受賞国：日本国

受賞国：台湾

受賞国：日本国

受賞国：中華人民共和国

受賞国：日本国

受賞国：日本国

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Abstract

Fibrotic diseases are involved in 45% of deaths in the United States. In particular, fibrosis of the kidney and lung are major public health concerns due to their high prevalence and lack of existing treatment options. Here, we harness the pathophysiological features of fibrotic diseases, namely leaky vasculature and aberrant extracellular matrix (ECM) protein deposition (i.e. collagen), to target an anti-fibrotic biologic and a small molecule drug to disease sites of fibrosis, thus improving their therapeutic potential in mouse models of lung and kidney fibrosis. First, we identify and validate collagen-targeting drug delivery systems that preferentially accumulate in the diseased organs: von Willebrand Factor’s A3 domain (VWF-A3) and decorin-derived collagen-binding peptide-conjugated micelles (CBP-micelles). We then engineer and recombinantly express novel candidate biologic therapies based on the anti-inflammatory cytokine IL-10: A3-IL-10 and A3-Serum Albumin-IL-10 (A3-SA-IL-10). Simultaneously, we stably encapsulate the potential anti-fibrotic water-insoluble drug, rapamycin, in CBP-micelles. We show that these novel formulations of therapeutics bind to collagen in vitro and that their efficacy in mouse models of lung and kidney fibrosis is improved, compared to free, untargeted drugs. Our results demonstrate that collagen-targeted anti-fibrotic drugs may be next generation therapies of high clinical potential.

DOI： 10.1101/2022.01.04.474747

担当区分：責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Induction of cellular immunity is important for effective cancer immunotherapy. Although various antigen carriers for cancer immunotherapy have been developed to date, balancing efficient antigen delivery to antigen presenting cells (APCs) and their activation via innate immune receptors, both of which are crucially important for the induction of strong cellular immunity, remains challenging. For this study, branched β-glucan was selected as an intrinsically immunity-stimulating and biocompatible material. It was engineered to develop multifunctional liposomal cancer vaccines capable of efficient interactions with APCs and subsequent activation of the cells. Hydroxy groups of branched β-glucan (Aquaβ) were modified with 3-methylglutaric acid ester and decyl groups, respectively, to provide pH-sensitivity and anchoring capability to the liposomal membrane. The modification efficiency of Aquaβ derivatives to the liposomes was significantly high compared with linear β-glucan (curdlan) derivatives. Aquaβ derivative-modified liposomes released their contents in response to weakly acidic pH. As a model antigenic protein, ovalbumin (OVA)-loaded liposomes modified with Aquaβ derivatives interacted efficiently with dendritic cells, and induced inflammatory cytokine secretion from the cells. Subcutaneous administration of Aquaβ derivative-modified liposomes suppressed the growth of the E.G7-OVA tumor significantly compared with curdlan derivative-modified liposomes. Aquaβ derivative-modified liposomes induced the increase of CD8+ T cells, and polarized macrophages to the antitumor M1-phenotype within the tumor microenvironment. Therefore, pH-sensitive Aquaβ derivatives can be promising materials for liposomal antigen delivery systems to induce antitumor immune responses efficiently.

DOI： 10.1039/d1tb00786f

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

The antimicrobial protein CAP18 (approximate molecular weight: 18 000), which was first isolated from rabbit granulocytes, comprises a C-terminal fragment that has negatively charged lipopolysaccharide binding activity. In this study, we found that CAP18 (106-121)-derived (sC18)2 peptides have macropinocytosis-inducible biological functions. In addition, we found that these peptides are highly applicable for use as extracellular vesicle (exosomes, EV)-based intracellular delivery, which is expected to be a next-generation drug delivery carrier. Here, we demonstrate that dimerized (sC18)2 peptides can be easily introduced on EV membranes when modified with a hydrophobic moiety, and that they show high potential for enhanced cellular uptake of EVs. By glycosaminoglycan-dependent induction of macropinocytosis, cellular EV uptake in targeted cells was strongly increased by the peptide modification made to EVs, and intriguingly, our herein presented technique is efficiently applicable for the cytosolic delivery of the biologically cell-killing functional toxin protein, saporin, which was artificially encapsulated in the EVs by electroporation, suggesting a useful technique for EV-based intracellular delivery of biofunctional molecules.

