Cleft Lip/Palate and Associated Syndromes Panel

Updated
Summary
  • Is a 22 gene panel that includes assessment of non-coding variants
  • Is ideal for patients with cleft lip and/or cleft palate, particularly those who have a positive family history for clefts or who are suspected to have an associated genetic syndrome.

Analysis methods
  • PLUS
Availability

4 weeks

Number of genes

22

Test code

MA3701

Panel size

Small

CPT codes
81479

Summary

The Blueprint Genetics Cleft Lip/Palate and Associated Syndromes Panel (test code MA3701):

ICD codes

Commonly used ICD-10 code(s) when ordering the Cleft Lip/Palate and Associated Syndromes Panel

ICD-10 Disease
Q89.8 Stickler syndrome
Q89.8 Kabuki syndrome
Q35.9 Cleft palate

Sample Requirements

  • Blood (min. 1ml) in an EDTA tube
  • Extracted DNA, min. 2 μg in TE buffer or equivalent
  • Saliva (Oragene DNA OG-500 kit/OGD-500 or OG-575 & OGD-575)

Label the sample tube with your patient's name, date of birth and the date of sample collection.

Note that we do not accept DNA samples isolated from formalin-fixed paraffin-embedded (FFPE) tissue. Read more about our sample requirements here.

Clefts of the lip and/or palate are common birth defects. Cleft lip (CL) with or without cleft palate is found in 1/700 – 1/1,000 births and cleft palate (CP) is found in about 1/1,500 births. In most cases CL and/or CP occur as an isolated malformation but they can be part of several genetic syndromes or chromosomal anomalies. The etiology of the non-syndromic clefts remains poorly understood and multifactorial inheritance is suspected but in some families predisposition to clefts may follow autosomal dominant inheritance with varying penetrance. There are many syndromes that can have clefts as a feature. Van der Woude syndrome (VWS) is caused by a pathogenic mutation in IRF6 or GRHL3 and is characterized by cleft lip with or without cleft palate or isolated cleft palate and lower-lip paramedian pits. It follows autosomal dominant inheritance, but penetrance is incomplete. Cleft palate can be a feature of Stickler syndrome, also known as hereditary arthro-ophthalmopathy, an inherited vitreoretinopathy characterized by the association of ocular signs with abnormalities affecting head and face, bone disorders, and sensorineural deafness. Stickler syndrome caused by mutations in COL2A1, COL11A1 or COL11A2 is inherited in an autosomal dominant manner. Autosomal recessive Stickler syndrome is rare and is caused by biallelic mutations in COL9A1, COL9A2 and COL9A3. Kabuki syndrome (KS) is a multiple congenital anomaly syndrome characterized by typical facial features, skeletal anomalies, mild to moderate intellectual disability and postnatal growth deficiency. Other findings may include cleft lip and/or palate. Kabuki syndrome is caused by mutations in KMT2D or KDM6A.

Genes in the Cleft Lip/Palate and Associated Syndromes Panel and their clinical significance

