Synthesis of Lipidated Ligands and Formulation of Glia-Specific LNPs for RNAi-Mediated BBB Protection.

Pro-inflammatory polarization of microglia and astrocytes results in neuroinflammation and blood-brain barrier (BBB) disruption after a primary traumatic brain injury (TBI). Herein, we demonstrate that the dual-ligand functionalized lipid nanoparticles (AM31 LNPs) were actively and specifically internalized by microglia and astrocytes via mannose receptor (MR)- and adenosine receptor (AR)-mediated endocytosis, respectively, in a mouse model of TBI. Systemic administration of AM31 LNPs carrying siRNA against p65 resulted in internalization by the glial cells in the peri-infarct region and a robust knockdown of p65 at both mRNA and protein levels in these cells, leading to significant down-regulation of key pro-inflammatory cytokines and up-regulation of key anti-inflammatory cytokines. AM31 LNP-mediated silencing of p65 ameliorated TBI-induced BBB disruption. Our data proved that AM 31 LNP is a promising vehicle for RNA therapeutics for targeting microglia and astrocytes in neural disorder.

Tumor-Targeted Codelivery of CpG and siRNA by a Dual-Ligand-Functionalized Curdlan Nanoparticle.

The simultaneous delivery of CpG oligonucleotide along with short interfering RNA (siRNA) has the potential to significantly boost the anticancer impact of siRNA medications. Our previous research demonstrated that Curdlan nanoparticles functionalized with adenosine are capable of selectively delivering therapeutic siRNA to cancerous cells through endocytosis mediated by adenosine receptors. Herein, we synthesized a dual-ligand-functionalized Curdlan polymer (denoted by CuMAN) to simultaneously target tumor cells and tumor-associated macrophages (TAMs). CuMAN nanoparticles containing CpG and siRNA demonstrated enhanced uptake by B16F10 tumor cells and bone marrow-derived macrophages, which are facilitated by AR on tumor cells and mannose receptor on macrophages. This led to increased release of pro-inflammatory cytokines in both in vitro and in vivo settings. The synergistic effect of CpG on TAMs and RNAi on tumor cells mediated by the CuMAN nanoparticle not only suppressed the tumor growth but also strongly inhibited the lung metastasis. Our findings indicate that the CuMAN nanoparticle has potential as an effective dual-targeting delivery system for nucleic acid therapeutics.

Astrocyte-targeted siRNA delivery by adenosine-functionalized LNP in mouse TBI model.

Traumatic brain injury (TBI) induces pro-inflammatory polarization of astrocytes and causes secondary disruption of the blood-brain barrier (BBB) and brain damage. Herein, we report a successful astrocyte-targeted delivery of small interfering RNA (siRNA) by ligand functionalized lipid nanoparticles (LNPs) formulated from adenosine-conjugated lipids and a novel ionizable lipid (denoted by Ad4 LNPs). Systemic administration of Ad4 LNPs carrying siRNA against TLR4 to the mice TBI model resulted in the specific internalization of the LNPs by astrocytes in the vicinity of damaged brain tissue. A substantial knockdown of TLR4 at both mRNA and protein levels in the brain was observed, which led to a significant decrease of key pro-inflammatory cytokines and an increase of key anti-inflammatory cytokines in serum. Dye leakage measurement suggested that the Ad4-LNP-mediated knockdown of TLR4 attenuated the TBI-induced BBB disruption. Together, our data suggest that Ad4 LNP is a promising vehicle for astrocyte-specific delivery of nucleic acid therapeutics.

Tumor targeted siRNA delivery by adenosine receptor-specific curdlan nanoparticles.

Aminated curdlan derivatives are highly effective nucleic acid carriers. Previously, we proved that the ligand-functionalized curdlan derivatives have greatly enhanced cell type specificity induced by receptor-mediated internalization in vitro. In this study, to improve biocompatibility and enhance tumor-targeting efficacy of the curdlan derivative, we pegylated the adenosine functionalized amino curdlan derivative (denoted by pAVC polymer). We confirmed that the uptake of pAVC polymer carrying siRNA by tumor cells was adenosine receptor (AR)-dependent and was specifically inhibited by AMP but not by GMP. The pAVC polymers not only preserved the receptor recognition and exhibited significantly decreased cytotoxicity but also showed remarkable tumor targeting efficiency in vivo. The nanoparticles formulated from siRNA (against STAT3) and pAVC4 polymer, which bears the highest degree of PEG substitution, delivered siRNA highly specifically to tumor tissue, knocked down STAT3, and inhibited tumor growth. The pAVC polymers may be a promising carrier for tumor specific delivery of nucleic acid drugs.

