Fetal globin expression is regulated by Friend of Prmt1

Abstract An estimated 6% to 7% of the earth's population carries a mutation affecting red blood cell function. The β-thalassemias and sickle cell disease are the most common monogenic disorders caused by these mutations. Increased levels of γ-globin ameliorate the severity of these diseases because fetal hemoglobin (HbF; α2γ2) can effectively replace adult hemoglobin (HbA; α2β2) and counteract polymerization of sickle hemoglobin (HbS; α2βS2). Therefore, understanding the molecular mechanism of globin switching is of biologic and clinical importance. Here, we show that the recently identified chromatin factor Friend of Prmt1 (FOP) is a critical modulator of γ-globin gene expression. Knockdown of FOP in adult erythroid progenitors strongly induces HbF. Importantly, γ-globin expression can be elevated in cells from β-thalassemic patients by reducing FOP levels. These observations identify FOP as a novel therapeutic target in β-hemoglobinopathies.

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Differences in Fetal Hemoglobin Induction in Sickle Cell Disease and beta-Thalassemia.

Blood ◽

10.1182/blood.v108.11.555.555 ◽

2006 ◽

Vol 108 (11) ◽

pp. 555-555 ◽

Cited By ~ 2

Author(s):

Hassana Fathallah ◽

Ali Taher ◽

Ali Bazarbachi ◽

George F. Atweh

Keyword(s):

Gene Expression ◽

Sickle Cell Disease ◽

Globin Gene ◽

Fetal Hemoglobin ◽

Mrna Levels ◽

Globin Genes ◽

Thalassemia Intermedia ◽

Cell Disease ◽

Globin Mrna ◽

Globin Gene Expression

Abstract A number of therapeutic agents including hydroxyurea, butyrate and decitabine have shown considerable promise in the treatment of sickle cell disease (SCD). However, the same agents have shown less clinical activity in β-thalassemia. As a first step towards understanding the molecular basis of the different clinical responses to these agents, we have studied the mechanisms of induction of fetal hemoglobin (HbF) by butyrate in BFU-E derived cells from 5 patients with SCD and 9 patients with β-thalassemia intermedia. Exposure to butyrate resulted in a dose-dependent augmentation of γ-globin mRNA levels in erythroid cells from patients with SCD. In contrast, induction of γ-globin expression in erythroid cells from patients with β-thalassemia intermedia was only seen at a high concentration of butyrate. The increase in γ-globin mRNA levels in patients with SCD and β-thalassemia intermedia was associated with opening of the DNA structure as manifested by decreased DNA methylation at the γ-globin promoters. Interestingly, butyrate exposure had markedly different effects on the expression of the β- and α-globin genes in the two categories of patients. Butyrate decreased the level of β-globin mRNA in 4 out of 5 patients with SCD (P = 0.04), while in β-thalassemia the levels of β-globin mRNA did not change in 7 patients and decreased in 2 patients after butyrate exposure (P = 0.12). Thus in patients with SCD, the effects of the induction of the γ-globin gene on the γ/(β+γ) mRNA ratios were further enhanced by the butyrate-mediated decreased expression of the β-globin gene. As a result, γ/(β+γ) mRNA ratios increased in all patients with SCD, with a mean increase of 31% (P = 0.002). In contrast, butyrate increased γ/(β+γ) mRNA ratios only in 4 out of 9 patients with β-thalassemia, with a more modest mean increase of 12% (P = 0.004). Interestingly, the decreased β-globin expression in patients with SCD was associated with closing of the DNA configuration as manifested by hypermethylation of DNA at the promoter of the β-globin gene while methylation of the same promoter did not change following butyrate exposure in patients with β-thalassemia intermedia. More surprisingly, the expression of the α-globin genes increased following butyrate exposure in 4 out of 9 patients with β-thalassemia, while the levels of α-globin mRNA decreased in 4 out of 5 patients with SCD. As a result, the favorable effects of the butyrate-induced increase in γ-globin gene expression on the α: non-α mRNA imbalance in patients with β-thalassemia intermedia were partly neutralized by the corresponding increase in α-globin gene expression. These differences may explain, at least in part, the more favorable effects of inducers of HbF in SCD than in β-thalassemia. Further studies are necessary to fully understand the molecular bases of the different responses to agents that induce HbF in patients with these disorders.

