PLS3 Sequencing in Childhood-Onset Primary Osteoporosis Identifies Two Novel Disease-Causing Variants

Osteoporos Int (2017) 28:3023–3032 DOI 10.1007/s00198-017-4150-9 ORIGINAL ARTICLE PLS3 sequencing in childhood-onset primary osteoporosis identifies two novel disease-causing variants A. J. Kämpe1,2 & A. Costantini1,2 & R. E. Mäkitie3 & N. Jäntti1,2 & H. Valta4 & M. Mäyränpää4 & H. Kröger5 & M. Pekkinen3 & F. Taylan 1,2 & H. Jiao6 & O. Mäkitie1,2,3,4 Received: 27 December 2016 /Accepted: 6 July 2017 /Published online: 26 July 2017 # The Author(s) 2017. This article is an open access publication Abstract osteoporosis. Cohort I comprised 31 patients with childhood- Summary Altogether 95 children with primary bone fragility onset primary osteoporosis of unknown etiology. Cohort II were screened for variants in PLS3, the gene underlying X- comprised 64 children who had sustained multiple fractures linked osteoporosis. Two children with multiple peripheral but were otherwise healthy. Clinical and radiological data and spinal fractures and low BMD had novel disease- were reviewed. Peripheral blood DNA was Sanger sequenced causing PLS3 variants. Children with milder phenotypes had for coding exons and flanking intronic regions of PLS3. no pathogenic variants. PLS3 screening is indicated in Results In two patients of cohort I, where other common ge- childhood-onset primary osteoporosis. netic causes had been excluded, we identified two novel Introduction The study aimed to determine the role of patho- disease-causing PLS3 variants. Patient 1 was a male with genic PLS3 variants in children’s bone fragility and to eluci- bilateral femoral fractures at 10 years, low BMD (Z-score date the associated phenotypic features. −4.1; 18 years), and multiple vertebral compression fractures. Methods Two cohorts of children with bone fragility were He had a novel nonsense variant in PLS3. Patient 2 was a girl screened for variants in PLS3, the gene underlying X-linked with multiple long bone and vertebral fractures and low BMD (Z-score −6.6 at 6 years). She had a de novo missense variant in PLS3; whole exome sequencing and array-CGH identified no other genetic causes. Iliac crest bone biopsies confirmed Electronic supplementary material The online version of this article low-turnover osteoporosis in both patients. In cohort II,no (doi:10.1007/s00198-017-4150-9) contains supplementary material, which is available to authorized users. pathogenic PLS3 variants were identified in any of the subjects. * A. J. Kämpe Conclusions Two novel disease-causing variants in PLS3 [email protected] were identified in a boy and a girl with multiple peripheral and spinal fractures and very low BMD while no pathogenic variants were identified in children with less severe skeletal 1 Department of Molecular Medicine and Surgery, Karolinska Institutet, Stockholm, Sweden fragility. PLS3 screening is warranted in male and female patients with childhood-onset primary osteoporosis. 2 Center for Molecular Medicine, Karolinska Institutet, Stockholm, Sweden 3 Keywords Children . Early-onset osteoporosis . Fractures . Folkhälsan Institute of Genetics and University of Helsinki, . Helsinki, Finland Osteogenesis imperfecta Plastin 3 X-Linked osteoporosis 4 Children’s Hospital, University of Helsinki and Helsinki University Hospital, Helsinki, Finland 5 Department of Orthopaedics and Traumatology, Bone and Cartilage Introduction Research Unit, Kuopio University Hospital and University of Eastern Finland, Kuopio, Finland Childhood-onset primary osteoporosis is a rare but clinically 6 Department of Biosciences and Nutrition and Science for Life important condition characterized by reduced bone strength Laboratory, Karolinska Institutet, Stockholm, Sweden andanelevatedriskoffractures[1]. Current definition 3024 Osteoporos Int (2017) 28:3023–3032 requires a clinically significant fracture history and BMD Z- Materials and methods score at or below −2.0. A clinically significant fracture history is defined as (1) two or more long bone fractures ≤10 years This study involves two patient cohorts assessed for primary and (2) three or more long bone fractures ≤19 years [2]. When skeletal fragility at the Children’s Hospital, Helsinki vertebral compression fractures are present, they alone can be University Hospital. The institutional Ethics Review Board grounds for the diagnosis even when BMD is normal [2]. The at Helsinki University Hospital approved the study protocol, diagnosis further requires that other nutritional or medical and all patients and/or their guardians gave a written informed causes for the child’s bone fragility have been excluded. consent before participation. With such a broad definition, it is easy to understand that childhood-onset primary osteoporosis includes a spectrum of skeletal diseases with different