Lesheng Teng

Clinical translation of folate receptor-targeted therapeutics.

Introduction: Folate receptor−α (FR−α) has been established as a membrane marker for ovarian cancer. In addition, it is frequently overexpressed in other major types of epithelial tumors. FR−α-based tumor-targeted therapy and drug carriers have been an active area of laboratory research for more than 20 years. Recently, there has been a great increase in the effort to finally translate this promising technology into the clinic and bring FR-targeted therapeutics into the market. Areas covered: Two FR-targeted therapeutic agents have moved into Phase III clinical trials, the monoclonal antibody farletuzumab and the low molecular weight vintafolide, combined with etarfolatide as a companion imaging agent, representing two alternative strategies for targeting the FR. Expert opinion: Each of the two strategies has advantages and disadvantages. Identification of the best target patient population is likely critical to the ultimate success of FR-targeted agents in the clinic. A successful clinical strategy may require the integration between FR expression analysis and an optimal combination of FR-targeted therapy and standard chemotherapy. Advancement into Phase III trials and the ongoing clinical development of several additional folate conjugates are likely to usher in a new era of clinical translation and validation of FR-targeted imaging and therapeutic agents.

A Polyethylenimine-Linoleic Acid Conjugate for Antisense Oligonucleotide Delivery

A novel antisense oligonucleotide (ASO) carrier, polyethylenimine conjugated to linoleic acid (PEI-LA), was synthesized and evaluated for delivery of LOR-2501 to tumor cells. LOR-2501 is an ASO targeting ribonucleotide reductase R1 subunit (RRM1). In this study, PEI-LA was synthesized by reacting PEI (Mw ~ 800) with linoleoyl chloride. Gel retardation assay showed complete complexation between PEI-LA and LOR-2501 at N/P ratio above 8. No significant cytotoxicity was observed with these complexes at the tested dosage levels. Interestingly, at N/P ratio of >6, levels of cellular uptake of PEI-LA/LOR-2501 were double that of PEI/LOR-2501 complexes of the same N/P ratio. PEI-LA/LOR-2501 induced downregulation of 64% and 70% of RRM1 at mRNA and protein levels, respectively. The highest transfection activity was shown by PEI-LA/LOR-2501 complexes at N/P ratio of 10. Finally, using pathway specific inhibitors, clathrin-mediated endocytosis was shown to be the principle mechanism of cellular internalization of these complexes. In conclusion, PEI-LA is a promising agent for the delivery of ASOs and warrants further investigation.

Insight into Mechanisms of Cellular Uptake of Lipid Nanoparticles and Intracellular Release of Small RNAs

PurposeUnderstanding mechanisms of cellular uptake and intracellular release would enable better design of nanocarriers for delivery of nucleic acids such as siRNA and microRNA (miRNA).MethodIn this study, we investigated cellular pharmacokinetics of siRNA by co-encapsulating fluorescently labeled siRNA and molecular beacon (MB) in four different formulations of cationic lipid nanoparticles (LNPs). A miRNA mimic was also used as a probe for investigating cellular pharmacokinetics, which correlated well with RNAi activities.ResultsWe tried to find the best LNP formulation based on the combination of DOTMA and DODMA. When the DOTMA/DODMA ratio was at 5/40, the LNP containing a luciferase siRNA produced the highest gene silencing activity. The superior potency of DOTMA/DODMA could be attributed to higher uptake and improved ability to facilitate siRNA release from endosomes subsequent to uptake.ConclusionsOur findings may provide new insights into RNAi transfection pathways and have implications on cationic LNP design.