DOI： 10.1021/acs.molpharmaceut.1c00244

PubMed

担当区分：筆頭著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Poor distribution of nanocarriers at the tumor site and insufficient drug penetration into the tissue are major challenges in the development of effective and safe cancer therapy. Here, we aim to enhance the therapeutic effect of liposomes by accumulating doxorubicin-loaded liposomes at high concentrations in and around the tumor, followed by heat-triggered drug release to facilitate low-molecular-weight drug penetration throughout the tumor. A cyclic RGD peptide (cRGD) was incorporated into liposomes decorated with a thermosensitive polymer that allowed precise tuning of drug release temperature (i.e., Polymer-lip) to develop a targeted thermosensitive liposome (cRGD-Polymer-lip). Compared with conventional thermosensitive liposomes, cRGD-Polymer-lip enhanced the binding of liposomes to endothelial cells, leading to their accumulation at the tumor site upon intravenous administration in tumor-bearing mice. Drug release triggered by local heating strongly inhibited tumor growth. Notably, tumor remission was achieved via multiple administrations of cRGD-Polymer-lip and heat treatments. Thus, combining the advantages of tumor neovascular targeting and heat-triggered drug release, these liposomes offer high potential for minimally invasive and effective cancer chemotherapy.

DOI： 10.1021/acs.molpharmaceut.1c00263

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Interleukin-4 (IL-4) suppresses the development of multiple sclerosis in a murine model of experimental autoimmune encephalomyelitis (EAE). Here, we show that, in mice with EAE, the accumulation and persistence in the lymph nodes and spleen of a systemically administered serum albumin (SA)-IL-4 fusion protein leads to higher efficacy in preventing disease development than the administration of wild-type IL-4 or of the clinically approved drug fingolimod. We also show that the SA-IL-4 fusion protein prevents immune-cell infiltration in the spinal cord, decreases integrin expression in antigen-specific CD4+ T cells, increases the number of granulocyte-like myeloid-derived suppressor cells (and their expression of programmed-death-ligand-1) in spinal cord-draining lymph nodes and decreases the number of T helper 17 cells, a pathogenic cell population in EAE. In mice with chronic EAE, SA-IL-4 inhibits immune-cell infiltration into the spinal cord and completely abrogates immune responses to myelin antigen in the spleen. The SA-IL-4 fusion protein may be prophylactically and therapeutically advantageous in the treatment of multiple sclerosis.

DOI： 10.1038/s41551-020-00627-3

PubMed

その他URL： http://www.nature.com/articles/s41551-020-00627-3

担当区分：筆頭著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

OBJECTIVE: Rheumatoid arthritis (RA) is a major autoimmune disease that causes synovitis and joint damage. Although clinical trials have been performed using interleukin-10 (IL-10), an antiinflammatory cytokine, as a potential treatment of RA, the therapeutic effects of IL-10 have been limited, potentially due to insufficient residence in lymphoid organs, where antigen recognition primarily occurs. This study was undertaken to engineer an IL-10-serum albumin (SA) fusion protein and evaluate its effects in 2 murine models of RA. METHODS: SA-fused IL-10 (SA-IL-10) was recombinantly expressed. Mice with collagen antibody-induced arthritis (n = 4-7 per group) or collagen-induced arthritis (n = 9-15 per group) were injected intravenously with wild-type IL-10 or SA-IL-10, and the retention of SA-IL-10 in the lymph nodes (LNs), immune cell composition in the paws, and therapeutic effect of SA-IL-10 on mice with arthritis were assessed. RESULTS: SA fusion to IL-10 led to enhanced accumulation in the mouse LNs compared with unmodified IL-10. Intravenous SA-IL-10 treatment restored immune cell composition in the paws to a normal status, elevated the frequency of suppressive alternatively activated macrophages, reduced IL-17A levels in the paw-draining LN, and protected joint morphology. Intravenous SA-IL-10 treatment showed similar efficacy as treatment with an anti-tumor necrosis factor antibody. SA-IL-10 was equally effective when administered intravenously, locally, or subcutaneously, which is a benefit for clinical translation of this molecule. CONCLUSION: SA fusion to IL-10 is a simple but effective engineering strategy for RA therapy and has potential for clinical translation.

DOI： 10.1002/art.41585

PubMed

その他URL： https://onlinelibrary.wiley.com/doi/full-xml/10.1002/art.41585

担当区分：筆頭著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

For the establishment of advanced medicines such as cancer immunotherapy, high performance carriers that precisely deliver biologically active molecules must be developed to target organelles of the cells and to release their contents there. From the viewpoint of antigen delivery, endosomes are important target organelles because they contain immune-response-related receptors and proteins of various types. To obtain carriers for precision endosome delivery, a novel type of polyamidoamine dendron-based lipid having pH-sensitive terminal groups was synthesized for this study. Liposomes were prepared using these pH-sensitive dendron-based lipids and egg yolk phosphatidylcholine. Their pH-responsive properties and performance as an endosome delivery carrier were investigated. pH-Sensitive dendron lipid-based liposomes retained water-soluble molecules at neutral pH but released them under weakly acidic conditions. Particularly, liposomes containing CHexDL-G1U exhibited highly sensitive properties responding to very weakly acidic pH. These dendron lipid-based liposomes released the contents specifically in the endosome. The timing of content release can be controlled by selecting pH-sensitive dendron lipids for liposome preparation. Significant tumor regression was induced in tumor-bearing mice by the administration of CHexDL-G1U-modified liposomes containing the model antigenic protein. Furthermore, CHexDL-G1U-modified liposomes induced WT1 tumor antigenic peptide-specific helper T cell proliferation. The results demonstrate that dendron lipid-based liposomes are useful as a potent vaccine for cancer immunotherapy.

