The new england journal of medicine

review article

medical progress

b-Thalassemia
Deborah Rund, M.D., and Eliezer Rachmilewitz, M.D.

t halassemia is a hereditary anemia resulting from defects in

hemoglobin production.1 b-Thalassemia, which is caused by a decrease in the
production of b-globin chains (Fig. 1), affects multiple organs and is associ-
ated with considerable morbidity and mortality.2 Accordingly, lifelong care is required,3
and financial expenditures for proper treatment are substantial.4
From the Hematology Department, He-
brew University–Hadassah Medical Cen-
ter, Ein Kerem, Jerusalem (D.R.); and the
Hematology Department, E. Wolfson
Medical Center, Holon (E.R.) — both in
Israel. Address reprint requests to Dr.
Rund at the Hematology Department, Ha-
Thalassemia is among the most common genetic disorders worldwide; 4.83 per- dassah University Hospital, Ein Kerem,
cent of the world’s population carry globin variants, including 1.67 percent of the pop- Jerusalem IL91120, Israel, or at rund@
ulation who are heterozygous for a-thalassemia and b-thalassemia. In addition, 1.92 cc.huji.ac.il.

percent carry sickle hemoglobin, 0.95 percent carry hemoglobin E, and 0.29 percent N Engl J Med 2005;353:1135-46.
carry hemoglobin C. Thus, the worldwide birth rate of people who are homozygous or Copyright © 2005 Massachusetts Medical Society.
compound heterozygous for symptomatic globin disorders, including a-thalassemia
and b-thalassemia, is no less than 2.4 per 1000 births, of which 1.96 have sickle cell
disease and 0.44 have thalassemias.5

molecular and cellular pathology

b-Thalassemia is caused by any of more than 200 point mutations and, rarely, by dele-
tions.1 Thalassemia is clinically heterogeneous because various genetic lesions vari-
ably impair globin-chain synthesis. However, genotypic variability at known loci is of-
ten insufficient to explain the disparate phenotypes of individual patients with the
same genotype. Disparity between genotypes and phenotypes is particularly marked in
thalassemia intermedia and hemoglobin E thalassemia (Table 1). However, the known
genetic factors are insufficient to account for the marked variability, and other genetic
modifiers may exist.6
Recently, an a-hemoglobin–stabilizing protein was identified that binds to and sta-
bilizes free a chains, thereby blocking the production of reactive oxygen species and
reducing oxidative damage to erythrocytes. This protein appears to modulate the clini-
cal picture of b-thalassemia in a murine model7 but in human studies has not been
found to modify thalassemia.
Hemolysis and ineffective erythropoiesis together cause the anemia that occurs in
thalassemia. The relative contributions of these two pathologic processes differ in var-
ious forms of thalassemia.8 Figure 2 illustrates the complex chain of events that occurs
in erythrocytes, resulting in their accelerated peripheral destruction.
The bone marrow of patients with thalassemia contains five to six times the number
of erythroid precursors as does the bone marrow of healthy controls,17 with 15 times
the number of apoptotic cells in the polychromatophilic and orthochromic stages.17,18
Accelerated apoptosis, the major cause of ineffective erythropoiesis, is caused by ex-
cess a-chain deposition in erythroid precursors.8 Although the exact mechanism is not
known, a death-receptor–mediated pathway seems to be involved with Fas–Fas ligand
interactions.19 In normal erythropoiesis, apoptotic mechanisms seem to play a regula-
tory role and are required for normal erythroid maturation.20 Accelerated apoptosis is

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 The new england journal of medicine

Figure 1. Management of Thalassemia and Treatment-Related Complications.

