Complex interaction of δ-globin variant HbA2-Famagusta combined with β-globin variant Hb Monroe in cis with the HBB:c.-92C>G mutation: hematological characteristics and molecular diagnostic insights.

Background: Hemoglobin (Hb) variants can cause clinical conditions like thalassemia. Understanding their molecular phenotypes and clinical profiles is essential for accurate diagnosis and genetic counseling.

Research design and methods: Blood samples from a 67-year-old Thai female and four relatives were analyzed using HPLC and capillary electrophoresis. PCR techniques and sequencing were used to identify globin gene mutations and β-globin haplotypes. Pathogenicity and transcription factor-binding sites were predicted. A rapid diagnostic method was developed and validated using 1,414 samples to assess the geographic distribution of the variants.

Results: Hb analysis revealed HbA, HbA₂, and a slower-migrating Hb fraction. Mutations identified included HBB:c.92 G > C(HbMonroe) in cis with HBB:c.-92C > G, and HBD:c.60C > A(HbA₂-Famagusta). This novel combination exhibited normal HbA₂ levels, potentially masking β-thalassemia. Combining HbMonroe with HbE leads to severe clinical symptoms. HbA₂-Famagusta was predicted to be benign, while HbMonroe was deleterious. The geographic distribution of HbMonroe was found to be 0.09% with an allele frequency of 0.00042 and 1.30% with an allele frequency of 0.00649 in β-thalassemia carriers and EF syndrome, respectively.

Conclusions: HbMonroe is a β⁰-thalassemic variant affecting splicing and structure, undetectable by standard Hb analysis. Including HbA₂ variants in total HbA₂ quantification is essential for accurate diagnosis and improved genetic counseling in thalassemia-endemic areas.