Gene discovery brings cystic fibrosis cure closer

Gene discovery brings cystic fibrosis cure closer

The recent discovery of the cystic fibrosis gene promises far-reaching advances in the detection, treatment and eventual cure of a devastating genetic disease.

Cystic fibrosis is the commonest inherited, potentially fatal disease among Caucasians, especially those of North European descent. In Canada, about one in 2,500 children is born with cystic fibrosis (CF) and about one in every 25 persons carries the flawed gene, often without knowing it and without any signs of the illness. If both parents are CF carriers, each child they bear has a one in four chance of being born with CF, i.e., getting a “double dose” (two copies) of the faulty gene which produces the disease.

Not in itself a killer, the genetic defect affects primarily the lungs, sweat glands and digestive system. Until about 10 years ago, all that was known of CF was its classic inheritance pattern as a single-gene, autosomal recessive disorder and the fact that it causes chronic lung and digestive problems linked to abnormal salt transport. Now, by a stunning piece of collaborative research, scientists at the Universities of Toronto and Michigan have located and cloned the gene responsible, worked out its chemical code in DNA (the genetic molecule inside chromosomes) and defined many of the mutations – variants from the normal – that produce CE (A gene is a segment of DNA inside a chromosome in a living cell. The gene has a precise sequence of chemical units or bases that provides the information for manufacturing a product, usually a protein.)

While the exact mechanism behind cystic fibrosis remains unknown, the good news is that cloning the gene has enabled scientists to identify and analyze the structure of the gene-controlled protein involved, locate its probable site in affected cells and theorize about the way it works – knowledge that will ultimately lead to more effective treatment, perhaps even a cure.

What exactly is cystic fibrosis?

Cystic fibrosis strikes with varying degrees of severity. It involves an inability to move salt or sodium chloride (particularly the chloride part) and water through the channels in cells that line certain organs. The flawed chloride transport and consequent dryness of cell surfaces result in abnormally thick secretions that hamper the normal function of organs such as the lungs, sweat glands, intestine and pancreas. The lung defect usually poses the greatest danger because thick, sticky mucus plugs the airways and leads to frequent respiratory infections. Many CF sufferers find life a constant battle against bacterial infections. Digestion may also be impeded because thick secretions obstruct the ducts, slow the flow and deplete the supply of pancreatic enzymes. The lack of digestive enzymes hinders food absorption, leading to malnutrition and fatty stools. People with pancreatic insufficiency must take many pills daily to prevent under-nourishment.

The first comprehensive report of the disease in 1936 described it as “cystic fibrosis of the pancreas” because the thick mucus that blocks the flow of secretions causes scaning or fibrosis of the pancreas. However the pancreatic problem isn’t usually the most dangerous aspect of CF, nor is the pancreas always seriously affected. (About 15 per cent of patients are classed as pancreas sufficient.) Nonetheless, the term cystic fibrosis has largely stuck; in a few countries it’s called mucoviscidosis. In the 1960s it became clear that the basic defect is faulty salt transport and that the sweat of CF sufferers is unusually salty. The sweat test, which detects the elevated salt content, has become the standard diagnostic tool for CF.

During the 1980s, scientists recognized chloride transport as the primary flaw. And in 1985, thanks largely to the work of researchers at Toronto’s Hospital For Sick Children, the gene responsible for CF was located on human chromosome number seven. In 1989, the molecular code of the gene (the sequence of chemical units in DNA) was accurately described. From that, scientists deduced the structure of the protein controlled by the gene – now named CFTR (TR stands for transmembrane conductance regulator). Recent work suggests that the CFRR protein resides in the so-called apical membrane of affected cells and probably plays some crucial part in regulating chloride movement through cell surfaces.

CF treatment has gradually improved, prolonging survival so that some now live well into their 40s. Many who reach adulthood lead near-normal lives – depending on the degree of organ damage. Whereas in 1960 only about four per cent of children born with CF lived beyond their teens, almost 80 per cent now reach adulthood, the current age of survival averaging 26 years. However, most sufferers need lifelong medication to combat lung infections and assist digestion. The 1990s will probably see clarification of the CF defect – the precise functional flaw (how salt regulation goes wrong) – development of more effective treatment and the establishment of comprehensive carrier screening tests.

