*1.5.1 Gene modification approach*

The hematopoietic stem cells of the affected individual are subjected to gene editing techniques ex vivo and then reinjected again to the patient for reconstitution [16].

To increase the production of γ-globin lentiviral vectors that express a zinc finger protein has been used in order to carry microRNAs that silence its repressors or interacts with the promoter of the γ-globin gene 80 [17].

Genome editing of the promoter of BCL11A can be accomplished by several nucleases, such as engineered zinc finger nucleases (ZFNs), transcription activatorlike effector nucleases (TALENs), and clustered regularly interspaced short palindromic repeats linked to Cas9 nucleases (CRISPR-Cas9) (**Figure 1**) [18]. Recently, it has been shown that ZFN-driven BCL11A enhancer ablation leads to increased production of HbF in erythroid progenitors derived hematopoietic stem cell from β-thalassemia patient which could be used for autologous transplantation [19]. Similarly, CRISPR-Cas9-mediated BCL11A enhancer inactivation in a human adultstage erythroid cell line can achieve the same results [20].

**Figure 1.** *Mechanism of gene editing.*

#### *1.5.2 Gene therapy*

Currently, gene therapy represents a novel therapeutic promise, after many years of extensive preclinical research for the optimization of gene transfer regimen. This is mediated through autologous transplantation of genetically modified hematopoietic stem cells, clinical trials being held worldwide have revealed that, by re-establishing effective hemoglobin production, patients may be rendered transfusion- and chelation-independent and escapes the immunological sequel that normally accompany allogeneic hematopoietic stem cell transplantation.

The approach of gene therapy has focused on two mechanisms: first, increasing the production of β-globin by the addition of a normal gene or correction of the mutated gene; and second, increasing the production of γ-globin by the addition of its gene, overexpression of its endogenous activating transcription factors, and silencing of its repressors. Studies of gene therapy have utilized mainly lentivirus vectors in experimental systems, including cultured CD34 HSCs from β-thalassemia patients and β-thalassemia mouse models. Yet the safety profile of such technologies is still uncertain [16].

Genomic editing has been demonstrated to modify the β-globin gene. Thus, TALEN-mediated gene correction has been used in induced HSCs from β-thalassemia patients [21].

#### *1.5.3 Allogeneic hematopoietic stem cell transplantation*

Currently, allogeneic stem cell transplant [allo-SCT] remains the only curative option for the majority of patients with β-thalassemia major before development of iron overload complications [22].

Patients with β-thalassemia major who have good risk features are reasonable to anticipate a greater than 90% chance of successful transplant outcome, even patients with high risk features, success rates are approaching 80%.

Challenging in allo-HSCT in high-risk patients is mainly related to graft rejection and risk of transplant-related mortality but nowadays novel modified or reduced-intensity conditioning regimens are used to improve the transplantation outcome in β-thalassemia patients with cheerful results [23].

Traditionally, completely matched human leukocyte antigen identical siblings have been used as donors, and on the other hand, matched unrelated donors have also been tried in patients with low risk. Bone marrow has been known as the preferred choice of stem cells in non-malignant hematological disorders to reduce the risk of GVHD but peripheral blood stem cell graft and cord stem cell when used have been reported to be associated with faster engraftment, lower requirement of blood product support in the peri-transplant period, and a low incidence of graft rejection in patients with low-risk [24, 25].

#### **1.6 Preventive strategies for thalassemia**

Despite the great advances in management tools used in β-thalassemia to improve the cure rate of the disease, yet the incidence rate of the disease is increasing especially in underdeveloped countries where the prevalence of consanguineous marriage is high and low level of standard and sometimes shortage in medical resources and supply that limit the early detection of carrier state. Therefore, the prevention of the homozygous state presents a big challenging issue. Prevention

**7**

**Figure 2.**

*therapeutic modalities are marked in blue.*

*Introductory Chapter: β-Thalassemia*

counseling is strongly needed.

and used by several groups [26, 27].

as well as other genetic diseases [28].

*1.6.1 Prenatal diagnosis*

*DOI: http://dx.doi.org/10.5772/intechopen.90946*

including prenatal diagnosis, carrier detections, molecular diagnosis, and genetic

Recently, prenatal diagnosis is carried out for couples at risk, either in first trimester through obtaining fetal material by chorionic villus sampling or in the

One of the main successful procedures in prenatal diagnosis is to study the fetal erythroid cells and detection of globin gene mutations. As the first primitive erythroblasts appear in embryonic bloodstream around the 4–5 weeks gestations, so obtaining fetal material by aspiration of coelomic fluid (celocentesis) followed by selection of embryo-fetal erythroid precursors by an anti-CD71 microbeads method or by direct micromanipulator pickup of the cells has been extensively improved

Nowadays, the possibility of cheaper and safer prenatal diagnosis facilities has emerged. Fetal-derived genetic material (cells or cell free DNA) can be obtained from the maternal blood and analyzed, which is considered a non-invasive maneuver with no risk of miscarriage and needs neither complicated procedures nor highly trained personnel for sampling. This allows future screening for thalassemia

Detection or exclusion of inherited fetal mutations is one of the most important approaches that focuses on detection of mutation that are absent from mother's

*Beta-thalassemia: causes, symptoms, and therapeutic modalities. Causes and symptoms are marked in red; and* 

second trimester through cordocentesis or amniocentesis.

including prenatal diagnosis, carrier detections, molecular diagnosis, and genetic counseling is strongly needed.

