Elucidating the role of free polycations in gene knockdown by siRNA polyplexes.

Unlabelled: Future improvements of non-viral vectors for siRNA delivery require better understanding of intracellular processing and vector interactions with target cells. Here, we have compared the siRNA delivery properties of a lipid derivative of bPEI 1.8kDa (DOPE-PEI) with branched polyethyleneimine (bPEI) with average molecular weights of 1.

EGFR Targeted Theranostic Nanoemulsion for Image-Guided Ovarian Cancer Therapy.

Purpose: Platinum-based therapies are the first line treatments for most types of cancer including ovarian cancer. However, their use is associated with dose-limiting toxicities and resistance. We report initial translational studies of a theranostic nanoemulsion loaded with a cisplatin derivative, myrisplatin and pro-apoptotic agent, C6-ceramide.

Radiofrequency ablation-induced upregulation of hypoxia-inducible factor-1α can be suppressed with adjuvant bortezomib or liposomal chemotherapy.

Purpose: To characterize upregulation of hypoxia-inducible factor (HIF)-1α after radiofrequency (RF) ablation and the influence of an adjuvant HIF-1α inhibitor (bortezomib) and nanodrugs on modulating RF ablation-upregulated hypoxic pathways.

Materials And Methods: Fisher 344 rats (n = 68) were used. First, RF ablation-induced periablational HIF-1α expression was evaluated in normal liver or subcutaneous R3230 tumors (14-16 mm).

Nanodrug-enhanced radiofrequency tumor ablation: effect of micellar or liposomal carrier on drug delivery and treatment efficacy.

Purpose: To determine the effect of different drug-loaded nanocarriers (micelles and liposomes) on delivery and treatment efficacy for radiofrequency ablation (RFA) combined with nanodrugs.

Materials/methods: Fischer 344 rats were used (n = 196). First, single subcutaneous R3230 tumors or normal liver underwent RFA followed by immediate administration of i.

On-demand intracellular amplification of chemoradiation with cancer-specific plasmonic nanobubbles.

Chemoradiation-resistant cancers limit treatment efficacy and safety. We show here the cancer cell-specific, on-demand intracellular amplification of chemotherapy and chemoradiation therapy via gold nanoparticle- and laser pulse-induced mechanical intracellular impact. Cancer aggressiveness promotes the clustering of drug nanocarriers and gold nanoparticles in cancer cells.

Targeted transferrin-modified polymeric micelles: enhanced efficacy in vitro and in vivo in ovarian carcinoma.

In this study, transferrin (Tf)-modified poly(ethylene glycol)-phosphatidylethanolamine (mPEG-PE) micelles loaded with the poorly water-soluble drug, R547 (a potent and selective ATP-competitive cyclin-dependent kinase (CDK) inhibitor), were prepared and evaluated for their targeting efficiency and cytotoxicity in vitro and in vivo to A2780 ovarian carcinoma cells, which overexpress transferrin receptors (TfR). At 10 mM lipid concentration, both Tf-modified and plain micelles solubilized 800 μg of R547. Tf-modified micelles showed enhanced interaction with A2780 ovarian carcinoma cells in vitro.

Phospholipid-modified polyethylenimine-based nanopreparations for siRNA-mediated gene silencing: implications for transfection and the role of lipid components.

Unlabelled: The clinical application of gene silencing mediated by small interfering RNA (siRNA) has been limited by the lack of efficient and safe carriers. Phospholipid modification of low molecular weight polyethylenimine (PEI 1.8 kDa) dramatically increased its gene down-regulation capacity while keeping cytotoxicity levels low.

Adjuvant liposomal doxorubicin markedly affects radiofrequency ablation-induced effects on periablational microvasculature.

Purpose: To evaluate the effects of radiofrequency (RF) ablation without and with adjuvant intravenous (IV) liposomal doxorubicin (Doxil) on microvessel morphology and patency and intratumoral drug delivery and retention.

Materials And Methods: There were 133 tumors/animals used in this experiment. First, single subcutaneous tumors (R3230 in Fischer rats and 786-0 in nude mice) were randomly assigned to receive RF ablation alone or no treatment and sacrificed 0-72 hours after treatment.

The effect of dual ligand-targeted micelles on the delivery and efficacy of poorly soluble drug for cancer therapy.

We prepared and evaluated transferrin (Tf) and monoclonal antibody (mAb) 2C5-modified dual ligand-targeted poly(ethylene glycol)-phosphatidylethanolamine micelles loaded with a poorly soluble drug, R547 (a selective adenosine triphosphate-competitive cyclin-dependent kinase inhibitor) for enhancement of targeting efficiency and cytotoxicity in vitro and in vivo to A2780 ovarian carcinoma compared to single ligand-targeted micelles. Micellar solubilization significantly improved the solubility of R547 from 1 to 800 μg/mL. The size of modified and non-modified micelles was 13-16 nm.

Immunomicelles for advancing personalized therapy.

