Long-read sequencing resolves complex CYP21A2 variants and identifies 2+0 carriers in 21-hydroxylase deficiency.

CYP21A2

Complex CYP21A2 structural variants arising from homology with the CYP21A1P pseudogene — deletions, duplications, chimeras and "2+0" genotypes resolved by long-read sequencing

Molecular diagnosis of 21-hydroxylase deficiency is complicated by the high homology between CYP21A2 and its pseudogene CYP21A1P. Twenty suspected patients with structural variants flagged on initial MLPA screening were reanalysed with CNVplex and SNaPshot, then with long-read sequencing. Long-read reliably characterised a novel large deletion and defined its boundaries, identified "2+0" carriers — a deletion masked by a duplication, a source of false negatives with standard techniques — and separated the CYP21A1P/CYP21A2 CH-4 and CH-9 chimera subtypes that the combined conventional assays could not tell apart. It also precisely characterised TNXA/TNXB chimeric deletions, frequently misclassified as CYP21A1P/CYP21A2 chimeras although they drive the diagnosis of CAH-X syndrome.

Synthesis written by Geno'X. For the full original abstract, please refer to the source publication.

What matters for genetic counselling is the "2+0" genotype: a relative falsely reassured about carrier status means a misestimated recurrence risk, and 21-hydroxylase deficiency is common enough for this to be encountered. The study covers twenty patients selected for their structural variants, which gives no estimate of how frequent these situations are in the general population. Long-read sequencing belongs here as a second-line test on discordant or incomplete genotypes, rather than up front.

Analysis by Dr Thibaut Benquey