DOI： 10.1039/d0bm01813a

PubMed

担当区分：責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

DOI： 10.1002/nano.202000218

その他URL： https://onlinelibrary.wiley.com/doi/full-xml/10.1002/nano.202000218

担当区分：筆頭著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

RNA interference (RNAi) using siRNA has gained much attention for use in therapies for cancer and genetic disorders. To establish RNAi-based therapeutics, the development of efficient siRNA nanocarriers is desired. Earlier, we developed polyamidoamine dendron-bearing lipids able to form complexes with nucleic acids as gene vectors. Especially, dendron lipids with unsaturated alkyl chains (DL-G1-U2) induced efficient endosomal escape by membrane fusion, leading to efficient transfection in vitro. For this study, dendron lipids having oleyl/linoleyl groups (DL-G1-U3) were designed to increase membrane fusogenic activity further. Indeed, DL-G1-U3/siRNA complexes achieved higher membrane fusogenic activity and knockdown of the target gene more efficiently than conventional DL-G1-U2/siRNA complexes did. A hydrophilic polymer, hyperbranched polyglycidol lauryl ester (HPG-Lau), was modified further on the surface of DL-G1-U3/siRNA complexes to provide colloidal stability. Surface modification of HPG-Lau increased the colloidal stability in a physiological condition more than complexes without HPG-Lau. Importantly, HPG-Lau-coated DL/siRNA complexes showed identical RNAi effects to those of parental DL/siRNA complexes, whereas the RNAi activity of poly(ethylene glycol)-bearing lipid (PEG-PE)-modified DL/siRNA complexes was hindered completely. Introduction of unsaturated bonds into dendron lipids and selection of suitable hydrophilic polymers for nanocarrier modification are important for obtaining efficient siRNA vectors toward in vivo siRNA delivery.

DOI： 10.1021/acs.bioconjchem.1c00035

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Exosomes (extracellular vesicles/EVs) participate in cell–cell communication and contain bioactive molecules, such as microRNAs. However, the detailed characteristics of secreted EVs produced by cells grown under low pH conditions are still unknown. Here, we report that low pH in the cell culture medium significantly affected the secretion of EVs with increased protein content and zeta potential. The intracellular expression level and location of stably expressed GFP-fused CD63 (an EV tetraspanin) in HeLa cells were also significantly affected by environmental pH. In addition, increased cellular uptake of EVs was observed. Moreover, the uptake rate was influenced by the presence of serum in the cell culture medium. Our findings contribute to our understanding of the effect of environmental conditions on EV-based cell–cell communication.

DOI： 10.1002/2211-5463.13107

PubMed

その他URL： https://onlinelibrary.wiley.com/doi/full-xml/10.1002/2211-5463.13107

担当区分：筆頭著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Specific delivery to antigen presenting cells (APC) and precise control of the intracellular fate of antigens are crucial to induce cellular immunity that directly and specifically attacks cancer cells. We previously achieved cytoplasmic delivery of antigen and activation of APC using carboxylated curdlan-modified liposomes, which led to the induction of cellular immunity in vivo. APCs express mannose receptors on their surface to recognize pathogen specifically and promote cross-presentation of antigen. In this study, mannose-residue was additionally introduced to carboxylated curdlan as a targeting moiety to APC for further improvement of polysaccharide-based antigen carriers. Mannose-modified curdlan derivatives were synthesized by the condensation between amino group-introduced mannose and carboxy group in pH-sensitive curdlan. Mannose residue-introduced carboxylated curdlan-modified liposomes showed higher pH-sensitivity than that of liposomes modified with conventional carboxylated curdlan. The introduction of mannose-residue to the liposomes induced aggregation in the presence of Concanavalin A, indicating that mannose residues were presented onto liposome surface. Mannose residue-introduced carboxylated curdlan-modified liposomes exhibited high and selective cellular association to APC. Furthermore, mannose residue-introduced carboxylated curdlan-modified liposomes promoted cross-presentation of antigen and induced strong antitumor effects on tumor-bearing mice. Therefore, these liposomes are promising as APC-specific antigen delivery systems for the induction of antigen-specific cellular immunity.