The inheritance pattern

Cystic fibrosis is inherited as an autosomal recessive disease, meaning that the faulty gene resides not on the X or Y sex chromosome but on one of the other 22 autosomal human chromosomes. Recessive means that a child must receive two copies of the gene for CF instead of its normal counterpart one from each parent – to produce disease. Two healthy carrier parents, each with one faulty and one normal gene, neither having any signs of CF, can each transmit the flawed (mutant) gene to their baby. If both mother and father’ are carriers there’s a one in four chance that any child conceived will have a disease-producing double dose of the defective gene. But that’s a matter of chance – a toss of two coins – and the risk of getting “heads” on both, two copies of the faulty gene. A child who gets only one faulty gene will be a carrier but won’t have the disease. The risk of two carrier parents having a second or subsequent affected child remains one in four. Successive pregnancies don’t lessen the risk.

Carriers are usually completely unaware that they carry the flawed gene as they have no symptoms of CF. Hence, the parents of an affected child may be bewildered and unable to understand why their baby has CF when they themselves have no signs of it. But CF is not due to diet, stress or any problems during pregnancy, nor can it be passed from one child to another: it’s entirely due to genes inherited from both parents.

How is cystic fibrosis usually detected or diagnosed?

Cystic fibrosis may show itself soon after birth – in an infant with bowel obstruction or a sickly child with abnormally fatty stools who fails to grow normally. Although present at birth, the disease is sometimes not detected until puberty or early adult life because the symptoms – such as breathing difficulties may be blamed on such diverse causes as asthma or bronchitis. If no sweat test is done, the symptoms may easily be attributed to something else. About 10 per cent of CF cases are discovered because of bowel obstruction (meconium ileus) right after birth or even before (by ultrasound), about 50 per cent because of lung infections, 30 per cent due to pancreatic insufficiency and failure to thrive.

Managing a devastating disease

Since there’s no cure at present, therapy must usually be continued throughout life – a demanding regime for CF sufferers, parents and doctors alike. Besides medical care, patients generally require psychological, occupational, social and genetic counselling.

* Managing the lung congestion. Treatment includes exercises to drain fluid, methods to assist in clearing mucus and antibiotics to combat recurrent bacterial infections.

* Dealing with pancreatic insufficiency and possible nutrient deficiencies. While not all CF sufferers have pancreatic insufficiency, most have some degree of abnormality in the production and supply of pancreatic enzymes and many need pills to make up the deficit.

* Improvement can be dramatic. Children with CF may improve so markedly, with excellent school records and participation in sports, that parents wonder whether their youngster really has CF. A child’s response depends largely on the severity of the illness when first detected.

How the gene was found

After a decade of intensive work, University of Toronto researchers working at the Hospital For Sick Children and their collaborators at Ann Arbor, Michigan, were among those who received the prestigious Gairdner Award in October 1990 for their outstanding contributions to medical science.

In contrast to the search for many other disease genes, the quest for the defective CF gene began without any physical clues no chromosome breaks or rearrangements – to help it along. Scientists only knew that the faulty gene resided somewhere in the 22 pairs of autosomal chromosomes (not on the X or Y sex chromosome) in a human cell. With no visible chromosome changes or “neon signs” to point the way, the scientists used positional cloning to find the gene. They had to narrow down the field from an immense area – amounting to about 100,000 genes or three billion bases along the DNA molecules – of which a tiny segment is the gene for CF. Trying to find one of these genes, knowing only its approximate location, is like looking for a needle in a haystack or – to use the analogy of one researcher – comparable to searching for a single burnt-out light bulb in a house somewhere in Canada ! The researchers traced the gene by using markers on the human chromosomes – close neighbours known to be co-inherited and travel along with CF from generation to generation – tracking them in the blood and cells of CF families. The search was advanced by the excellent records of CF families in Canada. After four painstaking years, the researchers realized that they were on the right track in 1985 when they’d narrowed down the probable gene site to the long ann of chromosome number seven. That’s like tracing the burnt-out light bulb to the right province. Eventually they isolated the gene’s probable position between two markers on chromosome seven – getting the burnt-out bulb into the right city.

Ultimately they homed in on the gene by “walking” along the chromosome, analyzing its DNA bit by bit. Pooling efforts with another research team, they also employed the innovative technique of chromosome jumping,” invented by a Michigan collaborator, to speed things up. Two years of chromosome “walking” and jumping” finally brought them really close – akin to finding the house with the burnt-out light bulb! But even when right on top of the gene, the researchers could only know they were “there” by finding a specific difference (a mutation or change from the normal DNA sequence) that was exclusive to chromosomes from CF families and never ever found in chromosomes from people who are neither CF patients nor carriers.