Near infrared spectroscopic (NIRS) analysis of drug-loading rate and particle size of risperidone microspheres by improved chemometric model

Microspheres have been developed as drug carriers in controlled drug delivery systems for years. In our present study, near infrared spectroscopy (NIRS) is applied to analyze the particle size and drug loading rate in risperidone poly(d,l-lactide-co-glycolide) (PLGA) microspheres. Various batches of risperidone PLGA microspheres were designed and prepared successfully. The particle size and drug-loading rate of all the samples were determined by a laser diffraction particle size analyzer and high performance liquid chromatography (HPLC) system. Monte Carlo algorithm combined with partial least squares (MCPLS) method was applied to identify the outliers and choose the numbers of calibration set. Furthermore, a series of preprocessing methods were performed to remove signal noise in NIR spectra. Moving window PLS and radical basis function neural network (RBFNN) methods were employed to establish calibration model. Our data demonstrated that PLS-developed model was only suitable for drug loading analysis in risperidone PLGA microspheres. Comparatively, RBFNN-based predictive models possess better fitting quality, predictive effect, and stability for both drug loading rate and particle size analysis. The correlation coefficients of calibration set (Rc(2)) were 0.935 and 0.880, respectively. The performance of optimum RBFNN models was confirmed by independent verification test with 15 samples. Collectively, our method is successfully performed to monitor drug-loading rate and particle size during risperidone PLGA microspheres preparation.

Enhanced delivery of Paclitaxel using electrostatically-conjugated Herceptin-bearing PEI/PLGA nanoparticles against HER-positive breast cancer cells.

We have developed a novel nanoparticle delivery system fabricated from polyethylenimine (PEI) and poly(d,l-lactide-co-glycolide) (PLGA), which were able to deliver the chemotherapeutic agent Paclitaxel, while the biomacromolecule Herceptin acted as a targeting ligand that was conjugated onto the surfaces of the nanoparticles via electrostatic interactions. In this study, these electrostatically-conjugated Herceptin-bearing PEI/PLGA nanoparticles (eHER-PPNs) were optimized and employed as vectors to target HER2-positive breast cancer cells. The eHER-PPNs had an average diameter of ∼ 280 nm and a neutral surface charge (1.00 ± 0.73 mV), which remained stable under physiological conditions. The anticancer effects of eHER-PPNs were investigated in HER2-positive BT474 cells and HER2-negative MCF7 cells. The eHER-PPNs showed enhanced cytotoxicity that was dependent on the receptor expression levels and the incubation time. These conjugated nanoparticles deliver Paclitaxel more efficiently (p<0.001) than unmodified PPNs, Herceptin and the combined effects of these two monotherapies. Furthermore, the chemically-conjugated Herceptin-bearing PEI/PLGA nanoparticles (cHER-PPNs) were fabricated as a comparison. The eHER-PPNs exhibited lower cell viability (46.7%) than that of cHER-PPNs (65.1%). The targeting ability of eHER-PPNs was demonstrated through confocal microscopy images and flow cytometry, which showed that eHER-PPNs displayed higher cellular uptake efficiency (p<0.001) in comparison with cHER-PPNs. Therefore, eHER-PPNs could provide promising platforms for the delivery of therapeutic drugs against HER2-positive breast cancers.

Enhanced antitumor efficacy of vitamin E TPGS-emulsified PLGA nanoparticles for delivery of paclitaxel

Nanoparticles are efficient delivery vehicles for cancer therapy such as paclitaxel (PTX). In this study, we formulated PTX into PLGA polymeric nanoparticles. Vitamin E TPGS was used as an emulsifier to stabilize the nanoparticle formulation. PTX was encapsulated in TPGS-emulsified polymeric nanoparticles (TENPs) by a nanoprecipitation method in ethanol-water system. The resultant PTX-TENPs showed a very uniform particle size (∼100 nm) and high drug encapsulation (>80%). The cytotoxicity of PTX-TENPs was examined in A549 lung cancer cell line. Preferential tumor accumulation of TENPs was observed in the A549 lung cancer xenograft model. Tumor growth was significantly inhibited by intravenous injection of PTX-TENPs. Our results suggested that the modified nanoprecipitation method holds great potential for the fabrication of the PTX loaded polymeric nanoparticles. TPGS can be used in the manufacture of polymeric nanoparticles for the controlled release of PTX and other anti-cancer drugs.

Functional exosome-mimic for delivery of siRNA to cancer: in vitro and in vivo evaluation.

