LONDON - An international team has pinpointed the gene for a rare inherited kidney disorder called nephropathic cystinosis.

The discovery will allow faster and more accurate prenatal diagnosis for affected families and will pave the way for development of more effective drugs to treat the disease.

Nephropathic cystinosis affects an estimated one in 200,000 people worldwide. About 400 people in the U.S. and a similar number in Europe are thought to suffer from it.

The disease, which is inherited in an autosomal recessive manner, results from the accumulation of the amino acid cystine in subcellular organelles called lysosomes. These are normally responsible for breaking down peptides and proteins and releasing their component amino acids back into the cytoplasm to re-enter metabolic pathways.

In affected children, the intracellular accumulation of cystine damages all tissues. In the kidneys, this damage results in severe fluid and electrolyte disturbance, which manifests itself with vomiting, poor growth and rickets by the age of six to 12 months. Without treatment, renal failure occurs by about age 10. Although this can be treated by renal transplantation, accumulation of cystine in other organs continues to cause significant health problems.

The gene encoding the lysosomal membrane cystine transporter, called cystinosin, has now been identified by a group of researchers in the U.K., France and Australia. The team was led by Margaret Town, research fellow at the Division of Medical and Molecular Genetics at the United Medical and Dental Schools, in London, and by Corinne Antignac, director of research at France's National Institute for Health and Medical Research (Institut National de la Santé et de la Recherche Médicale - INSERM), in Paris. The scientists report their results in a paper in the April issue of Nature Genetics, titled “A novel gene encoding an integral membrane protein is mutated in nephropathic cystinosis.“

Gene's Chromosomal Region Already Known

Town told BioWorld International, “Nephropathic cystinosis, although rare, is a devastating disease, and this is why we became involved in studying it. The discovery of the gene is the first step towards identifying the mechanism by which cystine is transported across the lysosomal membrane or - in the case of those with cystinosis-is not transported across the membrane. This will allow us in the future to have a rationale for the design of new therapies to treat this disease.“

In 1995, an international consortium mapped the gene to a region on chromosome 17p. Following this, while attempting to narrow the genetic interval for the cystinosis gene, Town, Antignac and their colleagues found a DNA marker, present in this candidate region, was deleted in some patients with cystinosis.

Town said, “We could not amplify this marker in some of the cystinosis patients, whereas we could amplify it in all the controls we tried. When we looked in the cystinosis families, we found that this apparent deletion segregated with the disease.“

This finding led the researchers to concentrate on the region containing the deletion. Using several methods, they determined that the extent of the deletion in the patients was 65 kilobases. They then carried out “exon trapping,“ a technique designed to identify the coding sequences within a gene. This allowed them to identify a novel gene which has been called CTNS.

The CTNS gene encodes a protein of 367 amino acids. With the help of specialized computer programs, Town and her colleagues predicted the tertiary structure of the protein, cystinosin. They found the molecule has six or seven membrane-spanning domains, and some features in common with known lysosomal membrane proteins.

“So this looked like it was an extremely good candidate for the lysosomal membrane cystine transporter,“ Town said.

To prove CTNS really is the gene responsible for cystinosis, however, the team needed to find mutations present in the gene in patients but not in controls.

After further studies, the group found an additional 11 small mutations in the gene in cystinosis patients. In each patient, both copies of chromosome 17 carried a mutation in CTNS, which is consistent with an autosomal recessive mode of inheritance. The mutations were not present in controls, and in affected families they segregated with the disease.

“Together with the large deletion already identified,“ Town concluded, “we believe that these data demonstrate that the gene we had found was indeed the gene responsible for cystinosis.“

Interestingly, the large deletion - though found in almost half the patients studied - was only present in patients of northern European origin, not in those from Africa, Asia or the Middle East.

Town said, “This might suggest a founder effect. If this was the case, you would expect to see a common haplotype - that is, a similar arrangement of genetic markers around this region - in individuals with the deletion. But we did not find any evidence of a common haplotype. This suggests that, if there is a founder effect, it would be a very ancient event for the different haplotypes to have arisen since then.“

An alternative explanation is that the deletion may have arisen several times independently, as a result of unstable genomic sequences.

Prenatal Testing May Be First Application

One immediate spin-off from the finding will be a more rapid and accurate prenatal test for families who require it.

In the longer term, Town said, the group wants to examine cystinosin in detail and try to identify its exact role in the transport of cystine across the lysosomal membrane. The researchers also want to identify other proteins which interact with it.

Another aim is to characterize mutations present in a larger group of patients. “We want to find out if there is any clustering of mutations in a particular part of the gene,“ Town said. “So far, all the mutations we have found are predicted to be null mutations which would result in loss of protein function. But we would like to find out if there are any mutations which cause altered protein structure or function rather than no protein at all. We would then try to correlate the severity and characteristics of the disease with the type of mutation.“

This will include a study of the rarer, late-onset form of the disease, in order to find out if the mutations in these patients differ significantly from the mutations in the nephropathic cystinosis group. *