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1.
Nano Lett ; 2024 Oct 07.
Artigo em Inglês | MEDLINE | ID: mdl-39373269

RESUMO

The delivery of mRNA molecules to organs beyond the liver is valuable for therapeutic applications. Functionalized lipid nanoparticles (LNPs) using exogenous mechanisms can regulate in vivo mRNA expression profiles from hepatocytes to extrahepatic tissues but lead to process complexity and cost escalation. Here, we report that mRNA expression gradually shifts from the liver to the spleen in an ionizable lipid tail length-dependent manner. Remarkably, this simple chemical strategy held true even when different ionizable lipid head structures were employed. As a potential mechanism underlying this discovery, our data suggest that 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) is enriched on the surface of mRNA/LNPs with short-tail lipids. This feature limits their interaction with biological components, avoiding their rapid hepatic clearance. We also show that spleen-targeting LNPs loaded with SARS-CoV-2 receptor-binding domain (RBD) mRNA can efficiently induce immune responses and neutralize activity following intramuscular vaccination priming and boosting.

2.
Immunity ; 42(1): 123-32, 2015 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-25557055

RESUMO

Host innate recognition triggers key immune responses for viral elimination. The sensing mechanism of hepatitis B virus (HBV), a DNA virus, and the subsequent downstream signaling events remain to be fully clarified. Here we found that type III but not type I interferons are predominantly induced in human primary hepatocytes in response to HBV infection, through retinoic acid-inducible gene-I (RIG-I)-mediated sensing of the 5'-ε region of HBV pregenomic RNA. In addition, RIG-I could also counteract the interaction of HBV polymerase (P protein) with the 5'-ε region in an RNA-binding dependent manner, which consistently suppressed viral replication. Liposome-mediated delivery and vector-based expression of this ε region-derived RNA in liver abolished the HBV replication in human hepatocyte-chimeric mice. These findings identify an innate-recognition mechanism by which RIG-I dually functions as an HBV sensor activating innate signaling and to counteract viral polymerase in human hepatocytes.


Assuntos
Produtos do Gene pol/antagonistas & inibidores , Vírus da Hepatite B/fisiologia , Hepatite B Crônica/imunologia , Hepatócitos/fisiologia , Fígado/fisiologia , Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , RNA Viral/imunologia , Animais , Pré-Escolar , Feminino , Células Hep G2 , Hepatócitos/transplante , Hepatócitos/virologia , Humanos , Imunidade Inata , Interferons/metabolismo , Fígado/virologia , Proteínas de Membrana/imunologia , Camundongos , Camundongos SCID , Proteínas do Tecido Nervoso/imunologia , RNA Viral/genética , Receptores de Superfície Celular , Transgenes/genética , Quimeras de Transplante , Replicação Viral/genética
3.
Biol Pharm Bull ; 47(10): 1726-1733, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39462587

RESUMO

Energy metabolism is crucial for cell polarity and pathogenesis. Mitochondria, which are essential for maintaining energy homeostasis within cells, can be targeted by drug delivery to regulate energy metabolism. However, there is a lack of research comparing how mitochondria control energy metabolism in different cell types derived from the neural crest. Understanding the effects of berberine (BBR), a compound that acts on mitochondria, on energy metabolism in neural crest-derived cells is important. This study reports how MITO-Porter, a mitochondria-targeted liposome, affects neuroblasts (Neuro2a cells) and normal human epidermal melanocytes (NHEMs) when loaded with BBR. We found that treatment with MITO-Porter containing BBR reduced mitochondrial respiration in Neuro2a cells, while it caused a slight increase in NHEMs. Additionally, the treatment shifted the ATP production pathway in Neuro2a cells to rely more on glycolysis, while in NHEMs, there was a slight decrease in the reliance on glycolysis. We also observed a significant decrease in ATP production in Neuro2a cells, while NHEMs showed a tendency to increase ATP production. Importantly, on the basis of the results of the Premix WST-1 assay, the study found that BBR treatment was not toxic to either cell type. It is important to take note of the varied effects of BBR treatment on different cell types derived from the neural crest. These findings necessitate attention when utilizing NHEMs as a cell model in the development of therapeutic strategies for neurodegenerative diseases, including the use of BBR for metabolic control.