AMP functionalized curdlan nanoparticles as a siRNA carrier: Synthesis, characterization and targeted delivery via adenosine A2B receptor.

Receptor-mediated endocytosis has been used for tissue targeted delivery of short interfering RNA (siRNA) drugs. Herein, we investigated adenosine receptor (AR) as a candidate for receptor-mediated siRNA internalization. We synthesized adenosine functionalized cationic curdlan derivatives (denote CuAMP polymers). One of these polymers, CuAMP4, efficiently delivered siRNA to breast cancer cells expressing high level of A2B receptor. The internalization of siRNA loaded CuAMP4 by cancer cells was inhibited by free AMP as well as endocytosis inhibitors. Moreover, knockdown of A2BR by siRNA, or pre-treatment of the cells with anti-A2BR antibody, strongly inhibited the cellular uptake of CuAMP4. Our findings confirmed that A2BR can be utilized for cell type specific siRNA delivery, and CuAMP4 NP may be a promising delivery system for cancer cell targeted delivery of therapeutic siRNAs.

Oxidized curdlan activates dendritic cells and enhances antitumor immunity.

Curdlan activates dendritic cells (DCs) and enhances DC-based antitumor immunity. However, hydrophobicity and heterogeneity of curdlan particulates hinder perfect binding of curdlan to dectin-1 receptor, resulting in the reduced activation of antigen presenting cells and limited antitumor effects. Herein, we synthesized partially oxidized curdlan derivative (ß-1,3-polyglucuronic acid, denote PGA). PGA-45 polymer, the reaction product prepared from curdlan by oxidation with 4-acetamido-TEMPO/NaClO/NaClO2 systems under acid conditions for 45 min, activated DCs, induced the expression of co-stimulatory molecules and cytokines, and promoted allogenic T cell proliferation as well as the expression of IL-2. Mechanistically, PGA-45 polymer strongly enhanced phosphorylation of IKK-ß and reduced the expression of phosphorylated Akt, suggesting that PGA-45 may activate multiple cell surface receptors such as TLR4 and dectin-1. Administration of tumor lysate pulsed DCs pre-treated with PGA-45 particles induced strong antitumor activity in B16F10 melanoma model. Our data suggest that PGA-45 have strong adjuvant effects for anti-cancer immunity and the design of PGA polymers may provide insights in the development of novel adjuvants for cancer immunotherapy.

Modulation of Microglia Polarization through Silencing of NF-κB p65 by Functionalized Curdlan Nanoparticle-Mediated RNAi.

Microglia polarization plays an important role in poststroke recovery. Inhibition of proinflammatory (M1) polarization and promotion of anti-inflammatory (M2) polarization of microglia are potential therapeutic strategies for inflammation reduction and neuronal recovery after stroke. Here, we evaluated the central nervous system (CNS)-targeted short interfering RNA (siRNA) delivery ability of functionalized curdlan nanoparticles (CMI) and investigated the nuclear factor-κB (NF-κB) p65 silencing efficiency of CMI-mediated siRNA in microglia, as well as the resulting neuroprotective effect of microglia polarization and neuroprotection in vitro and in vivo. The systemic delivery of NF-κB p65 siRNA (sip65) complexed to CMI nanoparticles in the mouse model of transient middle cerebral artery occlusion (tMCAO) resulted in the distribution of siRNA in microglia and significant silencing in NF-κB p65 in the peri-infarct region. Knockdown of NF-κB p65 resulted in M1 to M2 phenotypic transition of microglia, evidenced by the change in the expression pattern of signature cytokines as well as inducible nitric oxide synthase and CD206. Moreover, the CMI-mediated silencing of p65 increased the density of neurons and decreased pyknosis and edema in the peri-infarct region. Assessment of the neurological deficit score on the Bederson scale revealed a significantly reduced score in the mouse model of tMCAO treated with the sip65/CMI complex. Collectively, our data suggest that CMI nanoparticles are a promising CNS-targeting siRNA delivery system, and NF-κB p65 may be a potential therapeutic target for inflammation reduction and poststroke recovery.

Analysis and Identification of Tumorigenic Targets of MicroRNA in Cancer Cells by Photoreactive Chemical Probes.

