The patent covers the use of two AAV vectors to deliver DNA sequences that, once inside a cell, are used to produce or regulate a single protein. This approach allows AAV vectors to be used to deliver genes and their regulatory sequences that ordinarily would be too large to fit inside a single vector.
The patent is set to greatly broaden Targeted Genetics Corporation's range of molecular medicines for preventing and treating acquired and inherited diseases. The company currently has programs targeting AIDS and inflammatory arthritis as well as programs focused on hyperlipidemia, congestive heart failure, and Huntington's disease.
Adeno-associated virus (AAV) is a non-pathogenic parvovirus with a single-stranded DNA genome of 4680 nucleotides. The genome may be of either plus or minus polarity, and codes for two groups of genes, Rep and Cap. Inverted terminal repeats (ITRs), characterised by palindromic sequences producing a high degree of secondary structure, are present at both ends of the viral genome.
While other members of the parvovirus group replicate autonomously, AAV requires co-infection with a helper virus (i.e. adenovirus or herpes virus) for lytic phase productive replication. In the absence of a helper virus, wild-type AAV (wtAAV) establishes a latent, non-productive infection with long-term persistence by integrating into a specific locus on chromosome 19, AAVS1, of the host genome through a Rep-facilitated mechanism.
"To date we have been able to address the size constraint of AAV vectors for our product development programs by utilising small promoter elements or by modifying DNA sequences to reduce their size without affecting the protein that ultimately is produced," said Barrie Carter, executive vice president and chief scientific officer of Targeted Genetics.
"The technology covered by this patent significantly expands the potential application of AAV vectors by providing a more flexible method to deliver genes and their associated regulatory elements that are larger than the normal carrying capacity of these vectors," he added.
AAV may aid in the development of non-viral or viral-based gene delivery systems having increased efficiency. For example, therapeutic or prophylactic therapies in which the present vectors are useful include blood disorders (e.g., sickle cell anaemia, thalassemias, haemophilias, and Fanconi anaemia's), neurological disorders, such as Alzheimer's disease and Parkinson's disease, and muscle disorders involving skeletal, cardiac or smooth muscle.
In particular, therapeutic genes useful in the vectors of the invention include the â-globin gene, the ã-globin gene, the cystic fibrosis transmembrane conductance receptor gene (CFTR), the Fanconi anaemia complementation group, a gene encoding a ribozyme, an antisense gene, a low density lipoprotein (LDL) gene, a tyrosine hydroxylase gene (Parkinson's disease), a glucocerebrosidase gene (Gaucher's disease), an arylsulfatase A gene (metachromatic leukodystrophies) or genes encoding other polypeptides or proteins.
"We then engineered vectors in a manner that causes this assembly to join two pieces of RNA that together serve as a template for regulating a single protein. We recognised that this approach is very powerful because it allows us to double the size of the genes and their regulatory sequences that one can deliver with AAV vectors," he added.