Abnormal Genitalia/ Disorders of Sex Development Panel
Updated

Last modified: Mar 21, 2018

Summary

  • Is a 49 gene panel that includes assessment of non-coding variants
  • Is ideal for patients presenting with ambiguous genitalia, patients suspected to have a disorder of sexual development and patients suspected to have congenital adrenal hyperplasia (CAH).

Analysis methods

  • PLUS
  • SEQ
  • DEL/DUP

Availability

3-4 weeks

Number of genes

49

Test code

EN0201

CPT codes

SEQ 81405
SEQ 81406
SEQ 81407
DEL/DUP 81479

Summary

The Blueprint Genetics Abnormal Genitalia/ Disorders of Sex Development Panel (test code EN0201):

  • Is a 49 gene panel that includes assessment of selected non-coding disease-causing variants
  • Is available as PLUS analysis (sequencing analysis and deletion/duplication analysis), sequencing analysis only or deletion/duplication analysis only

ICD codes

Commonly used ICD-10 code(s) when ordering the Abnormal Genitalia/ Disorders of Sex Development Panel

ICD-10 Disease
Q56.0 Indeterminate sex and pseudohermaphroditism
Q56.1 Male pseudohermaphroditism
Q56.2 Female pseudohermaphroditism
E34.50 Androgen insensitivity syndrome
E25.0 Congenital adrenal hyperplasia

Sample Requirements

  • EDTA blood, min. 1 ml
  • Purified DNA, min. 3μg
  • Saliva (Oragene DNA OG-500 kit)

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.

Disorders of sex development (DSD) are a group of congenital conditions characterized by problems in the course of gender patterning, gonadal and sex development. It has been estimated that 1% – 2% of live births have some aspect of DSD. Approximately 5% of infants with DSD have ambiguous genitalia and indeterminate sex at birth. However, the vast majority of these patients do not require corrective surgery. Patients with 46,XY DSD have often impaired androgen synthesis or action and may have normal female external genitalia, while patients with 46,XX DSD conditions have often androgen excess. In 46,XX females, congenital adrenal hyperplasia (CAH) caused by 21-hydroxylase deficiency (21-OHD) is the most common cause of DSD. The estimated prevalence of CAH is 1:10,000 and 90%-95% of cases are due to mutations in CYP21A2. The severity of the condition often depends on the residual enzyme activity subdiving CYP21A2 mutations in severe (classic phenotype, enzyme activity 0%-10%) and mild (non-classic, enzyme activity 20%-50%). Androgen insensitivity syndrome (AIS), caused by mutations in AR, is characterized by feminization of external genitalia and atypical sexual development in 46,XY individuals. The condition may be complete, partial or mild, depending on the level of androgen insensitivity. Mutations in the AR gene explain up to 95% of cases with complete androgen insensitivity, while the proportions are lower for the partial and mild subtypes. The combined prevalence of various AIS subtypes is estimated to be 5:100,000.

Genes in the Abnormal Genitalia/ Disorders of Sex Development Panel and their clinical significance