Assuntos
Trifosfato de Adenosina , Berberina , Mitocôndrias , Crista Neural , Berberina/farmacologia , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Crista Neural/citologia , Crista Neural/efeitos dos fármacos , Crista Neural/metabolismo , Humanos , Trifosfato de Adenosina/metabolismo , Animais , Melanócitos/efeitos dos fármacos , Melanócitos/metabolismo , Camundongos , Metabolismo Energético/efeitos dos fármacos , Lipossomos , Linhagem Celular , Linhagem Celular Tumoral , Glicólise/efeitos dos fármacos
4.
Handb Exp Pharmacol ; 284: 329-339, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-37017791

RESUMO

The approval of mRNA-containing lipid nanoparticles (LNPs) for use in a vaccine against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the clinical utility of RNA-loaded nanocapsules has stimulated a rapid acceleration in research in this area. The development of mRNA-containing LNP vaccines has been rapid, not only because of regulatory adjustments, but also to the advances made in nucleic acid delivery as the result of efforts by many basic researchers. RNA functions, not only in the nucleus and cytoplasm, but also in mitochondria, which have their own genomic apparatus. Mitochondrial diseases caused by mutations or defects in the mitochondrial genome, mitochondrial DNA (mtDNA) are intractable and are mainly treated symptomatically, but gene therapy as a fundamental treatment is expected to soon be a reality. To realize this therapy, a drug delivery system (DDS) that delivers nucleic acids including RNA to mitochondria is required, but efforts in this area have been limited compared to research targeting the nucleus and cytoplasm. This contribution provides an overview of mitochondria-targeted gene therapy strategies and discusses studies that have attempted to validate mitochondria-targeted RNA delivery therapies. We also present the results of 'RNA delivery to mitochondria' based on the use of our mitochondria-targeted DDS (MITO-Porter) that was developed in our laboratory.


Assuntos
Mitocôndrias , RNA , Humanos , Mitocôndrias/genética , Lipossomos , Sistemas de Liberação de Medicamentos , DNA Mitocondrial/genética , RNA Mensageiro
5.
Int J Mol Sci ; 25(8)2024 Apr 12.
Artigo em Inglês | MEDLINE | ID: mdl-38673875

RESUMO

Photodynamic therapy is expected to be a less invasive treatment, and strategies for targeting mitochondria, the main sources of singlet oxygen, are attracting attention to increase the efficacy of photodynamic therapy and reduce its side effects. To date, we have succeeded in encapsulating the photosensitizer rTPA into MITO-Porter (MP), a mitochondria-targeted Drug Delivery System (DDS), aimed at mitochondrial delivery of the photosensitizer while maintaining its activity. In this study, we report the results of our studies to alleviate rTPA aggregation in an effort to improve drug efficacy and assess the usefulness of modifying the rTPA side chain to improve the mitochondrial retention of MITO-Porter, which exhibits high therapeutic efficacy. Conventional rTPA with anionic side chains and two rTPA analogs with side chains that were converted to neutral or cationic side chains were encapsulated into MITO-Porter. Low-MP (MITO-Porter with Low Drug/Lipid) exhibited high drug efficacy for all three types of rTPA, and in Low-MP, charged rTPA-encapsulated MP exhibited high drug efficacy. The cellular uptake and mitochondrial translocation capacities were similar for all particles, suggesting that differences in aggregation rates during the incorporation of rTPA into MITO-Porter resulted in differences in drug efficacy.


Assuntos
Interações Hidrofóbicas e Hidrofílicas , Mitocôndrias , Fotoquimioterapia , Fármacos Fotossensibilizantes , Porfirinas , Fármacos Fotossensibilizantes/química , Fármacos Fotossensibilizantes/farmacologia , Mitocôndrias/metabolismo , Mitocôndrias/efeitos dos fármacos , Humanos , Fotoquimioterapia/métodos , Porfirinas/química , Porfirinas/farmacologia , Nanopartículas/química , Sistemas de Liberação de Medicamentos/métodos , Linhagem Celular Tumoral , Oxigênio Singlete/metabolismo , Oxigênio Singlete/química
6.
PLoS Pathog ; 17(5): e1009228, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33979382