The anemia that is associated with thalassemia may be severe and is accompanied by ineffective erythropoiesis, with bone
expansion and extramedullary hematopoiesis in the liver, spleen, and other sites, such as paravertebral masses. Transfu-
sion therapy, which is the mainstay of treatment, allows for normal growth and development and suppresses ineffective
erythropoiesis. Transfusion-transmitted infections (primarily hepatitis B and C) are an important cause of death in coun-
tries where proper testing is not available. Iron overload results both from transfusional hemosiderosis and excess gas-
trointestinal iron absorption. Iron deposition in the heart, liver, and multiple endocrine glands results in severe damage
to these organs, with variable endocrine organ failure. The endocrinopathies can be treated with hormone replacement.
However, the most serious result of iron overload is life-threatening cardiotoxicity, for which chelation therapy is required.
Thalassemia can be cured by bone marrow transplantation. Experimental therapies to ameliorate the anemia that have
been or are currently under investigation include fetal hemoglobin modifiers and antioxidants. In the future, gene thera-
py or other molecular methods may be feasible.

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 medical progress

Table 1. Genetic Basis and Clinical Manifestations of Common b-Thalassemia Syndromes.

Feature Thalassemia Trait Thalassemia Intermedia Thalassemia Major Hemoglobin E Thalassemia

Genetic pathology One b-globin gene carry- Two b-globin genes carrying Two b-globin genes carrying One b-globin gene carrying
ing a thalassemia mu- a thalassemia mutation, a severe thalassemia mu- a thalassemia mutation
tation at least one of which is tation (mild or severe) in com-
mild; one b-globin thal- bination with a b-globin
assemia mutation in gene carrying the point
combination with excess mutation encoding he-
a-globin genes (less moglobin E
common)
Clinical manifesta- Mild or no anemia, with Mild to moderate anemia; Severe anemia requiring reg- Mild to severe anemia;
tions variable microcytosis relative independence ular transfusions begin- relative dependence
(mean corpuscular from transfusions; prom- ning in infancy; spleno- on transfusions; spleno-
volume, 60 to normal); inent splenomegaly and megaly and bone disease megaly and bony defor-
no splenomegaly; no bone deformities; varia- depending on the effica- mities; variable degree
bone disease ble degrees of iron over- cy of transfusion therapy; of iron overload, de-
load depending on sever- severe iron overload pending on severity of
ity of anemia and trans- anemia and transfusion
fusion requirement requirement
Severity Asymptomatic From asymptomatic to se- Lifelong supportive care re- From asymptomatic to se-
verely symptomatic quired verely symptomatic
Ameliorating genet- Presence of concurrent Presence of concurrent Presence of concurrent Mild b-thalassemia muta-
ic factors a-thalassemia a-thalassemia; elevated a-thalassemia; elevated tion; concurrent a-thal-
hemoglobin F hemoglobin F assemia; elevated he-
moglobin F
Exacerbating genet- Excess a-globin genes Number of excess a-globin None known Severe b-thalassemia
ic factors genes (five or more) mutation

associated with a rise in extracellular exposure of
phosphatidylserine (Fig. 2), an important signal for
removal by activated macrophages,14 whose num-
bers are increased in thalassemic bone marrow.21
clinical manifestations
an d supportive therapy
anemia and transfusion therapy
Regular transfusion therapy to maintain hemo-
globin levels of at least 9 to 10 g per deciliter allows
for improved growth and development and also
reduces hepatosplenomegaly due to extramedullary
hematopoiesis as well as bone deformities.2,3 Ta-
ble 2 summarizes some of the advances in transfu-
sion therapy.
endocrinopathies and bone disease
Impairment of growth and endocrinopathies, par-
ticularly hypogonadism, are common features of
thalassemia.2,23,24 Since these manifestations (Fig.
1) result from chronic anemia as well as from iron
overload, they are much more common in older
patients or those whose chelation therapy is insuffi-
cient (discussed below).2,23 Hormonal replace-
ment is indicated for residual endocrine insuffi-
ciency.24 Growth hormone therapy has had variable
success.23 Hypogonadotropic hypogonadism im-
pairs fertility but can be corrected with the use of
hormonal replacement in male patients. A small
number of female patients, including those with
thalassemia major or thalassemia intermedia, have
been able to become pregnant, either spontane-
ously (if they have received adequate chelation ther-
apy) or with assisted reproductive techniques.23
Pregnancy generally appears safe if baseline car-
diac function is good.25
Considerable morbidity in older patients re-
sults from bone disease due to osteopenia and os-
teoporosis, which is often accompanied by disabling
pain and fractures. The pathogenesis is complex
and multifactorial. Bone marrow expansion due to
ineffective erythropoiesis, endocrine dysfunction,
and complications of treatment all contribute to
the condition.26 Overly vigorous chelation is asso-
ciated with deferoxamine-induced bone dysplasia,
which can slow growth velocity in children and may
be only partially reversible.24 Bone-disease manage-
ment includes the careful monitoring of chelation,
lifestyle adjustments (increased calcium intake and
physical activity and refraining from smoking), hor-
monal therapy, and vitamin D therapy. Osteoclast

