Editing Dystrophin Genes

Researcher Carmen Bertoni is using DNA-like material to fix flawed genes

Carmen Bertoni says gene editing has the potential, when optimized, to "completely cure the disease."
Article Highlights:
  • Bertoni is experimenting with a gene-editing strategy that involves laboratory-engineered compounds that resemble DNA and stick to dystrophin genes, where they can trigger a cellular editing mechanism called mismatch repair.
  • The gene-editing approach is working in mice with a mutation in the dystrophin gene, but Bertoni says the level of correction needs to be improved before the strategy can be tested in patients with Duchenne muscular dystrophy.
  • Bertoni’s current MDA funding involves correcting dystrophin genes in muscle stem cells to provide a continuing supply of muscle repair kits.
  • This article is part of the Quest series titled Gene Therapy: The Next Generation (see below for the other stories included in the special package).
by Margaret Wahl on January 9, 2014 - 9:19am

Quest Winter 2014

When Carmen Bertoni came to the University of Tulane in New Orleans from her native Italy back in 1995 to study the burgeoning science of molecular genetics, she knew she wanted to study genetic disorders but not solely to advance the field. "I wanted to work on something that would potentially be curable within my lifetime," she says.

In addition, she knew she wanted to work on a genetic disorder that affects young children, not just for emotional reasons but because, she says, it's easier to isolate the genetic roots of a disease in the body before the effects of aging and environmental influences have muddied the waters too much.

After getting her doctoral degree in 1999, Bertoni did postdoctoral studies at Stanford University in California until 2006, and then came to the University of California, Los Angeles.

"I wanted a good climate and a good environment," she says. "The weather in California is great, and the United States is the best place to do science — no question about it."

Aiming at a cure through mismatch repair

Bertoni has watched gene-transfer therapy and gene-modification strategies for Duchenne muscular dystrophy (DMD) as they've developed since the early 1990s, and there's one thing that particularly worries her.

She isn't sure that the shortened versions of the dystrophin protein produced by inserting highly miniaturized dystrophin genes used in gene transfer or created by a gene-modification strategy called exon skipping will be fully functional in the body's muscles.

The current strategies, Bertoni says, can at best convert a very severe muscular dystrophy to a less severe one. On the other hand, she says that the strategy she's chosen to pursue — which she calls gene editing — has the potential, when optimized, "to completely cure the disease."

Bertoni's gene-editing strategy involves laboratory-engineered compounds that resemble DNA (but aren't exactly DNA) and are designed to target and stick to dystrophin genes, where they can trigger a natural cellular editing mechanism. (For those who want to know, they're called PNA-ssODNs, or peptide nucleic acid single-stranded oligodeoxynucleotides.)

Genes are made of strings of chemicals called nucleotides, and genes with one erroneous nucleotide are said to have point mutations. It's this type of mutation that Bertoni wants to fix.

She's been working with a dystrophin-deficient mouse that has a point mutation, using a compound that mirrors the mouse's dystrophin gene and sticks to it, as mirror images often do in biology. But it's not a perfect mirror image. Where the mouse Bertoni is studying has a flaw — the wrong nucleotide — at a specific point in the gene, the therapeutic compound Bertoni is using carries a corrective nucleotide.

When the gene-editing process takes place successfully, she says, the correction will cause the flawed dystrophin gene in the mouse to self-correct and then produce completely functional, full-length dystrophin protein.

The corrective mechanism — known as mismatch repair — is one that normally operates in people and other organisms, although it isn't perfect. (Without mismatch repair, Bertoni says, everyday phenomena like DNA damage from the sun would cause cancer in short order.)

In the mice, Bertoni says, her gene-editing approach is working, but "the level of correction is low. It's suboptimal, but we're trying to make it better. We need to increase the frequency of gene correction." Only then will she feel that her compounds should be tested in children with DMD.

A long-lasting fix

Another concern of Bertoni's is that many current therapeutic approaches that add new genes or modify existing ones will need to be given repeatedly throughout a person's life, potentially leading to complications.

Her method, however, could be a long-lasting fix — especially if the corrective compounds get into muscle stem cells and not just mature muscle cells. Muscle stem cells known as satellite cells naturally repair damaged muscle fibers in the body, but their repairs can only be fully effective if they carry a fully functional toolkit — including a corrected dystrophin gene.

A gene-editing technique that targets only mature muscle fibers will only last as long as the fibers do, but a technique that targets muscle stem cells can provide a continuing supply of repair kits. Targeting stem cells with gene editing is a focus of Bertoni's current MDA research grant.

Bertoni says she's not yet ready to make any promises, but she hopes to take her gene-editing strategy into clinical trials in the not-too-distant future.

This article is part of a special series titled Gene Therapy: The Next Generation, which includes these additional articles profiling researchers who are pursuing next-generation gene therapy for DMD:

For more information about Bertoni's gene therapy research, be sure to read:

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