RESUMO

Virus infection, such as hepatitis B virus (HBV), occasionally causes endoplasmic reticulum (ER) stress. The unfolded protein response (UPR) is counteractive machinery to ER stress, and the failure of UPR to cope with ER stress results in cell death. Mechanisms that regulate the balance between ER stress and UPR are poorly understood. Type 1 and type 2 interferons have been implicated in hepatic flares during chronic HBV infection. Here, we examined the interplay between ER stress, UPR, and IFNs using transgenic mice that express hepatitis B surface antigen (HBsAg) (HBs-Tg mice) and humanized-liver chimeric mice infected with HBV. IFNα causes severe and moderate liver injury in HBs-Tg mice and HBV infected chimeric mice, respectively. The degree of liver injury is directly correlated with HBsAg levels in the liver, and reduction of HBsAg in the transgenic mice alleviates IFNα mediated liver injury. Analyses of total gene expression and UPR biomarkers' protein expression in the liver revealed that UPR is induced in HBs-Tg mice and HBV infected chimeric mice, indicating that HBsAg accumulation causes ER stress. Notably, IFNα administration transiently suppressed UPR biomarkers before liver injury without affecting intrahepatic HBsAg levels. Furthermore, UPR upregulation by glucose-regulated protein 78 (GRP78) suppression or low dose tunicamycin alleviated IFNα mediated liver injury. These results suggest that IFNα induces ER stress-associated cell death by reducing UPR. IFNγ uses the same mechanism to exert cytotoxicity to HBsAg accumulating hepatocytes. Collectively, our data reveal a previously unknown mechanism of IFN-mediated cell death. This study also identifies UPR as a potential target for regulating ER stress-associated cell death.


Assuntos
Morte Celular , Antígenos de Superfície da Hepatite B/metabolismo , Hepatite B Crônica/complicações , Hepatócitos/patologia , Interferon-alfa/farmacologia , Falência Hepática Aguda/patologia , Resposta a Proteínas não Dobradas/efeitos dos fármacos , Animais , Chaperona BiP do Retículo Endoplasmático , Estresse do Retículo Endoplasmático , Antígenos de Superfície da Hepatite B/genética , Vírus da Hepatite B/isolamento & purificação , Hepatite B Crônica/patologia , Hepatite B Crônica/virologia , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Hepatócitos/virologia , Humanos , Falência Hepática Aguda/etiologia , Falência Hepática Aguda/metabolismo , Camundongos , Camundongos Transgênicos
7.
Biol Pharm Bull ; 46(12): 1648-1660, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-38044089

RESUMO

This review paper summarizes progress that has been made in the new field of "Controlled Intracellular Trafficking." This involves the development of new systems for delivering plasmid DNA (pDNA), small interfering RNA (siRNA), mRNA, proteins, their escape from endosomes, the mechanism for how they enter the nucleus, how they enter mithochondria and how materials subsequently function within a cell. In addition, strategies for delivering these materials to a selective tissue after intravenous administration was also intensively investigated not only to the liver but also to tumors, lungs, adipose tissue and the spleen. In 2020, a new mRNA vaccine was developed against coronavirus disease 2019 (COVID-19), where ionizable cationic lipids were used as a delivery system. Our strategy to identify an efficient ionizable cationic lipids (iCL) based on a lipid library as well as their applications concerning the delivery of siRNA/mRNA/pDNA is also described.


Assuntos
DNA , Nanomedicina , Distribuição Tecidual , RNA Interferente Pequeno , Lipídeos , RNA Mensageiro/metabolismo
8.
Microsc Microanal ; 29(3): 1228-1243, 2023 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-37749682

RESUMO

The prognosis of metastatic lung melanoma (MLM) has been reported to be poor. An increasing number of studies have reported the function of several immune cells in cancer regression. Although the function of mediastinal fat-associated lymphoid clusters (MFALCs) in the progression of inflammatory lung lesions has been previously reported, the association between MLM progression and MFALCs development has remained unexplored. Herein, we compared the microenvironmental changes in the lungs and MFALCs among phosphate-buffered saline (PBS) and cancer groups at early (1 week) and late (2 weeks) stages following the intravenous injection of B16-F10 melanoma cells into C57BL/6 mice. Except for lung CD4+ helper T-cells and Iba1+ macrophage populations of early stage, we observed a significant increase in the proliferating and immune cell (CD20+ B-lymphocytes, CD3+ T-lymphocytes, CD8+ cytotoxic T-cells, CD16+ natural killer (NK) cells populations, area of high endothelial venules, and lung lymphatic vessels in cancer groups at both the stages as compared with the PBS groups. Furthermore, a significant positive correlation was observed between immune cell populations in MFALCs and the lungs (B- and T-lymphocytes, and NK cells in both stages). Collectively, our findings suggest a promising cancer therapeutic strategy via targeting immune cells in MFALCs.


