Fatty Acid-Based Strategy for Efficient Brain Targeted Gene Delivery

Purpose

To investigate a fatty acid-based strategy for efficient brain targeted gene delivery and to understand mechanism(s) of this small molecule-mediated brain gene delivery strategy.

Methods

A series of fatty acids (FAs) were conjugated with polyethylenimine (PEI_25k). A near-infrared fluorescence probe, IR820, was used to study in vivo and ex vivo brain targeting ability of these fatty acid-PEI conjugates (FA-PEIs). Brain uptake of FA-PEI_25k/rhodamine-6-isothiocyanate (RITC)-labeled DNA nanoparticles was investigated via a fluorescence imaging method. Moreover, pEGFP was used as a model gene to study in vitro and in vivo transfection effect of the ideal FA-PEI_25k conjugate.

Results

FA modification did not have interference with the complexation between DNA and the PEI_25k. The FA-PEI_25k conjugates showed excellent brain targeting ability compared with unmodified PEI_25k. Among these FA-PEI_25k conjugates studied, myristic acid (MC)-PEI_25k showed sustained brain distribution profile and higher brain DNA uptake. Furthermore, MC-PEI_25k/pEGFP nanoparticles was able to achieve efficient in vitro and in vivo gene transfection. GFP expression was observed at different brain regions in vivo.

Conclusions

These results demonstrated that the small molecule fatty acid, particularly myristic acid-based brain gene delivery strategy, is promising to mediate efficient gene transfection in the brain.     

Folate and TAT Peptide Co-Modified Liposomes Exhibit Receptor-Dependent Highly Efficient Intracellular Transport of Payload In Vitro and In Vivo

Purpose

Using different chain lengths of PEG as linkers to develop a novel folate (FA) and TAT peptide co-modified doxorubicin (DOX)-loaded liposome (FA/TAT-LP-DOX) and evaluate its potential for tumor targeted intracellular drug delivery.

Methods

FA/TAT-LP-DOX was prepared by pH gradient method and post-insertion method and the optimal ligand density was screened by MTT assay. In vitro evaluation was systematically performed through cytotoxicity assay, cellular uptake studies, subcellular localization and cellular uptake mechanism in folate receptor (FR) over-expressing KB tumor cells. In vivo tumor targeted delivery of FA/TAT-LP-DOX was also studied by in vivo fluorescence imaging in a murine KB xenograft model.

Results

The particle size and zeta potential determination indicated that FA and TAT were successfully inserted into the liposome and cationic TAT peptide was completely shielded. With the optimal ligand density (5% of FA and 2.5% TAT), the FA/TAT-LP-DOX exhibited improved cytotoxity and cellular uptake efficiency compared with its single-ligand counterparts (FA-LP-DOX and PEG/TAT-LP-DOX). Competitive inhibition and uptake mechanism experiments revealed that FA and TAT peptide played a synergistic effect in facilitating intracellular transport of the liposome, and association between FA and FA receptors activated this transport process. In vivo imaging further demonstrated the superiority of FA/TAT-LP in tumor targeting and accumulation.

Conclusions

Folate and TAT peptide co-modified liposome using different chain lengths of PEG as linkers may provide a useful strategy for specific and efficient intracellular drug delivery.     

Phosphatidylinositol 3-kinase Inhibitor (PIK75) Containing Surface Functionalized Nanoemulsion for Enhanced Drug Delivery, Cytotoxicity and Pro-apoptotic Activity in Ovarian Cancer Cells

Purpose

Ovarian cancer is a debilitating disease, which needs multi-pronged approach of targeted drug delivery and enhanced efficacy with the use of combination therapeutics. In this study, we have examined the anticancer activity of PIK75 incorporated in surface functionalized nanoemulsions for targeted delivery to SKOV-3 cells. A pro-apoptotic molecule C₆-ceramide was also co-delivered to augment therapeutic efficacy.

Methods

EGFR and FR functionalized nanoemulsions incorporating PIK75 and C₆-ceramide were characterized for particle size, surface charge, entrapment efficiency and morphology. Fluorescence and quantitative uptake studies were conducted in SKOV-3 cells to determine intracellular distribution. Cell viability was assessed using MTT assay while mechanism of cytotoxicity was evaluated using capsase-3/7, TUNEL and hROS assay.

