Identification of genetic determinants underlying familial osteoporosis

Abstract

Introduction & Aim: Osteoporosis is a skeletal disease with a strong genetic influence. The study aimed to identify gene variants underlying early-onset osteoporosis in a nuclear Maltese pedigree. Methods: A two-generation family consisting of a 60-year-old affected father, a non-affected mother (54 years) and 2 affected male siblings (aged 29 and 32 years) was recruited through an interviewer-led questionnaire. Osteoporosis was defined by DXA measurements of the spine and hip, and secondary causes of osteoporosis were excluded through biochemical testing. The proband was a 29-year-old male with early-onset osteoporosis having sustained bilateral femur fractures before the age of 19 years. WGS was performed on the Illumina NovaSeq 6000 platform with different modes of inheritance investigated by sequential variant filtering and curation using human tissue and animal expression databases, population studies, databases of protein interactions and signalling cascades as well as in silico protein modelling. The shortlisted variants were genotyped in a population-based case-control collection of 1,045 Maltese postmenopausal women and three other independent affected Maltese families from the Malta Osteoporotic Fracture Study (MOFS) using real-time PCR. Genotyping data from the population study was assessed for genotype and allele frequencies, genotype-phenotype associations using the Mann-Whitney U test and exposure odds ratios by logistic regression adjusted for confounders. Results: A total of 9 conserved missense variants were identified following a dominant inheritance pattern, 2 of which were also predicted to alter pre-mRNA splicing, including METTL11B c.38G>T, NBEAL2 c.1948G>A, ADAM8 c.376G>A, TENM4 c.5934C>G, ACACB c.526G>A, PAPLN c.589G>T, ATP8B1 c.3017A>T, ABCA7 c.2326G>A, and PLIN3 c.319C>A. Two other variants, SUSD5 c.745C>A and SUSD5 c.172C>T were also detected following a compound heterozygous inheritance pattern. The alternative allele frequency of most variants was of <1% in line with gnomAD. Heterozygosity for TENM4 c.5934C>G was associated with a lower lumbar spine (LS) T-score (p=0.036) and LS BMD (p=0.034). Women with the GA genotype for the ABCA7 c.2326G>A had lower levels of serum calcium (p=0.045). Heterozygosity for SUSD5 c.745C>A was associated with lower total hip (TH) T-score (p=0.044), TH BMD (p=0.037) and femoral neck (FN) BMD (p=0.029). PLIN3 c.319C>A was associated with a protective effect on LS BMD (Adjusted OR: 0.36 [95% CI 0.15-0.85], p=0.02) which could be a result of the small sample size and genotyping bias. None of the variants were detected in the other Maltese families. Conclusion: The identified gene variants, alone or in combination, might play a role in the pathogenesis of early-onset osteoporosis with varying effect sizes. The encoded proteins are involved in pertinent bone signalling cascades, including Wnt/β-catenin, RANK-RANKL and Notch signalling, amongst others. Further genotyping in larger collections and extended families is warranted to confirm causality, whereas functional studies of the novel PAPLN c.589G>T and ATP8B1 c.3017A>T splicing variants is required.