Photoactive RNA probes have unique advantages in the identification of microRNA (miR) targets due to their ability for efficient conjugation to the target sequences by covalent crosslinking, providing stable miR-mRNA complexes for further analysis. Here, we report a highly efficient and straightforward method for miR target identification that is based on photo-reactive chemical probes and RNA-seq technology (denotes PCP-Seq). UV reactive probes were prepared by incorporating psoralen in the specific position of the seed sequence of miR. Cancer cells that were transfected with the miR probes were treated with UV, following the isolation of poly(A) RNA and sequencing of the transcriptome. Quantitative analysis of RNA-seq reads and subsequent validation by qPCR, dual luciferase assay as well as western blotting confirmed that PCP-Seq could highly efficiently identify multiple targets of different miRs in the lung cancer cell line, such as targets PTTG1 and PTGR1 of miR-29a and ILF2 of miR-34a. Collectively, our data showed that PCP-Seq is a robust strategy for miR targets identification, and unique in the identification of the targets that escape degradation by miRISC and maintain normal cellular level, although their translation is repressed.

Silencing of STAT3 via Peptidomimetic LNP-Mediated Systemic Delivery of RNAi Downregulates PD-L1 and Inhibits Melanoma Growth.

Cutaneous melanoma is the most aggressive skin cancer with notorious drug resistance. Inhibition of immune checkpoint molecules is one of the most promising approaches for cancer therapy. Herein, we show that RNAi mediated silencing of STAT3 expression in the tumor tissue robustly inhibit tumor growth in B16F10 mouse model of melanoma. We designed a peptidomimetic-based lipid nanoparticles (LNPs) for the delivery of siRNA in mouse model of melanoma. When systemically administered, the novel formulation (denote DoCh) preferentially delivered siRNA to the tumor tissue. Remarkably, sequential intravenous injections of siRNA against STAT3 induced profound silencing of STAT3 expression in tumor tissue, which resulted in significant downregulation of PD-L1, leading to significant inhibition of tumor growth through inhibition of tumor immune checkpoint. Moreover, DoCh-mediated siRNA delivery did not show noticeable damage to the major organs. Collectively, our data demonstrated that DoCh LNP is a promising tumor-targeted siRNA delivery system.

Design of curdlan-based pH-sensitive polymers with endosome buffering functionality for siRNA delivery.

Developing nucleic acid-based tools to control disease-relevant gene expression in human disorders, such as siRNAs, opens up potential opportunities for therapeutics. Because of their high molecular weight and polyanionic nature, synthetic siRNAs fail to cross biological membranes by passive diffusion and therefore, generally require transmembrane siRNA delivery technologies to access the cytoplasm of target cells. To create a biocompatible siRNA delivery agent, we chemically modified natural polysaccharide curdlan derivative 6AC-100 in a regioselective manner to introduce different ratios of imidazole rings in the amino units (denoted as Curimi) and evaluated their siRNA binding ability, cytotoxicity, endosome buffering capacity and siRNA transfection efficiency. The novel curdlan based Curimi polymers formed nanoparticles with siRNA at pH 7.4 in range of 85-105 nm and their size distribution increased along with decreasing pH condition. The zeta potential increased by lowering pH value as well. Curimi polymers showed lower toxicity and higher buffering capacity compared to 6AC-100, and efficiently delivered siRNA against to PLK1 into cancer cells, and subsequently, significantly inhibited target mRNA level. Our result suggested that novel curdlan based Curimi polymers may be used as efficient siRNA carrier for cancer therapy.

New ferrocene-triazole derivatives for multisignaling detection of Cu2+ in aqueous medium and their antibacterial activity.

Ferrocene-based naphthalene or quinoline receptors 1-4 linked by triazole were designed and synthesized. Their recognition properties of metal cations have been investigated systematically in aqueous environment. Upon addition 1 equiv. of Cu2+ ion, receptors 1 (C23H19FeN3O) and 2 (C22H18FeN4O) showed fluorescent turn-off, enhanced absorption and color variations. At the same time, receptor 1 also caused the perturbation of redox potential after addition 1 equiv. of Cu2+ ion. Therefore, receptors 1 and 2 behaved as naked-eye chemosensors and fluorescent probes for Cu2+ without interference by other ions and with low detection limit. In addition, receptor 1 could also be considered electrochemical sensor for Cu2+ having excellent sensitivity and selectivity. However, increasing the molecules flexibility resulted in the lower selectivity of ion recognition in the case of receptors 3 (C24H21FeN3O) and 4 (C23H20FeN4O). Furthermore, this series of compounds were nontoxicity and receptor 1 exhibited certain antibacterial activity.

