A Novel Plasma Heme Assay Reveals Disease Severity in Beta‐Thalassemia and Sickle Cell Anemia

Anemia results from imbalanced hemoglobin or red blood cell production and clearance. Hemolytic anemia, caused by premature red blood cell removal, can be intravascular (in blood) or extravascular (erythrophagocytosis). Hemolysis is common in Sickle Cell Disease (SCD) and Beta-Thalassemia anemia (β-thalassemia), the most prevalent inherited hemolytic anemias. Hemolysis severity is primarily assessed by measuring indirect serum biomarkers such as lactate dehydrogenase, released by cytolysis, bilirubin and haptoglobin. However, these markers do not directly indicate either the cause or the primary site of hemolysis. We introduced a novel plasma heme assay that quantifies all heme-related species in plasma, including hemoglobin, methemoglobin, heme, and hemopexin. Our findings revealed a more profound intravascular red blood cell destruction in SCD compared to β-thalassemia as demonstrated by higher values of plasma hemoglobin, respectively 6.20 and 2.52 μM (p < 0.001), with significant inter-individual variability. In contrast, β-thalassemia patients exhibited higher plasma heme values (11.00 μM vs. 1.51 μM; p < 0.0001) reflecting a probable mixed origin (dyserythropoiesis and hemolysis). Plasma hemopexin was negatively correlated with plasma heme in all patients. Plasma heme exceeded hemopexin scavenging capability in 72% of β-thalassemia and 36% of SCD patients. In β-thalassemia, plasma heme levels were significantly higher in transfusion-dependent compared to non-transfusion-dependent patients, indicating that excess heme reflects clinical severity. In SCD, elevated concentration of excess heme was associated with a significant increased risk of mortality compared to LDH or reticulocytes%. This novel spectral assay offered significant benefits for diagnosis, treatment, and patient management.