Gene Associated phenotypes Inheritance ClinVar HGMD
AMH Persistent Mullerian duct syndrome AR 5 54
AMHR2 Persistent Mullerian duct syndrome AR 6 33
ANOS1* Kallmann syndrome XL/Digenic 30 175
AR Androgen insensitivity XL 93 582
ARX Lissencephaly, Epileptic encephalopathy, Corpus callosum, agenesis of, with abnormal genitalia, Partington syndrome, Proud syndrome, Hydranencephaly with abnormal genitalia, Mental retardation XL 64 85
ATRX Carpenter-Waziri syndrome, Alpha-thalassemia/mental retardation syndrome, Holmes-Gang syndrome, Juberg-Marsidi syndrome, Smith-Fineman-Myers syndrome, Mental retardation-hypotonic facies syndrome XL 54 155
BCOR Microphthalmia, syndromic, Oculofaciocardiodental syndrome XL 29 48
CDKN1C Beckwith-Wiedemann syndrome, IMAGE syndrome AD 24 81
CEP41 Joubert syndrome AR/Digenic 7 10
CHD7 Isolated gonadotropin-releasing hormone deficiency, CHARGE syndrome AD 192 784
CREBBP Rubinstein-Taybi syndrome AD 138 340
CYP11A1 Adrenal insufficiency, congenital, with 46,XY sex reversal, partial or complete AD/AR 14 25
CYP11B1* Adrenal hyperplasia, congenital, due to 11-beta-hydroxylase deficiency, Glucocorticoid-remediable aldosteronism AD/AR 24 132
CYP17A1 Adrenal hyperplasia, congenital, due to 17-alpha-hydroxylase deficiency AR 35 120
CYP19A1 Aromatase deficiency AR 16 51
CYP21A2* Adrenal hyperplasia, congenital, due to 21-hydroxylase deficiency, Hyperandrogenism, nonclassic , due to 21-hydroxylase deficiency AR 45 299
DHCR7 Smith-Lemli-Opitz syndrome AR 64 214
DHH 46,XY partial gonadal dysgenesis, with minifascicular neuropathy, 46,XY sex reversal 7 AD/AR 4 16
DYNC2H1 Short -rib thoracic dysplasia with or without polydactyly type 1, Short -rib thoracic dysplasia with or without polydactyly type 3, Asphyxiating thoracic dysplasia (ATD; Jeune), SRPS type 2 (Majewski) AR/Digenic 46 101
ERCC3 Xeroderma pigmentosum, Trichothiodystrophy, photosensitive AR 10 17
FGF8 Hypogonadotropic hypogonadism AD/Digenic 12 28
FGFR1 Pfeiffer syndrome, Trigonocephaly, Hypogonadotropic hypogonadism, Osteoglophonic Dwarfism - Craniostenosis, Hartsfield syndrome AD/Digenic/Multigenic 61 239
FIG4 Amyotrophic lateral sclerosis, Polymicrogyria, bilateral occipital, Yunis-Varon syndrome, Charcot-Marie-Tooth disease AD/AR 26 58
FRAS1 Fraser syndrome AR 24 51
GATA4 Tetralogy of Fallot, Atrioventricular septal defect, Testicular anomalies with or without congenital heart disease, Ventricular septal defect, Atrial septal defect AD 36 120
GNRHR Hypogonadotropic hypogonadism AD/AR/Digenic 23 55
HSD3B2 3-beta-hydroxysteroid dehydrogenase, II deficiency AR 10 63
HSD17B3 17-Beta hydroxysteroid dehydrogenase III deficiency AR 15 58
IL17RD Hypogonadotropic hypogonadism AD/Digenic 5 8
IRF6 Orofacial cleft, Popliteal pterygium syndrome, van der Woude syndrome AD 36 336
KISS1R Precocious puberty, central 1 AR 7 35
LHCGR Precocious puberty, male, Leydig cell hypoplasia, Luteinizing hormone resistance, female AR 30 69
MAMLD1 Hypospadias 2, X-linked XL 5 19
MAP3K1 46,XY sex reversal 6 AD 8 20
MKRN3 Central precocious puberty AD 5 29
MKS1 Bardet-Biedl syndrome, Meckel syndrome AR 42 51
NR5A1 Adrenocortical insufficiency, Premature ovarian failure, 46,XY sex reversal AD/AR 26 165
NR0B1 Adrenal hypoplasia, congenital, 46,XY sex reversal XL 59 247
POR Disordered steroidogenesis due to cytochrome p450 oxidoreductase deficiency, Antley-Bixler syndrome AR 12 67
PROK2 Hypogonadism, hypogonadotropic, Kallmann syndrome AD/AR 7 20
PROKR2 Hypogonadotropic hypogonadism AD/AR 9 51
RSPO1 Palmoplantar hyperkeratosis with squamous cell carcinoma of skin and 46,XX sex reversal AR 3 3
SOX9 Campomelic dysplasia, 46,XY sex reversal, Brachydactyly with anonychia (Cooks syndrome) AD 34 139
SRD5A2 Steroid 5-alpha-reductase 2 deficiency AR 24 119
SRY 46,XX disorder of sex development, 46,XY disorder of sex development YL 24 106
STAR Lipoid adrenal hyperplasia AR 16 81
TACR3 Hypogonadotropic hypogonadism AR 8 32
WT1 Denys-Drash syndrome, Frasier syndrome, Wilms tumor AD 29 172
ZFPM2 46,XY sex reversal AD 8 44

* 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).

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 the panel

Gene Genomic location HG19 HGVS RefSeq RS-number
AMHR2 Chr12:53819426 c.622-47C>T NM_020547.2 rs200782636
AR ChrX:66764442 c.-547C>T NM_000044.3
AR ChrX:66942551 c.2450-118A>G NM_000044.3
CDKN1C Chr11:2905209 c.*5+20G>T NM_000076.2 rs760540648
CHD7 Chr8:61734568 c.2836-15C>G NM_017780.3
CHD7 Chr8:61757794 c.5051-15T>A NM_017780.3
CHD7 Chr8:61763035 c.5405-17G>A NM_017780.3 rs794727423
DYNC2H1 Chr11:103019205 c.2819-14A>G NM_001080463.1 rs781091611
GATA4 Chr8:11560864 c.-1407C>A NM_002052.3 rs769262495
GATA4 Chr8:11560787 c.-1484T>C NM_002052.3 rs372004083
GATA4 Chr8:11561369 c.-902G>T NM_002052.3
GATA4 Chr8:11561282 c.-989C>T NM_002052.3
IRF6 Chr1:209979367 c.-151G>A NM_006147.3
IRF6 Chr1:209975332 c.-19C>A NM_006147.3
IRF6 Chr1:209979435 c.-219C>T NM_006147.3
IRF6 Chr1:209975361 c.-48A>T NM_006147.3
POR Chr7:75544501 c.-5+4A>G NM_000941.2
STAR Chr8:38003676 c.466-11T>A NM_000349.2

