A. Overview

1. Deficient production of one hemoglobin (Hb) chain, either alpha (a) or ß
   1. ß-globin and related genes found on chromosome 11
   2. Promoters contain general and erythrocyte specific transcription factor binding sites
2. Most are due to insufficient production of a (4 genes) or ß (2 genes) chains only
   1. Can also have two genes affected: for example, HbS (sickle)/ß thalassemia
   2. Variation: abnormal chain produced in addition to reduction in normal chain
3. Increased production of the normal chain leading to tetramer precipitation
   1. Cells and/or membrane areas are removed in the spleen
   2. This leads to red blood cell (RBC) membrane damage and a high rate of cell death
4. Have major and minor forms (Mendelian)
   1. Major forms are homozygous / doubly heterozygous (for example, ß°-thalassemia)
   2. Minor forms are always singly heterozygous (or homozygous ß+-thalassemia
5. Anemia is the major problem
   1. Chronic blood transfusion is required leading to iron overload
   2. Iron overload requirs chelation therapy to prevent hemochromatosis [1]
6. Epidemiology [6]
   1. Overall, ~5% of world's population carry globin variant mutations
   2. ~1.9% carry HbSS; 0.2% have sickle cell anemia
   3. ~1% carry HbE
   4. ~0.3% carry HbC
   5. ~0.044 have thalassemia

B. ß-Thalassemia [6,7]

1. Widespread disease
   1. Mediterranean region
   2. Middle East
   3. Indian subcontinent and Burma
   4. Southeast Asia
   5. Indonesia
2. Severe ß-thalassemia usually occurs during first year of life
   1. Fetal Hb (HbF), composed of two alpha and two gamma chains, is used by newborns
   2. During first year of life, adult Hb (HbA) normally replaces the fetal (HbF) form
3. Clinical Syndromes (Phenotype)
   1. Four clinical syndromes
   2. Silent carrier state - asymptomatic, one mutant ß-globin allele
   3. ß-thalassemia trait - reduced RBC size, reduced Hb levels, one mutant ß-globin allele
   4. Thalassemia intermedia - compound heterozygotes, usually with ß+-thalassemia
   5. Thalassemia major - usually two null ß-globin alleles
   6. Hemoglobin E Thalassemia - mild or severe ß-globin mutation with one HbE chain
   7. Characteristic early onset of anemia, characteristic blood changes, and elevated HbF
   8. Additional clinical syndromes when HbS (sickle Hb) or HbC present
4. Genotype
   1. Genotypes are often named by restriction fragment length polymorphisms (RFLPs)
   2. RFLPs occur because of variance in the DNA sequences
   3. The presence of a specific RFLP defines a haplotype
   4. There are a limited number of haplotypes (RFLPs) found in each population
   5. About 80% of the mutations are associated with only 20% of the haplotypes (RFLPs)
   6. Over 200 different mutations have been found
   7. Overall poor correlation between mutations and clinical phenotype
5. Molecular Mechanisms of ß-Thalassemia
   1. Mutations in the promoter sequence or 5' untranslated region usually result in mild reductions in ß-globin production; this is called ß+-thalassemia (usually mild)
   2. Deletions of the 5' region which completely block transcription lead to the absence of ß-globin production; this is called ß°-thalassemia
   3. Mutations in splice sites and mutations in mRNA processing sites lead to variable reductions in ß-globin synthesis
   4. Thus, mutations in invariant dinucleotide intron/exon junctions lead to ß°-thalassemia
   5. Mutations in sequences flanking the intron/exon junctions lead to variable disease
   6. Mutations in the conserved AATAAA sequence in the 3' untranslated region lead to mild ß+-thalassemia
   7. Mutations in regions that control translation of the mRNA usually lead to severe ß°- thalassemia
   8. Unusual mutations in exon 3 leads to production of unstable ß-globin molecules which bind to alpha-globin chains and cause ß+-disease in a dominantly inherited fashion
6. Pathophysiology
   1. In untreated ß°-thalassemia, erythropoiesis is increased ~10 fold
   2. About 95% of this erythropoiesis is ineffective
   3. Ineffective erythropoiesis is central to the ß-thalassemias
   4. Ineffective erythropoiesis is due to excess of alpha-globin chains
   5. Excess alpha-globin chains leads to G1-cell cycle arrest and RBC precursor death
   6. Excess alpha-globin also causes abnormalities in the ratio of RBC skeletal proteins
   7. Bone marrow is expanded up to 30X normal with extramedullary hematopoiesis
   8. Marrow expansion leads to bone deformities, osteopenia, and microfractures
   9. HbF production is increased to 2-4gm/dL in patients with ß°-thalassemia
7. Clinical Manifestations
   1. Anemia - usually requiring transfusion therapy
   2. Endocrinopathies and bone disease - osteoporosis, infertility
   3. Iron overload
   4. Hypercoagulability - thromboembolic disease, both venous and arterial
8. Iron Overload
   1. Significant complication, primarily of thalassemia major
   2. May lead to liver injury, hepatic fibrosis, cirrhosis, hearing loss
   3. Complicated by hepatitis C virus (HCV) infection
   4. Iron removal treatment may reduce risk for liver injury
   5. Bone marrow transplantation may cure thalassemia and reverse cirrhosis [10]

