FANA Technology for RNA Silencing and Regulation

Overview

Some of the most important desired features in any gene-silencing or manipulation tool are:

(1) Efficient knockdown or regulation of the target RNA.
(2) Ability to bind to the target RNA (mRNA, miRNA, or lncRNA) in a high sequence specific manner.
(3) No toxicity.
(4) Most importantly – efficient delivery without an external source (e.g. without a transfection agent, formulation, conjugate or viral vector).

Antisense oligonucleotides designed using next-generation FANA technology can effectively and efficiently perform all these activities.

FANA-Antisense Oligonucleotides

Antisense oligonucleotides offer the ability to target and silence specific sequences of RNA. These molecules consist of a short segment of DNA flanked by two segments of RNA, linked with a phosphorothioate backbone. However, they are not without their own set of challenges, and can suffer from a lack of stability and/or affinity for their target. Both of these properties can be improved by the incorporation of a modified sugar in the RNA backbone, which changes the RNA segments to a similar but more effective molecule. This next-generation molecule is called Fluoroarabinonucleic acid, or 2’ F-ANA.

Structures of (a) 2’F-ANA and (b) ANA, in comparison with (c) DNA and (d) RNA oligomers.

FANA-ASOs effect gene silencing by finding mRNA complementary to their sequence. This FANA-RNA hybrid is recognized by a resident nuclease called RNAse H. RNAse H then cleaves the mRNA, preventing its translation into the protein it coded for. The FANA-ASO is then free to find and bind more mRNA, perpetuating mRNA target degradation and silencing the targeted gene. This process is illustrated and compared to a similar technology (siRNA) below.

FANA-ASO technology confers several key advantages over other RNA silencing technologies.

Can be Self-delivered:
FANA-ASOs are capable of gymnotic delivery, which is self-delivery to cells without requiring a delivery agent. No formulation, conjugate, or viral vector (like AAV) is necessary. Because of this, FANA-ASOs are ideal for primary cells or difficult to transfect cell lines, such as T- cells, B-cells, neuronal cultures, and others. Even without a delivery vehicle, FANA-ASOs exhibit remarkable resistance to degradation by serum and cellular nucleases, which significantly extends the duration of their activity to weeks or months.

Safe:
AUM LifeTech’s FANA-ASOs are proven to be non-toxic to a variety of different cells and tissues. They exhibit low non-specific protein binding and, unlike other technologies like siRNA, avoid RISC associated off-target effects. They likewise do not alter the biology of cells involved in the experiment, allowing for accurate analysis of gene expression and cell health.

Efficacious:
FANA-ASOs are capable of silencing in a range of in vitro and in vivo models, including mammals, insects, fish, and amphibians. Silencing is long-term and sustained, allowing FANA-ASOs to be used as an alternative for stable cell lines. Most importantly, FANA-ASOs bind to their RNA target with high affinity and specificity, and are effective in low concentrations. They offer high efficacy paired with high adaptability, and the ability to treat numerous gene-based diseases.

Cost-effective:
The FANA RNA silencing technology is cost-effective both as an experimental screen to understand the genetic underpinnings behind a disease and as a therapeutic to treat it. Experimentally, there is no need to grow an excessive number of cells to compensate for toxic transfection reagents, and FANA-ASOs are active at low concentrations with excellent reproducibility of silencing. Likewise, the same FANAs that were used to identify targets relevant to a disease are the same FANAs which can be used to silence and treat that disease. This saves significant time and resources, and improves cost efficiency of the technology.