Alkylation enhances biocompatibility and siRNA delivery efficiency of cationic curdlan nanoparticles.

Cationic curdlan derivatives are a class of promising carriers for nucleic acid delivery including short interfering RNA (siRNA). While our previous studies demonstrated the siRNA delivery efficiency of aminated curdlan derivatives, the associated cytotoxicity issue remained unsolved. To investigate the effects of alkylation on the toxicity as well as the transfection efficiency, we conjugated short alkyl chains to 6-amino-6-deoxy-curdlan (6AC-100). The cytotoxicity of alkylated 6AC-100 derivatives (denote CuVa polymers) decreased with the increase of the degree of substitution (DS). CuVa3, with the highest DS, showed a 50% decreased cytotoxicity compared to 6AC-100 to 6AC-100 at a concentration of 140 µg/mL. The CuVa polymers readily complexed with siRNA to form nanoparticles, and induced significant knockdown of a disease related gene (STAT3) in mouse melanoma cell line B16. However, B16 cells transfected with siSTAT3 complexed to CuVa3 showed the highest phenotypic changes. These findings suggest that CuVa polymers have significantly enhanced biocompatibility and may be a promising delivery system for delivery of therapeutic siRNAs.

Preparation and cell activities of lactosylated curdlan-triornithine nanoparticles for enhanced DNA/siRNA delivery in hepatoma cells.

A long-anticipated cancer therapy would deliver the right type of therapeutic agents to the target in control with minimal systemic toxicity. The purpose of this study was to prepare lactosylated curdlan-triornithine nanocarriers (CTOLs), and target deliver gene to hepatoma cells. Structures and biophysical properties had been elucidated with physical and chemical methods. The results revealed that those functionalized polymers can completely condense the gene into spherical nanoparticles. Cytotoxicity assay, GFP-pDNA and siRNA transfection in vitro were implemented successively. Observations showed that CTOL 20% with the highest lactose acid substitution degree targeted delivered gene into HepG2 cells over expressing ASGPR receptors and had pretty gene knockdown efficiency over 70%. Meanwhile, the carriers showed excellent biocompatibility. Our studies demonstrated that CTOLs with lower toxicity and higher gene binding capacity may serve as a potential valuable platform that can be tailored to target the liver cancer cells for therapeutic gene.

Design of Peptidomimetic Functionalized Cholesterol Based Lipid Nanoparticles for Efficient Delivery of Therapeutic Nucleic Acids.

Lipid nanoparticles (LNP) are the most potent carriers for the delivery of nucleic acid-based therapeutics. The first FDA approved a short interfering RNA (siRNA) drug that uses a cationic LNP system for the delivery of siRNA against human transthyretin (hTTR). However, preparation of such LNP involves tedious multi-step synthesis with relatively low yields. In the present study, we synthesized cationic peptidomimetic functionalized cholesterol (denote Chorn) in straightforward chemical approaches with high yield. When formulated with helper lipids, Chorn LNPs complexed with siRNA to form nanoparticles with an average diameter of 150 nm to 200 nm. Chorn LNP mediated transfection of a green fluorescence protein (GFP) expressing plasmid resulted in 60% GFP positive cells. Moreover, Chorn LNP delivered siRNA against polo-like kinase 1 (Plk1), a disease related gene in cancer cells and efficiently suppressed the expression of the gene, resulting in significant morphological changes in the cell nuclei. Our data suggested that cholesterol based cationic LNP, prepared through a robust chemical strategy, may provide a promising siRNA delivery system.

Receptor-mediated delivery of therapeutic RNA by peptide functionalized curdlan nanoparticles.

Natural carbohydrate polymer-based nanoparticles have great biocompatibility that is required for the safe delivery of various drugs including nucleic acid therapeutics. Herein, we designed curdlan-based nanoparticles for cancer cell targeted delivery of short interfering RNA (siRNA). iRGD peptide conjugated 6-amino-6-deoxy curdlan specifically delivered siRNA to integrin expressing cancer cells. Incubation of cancer cells with free iRGD peptide competitively blocked cellular uptake of the iRGD functionalized curdlan nanoparticles. Chloroquine but not nystatin inhibited cellular uptake of iRGD functionalized curdlan nanoparticles, indicating that the iRGD peptide conjugated curdlan nanoparticles were internalized through the receptor (clathrin)-mediated endocytosis. Moreover, a disease related gene Plk1 was substantially knocked down by siRNA carried by 6AC-iRGD nanoparticles in HepG2 cells. Our data suggested that iRGD functionalized curdlan may provide a biocompatible carrier for siRNA delivery.

