Osteogenesis Imperfecta via the FKBP10 Gene
- Summary and Pricing
- Clinical Features and Genetics
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The great majority of tests are completed within 18 days.
Due to limited publications, clinical sensitivity is currently unknown. In one publication, FKBP10 mutations were identified in 21 families affected with either autosomal recessive OI or Bruck syndrome (Schwarze et al. 2013). The mutation detection rate should be high, because all reported FKBP10 mutations are point mutations or small deletions and insertions, which can be detected by sequencing.
Osteogenesis imperfecta (OI) is a clinically and genetically heterogeneous skeletal disorder characterized by frequent bone fractures with or without minimal trauma. Clinical signs of OI can range from mild to severe. In addition to bone fractures, patients may have scoliosis, bowing of long bones, short stature, blue sclera, hearing loss, dentin defects, muscle weakness or joint laxity. The incidence is approximately 6-7/100,000 (Dijk et al. 2012). ~90% of clinically diagnosed OI is caused by mutations in the COL1A1 and COL1A2 genes, while ~10% is caused by mutations in the CRTAP, FKB10, LEPRE1, PLOD2, PPIB, SERPINF1, SERPINH1, SP7, WNT1, IFITM5, BMP1, TMEM38B and other undefined genes (Dijk et al. 2012; Valadares et al. 2014).
Mutations in the FKBP10 gene cause a rare severe form of autosomal recessive type XI OI, Bruck syndrome and Kuskokwim syndrome. Bruck syndrome, an osteogenesis imperfecta phenotypic spectrum disorder, is characterized by congenital contractures with pterygia, bone fractures in infancy or early childhood, postnatal short stature, severe limb deformity, and progressive scoliosis. Kuskokwim syndrome is defined by congenital joint contracture with minor bone fractures and occurs solely among populations in Central Alaskan Yupik. The FKBP65 protein coded by the FKBP10 gene belongs to a family of immunophilins and may serve as a peptidylprolyl isomerase required for lysyl hydroxylase activity in forming mature cross-links in bone collagen. To date, ~ 20 unique causative mutations have been identified. They are missense, nonsense, splicing site and small deletions/insertions. No large deletions or duplications have been reported (Shaheen et al. 2010; Barnes et al. 2013; Human Gene Mutation Database). A founder mutation c.948dup (p.Ile317Tyrfs*56) was found in patients from Samoa and nearby islands in the South Pacific (Schwarze (2013). Another founder mutation c.877_879delTAC (p.Tyr293del) was seen in Kuskokwim syndrome. ~ 3% of the population in Central Alaskan Yupik are carriers for this mutation (Barnes et al. 2013).
FKBP65 protein is coded by exons 1 to 10 of the FKBP10 gene on chromosome 17q21.2. Testing involves PCR amplification from genomic DNA and bidirectional Sanger sequencing of the coding exons and ~20bp of adjacent noncoding sequences. We will also sequence any single exon (Test #100) or pair of exons (Test #200) in family members of patients with known mutations or to confirm research results.
Indications for Test
Candidates for this test are patients with symptoms consistent with autosomal recessive OI type XI, Bruck syndrome type I and Kuskokwim syndrome, who have no mutations in the COL1A1 and COL1A2 genes and the family members of patients who have known FKBP10 mutations.
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- Genetic Counselor Team - email@example.com
- Juan Dong, PhD, FACMG - firstname.lastname@example.org
- Barnes AM, Duncan G, Weis M, Paton W, Cabral WA, Mertz EL, Makareeva E, Gambello MJ, Lacbawan FL, Leikin S, Fertala A, Eyre DR, et al. 2013. Kuskokwim Syndrome, a Recessive Congenital Contracture Disorder, Extends the Phenotype of FKBP10 Mutations. Human Mutation 34: 1279-1288. PubMed ID: 23712425
- Dijk FS Van, Byers PH, Dalgleish R, Malfait F, Maugeri A, Rohrbach M, Symoens S, Sistermans EA, Pals G. 2012. EMQN best practice guidelines for the laboratory diagnosis of osteogenesis imperfecta. European Journal of Human Genetics 20: 11-19. PubMed ID: 21829228
- Human Gene Mutation Database (Bio-base).
- Schwarze U, Cundy T, Pyott SM, Christiansen HE, Hegde MR, Bank RA, Pals G, Ankala A, Conneely K, Seaver L, Yandow SM, Raney E, et al. 2013. Mutations in FKBP10, which result in Bruck syndrome and recessive forms of osteogenesis imperfecta, inhibit the hydroxylation of telopeptide lysines in bone collagen. Hum Mol Genet 22: 1-17. PubMed ID: 22949511
- Shaheen R, Al-Owain M, Faqeih E, Al-Hashmi N, Awaji A, Al-Zayed Z, Alkuraya FS. 2011. Mutations in FKBP10 cause both Bruck syndrome and isolated osteogenesis imperfecta in humans. Am. J. Med. Genet. 155: 1448-1452. PubMed ID: 21567934
- Valadares ER, Carneiro TB, Santos PM, Oliveira AC, Zabel B. 2014. What is new in genetics and osteogenesis imperfecta classification? Jornal de Pediatria 90:536-41. PubMed ID: 25046257
Bi-Directional Sanger Sequencing
Nomenclature for sequence variants was from the Human Genome Variation Society (http://www.hgvs.org). As required, DNA is extracted from the patient specimen. PCR is used to amplify the indicated exons plus additional flanking non-coding sequence. After cleaning of the PCR products, cycle sequencing is carried out using the ABI Big Dye Terminator v.3.0 kit. Products are resolved by electrophoresis on an ABI 3730xl capillary sequencer. In most cases, sequencing is performed in both forward and reverse directions; in some cases, sequencing is performed twice in either the forward or reverse directions. In nearly all cases, the full coding region of each exon as well as 20 bases of non-coding DNA flanking the exon are sequenced.