Assuntos
Neoplasias Pulmonares , Melanoma , Camundongos , Animais , Camundongos Endogâmicos C57BL , Mediastino , Pulmão
9.
Int J Mol Sci ; 24(2)2023 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-36674418

RESUMO

Delivering drugs to mitochondria, the main source of energy in neurons, can be a useful therapeutic strategy for the treatment of neurodegenerative diseases. Berberine (BBR), an isoquinoline alkaloid, acts on mitochondria and is involved in mechanisms associated with the normalization and regulation of intracellular metabolism. Therefore, BBR has attracted considerable interest as a possible therapeutic drug for neurodegenerative diseases. While BBR has been reported to act on mitochondria, there are few reports on the efficient delivery of BBR into mitochondria. This paper reports on the mitochondrial delivery of BBR using a lipid nanoparticle (LNP), a "MITO-Porter" that targets mitochondria, and its pharmacological action in Neuro2a cells, a model neuroblastoma. A MITO-Porter containing encapsulated BBR (MITO-Porter (BBR)) was prepared. Treatment with MITO-Porter (BBR) increased the amount of BBR that accumulated in mitochondria compared with a treatment with naked BBR. Treatment with MITO-Porter (BBR) resulted in increased ATP production in Neuro2a cells, which are important for maintaining life phenomena, compared with treatment with naked BBR. Treatment with MITO-Porter (BBR) also increased the level of expression of mitochondrial ubiquitin ligase (MITOL), which is involved in mitochondrial quality control. Our findings indicate that increasing the accumulation of BBR into mitochondria is important for inducing enhanced pharmacological actions. The use of this system has the potential for being important in terms of the regulation of the metabolic mechanism of mitochondria in nerve cells.


Assuntos
Berberina , Sistemas de Liberação de Medicamentos , Sistemas de Liberação de Medicamentos/métodos , Berberina/farmacologia , Berberina/metabolismo , Lipossomos/metabolismo , Mitocôndrias/metabolismo
10.
Cancer Sci ; 113(5): 1855-1867, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-35266253

RESUMO

Tumor blood vessels play important roles in tumor progression and metastasis. Targeting tumor endothelial cells (TECs) is one of the strategies for cancer therapy. We previously reported that biglycan, a small leucine-rich proteoglycan, is highly expressed in TECs. TECs utilize biglycan in an autocrine manner for migration and angiogenesis. Furthermore, TEC-derived biglycan stimulates tumor cell migration in a paracrine manner leading to tumor cell intravasation and metastasis. In this study, we explored the therapeutic effect of biglycan inhibition in the TECs of renal cell carcinoma using an in vivo siRNA delivery system known as a multifunctional envelope-type nanodevice (MEND), which contains a unique pH-sensitive cationic lipid. To specifically deliver MEND into TECs, we incorporated cyclo(Arg-Gly-Asp-D-Phe-Lys) (cRGD) into MEND because αV ß3 integrin, a receptor for cRGD, is selective and highly expressed in TECs. We developed RGD-MEND-encapsulating siRNA against biglycan. First, we confirmed that MEND was delivered into OS-RC-2 tumor-derived TECs and induced in vitro RNAi-mediated gene silencing. MEND was then injected intravenously into OS-RC-2 tumor-bearing mice. Flow cytometry analysis demonstrated that MEND was specifically delivered into TECs. Quantitative RT-PCR indicated that biglycan was knocked down by biglycan siRNA-containing MEND. Finally, we analyzed the therapeutic effect of biglycan silencing by MEND in TECs. Tumor growth was inhibited by biglycan siRNA-containing MEND. Tumor microenvironmental factors such as fibrosis were also normalized using biglycan inhibition in TECs. Biglycan in TECs can be a novel target for cancer treatment.


Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Inibidores da Angiogênese , Animais , Biglicano/genética , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/terapia , Células Endoteliais , Humanos , Neoplasias Renais/genética , Lipossomos , Camundongos , RNA Interferente Pequeno/genética
11.
Biochem Biophys Res Commun ; 599: 24-30, 2022 04 09.
Artigo em Inglês | MEDLINE | ID: mdl-35168060

RESUMO

Inflammatory bowel disease (IBD) is a chronic inflammatory disease in the colon characterized by excessive activation of T cells. Glycosphingolipids (GSLs) are composed of lipid rafts in cellular membranes, and their content is linked to immune cell function. In the present study, we investigated the involvement of GSLs in IBD. Microarray data showed that in IBD patients, the expression of only UDP-glucose ceramide glucosyltransferase (UGCG) decreased among the GSLs synthases. Ad libitum access to dextran sulfate sodium (DSS) resulted in decreased UGCG and glucosylceramide (GlcCer) content in mesenteric lymph nodes and T cells from the spleen. Furthermore, the knockdown of Ugcg in T cells exacerbated the pathogenesis of colitis, which was accompanied by a decrease in Treg levels. Treatment with GlcCer nanoparticles prevented DSS-induced colitis. These results suggested that GlcCer in T cells is involved in the pathogenesis of IBD. Furthermore, GlcCer nanoparticles are a potential efficacious therapeutic target for IBD patients.