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Cucurbitacin D Upregulates Fetal Hemoglobin Expression in K562 Cells and Human Erythroid Progenitors.

Blood ◽

10.1182/blood.v110.11.3833.3833 ◽

2007 ◽

Vol 110 (11) ◽

pp. 3833-3833

Author(s):

Hongtao Xing ◽

Siwei Zhang ◽

H. Phillip Koeffler ◽

Ming Chiu Fung

Keyword(s):

Gene Expression ◽

Globin Gene ◽

Fetal Hemoglobin ◽

K562 Cells ◽

Negative Control ◽

Optimal Dosage ◽

Erythroid Progenitors ◽

Cucurbitacin D ◽

Globin Gene Expression ◽

Sodium Phenylbutyrate

Abstract The search for novel therapeutic candidates causing reactivation of fetal hemoglobin (a2g2; HbF) to reduce the imbalance of globin gene expression is important in order to develop effective approach for the clinical management of sickle cell anemia and b-thalassemia. For the first time, we have identified cucurbitacin D (CuD), a naturally occurring oxygenated tetracyclic triterpenoid, as a molecular entity inducing g-globin gene expression and HbF synthesis in K562 cells and human erythroid progenitors from either peripheral blood or bone marrow. The upregulation of HbF induced by CuD was dose- and time- dependent. CuD was compared to hydroxyurea (HU), 5-azacytidine, amifostine, recombinant human erythropoietin (rhEPO), and sodium phenylbutyrate. At their optimal dosage, CuD (12.5 ng/mL) and HU (25.0 μg/mL) induced nearly 70% K562 cells to express total hemoglobin after 6 days culture, which was higher than the induction by Amifostine (30%), 5-azacytidine (36%), rhEPO (16%), sodium phenylbutyrate (23%) at their optimal concentrations and negative control (11%). Fetal hemoglobin ELISA showed that CuD (12.5 ng/mL) and 5-azacytidine (400 ng/mL) induced higher levels of fetal hemoglobin in K562 cells (15.4 ng/μL and 29.3 ng/μL, respectively), compared to HU (10.3 ng/μL), amifostine (7.8 ng/μL), rhEPO (10.9 ng/μL), sodium phenylbutyrate (9.9 ng/μL) at their optimal concentrations and negative control (5.3 ng/μL). CuD induced a significantly higher fetal cell percentage than HU in K562 cells (65% vs 37% maximum) and primary erythroid progenitors (36% vs 21% maximum) based on the immunofluorescence imaging and flow cytometry analysis. Real-time PCR results showed that the amount of γ-globin mRNA increased from 2.5-fold in CuD-optimal-treated cells (12.5 ng/mL, 48 hours) compared with 1.5-fold in HU-optimal-treated cells (25.0 μg/mL, 48 hours). Growth inhibition assay (MTT) demonstrated that CuD at its optimal γ-globin inducing dosage (12.5 ng/mL) inhibited proliferation of K562 by less than 10% of untreated control cells; while hydroxyurea at its optimal dosage (25.0 μg/mL) inhibited 80% of cell division. The in vitro therapeutic index (calculated by dividing the dose inhibiting 50% cell growth (IC50) by dose inducing 50% maximal HbF production (ED50)) of CuD was 40-fold greater than HU. Taken together, the results suggest that CuD has the potential to be a therapeutic agent for treatment of sickle cell anemia and b-thalassemia.

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Loss of NRF2 Function Worsens the Pathophysiology of Sickle Cell Disease in a Transgenic Mouse Model

Blood ◽

10.1182/blood.v130.suppl_1.960.960 ◽

2017 ◽

Vol 130 (Suppl_1) ◽

pp. 960-960

Author(s):

Xingguo Zhu ◽

Betty S. Pace

Keyword(s):