Gene Associated phenotypes Inheritance ClinVar HGMD
ARHGAP29 AD 3 19
COL11A1 Marshall syndrome, Fibrochondrogenesis, Stickler syndrome type 2 AD/AR 34 94
COL11A2 Weissenbacher-Zweymuller syndrome, Deafness, Otospondylomegaepiphyseal dysplasia, Fibrochondrogenesis, Stickler syndrome type 3 (non-ocular) AD/AR 29 57
COL2A1 Avascular necrosis of femoral head, Rhegmatogenous retinal detachment, Epiphyseal dysplasia, with myopia and deafness, Czech dysplasia, Achondrogenesis type 2, Platyspondylic dysplasia Torrance type, Hypochondrogenesis, Spondyloepiphyseal dysplasia congenital (SEDC), Spondyloepimetaphyseal dysplasia (SEMD) Strudwick type, Kniest dysplasia, Spondyloperipheral dysplasia, Mild SED with premature onset arthrosis, SED with metatarsal shortening, Stickler syndrome type 1 AD 180 561
COL9A1 Stickler syndrome recessive type, Multiple epiphyseal dysplasia type 6 (EDM6) AR 9 6
COL9A2 Stickler syndrome, Multiple epiphyseal dysplasia type 2 (EDM2) AD/AR 7 12
COL9A3 Multiple epihyseal dysplasia type 3 (EDM3) AD/AR 10 14
CTNND1* Blepharocheilodontic syndrome 2, Cleft lip AD 4 16
FOXE1 Thyroid cancer, nonmedullary 4, Hypothyroidism, thyroidal, with spiky hair and cleft palate (Bamforth-Lazarus syndrome), Congenital hypothyroidism AD/AR 4 23
GRHL3 van der Woude syndrome 2 AD 12 27
IRF6 Orofacial cleft, Popliteal pterygium syndrome, van der Woude syndrome AD 45 338
KDM6A Kabuki syndrome XL 40 69
KMT2D Kabuki syndrome AD 350 670
MSX1 Orofacial cleft 5, Tooth agenesis, selective, 1, with/without orofacial cleft, Witkop syndrome AD 11 48
SATB2 Glass syndrome AD 62 106
SPECC1L* Facial clefting, oblique, 1, Opitz GBBB syndrome, type II AD 7 8
TBX2 AD 1 12
TBX22 Cleft palate with or without ankyloglossia XL 12 29
TGDS Catel-Manzke syndrome AR 6 7
TP63 Rapp-Hodgkin syndrome, Orofacial cleft, ADULT syndrome, Ectrodactyly, ectodermal dysplasia, and cleft lip/palate syndrome, Ankyloblepharon-ectodermal defects-cleft lip/palate, Split-hand/foot malformation, Limb-mammary syndrome AD 59 122
TXNL4A Burn-McKeown syndrome AR 19 13
ZSWIM6 Acromelic frontonasal dysostosis AD 4 2

* Some, or all, of the gene is duplicated in the genome. Read more.

# The gene has suboptimal coverage (means <90% of the gene’s target nucleotides are covered at >20x with mapping quality score (MQ>20) reads), and/or the gene has exons listed under Test limitations section that are not included in the panel as they are not sufficiently covered with high quality sequence reads.

The sensitivity to detect variants may be limited in genes marked with an asterisk (*) or number sign (#)

Gene refers to the HGNC approved gene symbol; Inheritance refers to inheritance patterns such as autosomal dominant (AD), autosomal recessive (AR), X-linked (XL), X-linked dominant (XLD) and X-linked recessive (XLR); ClinVar refers to the number of variants in the gene classified as pathogenic or likely pathogenic in this database (ClinVar); HGMD refers to the number of variants with possible disease association in the gene listed in Human Gene Mutation Database (HGMD). The list of associated, gene specific phenotypes are generated from CGD or Orphanet databases.

Non-coding variants covered by Cleft Lip/Palate and Associated Syndromes Panel

Gene Genomic location HG19 HGVS RefSeq RS-number
COL11A1 Chr1:103386637 c.3744+437T>G NM_080629.2
COL11A1 Chr1:103488576 c.1027-24A>G NM_080629.2
COL2A1 Chr12:48379984 c.1527+135G>A NM_001844.4
IRF6 Chr1:209975332 c.-19C>A NM_006147.3
IRF6 Chr1:209975361 c.-48A>T NM_006147.3
IRF6 Chr1:209979367 c.-151G>A NM_006147.3
IRF6 Chr1:209979435 c.-219C>T NM_006147.3
IRF6 Chr1:209989478 c.-10263dupT NM_006147.3
KMT2D Chr12:49428461 c.10356-12G>A NM_003482.3
TBX2 Chr17:59476302 c.-1236G>C NM_005994.3
TBX2 Chr17:59477201 c.-337C>T NM_005994.3
TBX2 Chr17:59477347 c.-191G>A NM_005994.3
TBX2 Chr17:59477352 c.-185delG NM_005994.3
TBX2 Chr17:59477371 c.-167G>A NM_005994.3
TBX22 ChrX:79270181 NM_001109878.1 rs1051260152
TBX22 ChrX:79278546 c.176-13C>A NM_016954.2 rs55760411
TXNL4A Chr18:77748580 c.-60-10913_-60-10880del34 NM_001305563.1 rs535089924
TXNL4A Chr18:77748603 c.-60-10936_-60-10903del34 NM_001305563.1 rs786205699