Assuntos
Glucosilceramidas/metabolismo , Glucosiltransferases/metabolismo , Doenças Inflamatórias Intestinais/patologia , Linfócitos T/metabolismo , Animais , Colite/induzido quimicamente , Colite/tratamento farmacológico , Colite/patologia , Sulfato de Dextrana/toxicidade , Modelos Animais de Doenças , Glucosilceramidas/administração & dosagem , Glucosilceramidas/genética , Glucosiltransferases/genética , Humanos , Doenças Inflamatórias Intestinais/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Nanopartículas/administração & dosagem , Nanopartículas/química , Linfócitos T/patologia
12.
Int J Mol Sci ; 23(12)2022 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-35743104

RESUMO

Despite recent advancements in therapeutic options for disorders of the central nervous system (CNS), the lack of an efficient drug-delivery system (DDS) hampers their clinical application. We hypothesized that liposomes could be optimized for retrograde transport in axons as a DDS from peripheral tissues to the spinal cord and dorsal root ganglia (DRGs). Three types of liposomes consisting of DSPC, DSPC/POPC, or POPC in combination with cholesterol (Chol) and polyethylene glycol (PEG) lipid were administered to sciatic nerves or the tibialis anterior muscle of mature rats. Liposomes in cell bodies were detected with infrared fluorescence of DiD conjugated to liposomes. Three days later, all nerve-administered liposomes were retrogradely transported to the spinal cord and DRGs, whereas only muscle-administered liposomes consisting of DSPC reached the spinal cord and DRGs. Modification with Cholera toxin B subunit improved the transport efficiency of liposomes to the spinal cord and DRGs from 4.5% to 17.3% and from 3.9% to 14.3% via nerve administration, and from 2.6% to 4.8% and from 2.3% to 4.1% via muscle administration, respectively. Modification with octa-arginine (R8) improved the transport efficiency via nerve administration but abolished the transport capability via muscle administration. These findings provide the initial data for the development of a novel DDS targeting the spinal cord and DRGs via peripheral administration.


Assuntos
Transporte Axonal , Gânglios Espinais , Animais , Grupos Diagnósticos Relacionados , Lipossomos , Fosfolipídeos , Ratos , Medula Espinal
13.
Mol Pharm ; 18(3): 878-888, 2021 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-33492961

RESUMO

This study describes the development of lipid nanoparticles (LNPs) for the efficient and selective delivery of plasmid DNA (pDNA) to the lungs. The GALA peptide was used as a ligand to target the lung endothelium and as an endosomal escape device. Transfection activity in the lungs was significantly improved when pDNA was encapsulated in double-coated LNPs. The inner coat was composed of dioleoylphsophoethanolamine and a stearylated octaarginine (STR-R8) peptide, while the outer coat was largely a cationic lipid, di-octadecenyl-trimethylammonium propane, mixed with YSK05, a pH-sensitive lipid, and cholesterol. Optimized amounts of YSK05 and GALA were used to achieve an efficient and lung-selective system. The optimized system produced a high gene expression level in the lungs (>107 RLU/mg protein) with high lung/liver and lung/spleen ratios. GALA/R8 modification and the double-coating design were indispensable for efficient gene expression in the lungs. Despite the fact that NPs prepared with 1-step or 2-step coating have the same lipid amount and composition and the same pDNA dose, the transfection activity was dramatically higher in the lungs in the case of 2-step coating. Surprisingly, 1-step or 2-step coatings had no effect on the amount of nanoparticles that were delivered to the lungs, suggesting that the double-coating strategy substantially improved the efficiency of gene expression at the intracellular level.