Personalized medicine, which ultimately seeks to afford tailored therapeutic regimens for individual patients, is quickly emerging as a new paradigm in the diagnosis and treatment of diseases. The idea of casting aside generic treatments in favor of patient-centric therapies has become feasible owing to advances in nanotechnology and drug delivery coupled with an enhanced knowledge of genomics and an understanding of disease at the molecular level. This review highlights polymeric immunomicelles as a class of nanocarriers that have the potential to combine diagnosis, targeted drug therapy, as well as imaging and monitoring of therapeutic response, to render a personalized approach to the management of disease.

Challenges in development of targeted liposomal therapeutics.

Liposomes, phospholipid vesicles with a bilayered membrane structure, have been widely used as pharmaceutical carriers for drugs and genes, in particular for treatment of cancer. To enhance the efficacy of the liposomal drugs, drug-loaded liposomes are targeted to the tumors by means of passive (enhanced permeability and retention mediated) targeting, based on the longevity of liposomes in blood and its accumulation in pathological sites with compromised vasculature, and active targeting, based on the attachment of specific ligands to the liposomal surface to bind certain antigens on the target cells. Antibody-targeted liposomes loaded with anticancer drugs demonstrate high potential for clinical applications.

Design, synthesis, and initial biological evaluation of a steroidal anti-estrogen-doxorubicin bioconjugate for targeting estrogen receptor-positive breast cancer cells.

As part of our program to develop breast cancer specific therapeutic agents, we have synthesized a conjugate agent that is a conjugate of the steroidal anti-estrogen and the potent cytotoxin doxorubicin. In this effort, we employed a modular assembly approach to prepare a novel 11β-substituted steroidal anti-estrogen functionalized with an azido-tetraethylene glycol moiety, which could be coupled to a complementary doxorubicin benzoyl hydrazone functionalized with a propargyl tetraethylene glycol moiety. Huisgen [3 + 2] cycloaddition chemistry gave the final hybrid that was evaluated for selective uptake and cytotoxicity in ER(+)-MCF-7 and ER(-)-MDA-MB-231 breast cancer cell lines.

Polyethyleneimine-lipid conjugate-based pH-sensitive micellar carrier for gene delivery.

A low molecular weight polyethyleneimine (PEI 1.8 kDa) was modified with dioleoylphosphatidylethanolamine (PE) to form the PEI-PE conjugate investigated as a transfection vector. The optimized PEI-PE/pDNA complexes at an N/P ratio of 16 had a particle size of 225 nm, a surface charge of +31 mV, and protected the pDNA from the action of DNase I.

Combination radiofrequency (RF) ablation and IV liposomal heat shock protein suppression: reduced tumor growth and increased animal endpoint survival in a small animal tumor model.

Background: To investigate the effect of IV liposomal quercetin (a known down-regulator of heat shock proteins) alone and with liposomal doxorubicin on tumor growth and end-point survival when combined with radiofrequency (RF) tumor ablation in a rat tumor model.

Methods: Solitary subcutaneous R3230 mammary adenocarcinoma tumors (1.3-1.

P-glycoprotein silencing with siRNA delivered by DOPE-modified PEI overcomes doxorubicin resistance in breast cancer cells.

Aims: Multidrug resistance (MDR) mediated by overexpression of drug efflux transporters such as P-glycoprotein (P-gp), is a major problem, limiting successful chemotherapy of breast cancer. The use of siRNA to inhibit P-gp expression in MDR tumors is an attractive strategy to improve the effectiveness of anticancer drugs.

Method: We have synthesized a novel conjugate between a phospholipid (dioleoylphosphatidylethanolamine) and polyethylenimine (PEI) for siRNA delivery, for the purpose of silencing P-gp to overcome doxorubicin resistance in MCF-7 human breast cancer cells.

Development of the novel PEG-PE-based polymer for the reversible attachment of specific ligands to liposomes: synthesis and in vitro characterization.

Surface grafting of liposomes with the wide variety of ligands including antibodies and other proteins is a promising approach for targeted delivery of therapeutics. In this paper, we describe a simple method of synthesizing a hydrazine-functionalized poly(ethylene glycol)-phosphatidylethanolamine (PEG-PE)-based amphiphilic polymer which can conjugate a variety of ligands via a reversible, pH-cleavable bond. In this method, the targeting ligand is attached to the distal end of the PEG chain, which facilitates its easy access to the targeted site of interaction.

Radiofrequency ablation combined with liposomal quercetin to increase tumour destruction by modulation of heat shock protein production in a small animal model.

Purpose: To investigate the effect of heat shock protein (HSP) modulation on tumour coagulation by combining radiofrequency (RF) ablation with adjuvant liposomal quercetin and/or doxorubicin in a rat tumour model.

Methods: Sixty R3230 breast adenocarcinoma tumours/animals were used in this IACUC-approved study. Initially, 60 tumours (n=6, each subgroup) were randomised into five groups: (1) RF alone, (2) intravenous (IV) liposomal quercetin alone (1 mg/kg), (3) IV liposomal quercetin followed 24 h later with RF, (4) RF followed 15 min later by IV liposomal doxorubicin (8 mg/kg), (5) IV liposomal quercetin 24 h before RF followed by IV liposomal doxorubicin 15 min post-ablation.

