Labs Find Microdystrophin Genes Effective

Good news about miniaturized (mini- and micro-) dystrophin genes has recently come from three labs, all of which have received MDA support. Dystrophin genes are the instructions for the muscle protein dystrophin, missing in boys with Duchenne muscular dystrophy (DMD).

The large dystrophin gene requires miniaturization before it can be given as gene therapy, a process that will result in smaller than normal dystrophin protein molecules. New strategies, such as exon skipping, that cause cells to skip over parts of the dystrophin gene that contain errors, will likewise lead to production of shortened forms of dystrophin.

Dongsheng Duan at the University of Missouri-Columbia and Jeffrey Chamberlain at the University of Washington in Seattle have shown that microdystrophin genes missing instructions for a section at one end, called the C terminal, and part of a midsection called the central rod domain, can provide effective treatment for mice with a severe disease resembling DMD.

Duan’s group, which published results online March 21 in Molecular Therapy, inserted microdystrophin genes originally developed in Chamberlain’s lab into transport vehicles made from adeno-associated viruses. They then injected them into the leg muscles of mice missing both dystrophin and utrophin.

These mice, known as double knockouts, develop a disease that more closely resembles human DMD than do mice missing dystrophin alone.

They found the new genes eliminated scarring and inflammation in the treated muscles, increased muscle force, reduced contraction-related damage, and restored muscle cell membrane proteins to their appropriate positions.

The highly truncated genes also allowed syntrophins and dystrobrevin — proteins thought to carry signals in muscle — to take their places near the cell membrane.

Chamberlain’s group went a step further by delivering the gene systemically to double knockout mice. They also used adeno-associated viral transport vehicles, but they delivered the genes intravenously.

Reporting their results at the New Directions in Skeletal Muscle Biology conference held in Dallas in April, the team said they saw dystrophin production in limb and respiratory muscles, increased muscle function, and a longer life span in the treated, compared to the untreated, mice.

Also this spring, researchers associated with the laboratory of Robert White at the University of Missouri-Kansas City, including MDA grantee Stephen Hauschka at the University of Washington, used an entirely different type of microdystrophin gene and found that it, too, conferred significant benefits.

This group bred double knockout mice to produce in their skeletal muscles a form of dystrophin normally found only in the eye’s retina. This form of dystrophin, known as Dp260, is missing the N terminal, at the opposite end of dystrophin from the C terminal, as well as some of the midsection of the protein. The N terminal is involved in anchoring dystrophin to the inside of the cell.

This team announced in the March issue of Neuromuscular Disorders that the mice bred to produce Dp260 developed only a very mild muscle disease, grew and gained weight normally, and had spinal curvatures like those seen in normal mice, in contrast to the severe curvatures that double knockout mice develop. They also lived longer than their untreated counterparts. (An earlier report from Chamberlain’s lab showed that Dp260 could partially protect dystrophin-deficient muscles in mice.)

To restore muscle cell function, two factors appear to be necessary: dystrophin’s dystroglycan binding domain, where it attaches to the dystroglycan protein in the muscle cell membrane, and at least some of the central rod domain.