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 The new england journal
inhibitors such as bisphosphonates have the po-
tential to reduce bone resorption and may be a valu-
able treatment approach, but further study is re-
quired before the routine use of these drugs can be
recommended.26
iron overload — pathogenesis,
measurement, and therapy
Iron overload causes most of the mortality and mor-
bidity associated with thalassemia. Iron deposi-
tion occurs in visceral organs (mainly in the heart,
liver, and endocrine glands), causing tissue dam-
age and ultimately organ dysfunction and failure
(Fig. 1). Cardiac events due to iron overload are still
the primary cause of death.27,28 Both transfusional
iron overload and excess gastrointestinal absorp-
tion are contributory. Paradoxically, excess gastro-
intestinal iron absorption persists despite massive
increases in total body iron load.
Hepcidin is a small peptide that inhibits iron ab-
sorption in the small bowel. Hepcidin levels nor-
mally increase when iron stores are elevated.29 Hep-
cidin levels were found to be inappropriately low in
patients with thalassemia intermedia and thalas-
semia major.30 Furthermore, serum from patients
with thalassemia inhibited hepcidin messenger
RNA expression in the HepG2 cell line, which sug-
gests the presence of a humoral factor that down-
regulates hepcidin.31 These observations suggest
that the administration of hepcidin or agents that
increase hepcidin expression may be therapeuti-
cally useful for the inhibition of inappropriate iron
absorption.
Accurate, preferably noninvasive measurement
of iron stores is crucial for the evaluation and man-
agement of chelation therapy. Serum ferritin is
most commonly measured as an indicator of iron
stores. Ferritin levels below 2500 mg per milliliter
are associated with improved survival, free of car-
diac disease.32 However, serum ferritin levels are
highly unreliable, particularly when liver disease
is present.33 Liver biopsy has often been used but is
invasive. Direct noninvasive measurement of he-
patic iron stores is possible with the technique of
magnetic susceptometry (with the use of a super-
conducting quantum-interference device) and is
either equivalent to or more accurate than the mea-
surement of hepatic iron by liver biopsy.28 However,
only four centers worldwide currently have this
capacity.
It is noteworthy that hepatic iron may not accu-
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of medicine
Figure 2 (facing page). Pathophysiological Features
of Hemolysis and Hypercoagulability.
Oxidation of a-, b-, or g-hemoglobin subunits leads to
the formation of hemichromes, whose rate of formation
determines the rate of hemolysis.9 Because a chains dis-
sociate into monomers more readily than do b or
g chains, they form hemichromes at a faster rate,10
which explains why b-thalassemia is clinically much
more severe than a-thalassemia. Precipitates of un-
paired b chains form single large inclusions known as
Heinz bodies.11 This process occurs at a later stage of
maturation than does the formation of smaller multiple
inclusions because of the precipitation of unpaired a
chains. Hemichromes bind to or modify various compo-
nents of the mature red-cell membrane, such as protein
band 3, protein 4.1, ankyrin, and spectrin.12 After precipi-
tation of hemichromes, heme disintegrates, and toxic
non–transferrin-bound iron species are released.13 The re-
sulting free iron catalyzes the formation of reactive oxygen
species.9 Iron-dependent oxidation of membrane proteins
and formation of red-cell “senescence” antigens such as
phosphatidylserine14 cause thalassemic red cells to be rig-
id and deformed and to aggregate, resulting in premature
cell removal.15 Phosphatidylserine is also involved in the
activation of the coagulation system.16
rately represent iron deposition in other vital organs
(such as the heart). Indeed, severe cardiac damage
has been observed in a few patients with presum-
ably adequate chelation, and myocardial iron and
left ventricular function apparently cannot be pre-
dicted from liver iron concentration, ferritin levels,
or both.34 Therefore, noninvasive techniques for
the measurement of cardiac iron levels are being
developed. Magnetic resonance imaging (MRI) for
the measurement of cardiac iron is technically prob-
lematic. However, the application of T2 gradient-
echo sequencing is more sensitive to hemosiderin
deposition and appears to be useful for the mea-
surement of myocardial iron in thalassemia,35 but
this approach requires further validation and long-
term studies to determine its usefulness in assess-
ing the effectiveness of chelation therapy.
Elevated tissue iron stores are only one com-
ponent of the damaging effects of iron overload.
A highly toxic form of iron, non–transferrin-bound
iron, is formed when the iron-binding capacity of
transferrin has been exceeded. Non–transferrin-
bound iron is highly toxic because it can catalyze
the formation of reactive oxygen species through
the Fenton reaction.9 A fraction of non–transfer-
rin-bound iron, the labile plasma iron, can be mea-
sured directly and may serve as a clinically useful
test for monitoring iron-chelation therapy.36
Iron-chelation therapy is largely responsible for
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 medical progress

