limit dextrin). Subsequently, structure, digestibility and emulsifying capacity of OSA ß-limit dextrin were measured. The FT-IR analysis confirmed formation of OSA ß-limit dextrin. When the addition of OSA was 1.0%, 2.0% and 3.0%, the DS of OSA ß-limit dextrin was 0.006, 0.011 and 0.019, respectively. The molecular weight, resistant starch content and emulsifying capacity of OSA ß-limit dextrin was higher than ß-limit dextrin and increased with the increasing of DS. In addition, the solubility of OSA ß-limit dextrin reached 100%. The emulsion prepared by OSA ß-limit dextrin was stable at pH 3.0-7.0 and became unstable at pH 8.0-11.0 or after addition of NaCl. These results suggested that OSA ß-limit dextrin was safe soluble resistant dextrin with emulsifying ability, which might be used in functional food and beverage products.
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To improve stability of peanut butter, rice bran wax (RBX) was added into peanut butter as a stabilizer by formation of organogel. Effects of addition of RBX, heating temperature, and cooling temperature on stabilization effect of peanut butter by RBX were investigated. The optimum conditions were as follow: addition of RBX at 4.0 wt%; heating temperature at 85 °C, and cooling temperature at 20 °C. Under the optimum conditions, the oil loss of peanut butter decreased from 12.19% to 4.04%, and the adhesiveness of peanut butter increased from 23.5 to 165.9 g·s. After storage for 25 weeks, the acid value (AV) of peanut butter prepared under the optimum conditions increased from 0.65 to 0.80 mg/g, and the peroxide value (PV) increased from 0.116 to 0.127 meq/kg. However, the AV of natural peanut butter increased to 1.73 mg/g, and PV increased to 0.178 meq/kg. The confocal laser scanning microscope images showed that the cooling temperature significantly affected crystallization of RBX and distribution of solid particles. When RBX formed needle-like crystals and peanut solid particles were evenly distributed in the oil phase, stable peanut butter was obtained. These results suggested that the RBX was an effective stabilizer for peanut butter. PRACTICAL APPLICATION: Oil separation often occurred to peanut butter during storage, which reduced the sensory quality of peanut butter and shortened its shelf life. This study stabilized peanut butter by addition of RBX based on the formation of organogel, which was of great practical significance to improve the shelf life of peanut butter.
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Arachis/química , Aditivos Alimentares/análise , Manipulação de Alimentos/métodos , Oryza/química , Extratos Vegetais/química , Ceras/análise , Cristalização , TemperaturaRESUMO
Annealing behavior of amylose and amylopectin was unclear. In this work, high purity amylose and amylopectin were extracted from rice starch, and structural properties of the retrograded rice starch, amylose, and amylopectin before and after annealing treatment were explored. It was found that the purity of the amylose and amylopectin was 95.64%⯱â¯2.69% and 94.98%⯱â¯0.97%, respectively. Their molecular weight was (2.93⯱â¯0.21)â¯×â¯106 Da and (5.90⯱â¯0.13)â¯×â¯107 Da, respectively. Besides, the relative crystallinities and ratios of 1047â¯cm-1/1022â¯cm-1 of the retrograded rice starch and amylose were significantly increased by annealing treatment, while that of retrograded amylopectin did not change. These results clarified that amylose was more sensitive to annealing treatment than amylopectin, and amylose was more responsible for annealing of starch than amylopectin. The findings contributed to a better understanding of the annealing behavior of starch.
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Amilopectina/química , Amilopectina/isolamento & purificação , Amilose/química , Amilose/isolamento & purificação , Oryza/química , Amido/química , Fracionamento Químico , Fenômenos Químicos , Peso Molecular , Análise EspectralRESUMO
Combinations of chemotherapy with immunotherapy have seen recent clinical success, including two approvals of anti-PD-1/L1 agents in combination with taxane-based chemotherapy in non-small cell lung cancer and triple-negative breast cancer. Here, we present a study on the combination activity and mechanistic rationale of a novel EphA2-targeted liposomal taxane (EphA2-ILs-DTXp) and anti-PD-1. This combination was highly active in mouse syngeneic tumor models, with complete responses observed in 3 of 5 models. In the EMT-6 tumor model, combination of EphA2-ILs-DTXp with anti-PD-1 resulted in a 60% complete response rate, with durable responses that were resistant to rechallenge. These responses were not observed in the absence of CD8+ T cells. Characterization of the immune infiltrates in EMT-6 tumors reveals increased CD8+ T cells, increased CD8+ IFNγ+ CTLs, and an increased CD8/regulatory T-cell (Treg) ratio. These immunomodulatory effects were not observed in mice treated with a combination of docetaxel and anti-PD-1. Pharmacokinetic analysis revealed that the AUC of docetaxel was increased 15 times, from 52.1 to 785 ng/mL/hour, when delivered by EphA2-ILs-DTXp. A dose reduction study of EphA2-ILs-DTXp showed a dose-response relationship for both tumor growth inhibition and the CD8/Treg ratio. Our data indicate that synergism between docetaxel and anti-PD-1 is achievable with nanoliposomal delivery.
