Katpak:m3d2

Background:

Ribonucleic acid interference (RNAi) has been recognized as both an insightful tool in research and a potential therapeutic. Briefly, the process of RNAi involves the targeted cleavage of messenger RNAs (mRNAs) via complementary recognition by the RNAi sequence. This consequently inhibits translation of the targeted mRNA and thus silences expression of the gene encoding that mRNA transcript. This concept can be applied to the development of pharmaceuticals that specifically recognize and inhibit expression of particular gene products which are expressed or over-expressed in disease-state individuals. Efforts to target such disease-state cellular expression are currently ongoing (i.e. in cancer therapy research). However, there are key barriers to the breakthrough of such new therapy application which must first be addressed. One problematic issue is the delivery of the interfering RNA into the body and subsequent recognition of target cells. To this end, several methods have been applied; however, few have been successful and none have been clinically approved.

Research Problems and Goals:

Problems: The problems we aim to address involve the current limitations of siRNA delivery for therapeutic purposes.

Goal: Our goal is to engineer a widely-applicable therapeutic agent that can be easily administered and can select target cells for delivery of siRNA.

We aim to produce a therapeutic agent that:

• Can be easily administered to patients
• Is highly specific for cell and gene targets
• Allows for modularity in siRNA design

Previous Efforts:

• Package siRNA inside liposomes - Liposomes protect the siRNA from degradation and kidney clearance, but tend to get

trapped in the liver, and so are mostly effective for targeting such conditions as hepatitis and hypercholesteremia.

• Conjugate the siRNA to a large targeting molecule – physically linking the siRNA to an antibody, aptamer, or cholesterol which allows for recognition by a cell membrane receptor.

• Complex the siRNA into a particle - Use electrostatic interaction

between the negatively charged RNA backbone and some positively charged molecule to form a complex which can target specific cells.

• Localizing delivery of siRNA - The first siRNA to be granted investigational new drug (IND) status by the US Food and Drug Administration (FDA) has to be administered by intravitrial injection (into the eye). By injecting into the eye, siRNAs are protected from nucleases and the immune system, and because of the compartment’s relatively small volume, can be delivered at high concentration.

• Delivery with a virus/plasmid – larger noncoding RNA called a microRNA (miRNA) expressed from a virus or plasmid and processed in vivo to produce the mature siRNA.

pH-Sensitive Drug Release

Delivery of drugs to cells is a relevant factor in all therapeutic design considerations. For instance, nano-sized vesicular systems (nanoparticles) can be used to load a drug and deliver it to cells via endocytosis,a natural cellular process. In this approach, it is important for the drug to be able to exit the endosome particle in order to be effective. A pH-sensitive drug delivery method has been proposed to target this issue. using N-acetyl histidine (NAcHis) conjugated to a hydrophilic polymer (glycol chitosan). At extracellular pH, the N-acetyl histidine (NAcHis) is hydrophobic and the conjugates form self-assembled nanoparticles. In slightly acidic environments, such as those in endosomes, the nanoparticles disassemble due to breakdown of the hydrophilic/hydrophobic balance by the protonation of the imidazole group of NAcHis and consequently release the drug into the cell.

Resources:

1. Perkel, J. Therapeutic RNAi: Delivering the Future? Science. November 2007.

2. Park, J.S., Han, T.H., Lee, K.Y., Han, S.S., Hwang, J.J., Moon, D.H., Kim, S.Y., Cho, Y.W. N-acetyl histidine-conjugated glycol chitosan self-assembled nanoparticles for intracytoplasmic delivery of drugs: endocytosis, exocytosis and drug release. Journal of Controlled Release. 115(1):37-45. September 2006.