REPORT Cole Disease Results from Mutations in ENPP1 Ori Eytan, 1,2,11 Fanny Morice-Picard, 3,4,11 Ofer Sarig, 1 Khaled Ezzedine, 3,4 Ofer Isakov, 5 Qiaoli Li, 6,7 Akemi Ishida-Yamamoto, 8 Noam Shomron, 5 Tomer Goldsmith, 1 Dana Fuchs-Telem, 1,2 Noam Adir, 9 Jouni Uitto, 6,7 Seth J. Orlow, 10 Alain Taieb, 3,4, * and Eli Sprecher 1,2, * The coexistence of abnormal keratinization and aberrant pigmentation in a number of cornification disorders has long suggested a mechanistic link between these two processes. Here, we deciphered the genetic basis of Cole disease, a rare autosomal-dominant geno- dermatosis featuring punctate keratoderma, patchy hypopigmentation, and uncommonly, cutaneous calcifications. Using a combina- tion of exome and direct sequencing, we showed complete cosegregation of the disease phenotype with three heterozygous ENPP1 mutations in three unrelated families. All mutations were found to affect cysteine residues in the somatomedin-B-like 2 (SMB2) domain in the encoded protein, which has been implicated in insulin signaling. ENPP1 encodes ectonucleotide pyrophosphatase/phosphodies- terase 1 (ENPP1), which is responsible for the generation of inorganic pyrophosphate, a natural inhibitor of mineralization. Previously, biallelic mutations in ENPP1 were shown to underlie a number of recessive conditions characterized by ectopic calcification, thus providing evidence of profound phenotypic heterogeneity in ENPP1-associated genetic diseases. Cole disease is a rare autosomal-dominant disorder that has been reported in four families since its original descrip- tion in 1976. 1 The disorder is characterized by congenital or early-onset punctate keratoderma associated with irreg- ularly shaped hypopigmented macules, which are typically found over the arms and legs, but not the trunk or acral regions. 1–4 Extracutaneous involvement has not been reported. The pathogenesis of the disorder has remained elusive. Skin biopsies obtained from palmoplantar lesions show hyperorthokeratosis, hypergranulosis, and acantho- sis and are therefore not indicative of any specific pathomechanism. 2,3 In contrast, upon histopathological examination, hypopigmented areas of the skin demon- strate a reduction in melanin content in keratinocytes, but not in melanocytes, as well as hyperkeratosis and a normal number of melanocytes. 4 Accordingly, ultrastruc- tural studies have revealed that melanocytes show a disproportionately large number of melanosomes in the cytoplasm and dendrites but that keratinocytes show a paucity of these organelles, suggestive of impaired melano- some transfer. 4 Attempts at identifying the molecular basis of Cole dis- ease through sequencing of candidate genes associated with overlapping phenotypes, such as epidermolysis bul- losa simplex with mottled pigmentation (MIM 131960) and Naegeli syndrome (MIM 161000), have failed to reveal causative mutations. 2 In the present report, using a genome-wide approach, we demonstrate that mutations in ENPP1 (MIM 173335), encoding ectonucleotide pyro- phosphatase/phosphodiesterase 1 (ENPP1), 5 underlie Cole disease. We studied three families affected by the disorder (Figure 1A). Families A and C are of French origin and have not been reported previously. Family B has been pre- viously described 2 and originates from the United States. All affected individuals displayed similar clinical and histo- pathological features, as detailed in Table 1. In brief, affected individuals developed hypopigmented macules mainly located over the extremities and hyperkeratotic papules over the palms and soles (Figures 1B–1C). Age of onset varied between 3 months and 1 year. All participants provided written informed consent to participate in this study according to a protocol reviewed and approved by our institutional review boards. Genomic DNA was extracted either from whole-blood samples with the 5 Prime ArchivePure DNA Blood Kit (5 PRIME) or from saliva collection kits (DNA Genotek). To identify the caus- ative mutations underlying Cole syndrome in our families, we used DNA samples from individuals II-1, II-3, III-2, and III-4 from family A and from individual II-1 from family B for whole-exome capture and next-generation sequencing. Exome sequencing was performed by Macrogen (family A) or by Galil Genetic Analysis (family B) with the same meth- odology. Whole-exome capture was done by in-solution hybridization with TruSeq (Illumina) and subsequent massive parallel sequencing (Illumina HiSeq2000) with 100 bp paired-end reads. Reads were aligned to GRCh37 (UCSC Genome Browser hg19) with the Burrows-Wheeler 1 Department of Dermatology, Tel Aviv Sourasky Medical Center, Tel Aviv 642395, Israel; 2 Department of Human Molecular Genetics & Biochemistry, Sack- ler Faculty of Medicine, Tel Aviv University, Ramat Aviv 69978, Israel; 3 Department of Dermatology and Pediatric Dermatology, National Center for Rare Skin Disorders, Saint Andre ´ Hospital, Bordeaux 33075, France; 4 Institut National de la Sante ´ et de la Recherche Me ´dicale 1035, University of Bordeaux 33075, France; 5 Department of Cell and Developmental Biology, Faculty of Medicine, Tel-Aviv University, Ramat-Aviv 69978, Israel; 6 Departments of Dermatology and Cutaneous Biology and Biochemistry and Molecular Biology, Jefferson Medical College, Thomas Jefferson University, Philadelphia, PA 19107, USA; 7 Jefferson Institute of Molecular Medicine, Thomas Jefferson University, Philadelphia, PA 19107, USA; 8 Department of Dermatology, Asahi- kawa Medical University, Asahikawa 078-8510, Japan; 9 Schulich Faculty of Chemistry, Technion-Israel Institute of Technology, Technion City, Haifa 32000, Israel; 10 The Ronald O. Perelman Department of Dermatology, New York University School of Medicine, New York, NY 10016, USA 11 These authors contributed equally to this work *Correspondence: alain.taieb@chu-bordeaux.fr (A.T.), elisp@tasmc.health.gov.il (E.S.) http://dx.doi.org/10.1016/j.ajhg.2013.08.007. Ó2013 by The American Society of Human Genetics. All rights reserved. 752 The American Journal of Human Genetics 93, 752–757, October 3, 2013