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 The new england journal
Table 2. Improvements in Supportive Care of b-Thalassemia.*
Treatment Approach
Transfusion
Definition of the optimal end point of transfusion
(a hemoglobin level of 9–10 rather than 10–12)
Use of leukodepletion techniques
Testing for viruses (including hepatitis B and C,
HIV, and HTLV-I)
Implantation of venous-access device
Chelation
Individualization of doses of deferoxamine
Development of oral chelators and combined
chelation therapy
Endocrine support
Administration of hormone replacement (sex,
thyroid, and growth hormones)
Administration of fertility agents
Administration of osteoclast inhibitors
* HIV denotes human immunodeficiency virus, and HTLV-I human T-cell lymphotropic virus type I.
doubling the life expectancy of patients with thal-
assemia major.28,32,37 Deferoxamine continues to
be the most common iron-chelating agent in use,
but it has several limitations: the need for paren-
teral administration (which is painful and reduces
compliance), side effects, and cost (which is pro-
hibitive in underdeveloped countries).28
Much effort has been invested in the develop-
ment of new orally active chelators. Deferiprone,
an orally administered chelator, was initially thought
to be an inadequate chelator that might worsen he-
patic fibrosis. However, cumulative worldwide ex-
perience indicates that the drug is safe and effec-
tive.38 Long-term administration of deferiprone
does not appear to be associated with liver dam-
age.39 Adverse effects of deferiprone include ar-
thralgia, nausea and other gastrointestinal symp-
toms, fluctuating liver enzyme levels, leukopenia,
and rarely agranulocytosis and zinc deficiency.40
Most of these effects can be monitored and con-
trolled.
Deferiprone has a number of advantages over
deferoxamine. It can penetrate the cell membrane
and chelate toxic intracellular iron species.41 In a
preliminary study, hemoglobin levels increased and
transfusion requirements decreased in a few patients
with hemoglobin E thalassemia who were treated
with deferiprone for an average of 50 weeks.42 Most
important, recent evidence suggests that defer-
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of medicine
Outcome
Less iron overload and alloimmunization
Fewer febrile nonhemolytic transfusion reactions, less
transmission of cytomegalovirus, less alloimmunization22
Substantial reduction in transfusion-transmitted infections
More effective therapy and improved patient comfort
and compliance
Fewer untoward effects (hearing loss, bone dysplasia)
Improved compliance and efficacy
Improvement in growth, development, and sexual maturation;
prevention of osteoporosis
Induction of spermatogenesis; achievement of pregnancy
Improvement in osteopenia, osteoporosis, and quality of life
iprone may be more effective than deferoxamine in
the removal of myocardial iron.34,43,44
An encouraging new approach to chelation
therapy is the sequential combined administration
of deferiprone and deferoxamine. Experimental evi-
dence suggests that intracellular iron chelated by
deferiprone is transferred in the plasma to the
more powerful chelator, deferoxamine (the so-called
“shuttle hypothesis”).45 As a consequence, more
iron is excreted with the use of combined therapy
than with the separate administration of each drug.
Furthermore, the compliance of patients was im-
proved with the use of combined therapy because
fewer painful injections of deferoxamine were re-
quired.43,46,47
A number of new oral iron chelators are under
development.25 Deferasirox (ICL670) is particu-
larly promising for its efficacy, which may be sim-
ilar to that of deferoxamine.44,48 Deferasirox is
administered once daily and appears to have an
acceptable side-effect profile.44,49,50 Toxic effects
that have been observed have been related mainly
to iron deprivation and transient gastrointestinal
symptoms. No cases of agranulocytosis have been
reported in several phase 2 trials involving several
hundred patients.49,50
In summary, a growing body of evidence sug-
gests that deferiprone is an acceptable alternative
for patients who are unable to receive deferoxa-
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 medical progress
mine.33 The combination of deferiprone and de-
feroxamine appears very promising but requires
further verification. The preliminary data on de-
ferasirox are encouraging, and long-term clinical
trials are still required. Finally, improved noninva-
sive technologies (including imaging and blood
tests) for quantitation of iron overload will provide
