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Osteogenesis Imperfecta (OI) -
Guidelines for Diagnostic Testing

OI is a genetically heterogeneous disorder characterized primarily by fragile bones presenting with fracture and bone deformity. The clinical severity and variation in age of onset and type of OI is determined by the particular gene mutation. A meeting in June 2010 of directors of European diagnostic laboratories to discuss strategies for laboratory diagnosis of OI was considered in designing our approach to testing:  EMQN best practice guidelines for the laboratory diagnosis of osteogenesis imperfecta 

Step 1: Type I Collagen Gene Sequencing - COL1A1 and COL1A2

About 90% of individuals with OI (perinatal lethal and non-lethal) have a mutation in one of the two type I collagen genes (COL1A1 and COL1A2).   In these families the condition is dominantly inherited.  Roughly half of the cases that we identify are new mutations. The first step in the lab evaluation of any form of OI is genomic sequencing of COL1A1 and COL1A2. The majority of the remaining individuals (without a type I collagen gene mutation) are likely to be homozygous or compound heterozygous for recessive gene mutations.

Step 2: Recessive OI Panel Gene Sequencing: FKBP10, CRTAP, LEPRE1, PPIB, SERPINH1, SP7, SERPINF1, AND PLOD2

To date, there are 8 additional genes (CRTAP, LEPRE1, PPIB, FKBP10, SERPINH1, SERPINF1, PLOD2, or OSX) determined to cause OI.  In these families, the disorder is recessively inherited.  As the clinical characteristics vary by gene and overlap, it is possible but impractical to choose a single gene test or sequence in a tiered fashion.   In our experience, it is easier to sequence all the recessive OI genes at one time saving time and money.  The recessive OI panel includes the 8 genes in which mutations have been identified and will be expanded as others are identified.

Recessive gene testing should be considered when a severe OI phenotype persists in the presence of normal COL1A1 and COL1A2 sequencing and

• When familial consanguinity is reported.
• In families with two or more affected children born to unaffected parents (although the greatest proportion of recurrence results from parental mosaicism for dominant gene mutations)
• When the affected individual is from a geographic or ethnic population known to have a "founder" mutation in one of the recessive genes. The populations include W. Africans, Vietnamese, Burmese, Samoan and First Nation Canadians.

The exception to this approach may be the family with a clear OI type I phenotype or dominant inheritance.  In such instances, study of gene copy number of COL1A1 may be indicated by array or MLPA to evaluate gene copy number.  Haploinsufficiency for COL1A1 is expected to result in OI type I but is seen in a very small group.  The same alteration in COL1A2 is thought to have no clinical correlation. 

Step 3: Studies of collagens synthesized by cultured fibroblasts

Abnormal type I collagen screening is an indication that an OI gene mutation is present and that it disrupts the synthesis, folding, excretion or assembly of collagen chains. There are several instances when "collagen screening" is recommended.

1. To look for evidence of a multi-exon deletion if exon by exon sequencing is normal in a patient with a clear OI phenotype.
2. To evaluate the protein consequence of an "unknown" sequence alteration identified by genomic sequencing.
3. When an adequate blood sample is not available for testing.

 ADDITIONAL CONSIDERATIONS

Research studies for new genes

If no mutation is found at this point, clinical review and analysis of cultured cells for abnormal type I procollagen production could be the first approach.  If no abnormalities are found to direct further mutation search, then DNA and/or cells from these families should be stored in preparation for identification of new genes and for whole exome/genome approaches to gene discovery.

Unexplained fractures and the consideration of abuse

There are about 400 infants born with OI in the US each year (an incidence of about 1/10,000).  In the age group in which abuse that results in fracture is most frequent (0-3 years) about 25,000 children are identified each year with fractures.  Thus we could expect that if there is no screening about 5% of those children with unexplained fractures would have OI, quite similar to the number we have identified in the course of studying children in whom the question of abuse had been raised. 

To date, all infants in which the recessive forms of OI have been identified have had clearly recognized skeletal alterations either at birth or within a very short time thereafter.  Among children with OI that results from mutations in type I collagen genes those with OI type II (the perinatal lethal form) and OI type III (the progressive deforming variety) can be recognized at birth from both their clinical presentation and the radiographic alterations.  The blue sclerae of infants with OI type I may be difficult to appreciate and so the diagnosis could be missed and at an early age, some of the clinical features of OI type IV might not be seen (tooth a sclearal hue abnormalities).

Given these considerations, we think that in this context, analysis of type I collagen genes by DNA sequence determination is sufficient such that a normal result diminishes the likelihood that the child has OI to well below 1%.  If the clinical indications of OI are still very strong, then analysis of the collagen produced by cultured cells should assist in the identification of the underlying alteration.  In this context, we do not recommend sequence analysis of the recessive OI genes.

References:

Osteogenesis Imperfecta - Diagnosis

Byers PH The Metabolic & Molecular Basis of Inherited Disease 8th Edition Volume IV 2000 Disorders of Collagen Biosynthesis and Structure p 5241-85.

Byers PH et al. Perinatal lethal osteogenesis imperfecta (OI type II): a biochemically heterogeneous disorder usually due to new mutations in the genes for type I collagen. Am J Hum Genet. 1988 Feb;42(2):237-48.

Sillence DO et al. Genetic heterogeneity in osteogenesis imperfecta J Med Genet 1979 Apr;16(2):101-16.

Wenstrup RJ et al. Distinct biochemical phenotypes predict clinical severity in nonlethal variants of osteogenesis imperfecta. Am J Hum Genet. 1990 May;46(5):975-82.

Glorieux FH et al. Type V osteogenesis imperfecta: a new form of brittle bone disease. J Bone Miner Res 2000 Sep;15(9):1650.

Glorieux FH et al. Osteogenesis imperfecta type VI: a form of brittle bone disease with a mineralization defect. J Bone Miner Res 2002 Jan;17(1):30-8.

Morello, R et al Cell 2006 Oct 20;127(2):291-304. CRTAP is required for prolyl 3- hydroxylation and mutations cause recessive osteogenesis imperfecta.

Barnes AM et al NEJM 2006 Dec 28;355(26):2757-64. Deficiency of cartilage-associated protein in recessive lethal osteogenesis imperfecta.

Cabral WA et al Nat Genet Prolyl 3-hydroxylase 1 deficiency causes a recessive metabolic bone disorder resembling lethal/severe osteogenesis imperfecta. 2007 Mar;39(3):359-65.

Van Dijk FS et al  Am J Hum Genet 2009 Oct;85(4):521-7 PPIB mutations cause severe osteogenesis imperfecta.

Barnes AM et al  NEJM 2010 Feb 11;362(6):521-8. Lack of cyclophilin B in osteogenesis imperfecta with normal collagen folding.

Christiansen HE et al  Am J Hum Genet 2010 Mar 12;86(3):389-98. Homozygosity for a missense mutation in SERPINH1, which encodes the collagen chaperone protein HSP47, results in severe recessive osteogenesis imperfecta.

Alanay Y et al  "American journal of human genetics." Am J Hum Genet. 2010 Apr 9;86(4):551-9. Mutations in the gene encoding the RER protein FKBP65 cause autosomal-recessive osteogenesis imperfecta.

Prenatal Diagnosis and Mode of Delivery:

Pepin M et al. Strategies and outcomes of prenatal diagnosis for osteogenesis imperfecta: a review of biochemical and molecular studies completed in 129 pregnancies. Prenat Diagn. 1997 Jun;17(6):559-70.

Cubert R et al. Osteogenesis imperfecta: mode of delivery and neonatal outcome. Obstet Gynecol. 2001 Jan;97(1):66-9.