ALOX12B, ALOXE3, and TGM1 Mutations in Scandinavian Pati - Core

ORIGINAL ARTICLE

Genotypic and Clinical Spectrum of Self-Improving Collodion Ichthyosis: ALOX12B, ALOXE3, and TGM1 Mutations in Scandinavian Patients Anders Vahlquist1, Anette Bygum2, Agneta Ga˚nemo1, Marie Virtanen1, Maritta Hellstro¨m-Pigg3, Gitte Strauss4, Flemming Brandrup2 and Judith Fischer5 Infants born with autosomal recessive congenital ichthyosis (ARCI) are often encapsulated in a collodion membrane, which shows a lamellar or erythrodermic type of ichthyosis upon shedding. However, some babies show a nearly normal underlying skin after several weeks, a phenotype called ‘‘self-healing collodion baby’’ (SHCB). Mutations in two genes, TGM1 and ALOX12B, have previously been implicated in the etiology of SHCB, but the full genotypic spectrum remains to be determined. DNA sequencing in 11 Swedish and 4 Danish SHCB patients showed ALOX12B mutations in eight cases, ALOXE3 mutations in three cases, and TGM1 mutations in one case. In three patients, we could not find mutations in any of the known ARCI genes. In all cases, a spontaneous shedding of the collodion membrane occurred 2–4 weeks after birth. When re-examined at 2–37 years of age, the patients showed skin xerosis, a mild or focal scaling, palmar hyperlinearity with keratoderma, and a frequent appearance of red cheeks and anhidrosis. Thus, we propose replacing SHCB with the term ‘‘self-improving collodion ichthyosis’’ (SICI). In conclusion, ALOX12B mutations are the leading cause of SICI in Scandinavia, followed by ALOXE3 mutations, which have not been previously associated with this variant of ARCI. Journal of Investigative Dermatology (2010) 130, 438–443; doi:10.1038/jid.2009.346; published online 5 November 2009

INTRODUCTION Infants born with a glue-like hyperkeratotic membrane covering the entire body are called ‘‘collodion babies.’’ They frequently also show everted eye lids (ectropion) and lips (eclabium). The collodion membrane is associated with an impaired skin barrier function, with increased transepidermal water loss. Potential complications are hypothermia, dehydration, and hypernatremia, usually requiring observation and treatment in an intensive neonatal care unit. The collodion membrane is usually shed within 1–3 weeks, showing an underlying lamellar ichthyosis (LI) or congenital ichthyosiform erythroderma (CIE), the most common phenotypes of autosomal recessive congenital ichthyosis (ARCI). However, in about 10–25% of all cases, 1

Department of Medical Sciences, Uppsala University, Uppsala, Sweden; Department of Dermatology and Allergy, Odense University Hospital, Odense, Denmark; 3Department of Genetics and Pathology, Uppsala University, Uppsala, Sweden; 4Department of Dermatology, Gentofte Hospital, Copenhagen, Denmark and 5CEA, Institut de Geńomique, Centre National de Geńotypage, Evry, France

2

Correspondence: Anders Vahlquist, Department of Dermatology, University Hospital, Uppsala SE-75185, Sweden. E-mail: [email protected] or Judith Fischer, CEA, Institut de Geńomique, Centre National de Geńotypage, Evry, France. E-mail: [email protected] Abbreviations: ARCI, autosomal recessive congenital ichthyosis; CIE, congenital ichthyosiform erythroderma; LI, lamellar ichthyosis; SHCB, self-healing collodion baby; SICI, self-improving collodion ichthyosis Received 16 July 2009; revised 1 September 2009; accepted 17 September 2009; published online 5 November 2009

