Clinical Information

Next-generation sequencing is a methodology that can interrogate large regions of genomic DNA in a single assay. The presence and pattern of gene variants can provide critical diagnostic, prognostic, and therapeutic information for managing physicians.

Erythrocytosis (ie, increased red blood cell [RBC] mass or polycythemia) may be primary, due to an intrinsic defect of bone marrow stem cells (ie, polycythemia vera: PV), or secondary, in response to increased serum erythropoietin (EPO) levels. Secondary erythrocytosis is associated with a number of disorders including chronic lung disease, chronic increase in carbon monoxide (due to smoking), cyanotic heart disease, high-altitude living, renal cysts and tumors, hepatoma, and other EPO-secreting tumors. When these common causes of secondary erythrocytosis are excluded, a heritable cause involving hemoglobin or erythrocyte regulatory mechanisms may be suspected.

Unlike polycythemia vera, hereditary erythrocytosis is not associated with the risk of clonal evolution and should present with isolated erythrocytosis that has been present since birth. A small subset of cases is associated with neoplasia (ie, pheochromocytoma and/or paraganglioma formation). It is caused by variations in several genes and may be inherited in either an autosomal dominant or autosomal recessive manner. A family history of erythrocytosis would be expected in these cases, although it is possible for new variants to arise in an individual.

The genes coding for hemoglobin, beta globin and alpha globin (high-oxygen-affinity hemoglobin variants), hemoglobin-stabilization proteins (2,3 bisphosphoglycerate mutase: BPGM), and the erythropoietin receptor (EPOR) and oxygen-sensing pathway enzymes (hypoxia-inducible factor [HIF2A/EPAS1], prolyl hydroxylase domain 2 [PHD2/EGLN1], and von Hippel Lindau [VHL]) can result in hereditary erythrocytosis. High-oxygen-affinity hemoglobin variants and BPGM abnormalities result in a decreased p50 result, whereas those affecting EPOR, HIF2A, PHD2, and VHL have normal p50 results. The true prevalence of hereditary erythrocytosis-causing variants is unknown. Due to high homology, hemoglobin genes, HBA1/HBA2 and HBB, are not interrogated in this panel.

The oxygen-sensing pathway functions through an enzyme, HIF, which regulates RBC mass. A heterodimer protein comprised of alpha and beta subunits, HIF functions as a marker of depleted oxygen concentration. When present, oxygen becomes a substrate mediating HIF-alpha subunit degradation. In the absence of oxygen, degradation does not take place, and the alpha protein component is available to dimerize with a HIF-beta subunit. The heterodimer then induces transcription of many hypoxia response genes including EPO. HIF-alpha is regulated by VHL protein-mediated ubiquitination and proteasomal degradation, which requires prolyl hydroxylation of HIF proline residues. The HIF-alpha subunit is encoded by the HIF2A (EPAS1) gene. Enzymes important in the hydroxylation of HIF-alpha are the prolyl hydroxylase domain proteins, of which the most significant isoform is PHD2, which is encoded by the PHD2 (EGLN1) gene. Variations resulting in altered HIF-alpha, PHD2, and VHL proteins can lead to clinical erythrocytosis. A small subset of variants, in PHD2/EGLN1 and HIF2A/EPAS1, have also been detected in erythrocytic patients presenting with paragangliomas or pheochromocytomas. Truncating variants in the EPOR gene coding for the erythropoietin receptor can result in erythrocytosis through loss of the negative regulatory cytoplasmic SHP-1 binding domain leading to EPO hypersensitivity. All currently known variants have been localized to exon 8 and are heterozygous truncating variants. EPOR variants are associated with decreased EPO levels and normal p50 values. Gain of function variants in EPO have also been associated with hereditary erythrocytosis.