Gene Therapy Reaches Human Trials in Peripheral Nerve, Muscle

SAN FRANCISCO, CALIF. – Preliminary human trials of gene therapy are showing hints of benefit for disorders of the peripheral nerves and muscles but also are uncovering some unexpected barriers.

Investigators are testing a variety of ways to deliver potentially therapeutic genes through direct injection of vascular endothelial growth factor (VEGF) DNA into muscles or via vectors created from adeno-associated virus (AAV) or herpes simplex virus (HSV).

Although these small phase I or II trials are focused on safety outcomes, they have produced tantalizing suggestions of improvements in painful diabetic neuropathy and increased muscle size in some patients with muscular dystrophy. The muscular dystrophy studies also have identified surprising immune barriers to gene transfer in humans that have not been seen in animal studies.

The investigators described preliminary results and new insights from these early and ongoing trials in a series of talks at the annual meeting of the American Neurological Association.

Dr. Allan H. Ropper, professor of neurology at Harvard Medical School, Boston, led a randomized, blinded trial of gene therapy in 50 patients with diabetic neuropathy who received intramuscular injections of a plasmid containing VEGF or injections of saline serving as placebo. Three sets of injections were given at eight sites next to the sciatic, peroneal, and tibial nerves of one leg.

In the group of 39 patients who received the VEGF plasmid, Dr. Ropper and his associates detected modest improvement in the trial’s primary outcomes – changes in symptom scores at 6 months and a prespecified overall clinical and electrophysiological improvement score – when compared with 11 patients who received placebo. These comparisons were adjusted for any changes in the untreated leg. Overall improvement was seen in 12 of the 39 patients who got plasmid VEGF and 2 of 11 who got placebo. Improvements were seen in only two of five secondary outcomes (Ann. Neurol. 2009;65:386-93).

“Our interpretation was that this approach was promising but not definitive,” Dr. Ropper said. He called the improvements “small but genuine.”

The treatment was associated with a higher rate of serious adverse events, which did not reach statistical significance. Overall, the therapy warrants further investigation, he said.

Other promising data have come from animal studies of VEGF gene transfer before chemotherapy with thalidomide and cisplatin to prevent neuropathies associated with those drugs, Dr. Ropper noted. Human trials to prevent neuropathy of chemotherapy are being considered.

Dr. David J. Fink, who chaired the session at the meeting, described non-replicating HSV vectors for gene therapy that he and his colleagues are developing. Cutaneous injection of the vectors may deliver genes to specific targets in order to interrupt nociceptive neurotransmission selectively and better control chronic pain.

In an ongoing phase I clinical trial of patients with intractable pain from end-stage cancer, Dr. Fink and his colleagues are testing a replication defective HSV vector that carries the gene for preproenkephalin. In animal studies, the HSV vector containing the preproenkephalin gene produced enkephalin, which provided a continuous analgesic effect that was additive with morphine and persisted despite tolerance to morphine.

In the human trial, patients receive 10 injections of the vector in 100-mcL doses in the dermatome that corresponds to the radicular distribution of the pain. No serious adverse events have occurred so far, said Dr. Fink, professor and chair of neurology at the University of Michigan, Ann Arbor.

“We have only one more patient to enroll. We’ve seen no serious adverse events so far,” he added.

Diamyd Medical, which makes the vector, has agreed to start a randomized phase II trial in the fall of 2010.

A separate randomized, placebo-controlled, phase I/II study also should start in late 2010 to treat 60 patients with painful diabetic neuropathy using a different HSV vector that Dr. Fink and his associates have developed. The HSV vector expresses glutamic acid decarboxylase, the enzyme that catalyzes synthesis of the neurotransmitter gamma-aminobutyric acid (GABA). Other HSV vectors also are being explored.

For patients with Duchenne muscular dystrophy, Dr. Jerry R. Mendell reported that the concept of using a virus as a vehicle to replace a defective gene might run into unanticipated barriers presented by the human immune system. He and his associates have focused on the relatively nonpathogenic AAV, which regularly produces “therapeutic triumphs” as a gene vector in rodent studies, he said.

In humans, however, size is a problem. Duchenne muscular dystrophy is caused by mutations in the very large dystrophin gene, which is too large to transfer when incorporated into the tiny AAV. To get around this problem, the researchers deleted portions of the gene to create a “mini-dystrophin” that could be transferred using the AAV.