Assuntos
DNA/administração & dosagem , Lipídeos/química , Pulmão/efeitos dos fármacos , Nanopartículas/química , Peptídeos/química , Plasmídeos/administração & dosagem , Animais , Linhagem Celular , Feminino , Expressão Gênica/efeitos dos fármacos , Técnicas de Transferência de Genes , Humanos , Concentração de Íons de Hidrogênio , Fígado/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos ICR , Oligopeptídeos/administração & dosagem , Transfecção/métodos
14.
Mol Pharm ; 18(9): 3281-3289, 2021 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-34351769

RESUMO

Lipid-based formulations, such as self-microemulsifying drug-delivery systems (SMEDDSs), are promising tools for the oral delivery of poorly water-soluble drugs. However, failure to maintain adequate aqueous solubility after coming into contact with gastrointestinal fluids is a major drawback. In this study, we examined the use of a novel cinnamic acid-derived oil-like material (CAOM) that binds drugs with a high affinity through π-π stacking and hydrophobic interactions, as an oil core in a SMEDDS for the oral delivery of fenofibrate in rats. The use of the CAOM in the SMEDDS resulted in an unprecedented enhancement in fenofibrate bioavailability, which exceeded the bioavailability values obtained using SMEDDSs based on corn oil, a conventional triglyceride oil, or Labrasol, an enhancer of intestinal permeation. Further characterization revealed that the CAOM SMEDDS does not alter the intestinal permeability and has no inhibitory activity on P-glycoprotein-mediated drug efflux. The results reported herein demonstrate the strong potential of CAOM formulations as new solubilizers for the efficient and safe oral delivery of drugs that have limited water solubility.


Assuntos
Sistemas de Liberação de Medicamentos/métodos , Emulsões/química , Excipientes/química , Fenofibrato/farmacocinética , Lipídeos/química , Administração Oral , Animais , Disponibilidade Biológica , Química Farmacêutica , Óleo de Milho/química , Cães , Composição de Medicamentos/métodos , Liberação Controlada de Fármacos , Fenofibrato/administração & dosagem , Glicerídeos/química , Mucosa Intestinal/metabolismo , Células Madin Darby de Rim Canino , Masculino , Modelos Animais , Ratos , Solubilidade , Água/química
15.
Int J Mol Sci ; 23(1)2021 Dec 22.
Artigo em Inglês | MEDLINE | ID: mdl-35008537

RESUMO

The development of drug delivery systems for use in the treatment of cardiovascular diseases is an area of great interest. We report herein on an evaluation of the therapeutic potential of a myocardial mitochondria-targeting liposome, a multifunctional envelope-type nano device for targeting pancreatic ß cells (ß-MEND) that was previously developed in our laboratory. Resveratrol (RES), a natural polyphenol compound that has a cardioprotective effect, was encapsulated in the ß-MEND (ß-MEND (RES)), and its efficacy was evaluated using rat myocardioblasts (H9c2 cells). The ß-MEND (RES) was readily taken up by H9c2 cells, as verified by fluorescence-activated cell sorter data, and was observed to be colocalized with intracellular mitochondria by confocal laser scanning microscopy. Myocardial mitochondrial function was evaluated by a Seahorse XF Analyzer and the results showed that the ß-MEND (RES) significantly activated cellular maximal respiratory capacity. In addition, the ß-MEND (RES) showed no cellular toxicity for H9c2 cells as evidenced by Premix WST-1 assays. This is the first report of the use of a myocardial mitochondria-targeting liposome encapsulating RES for activating mitochondrial function, which was clearly confirmed based on analyses using a Seahorse XF Analyzer.


Assuntos
Respiração Celular/efeitos dos fármacos , Lipossomos/química , Mitocôndrias Cardíacas/efeitos dos fármacos , Miócitos Cardíacos/efeitos dos fármacos , Resveratrol/farmacologia , Animais , Linhagem Celular , Células Secretoras de Insulina/efeitos dos fármacos , Nanopartículas/química , Polifenóis/química , Ratos , Resveratrol/química
16.
Biochem Biophys Res Commun ; 525(2): 313-318, 2020 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-32087973

RESUMO

For achieving efficient cancer treatment, it is important to elucidate the mechanism responsible for the accumulation of nanoparticles in tumor tissue. Recent studies suggest that nanoparticles are not delivered merely through gaps between tumor endothelial cells. We previously reported that the maturation of the vascular structure by the vascular endothelial cell growth factor receptor 2 (VEGFR2) using a previously developed siRNA delivery technology (RGD-MEND) significantly enhanced the accumulation of nanoparticles in types of cancers that area vessel-rich (renal cell carcinoma). This result was completely inconsistent with the generally accepted theory of the enhanced permeability and retention (EPR) effect. We hypothesized that a caveolin-1 (Cav1)-mediated transcellular route would be involved with the penetration of nanoparticles into tumor vasculature. To reveal the exact mechanism responsible for this enhancement, we observed the delivery of long-circulating liposomes (LPs) after Cav1 was co-suppressed by RGD-MEND with VEGFR2. The enhanced delivery of LPs by siRNA against VEGFR2 (siVEGFR2) was accompanied by the elevated expression of the Cav1 protein. In addition, Cav1 knockdown by siRNA against Cav1 (siCav1) canceled the enhanced delivery of LPs by siVEGFR2. The injection of siCav1 had no effect on the formation of alpha smooth muscle actin or vascular endothelial cell adhesion molecules. These results suggest that a Cav1-induced transcellular route and not a paracellular route, at least partially, contributes to the accumulation of nanoparticles in tumors.