Added and removed genes from the panel

Genes added Genes removed
ARHGAP29
CTNND1
SPECC1L
TBX2
TXNL4A
ZSWIM6

Test Strengths

The strengths of this test include:
  • CAP accredited laboratory
  • CLIA-certified personnel performing clinical testing in a CLIA-certified laboratory
  • Powerful sequencing technologies, advanced target enrichment methods and precision bioinformatics pipelines ensure superior analytical performance
  • Careful construction of clinically effective and scientifically justified gene panels
  • Our Nucleus online portal providing transparent and easy access to quality and performance data at the patient level
  • Our publicly available analytic validation demonstrating complete details of test performance
  • ~2,000 non-coding disease causing variants in our clinical grade NGS assay for panels (please see ‘Non-coding disease causing variants covered by this panel’ in the Panel Content section)
  • Our rigorous variant classification scheme
  • Our systematic clinical interpretation workflow using proprietary software enabling accurate and traceable processing of NGS data
  • Our comprehensive clinical statements

Test Limitations

Genes with partial, or whole gene, segmental duplications in the human genome are marked with an asterisk (*) if they overlap with the UCSC pseudogene regions. The technology may have limited sensitivity to detect variants in genes marked with these symbols (please see the Panel content table above).

This test does not detect the following:
  • Complex inversions
  • Gene conversions
  • Balanced translocations
  • Mitochondrial DNA variants
  • Repeat expansion disorders unless specifically mentioned
  • Non-coding variants deeper than ±20 base pairs from exon-intron boundary unless otherwise indicated (please see above Panel Content / non-coding variants covered by the panel).
This test may not reliably detect the following:
  • Low level mosaicism (variant with a minor allele fraction of 14.6% is detected with 90% probability)
  • Stretches of mononucleotide repeats
  • Indels larger than 50bp
  • Single exon deletions or duplications
  • Variants within pseudogene regions/duplicated segments

The sensitivity of this test may be reduced if DNA is extracted by a laboratory other than Blueprint Genetics.

For additional information, please refer to the Test performance section and see our Analytic Validation.

The genes on the panel have been carefully selected based on scientific literature, mutation databases and our experience.

Our panels are sectioned from our high-quality, clinical grade NGS assay. Please see our sequencing and detection performance table for details regarding our ability to detect different types of alterations (Table).

Assays have been validated for various sample types including EDTA-blood, isolated DNA (excluding from formalin fixed paraffin embedded tissue), saliva and dry blood spots (filter cards). These sample types were selected in order to maximize the likelihood for high-quality DNA yield. The diagnostic yield varies depending on the assay used, referring healthcare professional, hospital and country. Plus analysis increases the likelihood of finding a genetic diagnosis for your patient, as large deletions and duplications cannot be detected using sequence analysis alone. Blueprint Genetics’ Plus Analysis is a combination of both sequencing and deletion/duplication (copy number variant (CNV)) analysis.

Performance of Blueprint Genetics high-quality, clinical grade NGS sequencing assay for panels.

Sensitivity % (TP/(TP+FN) Specificity %
Single nucleotide variants 99.89% (99,153/99,266) >99.9999%
Insertions, deletions and indels by sequence analysis
1-10 bps 99.2% (7,745/7,806) >99.9999%
11-50 bps 99.13% (2,524/2,546) >99.9999%
Copy number variants (exon level dels/dups)
1 exon level deletion (heterozygous) 100% (20/20) NA
1 exon level deletion (homozygous) 100% (5/5) NA
1 exon level deletion (het or homo) 100% (25/25) NA
2-7 exon level deletion (het or homo) 100% (44/44) NA
1-9 exon level duplication (het or homo) 75% (6/8) NA
Simulated CNV detection
5 exons level deletion/duplication 98.7% 100.00%
     
Size range (0.1-47 Mb) 100% (25/25)
     