When investigators transferred AAV with mini-dystrophin to the biceps of six patients with Duchenne muscular dystrophy, it provoked a new and unexpected immune response when it incorporated itself into the domain of the deleted endogenous gene and was expressed. Muscle fibers that expressed the mini-dystrophin gene also developed new epitopes that were immunogenic, making it difficult for the transferred gene to escape T-cell immunity, said Dr. Mendell, director of the Center for Gene Therapy at Nationwide Children’s Hospital and professor of pediatrics and neurology at Ohio State University, both in Columbus.

In future trials, “patients should be checked for preexisting immunity to dystrophin prior to gene transfer” and the transgene should not be expressed in an area of gene deletion, he suggested.

In a separate randomized, double-blind study, Dr. Mendell and his associates successfully transferred the large limb-girdle muscular dystrophy type 2D gene via the AAV by using a muscle-specific promoter to avoid off-target gene expression (Ann. Neurol. 2009;66:290-7).

New data on 6-month follow-up in six patients who received the gene therapy injections in the extensor digitorum brevis muscle showed persistent gene expression and muscle effects in five patients, but one patient showed no increased gene expression and developed a significant immune response, he reported.

“Preexisting immunity to AAV is a potential barrier to success,” Dr. Mendell said.

Disclosures: Diamyd Medical funded Dr. Fink’s trial and provided the human-grade vectors for it. Dr. Fink has no financial relationship with Diamyd, but he is the inventor on patents that have been licensed by Diamyd from Dr. Fink’s institution and two others. Dr. Ropper and Dr. Mendell said they have no pertinent conflicts of interest.

SAN FRANCISCO, CALIF. – Preliminary human trials of gene therapy are showing hints of benefit for disorders of the peripheral nerves and muscles but also are uncovering some unexpected barriers.

Investigators are testing a variety of ways to deliver potentially therapeutic genes through direct injection of vascular endothelial growth factor (VEGF) DNA into muscles or via vectors created from adeno-associated virus (AAV) or herpes simplex virus (HSV).

Although these small phase I or II trials are focused on safety outcomes, they have produced tantalizing suggestions of improvements in painful diabetic neuropathy and increased muscle size in some patients with muscular dystrophy. The muscular dystrophy studies also have identified surprising immune barriers to gene transfer in humans that have not been seen in animal studies.

The investigators described preliminary results and new insights from these early and ongoing trials in a series of talks at the annual meeting of the American Neurological Association.

Dr. Allan H. Ropper, professor of neurology at Harvard Medical School, Boston, led a randomized, blinded trial of gene therapy in 50 patients with diabetic neuropathy who received intramuscular injections of a plasmid containing VEGF or injections of saline serving as placebo. Three sets of injections were given at eight sites next to the sciatic, peroneal, and tibial nerves of one leg.

In the group of 39 patients who received the VEGF plasmid, Dr. Ropper and his associates detected modest improvement in the trial’s primary outcomes – changes in symptom scores at 6 months and a prespecified overall clinical and electrophysiological improvement score – when compared with 11 patients who received placebo. These comparisons were adjusted for any changes in the untreated leg. Overall improvement was seen in 12 of the 39 patients who got plasmid VEGF and 2 of 11 who got placebo. Improvements were seen in only two of five secondary outcomes (Ann. Neurol. 2009;65:386-93).

“Our interpretation was that this approach was promising but not definitive,” Dr. Ropper said. He called the improvements “small but genuine.”

The treatment was associated with a higher rate of serious adverse events, which did not reach statistical significance. Overall, the therapy warrants further investigation, he said.

Other promising data have come from animal studies of VEGF gene transfer before chemotherapy with thalidomide and cisplatin to prevent neuropathies associated with those drugs, Dr. Ropper noted. Human trials to prevent neuropathy of chemotherapy are being considered.

Dr. David J. Fink, who chaired the session at the meeting, described non-replicating HSV vectors for gene therapy that he and his colleagues are developing. Cutaneous injection of the vectors may deliver genes to specific targets in order to interrupt nociceptive neurotransmission selectively and better control chronic pain.

In an ongoing phase I clinical trial of patients with intractable pain from end-stage cancer, Dr. Fink and his colleagues are testing a replication defective HSV vector that carries the gene for preproenkephalin. In animal studies, the HSV vector containing the preproenkephalin gene produced enkephalin, which provided a continuous analgesic effect that was additive with morphine and persisted despite tolerance to morphine.