DOI： 10.3390/pharmaceutics12080754

PubMed

担当区分：責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Background: Gene delivery to target cells is crucially important to establish gene therapy and regenerative medicine. Although various virus-based and synthetic molecule-based gene vectors have been developed to date, selective transfection in a site or a cell level is still challenging. For this study, both light-responsive and temperature-responsive synthetic gene vectors were designed for spatiotemporal control of a transfection system. Methods: 11-Mercaptoundecanoic acid-coated gold nanorods were mixed with polyamidoamine dendron-bearing lipids of two types having amino-terminus or ethoxydiethylene glycol-terminus to obtain hybrid vectors. Hybrid vectors were mixed further with pDNA. Then we investigated their physicochemical properties and transfection efficacy with or without near infrared laser irradiation. Results: Hybrid vectors formed complexes with pDNA and exhibited enhanced photothermal property under near infrared laser irradiation compared with parent gold nanorods. Transfection efficacy of complexes was promoted considerably by brief laser irradiation soon after complex application to the cells. Analysis of intracellular distribution revealed that laser irradiation promoted the adsorption of complexes to the cells and cytosolic release of pDNA, which is derived from the change in surface hydrophobicity of complexes through dehydration of temperature-responsive groups. Conclusions: Hybrid vector is promising as a light-activatable transfection system.

DOI： 10.3390/pharmaceutics12030239

PubMed

担当区分：責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Temperature-responsive nanocarrier systems using external stimuli are one of the most widely investigated stimuli-responsive strategies because heat is easy and safe to use for hyperthermia and controlled drug delivery. Polyamidoamine dendron lipids (PAMAM-DLs) composed of PAMAM dendron as head group and two alkyl chains can exhibit temperature-responsive morphological change through the attachment of suitable moieties to terminal of PAMAM dendron. In this study, oligo(ethylene glycol)s including ethoxy- or methoxy-diethylene glycols were attached to the terminals of PAMAM-DL, and temperature-responsive properties of their self-assemblies were evaluated by calorimetric and turbidity measurements. In the evaluation of temperature-responsive properties, ethoxy diethylene glycol (EDEG)-attached PAMAM-DL composed of two saturated alkyl chains and PAMAM dendron with 1st generation had lipid bilayer structure and suitable cloud point for the application as drug carrier. In vitro performances of the assemblies combining EDEG-attached PAMAM-DLs with cholesteryl-oxy-poly(ethylene glycol) (PEG-Chol) was evaluated using doxorubicin (DOX) as an anticancer drug. Cellular uptake of DOX-loaded EDEG-attached PAMAM-DL/PEG-Chol assemblies was promoted at 42 ◦C rather than 37 ◦C, resulting in an effective decrease in cell viability.

DOI： 10.3390/jfb11010016

PubMed

担当区分：責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Functional dendrimer-gold nanorod hybrid for combination of anticancer drugs and laser hyperthermia towards efficient cancer treatment with less-adverse effects.

DOI： 10.1039/c9tb02163a

PubMed

担当区分：筆頭著者,　最終著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Liposomes are a promising nanocarrier for drug delivery because of their biocompatibility and the encapsulation capacity of drugs. Liposomes can be functionalized easily by introduction of functional materials such as stimulus-responsive materials. Temperature-responsive liposomes and pH-responsive liposomes are representative stimulus-responsive liposomes that can deliver drugs to locally heated target tissues and intracellular organelles. Here, temperature-responsive liposomes for the selective release of cargo and pH-responsive liposomes for the induction of antigen-specific immunity are overviewed. Temperature-responsive polymer-modified liposomes immediately released drugs in response to heating, which achieved selective drug release at a tumour after topical heating of tumour-bearing mice. Introduction of MR-detectable molecules enabled the tracing of liposome accumulation into target sites to optimize the heating timing. These liposomes can also be combined with magnetic nanoparticles or carbon nanomaterials to attain magnetic field-responsive, electric field-responsive and light-responsive properties to support on-demand drug release or control of biological reactions using these external stimuli. pH-Responsive liposomes were produced by modification of poly(carboxylic acid) derivatives or by pH-responsive amphiphiles. These liposomes delivered antigenic proteins into the cytosol of antigen presenting cells, which induced cross-presentation and antigen-specific cellular immunity. Adjuvant molecules or bioactive polysaccharide-based pH-responsive polymers improved their immunity-inducing effect further, leading to tumour regression in tumour-bearing mice. Precise design and control of the structures of stimulus-responsive materials and combination with functional materials are expected to create novel methodologies to control biological functions and to produce highly potent liposomal drugs that can achieve selective release of bioactive molecules.