The performance presented above reached by Blueprint Genetics high-quality, clinical grade NGS sequencing assay with the following coverage metrics
     
Mean sequencing depth 143X
Nucleotides with >20x sequencing coverage (%) 99.86%

Bioinformatics

The target region for each gene includes coding exons and ±20 base pairs from the exon-intron boundary. In addition, the panel includes non-coding and regulatory variants if listed above (Non-coding variants covered by the panel). Some regions of the gene(s) may be removed from the panel if specifically mentioned in the ‘Test limitations” section above. The sequencing data generated in our laboratory is analyzed with our proprietary data analysis and annotation pipeline, integrating state-of-the art algorithms and industry-standard software solutions. Incorporation of rigorous quality control steps throughout the workflow of the pipeline ensures the consistency, validity and accuracy of results. Our pipeline is streamlined to maximize sensitivity without sacrificing specificity. We have incorporated a number of reference population databases and mutation databases including, but not limited, to 1000 Genomes Project, gnomAD, ClinVar and HGMD into our clinical interpretation software to make the process effective and efficient. For missense variants, in silico variant prediction tools such as  SIFT, PolyPhen, MutationTaster are used to assist with variant classification. Through our online ordering and statement reporting system, Nucleus, ordering providers have access to the details of the analysis, including patient specific sequencing metrics, a gene level coverage plot and a list of regions with <20X sequencing depth if applicable. This reflects our mission to build fully transparent diagnostics where ordering providers can easily visualize the crucial details of the analysis process.

Clinical interpretation

We provide customers with the most comprehensive clinical report available on the market. Clinical interpretation requires a fundamental understanding of clinical genetics and genetic principles. At Blueprint Genetics, our PhD molecular geneticists, medical geneticists and clinical consultants prepare the clinical statement together by evaluating the identified variants in the context of the phenotypic information provided in the requisition form. Our goal is to provide clinically meaningful statements that are understandable for all medical professionals regardless of whether they have formal training in genetics.

Variant classification is the corner stone of clinical interpretation and resulting patient management decisions. Our classifications follow the Blueprint Genetics Variant Classification Schemes based on the ACMG guideline 2015. Minor modifications were made to increase reproducibility of the variant classification and improve the clinical validity of the report. Our experience with tens of thousands of clinical cases analyzed at our laboratory allowed us to further develop the industry standard.

The final step in the analysis is orthogonal confirmation. Sequence variants classified as pathogenic, likely pathogenic and variants of uncertain significance (VUS) are confirmed using bi-directional Sanger sequencing when they do not meet our stringent NGS quality metrics for a true positive call.
Reported heterozygous and homo/hemizygous copy number variations with a size <10 and <3 target exons are confirmed by orthogonal methods such as qPCR if the specific CNV has been seen and confirmed less than three times at Blueprint Genetics.

Our clinical statement includes tables for sequencing and copy number variants that include basic variant information (genomic coordinates, HGVS nomenclature, zygosity, allele frequencies, in silico predictions, OMIM phenotypes and classification of the variant). In addition, the statement includes detailed descriptions of the variant, gene and phenotype(s) including the role of the specific gene in human disease, the mutation profile, information about the gene’s variation in population cohorts and detailed information about related phenotypes. We also provide links to the references, abstracts and variant databases used to help ordering providers further evaluate the reported findings if desired. The conclusion summarizes all of the existing information and provides our rationale for the classification of the variant.

Identification of pathogenic or likely pathogenic variants in dominant disorders or their combinations in different alleles in recessive disorders are considered molecular confirmation of the clinical diagnosis. In these cases, family member testing can be used for risk stratification. We do not recommend using variants of uncertain significance (VUS) for family member risk stratification or patient management. Genetic counseling is recommended.

Our interpretation team analyzes millions of variants from thousands of individuals with rare diseases. Our internal database and our understanding of variants and related phenotypes increases with every case analyzed. Our laboratory is therefore well-positioned to re-classify previously reported variants as new information becomes available. If a variant previously reported by Blueprint Genetics is re-classified, our laboratory will issue a follow-up statement to the original ordering health care provider at no additional cost.

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