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Hidrocarbonetos Aromáticos com Pontes/uso terapêutico , Neoplasias/tratamento farmacológico , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor EphA2/metabolismo , Taxoides/uso terapêutico , Animais , Hidrocarbonetos Aromáticos com Pontes/farmacologia , Modelos Animais de Doenças , Feminino , Humanos , Camundongos , Neoplasias/patologia , Taxoides/farmacologiaRESUMO
Ephrin receptor A2 (EphA2) is a member of the Ephrin/Eph receptor cell-to-cell signaling family of molecules, and it plays a key role in cell proliferation, differentiation, and migration. EphA2 is overexpressed in a broad range of cancers, and its expression is in many cases associated with poor prognosis. We recently developed a novel EphA2-targeting antibody-directed nanotherapeutic encapsulating a labile prodrug of docetaxel (EphA2-ILs-DTXp) for the treatment of EphA2-expressing malignancies. Here, we characterized the expression of EphA2 in bladder cancer using immunohistochemistry in 177 human bladder cancer samples and determined the preclinical efficacy of EphA2-ILs-DTXp in four EphA2-positive patient-derived xenograft (PDX) models of the disease, either as a monotherapy, or in combination with gemcitabine. EphA2 expression was detected in 80-100% of bladder cancer samples and correlated with shorter patient survival. EphA2 was found to be expressed in tumor cells and/or tumor-associated blood vessels in both primary and metastatic lesions with a concordance rate of approximately 90%. The EphA2-targeted antibody-directed nanotherapeutic EphA2-ILs-DTXp controlled tumor growth, mediated greater regression, and was more active than free docetaxel at equitoxic dosing in all four EphA2-positive bladder cancer PDX models. Combination of EphA2-ILs-DTXp and gemcitabine in one PDX model led to improved tumor growth control compared to monotherapies or the combination of free docetaxel and gemcitabine. These data demonstrating the prevalence of EphA2 in bladder cancers and efficacy of EphA2-ILs-DTXp in PDX models support the clinical exploration of EphA2 targeting in bladder cancer.
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Retinoic acid (RA), the bioactive metabolite of retinol, is essential for robust humoral immunity in animals and humans. Recent interest in RA as a vaccine adjuvant has been encouraged by reports showing cooperative enhancement of antibody responses to tetanus toxin in rodents by all-trans RA (ATRA) and a Toll-like receptor-3 agonist. We hypothesized that RA would augment the antibody response to a co-delivered lipopeptide immunogen derived from the membrane proximal region (MPR) of HIV-1 gp41. The MPR is weakly immunogenic and could benefit from potent new humoral adjuvants. When co-formulated in liposomes and administered to BALB/C mice, ATRA alone did not elicit serum anti-peptide antibodies to an MPR-derived lipopeptide. However, addition of ATRA, but not 13-cis RA, to a liposomal formulation containing the Toll-like receptor-4 agonist monophosphoryl lipid A resulted in a fourfold enhancement of serum anti-peptide IgG titers as compared with a formulation containing lipid A alone (P=0.00039). The difference did not arise from biophysical changes in the liposome formulation, including vesicle size, vesicle charge and liposome association of antigen. Thus, ATRA warrants further study as a vaccine adjuvant.
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Adjuvantes Imunológicos/administração & dosagem , Anticorpos Antivirais/imunologia , Proteína gp41 do Envelope de HIV/imunologia , HIV-1/imunologia , Lipídeo A/administração & dosagem , Lipopeptídeos/imunologia , Tretinoína/administração & dosagem , Animais , Sinergismo Farmacológico , Feminino , Lipídeo A/imunologia , Lipossomos , Camundongos , Camundongos Endogâmicos BALB C , Estrutura Molecular , Tretinoína/químicaRESUMO
Nitrilotriacetic acid (NTA) has moderate affinity (10 µM) for hexahistidine (His6) and is widely used in the purification of His6-tagged proteins. The affinity can be increased significantly (10 nM) through multivalency such as using a tris-NTA. We show that the binding affinity of tris-NTA is dependent on the flexibility and length of the spacer between the mono-NTA and the scaffold: the shorter the spacer, the higher the affinity. A series of biotinylated tris-NTA having different spacers were synthesized and used to prepare tris-NTA sensor chips for surface plasmon resonance measurement of binding affinity. Subnanomolar affinity can be achieved with a short spacer. The new high-affinity tris-NTA enables the formation of stable complexes with hexahistidine containing molecules and provides a convenient method to noncovalently attach proteins to various surfaces.
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Histidina/química , Nanoestruturas/química , Ácido Nitrilotriacético/química , Oligopeptídeos/química , Biotina/química , Estrutura Molecular , Fragmentos de Peptídeos/síntese química , Fragmentos de Peptídeos/química , Estereoisomerismo , Ressonância de Plasmônio de SuperfícieRESUMO
We evaluated gene transfer using PEGylated bioresponsive nanolipid particles (NLPs) containing plasmid DNA administered by convection-enhanced delivery (CED) into orthotopically implanted U87-MG tumors in rat brain. We hypothesized that attachment of the human immunodeficiency virus trans-acting transcriptional activator peptide (TATp) to pH-sensitive, reduction-sensitive NLPs would increase gene transfer. TATp was attached either directly to a phospholipid (TATp-lipid) or via a 2-kd polyethylene glycol (PEG) to a lipid (TATp-PEG-lipid). Incorporation of 0.3 mol% TATp-PEG into pH-sensitive NLPs improved transfection 100,000-fold compared to NLPs in culture. In the brain or implanted tumors, the TATp-PEG restricted NLP convection to regions adjacent to the infusion catheter. Gene transfer in the brain from TATp-PEG NLPs, measured by green fluorescent protein (GFP) expression, was substantially greater than from NLPs adjacent to the catheter. Gene transfer using TATp-PEG NLPs, measured by luciferase expression, was 8-12-fold greater than from a 1,2-dioleoyl-3-trimethylammonium-propane/cholesterol cationic lipoplex but 13-27-fold less than from the NLPs. Brain luciferase expression was localized in perivascular macrophages. Thus a cationic ligand, such as the TATp-PEG-lipid, can dramatically increase gene expression in culture, in the normal brain, and in implanted tumors; however, restriction of NLP distribution to the vicinity of the infusion catheter reduces the absolute level of gene transfer.