Molecular Diagnoses of X-Linked and Other Genetic Hypophosphatemias: Results From a Sponsored Genetic Testing Program

Abstract

X‐linked hypophosphatemia (XLH), a dominant disorder caused by pathogenic variants in the PHEX gene, affects both sexes of all ages and results in elevated serum fibroblast growth factor 23 (FGF23) and below‐normal serum phosphate. In XLH, rickets, osteomalacia, short stature, and lower limb deformity may be present with muscle pain and/or weakness/fatigue, bone pain, joint pain/stiffness, hearing difficulty, enthesopathy, osteoarthritis, and dental abscesses. Invitae and Ultragenyx collaborated to provide a no‐charge sponsored testing program using a 13‐gene next‐generation sequencing panel to confirm clinical XLH or aid diagnosis of suspected XLH/other genetic hypophosphatemia. Individuals aged ≥6 months with clinical XLH or suspected genetic hypophosphatemia were eligible. Of 831 unrelated individuals tested between February 2019 and June 2020 in this cross‐sectional study, 519 (62.5%) individuals had a pathogenic or likely pathogenic variant in PHEX (PHEX‐positive). Among the 312 PHEX‐negative individuals, 38 received molecular diagnoses in other genes, including ALPL, CYP27B1, ENPP1, and FGF23; the remaining 274 did not have a molecular diagnosis. Among 319 patients with a provider‐reported clinical diagnosis of XLH, 88.7% (n = 283) had a reportable PHEX variant; 81.5% (n = 260) were PHEX‐positive. The most common variant among PHEX‐positive individuals was an allele with both the gain of exons 13–15 and c.*231A>G (3′UTR variant) (n = 66/519). Importantly, over 80% of copy number variants would have been missed by traditional microarray analysis. A positive molecular diagnosis in 41 probands (4.9%; 29 PHEX positive, 12 non‐PHEX positive) resulted in at least one family member receiving family testing. Additional clinical or family member information resulted in variant(s) of uncertain significance (VUS) reclassification to pathogenic/likely pathogenic (P/LP) in 48 individuals, highlighting the importance of segregation and clinical data. In one of the largest XLH genetic studies to date, 65 novel PHEX variants were identified and a high XLH diagnostic yield demonstrated broad insight into the genetic basis of XLH. © 2021 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

Novel PHEX gene locus‐specific database: Comprehensive characterization of vast number of variants associated with X‐linked hypophosphatemia (XLH)

“…ClinVar reports 814 XLH‐related variants in PHEX (accessed May 24, 2021), a recently updated count which includes 287 variants identified in the hypophosphatemia genetic testing program and reported to ClinVar (Rush et al, 2021 ). The locus‐specific database contributes another 50 novel XLH‐related PHEX variants which are presented here for the first time (Table S2 ).…”

“…The most prevalent variant in the database is a c.*231A>G substitution in the 3′‐UTR, which was frequently reported co‐occurring with an exon 13–15 duplication, suggesting that these two variants co‐segregate and constitute a single allele. In an analysis reported from the hypophosphatemia gene panel program, these variants were found together in both males and females in 65 of 66 probands and were shown to be in cis in all 51 individuals for whom phasing information was available (Rush et al, 2021 ).…”

“…Although the overall pattern of variants is consistent with previous studies, there has been a large increase in the number of CNV reported, which is likely a reflection of improved genetic testing methods for CNV. In the hypophosphatemia genetic testing program, 81% of the CNV events detected were sub‐genic and would be missed by traditional microarray analysis (Rush et al, 2021 ). However, despite the increased detection of CNV from more recent reports, most of the routine clinical testing methods employed so far, including those used in the hypophosphatemia genetic testing program, cannot detect deep intronic variants or large structural variants with breakpoints outside of the PHEX coding region.…”

“…The database was developed by integrating PHEX variants from four different sources: (1) a now inactive McGill University PHEX locus‐specific database (Sabbagh et al, 2000 ), last updated April, 2017, (2) genetic testing results from burosumab clinical trials, (3) a comprehensive literature review, and (4) a sponsored hypophosphatemia genetic testing program initiated by Ultragenyx Pharmaceutical Inc. and Invitae Corporation, which provided no‐charge next‐generation sequencing with a multi‐gene panel to confirm a clinical XLH diagnosis or to aid diagnosis of suspected XLH or other genetic hypophosphatemia (Rush et al, 2021 ). The hypophosphatemia genetic testing program was designed to detect single nucleotide variants (SNV), small and large insertions/deletions (indels), sub‐genic structural variants, and exon‐level copy number variants (CNV).…”

“…A new locus‐specific database for PHEX gene variants, PHEX LSDB, was established to collect and disseminate information to the scientific community and affected families about disease‐causing PHEX variants ( https://www.rarediseasegenes.com/ ). The database compiles variants from four sources: an older, archived PHEX locus‐specific variants database (Sabbagh et al, 2000 ), variants identified in a recent hypophosphatemia genetic testing program (Rush et al, 2021 ), unpublished variants identified in previous XLH clinical studies, and previously published variants identified from a comprehensive literature review. The PHEX LSDB will be updated regularly with new information on PHEX variants as reported in the literature or submitted directly to the database website.…”