Gabriele Richard, MD, FACMG, Medical Director and Vice President, GeneDX, Gaithersburg, MD

Dr. Gabriel Richard

Ichthyosis and related disorders represent a group of more than 28 distinct disorders with different modes of inheritance and different causes.  All these disorders primarily affect the outermost layer of the skin, the epidermis, and can be summarized as disorders of cornification. The epidermis provides our barrier to the environment. It is a highly specialized epithelium designed to protect the human body from water loss and physical, chemical, and mechanical insults. In order to establish and constantly maintain this barrier, the epidermal cells, known as keratinocytes, undergo a complex, highly organized, and tightly controlled process of changes leading to cornification. During this process, cells migrate to the surface, where they form the horny or co  rnified layer and ultimately are cast off. The human body sheds approximately 2 billion cells during the course of a day. Under normal conditions, cell proliferation and desquamation are in equilibrium. Any external or internal condition that disturbs this balance is bound to impair the barrier function of the skin, eventually resulting in disease. As indicated by the name ‘ichthyosis,’ that stems from a Greek root meaning fish, these disorders often are characterized by scaling of the skin. Visible scales are produced by shedding of clumps of 100 to 500 or more cells that stick together. Scaling is usually associated with thickening of the cornified layer, called ‘hyperkeratosis,’ due to an increased number of cells produced by the epidermis, a delay in cell shedding, or as in most circumstances, a combination of both mechanisms.

In the last decade, enormous progress has been made in deciphering the molecular causes of inherited skin disorders. Genetic research uncovered over 120 different genes,which, when altered, disturb the normal structure and/or function of the skin and result in disease. Among them are 18 genes causing generalized ichthyosis, 19 genes implicated in palmoplantar keratoderma (the thickening of palms and soles), 4 genes linked to erythrokeratoderma (usually characterized by localized redness and thickening of skin), and 9 genes associated with other disorders such as Darier disease and Hailey-Hailey disease. This article highlights some of the most recent advances in genetic research of autosomal recessive ichthyoses, which already has considerably enhanced our understanding of the disease mechanisms and laid the foundation for molecular diagnoses and future development of new treatment modalities.

Autosomal recessive ichthyoses occur in approximately 1 in 100,000 to 1 in 300,000 individuals. Both genders are affected equally. Most autosomal recessive ichthyoses have been observed worldwide without ethnic clustering, although they are more common in populations with a high degree of blood relations (consanguinity). One of the best-known types is lamellar ichthyosis (LI), which manifests at birth. Babies are usually born encased in a tight, shiny covering of hardened skin called collodion membrane. After shedding, large, dark- brown scales develop over the entire body. The facial skin is tight and may pull eyelids and lips outward (ectropion and eclabion), which can result in secondary damage to the outer parts of the eyes. Other problems include reduced or absent sweating and heat intolerance as well as skin infections and scarring hair loss. The major gene responsible for LI, TGM1, is located on the long arm of chromosome 14 and determines the production of an enzyme called transglutaminase-1. As many as 50% of LI patients carry a small change in the DNA code (mutations) in each of the 2 gene copies of TGM1. These mutations lead to reduced or absent enzyme production or diminished enzyme activity. To date, over 50 different mutations have been identified. One mutation (designated 2526A->G) appears to be common among individuals of Northern European descent and likely originates from a German ancestor (founder effect). It leads to the making of incorrect, nonfunctional transglutaminase-1. The enzyme is essential for forming the horny layers of the skin. Transglutaminase-1 normally functions as super glue that cross-links numerous structural proteins in the epidermis facilitating the collapse of cells to cornified remnants. However, it is also necessary for attaching epidermis-specific fats (lipids) to this protein envelope. The diminished or absent transglutaminase-1 activity in patients with LI seriously disturbs the complex process of cornification and results in scaling. Comparisons of patients with and without detectable TGM1 mutations revealed that TGM1 defects are most commonly associated with ‘classical’ LI, although some patients with mild to moderate redness or white, gray and smaller scales carry TGM1 defects. The etiology of LI in patients without TGM1 mutations remains elusive. Two other genes have been mapped to chromosomes 2q33-q35 and 19p12-q12, respectively, but their identity and functions of the corresponding proteins are still unknown.

