Molecular Genetics Autosomal Dominant Osteogenesis Imperfecta Panel

Osteogenesis Imperfecta (OI) is a group of disorders that affect bone fragility and formation. OI is characterized by fractures with minimal or absent trauma, short stature, blue sclera, and dentinogenesis imperfecta. There are 19 known types of this disorder with a wide range of features and severities. A majority of patients with OI have a mutation in either the COL1A1 or COL1A2 gene, which are the genes that code for alpha 1 and alpha 2 chains of collagen type I. Mutations in COL1A1 and COL1A2 are associated with autosomal dominant inheritance and can present with a wide phenotypic spectrum from perinatal lethality to mild predisposition to fractures.

Another autosomal dominant form of OI is caused by mutations in IFITM5. Mutations in this gene are associated with OI type V, which also has a wide phenotypic range, but does have distinct clinical and radiological features. Autosomal recessive OI is less common.

The results from genetic tests can facilitate assessment of levels of risk for patients and lead to more efficient and appropriate medical management. It is recommended that this testing be accompanied by a complete family history and genetic counseling.

Reasons for Referral:

• Molecular confirmation of a clinical diagnosis of Osteogenesis Imperfecta
• To aid in decision-making for treatment and management of individuals with Osteogenesis Imperfecta.
• Testing for individuals with a positive family history for Osteogenesis Imperfecta (targeted testing is available if the familial mutation is known).

Genomic DNA is analyzed using next-generation sequencing (NGS) on the Illumina NextSeq 2000 platform, with target enrichment performed using hybridization-based probes to capture exonic (coding) regions of the gene(s). Single nucleotide variants (SNVs) and small insertions or deletions (INDELs) are identified using the Illumina DRAGEN Enrichment Workflow, executed onboard the NextSeq2000. This pipeline combines software and hardware acceleration to generate high-confidence germline haplotype calls. Clinical and analytical validation of DRAGEN was performed in our laboratory. Based on validation study results, for SNVs, this assay achieves >96% analytical sensitivity and >99% positive predictive value (PPV). For INDELs <50 bp, the analytical sensitivity is >87% and the PPV is >97%. INDELs >50 bp may be detected but the sensitivity for these is reduced.

Exon-level copy number variants (CNVs) are detected using the Germline Copy Number Variation Best Practices pipeline from GATK. A Bayesian model, clinically validated in our laboratory, enables detection of deletions and duplications involving three or more contiguous exons in genes with adequate probe coverage and without complicating factors (e.g. pseudogene homology, short tandem repeats, segmental duplications). Please note that exon-centric microarray remains the gold standard for exonic copy number variant calling. If exon-centric microarray is of interest, please contact the laboratory for additional information.

This test is not designed to detect polynucleotide repeats, low-level mosaicism, structural rearrangements or balanced alterations (e.g. inversions, gene conversion events, translocations, etc.) or variants in difficult regions. Additionally, variants located in regions of insufficient coverage, including introns and promoter regions; pseudogenes; where the reference genome is inaccurate or contains gaps and insertions; and of high GC content may not be detected. This test does not provide complete coverage of all exons and noncoding regions may have limited information and ability to interpret. Variants in introns that are greater than 10 bp from the intron-exon junction may be analyzed. Please contact the laboratory if interrogation of intronic sequence greater than 10 bp from the intron-exon boundary is desired. 

COL1A1, COL1A2, IFITM5, LRP5

Blood: EDTA or ACD (Solution A or B):

• Adult: 5 mL
• Child: 5 mL
• Infant: 2-3 mL

Saliva: 2 ORAgene™ Saliva Collection Kit(s) (OGR-500) used according to manufacturer instructions. Please contact KDL Client Services for a Saliva Collection Kit for patients that cannot provide a blood sample.

Assisted Saliva: 4 ORAgene™ Assisted Saliva Collection Kits (OGR-575) used according to manufacturer instructions.  Please contact KDL Client Services for an Assisted Saliva Collection Kit for patients that cannot provide a blood sample.

Buccal Cells: 4 CytoSoft™ Cytology Brush (Medical Packaging CYB-1) used according to manufacturer instructions.  Please contact KDL Client Services for a Buccal Collection Kit for patients that cannot provide a blood sample.

Skin Fibroblast: Punch Biopsy (Cell cultures will be prepared at KDL and used for testing), or 2 T-25 confluent flasks

Prenatal: 

• Direct Amniotic Fluid (10-20mL)
• Direct CVS
• Cultured Amniocytes (2-T25 flasks)
• Cultured CVS (2-T25 flasks)
• Cultured Fetal Tissue: Product of Conception (2-T25 flasks)
• Cord Blood (1-2-mL)

DNA: 5-10µg at a minimum of 60-100ng/µL (DNA must be extracted in a CLIA-certified laboratory or a laboratory meeting equivalent requirements as determined by the CAP and/or CMS)

Maternal cell rule-out testing will be performed on all prenatal specimens received. Please provide maternal blood (or saliva) in addition to the fetal specimen. Additional charges apply for the maternal cell rule-out test.

For routine testing of blood and saliva (or DNA extracted from them), KDL does NOT accept samples from patients within two (2) weeks of a packed cell/platelet transfusion or within four (4) weeks of a whole blood transfusion.  For extraordinary circumstances, where testing must be performed within the above windows, please contact our lab.

REQUISITION FORM MUST ACCOMPANY ALL SAMPLES.  Please include detailed clinical information, including ethnicity, clinical history, and family history.

Weekly

6-8 weeks

• Package and ship specimen to remain cold, but not frozen. 
• Ship via overnight express, using the FedEx priority overnight label provided. 
• Contact Client Services for shipping kits and instructions at (855) 535-1522.

1. Genetics Home Reference https://ghr.nlm.nih.gov/condition/osteogenesis-imperfecta
2. Uncommon IFITM5 mutation associated with severe skeletal deformity in osteogenesis imperfecta. Rodriguez Celin M1, Moosa S2, Fano V1. https://www.ncbi.nlm.nih.gov/pubmed/30039845
3. Genotype-Phenotype Correlations in Autosomal Dominant Osteogenesis Imperfecta Mouna Ben Amor, Francis H. Glorieux, and Frank Rauch Shriners Hospital for Children and McGill University, Montreal, QC, Canada https://doi.org/10.4061/2011/540178