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Gene therapy researchers repair rather than replace damaged genes
  
By Bijal P. Trivedi

Researchers have found a way to correct single letter spelling mistakes within genes. Once perfected, the approach could provide the ultimate treatment for genetic diseases caused by single letter, or point, mutations.

Recent gene therapy experiments have focused on delivering a healthy gene rather than repairing a damaged copy, but the problem with this approach is that the replacement gene rarely functions correctly after being randomly inserted into a chromosome. Genes are regulated by genetic switches, which in some cases lie far away on the chromosome. A gene that is delivered without its chromosomal context usually does not function normally. This approach is also accompanied by the danger that insertion of the therapeutic gene into a chromosome will disrupt the activity of other genes. Repairing avoids both these problems.

David Russell, of the University of Washington in Seattle, and colleagues tested their method of gene correction by introducing point mutations into the hypoxanthine phosphoribosyl transferase (HPRT) gene in human cells.

Russell's team used six viruses each carrying an HPRT gene with a different point mutation. They observed that the viral gene targets the healthy version in the cells and, through a mechanism that is not yet understood, transfers the mutation.

The researchers took cells containing the healthy version of HPRT and used the viruses to create six mutants. They altered one of two bases—adenine (A) or guanine (G)—at a unique location within the gene and created six other combinations of AG: TG, AA, AT, CG, AC, and GG. The experiments illustrate that a single nucleotide among the three billion others in the genome can be specifically corrected.

In a second experiment, the researchers created five different mutants of an alkaline phosphatase gene and used gene therapy to repair each mutant.

Russell and his colleagues are not exactly certain how the repair occurs. A virus carries a healthy copy of the gene into the cell and aligns itself with the mutated copy on the chromosome. This healthy gene might serve as a template that the cell uses to repair its mutated gene. Or, the healthy gene carried by the virus might be directly incorporated into the chromosome. The process could also be a combination of the two, according to Russell.

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Inoue, N. et al. Introduction of single base substitutions at homologous chromosomal sequences by adeno-associated virus vectors. Mol Ther 3, 526-530 (April 2001).
 
Porter, A.C.G. Correcting a deficiency. Mol Ther 3, 423-424 (April 2001).
 

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