DOI： 10.1039/c9tb02470k

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

The tumor microenvironment strongly influences clinical outcomes of immunotherapy. By transfecting genes of relevant cytokines into tumor cells, we sought to manipulate the microenvironment so as to elicit activation of T helper type 1 (Th1) responses and the maturation of dendritic cells (DCs). Using a synthetic vehicle, the efficiency of in vivo transfection of GFP-cDNA into tumor cells was about 7.5% by intratumoral injection and about 11.5% by intravenous injection. Survival was significantly improved by both intratumoral and intravenous injection of the plasmid containing cDNA of interferon-gamma, followed by intratumoral injection of DCs presenting the tumor antigens. Also, tumor growth was inhibited by these treatments. A more significant effect on survival and tumor growth inhibition was observed following injection of the plasmid containing cDNA of CD40 ligand, which is a potent inducer of DC-maturation. Furthermore, the co-injection of both IFNγ- and CD40 ligand-encoding cDNA-plasmids, followed by DC treatment, gave rise to further marked and enhancement, including 100% survival and more than 50% complete remission. This treatment regimen elicited significant increases in mature DCs and types of cells contributing to Th1 responses, and significant decreases in immune suppressor cells in the tumor. In the spleen, the treatment significantly increased activities of tumor-specific killer and natural killer cells, but no alteration was observed in mature DCs or suppressor cells. These results indicate that transfection of these cytokine genes into tumor cells significantly alter the tumor microenvironment and improve the therapeutic results of DC-based immunotherapy.

DOI： 10.1096/fba.2019-00052

PubMed

掲載種別：論文集(書籍)内論文   共著区分：共著   国際・国内誌：国際誌  

Poly(L-lysine)-bearing multi-arm poly(ethylene glycol) terminary (maPEG-PLL) can form nanorod and nanofiber polyplexes with plasmid DNA (pDNA) depending on the size (exclusion volume) of maPEG. Nanorod and nanofiber polyplexes were prepared using maPEG-PLL bearing different maPEG sizes. Gene expression of nanofiber and nanorod polyplexes was comparable, although the cellular uptake of nanofiber polyplexes was less than that of nanorod polyplexes. This suggests that there is a difference in cellular activities after polyplex enters the cell. The release of pDNA from polyplexes through the exchange with heparin occurred more easily in nanorod polyplexes than it did in nanofiber polyplexes. However, it was confirmed by real-time PCR measurements that pDNA molecules in nanofiber polyplexes exhibit high reactivity with polymerase compared with that in nanorod polyplexes. Further, higher reactivity of pDNA with polymerase in nanofiber polyplexes was maintained even in the presence of crowding cosolutes including Ficoll and dextran. This suggests that pDNA in nanofiber polyplex might reflect a higher efficiency of transcription process in the cell compared with pDNA in nanorod polyplex. These results provide valuable information in the design of nonviral gene vectors using cationic polymers.

DOI： 10.1021/bk-2020-1350.ch002

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Background: Although disease in a majority of rheumatoid arthritis (RA) patients is often initially limited to one or a few joints, currently approved medications including anti-tumor necrosis factor-α antibody (α-TNF) are injected systemically. Given that α-TNF systemic injection typically does not cure RA and involves risk of treatment-related adverse events, one possible approach to enhance therapeutic efficacy and reduce α-TNF systemic exposure is to retain the antibodies in arthritic joints after local administration. The aim of this study was to evaluate the approach of conferring extracellular matrix (ECM) binding affinity to α-TNF antibodies in a RA model. Methods: α-TNF was chemically conjugated with a promiscuous ECM-binding peptide derived from placenta growth factor 2 (PlGF-2123-144). The binding activity of PlGF-2123-144-conjugated α-TNF (PlGF-2123-144-α-TNF) against ECM proteins was assessed by ELISA and by immunostaining on human cartilage specimens. The effect of conjugation on antibody function was assessed as a neutralizing activity against osteoclast differentiation. Retention at the injection site and therapeutic efficacy of PlGF-2123-144-α-TNF were tested in a collagen antibody-induced arthritis (CAIA) model in the mouse. Results: PlGF-2123-144 peptide conjugation conferred α-TNF with affinity to ECM proteins without impairment of antigen recognition. PlGF-2123-144-α-TNF locally injected at a paw in the CAIA model was retained for at least 96 h at the injection site, whereas unmodified α-TNF was dispersed rapidly after injection. Local treatment with unmodified α-TNF did not suppress the arthritis score relative to isotype controls. By contrast, local administration of PlGF-2123-144-α-TNF suppressed arthritis development almost completely in the treated paw even at a 1000× lower dose. Conclusion: These data demonstrate that retention of α-TNF in arthritic joints can suppress arthritis development and enhance therapeutic efficacy. This simple bioengineering approach of ECM-binding peptide conjugation offers a powerful and clinically translational approach to treat RA.