Oxidative Stress ◽

Gene Expression ◽

Globin Gene ◽

Critical Role ◽

Fetal Hemoglobin ◽

Oxygen Species ◽

Cell Disease ◽

Nrf2 Knockout ◽

Globin Gene Expression

The basic leucine zipper transcription factor, nuclear factor (erythroid-derived 2)-like 2 (NRF2) plays a critical role in the cellular antioxidant response to control oxidative stress. We and others previously demonstrated that NRF2 activation enhances γ-globin gene transcription and fetal hemoglobin expression in human primary erythroid progenitors through changes in chromatin structure (Zhu et al., Haematologica 2017). In this study, we investigated the protective role of NRF2 in reversing the pathophysiology of sickle cell disease (SCD) in a SCD/NRF2 knockout (SCD/NRF2KO) transgenic model created in our lab by crossbreeding Townes SCD mice (Ryan et al., Science 1997) and NRF2 knockout mice (Kuroha et al., J Biochem 1998). The NRF2 gene is transmitted through autosomal recessive Mendelian genetics for wild-type, heterozygote and NRF2KO pups. By contrast, the SCD/NRF2KO genotype was present in <2% of pups. In addition, the fertility and litter size of SCD/NRF2KO females were lower than SCDWT mice, suggesting a critical role of NRF2 in the survival of pups during gestation. To determine the hematopoietic effect of NRF2KO in SCD, we monitored the complete blood count with differential and reticulocyte count. There was no significant change in any parameters except higher total white blood cell counts in the SS/NRF2KO mouse suggesting increased inflammation. Examination of globin gene expression patterns in SS/NRF2KO mice showed reduced γ-globin gene expression during erythroid differentiation in the E13.5 and E18.5 fetal liver, adult spleen and bone marrow. In addition, peripheral blood red cells had a 33% increase (p<0.05) in reactive oxygen species and a significant 38% increase in sickling under in vitro hypoxic conditions. We next characterized the effects of NRF2 loss on organ pathology. The SCD/NRF2KO mice displayed greater splenomegaly indicating exaggerated hemolysis most likely due to high levels of reactive oxygen species. By 8-10 weeks of age, the SCD/NRF2KO mouse showed significant inflammation by hematoxylin-eosin staining of the spleen, lung and liver when compared to SCD/NRF2WT mice. Protein expression profiling by western blotting using adult spleen demonstrated downregulation of the antioxidant proteins heme oxygenase 1 (HMOX1), NADPH: quinone oxidoreductase 1 (NQO1) and catalase by 31%, 60%, and 48% respectively (p<0.05). To further support a severe disease phenotype, the expression of intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1) were increased by 1.7-fold and 2.3-fold (p<0.05) while vascular endothelial growth factor (VEGF) levels were not changed. Lastly, the expression of interleukin 1β (IL-1β), IL-6, tumor necrosis factor α (TNF-α), monocyte chemoattractant protein (MCP-1) and macrophage migration inhibitory factor (MIF-1) was elevated in SCD/NRF2KO mice compared to SCDWT mice. These data validate a critical role of NRF2 in ameliorating the phenotypic severity of SCD by protecting against red blood cell sickling, oxidative stress and tissue inflammation. Furthermore, the ability of NRF2 to mediate fetal hemoglobin induction provides a rationale for the development of therapeutic agents that activate NRF2 expression. Disclosures No relevant conflicts of interest to declare.

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Analysis of hemoglobin F production in Saudi Arabian families with sickle cell anemia

Blood ◽

10.1182/blood.v70.3.716.bloodjournal703716 ◽

1987 ◽

Vol 70 (3) ◽

pp. 716-720 ◽

Cited By ~ 1

Author(s):

BA Miller ◽

M Salameh ◽

M Ahmed ◽

N Olivieri ◽

G Antognetti ◽

...

Keyword(s):