The clinical features of LI widely overlap with non-bullous Congenital Ichthyosiform Erythroderma (CIE), another autosomal recessive ichthyosis. Most affected individuals are born with a collodion membrane, which subsequently evolves into generalized scaling and redness of the skin. In contrast to ‘classic’ LI, scales are usually white, fine and powdery, although they may become larger and darker on the lower legs. Extensive thickening of the skin of the palms and soles is also a common feature of CIE. In the vast majority of families, CIE is inherited as an autosomal recessive trait, although autosomal dominant inheritance has been occasionally observed. Similar to LI, CIE can have several different causes. A small group of patients, especially those with less intense redness of the skin, carry inactivating mutations in TGM1 as discussed above. Other patients originating from the Mediterranean basin were found to harbor mutations in one of two functionally related genes, ALOXE3 and ALOX12B, both located on the short arm of chromosome 17. These genes produce 2 enzymes belonging to the lipoxygenase family that are involved in the metabolism of skin fats (lipids), such as polyunsaturated fatty acids, phospholipids and triglycerides. The reported mutations result in loss or inactivation of the enzymes. Although the specific functions of each enzyme are still unknown, they are assumed to operate jointly in a common metabolic pathway crucial for maintaining the lipid barrier of the epidermis. In the future it will be necessary to determine if ALOXE3 and ALOX12B mutations play a role in CIE patients of other geographic and ethnic backgrounds or if other genes might be involved. Until then, the underlying genetic basis of CIE in most patients awaits identification.

Sjögren-Larsson syndrome (SLS) is an autosomal recessive disorder characterized by congenital ichthyosis, spasticity, and mental retardation. At birth, the skin may show varying degrees of redness and scaling with accentuated skin markings. While the redness subsides over time, thickening and scaling of the skin tend to worsen, especially on the abdomen, in large skin folds and on palms and soles. In contrast to many other ichthyoses, the skin is very itchy. During early childhood neurological problems, such as stiffness of arms and/or legs and difficulties with walking, develop. Another characteristic sign are glistening white dots in the back of the eyes. The involvement of the central nervous system slowly progresses and results in developmental delay, speech defects, severe mental retardation and a host of other problems. SLS is caused by deleterious mutations in the gene FALDH on the short arm of chromosome 17. The gene produces another enzyme involved in lipid metabolism, Fatty Aldehyde Dehydrogenase. This enzyme is part of a pathway that produces fatty acids, which are important for the making of epidermal lipids as well as the degradation of certain phospholipids in the brain. The majority of the more than 50 different mutations detected in the SLS gene are unique to each family. Nevertheless, research revealed a few mutations that are common among individuals of Northern European and Middle Eastern origin. This knowledge can be very helpful for a fast molecular diagnosis based on DNA tests. Other specific and reliable diagnostic tests are the measurement of enzyme activity and the detection of elevated free fatty alcohols in cultured skin cells (fibroblasts) or the blood.

A final example is Comèl-Netherton syndrome (NTS). This autosomal recessive ichthyosis is associated with hair abnormalities (bamboo hair, alopecia) as well as abnormalities of the immune system, including elevated levels of immunoglobulin E in the blood, susceptibility to skin and respiratory tract infections and allergic reactions. A recent study revealed that almost 1 in 5 of all babies born with generalized red and scaly skin have NTS. In the newborn period, affected children may suffer from fluid and electrolyte imbalances, failure to gain weight and thrive, and life-threatening infections. Later, the skin disorder in NTS either remains generalized closely resembling CIE,or change into scaling, itchy plaques with prominent borders. The latter variant is named ‘ichthyosis linearis circumflexa’. Despite these clinical differences, both variants are caused by recessive mutations in the gene SPINK5 on the long arm of chromosome 5. This relatively large gene contains the genetic blue print for producing an inhibitor of protein-degrading enzymes named LEKTI (Lympho-Epithelial Kazal-Type Inhibitor). Over 50 mutations of different types and location have been detected so far, 65 percent of which result in the complete loss of LEKTI enzyme due to disrupted transcription from DNA into protein, while the remainder are thought to compromise enzyme function. A few mutations seem to be more common among Turkish or Arab populations, but similar to LI, most families carry unique mutations. The LEKTI enzyme is expressed in the skin, mucous membranes, tonsils and thymus, and when needed, can be quickly cleaved into 15 active domains that suppress enzymes like trypsin. The specific biological targets of LEKTI in human tissues, however, are currently unknown. In principal, loss of LEKTI will lead to uncontrolled and prolonged activity of destructive, protein-degrading enzymes. In the skin, this process disturbs the delicate balance between lipid-processing enzymes and hastens disintegration and shedding of the horny cells, thus severely disrupting the barrier function. In addition, the skin and mucous membranes lose protection against invading microorganisms and inflammation, which further contributes to the disease.

As illustrated with these five examples, molecular genetic research of the ichthyoses has been incredibly successful in unveiling the underlying defects of numerous single gene disorders, although the list of disease genes is far from complete. Often the question is asked: ‘What can modern genetics do for us?’ The previous sections aimed to demon strate how gene discovery and molecular biological research have advanced our insight into the different mechanisms leading to disturbed cornification and skin barrier function. It is now possible to categorize ichthyoses not only based on their clinical symptoms but also based on the underlying genetic defects. This new information also provides an invaluable basis, from which the biological consequences of mutations and the pathways of disease can be explored in detail, such as the lipid metabolism of the skin.

