Atelocollagen-mediated systemic DDS for nucleic acid medicines.

The goal of our research is to provide a practical platform for drug delivery in oligonucleotide therapy. We report here the efficacy of an atelocollagen-mediated oligonucleotide delivery system applied to systemic siRNA and antisense oligonucleotide treatments in animal disease models. Atelocollagen and oligonucleotides formed a complex of nanosized particles, which was highly stable against nucleases. The complex allowed oligonucleotides to be delivered efficiently into several organs and tissues via intravenous administration. In a tumor metastasis model, the complex successfully delivered siRNA to metastasized tumors in bone tissue and inhibited their growth. We also demonstrated that a single intravenous treatment of the antisense oligodeoxynucleotide complex suppressed ear dermatitis in a contact hypersensitivity model. These results indicate the strong potential of the atelocollagen-mediated drug delivery system for practical therapeutic technology.

Atelocollagen-mediated synthetic small interfering RNA delivery for effective gene silencing in vitro and in vivo.

Silencing gene expression by siRNAs is rapidly becoming a powerful tool for the genetic analysis of mammalian cells. However, the rapid degradation of siRNA and the limited duration of its action call for an efficient delivery technology. Accordingly, we describe here that Atelocollagen complexed with siRNA is resistant to nucleases and is efficiently transduced into cells, thereby allowing long-term gene silencing. Site-specific in vivo administration of an anti-luciferase siRNA/Atelocollagen complex reduced luciferase expression in a xenografted tumor. Furthermore, Atelocollagen-mediated transfer of siRNA in vivo showed efficient inhibition of tumor growth in an orthotopic xenograft model of a human non-seminomatous germ cell tumor. Thus, for clinical applications of siRNA, an Atelocollagen-based non-viral delivery method could be a reliable approach to achieve maximal function of siRNA in vivo.

Potential of atelocollagen-mediated systemic antisense therapeutics for inflammatory disease.

To study the possibility of using atelocollagen as an oligonucleotide (ODN) delivery carrier in vivo, the activity of formulated antisense ODN targeted against the intercellular adhesion molecule-1 (ICAM-1) mRNA was investigated in an allergic dermatitis model in mice. The allergic dermatitis was elicited in one ear of animals sensitized by treatment with 2,4-dinitrofluorobenzene. Antisense ODN was given to the animals as a single intravenous injection of formulation containing atelocollagen. Antisense activity was determined by measurement of ear thickness, histopathology, and immunohistochemistry 24 hr after the initiation of the dermatitis. Antisense activity was found to increase according to the concentration of atelocollagen in the formulation. The effect mediated by the ODN formulated with 0.05% atelocollagen was more than 50 times greater than that provided by ODN infusion, although the levels of ODN formulated with atelocollagen dropped below that of the 24-hr infusion group within 30 min. The formulated ODN could suppress inflammatory progression by treatment at 8 hr after the ear challenge when inflammation had already commenced at the challenged site. Moreover, antisense activity was noted even when the formulated ODN was injected 3 days before the initiation of inflammation. These data demonstrate that atelocollagen can enhance antisense activity remarkably and that the sustainable antisense activity mediated by the formulation of ODN with atelocollagen could completely change the strategy of antisense therapeutics.

Efficient delivery of small interfering RNA to bone-metastatic tumors by using atelocollagen in vivo.

Silencing of gene expression by small interfering RNAs (siRNAs) is rapidly becoming a powerful tool for genetic analysis and represents a potential strategy for therapeutic product development. However, there are no reports of systemic delivery for siRNAs toward treatment of bone-metastatic cancer. Accordingly, we report here that i.v. injection of GL3 luciferase siRNA complexed with atelocollagen showed effective reduction of luciferase expression from bone-metastatic prostate tumor cells developed in mouse thorax, jaws, and/or legs. We also show that the siRNA/atelocollagen complex can be efficiently delivered to tumors 24 h after injection and can exist intact at least for 3 days. Furthermore, atelocollagen-mediated systemic administration of siRNAs such as enhancer of zeste homolog 2 and phosphoinositide 3'-hydroxykinase p110-alpha-subunit, which were selected as candidate targets for inhibition of bone metastasis, resulted in an efficient inhibition of metastatic tumor growth in bone tissues. In addition, upregulation of serum IL-12 and IFN-alpha levels was not associated with the in vivo administration of the siRNA/atelocollagen complex. Thus, for treatment of bone metastasis of prostate cancer, an atelocollagen-mediated systemic delivery method could be a reliable and safe approach to the achievement of maximal function of siRNA.