Results

Cytotoxicity assay showed 57% decrease in IC₅₀ value of PIK75 following treatment with EGFR targeted nanoemulsion and 40% decrease following treatment with FR targeted nanoemulsion. Combination therapy with PIK75 and ceramide enhanced the cytotoxicity of PIK75 compared to therapy with individual formulations. The increase in cytotoxicity was attributed to increase in cellular apoptosis and hROS activity.

Conclusion

The results of this study showed that the targeted system improved cytotoxicity of PIK75 compared to the non-targeted system. Combination therapy with ceramide augmented PIK75’s therapeutic activity.     

Solidified Self-Nanoemulsifying Formulation for Oral Delivery of Combinatorial Therapeutic Regimen: Part II In vivo Pharmacokinetics,Antitumor Efficacy and Hepatotoxicity

Purpose

The present work focuses on the in vivo evaluation of tamoxifen and quercetin combination loaded into solid self-nanoemulsifying drug delivery system (s-Tmx-QT-SNEDDS).

Methods

Lyophilization was employed to prepare s-Tmx-QT-SNEDDS using Aerosil 200 as carrier. The developed formulation was evaluated for in vitro cell cytotoxicity, in vivo pharmacokinetics, antitumor efficacy and toxicity studies.

Results

In vivo pharmacokinetics revealed ~8-fold and ~4-fold increase in oral bioavailability of tamoxifen and quercetin, respectively as compared to free counterparts. s-Tmx-QT-SNEDDS exhibited significantly higher cell cytotoxicity, as compared to free drug combination revealing ~32-fold and ~22-fold higher dose reduction index for tamoxifen and quercetin, respectively estimated using median effect dose analysis. s-Tmx-QT-SNEDDS could suppress tumor growth in DMBA induced tumor bearing animals by ~80% in contrast to ~35% observed with tamoxifen citrate. The significant appreciation in antitumor efficacy was further supported by normalized levels of tumor angiogenesis markers (MMP-2 and MMP-9). Finally, complete obliteration in tamoxifen induced hepatotoxicity was observed upon administration of developed formulation in contrast to that of clinically available tamoxifen citrate when measured as function of hepatotoxicity markers and histopathological changes.

Conclusions

In nutshell, co-encapsulation of quercetin with tamoxifen in solid SNEDDS poses great potential in improving the therapeutic efficacy and safety of tamoxifen.     

Bioreducible Polypeptide Containing Cell-Penetrating Sequence for Efficient Gene Delivery

Purpose

To design excellent polypeptide-based gene vectors and determine the gene delivery efficiency.

Methods

Polypeptides (designated as xPolyK₆, xPolyK₆-R₈1 and xPolyK₆-R₈2), comprising the DNA condensing and buffering peptide HK₆H as well as cell penetrating peptide (CPP) R₈ were obtained by the oxidative polymerization of CHK₆HC and CR₈C at different molar ratios in 4 mL phosphate-buffered saline (PBS) containing 30% (v/v) DMSO at room temperature for 96 h. The cytotoxicity of vectors was studied by MTT assay. Moreover, particle size, zeta potential and morphology along with the in vitro transfection efficiency and cellular uptake of vector/plasmid DNA (pDNA) complexes were characterized at various w/w ratios to determine their potential in gene therapy.

Results

All the vectors presented excellent ability of binding and condensing pDNA, additionally with low cytotoxicity. Simultaneously, transfection efficiency of the vectors appeared apparent dependence on the vector composition. The distinct correlation between the content of CR₈C with the transfection efficiency demonstrated the effective improvement in transfection efficacy by the oxidative polymerization. Particularly, xPolyK₆-R₈2 possessed the highest transfection efficiency at a w/w ratio of 50. Furthermore, xPolyK₆-R₈2 also presented the best cellular uptake capability demonstrated by confocal microscopy and flow cytometry.