## *1.6.1 Prenatal diagnosis*

*Beta Thalassemia*

*1.5.2 Gene therapy*

transplantation.

is still uncertain [16].

β-thalassemia patients [21].

iron overload complications [22].

rejection in patients with low-risk [24, 25].

**1.6 Preventive strategies for thalassemia**

*1.5.3 Allogeneic hematopoietic stem cell transplantation*

Currently, gene therapy represents a novel therapeutic promise, after many years of extensive preclinical research for the optimization of gene transfer regimen. This is mediated through autologous transplantation of genetically modified hematopoietic stem cells, clinical trials being held worldwide have revealed that, by re-establishing effective hemoglobin production, patients may be rendered transfusion- and chelation-independent and escapes the immunological sequel that normally accompany allogeneic hematopoietic stem cell

The approach of gene therapy has focused on two mechanisms: first, increasing the production of β-globin by the addition of a normal gene or correction of the mutated gene; and second, increasing the production of γ-globin by the addition of its gene, overexpression of its endogenous activating transcription factors, and silencing of its repressors. Studies of gene therapy have utilized mainly lentivirus vectors in experimental systems, including cultured CD34 HSCs from β-thalassemia patients and β-thalassemia mouse models. Yet the safety profile of such technologies

Genomic editing has been demonstrated to modify the β-globin gene. Thus, TALEN-mediated gene correction has been used in induced HSCs from

Currently, allogeneic stem cell transplant [allo-SCT] remains the only curative option for the majority of patients with β-thalassemia major before development of

Patients with β-thalassemia major who have good risk features are reasonable to anticipate a greater than 90% chance of successful transplant outcome, even

Challenging in allo-HSCT in high-risk patients is mainly related to graft rejection and risk of transplant-related mortality but nowadays novel modified or reduced-intensity conditioning regimens are used to improve the transplantation

Traditionally, completely matched human leukocyte antigen identical siblings have been used as donors, and on the other hand, matched unrelated donors have also been tried in patients with low risk. Bone marrow has been known as the preferred choice of stem cells in non-malignant hematological disorders to reduce the risk of GVHD but peripheral blood stem cell graft and cord stem cell when used have been reported to be associated with faster engraftment, lower requirement of blood product support in the peri-transplant period, and a low incidence of graft

Despite the great advances in management tools used in β-thalassemia to improve the cure rate of the disease, yet the incidence rate of the disease is increasing especially in underdeveloped countries where the prevalence of consanguineous marriage is high and low level of standard and sometimes shortage in medical resources and supply that limit the early detection of carrier state. Therefore, the prevention of the homozygous state presents a big challenging issue. Prevention

patients with high risk features, success rates are approaching 80%.

outcome in β-thalassemia patients with cheerful results [23].

**6**

Recently, prenatal diagnosis is carried out for couples at risk, either in first trimester through obtaining fetal material by chorionic villus sampling or in the second trimester through cordocentesis or amniocentesis.

One of the main successful procedures in prenatal diagnosis is to study the fetal erythroid cells and detection of globin gene mutations. As the first primitive erythroblasts appear in embryonic bloodstream around the 4–5 weeks gestations, so obtaining fetal material by aspiration of coelomic fluid (celocentesis) followed by selection of embryo-fetal erythroid precursors by an anti-CD71 microbeads method or by direct micromanipulator pickup of the cells has been extensively improved and used by several groups [26, 27].

Nowadays, the possibility of cheaper and safer prenatal diagnosis facilities has emerged. Fetal-derived genetic material (cells or cell free DNA) can be obtained from the maternal blood and analyzed, which is considered a non-invasive maneuver with no risk of miscarriage and needs neither complicated procedures nor highly trained personnel for sampling. This allows future screening for thalassemia as well as other genetic diseases [28].

Detection or exclusion of inherited fetal mutations is one of the most important approaches that focuses on detection of mutation that are absent from mother's

#### **Figure 2.**

*Beta-thalassemia: causes, symptoms, and therapeutic modalities. Causes and symptoms are marked in red; and therapeutic modalities are marked in blue.*

genome that requires DNA quantifications with high sensitivity. Even when the parents have the identical mutation, the relative mutation/haplotype approach might detect this fetal mutation [29].