Traditional Mongolian medicine Eerdun Wurile improves stroke recovery through regulation of gene expression in rat brain.

ETHNOPHARMACOLOGICAL RELEVANCE: Eerdun Wurile (EW) is one of the key Mongolian medicines for treatment of neurological and cardiological disorders. EW is ranked most regularly used Mongolian medicine in clinic. Components of EW which mainly originate from natural products are well defined and are unique to Mongolian medicine. AIM OF THE STUDY: Although the recipe of EW contains known neuroactive chemicals originated from plants, its mechanism of action has never been elucidated at molecular level. The objective of the present study is to explore the mechanism of neuroregenerative activity of EW by focusing on the regulation of gene expression in the brain of rat model of stroke. MATERIALS AND METHODS: Rat middle cerebral artery occlusion (MCAO) models were treated with EW for 15 days. Then, total RNAs from the cerebral cortex of rat MCAO models treated with either EW or control (saline) were extracted and analyzed by transcriptome sequencing. Differentially expressed genes were analyzed for their functions during the recovery of ischemic stroke. The expression level of significantly differentially expressed genes such as growth factors, microglia markers and secretive enzymes in the lesion was further validated by RT-qPCR and immunohistochemistry. RESULTS: Previously identified neuroactive compounds, such as geniposide (Yu et al., 2009), myristicin (Shin et al., 1988), costunolide (Okugawa et al., 1996), toosendanin (Shi and Chen, 1999) were detected in EW formulation. Bederson scale indicated that the treatment of rat MCAO models with EW showed significantly lowered neurological deficits (p < 0.01). The regional cerebral blood circulation was also remarkably higher in rat MCAO models treated with EW compared to the control group. A total of 186 genes were upregulated in the lesion of rat MCAO models treated with EW compared to control group. Among them, growth factors such as Igf1 (p < 0.05), Igf2 (p < 0.01), Grn (p < 0.01) were significantly upregulated in brain after treatment of rat MCAO models with EW. Meanwhile, greatly enhanced expression of microglia markers, as well as complementary components and secretive proteases were also detected. CONCLUSION: Our data collectively indicated that EW enhances expression of growth factors including Igf1 and Igf2 in neurons and microglia, and may stimulate microglia polarization in the brain. The consequences of such activity include stimulation of neuron growth, hydrolysis and clearance of cell debris at the lesion, as well as the angiogenesis.

Design of Mannose-Functionalized Curdlan Nanoparticles for Macrophage-Targeted siRNA Delivery.

6-Amino-6-deoxy-curdlan is a promising nucleic acid carrier that efficiently delivers plasmid DNA as well as short interfering RNA (siRNA) to various cell lines. The highly reactive C6-NH2 groups of 6-amino-6-deoxy-curdlan prompt conjugation of various side groups including tissue-targeting ligands to enhance cell-type-specific nucleic acid delivery to specific cell lines. Herein, to test the primary-cell-targeting efficiency of the curdlan derivative, we chemically conjugated a macrophage-targeting ligand, mannose, to 6-amino-6-deoxy-curdlan. The resulting curdlan derivative (denoted CMI) readily complexed with siRNA and formed nanoparticles with a diameter of 50-80 nm. The CMI nanoparticles successfully delivered a dye-labeled siRNA to mouse peritoneal macrophages. The delivery efficiency was blocked by mannan, a natural ligand for a macrophage surface mannose receptor (CD206), but not by zymosan, a ligand for the dectin-1 receptor, which is also present on the surface of macrophages. Moreover, CMI nanoparticles were internalized by macrophages only at 37 °C, suggesting that the cellular uptake of CMI nanoparticles was energy-dependent. Furthermore, CMI nanoparticle efficiently delivered siRNA against tumor necrosis factor α (TNFα) to lipopolysaccharide-stimulated primary mouse peritoneal macrophages. In vivo experiments demonstrated that CMI nanoparticles successfully delivered siTNFα to mouse peritoneal macrophages, liver, and lung and induced significant knockdown of the TNFα expression at both messenger RNA and protein levels. Therefore, our design of CMI may be a promising siRNA carrier for targeting CD206-expressing primary cells such as macrophage and dendritic cells.