Sickle Cell Disease ◽

Sickle Cell ◽

Sickle Cell Anemia ◽

Globin Gene ◽

Fetal Hemoglobin ◽

Cell Disease ◽

Erythroid Progenitors ◽

Hemoglobin F ◽

Gamma Globin ◽

Hemoglobin Production

Erythrocytes and progenitor-derived erythroblasts of sickle cell anemia patients from the Eastern Province of Saudi Arabia contain increased fetal hemoglobin and G gamma globin. A distinctive DNA polymorphism haplotype in the beta globin gene cluster (++- +-), tightly coupled to a C----T substitution at position -158 5′ to the cap site of the G gamma globin gene, is strongly associated with sickle cell disease in this region. To determine whether the increased fetal hemoglobin production and/or elevated G gamma globin content are tightly linked to this haplotype, we studied 55 members of five Saudi families in which sickle cell disease is present. The results did not suggest a tight linkage of the haplotype to increased fetal hemoglobin production. On the other hand, several sickle trait family members heterozygous for the haplotype had normal fetal hemoglobin production in culture but elevated G gamma to A gamma ratios in peripheral blood. This observation suggests that in this genetic background increased expression of the G gamma globin gene may occur without a measurable increase in total fetal hemoglobin production. The family studies also clearly demonstrate that increased fetal hemoglobin production by erythroid progenitors is dependent on zygosity for the sickle gene in this population. These findings strongly suggest that other factors, such as the products of genes stimulated by hemolytic stress or other genetic determinants associated with the Saudi beta S chromosome, may interact with the -158 C----T substitution and influence gamma globin gene expression in this population.

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Analysis of hemoglobin F production in Saudi Arabian families with sickle cell anemia

Blood ◽

10.1182/blood.v70.3.716.716 ◽

1987 ◽

Vol 70 (3) ◽

pp. 716-720 ◽

Cited By ~ 36

Author(s):

BA Miller ◽

M Salameh ◽

M Ahmed ◽

N Olivieri ◽

G Antognetti ◽

...

Keyword(s):

Sickle Cell Disease ◽

Sickle Cell ◽

Sickle Cell Anemia ◽

Globin Gene ◽

Fetal Hemoglobin ◽

Cell Disease ◽

Erythroid Progenitors ◽

Hemoglobin F ◽

Gamma Globin ◽

Hemoglobin Production

Abstract Erythrocytes and progenitor-derived erythroblasts of sickle cell anemia patients from the Eastern Province of Saudi Arabia contain increased fetal hemoglobin and G gamma globin. A distinctive DNA polymorphism haplotype in the beta globin gene cluster (++- +-), tightly coupled to a C----T substitution at position -158 5′ to the cap site of the G gamma globin gene, is strongly associated with sickle cell disease in this region. To determine whether the increased fetal hemoglobin production and/or elevated G gamma globin content are tightly linked to this haplotype, we studied 55 members of five Saudi families in which sickle cell disease is present. The results did not suggest a tight linkage of the haplotype to increased fetal hemoglobin production. On the other hand, several sickle trait family members heterozygous for the haplotype had normal fetal hemoglobin production in culture but elevated G gamma to A gamma ratios in peripheral blood. This observation suggests that in this genetic background increased expression of the G gamma globin gene may occur without a measurable increase in total fetal hemoglobin production. The family studies also clearly demonstrate that increased fetal hemoglobin production by erythroid progenitors is dependent on zygosity for the sickle gene in this population. These findings strongly suggest that other factors, such as the products of genes stimulated by hemolytic stress or other genetic determinants associated with the Saudi beta S chromosome, may interact with the -158 C----T substitution and influence gamma globin gene expression in this population.

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δ-Aminolevulinate induces fetal hemoglobin expression by enhancing cellular heme biosynthesis

Experimental Biology and Medicine ◽

10.1177/1535370219872995 ◽

2019 ◽

Vol 244 (14) ◽

pp. 1220-1232

Author(s):

Li Liu ◽

Xingguo Zhu ◽

Alexander Yu ◽

Christina M Ward ◽

Betty S Pace

Keyword(s):

