After some tough times last year due to investor concern over thepace of technology development, gene-therapy companies are on therebound. Last month's grant of a broad patent on ex vivo therapy toGenetic Therapy Inc. has boosted expectations further. (SeeBioWorld Today, March 22, 11995, p. 1.)

But delivering the therapeutic gene is still a difficult challenge; onethat can differ markedly from one disease to the next. Long-termmaintenance of transgene expression also requires furtherdevelopment.

In the April issue of Nature Medicine, the Pasteur Institute reports afurther advance in gene delivery technology. A paper titled "Long-term delivery of a lysosomal enzyme by genetically modifiedfibroblasts in dogs," describes the efforts of Philippe Moullier and hisco-authors to devise viable gene therapy for Hurler's disease.

They showed that intraperitoneal implants _ so-called "neo-organs"_ of fibroblasts transfected with the human b-glucuronidase gene,and grown in collagen lattices, could deliver therapeutically effectivedoses of the human protein to sizable recipient canines. The neo-organs remained well-vascularized with no apparent localinflammation. This feat extended the group's previous successfulconstruction of neo-organs in mice.

Next: Hurler's Disease Therapy In Humans

Gene therapist and senior author Jean Michel Heard told BioWorldToday: "Now that we have been able to show secretion of b-glucuronidase in dogs, an animal about the size of a child, at levelsbetween about 1 and 3 percent of normal for up to one year, our nextstep is to move into clinical trials. We will use essentially the sameapproach in an attempt to treat children with Hurler's disease. We arehopeful that there will be a beneficial effect because we already knowthat these affected children only need serum b-glucuronidase levelsthat are 0.5 percent of normal to reverse their disease."

The Pasteur Institute has submitted these gene-therapy protocols to the five appropriate regulatory agencies inFrance, two of which have already approved them. "We expect toreceive the remaining approvals and be able to begin the trials by thissummer," Heard said.

Selected Vectors For Specific Delivery Strategies

He explained: "The gene therapy techniques we used are verydependent on the disease one is treating. In general, retroviral vectorshave been used for blood cells; adenoviral vectors and liposomes forreimplanted cells, or direct in vivo delivery."

National Institutes of Health Clinician Michael Blaese, a genetherapy pioneer and co-inventor on Genetic Therapy Inc.'s new exvivo gene therapy patent, told BioWorld Today: "Retroviral vectorsintegrate, so all daughter cells get a copy of the inserted gene.Adenoviral vectors remain episomal and therefore do not integrate.So, division of cells transfected with adenoviral vectors dilutes outthe gene. But high levels of transfer usually means that each cell getsmultiple copies of the adenoviral vector."

Blaese elaborated, "As a result, retroviral vectors tend to be used fortarget cells that will divide, and adenoviral vectors tend to be used forquiescent target cells. The clinical setting and characteristics of theparticular hereditary disease being treated, determine the systemsused for gene therapy."

Gene Silencing In Reimplanted Cells

Heard said, "A major concern is whether expression of the gene willeventually be extinguished, given the slow decline seen over time inour dogs. In mice, where we inserted the mouse gene and thereforedid not have any immune response to complicate matters, we still sawthis gradual loss of expression. The latter indicates that the slow lossof gene expression is a general phenomenon."

Blaese concurred: "Whether the genes will continue to express whencells are put back into the body," he said, "is always a gene therapyproblem. Cells express incorporated genes in culture when they aregrowing. But, when placed in the body, cell growth slows, andexpression is lost over time.

"We see this in T cell therapy for adenosine deaminase [ADA]deficiency. When the T cells are removed from these patients after aperiod of time in their body, they grow again in cell culture and alsoincrease their expression of ADA."

But overall, Blaese is hopeful for the clinical success of neo-organs."The potential need for yearly treatment with additional implantsshould probably not be a problem," he surmised. "However, becausescar formation can occur in the intraperitoneal sites, other sites, suchas the abdominal wall muscle, may be preferable. Andreimplantation, at least in some cases, can be a solution to the gradualloss of gene expression." n

-- Chester Bisbee Special To BioWorld Today

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