Taking the chance out of CF diagnostics

Cystic Fibrosis is currently diagnosed with a sweat test – but is this the best way?

Cystic Fibrosis (CF) is an extremely serious condition which affects multiple organs and shortens the lifespan significantly. The median age for survival is around 30 years and the leading cause of death is a progressive decline in pulmonary functions due to excessive secretions and chronic microbial infections. Other complications include exocrine pancreas insufficiency affecting roughly 85% of patients, meconium ileus in 15% of infants, CF-related diabetes mellitus also in approximately 15% of patients and severe liver disease in 5%. Most males affected by the disease are infertile. A progressive deterioration in quality of life starts in childhood years and continues throughout the sufferer’s lifetime with amelioration basically only possible if disease therapy is allowed to start as soon as possible.

The disease affects the Caucasian population with 4% of Europeans carrying one CF allele. As cystic fibrosis is an autosomal regressive disease most people do not suffer from the disease since one normally functioning allele will prevent development of the disease. In other words, only one working copy of the CFTR gene is needed to prevent cystic fibrosis.  The CFTR gene regulates transmembrane conductance which directly impacts the production of components of mucous, sweat and digestive juices. When a malfunction occurs, secretions are over-produced and are of a very thickened consistency resulting in the typical associated symptoms.

Over 1,500 mutations in this gene have been identified to date in different populations and the Astra Biotech microarray-on-a-chip includes 25 of the most commonly found ones according to the 2009 Best Practice Guidelines for molecular genetic diagnosis of cystic fibrosis and CFTR-related disorders (updated European recommendations): 17 of these mutations are particularly common in Western European, others are found in Eastern European populations. Thus, Astra Biotech’s microarray panel is designed for the European area including Central European and Slavic population groups. One mutation includes a commonly appearing gene in Turkish populations.

“The median age for survival is around 30 years and the leading cause of death is a progressive decline in pulmonary functions due to excessive secretions and chronic microbial infections”

Due to nature of the disease and the relatively high rate of carriers, the World Health Organisation recommends Caucasian individuals be tested, particularly in cases of family history of the disease and in cases of couples of Caucasian descent or with a family history. The most common tests carried out to detect the presence of the disease are sweat tests, and in newborns a screening for immune-reactive trypsinogen is performed. Tests are routinely carried out in many countries after birth in cases of risk or where symptoms of the disease are already apparent.

However, the most common of tests, the sweat test, is not in all cases reliable. The test is based on the fact that sufferers’ sweat will have a higher than normal amount of chloride or sodium in their sweat. In cases of mild mutations test results may be borderline or not alert healthcare staff, or may turn out false positive or false negative results. Especially due to false positives and false negatives, neonatal screenings can be considered controversial in some parts of the world. Not all countries screen this early, thus allowing over 10% of sufferers to be diagnosed after childhood when all symptoms have already done considerable damage to the individual’s organs. To counteract damage to vital organs, the trend in healthcare facilities is clearly towards more proactive and therefore earlier treatment of the patients; equally, the trend in diagnosing the disease should be similar. The earlier the diagnosis, the earlier disease management may be commenced. In keeping with these facts, the earliest and most reliable diagnosis is a genetic test.  If one or both parents are found to be carriers, a test on the foetus can be performed. The test itself is DNA-based and uses reverse hybridisation. It can be carried out on people with a family history of cystic fibrosis, on newborns and prenatal diagnostics, and in family planning.

The Astra Biotech microarray consists of a biochip with the named CFTR mutations, wild-type sequences and the respective mutation sequence. In positive cases for the mutation corresponding signals will appear in the mutation area. The results are read by a biochip scanner. If only the wild-types produce fluorescent signals after hybridisation, test results are negative.

A variety of features of the Astra Biotech microarray-on-a-chip contribute to better interpretation of results and are easier to use in laboratories. Specially treated slides with individual hybridisation chambers are used and make use of a spraying technology so as to achieve an excellent surface homogeneity. This technology in turn contributes to very low background noise and superior spot morphology. The precision of the data interpretation increases as a result. The slides themselves support all standard glass substrate slide microarrays, are usable within a wide range of temperatures, thus eliminating the need for cool environments and can be conveniently stacked either in storage eliminating wastage of storage space or during use in a water bath which ensure a uniform reaction temperature among multiple cassettes.

“The most common tests carried out to detect the presence of the disease are sweat tests, and in newborns a screening for immune-reactive trypsinogen is performed”

In all cases where there is suspicion of risk, close consultation with doctors prior to performing a genetic test is important as results must be interpreted in accordance with the latest officially recognised and peer-reviewed guidelines, whether the test is performed pre- or postnatally.

There is no doubt cystic fibrosis is a major debilitating disease and early recognition with treatment is essential if quality of life is to be improved. Genetic screening of patients is now recognised as the moist effective method for identifying the illness and the latest microarray technology is in the front line for accurate diagnosis.