In the human trial, patients receive 10 injections of the vector in 100-mcL doses in the dermatome that corresponds to the radicular distribution of the pain. No serious adverse events have occurred so far, said Dr. Fink, professor and chair of neurology at the University of Michigan, Ann Arbor.

“We have only one more patient to enroll. We’ve seen no serious adverse events so far,” he added.

Diamyd Medical, which makes the vector, has agreed to start a randomized phase II trial in the fall of 2010.

A separate randomized, placebo-controlled, phase I/II study also should start in late 2010 to treat 60 patients with painful diabetic neuropathy using a different HSV vector that Dr. Fink and his associates have developed. The HSV vector expresses glutamic acid decarboxylase, the enzyme that catalyzes synthesis of the neurotransmitter gamma-aminobutyric acid (GABA). Other HSV vectors also are being explored.

For patients with Duchenne muscular dystrophy, Dr. Jerry R. Mendell reported that the concept of using a virus as a vehicle to replace a defective gene might run into unanticipated barriers presented by the human immune system. He and his associates have focused on the relatively nonpathogenic AAV, which regularly produces “therapeutic triumphs” as a gene vector in rodent studies, he said.

In humans, however, size is a problem. Duchenne muscular dystrophy is caused by mutations in the very large dystrophin gene, which is too large to transfer when incorporated into the tiny AAV. To get around this problem, the researchers deleted portions of the gene to create a “mini-dystrophin” that could be transferred using the AAV.

When investigators transferred AAV with mini-dystrophin to the biceps of six patients with Duchenne muscular dystrophy, it provoked a new and unexpected immune response when it incorporated itself into the domain of the deleted endogenous gene and was expressed. Muscle fibers that expressed the mini-dystrophin gene also developed new epitopes that were immunogenic, making it difficult for the transferred gene to escape T-cell immunity, said Dr. Mendell, director of the Center for Gene Therapy at Nationwide Children’s Hospital and professor of pediatrics and neurology at Ohio State University, both in Columbus.

In future trials, “patients should be checked for preexisting immunity to dystrophin prior to gene transfer” and the transgene should not be expressed in an area of gene deletion, he suggested.

In a separate randomized, double-blind study, Dr. Mendell and his associates successfully transferred the large limb-girdle muscular dystrophy type 2D gene via the AAV by using a muscle-specific promoter to avoid off-target gene expression (Ann. Neurol. 2009;66:290-7).

New data on 6-month follow-up in six patients who received the gene therapy injections in the extensor digitorum brevis muscle showed persistent gene expression and muscle effects in five patients, but one patient showed no increased gene expression and developed a significant immune response, he reported.

“Preexisting immunity to AAV is a potential barrier to success,” Dr. Mendell said.

Disclosures: Diamyd Medical funded Dr. Fink’s trial and provided the human-grade vectors for it. Dr. Fink has no financial relationship with Diamyd, but he is the inventor on patents that have been licensed by Diamyd from Dr. Fink’s institution and two others. Dr. Ropper and Dr. Mendell said they have no pertinent conflicts of interest.

SAN FRANCISCO, CALIF. – Preliminary human trials of gene therapy are showing hints of benefit for disorders of the peripheral nerves and muscles but also are uncovering some unexpected barriers.

Investigators are testing a variety of ways to deliver potentially therapeutic genes through direct injection of vascular endothelial growth factor (VEGF) DNA into muscles or via vectors created from adeno-associated virus (AAV) or herpes simplex virus (HSV).

Although these small phase I or II trials are focused on safety outcomes, they have produced tantalizing suggestions of improvements in painful diabetic neuropathy and increased muscle size in some patients with muscular dystrophy. The muscular dystrophy studies also have identified surprising immune barriers to gene transfer in humans that have not been seen in animal studies.

The investigators described preliminary results and new insights from these early and ongoing trials in a series of talks at the annual meeting of the American Neurological Association.

Dr. Allan H. Ropper, professor of neurology at Harvard Medical School, Boston, led a randomized, blinded trial of gene therapy in 50 patients with diabetic neuropathy who received intramuscular injections of a plasmid containing VEGF or injections of saline serving as placebo. Three sets of injections were given at eight sites next to the sciatic, peroneal, and tibial nerves of one leg.