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Journal of Investigative Dermatology (2010), Volume 130

the membrane is shed without showing residual signs of ichthyosis—a condition known as ‘‘self-healing collodion baby’’ (SHCB) (Dean, 1921; Frenk and de Techtermann, 1992; Van Gysel et al., 2002). For reasons given in this paper, we propose to complement SHCB by the more adapted term ‘‘self-improving collodion ichthyosis’’ (SICI), also suitable for older children and adult patients who show some remaining skin symptoms. Over the last decade at least six different genes have been implicated in the etiology of ARCI, all of which encode proteins essential for the formation of the skin barrier (Huber et al., 1995; Russell et al., 1995; Jobard et al., 2002; Lefevre et al., 2003, 2004, 2006; Fischer, 2009). The most common cause of LI/CIE is inactivating mutations in the TGM1 gene, which encodes transglutaminase-1, an enzyme that crosslinks proteins in the cornified cell envelope (Russell et al., 1995; Farasat et al., 2009). Two unusual point mutations in TGM1, which reduced the enzyme activity in utero but not in vivo, were the first causes of SHCB/SICI to be reported (Raghunath et al., 2003). Additional ARCI-associated genes encode enzymes or transport proteins in the lipoxygenase pathway, which produces metabolites required for a proper deposition of lipids between the horny cells (Jobard et al., 2002; Brash et al., 2007; Epp et al., 2007). For example, ALOX12B and ALOXE3 gene mutations have been associated with a mild type of ichthyosis, which does not always fit into the traditional LI/CIE classification (Jobard et al., 2002). This & 2010 The Society for Investigative Dermatology

A Vahlquist et al. Self-Improving Collodion Ichthyosis

phenotype is sometimes described as congenital ichthyosis with fine or focal scaling (Ganemo et al., 2003; Eckl et al., 2009). Recently, Harting et al. (2008) described two American cases of SHCB/SICI with previously unreported mutations in the ALOX12B gene; the collodion phenotype in these cases later changed into a mild ichthyosis. In this study, we successfully determined the genotype–phenotype relationship in 12 of 15 SICI patients identified by nationwide screenings for ARCI in Sweden and Denmark, including over 250 ichthyosis patients. Besides several types of ALOX12B and TGM1 mutations, we report for the first time ALOXE3 mutations as a cause of SICI, and further elaborate the spectrum of persisting skin symptoms in this group of patients. RESULTS Of the 15 SICI patients (9 females and 6 males), 12 showed homozygous or compound heterozygous mutations in at least one of the ARCI genes investigated (Table 1). Eight of the patients had ALOX12B mutations, three had ALOXE3 mutations, and one patient had TGM1 mutations. There was no consanguinity reported. Prematurity (3–5 weeks) was noted in four cases. At birth, all babies were covered by a variably thick collodion membrane and ectropion was present in eight cases (Table 1). The membrane was spontaneously shed after 2–4 weeks, exposing an almost normal-looking skin, which however later developed signs of mild ichthyosis (Figure 1a–d). When re-examined at 2–37 years of age, all patients showed varying degrees of dry skin with fine or focal scaling (Table 1). Typical features were scaling in the armpits (Figure 1e), acral hyperkeratosis and keratoderma (Figure 1f), coarse scales on the scalp (Figure 1g), and a fine scaling around the neck (Figure 1h), together with xerotic extremities and red cheeks that were stinging when exposed to sea water or certain emollients (Figure 1i). All patients, except the one with TGM1 mutations, had moderate-to-severe anhidrosis, which was manifested as fatigue and intense skin erythema during exercise and in a hot environment. Some patients occasionally noted sweating on a small patch in the face. Besides these symptoms, the patients were healthy except for minor psychomotor problems and a sunlight sensitive skin in two cases each. Five patients carried the same ALOX12B mutation, which replaces an A to a G in position 1,562, leading to a replacement of tyrosine with cysteine at aa position 521 (p.Tyr521Cys). Four of them (patients P3, P4, P6, and P7) were homozygous for this mutation, and one patient (P2) was a compound heterozygote for this mutation, and a second missense mutation (c.199A4T; p.Ile67Phe). Another patient (P1) was homozygous for a missense mutation in exon 11 (c.1385G4A; p.Gly462Asp), and one further patient (P5) was a compound heterozygote for p.[Val527Met] þ [Ala597Glu] in exon 12 and 14 respectively. Finally, one Danish patient (P8) was homozygous for a known splice site mutation at position 1654 þ 3A4G of exon 12 (Table 1). Three SICI patients presented mutations in ALOXE3; one Swedish patient (P10) was homozygous for a T-to-C mutation at position 1,280 in exon 9 (p.Leu427Pro). Two other patients