Therapeutic Strategy ◽

Clinical Symptoms ◽

Globin Gene ◽

Fetal Hemoglobin ◽

Heme Biosynthesis ◽

Erythroid Cell ◽

Nuclear Factor ◽

Cell Disease ◽

Erythroid Progenitors ◽

Novel Therapeutic

Sickle cell disease (SCD) and β-thalassemia are inherited blood disorders caused by genetic defects in the β-globin gene on chromosome 11, producing severe disease in people worldwide. Induction of fetal hemoglobin consisting of two α-globin and two γ-globin chains ameliorates the clinical symptoms of both disorders. In the present study, we investigated the ability of δ-aminolevulinate (ALA), the heme precursor, to activate γ-globin gene expression as well as its effects on cellular functions in erythroid cell systems. We demonstrated that ALA induced γ-globin expression at both the transcriptional and protein levels in the KU812 erythroid cell line. Using inhibitors targeting two enzymes in the heme biosynthesis pathway, we showed that cellular heme biosynthesis was involved in ALA-mediated γ-globin activation. Moreover, the transcription factor NRF2 (nuclear factor [erythroid-derived 2]-like 2), a critical regulator of the cellular antioxidant response, was activated by ALA and contributed to mechanisms of γ-globin activation; ALA did not affect cell proliferation and was not toxic to cells. Subsequent studies demonstrated ALA-induced γ-globin activation in erythroid progenitors generated from normal human CD34+ stem cells. These data support future study to explore the potential of stimulating intracellular heme biosynthesis by ALA or similar compounds as a novel therapeutic strategy for treating SCD and β-thalassemia. Impact statement Inherited mutations in the β-globin-like genes result in the most common forms of genetic blood disease including sickle cell disease (SCD) and β-thalassemia worldwide. Therefore, effective inexpensive therapies that can be distributed widely are highly desirable. Currently, drug-mediated fetal hemoglobin (HbF) induction can ameliorate clinical symptoms of SCD and β-thalassemia and is the most effective strategy for developing new therapeutic options. In the current study, we confirmed that δ-Aminolevulinate (ALA), the precursor of heme, induces γ-globin expression at both the transcriptional and translational levels in primary human erythroid progenitors. Moreover, the results indicate activation of the transcription factor NRF2 (nuclear factor (erythroid-derived 2)-like 2) by ALA to enhance HbF expression. These data support future study to explore the potential of stimulating intracellular heme biosynthesis by ALA or similar compounds as a novel therapeutic strategy for treating SCD and β-thalassemia.

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NRF2 mediates γ-globin gene regulation through epigenetic modifications in a β-YAC transgenic mouse model

Experimental Biology and Medicine ◽

10.1177/1535370220945305 ◽

2020 ◽

Vol 245 (15) ◽

pp. 1308-1318

Author(s):

Xingguo Zhu ◽

Caixia Xi ◽

Alexander Ward ◽

Mayuko Takezaki ◽

Huidong Shi ◽

...

Keyword(s):

Sickle Cell Disease ◽

Mouse Model ◽

Sickle Cell ◽

Globin Gene ◽

Fetal Hemoglobin ◽

Dimethyl Fumarate ◽

Cell Disease ◽

Nrf2 Activator ◽

Globin Gene Regulation ◽

Globin Gene Expression

NRF2 is the master regulator for the cellular oxidative stress response and regulates γ-globin gene expression in human erythroid progenitors and sickle cell disease mice. To explore NRF2 function, we established a human β-globin locus yeast artificial chromosome transgenic/NRF2 knockout (β-YAC/NRF2−/−) mouse model. NRF2 loss reduced γ-globin gene expression during erythropoiesis and abolished the ability of dimethyl fumarate, an NRF2 activator, to enhance γ-globin transcription. We observed decreased H3K4Me1 and H3K4Me3 chromatin marks and association of TATA-binding protein and RNA polymerase II at the β-locus control region (LCR) and γ-globin gene promoters in β-YAC/NRF2−/− mice. As a result, long-range chromatin interaction between the LCR DNase I hypersensitive sites and γ-globin gene was decreased, while interaction with the β-globin was not affected. Further, NRF2 loss silenced the expression of DNA methylcytosine dioxygenases TET1, TET2, and TET3 and inhibited γ-globin gene DNA hydroxymethylation. Subsequently, protein-protein interaction between NRF2 and TET3 was demonstrated. These data support the ability of NRF2 to mediate γ-globin gene regulation through epigenetic DNA and histone modifications. Impact statement Sickle cell disease is an inherited hemoglobin disorder that affects over 100,000 people in the United States causing high morbidity and early mortality. Although new treatments were recently approved by the FDA, only one drug Hydroxyurea induces fetal hemoglobin expression to inhibit sickle hemoglobin polymerization in red blood cells. Our laboratory previously demonstrated the ability of the NRF2 activator, dimethyl fumarate to induce fetal hemoglobin in the sickle cell mouse model. In this study, we investigated molecular mechanisms of γ-globin gene activation by NRF2. We observed the ability of NRF2 to modulate chromatin structure in the human β-like globin gene locus of β-YAC transgenic mice during development. Furthermore, an NRF2/TET3 interaction regulates γ-globin gene DNA methylation. These findings provide potential new molecular targets for small molecule drug developed for treating sickle cell disease.

