Hemoglobin profile and molecular characteristics of the complex interaction of hemoglobin Doi-Saket [α9(A7) asn > lys, HBA2:c.30C > a], a novel α2α1 hybrid globin variant, with hemoglobin E [ß26(B8) Glu > lys, HBB:c.79G > A] and deletional α+-thalassemia in a Thai family.

BACKGROUND: An increasing number of α-hemoglobin (Hb) variants is causing various clinical symptoms; therefore, accurate identification of these Hb variants is important. OBJECTIVE: This study aimed to describe the molecular and hematological characteristics of novel Hb Doi-Saket that gives rise to a typical α+-thalassemia phenotype in carriers with and without other hemoglobinopathies. MATERIALS AND METHODS: Biological samples from a proband and his family members were analyzed. Hematological profiles were analyzed using a standard automated cell counter. Hb was analyzed by capillary electrophoresis and high-performance liquid chromatography. Mutations and globin haplotype were identified by DNA analysis. Novel diagnostic tools based on allele-specific polymerase chain reaction (PCR) and PCR-restriction fragment length polymorphism were developed. RESULTS: Hb analysis showed a major abnormal Hb fraction, moving slower than HbA, and a minor Hb fraction alongside HbA2 in the proband, his father, and son. DNA analysis of the α-globin gene identified the -α3.7 deletion and in cis the C > A mutation on codon 9 of the α2α1 gene, corresponding to Hb Doi-Saket [α9(A7) Asn > Lys]. This mutation could be identified using newly developed allele-specific PCR-based assays. The Hb Doi-Saket al.lele was significantly associated with haplotype [- + M + + 0 -]. Interaction of αDoi-Saket with ßE globin chains led to a new Hb variant (HbE Doi-Saket). Phenotypic expression was clinically silent in heterozygotes and might present slight microcytosis. CONCLUSIONS: Hb Doi-Saket emphasizes a great diversity present in α-globin gene. The mutation in this family from Thailand was linked to -α3.7 and caused mild microcytosis in the carriers. The combination of this variant with deletions in α genes might cause a severe clinical phenotype. Different methods of separation can provide useful information in diagnosis, and a complete molecular approach is needed for confirmation before considering patient management.

The Hb Doi-Saket is a novel α-globin variant mutation occurring in the α2-globin gene in cis to the -α^3.7 kb chromosome.The carrier of Hb Doi-Saket may present slight microcytosis and have severe clinical entities when it interacts with deletions in α-globin genes.Hb analysis with the HPLC system could completely separate Hb Doi-Saket and its derivative from other Hbs.

Analysis of Deletional Hb H Diseases in Samples with Hb A2-Hb H and Hb A2-Hb Bart's on Capillary Electrophoresis.

The capillary electrophoresis (CE) system allows the quantification of Hb Bart's (γ4) and Hb H (ß4) that is used for screening of Hb H disease. However, Hb Bart's hydrops fetalis and Hb H are not always codetected in patients with Hb H disease. In this study, 35 samples were analyzed for the α0-thalassemia (α0-thal) [- -SEA (Southeast Asian) and - -THAI (Thailand)] deletions and the α+-thal [-α3.7 (rightward) and -α4.2 (leftward)] type deletions using real time-polymerase chain reaction (real time-PCR) with SYBR Green1 and high-resolution melting (HRM) analysis and conventional gap-PCR techniques, respectively. Results showed that 28 of 29 (96.6%) samples with the Hb A2-Hb H phenotype on CE electrophoregrams presented the genotype of - -SEA/-α3.7, while the - -SEA/-α4.2 made up the remainder. The - -SEA/-α3.7 genotype was also found in all six samples (100.0%) with Hb A2-Hb Bart's on CE electrophoregrams. Thus, for genetic counseling, prevention and control programs of Hb Bart's hydrops fetalis and Hb H disease, α-thal genotype analysis is required.