Penn Medicine studies unlock new insights into gene therapy

The studies on AAV-based gene therapies in non-human primates suggest that integration into human DNA is unlikely to drive cancer mutations.

Gene therapy adeno-associated viruses (AAVs)—viruses that can be engineered to deliver DNA to target cells—are unlikely to cause cancer-triggering insertions in humans or monkeys and may contribute to long-term efficacy, according to new research from the University of Pennsylvania’s Gene Therapy Program (GTP). AAVs that are used to deliver gene therapies sometimes end up being inserted into chromosomal DNA, which has led to concerns that such insertions could disrupt the host genome, potentially causing cellular dysfunction or the development of cancer.

String of colorful DNA.
Image: iStock/natrot

Two large companion studies in non-human primates indicate that vector integrations in primate liver following AAV gene therapy may be an important mechanism for achieving durable expression and are unlikely to induce cancer mutations in humans. The two new studies are both published in Nature Biotechnology and Human Gene Therapy.

Viruses evolved to break into cells, and gene therapy researchers have long viewed them as the go-to carriers or “vectors” for therapeutic transgenes. AAVs are the most commonly used viral vectors for gene therapies because they can enter both dividing and non-dividing cells, don’t cause viral illness in humans, and usually don’t trigger an immune response—which would swiftly degrade their therapeutic impact.

AAVs and the therapeutic transgenes they can carry normally exist “episomally”—free-floating in the cell nucleus and not integrated into chromosomal DNA. However, scientists now recognize that engineered AAVs used for gene therapies do sometimes end up in chromosomal DNA, perhaps mainly as a result of DNA repair processes that inadvertently stitch them into the genome.

In the Human Gene Therapy study, Penn researchers examined tissue samples from 86 macaque monkeys that had been treated with AAV-based gene therapies in preclinical tests, and 253 macaques and humans that had never been exposed to engineered AAVs. They found that the engineered AAVs in the gene therapy recipients, and endemic natural (“wild-type”) AAVs in the other group, were inserted at low rates in mostly random distributions across the genome with a low risk for expansion, even in a monkey 15 years after treatment.

In the Nature Biotechnology study, Wilson’s team followed 12 macaques for over 2 years after they received AAV-based gene therapies targeting liver cells. They found the presence of complex structures of AAV genomes existing outside of the chromosome that persist but appear to be rapidly inactivated soon after vector delivery.

“These studies inform new approaches for improving liver gene therapy that focus more on expression rather than delivery,” says study senior author James M. Wilson, director of the Gene Therapy Program, Rose H. Weiss Professor and Director of the Orphan Disease Center, and professor in the Departments of Medicine and Pediatrics at the Perelman School of Medicine. “Our data also highlight the potential advantages of genome editing for liver in which insertions are directed to safe harbor regions of the chromosome above the background of more widely distributed integrations that occur with vector alone.”’

Read more at Penn Medicine News.