DOI： 10.1186/s13075-019-2075-8

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

BACKGROUND/AIM: Extracellular vesicles (exosomes, EVs) (30-200 nm in diameter) are secreted by various cells in the body. Owing to the pharmaceutical advantages of EVs, an EV-based drug delivery system (DDS) for cancer therapy is expected to be the next-generation therapeutic system. However, preservation methods for functional and therapeutic EVs should be developed. Here, we developed the method of lyophilization of arginine-rich cell penetrating peptide (CPP)-modified EVs and investigated the effects of lyophilization on the characteristics of EVs. MATERIALS AND METHODS: Particle size, structure, zeta-potential, and cellular uptake efficacy of the arginine-rich CPP-modified EVs were analyzed. The model protein saporin (SAP), having anti-cancer effects, was encapsulated inside the EVs to assess the cytosolic release of EV content after cellular uptake. RESULTS: Lyophilization of the EVs did not affect their particle size, structure, zeta-potential, and cellular uptake efficacy; however, the biological activity of the encapsulated SAP was inhibited by lyophilization. CONCLUSION: Lyophilization of EVs may affect SAP structures and/or reduce the cytosolic release efficacy of EV's content after cellular uptake and needs attention in EV-based DDSs.

DOI： 10.21873/anticanres.13885

PubMed

担当区分：責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Cancer immunotherapy has gained much attention because of the recent success of immune checkpoint inhibitors. Nevertheless, clinical therapeutic effects of immune checkpoint inhibitors remain limited, probably because most patients have other immune checkpoint molecules or because they lack cancer-specific cytotoxic T lymphocytes. Induction of cancer-specific cytotoxic T lymphocytes requires efficient antigen delivery systems that can convey cancer antigens specifically to antigen presenting cells, promote the endosomal escape of antigen into cytosol, and activate immune cells. Earlier, we reported cytoplasmic delivery systems of antigen using pH-sensitive polymer-modified liposomes. Adjuvant molecules were further incorporated into these liposomes to provide activation properties of cellular immune responses. This study further introduced cell specificity to these liposomal systems using hyaluronic acid-based pH-sensitive polymers, which are recognized by CD44 expressing on antigen presenting cells. pH-Sensitive hyaluronic acid derivative-modified liposomes showed much higher cellular association to antigen presenting cells than to fibroblasts with less CD44 expression. These liposomes achieved the delivery of model antigenic proteins into cytosol of dendritic cells and promoted Th1 cytokine production from the cells. Subcutaneous administration of these liposomes to mice induced antigen-specific cellular immune response in the spleen, leading to tumor regression in tumor-bearing mice. The results show that pH-sensitive hyaluronic acid derivative-modified liposomes are promising as multifunctional antigen carriers having cell-specificity, cytoplasmic antigen delivery performance, and adjuvant property to induce antigen-specific cellular immunity.

DOI： 10.1021/acsbiomaterials.9b01278

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Controlling the aspect ratio of polyplexes prepared by mixing pDNA with a polycation mixture of a poly(l-lysine) (PLL) homopolymer and PLL terminally bearing a multiarm poly(ethylene glycol) (PEG) part (maPEG-PLL) was examined. By varying the maPEG-PLL content in the polycation mixture, the condensation of pDNA accompanied by polyplex formation and the morphology of the polyplexes were evaluated by a dye exclusion assay and AFM observations, respectively. Increasing the maPEG-PLL content caused elongation of the polyplex, and polyplexes with aspect ratios from 2 to 10 were prepared successfully by controlling the maPEG-PLL content. The reactivity of pDNA in the polyplexes with varying aspect ratios against DNase I and polymerase were examined by agarose gel electrophoresis and real-time PCR measurements, respectively. Moreover, cellular uptake and transfection efficiency of the polyplexes by HeLa cells was evaluated. The results revealed that an increase in aspect ratio of the polyplexes caused an increase in PCR efficiency with a concomitant decrease in cellular uptake.

DOI： 10.1021/acsbiomaterials.8b01498

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Enhancing the therapeutic efficacy of drugs for inflammatory diseases is of high demand. One possible approach is targeting drugs to the extracellular matrix of the inflamed area. Here, we target collagens in the matrix, which are inaccessible in most tissues yet are exposed to the bloodstream in the inflamed area because of vascular hyperpermeability. We conferred collagen affinity to anti–tumor necrosis factor-α (α-TNF) antibody by conjugating a collagen-binding peptide (CBP) derived from the sequence of decorin. CBP–α-TNF accumulated in the inflamed paw of the arthritis model, and arthritis development was significantly suppressed by treatment with CBP–α-TNF compared with the unmodified antibody. Similarly, CBP–anti-transforming growth factor-β (α–TGF-β) accumulated in the inflamed lung of pulmonary fibrosis model and significantly suppressed pulmonary fibrosis compared with the unmodified antibody. Together, collagen affinity enables the anticytokine antibodies to target arthritis and pulmonary fibrosis accompanied by inflammation, demonstrating a clinically translational approach to treat inflammatory diseases.