reliable information for the assessment of the effi-
cacy of present and future therapies.
hypercoagulability
Thromboembolic phenomena, both venous and
arterial, are not uncommon in patients with thalas-
semia, particularly in patients who have undergone
splenectomy and who undergo transfusion infre-
quently.51 Abnormalities in the levels of coagula-
tion factors and their inhibitors have been report-
ed, resulting in what can be defined as a chronic
hypercoagulable state.16
Erythrocyte-membrane abnormalities contrib-
ute to hypercoagulability (Fig. 2). Membrane lipid
peroxidation increases the surface expression of an-
ionic phospholipids such as phosphatidylserine.14
Exposure of phosphatidylserine on the erythrocyte
was highly correlated with the expression of plate-
let activation markers.16 Erythrocytes that are ex-
posed to phosphatidylserine may also contribute
directly to the vascular damage observed in thalas-
semia.52 In addition, erythrocytes and platelets from
patients with thalassemia contain higher levels of
reactive oxygen species and lower levels of intracel-
lular glutathione than do normal erythrocytes and
platelets, and this finding may be attributed to con-
tinuous exposure to oxidative insults.53 Further
studies will be required before conclusive recom-
mendations can be made for prophylactic antico-
agulation, antiplatelet therapy, or both for patients
with thalassemia who are at risk (during pregnancy
or in the postoperative period) or on a routine basis,
particularly in patients who have undergone sple-
nectomy.
hematopoietic stem- cell
transplantation
Although hematopoietic stem-cell transplantation
is the only available curative approach for thalasse-
mia, it has been limited by the high cost and the
scarcity of HLA-matched, related donors. The past
several years have brought progress in the realms
of conditioning regimens, donor identification and
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selection, and the development of alternative sourc-
es of hematopoietic stem cells.
Low-risk patients (those with thalassemia termed
class 1 or class 2 by the Lucarelli classification, which
is used to assess risk factors that predict outcome
and prognosis and addresses the degree of hepa-
tomegaly, the presence of portal fibrosis on liver
biopsy, and the effectiveness of chelation therapy
before transplantation) have had excellent results
after bone marrow transplantation; however, pa-
tients with class 3 disease (with extensive liver dam-
age from iron overload) have had poor results in
the past, primarily because of the 30 percent rate of
graft rejection due to attenuated conditioning.54 A
new preparatory regimen (including hydroxyurea,
azathioprine, fludarabine, busulfan, and cyclophos-
phamide) has substantially improved the results in
patients with class 3 disease who are younger than
17 years of age. The survival rate among these pa-
tients was 93 percent, and the rejection rate fell to
8 percent.55
Because of the difficulty of eradicating the en-
dogenous thalassemic bone marrow, it has been
considered essential to administer full myeloab-
lative conditioning regimens for transplantation.
Nonmyeloablative regimens have rarely been at-
tempted.56,57 Follow-up is short, and it is unclear
whether this approach will be beneficial. However,
in contrast to what may occur after bone marrow
transplantation for cancer, long-term mixed chi-
merism can sometimes result in an acceptable clin-
ical outcome in thalassemia as long as the anemia
is corrected.58
An increase in the available donor pool for bone
marrow transplantation has been achieved by us-
ing matched, unrelated donors.59 Extended haplo-
typing has been developed for unrelated donors,
and the resulting outcomes were similar to those
using matched, related donors.60 The use of relat-
ed or unrelated umbilical-cord blood further in-
creases the donor pool. However, cord-blood trans-
plantations have often been unsuccessful in the
treatment of thalassemia because large numbers of
transplanted cells need to be administered to sus-
tain hematopoiesis and prevent graft rejection. In
one study using related cord blood,61 7 of 33 pedi-
atric patients rejected grafts and 3 others exhibited
stable mixed chimerism. Another study reported a
high rate of nonengraftment and secondary rejec-
tion: only four of nine children were transfusion-
independent at a median follow-up period of 49
months.62 Rare case reports have described a suc-
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 The new england journal of medicine