As of March 2016, we compared 17.37 Mb of Sanger DNA sequence generated at PreventionGenetics to NextGen sequence generated in other labs. We detected only 4 errors in our Sanger sequences, and these were all due to allele dropout during PCR. For Proficiency Testing, both external and internal, in the 12 years of our lab operation we have Sanger sequenced roughly 8,800 PCR amplicons. Only one error has been identified, and this was due to sequence analysis error.
Our Sanger sequencing is capable of detecting virtually all nucleotide substitutions within the PCR amplicons. Similarly, we detect essentially all heterozygous or homozygous deletions within the amplicons. Homozygous deletions which overlap one or more PCR primer annealing sites are detectable as PCR failure. Heterozygous deletions which overlap one or more PCR primer annealing sites are usually not detected (see Analytical Limitations). All heterozygous insertions within the amplicons up to about 100 nucleotides in length appear to be detectable. Larger heterozygous insertions may not be detected. All homozygous insertions within the amplicons up to about 300 nucleotides in length appear to be detectable. Larger homozygous insertions may masquerade as homozygous deletions (PCR failure).
In exons where our sequencing did not reveal any variation between the two alleles, we cannot be certain that we were able to PCR amplify both of the patient’s alleles. Occasionally, a patient may carry an allele which does not amplify, due for example to a deletion or a large insertion. In these cases, the report contains no information about the second allele.
Similarly, our sequencing tests have almost no power to detect duplications, triplications, etc. of the gene sequences.
In most cases, only the indicated exons and roughly 20 bp of flanking non-coding sequence on each side are analyzed. Test reports contain little or no information about other portions of the gene, including many regulatory regions.
In nearly all cases, we are unable to determine the phase of sequence variants. In particular, when we find two likely causative mutations for recessive disorders, we cannot be certain that the mutations are on different alleles.
Our ability to detect minor sequence variants, due for example to somatic mosaicism is limited. Sequence variants that are present in less than 50% of the patient’s nucleated cells may not be detected.
Runs of mononucleotide repeats (eg (A)n or (T)n) with n >8 in the reference sequence are generally not analyzed because of strand slippage during PCR and cycle sequencing.
Unless otherwise indicated, the sequence data that we report are based on DNA isolated from a specific tissue (usually leukocytes). Test reports contain no information about gene sequences in other tissues.
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- The test can be added to your online orders in the Summary and Pricing section.
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- A completed requisition form must accompany all specimens.
- Billing information along with specimen and shipping instructions are within the requisition form.
- All testing must be ordered by a qualified healthcare provider.
(Delivery accepted Monday - Saturday)
- Collect 3 ml -5 ml (5 ml preferred) of whole blood in EDTA (purple top tube) or ACD (yellow top tube). For Test #500-DNA Banking only, collect 10 ml -20 ml of whole blood.
- For small babies, we require a minimum of 1 ml of blood.
- Only one blood tube is required for multiple tests.
- Ship blood tubes at room temperature in an insulated container. Do not freeze blood.
- During hot weather, include a frozen ice pack in the shipping container. Place a paper towel or other thin material between the ice pack and the blood tube.
- In cold weather, include an unfrozen ice pack in the shipping container as insulation.
- At room temperature, blood specimen is stable for up to 48 hours.
- If refrigerated, blood specimen is stable for up to one week.
- Label the tube with the patient name, date of birth and/or ID number.
(Delivery accepted Monday - Saturday)
- Send in screw cap tube at least 5 µg -10 µg of purified DNA at a concentration of at least 20 µg/ml for NGS and Sanger tests and at least 5 µg of purified DNA at a concentration of at least 100 µg/ml for gene-centric aCGH, MLPA, and CMA tests, minimum 2 µg for limited specimens.
- For requests requiring more than one test, send an additional 5 µg DNA per test ordered when possible.
- DNA may be shipped at room temperature.
- Label the tube with the composition of the solute, DNA concentration as well as the patient’s name, date of birth, and/or ID number.
- We only accept genomic DNA for testing. We do NOT accept products of whole genome amplification reactions or other amplification reactions.
(Delivery preferred Monday - Thursday)
- PreventionGenetics should be notified in advance of arrival of a cell culture.
- Culture and send at least two T25 flasks of confluent cells.
- Some panels may require additional flasks (dependent on size of genes, amount of Sanger sequencing required, etc.). Multiple test requests may also require additional flasks. Please contact us for details.
- Send specimens in insulated, shatterproof container overnight.
- Cell cultures may be shipped at room temperature or refrigerated.
- Label the flasks with the patient name, date of birth, and/or ID number.
- We strongly recommend maintaining a local back-up culture. We do not culture cells.