DOI： 10.1126/sciadv.aay1971

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Dendritic cells (DCs) present exogenous antigens on major histocompatibility complex (MHC) class I molecules, thereby activating CD8+ T cells, contributing to tumor elimination through a mechanism known as antigen cross-presentation. A variety of factors such as maturation state of DCs, co-stimulatory signals, T-cell microenvironment, antigen internalization routes and adjuvants regulate the process of DC-mediated antigen cross-presentation. Recently, the development of successful cancer immunotherapies may be attributed to the ability of DCs to cross-present tumor antigens. In this review article, we focus on the underlying mechanism of antigen cross-presentation and ways to improve antigen cross-presentation in different DC subsets. We have critically summarized the recent developments in the generation of novel nanovaccines for robust CD8+ T-cell response in cancer. In this context, we have reviewed nanocarriers that have been used for cancer immunotherapeutics based on antigen cross-presentation mechanism. Additionally, we have also expressed our views on the future applications of this mechanism in curing cancer.

DOI： 10.1042/BSR20193220

PubMed

担当区分：責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Induction of cancer-specific cytotoxic T lymphocytes is crucially important to complement therapeutic effects of immune checkpoint inhibitors and to achieve efficient cancer immunotherapy. To induce cancer-specific cytotoxic T lymphocytes, cancer antigen carriers must have multiple functions to deliver cancer antigens to antigen presenting cells, release antigens into cytosol, and promote the maturation of these cells. We earlier achieved cytosolic delivery of antigens and induction of antigen-specific cytotoxic T lymphocytes using carboxylated polyglycidol or polysaccharide derivative-modified liposomes that can induce membrane fusion with endosomes in response to weakly acidic pH. Furthermore, pH-sensitivity and adjuvant properties of these polymers were enhanced strongly by introduction of hydrophobic carboxylated units to dextran. Against our expectations, these polymer-modified liposomes only slightly induce cancer immunity, probably because of the high hydrophobicity of spacer units. This study used a polysaccharide with charged groups (chondroitin sulfate) instead of dextran as a backbone to reduce hydrophobicity. Chondroitin sulfate derivative-modified liposomes showed almost equal pH-sensitivity to that of dextran derivative-modified liposomes and achieved selective delivery to dendritic cells, whereas dextran derivative-modified liposomes were highly taken up by both dendritic cells and fibroblasts. Chondroitin sulfate derivative-modified liposomes delivered model antigenic proteins into cytosol of dendritic cells and promoted cytokine production from the cells, leading to tumor regression on tumor-bearing mice after subcutaneous administration. Results demonstrate that charged groups having polysaccharide as a backbone can be used in an effective strategy to balance strong hydrophobicity of spacer units with their utilization for immunity-inducing systems.

DOI： 10.1021/acs.bioconjchem.9b00221

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Cerebral ischemia is a deadly condition that arises due to blockage of the blood vessels in the brain leading to oxygen deficiency thereby arresting brain functions and resulting in death or permanent impairment. Though a myriad of factors has been proposed to cause cerebral ischemia, it has been generally regarded as an old age-associated malady. However, unhealthy diet, stressful lifestyle and deteriorating environment quality has dramatically reduced the age on onset as well as the number of victims in recent years. Hence, there exists a need for prompt and effective therapeutic strategies for immediate as well as long-term damage control and maintenance of the functions of the brain, as well as early and accurate diagnosis of the risk of stroke or extent of damage after stroke. The physiological barriers further complicate the development of therapeutic and diagnostic interventions for cerebral stroke. The advent of nanotechnology has initiated new vistas for more effective and superior therapeutic and imaging modalities for management of cerebral ischemia. This review provides an overview on the current knowledge on the mechanism and causative factors of cerebral ischemia, drawbacks of conventional therapy as well as molecular targets that are being explored for stroke therapy. The review also discusses in detail the advances made using nano-interventions for therapy and imaging of stroke-affected regions along with their pros and cons. Emergent multi-functional nanoparticles for stroke management have also been reviewed.

DOI： 10.1016/j.jconrel.2019.02.031

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

Sonodynamic therapy (SDT) is a novel promising noninvasive therapy involving utilization of low-intensity ultrasound and sonosensitizer, which can generate reactive oxygen species (ROS) by sonication. In SDT, a high therapeutic effect is achieved by intracellular delivery and accumulation at the target sites of sonosensitizer followed by oxidative damage of produced ROS by sonication. Here, pH- and redox-responsive hollow nanocapsules are prepared through the introduction of disulfide cross-linkages to self-assembled polymer vesicles formed from polyamidoamine dendron-poly(l-lysine) for the efficient delivery of sonosensitizer. As sonosensitizer, doxorubicin (DOX), an anticancer drug accumulating into cell nucleus, is selected. Also, the conjugate of DOX and triphenylphosphonium (TPP-DOX) is synthesized as sonosensitizer with mitochondrial targeting ability. DOX and TPP-DOX are delivered to nucleus and mitochondria by nanocapsules. Furthermore, DOX- or TPP-DOX-loaded nanocapsules exhibit in vitro sonodynamic therapeutic effect to HeLa cells with sonication, which might be through oxidative damage to nucleus and mitochondria.

