Genetic Markers of Quantitative Trait Loci for Phenotypic Manifestation of Thalassemia Major Disease

The genetic predisposition of fetal hemoglobin and modifier genes responsible for regulating the balance of alpha/beta globin chains required to improve the phenotype of beta thalassemia. This condition can prevent the severe phenotype of the disease to occur. Subsequently, β-thalassemia patients presented with diverse clinical manifestation ranging from the severe form of transfusion-dependent thalassemia major to non-transfusion dependent form of asymptomatic thalassemia disease. It was already acknowledged that TM patients represented with higher levels of fetal hemoglobin [HbF] are those patients in which less severe disease phenotype were noted. Currently, the main attention is on secondary & tertiary genetic modifiers factors that can prove to show betterment in the clinical phenotype of β-thalassemia syndromes. The most common quantitative trait loci (QTLs) known for HbF are variations on genes like: B-cell lymphoma/leukemia 11A (BCL11A), HBS1L-MYB intergenic region and Krueppel-like factor 1 (KLF1). To understand the phenotypic variations in thalassemia it is necessary to be looked into the responsible genes and their genetic modifiers factors regulating the balance of the alpha –beta globin gene clusters.

Keywords: Thalassemia, gene, diagnosis, fetal-hemoglobin

Beta thalassemia (BT) is an inherited, autosomal recessive, monogenic genetic disorder, BT mainly caused by a reduced or absent synthesis of one or more of the alpha or beta globin chains. Mainly two forms of thalassemia are well-known i.e., alpha and beta thalassemias which are mainly caused by either reduction or complete absence of the synthesis of either alpha or beta globin chains respectively. The severe complications of iron overload, together with the sequels of the anemia and ineffective erythropoiesis, and the chelation therapy are major reasons of morbidity and mortality in patients of transfusion dependent β-thalassemias. It is mostly prevalent in populations of regions the Mediterranean, Middle East, Transcaucasus, Central Asia, Indian subcontinent, and Far East. Beta thalassemia (β-thal) disease is caused by mutation heamoglobin (HBB) gene, till date more than 400-point mutations are reported that ultimately results in various clinical conditions of Thalassemia [1].

Potential genetic factors that influence the severity of anemia in thalassemia patients probably will be inherited or non-inherited genetic factors. The most commonly known inherited genetic factors considered for the different forms of β-thalassemia are coinheritance of alpha thalassemia (α thalassemia) with mutation in HBB gene and other genetic factors outside of beta globin gene that stimulate and play important role in production of fetal hemoglobin (HbF). [2, 3].

HbF is the dominant hemoglobin found especially high in the fetus. After birth, HbF is nearly or totally replaced or switch over by HbA and finally the “adult” levels of HbA are nearly attained by the age of 6 months. In some cases of the β hemoglobinopathies, in patients at the age of 5 years stable adult levels of HbA are not still produced. The molecular mechanism of switching from HbF to HbA comprises repression of HBG2 and HBG1 followed by an up-regulation of HBB expression [4, 6]. This process of switch over is never totally ceases the production of HBF and some clones of erythroid precursors are still continuing to make progeny that are enough for expression of the HbF genes. Usually less than 1 % HBF; a very less amounts of HbF still express in normal adults [3].

HbF gene expression includes interaction and binding of many different proteins on promoter of gene, which includes the transcription factors (TF) KLF1 (Krueppel-like factor 1) and BCL11A (B-cell lymphoma/leukemia 11A), and the hematopoietic regulatory factor MYB and other co-repressor complexes that implicate chromatin-modeling and epigenetic modifiers [4]. As KLF1 is a known transcription regulator of BCL11A and globin switching, polymorphisms in these genes might be associated with HbF levels [17]. In beta-thalassemia, patients who have higher levels of fetal hemoglobin have shown to observe less severe disease.

Figure 1: Gene regulating Hb switching

The most important QTLs (quantitative trait loci) (modifier genes) for HbF and F-cells that have been widely researched and studied are shown in Figure1 and mostly reported to be found in three loci, one is in-cis with the β-globin gene cluster and two are in-trans with the β-globin gene cluster, the others include HBS1L-MYB intergenic region and BCL11A gene. Any deletions in BCL11A-binding motifs were believed to be associated with increased HbF levels as BCL11A plays an important role in heamoglobin switching [5, 7]. BCL11A mainly binds to the upstream locus control region (LCR), γ-globin and the intergenic regions present between β-globin and γ- globin genes, working together with SOX6 gene that is well known for reconfiguration of the β-globin cluster by selectively modulating chromosomal loop formation, which ultimately leads to transcriptional silencing of the γ-globin genes [16]. Numerous studies have verified that patients presented with sequence variations in the BCL11A or HSB1L-MYB genes have higher levels of fetal heamoglobin and less severe beta-thalassemia [8].

To better understand the clinical or molecular interactions, it is essential to know or investigate any factor which play important role in of reducing the alpha/non alpha globin chain imbalance and may show an ameliorating effect on the clinical picture of the disease [9]. Also, some of the reported mutations present within the β-globin promoter region are observed to be associated with increased γ-globin chain expression. These are mainly known as primary modifiers. Presently, the more focus shifts towards the research on other genetic variants that modulate HbF levels, but located outside of the HBB genes (known as Secondary modifiers) have also been reported to play center role. These modifiers commonly called as secondary modifiers; they primarily act directly to modify the known pathophysiology of the disease [10].

Secondary modifiers comprise of alterations in the genes affecting α/β globin chain balance such results in α/γ-globin genes expression, besides this more research is required on those genes which are involved in the γ-globin gene expression (HBS1MYB, BCL11A,) [11]. In the upcoming years, there has been significant advancement in the fields research on secondary genetic modifiers that were found to ameliorate the clinical phenotype of β-thalassemia disease. Absolutely, any increase in the production of fetal hemoglobin (HbF) throughout adulthood period could ameliorate the severity of β-thalassemia phenotype; as γ-globin polypeptide chains pay off the role in place of the functional β-globin polypeptide chains [10, 12]. Therefore, γ-globin genes, together with other secondary modifiers, signified as the most common targets for current developmental therapeutic strategies. Individually, BCL11A loci account for approximately 50% of the HbF level variation, indicative of fact that other factors are also involved [9].

In northern Europeans countries it was reported that presence of variants within HBS1LMYB intragenic region, located on chromosome 6q23, reason for more than 20% of the increased HbF level [13]. Hematopoiesis is up-regulated by MYB gene and accountable for modulation in HbF expression indirectly through modification of the kinetics of erythroid differentiation; also, directly via activation of KLF1 and other repressors. On inducing reduction in MYB levels erythroid differentiation is enhanced leading to release of early erythroid progenitor cells that are still synthesizing predominantly HbF [14, 15].

Fetal hemoglobin (HbF) is regulated by multigenic traits or factors. The presence of HBS1L-MYB intergenic polymorphism (HMIP) on chromosome 6q23 and the polymorphism on BCL11 A gene located on chromosome 2p16.5 that can modify HbF expression and ameliorate disease severity in β-thalassemia, sickle cell anemia. Among different populations the relative contributions of these polymorphisms to HbF regulation changes variably. Genetic heterogeneity related to fetal hemoglobin (HbF) production has been reported as an influencing phenotypic factor of β-thalassemia (β-thal) so it is current demand to do more research in this direction.