cessful outcome with unrelated cord-blood trans- erythropoiesis” without raising fetal hemoglobin.
plantation.63,64 In the future, umbilical-cord-blood The effect appeared to be dose-dependent and was
transplantation may be more successful if stem cells observed primarily in patients with b-thalassemia
can be expanded ex vivo. intermedia who had undergone splenectomy.71 Re-
In summary, hematopoietic stem-cell transplan- cently, long-acting darbepoetin alfa was shown to
tation with the use of related or unrelated donors increase hemoglobin levels substantially in patients
is a viable alternative that generally results in an ex-
with hemoglobin E–b-thalassemia disease.72 Two
cellent outcome for low-risk patients. If transplan- important obstacles to the use of recombinant hu-
tation is successful, transfusions, and usually chela- man erythropoietin are its relatively high cost and its
tion therapy, are no longer necessary. There is a subcutaneous administration route, which restrict
small risk of serious complications (death, graft its use in developing countries. Appropriate clinical
failure or rejection, and graft-versus-host disease). protocols are needed to delineate the role of recom-
Furthermore, growth failure and endocrinopathies, binant human erythropoietin (alone or in combina-
particularly gonadal dysfunction, can still occur.23,65tion with the aforementioned drugs) in the treat-
All these factors, in addition to the availability of ad-
ment of b-thalassemia.
equate supportive care in various geographical re- Screening for new compounds that augment fe-
gions, must be considered when deciding whether tal hemoglobin production can be performed with
to perform transplantation in any given patient. the use of cell-culture techniques, and various ani-
mal models have been useful in the evaluation of
experimental therapies potential fetal hemoglobin inducers.73 Further-
more, perturbations of growth-related signaling
cellular and molecular modifiers appear to activate tissue-specific fetal genes.74 Ex-
Augmenting the synthesis of fetal hemoglobin ploration of these signaling pathways may lead to
should ameliorate the severity of b-thalassemia.1 the discovery of new pharmacologic agents for the
Drugs such as 5-azacytidine, hydroxyurea, and var- treatment of thalassemia.
ious butyrate derivatives have been used for this pur-
pose.66 Hydroxyurea has shown substantial ben- potential role of antioxidants
efits in a subgroup of patients with sickle cell Because reactive oxygen species play an important
anemia67 but has been used less often in thalasse- role in the pathophysiology of thalassemia9 (Fig. 2),
mia. In a few pediatric patients with thalassemia, antioxidants may be an effective therapy. Patients
transfusion requirements were eliminated after with thalassemia have very high plasma levels of
treatment with hydroxyurea for approximately 20 malonyldialdehyde, a by-product of lipid peroxida-
months.68 In general, preliminary results among a tion. Malonyldialdehyde levels correlate positively
relatively small number of patients have been incon- with serum iron and with non–transferrin-bound
sistent, and thus the role of hydroxyurea in thalas- iron.75 Elevated levels of reactive oxygen species
semia therapy remains uncertain.66 tended to normalize in response to oral therapy with
One possible explanation for the differential ef- vitamin E, with patients exhibiting improvement in
fects of hydroxyurea in sickle cell anemia as com- the antioxidant–oxidant balance in plasma and
pared with thalassemia is that many patients with decreased lipid peroxidation in erythrocytes.76 Plant
thalassemia are transfusion-dependent. Hypertrans- flavonoids (including rutin and curcumin) are an-
fusion suppresses endogenous erythropoiesis, par- other group of antioxidants with therapeutic po-
ticularly of hydroxyurea-responsive types of cells.67 tential in thalassemia. These compounds have salu-
Therefore, despite the beneficial effects of hy- tary effects on erythrocytes that have been damaged
droxyurea on erythropoiesis,66 it will be difficult to by oxidation. Polyphenols (a major component of
correct the anemia associated with thalassemia with tea) bind to ferric iron and could also protect thal-
the use of hydroxyurea. Moreover, genetic predis- assemic erythrocytes from oxidation.77 However,
position, such as the presence of Xmnl polymor- despite their apparent salutary effects on erythro-
phism,69 and the type of thalassemia, such as hemo- cytes, antioxidants have not yet been shown to ame-
globin E thalassemia,70 may determine the response liorate the anemia of patients with thalassemia.
to hydroxyurea treatment. Antioxidants may be more effective if used in
Recombinant human erythropoietin was shown combination — for example, as a lipid antioxidant
to provide the benefit of increasing “thalassemic like vitamin E, together with N-acetylcysteine, which