Conclusions

Bioreducible polypeptides incorporating with proper amount of CPP are promising as effective non-viral gene vectors in gene therapy.     

Asymmetric Peptide Dendrimers are Effective Linkers for Antibody-Mediated Delivery of Diverse Payloads to B Cells in Vitro and in Vivo

Purpose

Safe, targeted delivery of therapeutics remains a focus of drug/gene delivery, the aim being to achieve optimal efficacy while minimising off-target delivery. Dendrimers have a vast array of potential applications and have great potential as gene and drug delivery tools. We previously reported the development of peptide dendrimers that effectively complexed DNA and that have distinct advantages over conventional spherical dendrimers. Here, to expand the application of peptide-based low generation dendrimers we tested their capacity to be transformed into linkers for antibody-based targeting of diverse payloads.

Methods

Peptide-based low-generation asymmetric dendrimers were generated and conjugated to partially-reduced antibodies specific for B cell surface antigens or an irrelevant antigen. Preservation of antigen binding by the antibodies and targeting of the conjugated dendrimers carrying a small molecule (biotin) or plasmid DNA payloads was tested.

Results

Peptide-based low generation dendrimers were efficiently and site-specifically conjugated to antibodies with retention of antigen-binding capacity. Altering the branching termini of dendrimers facilitated delivery of diverse payloads in vitro and in vivo.

Conclusions

We propose that safe, non-toxic peptide dendrimers, which are readily synthesised and modifiable for a variety of applications, form the basis of a new family of biocompatible “linkers” with substantial potential for targeted delivery applications.     

Improving Intracellular Doxorubicin Delivery Through Nanoliposomes Equipped with Selective Tumor Cell Membrane Permeabilizing Short-Chain Sphingolipids

Purpose

To improve nanoliposomal-doxorubicin (DoxNL) delivery in tumor cells using liposome membrane-incorporated short-chain sphingolipids (SCS) with selective membrane-permeabilizing properties. DoxNL bilayers contained synthetic short-chain derivatives of known membrane microdomain-forming sphingolipids; C₈-glucosylceramide (C₈-GluCer), C₈-galactosylceramide (C₈-GalCer) or C₈-lactosylceramide (C₈-LacCer).

Methods

DoxNL enriched with C₈-GluCer or C₈-GalCer were developed, optimized and characterized with regard to size, stability and drug retention. In vitro cytotoxic activity was studied in a panel of human tumor cell lines and normal cells. Intracellular Dox delivery was measured by flow cytometry and visualized by fluorescence microscopy. For a further understanding of the involved drug delivery mechanism confocal microscopy studies addressed the cellular fate of the nanoliposomes, the SCS and Dox in living cells.

Results

C₈-LacCer-DoxNL aggregated upon Dox loading. In tumor cell lines SCS-DoxNL with C₈-GluCer or C₈-GalCer demonstrated strongly increased Dox delivery and cytotoxicity compared to standard DoxNL. Surprisingly, this effect was much less pronounced in normal cells. Nanoliposomes were not internalized, SCS however transfered from the nanoliposomal bilayer to the cell membrane and preceded cellular uptake and subsequent nuclear localization of Dox.

Conclusion

C₈-GluCer or C₈-GalCer incorporated in DoxNL selectively improved intracellular drug delivery upon transfer to tumor cell membranes by local enhancement of cell membrane permeability.     

Modulation of Polyplex Release from Biodegradable Microparticles through Poly(ethylenimine) Modification and Varying Loading Concentration

Purpose

This work investigates the effects of hyaluronic acid (HA) conjugated onto branched poly(ethylenimine) (bPEI) and varying loading concentrations of these polymers complexed with DNA on their release from poly(DL-lactic-co-glycolic acid) (PLGA) microparticles and the transfection of target cells.

Methods

To examine the effect of alteration of the gene delivery polymer on the system, we observed the morphology, size, loading efficiency, polymer and DNA release, and the transfection efficiency for the microparticles formed with three internal phase loading concentrations during microparticle formation.