Added and removed genes from the panel

Genes added Genes removed
CYP11A1
DHH
ERCC3
FGF8
FGFR1
KISS1R
MAMLD1
MAP3K1
MKRN3
PROK2

Test strength

The strengths of this test include:
  • CAP and ISO-15189 accreditations covering all operations at Blueprint Genetics including all Whole Exome Sequencing, NGS panels and confirmatory testing
  • 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 publically available analytic validation demonstrating complete details of test performance
  • 1479 non-coding disease causing variants in Blueprint WES assay (please see below ‘Non-coding disease causing variants covered by this panel’)
  • Our rigorous variant classification based on modified ACMG 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

Although the CYP21A2 and CYP11B1 genes have segmentally duplicated pseudogenes that reduce sensitivity of NGS diagnostics in general, Blueprint Genetics custom assay has good coverage (>20x) with high mapping rates (mapping quality >40) for target regions of these genes: CYP21A2 98.6% and CYP11B1 100.0%. Our validation showed mean coverage of 105x for CYP21A2 gene and 244x for CYP11B1. Thus, our NGS Panel is not expected to have major limitations in detecting sequence variants in these genes. 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
  • 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 Blueprint Genetics abnormal genitalia/ disorders of sex development panel covers classical genes associated with indeterminate sex and pseudohermaphroditism, male pseudohermaphroditism, female pseudohermaphroditism, Androgen insensitivity syndrome, Persistent Mullerian duct syndrome and congenital adrenal hyperplasia. The genes on the panel have been carefully selected based on scientific literature, mutation databases and our experience.

Our panels are sliced from our high-quality whole exome sequencing data. Please see our sequencing and detection performance table for different types of alterations at the whole exome level (Table).

Assays have been validated for different starting materials including EDTA-blood, isolated DNA (no FFPE), saliva and dry blood spots (filter card) and all provide high-quality results. The diagnostic yield varies substantially depending on the assay used, referring healthcare professional, hospital and country. Blueprint Genetics' Plus Analysis (Seq+Del/Dup) maximizes the chance to find a molecular genetic diagnosis for your patient although Sequence Analysis or Del/Dup Analysis may be a cost-effective first line test if your patient's phenotype is suggestive of a specific mutation type.

Performance of Blueprint Genetics Whole Exome Sequencing (WES) assay. All individual panels are sliced from WES data.

Sensitivity % (TP/(TP+FN) Specificity %
Single nucleotide variants 99.65% (412,456/413,893) >99.99%
Insertions, deletions and indels by sequence analysis
1-10 bps 96.94% (17,070/17,608) >99.99%
11-50 bps 99.07% (957/966) >99.99%
Copy number variants (exon level dels/dups)
Clinical samples (small CNVs, n=52)
1 exon level deletion 92.3% (24/26) NA
2 exons level deletion/duplication 100.0% (11/11) NA
3-7 exons level deletion/duplication 93.3% (14/15) NA
Microdeletion/-duplication sdrs (large CNVs, n=37))
Size range (0.1-47 Mb) 100% (37/37)
Simulated CNV detection
2 exons level deletion/duplication 90.98% (7,357/8,086) 99.96%
5 exons level deletion/duplication 98.63% (7,975/8,086) 99.98%
     
The performance presented above reached by WES with the following coverage metrics
     
Mean sequencing depth at exome level 174x
Nucleotides with >20x sequencing coverage (%) 99.4%

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 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 such as, 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, the customer has an access to details of the analysis, including patient specific sequencing metrics, a gene level coverage plot and a list of regions with inadequate coverage if present. This reflects our mission to build fully transparent diagnostics where customers have easy access to 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 of sequence variants is confirmation of variants classified as pathogenic or likely pathogenic using bi-directional Sanger sequencing. Variant(s) fulfilling all of the following criteria are not Sanger confirmed: 1) the variant quality score is above the internal threshold for a true positive call, 2) an unambiguous IGV in-line with the variant call and 3) previous Sanger confirmation of the same variant at least three times at Blueprint Genetics. Reported variants of uncertain significance are confirmed with bi-directional Sanger sequencing only if the quality score is below our internally defined quality score for true positive call. Reported copy number variations with a size <10 exons are confirmed by orthogonal methods such as qPCR if the specific CNV has been seen 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 used, congress abstracts and mutation databases to help our customers 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 within the family. In the case of variants of uncertain significance (VUS), we do not recommend family member risk stratification based on the VUS result. Furthermore, in the case of VUS, we do not recommend the use of genetic information in patient management or genetic counseling. For eligible cases, Blueprint Genetics offers a no charge service to investigate the role of reported VUS (VUS Clarification Service).

Our interpretation team analyzes millions of variants from thousands of individuals with rare diseases. Thus, our database, and our understanding of variants and related phenotypes, is growing by leaps and bounds. 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.