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Investigation of miRNA-29b As a Fetal Hemoglobin Inducer in Sickle Cell Disease

Blood ◽

10.1182/blood-2021-153290 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 2023-2023

Author(s):

Athena Starlard-Davenport ◽

Alana Smith ◽

Qingqing Gu ◽

Umapathy N Siddaramappa ◽

Chithra Palani ◽

...

Keyword(s):

Sickle Cell ◽

Globin Gene ◽

Fetal Hemoglobin ◽

Erythroid Differentiation ◽

Cell Disease ◽

Dna Hypomethylation ◽

Erythroid Progenitors ◽

Protein Levels ◽

F Cells

Abstract Introduction: Therapeutic intervention aimed at inducing fetal hemoglobin (HbF) expression is an effective approach for ameliorating the clinical severity of sickle cell disease (SCD). Hydroxyurea (HU) is the only FDA-approved drug with proven efficacy for inducing HbF in SCD. Recently, DNA methyltransferase (DNMT) inhibitors were shown to reactivate γ-globin gene expression via DNA hypomethylation. However, alternate approaches involve development of small non-coding microRNAs (miR) to silence major repressors of γ-globin transcription. Previous studies from our group showed that miR-29b inhibits DNA methylation by binding the 3' untranslated region of DNMT3A/B (Starlard-Davenport A et al., J Carcinog 12:15; 2013). Subsequent, studies demonstrated that increased levels of miR-29b are associated with high HbF levels in patients with SCD. To gain insights into mechanisms, studies performed in KU812 cells demonstrated DNMT3A/B silencing with ү-globin gene activation. Moreover, miR29b increased HbF expression in erythroid progenitors generated from normal adult CD34 + stem cells (Starlard-Davenport A et al., Br J Haematol 186:91-100; 2019). Therefore, we tested the hypothesis that miR-29b activates γ-globin transcription via DNA hypomethylation in normal and sickle erythroid progenitors and Townes SCD mouse model. Methods: Normal CD34 + cells (n=3) and peripheral blood mononuclear cells (n=2) isolated from SCD patients, were cultured using a two-phase erythroid culturing system. Cells were electroporated with miR-29b mimic (50nM and 100nM) or negative Scrambled (Scr) control on day 8 and collected after 24 h. Erythroid differentiation was assessed using Giemsa staining, and flow cytometry was used to measure the % HbF positive cells (F-cells). Real-time PCR was used to quantify expression of miR-29b, γ-globin, and β-globin genes, and Western blot was performed to measure DNMT, HbF and HbS protein levels. In vivo studies were performed in Townes SCD mice (3 per group) treated for 28 days by continuous infusion with subcutaneous mini-osmotic pumps. The treatment groups included miR-29b (2mg/kg/day and 3mg/kg/day) and corresponding doses of Scr control mimics. Results: We observed erythroid differentiation of untreated normal and SCD erythroid progenitors at day 7, 12, and 14 of culture assessed by Giemsa stain. Treatment with miR-29b increased the ү/ү+β mRNA ratio by 2.5-fold (p<0.01) and F-cell levels increased from ~6.0% (Scr) to 15% with miR-29b (100nM); we observed lower DNMT3 mRNA and protein levels after 100nM miR-29b treatment. To confirm HbF induction by miR-29b in sickle progenitors we treated cells on day 8 with 50nM and 100nM miR-29b and observed >85% increase in %F-cells compared to Scr cells. Subsequent miR-29b treatment was conducted for 28 days in Townes SCD mice, which was well tolerated documented by good weight gain and no deaths. There were minimal effects on hematopoiesis with a decrease in total white blood count and an increase in neutrophils. Total hemoglobin, reticulocyte and platelet counts remained stable. By week 4, we observed a 0.5 fold and 2.3-fold (p<0.005) increase in %F-cells at the 2mg/kg and 3 mg/kg miR-29b doses respectively compared to corresponding Scr controls. By week 4, miR29b increased HbF levels 2.1-fold by Western blot, and sickle cell levels under hypoxic conditions decreased 32% (p<0.01). Conclusions: Our findings support the ability of miR-29b to induce HbF in normal and sickle erythroid progenitors without significant toxicity in vitro and in SCD mice in vivo. This research highlights a novel miRNA-based epigenetic approach to induce HbF supporting discovery of new drugs to expand treatment options for SCD. Disclosures Pace: Imara Inc.: Consultancy.

