Pharmacological exon skipping in brains of mice
Jon Moulton
Tuesday, 21 October 2008 14:34 UTC
The mdx mouse is a model of Duchenne muscular dystrophy, carrying a premature termination codon in exon 23 of dystrophin. Morpholino oligos targeting dystrophin exon 23 were delivered from a micro-osmotic pump via cannula intracerebroventricularly. The unmodified Morpholino oligos triggered excision of dystrophin exon 23 and sometimes both exons 22 and 23, removing the premature termination codon. While a small fraction of the transcripts underwent modified splicing, this was sufficient to ameliorate behavioral abnormalities of the mdx mice.
RT-PCR of mRNA isolated from (mostly) the cerebral cortex, amygdala and hippocampus predominantly revealed the wild-type band with exons 22-23-24, though dystrophin transcripts missing exon 23 and others missing exons 22 and 23 were visibly present in lanes from antisense treated mice but not sense treated mice. Western blots revealed that, after Morpholino treatment, the transcript downstream of exon 23 was translated in the forebrain.
Sekiguchi M, Zushida K, Yoshida M, Maekawa M, Kamichi S, Yoshida M, Sahara Y, Yuasa S, Takeda S, Wada K. A deficit of brain dystrophin impairs specific amygdala GABAergic transmission and enhances defensive behaviour in mice. Brain. 2008 Oct 16. [Epub ahead of print]
-
Replies
-
So what’s your take on this result, Jon? Your interpretation would be interesting. (Abstract a bit technical.)
-
Hi Maxine,
Dystrophin does a lot of jobs in different cells. The requirement for dystrophin in muscle, and the consequences of dystrophin malformation (Duchenne and Becker muscular dystrophies), are well known. The effect of Dystrophin malformation in nerve cells is a less popular topic, but critically important for parents of dystrophic children who note behavioral abnormalities and struggle to determine how to improve their childrens’ quality of life.
The use of exon-skipping oligo to treat DMD is a research focus of many molecular and clinical labs as well as several pharma companies. While the intended target of exon-skipping oligos are the muscle cells, there is the possibility that nerve cells could also benefit from dystrophin splice modification.
This paper addresses exon skipping in the brains of dystrophic mice, describing promising molecular as well as behavioral results. While intracerebroventricular cannulation is not likely to be a popular therapeutic method, the study was performed with unconjugated Morpholino oligos; advances in the delivery of Morpholinos to the cytosol by conjugation with delivery moieties (e.g. cell penetrating peptides, octaguanidinium dendrimers) suggest that much smaller doses might be therapeutically useful if delivery-enabled Morpholino conjugates can be administered to the brain side of the blood-brain barrier.
The other promising result reported in this paper is that modifying splicing of a small fraction of the total dystrophin RNA is sufficient to see a shift of behavior toward the wild-type condition. You don’t have to fix all the dystrophin to see therapeutic benefits, making development of a useful therapeutic more readily attainable.
-