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 medical progress
is a protein antioxidant that improves several mea-
sures in oxidized sickle erythrocytes,78 and an iron
chelator such as deferiprone. This approach, if
successful, could be particularly useful in countries
with limited financial resources.
experimental molecular therapies
Initial efforts at gene therapy were directed against
diseases of the b-globin gene. This therapeutic
strategy involves the insertion of a normally func-
tioning g-globin or b-globin gene into the patient’s
autologous hematopoietic stem cells. Although the
concept is relatively straightforward, it has met with
two decades of seemingly insurmountable hurdles
that have been well summarized elsewhere.79,80
The major problems with this type of gene thera-
py have been related to vector construction. The ge-
netic elements of the vector that are necessary for ap-
propriate regulation of the inserted gene have been
defined.79 However, the therapeutic gene must be
inserted into a hematopoietic stem cell and must be
expressed at high levels, over an extended period, in
an erythroid-specific manner. In addition, the vector
must be safe from recombination or mutagenesis.
Oncoretroviral and adenoviral vectors have been
found to be unsuitable for various reasons.80
The introduction of lentiviral vectors was an
important advance, since these viruses do not re-
quire cell division for entry into eukaryotic cells
and can stably hold larger DNA inserts without re-
arrangements.81,82 Self-inactivating lentiviral vec-
tors were constructed to address safety issues.83
Longer locus-control-region elements84 may abro-
gate position effects that reduce expression of the
therapeutic gene. The problem of gene silencing of
the transduced gene85 has been approached by the
use of insulators, which are DNA sequences that
are thought to function as boundary elements pro-
tecting against chromatin-dependent repression
of genetic activity.86,87 A lentiviral vector carrying
both the b-globin gene and an insulator stably cor-
rected the b-thalassemia phenotype in human b-
thalassemia progenitor cells that were transplant-
ed into immune-deficient mice.88 However, a re-
cent report demonstrated that lentiviral constructs
preferentially integrated at intragenic sites, often
within genes that regulate hematopoiesis.89 Be-
sides being associated with lower expression lev-
els,87 intragenic integration raises new concerns
regarding the safety of these vectors.89
Small interfering RNA is the basis for a new
strategy to augment transduced b-globin expres-
n engl j med 353;11 www.nejm.org
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sion. Small interfering RNA corresponding to tran-
scripts of BP1 (a protein that negatively regulates
b-globin expression by binding to its upstream re-
gion) enhanced b-globin promoter activity in eryth-
roid cells.90
Another new molecular approach uses so-called
antisense technology for correction of the molec-
ular defect caused by the thalassemia mutation. Syn-
thesis of hemoglobin A was restored and alternate
splicing corrected with the use of antisense oligo-
nucleotides that blocked splicing at the mutant
alternate splice site, thereby forcing the splicing
machinery to select the normal splice site.91 In addi-
tion, lentiviral vectors carrying an altered U7 small
nuclear RNA gene encoding an antisense RNA have
been shown to correct splicing defects caused by
thalassemia mutations.92
Clearly, these types of therapies remain highly
experimental; accordingly, their clinical potential
remains uncertain. Nevertheless, these methods
may be useful molecular approaches for the devel-
opment of new therapies for thalassemias.
progress in prenatal diagnosis
Polymerase-chain-reaction (PCR) technology has
been used for more than a decade to detect point
mutations or deletions in chorionic-villus samples,
enabling first-trimester, DNA-based testing for thal-
assemia. However, because pregnancy termination
is unacceptable to some persons (even when the
fetus is affected), methods were developed, begin-
ning in the early 1990s, to perform diagnostic test-
ing before implantation. Preimplantation genetic
diagnosis93 involves performing conventional in
vitro fertilization, followed by removal of one or
two cells from the resulting blastomeres on day 3.
PCR is then used to detect thalassemia mutations
within the cells that have been removed so that un-
affected blastomeres may be selected for implanta-
tion. Preimplantation genetic diagnosis requires a
high degree of technical expertise. Furthermore,
the phenomenon of “allele dropout” (failure to am-
plify one of the two alleles in a heterozygous cell)
can result in diagnostic errors.94 Nonetheless, this
technology has been successful, and improvements
in outcome have led to its use in many countries.
Recently, preimplantation genetic diagnosis has
been extended to HLA typing on embryonic biop-
sies, which allows the selection of an embryo that
is not affected by thalassemia and that may also
serve as a stem-cell donor for a previously affected
september 15, 2005 1143
 The new england journal of medicine