Results

Addition of HA to this vector allowed for increased loading concentration within these systems and significantly altered release kinetics without changing the morphology of the particles. The incorporation of HA onto the bPEI backbone significantly increased the transfection efficiency of the complexes released from the corresponding microparticle formulation.

Conclusions

The results show that the modification of bPEI with HA and the concentration of loaded polymer/DNA complexes can significantly alter the entrapment and release profiles from PLGA microparticles. This is significant in that it offers insight into the effects of modification of gene delivery vectors on a controlled release system designed to achieve a sustained therapeutic response.     

EphA2 Targeting Pegylated Nanocarrier Drug Delivery System for Treatment of Lung Cancer

Purpose

Evaluation of tumor targeting pegylated EphA2 peptide coated nanoparticles (ENDDs) of a novel anticancer agent DIM-C-pPhC₆H₅ (DIM-P) and Docetaxel (DOC) and investigate its antitumor activity and potential for treatment of lung cancer.

Methods

Nanoparticles were prepared with DIM-P and DOC (NDDs) using Nano-DeBEE. ENDDs were prepared by conjugating NDDs with 6His-PEG2K-EphA2 peptide and characterized for physicochemical properties, binding assay, cytotoxicity, cellular uptake studies, drug release and pharmacokinetic parameters. Anti-tumor activity of ENDDs was evaluated using a metastatic H1650 and orthotopic A549 tumor models in nude mice and tumor tissue were analyzed by RT-PCR and immunohistochemistry.

Results

Particle size and entrapment efficiency of ENDDs were 197?±?21 nm and 95?±?2%. ENDDs showed 32.5?±?3.5% more cellular uptake than NDDs in tumor cells. ENDDs showed 23?±?3% and 26?±?4% more tumor reduction compared to NDDs in metastatic and orthotopic tumor models, respectively. In-vivo imaging studies using the Care stream MX FX Pro system showed (p?Conclusions The results emanating from these studies demonstrate anti-cancer potential of DIM-P and the role of ENDDs as effective tumor targeting drug delivery systems for lung cancer treatment.     

In Vitro and In Vivo Characterisation of PEG-Lipid-Based Micellar Complexes of Salmon Calcitonin for Pulmonary Delivery

Purpose

To investigate DSPE-PEG₂₀₀₀-based micellar formulations of salmon calcitonin (sCT) for their ability to improve pulmonary delivery.

Methods

Micelles were characterised by DLS and ³¹P-NMR spectroscopy. Stability against sCT degrading peptidases, trypsin, ??-chymotrypsin and neutrophil elastase as well as their influence on transepithelial absorption was investigated in vitro. In vivo perfomance of sCT micelles was studied in an experimental model of intratracheal aerosolisation into rats.

Results

Micelles with a mean hydrodynamic diameter of 12?nm spontaneously assembled, when a total concentration of 0.02?mM of PEG-lipid and sCT (at 1:1 molar ratio) was exceeded. Nuclear magnetic resonance confirmed the presence of small micellar structures. The micellar formulation showed increased stability against enzymatic digestion. In vitro studies also showed that sCT micelles were able to enhance transepithelial absorption. Data obtained from in vivo experiments provided evidence of significantly (P?AUC _inf and a relative bioavailability of 160?±?55% when compared to plain sCT solution.

Conclusions

The herein described PEG-lipid micelles are promising carriers for enhanced pulmonary delivery of sCT.     

Drug Delivery Using Platelet Cancer Cell Interaction

Purpose

To develop an efficient biocompatible and targeted drug delivery system in which platelets, an essential blood component having a natural affinity for cancer cells, are used as carrier of anticancer drug as delivery of drug to the targeted site is crucial for cancer treatment.

Methods

Doxorubicin hydrochloride, a potent anti cancer drug, was delivered in lung adenocarcinoma cell line (A549) using platelet as a delivery agent. This delivery mode was also tested in Ehrlich ascites carcinoma (EAC) bearing mice in presence and absence of platelets.

Results

The results show that platelets can uptake the drug and release the same upon activation. The efficiency of drug loaded platelets in inducing cytotoxicity was significantly higher in both in vitro and in vivo model, as compared to the free drug.