Finally, the researchers found a segment of DNA from a CF sufferer that was distinctly different from its normal counterpart. The searchers knew they’d found the gene for CF (the right segment of DNA) by identifying a difference in the DNA sequence of chromosomes from CF sufferers and those from the cells of people unaffected by CE The change or mistake was a deletion of three bases (triplet deletion) in DNA, unique to CF families. The gene discovery, reported in the September 1989 issue of Science, Was hailed as a great achievement.

Cure prospects slowly coming nearer

Until the precise biochemical flaw in CF is known and corrected, treatment can only slow down, not halt the disease. Since the 1989 discovery, researchers have made great strides towards defining the three dimensional, folded structure of CFTR and have made antibodies to it. They are also comparing the human CFfR molecule to similar protein transporters in other animals – for instance in sharks, yeast and bacteria – to find out exactly how things go wrong in CF. Discovery of the CF-causing gene and its protein-product opens up the chance of developing “designer” drugs that could be deliberately designed to stop it from hindering salt movement. In addition, gene therapy may some day provide a cure for this disease. Inserting a normal gene into CF cells to replace the defective one could overcome cystic fibrosis. The first step has already been achieved – transferring a normal version of the gene into defective pancreatic and airway cells in a culture dish, thereby regularizing chloride/water transport so that the treated cells behave normally. And just recently the first successful experiment in gene replacement in a living patient (with the immune system disorder, adenosine deaminase deficiency) was reported. While many hurdles must still be tackled, this success may pave the way to replacement of the defective gene in the affected tissues of CF patients.

Genetic screening and prenatal diagnosis now possible

The discovery and cloning of the gene sets the stage for the development of CF carrier tests. There is however a vigorous debate as to whether or not widespread, universal screening for CF carriers should be performed on the population before all the existing mutations are known. Since all the 60 CF mutations reported to date only add up to 85 per cent of the possible gene errors, mass screening done now would miss some CF carriers. While the benefit of identifying carriers (one in 25 people) is clear, most experts argue against wide scale screening until at least 95 per cent of the possible CF mutations are known. (Some biotechnology companies have already developed CF carrier test kits and are prepared to start commercial testing, even though such tests would miss some carriers.)

In known CF families, where the particular mutation has been characterized, carrier screening is already widely done. Before the gene discovery, parents only became alerted to the possibility of having a child with CF if they’d already had one or more affected children. Now, carriers can be detected with reasonable accuracy in CF families. And genetic tests on unborn babies (examining cells removed from the chorionic villi or amniotic fluid around the fetus) permit prenatal diagnosis as early as two months after conception. The DNA from fetal cells can be examined to see if the fetus carries any known mutation. If the flawed gene sequence is found, the fetus has CF and the parents can decide whether or not to terminate the pregnancy.

Many ethical dilemmas to solve

Even when carrier screening tests can identify 95 per cent or more of the possible CF mutations and become generally available, they will still raise moral, social, financial and ethical dilemmas. For instance, how and by whom should the tests be done? When should they be administered? Should they be given to teenagers – risking possible hardship or stigma if a youngster turns out to be a CF carrier? Or, should the carrier test be offered to couples contemplating marriage, or at a woman’s first obstetric visit? If a genetic test shows that a fetus has CF, is that sufficient grounds for terminating the pregnancy? Such difficult questions have no simple answers. But pilot studies done in Britain and Montreal wrong students and young people brought an overwhelming “yes” in favour of CF screening, even though the test isn’t yet 100 per cent foolproof and would miss some carriers. Initiating such tests would entail an immense public education effort in helping people to make informed decisions. But as therapy improves and cure prospects come nearer, CF would no longer be a lethal disease, making some experts question the wisdom of mounting a costly mass screening effort.

* Clinical features: possible alerting signs

* Intestinal (bowel) blockage in infancy, which causes abdominal swelling, vomiting and dehydration, often requiring surgery;

* unusually salty taste- of baby’s skin when kissed;

* inability of the pancreas to deliver necessary digestive enzymes and failure to gain weight despite a normal or unusually large appetite; fatty stools; frequent respiratory infections;

* finger clubbing (abnormal finger-nail growth) – also seen in other chronic illnesses;

* sinusitis; nasal polyps;

* abnormal glucose test and diabetes mellitus because of damage to the pancreas and insulin insufficiency;

* delayed puberty, reproductive abnormalities especially male sterility (over 90 per cent of males with CF are infertile).

COPYRIGHT 1990 Strategic Inc. Communications Ltd.

COPYRIGHT 2008 Gale, Cengage Learning