Congenital Adrenal Hyperplasia (CAH) Panel

Congenital adrenal hyperplasia (CAH) is a group of autosomal recessive disorders each due to defects in a single gene involved in different steps of cortisol biosynthesis (Hannah-Shmouni et al. 2017. PubMed ID: 28476231; Merke and Bornstein. 2005. PubMed ID: 15964450). The clinical consequence of deficient cortisol biosynthesis represents a continuous phenotypic spectrum depending on causative gene, genotype, residual activity and age of presentation. Clinical features of CAH include adrenal insufficiency, genital ambiguity or disordered sex development, infertility, short stature, hypertension, and an increased risk of metabolic syndrome during adolescence and adulthood.

Over 95% of CAH cases are caused by 21- hydroxylase deficiency (21-OHD). The second most common form of CAH is due to 11-hydroxylase deficiency (11-OHD) which accounts for approximately 5% of cases. Other less common forms of CAH include 3β-hydroxysteroid dehydrogenase type 2 deficiency, 17α-hydroxylase deficiency, congenital lipoid adrenal hyperplasia, side-chain cleavage enzyme deficiency, and cytochrome P450 oxidoreductase deficiency (ORD).

CAHs are autosomal recessive disorders. The known causative genes are CYP21A2, CYP11B1, HSD3B2, CYP17A1, POR, STAR and CYP11A1. Documented pathogenic variants in these genes include various types of genetic defects, including truncating changes (nonsense, splice variants and frameshift small deletion/insertions), missense substitutions and intragenic large deletions. Complex gene rearrangements are commonly found in the CYP21A2 gene.

Over 95% of CAH cases are caused by 21- hydroxylase deficiency (21-OHD) due to genetic defects in the CYP21A2 gene. Genetic analysis of this gene is complicated by its highly similar pseudogene CYP21A1P and the nature of local surrounding sequence. This region is characterized by relatively frequent homologous recombination events due to the existence of highly paralogous pseudogenes in tandem. Consequently, the pathogenic variants of CYP21A2 include chimeric CYP21A1P/CYP21A2 genes (alternatively called 30 kb deletions in the literature), common pseudogene variants transferred via gene conversions and other rare pathogenic variants. For more details, please see the CYP21A2 single-gene test description.

The second most common form of CAH (~5% of cases) is 11-hydroxylase deficiency (11-OHD) due to genetic defects in the CYP11B1 gene. 11-hydroxylase is a P450 type I mitochondrial enzyme which is responsible for the conversion of 11-deoxycortisol to cortisol, and 11-deoxycorticosterone (DOC) to corticosterone. Due to high sequence similarity with the CYP11B2 gene, which encodes aldosterone synthase, rare chimeric CYP11B2/CYP11B1 genes can also result in CAH due to 11-OHD (Hampf et al. 2001. PubMed ID: 11549691; Duan et al. 2018. PubMed ID: 29703198).

Other less common forms of CAH include 3β-hydroxysteroid dehydrogenase type 2 (HSD3B2) deficiency, 17α-hydroxylase (CYP17A1) deficiency, congenital lipoid adrenal hyperplasia (STAR), side-chain cleavage enzyme (CYP11A1) deficiency, and cytochrome P450 oxidoreductase (POR) deficiency.

Founder pathogenic variants in different geographic regions worldwide are evident in all of these genes. See individual gene test descriptions for information on the molecular biology of gene products and spectra of pathogenic variants.

Since our test utilizes an integrative strategy via Sanger sequencing to perform a comprehensive evaluation for CYP21A2 (see Testing Strategy in the CYP21A2 test description), the detection rate of pathogenic CYP21A2 variants is expected to be approximately 98% (Krone et al. 2000. PubMed ID: 10720040; Stikkelbroeck et al. 2003. PubMed ID: 12915679; Finkielstain et al. 2011. PubMed ID: 20926536; Krone et al. 2013. PubMed ID: 23337727).

In a study of 28 patients from 24 Turkish families affected by 11-hydroxylase deficiency (26 classic and 2 late-onset), using the strategy of specifically amplifying CYP11B1 gene fragments, all patients received a molecular diagnosis (Kandemir et al. 2017. PubMed ID: 26956189). Similar clinical sensitivity was observed by Bas et al. (2018. PubMed ID: 29626607). These studies indicated intragenic large deletions and chimeric CYP11B2/CYP11B1 genes are rare. Therefore, our test is expected to be able to detect the vast majority of pathogenic variants.

Other genes in this panel have no paralogous sequences in the genome. Therefore, this panel is able to detect over 98% of pathogenic variants for congenital adrenal hyperplasia patients overall.

This panel provides full coverage of all coding exons of the genes plus 10 bases of flanking noncoding DNA in all available transcripts along with other non-coding regions in which pathogenic variants have been identified at PreventionGenetics or reported elsewhere. We define coverage as ≥20X NGS reads or Sanger sequencing. PGnome panels typically provide slightly increased coverage over the PGxome equivalent. PGnome sequencing panels have the added benefit of additional analysis and reporting of deep intronic regions (where applicable).

We utilize a long range PCR strategy to specifically amplify the CYP21A2 gene. This “one-stop” strategy is able to capture chimeric CYP21A1P/CYP21A2 genes (alternatively called 30kb deletions in the literature), common pseudogene variants transferred via gene conversions and other rare pathogenic variants. After this test, we don’t recommend additional testing (such as MLPA for deletions), except family follow-up tests. Please see details in the CYP21A2 single-gene test description.

Using the strategy of specifically amplifying CYP11B1 (to avoid the paralogous gene CYP11B2), our strategy is NOT designed to detect rare chimeric CYP11B2/CYP11B1 genes. However, our assay is expected to capture a large portion of small gene-conversion events.

LIMITATIONS OF THIS TEST: A duplicated CYP21A2 gene or a CYP21A2-like gene next to the pseudogene TNXA in the middle of the RCCX module, which is uncommon, CANNOT be detected via the current strategy (Koppens et al. 2002. PubMed ID: 12384784; Kleinle et al. 2011. PubMed ID: 19773403; Tsai et al. 2011. PubMed ID: 21324303). Therefore, test results via the current strategy should always be interpreted in context of clinical findings, family history and other laboratory data.

Our assay also will generally not detect rare chimeric CYP11B2/CYP11B1 genes.

Dependent on the sequencing backbone selected for this testing, discounted reflex testing to any other similar backbone-based test is available (i.e., PGxome panel to whole PGxome; PGnome panel to whole PGnome).