(P9 and P11) were compound heterozygotes; they shared the same missense mutation at nucleotide position 1,889 (p.Pro630Leu), and they had a different second mutation (p.Cys109X and p.Arg234X). The last patient (P12) showed compound heterozygous mutations in exon 6 of TGM1: one splice site mutation c.877-2A4G and one already known missense mutation c.919C4G, which corresponds to p.Arg307Gly on the protein level. These mutations probably result in a similar effect on the transglutaminase enzyme activity as previously reported in a German SICI patient (Raghunath et al., 2003). Her persistent skin symptoms were mild (see Figure 1h) and she had a normal sweating ability. DISCUSSION To the best of our knowledge, this is the largest study of SICI etiology reported to date. It confirms that ALOX12B gene mutations are a common cause of this mostly transitory form of ARCI (Harting et al., 2008), accounting for 67% of 12 genotyped cases in Scandinavia. Many of these patients were either homozygous for the p.Tyr521Cys mutation or carried at least one such mutation. Speculatively, this type of missense mutation in ALOX12B could result in a protein misfolding that negatively affects the enzyme activity only under in utero conditions, thus explaining a dramatic improvement of epidermal cornification after birth. Three out of the six ALOX12B mutations have been described previously: p.Tyr521Cys (Eckl et al., 2005; Eckl et al., 2009), pVal527Met (Lesueur et al., 2007), and c.[1654 þ 3A4G] (Eckl et al., 2009). Interestingly, none of these former patients have been described with a SHCB phenotype, suggesting that other factors may have a role in the developing of this phenotype. However, all seven of the previously described patients show a mild phenotype including fine scaling and mild erythema. The same phenomenon has also been observed in patients with specific TGM1 mutations, which leads to SICI in the majority of cases, but in some cases to the more common LI. The second most common cause of SICI in our study was ALOXE3 mutations. This gene has been previously associated with ARCI (Jobard et al., 2002), but has never been reported as a cause of SHCB/SICI. Two of the three mutations have been previously described in adult patients without the SICI phenotype: p.Arg234X (Jobard et al., 2002) and p.Pro630Leu (Eckl et al., 2005). As ALOXE3 and ALOX12B encode enzymes operating in the same metabolic pathway (Jobard et al., 2002; Brash et al., 2007; Epp et al., 2007) and mutations in both genes are relatively common as causes of ARCI (Eckl et al., 2009; Fischer, 2009), our finding of ALOXE3 mutations in 25% of Scandinavian SICI patients may not be so surprising. Only one of our SICI patients had compound heterozygous TGM1 mutations, confirming a similar rare finding in a German patient (Raghunath et al., 2003). The first mutation p.Arg307Gly has been described in patients with SICI, whereas mutations of the same arginine codon 307 replaced by a tryptophan codon (p.Arg307Trp) are known to be associated with traditional LI (Farasat et al., 2009). In addition, the second mutation (c.877-2A4C), which is a splice site www.jidonline.org

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Table 1. Clinical and genotypic features of 12 patients who were born as collodion babies but later showed only minimal signs of ichthyosis Patient no., origin Age1 Sex

Ectropion Current type at birth of ichthyosis

Palmoplantar Special clinical keratoderma Anhidrosis features

Gene, mutation

Effect

Exon

ALOX12B P1 SV

12

M

?

Fine scaling with Moderate focal accentuation

+

Light sensitivity, minor deafness, and slight mental retardation

c.[1385G4A] +[=]

p.[Gly462Asp] +[=]

11

P2 SV

14

F

+

Fine scaling, acral keratosis

Mild

+

ADHD

c.[199A4T] +[1562A4G]

p.[Ile67Phe] +[Tyr521Cys]

2 12

P3 SV

14

M

+

Xerosis

Mild

+

Red cheeks, stinging in contact with sea water

c.[1562A4G] +[=]

p.[Tyr521Cys] +[=]

12

P4 SV

9

M

+

Fine scaling, slight erythema

Mild

+

Red cheeks, prematurity2, c.[1562A4G] and mild autism +[=]

p.[Tyr521Cys] +[=]