Knowledge of the genes and mechanisms of disease will be crucial for conquering the next great challenge of developing new therapeutic approaches specifically tailored to these disorders. Animal models of human ichthyoses are being developed, such as mice deficient for transglutaminase-1, which exhibit a skin disorder similar to LI with impaired barrier function leading to water loss and abnormal absorption through the skin. Such models allow the systematic evaluation of the functions of specific epidermal proteins as well as testing of new therapeutic modalities. In parallel, several approaches are being explored to replace the lost enzyme activity of transglutaminase-1 or LEKTI in the epidermis or to deliver normal gene copies directly to the skin. While much more work, time and money will be needed to develop practicable and effective new treatments, molecular genetics has paved the way for these developments.

One of the immediate benefits of genetic research, however, is the development of molecular tests to confirm a clinical diagnosis, allow prenatal or presymptomatic testing, determine the risk of disease for members of families and improve genetic counseling. Establishing or ruling out a clinical diagnosis can be especially helpful in those types of ichthyoses, in which scaling of the skin occurs before the onset of other organ manifestations, as in neutral lipid storage disease or SLS. In other disorders, such as Netherton syndrome, severe health problems may develop soon after birth and during infancy, while diagnostic clinical features or laboratory results will not be available until later. Thus molecular testing can help to recognize the correct diagnosis and to make predictions about the course and progression of disease. Nevertheless, since since the treatment of most ichthyoses is still limited to easing symptoms, the outcome of molecular testing often has little immediate impact on the treatment of patients treatment of patients with an inherited ichthyosis.

Knowing exactly which DNA mutations cause the disorder in a patient or family also offers the possibility of testing a baby before it is born (prenatal testing) or determining the carrier status of family members. Reliable and early prenatal molecular testing has been well established for some disorders with severe and sometimes fatal outcomes, such as NTS and SLS, and can be performed using chorionic villus sampling as well as amniocentesis. In LI and CIE, DNA-based testing can replace difficult and invasive fetal skin biopsies, but currently remains limited to detecting disease-causing mutations in TGM1.

Biological material for molecular testing can be relatively easy to obtain. Most tests are performed from DNA, which is extracted from a small venous blood sample or buccal swabs. For screening of large genes, such as TGM1, or if DNA mutations alter the size of the DNA transcript, it might be necessary to extract messenger RNA from a small skin biopsy. In addition, certain biochemical assays (for example in SLS) or the detection of a protein deficiency in the skin (for example, loss of transglutaminase-1 in LI) requires a skin biopsy.

Most ichthyoses are caused by distinct DNA mutations that are specific to the affected individual or family. Finding these changes is often costly and labor intensive. Therefore, these specialized tests are only performed in very few research and commercial laboratories worldwide. Multiple different methods and strategies have been developed to identify DNA mutations, each of which has its advantages and limitations. Direct sequencing of the DNA sequence of a gene is the most sensitive but often also the most costly approach. Depending on the nature and location of mutations, it is possible to identify up to 95% of all disease-causing mutations. Faster and less expensive methods, such as heteroduplex analysis, are often used to screen large genes for unknown mutations. The gene is divided into multiple overlapping fragments, which are a million times multiplied by a method called polymerase chain reaction (PCR), and analyzed for differences in the DNA sequence. Once aDNA change is suspected in such a gene fragment, the exact mutation will be determined by DNA sequencing.




Despite the great advances in technology and molecular testing, the widespread application of diagnostic testing is curbed by a number of limitations. For some disorders, tests might not be currently available. Many disorders, including LI and CIE, are caused by mutations in more than one gene and not all of these genes have yet been identified. Before testing, there are no clinical or biochemical signs that would allow predicting if a patient carries mutations in TGM1 or in another gene. Another problem already mentioned is the fact that no assay is sensitive enough to detect all mutations. Depending on the nature of the tests performed, approximately 5% to 20% of mutations might be missed, and this number might be even higher in certain disorders, such as Darier disease and Hailey-Hailey disease. For all single gene disorders there are exceptions to the rules of traditional Mendelian inheritance, which complicate or defy test interpretations. Examples are reduced gene expression leading to differences between the genotype (mutation detected) and the phenotype (no signs of disease), and genetic mosaicism, in which case some cells of the body carry a mutation while others do not. Finally, moral and ethical considerations especially for prenatal testing or testing of individuals before they show symptoms of disease (presymptomatic testing) should also be mentioned here. To evaluate availability, options and limitations of DNA-based testing, seeking genetic counseling by trained professionals in medical genetics is advisable.

In summary, striking progress has been made in our understanding of the ichthyoses on a molecular level. Molecular genetic testing has become available for a number of ichthyoses and may aid and complement the clinical diagnosis, although multiple limitations require careful consideration. Research now faces the next challenge of developing effective treatments for affected individuals.


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