Palmitoyl ascorbate liposomes and free ascorbic acid: comparison of anticancer therapeutic effects upon parenteral administration.

Purpose: To evaluate and compare anticancer therapeutic effect of palmitoyl ascorbate liposomes (PAL) and free ascorbic acid (AA).

Methods: Liposomes incorporating palmitoyl ascorbate (PA) were prepared and evaluated for PA content by HPLC. To elucidate mechanism of action of cell death in vitro, effect of various H(2)O(2) scavengers and metal chelators on PA-mediated cytotoxicity was studied.

Design and synthesis of novel functional lipid-based bioconjugates for drug delivery and other applications.

The modification of biologicals such as proteins/peptides, small molecules, and other polymers with lipids provides an efficient method for mediating their insertion into liposomes and lipid-core micellar nanocarriers. In this chapter, we describe several representative protocols developed in our laboratory for the bioconjugation of liposomes and lipid-core micelles for drug/gene delivery and diagnostic imaging applications.

Cell-penetrating TAT peptide in drug delivery systems: proteolytic stability requirements.

The stability and activity of the HIV cell-penetrating TAT peptide (TATp) on the surface of TATp-modified micelles and liposomes in relation to its proteolytic cleavage was investigated. TATp moieties were attached to the surface of these nanocarriers using TATp modified with a conjugate of phosphatidyl ethanolamine with a 'short' PEG (PEG-PE). Following pre-incubation with trypsin, elastase, or collagenase, the proteolytic stability of TATp on the surface of these modified carriers was studied by HPLC with fluorescence detection using fluorenylmethyl chloroformate (FMOC) labeling.

Surface modification of liposomes with rhodamine-123-conjugated polymer results in enhanced mitochondrial targeting.

A novel mitochondrial-targeted liposomal drug-delivery system was prepared by modification of the liposomal surface with a newly synthesized polymer, rhodamine-123 (Rh123)-PEG-DOPE inserted into the liposomal lipid bilayer. This novel polymer was synthesized by conjugating the mitochondriotropic dye Rh123, with the amphiphilic polyethylene glycol-phosphatidylethanolamine (PEG-PE) conjugate. The modified liposomes showed better uptake by cells (HeLa, B16F10) estimated by fluorescence microscopy and FACS analysis.

Phospholipid-polyethylenimine conjugate-based micelle-like nanoparticles for siRNA delivery.

Gene silencing using small interfering RNA (siRNA) is a promising therapeutic strategy for the treatment of various diseases, in particular, cancer. Recently, our group reported on a novel gene carrier, the micelle-like nanoparticle (MNP), based on the combination of a covalent conjugate of phospholipid and polyethylenimine (PLPEI) with polyethylene glycol (PEG) and lipids. These long-circulating MNPs loaded with plasmid DNA-mediated gene expression in distal tumors after systemic administration in vivo.

Multifunctionality of lipid-core micelles for drug delivery and tumour targeting.

Phospholipid micelles have proven to be the versatile pharmaceutical nanocarrier of choice for the delivery of poorly soluble chemotherapeutics for cancer therapy using various treatment modalities. Phospholipid micelles are typically expected to increase the accumulation of the loaded drugs in tumour tissues by taking advantage of the enhanced permeability and retention effect and by ligand-mediated active targeting. Furthermore, by tailoring the composition of the micelles, it is possible to enhance the intracellular delivery of the cargo.

Do liposomal apoptotic enhancers increase tumor coagulation and end-point survival in percutaneous radiofrequency ablation of tumors in a rat tumor model?

Purpose: To characterize effects of combining radiofrequency (RF) ablation with proapoptotic intravenous liposome-encapsulated paclitaxel and doxorubicin on tumor destruction, apoptosis and heat-shock protein (HSP) production, intratumoral drug accumulation, and end-point survival.

Materials And Methods: R3230 mammary adenocarcinomas (n = 177) were implanted in 174 rats in this animal care committee-approved study. Tumors received (a) no treatment, (b) RF ablation, (c) paclitaxel, (d) RF ablation followed by paclitaxel (RF ablation-paclitaxel), (e) paclitaxel before RF ablation (paclitaxel-RF ablation), (f) RF ablation followed by doxorubicin (RF ablation-doxorubicin), (g) paclitaxel followed by doxorubicin without RF ablation (paclitaxel-doxorubicin), or (h) paclitaxel before RF ablation, followed by doxorubicin (paclitaxel-RF ablation-doxorubicin).

Palmitoyl ascorbate-loaded polymeric micelles: cancer cell targeting and cytotoxicity.

Purpose: To evaluate the potential of palmitoyl ascorbate (PA)-loaded micelles for ascorbate-mediated cancer cell targeting and cytotoxicity.

Methods: PA was incorporated in polyethylene glycol-phosphatidyl ethanolamine micelles at varying concentrations. The formulations were evaluated for PA content by RP-HPLC.