Antisense promised major advances in treating a broad range of intractable diseases, but in recent years progress has been stymied by technical problems, most notably inadequate specificity, ineffective delivery into the proper subcellular compartment, and unpredictable activity within cells. Herein is an overview of the design, preparation, and properties of Morpholino oligos, a novel antisense structural type that solves the sequence specificity problem and provides high and predictable activity in cells. Morpholino oligos also exhibit little or no nonantisense activity, afford good water solubility, are immune to nucleases, and are designed to have low production costs.

Morpholino antisense oligomers: design, preparation, and properties.

Conceptual simplicity, the possibility of rational design, relatively inexpensive cost, and developments in the sequencing of human genome have led to the use of short fragments of nucleic acid, commonly called oligonucleotides, either as therapeutic agents or as tools to study gene function

https://mct.aacrjournals.org/content/molcanther/1/5/347.full.pdf

Antisense Oligonucleotides: Basic Concepts and Mechanisms

Locked nucleic acid (LNA): fine-tuning the recognition of DNA and RNA

https://www.gene-tools.com/sites/default/files/bba%20review.pdf

Morpholino antisense oligomers: the case for an RNase H-independent structural type.

Morpholino oligos: making sense of antisense?

Oligomers possessing the Morpholino phosphorodiamidate backbone were evaluated for resistance to a variety of enzymes and biologic fluids. A 25-mer was incubated with nucleases, proteases, esterase...

Resistance of morpholino phosphorodiamidate oligomers to enzymatic degradation.

The phosphorodiamidate Morpholino oligomers (PMO) are a new class of antisense agents that inhibit gene expression by binding to RNA and sterically blocking processing or translation. In a search for a Morpholino agent that would inhibit cell proliferation, it was found that oligomers directed against c-myc, a gene involved in control of the cell cycle, were effective. The sequence specificity and mechanism of action of one agent were determined. The 20-mer 126 lowers c-myc protein levels in treated cells and arrests cells in G0/G1 of the cell cycle. It also acts at the RNA level to inhibit normal pre-mRNA splicing and instead produces an aberrantly spliced mRNA. Irrelevant and mispair control oligomers indicated that the observed antiproliferative effect was sequence specific. This was confirmed in a reporter gene model system using a c-myc 5‵-untranslated region (5‵-UTR) fused to a cDNA copy of the insect luciferase gene. We conclude that 126 is acting through an antisense mechanism involving Watson-Cri...

Antiproliferative effects of steric blocking phosphorodiamidate morpholino antisense agents directed against c-myc.

Antisense compositions targeted against an mRNA sequence coding for a selected protein, at a region having its 5′ end from 1 to about 25 base pairs downstream of a normal splice acceptor junction in the preprocessed mRNA, are disclosed. The antisense compound is RNase-inactive, and is preferably a phosphorodiamidate-linked morpholino oligonucleotide. Such targeting is effective to inhibit natural mRNA splice processing, produce splice variant mRNAs, and inhibit normal expression of the protein.

Splice-region antisense composition and method

L'invention concerne des compositions anti-sens ciblees contre une sequence d'ARN messager codant pour une proteine selectionnee, au niveau d'une region dont l'extremite 5' se situe a 1 a environ 25 paires de base en aval d'un accepteur d'epissage normal dans l'ARN messager pretraite. Ce compose anti-sens est inactif par rapport a la RNase, et se presente de preference sous la forme d'un morpholino-oligonucleotide lie a un phosphorodiamidate. Ce ciblage est efficace pour inhiber le traitement d'epissage d'ARN messager naturel, pour produire des variantes d'ARN messagers d'epissage, et pour inhiber l'expression normale de la proteine.

Robert M. Hudziak

Papers

Resistance of morpholino phosphorodiamidate oligomers to enzymatic degradation.

Antiproliferative effects of steric blocking phosphorodiamidate morpholino antisense agents directed against c-myc.

Splice-region antisense composition and method

Composition anti-sens a region d'epissage et methode associee