Genotype-Phenotype Correlation of Seven Known and Novel β-Globin Gene Variants

Variants of uncertain significance (VUS) are often challenging for genetic counseling and require additional data for accurate variant classification. This study aims to describe the genotype-phenotype correlation of the seven β-globin gene variants found in Thailand. Retrospective data in a total of 45,914 subjects encountered at our diagnostic laboratory from January 2012 to December 2024 were reviewed. A total of 33 leftover EDTA blood specimens, suspected of having β-globin gene defects, were included. Eighty-nine normal subjects were also analyzed to confirm phenotypic expression of the variants. The whole β-globin and Krüppel-like factor 1 (KLF1) genes were examined using PCR-based methods. Seven nucleotide variants were identified among 33 suspected subjects, including a novel (β^-206(C>G)), four hitherto undescribed in Thailand [β^-198(A>G), β^{IVSII-180(T>C)}, β^{IVSII-337(A>G)}, and β^*233(G>C)], and two known variants [β^-50(G>A) and β^{IVSII-258(G>A)}]. The β^-198(A>G) and β^*233(G>C) variants were also identified in 1.69% of normal subjects, indicating neutral DNA polymorphisms. All subjects of β^-198(A>G), β^{IVSII-180(T>C)}, β^{IVSII-258(G>A)}, and β^{IVSII-337(A>G)} with borderline Hb A₂ levels had KLF1 mutations. Compound heterozygous β^-206(C>G) and known β⁺-thalassemia trait revealed β-thalassemia trait phenotype. In silico pathogenicity prediction showed that the β^-206(C>G), β^-198(A>G), β^{IVSII-180(T>C)}, β^{IVSII-258(G>A)}, β^{IVSII-337(A>G)}, and β^*233(G>C) were associated with benign variants. It was found that heterozygous β^-50(G>A) had elevated Hb A₂ levels resembling those of β-thalassemia trait. However, the association of the β^-50(G>A) and Hb E or β-thalassemia revealed a phenotype of Hb E or β-thalassemia trait. Most prediction tools indicate that the β^-50(G>A) is associated with benign variants; however, PromoterAI revealed that the β^-50(G>A) is associated with under-expression of the β-globin gene with high sensitivity. Based on these findings, the β^-50(G>A) is most likely a very mild β⁺-thalassemia allele. This study described the genotype-phenotype correlation of known and novel β-globin gene variants found in Thailand. The data should prove useful for accurate variant classification, genetic counseling, and a prevention and control program of severe thalassemia diseases in Thailand.