January 16, 2007
Nanoparticles Continue to Yield Delivery Tools for Therapeutic Nucleic Acids
Cancer researchers hold great hope that nucleic acid-based therapeutics, such as anticancer genes, antisense oligonucleotides, and small interfering RNA (siRNA) molecules, will prove to be powerful antitumor agents. One reason for that optimism is that a variety of nanoscale materials are showing their mettle as delivery agents for these potent but biologically unstable molecules. Three new reports highlight progress in this field.
One study, published in the journal Gene Therapy by Hideyoshi Harashima, Ph.D., and colleagues at Hokkaido University in Japan, describes the development of what the researchers call a multifunctional envelope-type nano device (MEND) designed to shed a protective coating only when it comes in contact with tumor cells. With this coating removed, the nanoparticle enters tumor cells and safely delivers its nucleic acid payload.
When injected into the bloodstream, the nanoscale device contains a coating of poly(ethylene glycol), or PEG, a biocompatible polymer that protects the nanoparticle from rapid clearance from the body. However, this PEG coating interferes with the ability of therapeutic genes contained within the MEND from expressing within tumor cells and exerting their intended therapeutic effect. The investigators resolved this conflict by adding a linker between the PEG coating and the surface of the MEND. An enzyme known as matrix metalloproteinase (MMP), found in the spaces between tumor cells, can cleave this linker, freeing the MEND of its PEG coating.
Studies with cultured tumor cells showed that this PEG-sloughing mechanism works as intended only when the tumor cells produce MMP. When MMP is present, MEND particles effectively deliver an antitumor gene that then expresses within the targeted cells. However, when the researchers added this MEND to a second tumor cell type, one that does not produce MMP, the level of gene expression was 95 percent less than in the MMP-positive cells. Subsequent studies in mice bearing MMP-expressing tumors confirmed these results – the level of anticancer gene expression in tumors was 100-fold higher in mice dosed with the PEG-sloughing MEND compared to the level observed in mice that received nanoparticles whose PEG coating remained intact.
Taking a different approach, a research team headed by Uwe Zangemeister-Wittke, Ph.D., at the University of Zürich in Switzerland has developed a lipid-based nanoparticle designed to bind to a cancer cell protein known as epithelial cell adhesion molecule (EpCAM). As binding occurs, the nanoparticle is quickly taken up by the tumor cells. Once inside the cells, the nanoparticle releases an antisense oligonucleotide that shuts down production of two proteins known as bcl-2 and bcl-xL. Without these proteins, the treated tumor cells become sensitive to the anticancer drug doxorubicin.
This study, which was published in the journal Molecular Cancer Therapeutics, started with the investigators developing an antibody fragment that binds to human EpCAM. They then attached this antibody fragment to a lipid-based nanoparticle and showed that these nanoparticles were taken up efficiently by EpCAM-expressing tumor cells and not at all by cells that do not express EpCAM. Based on these results, the investigators then loaded the targeted nanoparticles with the therapeutic antisense agent and added it to EpCAM-expressing human solid tumor cells. Production of bcl-2 and bcl-xL fell by 70 percent and 60 percent, respectively. In contrast, non-targeted nanoparticles and EpCAM-targeted nanoparticles loaded with a random oligonucleotide had no effect on protein production.
Tumor cells that produce bcl-2 and bcl-xL are resistant to many anticancer agents because these proteins prevent cells from undergoing apoptosis in response to chemotherapy. But after treatment with the targeted nanoparticles, tumor cells succumbed rapidly to doxorubicin. The investigators calculated that their nanoparticle increased tumor cell sensitivity to the lethal effects of doxorubicin by as much as five-fold.
Meanwhile, a group of investigators led by Yong Zhang, Ph.D., at the National University of Singapore, have developed a quantum dot-based nanoparticle that allowed the researchers to deliver siRNA molecules to tumor cells and track that the nanoparticles successfully delivered their cargo. This work is reported in the journal Biomaterials.
The primary goal of Zhang’s group was to develop a nanoscale device that would enable them to monitor the delivery of siRNA to tumor cells. To create this device, the investigators packed fluorescent quantum dots inside chitosan nanoparticles, which many groups have been developing as tumor-targeting drug delivery agents. The researchers also incorporated an siRNA molecule designed to shut down production of the protein HER2, which plays a central role in the formation of some forms of breast cancer.
Experiments with this construct showed that they were readily taken up by HER2-positive tumor cells. Thanks to the presence of the brightly shining quantum dots, the investigators were able to observe this uptake process easily using a confocal fluorescence microscope. Once inside the tumor cells, the siRNA agent shut down production of the HER2 protein.
The work with PEG-sloughing nanoparticles is detailed in a paper titled, “Development of a novel systemic gene delivery system for cancer therapy with a tumor-specific cleavable PEG-lipid.” Investigators from the Japan Science and Technology Agency, the University of Tsukuba, Toyama Medical and Pharmaceutical University, and Daiichi Pharmaceutical Co., all in Japan, also participated in this study. An abstract of this paper is available at the journal’s Website.
The work with EpCAM-targeted nanoparticles is detailed in a paper titled, “Chemosensitization of carcinoma cells using epithelial cell adhesion molecule-targeted liposomal antisense against bcl-2/bcl-xL.” An investigator from the University of Alberta, in Canada, also participated in this study. An abstract of this paper is available through PubMed.
The work with the HER2-silencing nanoparticles is detailed in a paper titled, “Quantum-dot based nanoparticles for targeted silencing of HER2/neu gene via RNA interference.” An abstract of this paper is available through PubMed.