DOI： 10.1002/mabi.201800365

PubMed

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

DOI： 10.1021/acsbiomaterials.8b01254

担当区分：筆頭著者,　最終著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：単著   国際・国内誌：国内誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国内誌  

担当区分：筆頭著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：筆頭著者,　最終著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：単著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/16174

担当区分：責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/16172

担当区分：筆頭著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/16173

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：筆頭著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/15729

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：筆頭著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/15730

担当区分：責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/15731

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：筆頭著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/15728

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：筆頭著者,　最終著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：単著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/15732

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：筆頭著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/00017521

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/00017520

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：筆頭著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/00017519

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：筆頭著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/00017518

担当区分：筆頭著者,　責任著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/00017517

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：筆頭著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：筆頭著者   掲載種別：研究論文（研究会，シンポジウム資料等）   共著区分：共著   国際・国内誌：国内誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：筆頭著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/12482

担当区分：筆頭著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/12481

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

担当区分：筆頭著者   掲載種別：研究論文（学術雑誌）   共著区分：共著   国際・国内誌：国際誌  

リポジトリURL： http://hdl.handle.net/10466/12480

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担当区分：筆頭著者,　最終著者,　責任著者   掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著   国際・国内誌：国内誌  

免疫療法の発展のために、免疫担当細胞に抗原を運搬すると同時に活性化し、抗原特異的な細胞性免疫を誘導できる抗原キャリアが求められている。筆者らは、弱酸性pHに応答して膜融合性を発現する機能性高分子を抗原封入リポソームに修飾することで、細胞性免疫を効果的に誘導できる抗原キャリアを開発した。pH応答性高分子修飾リポソームは樹状細胞に効果的に取り込まれ、エンドソームの弱酸性pHに応答して膜融合性となりサイトゾルに抗原を運搬することで、抗原特異的な細胞性免疫を誘導した。さらに、アジュバント分子の組み込みや、生理活性多糖を基盤としたpH応答性高分子の利用により、担がんマウスにおいて強力な抗腫瘍免疫の誘導に成功した。(著者抄録)

J-GLOBAL

担当区分：筆頭著者,　最終著者,　責任著者   掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：共著   国際・国内誌：国内誌  

担当区分：筆頭著者,　最終著者,　責任著者   掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著   国際・国内誌：国内誌  

担当区分：筆頭著者,　責任著者   掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：共著   国際・国内誌：国内誌  

担当区分：筆頭著者,　責任著者   掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：共著   国際・国内誌：国内誌  

担当区分：筆頭著者,　最終著者,　責任著者   掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著   国際・国内誌：国内誌  

担当区分：筆頭著者,　最終著者,　責任著者   掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著   国際・国内誌：国内誌  

担当区分：筆頭著者,　最終著者,　責任著者   掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：単著   国際・国内誌：国内誌  

担当区分：筆頭著者   掲載種別：記事・総説・解説・論説等（商業誌、新聞、ウェブメディア）   共著区分：共著   国際・国内誌：国内誌  

掲載種別：記事・総説・解説・論説等（学術雑誌）   共著区分：共著   国際・国内誌：国内誌  

掲載種別：記事・総説・解説・論説等（商業誌、新聞、ウェブメディア）   共著区分：共著   国際・国内誌：国内誌  

会議種別：口頭発表（招待・特別）  

会議種別：口頭発表（招待・特別）  

会議種別：口頭発表（招待・特別）  

会議種別：口頭発表（招待・特別）  

会議種別：ポスター発表  

会議種別：口頭発表（招待・特別）  

会議種別：口頭発表（招待・特別）  

会議種別：口頭発表（招待・特別）  

会議種別：口頭発表（招待・特別）  

会議種別：口頭発表（招待・特別）  

会議種別：口頭発表（招待・特別）  

会議種別：口頭発表（招待・特別）  

会議種別：口頭発表（招待・特別）  

会議種別：口頭発表（招待・特別）  

会議種別：口頭発表（招待・特別）  

会議種別：口頭発表（招待・特別）  

会議種別：口頭発表（招待・特別）  

産業財産権の種類：特許権 

特許番号/登録番号：特許第5526318号  

産業財産権の種類：特許権 

特許番号/登録番号：特許第5429710号