C. Alpha-Thalassemia [7]

1. Normal individuals have two alpha-globin genes on each chromosome 16
2. There are many carriers of mutations at the alpha globin loci
   1. If both genes are lost, no alpha globin chains are made (a°-thalassemia trait)
   2. If one gene is lost, about 50% of alpha globin is made (a+-thalassemia trait)
3. Inheritance is therefore complex
   1. Inheritance of two a°-thalassemia traits leads to severe thalassemia
   2. Inheritance of one a+- and one a°-thalassemia trait leads moderately severe disease
   3. Impaired alpha-globin production leads to excess gamma and ß chains
   4. These chains form unstable and useless tetramers
   5. Hemoglobin (Hb) H is an a+/a° thalassemia
   6. Hb Barts is an a°/a° thalassemia
4. Hemoglobin H Disease (a+/a° thalassemia) [9]
   1. Moderately severe hemoloytic anemia
   2. RBC show inclusions of precipited Hb H when deoxygenated
   3. Due to deletion or mutation of 3 of the 4 alpha-thalassemia genes
   4. Leads to formation of tetramers of ß-globin genes
   5. In Chinese persons, nondeletional type Hb H disease is more severe than deletional type
   6. Iron overload is the major cause of morbidity in all forms of Hb H disease
5. Hb Barts Disease
   1. Extremely severe disease incompatible with life
   2. Hydrops fetalis syndrome develops
   3. This is stillbirth of severely hydropic infant in last half of pregnancy
   4. Hb Barts is formed from 4 gamma-globin tetramers (a°/a° thalassemia)
6. Mild Forms of Alpha-Thalassemia
   1. With a°-thalassemia - mild hypochromic anemia with normal HbA2 levels
   2. With a+-thalassemia - completely normal appearing or very mild hypochromia
   3. Homozygous a+-thalassemia resuls in mild hypochromic anemia with normal HbA2 levels
   4. In general, genetic testing is required for diagnosis of these forms of thalassemia
7. Alpha-Thalassemia - Retardation (ATR) Syndrome
   1. ATR is not typically found in tropical populations
   2. Both X-linked and chromosome 16 forms of the disease exist
   3. X-linked form due to mutations of a gene, XH2, on Xq (regulates alpha-globin genes)
   4. Autosomal form due to deletions of the tip of chromosome 16

D. Diagnosis

1. Blood Smear
   1. Microcytic, hypochromic cells, with target cells
   2. Nucleated red cells and reticulocytosis present due to insufficient oxygenation
2. Molecular diagnosis now possible for ß-thalassemias [4]
3. Elevated HbF levels

E. Therapy [6]

1. Blood Transfusions
   1. Maintain Hb levels >11gm/dL
   2. However, Hb prior to transfusion should not exceed 9.5gm/dL
   3. Transfuse to decrease nucleated RBC in periphery
   4. Reticulocyte counts are low because of killing in bone marrow medulla
2. Frequent transfusions leads to iron overload
   1. This will progress to hemochromatosis unless treated
   2. Treatment is chelation of iron which is effective and improves survival [1,2]
3. Chelation Therapy [1,2,6]
   1. Deferoxamine B mesylate (subcutaneous) or oral deferiprone available
   2. Deferoxamine was standard of care and is very effective
   3. Deferaserox (Exjade®), an oral iron chelator, is now approved for iron overload
   4. Ferritin levels have classically been used to monitor body iron loads
   5. However, hepatic iron stores are probably a better monitoring system
   6. Determination of hepatic iron stores requires a liver biopsy
   7. Hepatic iron stores accurately reflect total body iron (in thalassemia patients) [8]
   8. ICL670 is an oral iron chelator with very promising early clinical data [12]
4. Oral Iron Chelators
   1. Deferiprone and deferasirox are oral iron chelators
   2. Deferiprone more effective than deferoxamine in removing cardiac iron in ß-thalassemia patients [11]
   3. Patients on qd deferiprone had >5X increased risk of hepatic fibrosis (versus tid) [5]
   4. Increased risk (overall low) of agranulocytosis [3]
   5. Therefore, monitoring is required with long term use of deferiprone
   6. Thrice daily deferiprone should be used (not daily)
   7. Oral deferasirox (Exjade®) now available for age over 2 years for iron overload [13]
   8. Initial dose deferasirox is 20mg/kg oral initially, adjusted every 3-6 months; discontinue temporarily if ferritin remains <500µg/L [13]
   9. Agranulocytosis does not appear to be a concern with deferasirox
5. Hematopoietic Stem Cell Transplant
   1. Reserved for patients with severe ß-thalassemias
   2. Marrow obtained from HLA-identical donors
   3. Outcomes are good if liver damage from iron overload has not occurred
   4. However, reversal of liver damage after BMT and iron removal reported [10]
6. Novel Treatments
   1. Induction of HbF
   2. Gene Therapy
   3. Unclear if anticoagulant or antiplatelet therapy are net beneficial in ß-thalassemia

References

1. Brittenham GM, Griffith PM, Nienhuis AW, et al. 1994. NEJM. 331(9):567
2. Olivieri NF, Nathan DG, MacMillan JH, et al. 1994. NEJM. 331(9):574
3. Olivieri NF, Brittenham GM, Matusi D, et al. 1995. NEJM. 332(14):918
4. Cao A, Saba L, Galanello R, Rosatelli MC. 1997. JAMA. 278(15):1273
5. Rund D and Rachmilewitz E. 2005. NEJM. 353(11):1135
6. Olivieri NF. 1999. NEJM. 341(2):99
7. Weatherall DJ and Provan AB. 2000. Lancet. 355(9120):1169
8. Angelucci E, Brittenham GM, McLaren CE, et al. 2000. NEJM. 343(5):327
9. Chen FE, Ooi C, Ha SY, et al. 2000. NEJM. 343(8):544
10. Muretto P, Angelucci E, Lucarelli G. 2002. Ann Intern Med. 136(9):667
11. Anderson LJ, Wonke B, Prescott E, et al. 2002. Lancet. 360(9332):516
12. Nisbet-Brown E, Olivieri NF, Giardina PJ, et al. 2003. Lancet. 361(9369):1597
13. Deferasirox. 2006. Med Let. 48(1233):35