molecular causes. Most affect- Cohort characteristics ed children are classified as having osteogenesis imperfecta (OI), in which the great majority of cases are due to pathogen- Cohort 1 (Bprimary osteoporosis^) consisted of 31 patients ic variants in either COL1A1 or COL1A2, the two genes (17 boys and 14 girls), who had been referred to the encoding type I collagen [3, 4]. However, during the last Metabolic Bone Clinic, Children’s Hospital, Helsinki during 10 years, studies have shown that pathogenic variants in 20 the years 2003–2013 for investigation for childhood-onset different genes can cause childhood-onset primary osteoporo- primary osteoporosis but where the investigation did not re- sis [5–11]. veal a molecular cause of their disease. The inclusion criteria PLS3, located on the X chromosome, is one of the most were (1) exclusion of type I collagen-related OI, either clini- recently described genes underlying childhood-onset primary os- cally or by genetic testing, and (2) exclusion of secondary teoporosis. PLS3 osteoporosis was first described in five families osteoporosis with biochemical and individually determined with apparent X-linked osteoporosis in 2013, and the gene was clinical evaluations. Further, all had been screened and found subsequently shown to be of importance in bone metabolism negative for pathogenic variants in WNT1 and LRP5 and sev- [12]. PLS3 codes for the protein Plastin3, which is widely eral had undergone more extensive genetic testing before in- expressed in solid tissues and thought to be involved in cytoskel- clusion. At the time of referral, all the patients were between eton remodeling. Plastin3 contains two actin-binding sites and the ages of 4 and 17 years, displayed clinically an osteoporotic two calcium-binding sites, andininvitroexperiments,itcan phenotype, and several of them had similarly affected family form bundles of actin by crosslinking single filaments to each members. Most of the patients (25/31) fulfilled the ISCD other [13, 14]. In bone, Plastin3 has been suggested to either be criteria [2] for pediatric osteoporosis. Six patients only had a part of the osteocytes’ mechanosensing apparatus or play a role history of increased non-spinal fractures and a BMD Z-score in the mineralization process [12, 15]. Plastin3 has also been >−2.0 but had other additional features (e.g., osteopenic ap- reported as a protective modifier in spinal muscular atrophy pearance on skeletal radiographs, fragile bone on orthopedic (SMA) and to be important in axonogenesis [16, 17]. A PLS3 surgery) suggesting osteoporosis. knockout mouse model shows decreased BMD and knockdown Cohort II (Bfracture prone^) consisted of 64 otherwise of PLS3 in Zebrafish results in craniofacial dysplasia in the de- healthy children who had sustained multiple fractures (43 veloping Zebrafish larvae [12, 18]. However, as of yet, the direct boys and 21 girls), all recruited at the Children’s Hospital, function of Plastin3, the mechanism of its action, and pathogen- Helsinki, Finland during a prospective epidemiological study esis of PLS3 osteoporosis are still undetermined. [20]. Over a 12-month period, all children (n = 1412) aged 4– Because of PLS3’s location on the X chromosome, males 15 years who had been treated for an acute and radiographi- are in general more severely affected by loss of function var- cally confirmed fracture were assessed for fracture history. iants than females. PLS3 osteoporosis is foremost character- The trauma mechanisms and previous medical histories were ized by vertebral compression fractures, frequent peripheral available for 1361 (96%) of the children. The inclusion criteria fractures, and a low BMD [12, 19]. Other traits common in for cohort II were (1) age 4–15 years, (2) ≥2 low-energy long classical OI, such as blue sclerae, joint hyperlaxity, and short bone fractures before age 10 years, (3) ≥3low-energylong stature, are usually not present. However, only eight families bone fractures before age 16 years, or (4) ≥1low-energyver- with childhood-onset primary osteoporosis due to pathogenic tebral fracture (loss of ≥20% vertebral height). Children with a variants in PLS3 have been described in the literature, and thus diagnosis or suspicion of OI were excluded, as well as chil- the features of PLS3 osteoporosis have not been fully charac- dren with an underlying disease that could explain their bone terized [12, 15, 19]. fragility. Altogether 71 patients of the >1400 children fulfilled In this study, we have attempted (1) to determine the over- these criteria; DNA from peripheral blood was available for all role of pathogenic variants in PLS3 in children with bone 64 of them and they comprised cohort II. Data on blood bio- fragility and (2) to increase the understanding of the clinical chemistry, spinal radiographs,

Load more