Conclusions

The proposed drug delivery strategy may lead to clinical improvement in the management of cancer treatment as lower drug concentration can be used in a targeted mode. Additionally the method can be personalized as patient's own platelet can be used for deliver various drugs.     

Hyaluronidase Enzyme-responsive Targeted Nanoparticles for Effective Delivery of 5-Fluorouracil in Colon Cancer

Purpose

In this study, we have successfully prepared the hyaluronic acid (HA)-conjugated mesoporous silica nanoparticles loaded with 5-fluorouracil (5-FU) to increase the anticancer efficacy in colon cancers.

Methods

The particles were nanosized and perfectly spherical. In vitro release kinetics clearly showed the enzyme-sensitive release of 5-FU from HA-conjugated 5-FU loaded mesoporous silica nanoparticles (HA/FMSN).

Results

The presence of HA on the surface of nanoparticles targeted the CD44 receptors overexpressed in the colon cancer cells In vitro cell viability and apoptosis assay clearly showed the superior anticancer effect of HA/FMSN in HT29 colon cancer cells. HA/FMSN exhibited a remarkably higher 43% of cells in early apoptosis phase and 55% of cells in late apoptosis phase indicating the superior anticancer effect of HA/FMSN. HA/FMSN exhibited a significant reduction in the tumor burden compared to that of any group. HA/FMSN was 3-fold more effective than free drug and 2-fold more effective than -FU loaded mesoporous silica nanoparticles (FMSN).

Conclusions

Overall, results suggest that the novel delivery strategy could hold enormous potential in colon cancer targeting.

     

Cellular Uptake Mechanism and Therapeutic Utility of a Novel Peptide in Targeted-Delivery of Proteins into Neuronal Cells

Purpose

The peptide-based delivery system constitutes a potent approach to overcome the limitations of drug delivery in vitro and in vivo. We recently proposed a novel peptide RDP, which enables brain-targeting delivery of proteins into neuronal cells. Here we investigate the possible internalization mechanism of RDP, and identify the therapeutic effects of functional proteins when linked with RDP in brain disease.

Methods

The RDP fusion proteins are produced through recombinant DNA technology, and cell culture is used to investigate the uptake mechanism of RDP and its fusion protein. Experimental Parkinson’s disease (PD) model is prepared in mice by intra-striatal injection of 6-hydroxydopamine, and is tested by apomorphine- and amphetamine-induced rotation.

Results

The results suggest that the possible route for RDP cellular uptake might involve GABA receptor-dependent, clathrin-mediated endocytosis pathway. Additionally, the conjugate of RDP and glial cell-derived neurotrophic factor (GDNF) exhibits the neuroprotective effect in experimental PD animals, including reduction of apomorphine- and amphetamine-induced rotation following toxin administration.

Conclusions

RDP may become an effective tool for the targeted delivery of proteins into brain for disease treatment.     

Hyaluronic Acid Layer-By-Layer (LbL) Nanoparticles for Synergistic Chemo-Phototherapy

Purpose

The aim of this study was to design hyaluronic acid (HA) layer-by-layer (LbL) nanoparticles, which carried paclitaxel (PTX) and Indocyanine green (ICG) to both tumor cells and tumor associated cells to achieve synergistic chemo-photothermal therapeutic effect.

Methods

The LbL-engineered nanoparticles (PDIH) were prepared by dopamine self-polymerization on PTX nanocrystal to form thin, surface-adherent polydopamine (PDA) films, which subsequently absorbed ICG and HA. The tumor cell and tumor associated cell targeting and antitumor efficacy of PDIH were investigated both in vitro an in vivo using 4 T1 murine mammary cancer cell lines and mice bearing orthotopic 4 T1 breast tumor.

Results

PDIH presented a long-rod shape in TEM and showed enhanced photothermal effect and cytotoxicity upon NIR laser irradiation both in vitro and in vivo. PDIH also displayed high target ability to CD44 overexpressed tumor cells and tumor associated cells mediated by HA. In vivo antitumor study indicated that PDIH therapeutic strategy could achieve remarkable antitumor efficacy.