Assuntos
Caveolina 1/fisiologia , Lipossomos/metabolismo , Neoplasias/metabolismo , Transcitose , Animais , Carcinoma de Células Renais/irrigação sanguínea , Carcinoma de Células Renais/metabolismo , Caveolina 1/efeitos dos fármacos , Humanos , Nanopartículas/metabolismo , Neoplasias/irrigação sanguínea , RNA Interferente Pequeno/farmacologia , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores
17.
Mol Pharm ; 17(10): 3773-3782, 2020 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-32881529

RESUMO

The intravenous administration of drug-loaded nanoparticles (NPs) is needed to achieve passive or active targeting in disease tissues. However, when the loaded drug is a hydrophobic small molecule, the NPs fail to reach adequate plasma drug concentrations mainly because of premature drug release. The pharmacokinetics of such drugs can be controlled by covalent modification, but this approach could compromise the safety or potency of the drug. In this study, we investigated two formulation parameters that could be used to improve the plasma concentrations of unmodified drugs that are loaded in a nanoemulsion (NE), a core-shell type NP. The first parameter is the loading ratio, and the second is the affinity of the drug to the core. Optimized NEs with reduced drug loading and with a high drug-core affinity resulted in a 12.4- and 11.2-fold increase in the plasma retention of curcumin and paclitaxel, respectively. Our strategy for enhancing the drug-core interaction affinity relied on mixing oils and surfactants to achieve cooperativity in noncovalent interactions, such as hydrophobic interactions, hydrogen bonding, and π-π stacking, which was further confirmed by theoretical calculations of interaction affinities. Finally, we report on the development of a cinnamic acid-derived oil-like material as a novel drug vehicle with exceptional solubilizing ability that could be used in intravenous formulations of NEs.


Assuntos
Antineoplásicos/farmacocinética , Cinamatos/química , Portadores de Fármacos/química , Excipientes Farmacêuticos/química , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Curcumina/administração & dosagem , Curcumina/química , Curcumina/farmacocinética , Liberação Controlada de Fármacos , Emulsões , Feminino , Interações Hidrofóbicas e Hidrofílicas , Injeções Intravenosas , Camundongos , Camundongos Endogâmicos ICR , Modelos Animais , Nanopartículas/química , Óleos/química , Paclitaxel/administração & dosagem , Paclitaxel/química , Paclitaxel/farmacocinética , Solubilidade
18.
Mol Pharm ; 17(3): 944-953, 2020 03 02.
Artigo em Inglês | MEDLINE | ID: mdl-31990567

RESUMO

Because the lymph node (LN) is a critical organ for inducing immune responses against pathogens and cancers, the transport of immune functional molecules such as antigens and adjuvants to LNs by delivery systems is a useful strategy for the effective outcome of an immune response. The size and charge of a delivery system largely affect the transitivity to and distribution within LN. Although pH-sensitive lipid nanoparticles (LNPs) prepared by microfluidic mixing are the latest delivery system to be applied clinically, the effects of their size and charge on the transitivity to and distribution within LN are currently unknown. We investigated the size and charge effect of LNPs prepared by microfluidic mixing on transitivity to and distribution within LNs. A 30 nm-sized LNP (30-LNP) was efficiently translocated to LNs and was taken up by CD8+ dendritic cells, while the efficiency was drastically decreased in the cases of 100 and 200 nm-sized LNPs. Furthermore, a comparative study between neutral, positively, and negatively charged 30-LNP revealed that the negative 30-LNP moved to the LN more efficiently than the other LNPs. Interestingly, the negative 30-LNP reached the deep cortex, namely, the T cell zone. Our findings provide informative insights for designing LN-targeting LNPs prepared by microfluidic mixing and for the translocation of nanoparticles in LNs.