In the group of 39 patients who received the VEGF plasmid, Dr. Ropper and his associates detected modest improvement in the trial’s primary outcomes – changes in symptom scores at 6 months and a prespecified overall clinical and electrophysiological improvement score – when compared with 11 patients who received placebo. These comparisons were adjusted for any changes in the untreated leg. Overall improvement was seen in 12 of the 39 patients who got plasmid VEGF and 2 of 11 who got placebo. Improvements were seen in only two of five secondary outcomes (Ann. Neurol. 2009;65:386-93).

“Our interpretation was that this approach was promising but not definitive,” Dr. Ropper said. He called the improvements “small but genuine.”

The treatment was associated with a higher rate of serious adverse events, which did not reach statistical significance. Overall, the therapy warrants further investigation, he said.

Other promising data have come from animal studies of VEGF gene transfer before chemotherapy with thalidomide and cisplatin to prevent neuropathies associated with those drugs, Dr. Ropper noted. Human trials to prevent neuropathy of chemotherapy are being considered.

Dr. David J. Fink, who chaired the session at the meeting, described non-replicating HSV vectors for gene therapy that he and his colleagues are developing. Cutaneous injection of the vectors may deliver genes to specific targets in order to interrupt nociceptive neurotransmission selectively and better control chronic pain.

In an ongoing phase I clinical trial of patients with intractable pain from end-stage cancer, Dr. Fink and his colleagues are testing a replication defective HSV vector that carries the gene for preproenkephalin. In animal studies, the HSV vector containing the preproenkephalin gene produced enkephalin, which provided a continuous analgesic effect that was additive with morphine and persisted despite tolerance to morphine.

In the human trial, patients receive 10 injections of the vector in 100-mcL doses in the dermatome that corresponds to the radicular distribution of the pain. No serious adverse events have occurred so far, said Dr. Fink, professor and chair of neurology at the University of Michigan, Ann Arbor.

“We have only one more patient to enroll. We’ve seen no serious adverse events so far,” he added.

Diamyd Medical, which makes the vector, has agreed to start a randomized phase II trial in the fall of 2010.

A separate randomized, placebo-controlled, phase I/II study also should start in late 2010 to treat 60 patients with painful diabetic neuropathy using a different HSV vector that Dr. Fink and his associates have developed. The HSV vector expresses glutamic acid decarboxylase, the enzyme that catalyzes synthesis of the neurotransmitter gamma-aminobutyric acid (GABA). Other HSV vectors also are being explored.

For patients with Duchenne muscular dystrophy, Dr. Jerry R. Mendell reported that the concept of using a virus as a vehicle to replace a defective gene might run into unanticipated barriers presented by the human immune system. He and his associates have focused on the relatively nonpathogenic AAV, which regularly produces “therapeutic triumphs” as a gene vector in rodent studies, he said.

In humans, however, size is a problem. Duchenne muscular dystrophy is caused by mutations in the very large dystrophin gene, which is too large to transfer when incorporated into the tiny AAV. To get around this problem, the researchers deleted portions of the gene to create a “mini-dystrophin” that could be transferred using the AAV.

When investigators transferred AAV with mini-dystrophin to the biceps of six patients with Duchenne muscular dystrophy, it provoked a new and unexpected immune response when it incorporated itself into the domain of the deleted endogenous gene and was expressed. Muscle fibers that expressed the mini-dystrophin gene also developed new epitopes that were immunogenic, making it difficult for the transferred gene to escape T-cell immunity, said Dr. Mendell, director of the Center for Gene Therapy at Nationwide Children’s Hospital and professor of pediatrics and neurology at Ohio State University, both in Columbus.

In future trials, “patients should be checked for preexisting immunity to dystrophin prior to gene transfer” and the transgene should not be expressed in an area of gene deletion, he suggested.

In a separate randomized, double-blind study, Dr. Mendell and his associates successfully transferred the large limb-girdle muscular dystrophy type 2D gene via the AAV by using a muscle-specific promoter to avoid off-target gene expression (Ann. Neurol. 2009;66:290-7).

New data on 6-month follow-up in six patients who received the gene therapy injections in the extensor digitorum brevis muscle showed persistent gene expression and muscle effects in five patients, but one patient showed no increased gene expression and developed a significant immune response, he reported.

“Preexisting immunity to AAV is a potential barrier to success,” Dr. Mendell said.

Disclosures: Diamyd Medical funded Dr. Fink’s trial and provided the human-grade vectors for it. Dr. Fink has no financial relationship with Diamyd, but he is the inventor on patents that have been licensed by Diamyd from Dr. Fink’s institution and two others. Dr. Ropper and Dr. Mendell said they have no pertinent conflicts of interest.