Exosomes, the smallest subgroup of extracellular vesicles, have been recognized as extracellular organelles that contain genetic and proteomic information for long distance intercellular communication. Exosome-based drug delivery is currently a subject of intensive research. Here, we report a novel strategy to produce nanoscale exosome-mimics (EMs) in sufficient quantity for gene delivery in cancer both in vitro and in vivo. Size-controllable EMs were generated at a high yield by serial extrusion of non-tumorigenic epithelial MCF-10A cells through filters with different pore sizes. siRNA was then encapsulated into the EMs by electroporation. Biosafety and uptake efficiency of the EMs were evaluated both in vitro and in vivo. The mechanism underlying their cellular endocytosis was also studied.

Nanotechnology for the delivery of phytochemicals in cancer therapy.

The aim of this review is to summarize advances that have been made in the delivery of phytochemicals for cancer therapy by the use of nanotechnology. Over recent decades, much research effort has been invested in developing phytochemicals as cancer therapeutic agents. However, several impediments to their wide spread use as drugs still have to be overcome. Among these are low solubility, poor penetration into cells, high hepatic disposition, and narrow therapeutic index. Rapid clearance or uptake by normal tissues and wide tissue distribution result in low drug accumulation in the target tumor sites can result in undesired drug exposure in normal tissues. Association with or encapsulation in nanoscale drug carriers is a potential strategy to address these problems. This review discussed lessons learned on the use of nanotechnology for delivery of phytochemicals that been tested in clinical trials or are moving towards the clinic.

Human serum albumin-coated lipid nanoparticles for delivery of siRNA to breast cancer.

UNLABELLED Human serum albumin (HSA)-coated lipid nanoparticles (HSA-LNPs) loaded with phrGFP-targeted siRNA (HSA-LNPs-siRNA) were prepared and evaluated for gene downregulation effect in phrGFP-transfected breast cancer cells and the corresponding xenograft tumor model. HSA-LNPs-siRNA were successfully prepared with a particle size of 79.5±5.5 nm. In phrGFP-transfected MCF-7 cells, HSA-LNPs-siRNA significantly decreased cell fluorescence even in the presence of fetal bovine serum (FBS). Moreover, cell fluorescence and phrGFP mRNA expression were significantly downregulated by HSA-LNPs-siRNA in phrGFP-transfected MCF-7, MDA-MB-231, and SK-BR-3 cells in comparison with control or HSA-LNPs-siRNA (scrambled). In phrGFP-transfected MCF-7 xenograft tumor model, tumor fluorescence was significantly decreased after three IV administrations of HSA-LNPs-siRNA at a dose of 3 mg/kg in comparison with siRNA alone. HSA-LNPs-siRNA demonstrated a superior pharmacokinetic profile in comparison with siRNA at a dose of 1mg/kg. These results show that the novel nonviral carrier, HSA-LNPs, may be used for the delivery of siRNA to breast cancer cells. FROM THE CLINICAL EDITOR Targeted delivery of siRNA to cancer cells may be a viable anti-cancer strategy with low toxicity. In this study the novel nonviral carrier, human serum albumin-coated lipid nanoparticles (HSA-LNP) were demonstrated as an efficient delivery agent of siRNA to breast cancer cells.

A Novel Isoquinoline Derivative Anticancer Agent and Its Targeted Delivery to Tumor Cells Using Transferrin-Conjugated Liposomes

We have screened 11 isoquinoline derivatives and α-methylene-γ-butyrolactones using the 3-(4,5-dimethylthi-azol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cytotoxicity assay in HeLa and HEK-293T cells. Compound 2 was identified as potential anticancer agent. To further improve its therapeutic potential, this agent was incorporated into transferrin (Tf)-conjugated liposomes (LPs) for targeted delivery to tumor cells. We have demonstrated Tf-LP-Compound 2 have superior antitumor activity compared to non-targeted controls and the free drug. These data show Tf-LP-Compound 2 to be a promising agent that warrants further evaluation.