Fabrication of functional hollow microspheres constructed from MOF shells: Promising drug delivery systems with high loading capacity and targeted transport.

An advanced multifunctional, hollow metal-organic framework (MOF) drug delivery system with a high drug loading level and targeted delivery was designed and fabricated for the first time and applied to inhibit tumour cell growth. This hollow MOF targeting drug delivery system was prepared via a simple post-synthetic surface modification procedure, starting from hollow ZIF-8 successfully obtained for the first time via a mild phase transformation under solvothermal conditions. As a result, the hollow ZIF-8 exhibits a higher loading capacity for the model anticancer drug 5-fluorouracil (5-FU). Subsequently, 5-FU-loaded ZIF-8 was encapsulated into polymer layers (FA-CHI-5-FAM) with three components: a chitosan (CHI) backbone, the imaging agent 5-carboxyfluorescein (5-FAM), and the targeting reagent folic acid (FA). Thus, an advanced drug delivery system, ZIF-8/5-FU@FA-CHI-5-FAM, was fabricated. A cell imaging assay demonstrated that ZIF-8/5-FU@FA-CHI-5-FAM could target and be taken up by MGC-803 cells. Furthermore, the as-prepared ZIF-8/5-FU@FA-CHI-5-FAM exhibited stronger cell growth inhibitory effects on MGC-803 cells because of the release of 5-FU, as confirmed by a cell viability assay. In addition, a drug release experiment in vitro indicated that ZIF-8/5-FU@FA-CHI-5-FAM exhibited high loading capacity (51%) and a sustained drug release behaviour. Therefore, ZIF-8/5-FU@FA-CHI-5-FAM could provide targeted drug transportation, imaging tracking and localized sustained release.

Design of Highly Potent Lipid-Functionalized Peptidomimetics for Efficient in Vivo siRNA Delivery.

RNA interference (RNAi) is a highly efficient approach for gene silencing. Regulation of gene expression at post-transcriptional level provides great potential for curing diseases caused by abnormal overexpression of disease-related genes. However, the application of RNAi in the clinic has been hindered by the lack of efficient and biocompatible delivery systems. Therefore, the development of a safe and tissue-targeted double-stranded interfering RNA (siRNA) carrier for clinical application is urgently needed. Here we report the discovery of a highly efficient liposomal siRNA delivery agent based on a novel peptidomimetic built from natural amino acids. Fine tuning of the composition of amino acids, the type of amide linkage in the peptidomimetic, as well as the formulation and the physicochemical parameters of the novel lipoplex resulted in a lipid nanoparticle (LNP) that efficiently encapsulates and carries siRNA to mouse liver. In vivo experiments showed that a single injection of unmodified siRNA complexed to one of the peptidomimetics at a clinically feasible dose induced significant RNAi in mouse liver, resulting in a 90% decrease in apolipoprotein B (ApoB) mRNA level, as well as a 60% decrease in serum ApoB protein level. Analysis of mouse serum by ELISA indicated that the novel peptidomimetic based lipoplex did not elevate the level of liver enzymes (ALT, AST) in the serum. Our novel peptidomimetic-based lipoplex demonstrated great potential for the development of a safe and efficient siRNA delivery agent for clinical applications.

PEGylation of 6-amino-6-deoxy-curdlan for efficient in vivo siRNA delivery.

RNA interference (RNAi) is an evolutionarily conserved gene-silencing phenomenon that shows great promise for developing new therapies. However, the development of small interfering RNA (siRNA)-based therapies need to establish efficient delivery system that silences target genes with siRNA doses that is clinically feasible in humans. Here we report synthesis and in vivo study of a novel PEGylated curdlan-based nanoparticle, designated as 6AC-100PEG, obtained by conjugation of mPEG 2000 to 6-amino-6-deoxy-curdlan. The complex of siRNA/6AC-100PEG showed homogenous nanoparticles with an average diameter of 200nm. MTT assay indicated that 6AC-100PEG does not have apparent cytotoxicity. Systemic administration of a complex of siapoB/6AC-100PEG significantly reduced the level of apoB mRNA in mouse liver, indicating that 6AC-100PEG can efficiently deliver siRNA to mouse liver and induce RNAi. Administration of siRNA/6AC-100PEG to mouse did not elevate liver enzyme level in the serum, indicating that 6AC-100PEG nanoparticle is a promising in vivo siRNA delivery agent.