Conclusion

PDIH showed excellent tumor-targeting property and chemo-photothermal therapeutic efficacy.

     

Targeted Delivery of Proteins into the Central Nervous System Mediated by Rabies Virus Glycoprotein-Derived Peptide

Purpose

Delivery of therapeutic proteins across the blood-brain barrier (BBB) is severely limited by their size and biochemical properties. Here we showed that a 39-amino acid peptide derived from the rabies virus glycoprotein (RDP) was exploited as an efficient protein carrier for brain-targeting delivery.

Methods

Three proteins with different molecular weight and pI, ??-galactosidase (??-Gal), luciferase (Luc) and brain-derived neurotrophic factor (BDNF), were fused to RDP and intravenously injected into the mice respectively. The slices of different tissues with X-Gal staining were used to examine whether RDP could deliver ??-Gal targeted into the CNS. The time-course relationship of RDP-Luc was studied to confirm the transport efficiency of RDP. The neuroprotective function of RDP-BDNF was examined in mouse experimental stroke to explore the pharmacological effect of RDP fusion protein.

Results

The results showed that the fusion proteins rapidly and specific entered the nerve cells in 15?min, and the t_1/2 was about 1?hr. Furthermore, RDP-BDNF fusion protein showed the neuroprotective properties in mouse experimental stroke including reduction of stroke volume and neural deficit.

Conclusions

RDP provides an effective approach for the targeted delivery of biological active proteins into the central nervous system.     

Design and <Emphasis Type="BoldItalic">In Vitro</Emphasis> Evaluation of Bispecific Complexes and Drug Conjugates of Anticancer Peptide,LyP-1 in Human Breast Cancer

Purpose

LyP-1, a nine-amino-acid tumor homing peptide, selectively binds to its cognate receptor, p32. Overexpression of p32 in certain tumors should allow use of LyP-1 as a targeting agent for the delivery of therapeutic or diagnostic agents. Peptide conjugates are developed for enhanced pre-targeting of MDA-MB-231 breast cancer cells with peptide-antibody bispecific complexes and targeting with multiple-drug/-fluorophore-conjugated nano-polymers.

Methods

LyP-1-anti-DTPA bispecific antibody complexes (LyP-1-bsAbCx) were generated by conjugation of anti-DTPA antibody and LyP-1. LyP-1–doxorubicin (Dox), Dox-DTPA-succinyl-polylysine (Dox-DSPL), Dox-DSPL-LyP-1, DTPA-Dox-poly glutamic acid (D-Dox-PGA) or DTPA-rhodamine conjugated polylysine (DSPL-RITC) were prepared. In vitro therapeutic efficacy and targeting by immunofluorescence in MDA-MB-231 breast cancer cells were assessed with Dox-LyP-1. Immunofluorescence visualization of cancer cells was evaluated after pretargeting with LyP-1-bsAbCx and targeting with DSPL-RITC.

Results

Cytotoxicity of Dox-LyP-1 conjugates was significantly greater than free doxorubicin (p?<?0.0001). For fluorescent-labeled LyP-1, internalization occurred in 30 min in tumor cells. Fluorescence intensity of two-step targeted cells showed that pretargeting with LyP-1-bsAbC, followed by targeting with DSPL-RITC was greater than non-pretargeted DSPL-RITC (p?<?0.05).

Conclusions

Peptide-conjugates are effective targeting agents for MDA-MB-231 breast cancer cells in culture. LyP-1-bsAbCx and Dox-LyP-1 conjugates may allow development of novel targeted cancer therapy and diagnosis.

     

Co-transfection Gene Delivery of Dendritic Cells Induced Effective Lymph Node Targeting and Anti-tumor Vaccination

Purpose

Successful genetically engineered Dendritic Cell (DC) can enhance DC’s antigen presentation and lymph node migration. The present study aims to genetically engineer a DC using an efficient non-viral gene delivery vector to induce a highly efficient antigen presentation and lymph node targeting in vivo.