12

P5 SV

4

F

+

Scaling around armpits

Moderate

+

Stiff fingers, scalp c.[1579G4A] hyperkeratosis, red cheeks, +[1790C4A] and prematurity2

p.[Val527Met] + [Ala597Glu]

12 14

P6 SV

2

F

+

Scaling around armpits, xerosis

Mild

+

Red cheeks, stinging in contact with sea water, and pruritus

c.[1562A4G] +[=]

p.[Tyr521Cys] +[=]

12

P7 DK

12

F

+

Fine scaling, scalp keratosis

Palmar hyperlinearity

+

Acral lichenification, red cheeks, and prematurity2

c.[1562A4G] +[=]

p.[Tyr521Cys] +[=]

12

P8 DK

37

F

?



+

Light sensitivity

c.[1654+3A4G] +[=]

splice site +[=]

12

ALOXE3 P9 DK

6

F

+



Heat Allergic rhinitis and intolerance prematurity2

c.[327C4A] +[1889C4T]

p.[Cys109X] +[Pro630Leu]

2 14

P10 SV

7

F

Fine scaling

—

+

Pruritus

c.[1280T4C] +[=]

p.[Leu427Pro] +[=]

9

P11 SV

22

M

Fine scaling

—

+

Syndactyly of two fingers

c.[700C4T] +[1889C4T]

p.[Arg234X] +[Pro630Leu]

6 14

P12 SV

18

F

+

Fine scaling, xerosis

Fissures

Much better in summer

c.[877-2A4G] +[919C4G]

Splice site p.[Arg307Gly]

6 6

TGM1

ADHD, attention-deficit hyperactivity disorder; DK, Danish origin; F, female; M, male; SV, Swedish origin. Age in years at last examination. 2 Prematurity of 3-5 weeks. The symbol ‘‘+[=]’’ indicates homozygous mutations, according to the Human Genome Variation Society nomenclature. 1

mutation, has already been described (Farasat et al., 2009). However, TGM1 mutations of other types are found in nearly 50% of Scandinavian collodion babies who eventually 440


develop LI or CIE (Ganemo et al., 2003; Pigg et al., unpublished). Clearly, TGM1 mutations underlying SICI are extremely rare events compared with ALOXE3 and


Figure 1. Examples of phenotypic variants of self-improving collodion ichthyosis. Patient P5 with ALOX12B mutations (a) at birth showing collodion membrane and ectropion, (b) at the age of 2 months showing normal-appearing skin, and (c and d) at the age of 6 years showing acral hyperkeratosis and xerosis. Other examples of skin symptoms in patient P6 with ALOX12B mutations at the age of 2–18 months (e and f), in patient P10 (at age 7 months) with ALOXE3 mutations (g), in the neck of patient P12 (at age 18) with TGM1 mutations (h), and in patient P2 (at age 14) with ALOX12B mutations (i). For further details, see Table 1. Patients’ parents gave permission for the use of identifiable photographs.

ALOX12B mutations, which seem to be the leading cause of SICI. Five of the mutations identified in this study have not yet been described in the literature; they include one nonsense mutation (c.Cys109X) and three missense mutations (p.Ile67Phe, p.Gly462Asp, and p.Ala597Glu) in ALOX12B, and one missense mutation in ALOXE3 (p.Leu427Pro). None

of the mutations in our 12 patients were found in 280 chromosomes from a Caucasian control population. ICHTHYIN mutations are another relatively common cause of ARCI in Scandinavia (Dahlqvist et al., 2007), but this genotype is seldom associated with a collodion membrane at birth (A. Vahlquist et al., unpublished observation); none of our SICI patients had this genotype. www.jidonline.org