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Epigenetic Insights and Potential Modifiers as Therapeutic Targets in β–Thalassemia

Biomolecules ◽

10.3390/biom11050755 ◽

2021 ◽

Vol 11 (5) ◽

pp. 755

Author(s):

Nur Atikah Zakaria ◽

Md Asiful Islam ◽

Wan Zaidah Abdullah ◽

Rosnah Bahar ◽

Abdul Aziz Mohamed Yusoff ◽

...

Keyword(s):

Gene Expression ◽

Clinical Presentation ◽

Globin Gene ◽

Fetal Hemoglobin ◽

Dna Hypomethylation ◽

Considerable Research ◽

Non Coding Rna ◽

Hbf Level ◽

Long Non Coding Rna ◽

Globin Gene Expression

Thalassemia, an inherited quantitative globin disorder, consists of two types, α– and β–thalassemia. β–thalassemia is a heterogeneous disease that can be asymptomatic, mild, or even severe. Considerable research has focused on investigating its underlying etiology. These studies found that DNA hypomethylation in the β–globin gene cluster is significantly related to fetal hemoglobin (HbF) elevation. Histone modification reactivates γ-globin gene expression in adults and increases β–globin expression. Down-regulation of γ–globin suppressor genes, i.e., BCL11A, KLF1, HBG-XMN1, HBS1L-MYB, and SOX6, elevates the HbF level. β–thalassemia severity is predictable through FLT1, ARG2, NOS2A, and MAP3K5 gene expression. NOS2A and MAP3K5 may predict the β–thalassemia patient’s response to hydroxyurea, a HbF-inducing drug. The transcription factors NRF2 and BACH1 work with antioxidant enzymes, i.e., PRDX1, PRDX2, TRX1, and SOD1, to protect erythrocytes from oxidative damage, thus increasing their lifespan. A single β–thalassemia-causing mutation can result in different phenotypes, and these are predictable by IGSF4 and LARP2 methylation as well as long non-coding RNA expression levels. Finally, the coinheritance of β–thalassemia with α–thalassemia ameliorates the β–thalassemia clinical presentation. In conclusion, the management of β–thalassemia is currently limited to genetic and epigenetic approaches, and numerous factors should be further explored in the future.

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Genome editing using CRISPR-Cas9 to create the HPFH genotype in HSPCs: An approach for treating sickle cell disease and β-thalassemia

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1612075113 ◽

2016 ◽

Vol 113 (38) ◽

pp. 10661-10665 ◽

Cited By ~ 79

Author(s):

Lin Ye ◽

Jiaming Wang ◽

Yuting Tan ◽

Ashley I. Beyer ◽

Fei Xie ◽

...

Keyword(s):

Sickle Cell Disease ◽

Genome Editing ◽

Sickle Cell ◽

Globin Gene ◽

Autologous Transplantation ◽

Clinical Manifestations ◽

Fetal Hemoglobin ◽

Compound Heterozygous ◽

Cell Disease ◽

Hematopoietic Stem

Hereditary persistence of fetal hemoglobin (HPFH) is a condition in some individuals who have a high level of fetal hemoglobin throughout life. Individuals with compound heterozygous β-thalassemia or sickle cell disease (SCD) and HPFH have milder clinical manifestations. Using RNA-guided clustered regularly interspaced short palindromic repeats-associated Cas9 (CRISPR-Cas9) genome-editing technology, we deleted, in normal hematopoietic stem and progenitor cells (HSPCs), 13 kb of the β-globin locus to mimic the naturally occurring Sicilian HPFH mutation. The efficiency of targeting deletion reached 31% in cells with the delivery of both upstream and downstream breakpoint guide RNA (gRNA)-guided Staphylococcus aureus Cas9 nuclease (SaCas9). The erythroid colonies differentiated from HSPCs with HPFH deletion showed significantly higher γ-globin gene expression compared with the colonies without deletion. By T7 endonuclease 1 assay, we did not detect any off-target effects in the colonies with deletion. We propose that this strategy of using nonhomologous end joining (NHEJ) to modify the genome may provide an efficient approach toward the development of a safe autologous transplantation for patients with homozygous β-thalassemia and SCD.

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