Assuntos
Células Dendríticas Foliculares/metabolismo , Sistemas de Liberação de Medicamentos/métodos , Lipídeos/química , Lipídeos/farmacocinética , Microfluídica/métodos , Nanopartículas/química , Nanopartículas/metabolismo , Tamanho da Partícula , Adjuvantes Imunológicos/administração & dosagem , Animais , Antígenos CD8/metabolismo , Feminino , Concentração de Íons de Hidrogênio , Camundongos , Camundongos Endogâmicos C57BL , Células RAW 264.7 , RNA Interferente Pequeno/química , Distribuição Tecidual
19.
Mol Pharm ; 17(4): 1397-1404, 2020 04 06.
Artigo em Inglês | MEDLINE | ID: mdl-32091909

RESUMO

Peptide modification is a popular strategy for developing an active targeting lipid nanoparticle (LNP). In modifying the surface of an LNP with a peptide, the sequence and structure of the peptide strongly affects the formation of the LNP. Specifically, a peptide with a high hydrophobicity can induce coarsening and aggregation of the LNP. In an attempt to prevent this from occurring, we incorporated monoacyl and diacyl group-conjugated poly(ethylene glycol) (PEG) into a LNP. We previously developed an original LNP, a multifunctional envelope type nanodevice (MEND) modified with an Epi-1 peptide, a ligand with a high affinity for the epithelial cell adhesion molecule (EpCAM). Using this peptide-modified MEND, the efficiency of delivery of a small interfering RNA (siRNA) encapsulated in the MEND was significantly improved. Although increasing the ratio of modification enhanced cellular uptake, the increase also induced aggregation of the LNP, particularly in the case of a large scale preparation. Our results indicate that a monoacyl PEG-lipid can prevent aggregation, even when the LNP is modified with higher molar ratios of peptide, but that this also results in a decrease in delivery efficiency. Moreover, the Epi-1-modified MEND exhibited a strong silencing effect in an ovarian cancer peritoneal dissemination model. Our results suggest that the simple incorporation of a monoacyl derivative into the PEG-lipid resulted in the formation of a peptide-modified LNP with improved characteristics.


Assuntos
Ácidos Graxos/química , Lipídeos/química , Nanopartículas/química , Polietilenoglicóis/química , Animais , Linhagem Celular Tumoral , Sistemas de Liberação de Medicamentos/métodos , Molécula de Adesão da Célula Epitelial/metabolismo , Feminino , Inativação Gênica/efeitos dos fármacos , Células HCT116 , Humanos , Ligantes , Camundongos , Camundongos Endogâmicos ICR , Camundongos SCID , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/metabolismo , Peptídeos/química , RNA Interferente Pequeno/administração & dosagem
20.
Mol Pharm ; 17(4): 1237-1247, 2020 04 06.
Artigo em Inglês | MEDLINE | ID: mdl-32129629

RESUMO

DNA vaccinations are promising strategies for treating diseases that require cellular immunity (i.e., cancer and protozoan infection). Here, we report on the use of a liposomal nanocarrier (lipid nanoparticles (LNPs)) composed of an SS-cleavable and pH-activated lipidlike material (ssPalm) as an in vivo DNA vaccine. After subcutaneous administration, the LNPs containing an ssPalmE, an ssPalm with vitamin E scaffolds, elicited a higher gene expression activity in comparison with the other LNPs composed of the ssPalms with different hydrophobic scaffolds. Immunization with the ssPalmE-LNPs encapsulating plasmid DNA that encodes ovalbumin (OVA, a model tumor antigen) or profilin (TgPF, a potent antigen of Toxoplasma gondii) induced substantial antitumor or antiprotozoan effects, respectively. Flow cytometry analysis of the cells that had taken up the LNPs in draining lymph nodes (dLNs) showed that the ssPalmE-LNPs were largely taken up by macrophages and a small number of dendritic cells. We found that the transient deletion of CD169+ macrophages, a subpopulation of macrophages that play a key role in cancer immunity, unexpectedly enhanced the activity of the DNA vaccine. These data suggest that the ssPalmE-LNPs are effective DNA vaccine carriers, and a strategy for avoiding their being trapped by CD169+ macrophages will be a promising approach for developing next-generation DNA vaccines.


Assuntos
Lipídeos/química , Nanopartículas/química , Infecções por Protozoários/imunologia , Vacinas de DNA/química , Vacinas de DNA/imunologia , Vitamina E/imunologia , Animais , Vacinas Anticâncer/química , Vacinas Anticâncer/imunologia , DNA/imunologia , Células Dendríticas/imunologia , Feminino , Concentração de Íons de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Imunidade Celular/imunologia , Imunização/métodos , Lipossomos/química , Lipossomos/imunologia , Linfonodos/imunologia , Macrófagos/imunologia , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Neoplasias/imunologia , Ovalbumina/imunologia , Plasmídeos/imunologia , Vitamina E/química
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