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However, one of the patients in this study (P9), who was a compound heterozygote for ALOXE3 mutations, also carried one heterozygous ICHTHYIN mutation. Whether this mutation acts as a disease modifier in SICI is presently unknown. Of the 15 SICI patients, 3 did not show any mutations in the investigated ARCI genes (ALOXE3, ALOX12B, ABCA12, CGI-58, CYP4F22, ICHTHYIN, and TGM1). We therefore conclude that mutations in other genes that remain to be identified may also lead to a SICI phenotype. Our study suggests that the prevalence of SICI in Sweden and Denmark is around 1 per million (15 cases in a population of 14 million). However, this is probably a low estimate, because many SICI patients will never be correctly diagnosed because of the transient nature of the skin problems. For example, an elderly ARCI patient with no detailed knowledge about the events surrounding his/her birth is unlikely ever to be diagnosed as having SICI. Even today, when a SICI baby is born in a smaller hospital and there is no dermatological expertise available, a thin collodion membrane may be misinterpreted as post-term skin dryness or debris of vernix caseosa remaining on the skin. In addition, male infants with X-linked recessive ichthyosis sometimes show a thin collodion-like membrane at birth that may confuse diagnosis (Hoyer et al., 1986; Hernandez-Martin et al., 1999; A. Vahlquist, unpublished observation). In fact, the term ‘‘SHCB’’ may itself be confounding. Although when seeing a collodion baby in the neonatal period the term is quite useful to distinguish one ARCI subtype with good prognosis from another subtype with worse prognosis, the word ‘‘self-healing’’ spuriously implies that the skin of the patient will always look normal after the collodion has been shed. On the contrary, when our SICI patients were re-examined at 2–37 years of age, all showed mild signs of ichthyosis, such as xerosis and a fine scaling especially in the axillae and around the neck, as well as red cheeks and a varying degree of anhidrosis associated with heat intolerance. Similarly, Harting et al. (2008) noted persistent skin symptoms in the two SICI cases they described. A further drawback of the previous term ‘‘SHCB’’ is that it implies that only babies are affected by this subtype of ARCI, making it unsuitable as a diagnosis when the patient is older. Instead, we propose ‘‘SICI’’ as a better or complementary name for this pleiomorphic form of ichthyosis. In conclusion, SICI illustrates the genetic diversity and clinical overlap between different types of ARCI. Only a careful dissection of clinical and genetic findings can define the SICI subtype of ARCI. It is important for clinicians to be aware of cases of congenital ichthyosis, in which a spontaneous transition from a severe to a mild phenotype occurs and to understand the recessive nature of this disease when asked for genetic counseling. MATERIALS AND METHODS The patients were recruited by a national survey of ARCI that has been ongoing in Sweden since 1997 (Vahlquist et al., 2003) and in Denmark since 2004 (A. Bygum et al., unpublished observation). 442


The study has been approved by the regional ethics committees (EPN) in Uppsala and Odense, and carried out in accordance with the Declaration of Helsinki Principles. Briefly, the survey involved a team of doctors and nurses examining the patients in a standardized manner and collecting data from other hospital departments where the patients had received previous medical care for ichthyosis. After informed consent, EDTA blood sample was collected from the patient and the parents if available. Among nearly 260 ARCI patients, 15 cases of SICI were identified. Some of these patients were examined by us soon after birth and followed up prospectively for several years. Other patients (children and adults) with typical features of SICI at birth were identified retrospectively by interviewing the patient or parents about the neonatal period, by looking at photos taken soon after delivery, and by consulting the patients’ hospital records. The age at the last examination is indicated in Table 1, together with other characteristics of the patients. Blood DNA was initially screened for TGM1 and ICHTHYIN mutations at the Department of Clinical Genetics (Uppsala University), using the same approaches as previously described (Pigg et al., 1998; Dahlqvist et al., 2007). The STS gene was investigated in boys to exclude X-linked recessive ichthyosis with neonatal onset. Samples without TGM1 or ICHTHYIN mutations were sent to Evry (JF, CNG) for further sequencing of the ALOX genes and other ARCI-related genes (Jobard et al., 2002; Lefevre et al., 2003, 2006; Lesueur et al., 2007; Fischer, 2009). CONFLICT OF INTEREST The authors state no conflict of interest.

ACKNOWLEDGMENTS This study was supported by grants from the Swedish Research Council, the Welander-Finsen Foundations, Kgl. Hofbuntmager Aage Bangs Fond, the EU Geneskin project (EC grant LSHM-CT-2005-512117), and the Centre National de Geńotypage.

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