Methods

Spermine-dextran (SD), a cationic polysaccharide vector, was used to prepare a gene delivery system for DC engineering. Transfection efficiency, nuclear trafficking, and safety of the SD/DNA complex were evaluated. A vaccine prepared by engineering DC with SD/gp100, a plasmid encoding melanoma-associated antigen, was injected subcutaneously into mice to evaluate the tumor suppression. The migration of the engineered DCs was also evaluated in vitro and in vivo.

Results

SD/DNA complex has a better transfection behavior in vitro than commercially purchased reagents. The DC vaccine co-transfected with plasmid coding CCR7, a chemokine receptor essential for DC migration, and plasmid coding gp100 displayed superior tumor suppression than that with plasmid coding gp100 alone. Migration assay demonstrated that DC transfected with SD/CCR7 can promote DC migration capacity.

Conclusions

The study is the first to report the application of nonviral vector SD to co-transfect DC with gp100 and CCR7-coding plasmid to induce both the capacity of antigen presentation and lymph node targeting.     

Fucosylated Multiwalled Carbon Nanotubes for Kupffer Cells Targeting for the Treatment of Cytokine-Induced Liver Damage

Purpose

To develop, characterize and exploring the sulfasalazine loaded fucoyslated multi walled carbon nanotubes for Kupffer cell targeting for effective management of cytokine-induce liver damage.

Methods

Sulfasalazine was loaded into the fucosylated MWCNTs after subsequential functionalization (carboxylation, acylation and amidation) using dialysis membrane technique. The in vitro, in vivo studies were performed on macrophages J 774 cell line for Kupffer cells targeting for the treatment of cytokine-induced liver damage.

Results

The loading of SSZ into SSZ-FUCO-MWCNTs was 87.77?±?0.11% (n?=?3). Sustained release was obtained from SSZ-FUCO-MWCNTs, with 89.12?±?0.71% of SSZ released into medium at 48th hr. SSZ-FUCO-MWCNTs showed the 9.01?±?0.23% hemolysis was drastically reduced from 21.62?±?0.24% SSZMWCNTs 21.62?±?0.24%. In SRB assay, SSZ-FUCO-MWCNTs showed more cytotoxicity than raw and SSZ-MWCNTs. In cytokine assay, SSZ- FUCO-MWCNTs exhibited significantly higher inhibition of IL-12 p40 secretion. In Western blot assay, SSZ-FUCO-MWCNTs significantly inhibit NF-κB activation.

Conclusion

The results suggested that the SSZ-FUCO-MWCNTs may be useful nano-carriers for targeted delivery to Kupffer cells in the treatment of cytokine-induced liver damage.     

Nanobubble Ultrasound Contrast Agents for Enhanced Delivery of Thermal Sensitizer to Tumors Undergoing Radiofrequency Ablation

Purpose

Pluronic has been shown to sensitize various tumor cell lines to chemotherapy and hyperthermia by altering the membrane fluidity, depleting ATP, and modulating the heat shock protein 70 expression. In our prior work, Pluronic was also used to formulate nanosized ultrasound contrast agents. In the current study we evaluate the use of these contrast agents as vehicles for image-guided delivery of Pluronic to improve outcomes of tumor radiofrequency (RF) ablation.

Methods

Lipid-shelled Pluronic nanobubbles were prepared and examined for size distribution, zeta potential, stability, biodistribution, accumulation of nanobubbles in the tumor, and treatment efficacy. LS174-T xenograft tumor-bearing mice were used to evaluate tumor growth suppression and measure treatment efficacy after RF ablation.

Results

The average diameter of Pluronic bubbles was 230 nm, and initial bubble echogenicity was 16 dB. In vitro, cells exposed to Pluronic nanobubbles exhibited low cytotoxicity in the absence of ultrasound, even if heat (43?ºC) was applied. When the cells were exposed to Pluronic nanobubbles, heat, and ultrasound; viability was significantly reduced. In vivo, tumors treated with ultrasound-modulated nanobubbles prior to RF ablation showed a significant reduction in growth compared to the RF alone (P?<?0.05).

Conclusion

Lipid and Pluronic-shelled, echogenic nanobubbles combined with ultrasound modulation can serve as an effective theranostic method for sensitization of tumors to RF ablation.