child within the same family. Recent reports95,96
have confirmed that this approach is feasible. How-
ever, serious ethical concerns have been raised.93
Although it is considered ethical not to implant an
embryo that is affected with a serious genetic dis-
order, in certain countries it is forbidden to select
an embryo on the basis of its designated role as a
potential stem-cell donor.
Future prenatal diagnosis may be performed
noninvasively, with the use of maternal blood sam-
ples to isolate either fetal cells97 or fetal DNA98 for
analysis. These techniques are feasible but have
not yet been perfected. Furthermore, techniques
using DNA in maternal plasma to exclude thalas-
semia in the fetus are applicable only to couples
for whom the paternal and maternal mutations
are different.
treatment in the developed
versus the underdeveloped
world
In developed parts of the world, such as the United
States and Europe, there are approximately 10,000
homozygous patients with thalassemia, and the
number of new cases has been progressively de-
creasing because of effective prevention methods.99
Comprehensive, high-quality medical care is avail-
able in these countries, with longer life expectancy
and a relatively good quality of life.27,37 Western
countries emphasize curative forms of therapy, such
as bone marrow transplantation and gene therapy,
which require compliant patients with access to the
newest medications and sophisticated scientific fa-
cilities. Western cultures need to develop improved
support for patients with thalassemia and their fam-
ilies. Doing so will prevent psychosocial issues from
taking their heavy toll due to noncompliance.100
In contrast, the treatment of thalassemia is en-
tirely different in less developed countries, where
most of the patients with this disease reside. Safe
transfusion (with the use of filtration and the viral
testing of blood) and chelation are not universally
available. Consequently, many patients with thal-
assemia in underdeveloped nations die in child-
hood or adolescence. Programs that provide ac-
ceptable care, including transfusion of safe blood
and supportive therapy including chelation, must
be established. Thalassemia-prevention protocols
must be developed in these countries, with the use
of better education and screening and improved
access to prenatal diagnosis.101,102 The challenge
for the future is to ensure that people who are
born with a severe form of thalassemia will con-
tinue to thrive, while effective prevention eventu-
ally decreases the number of severely affected pa-
tients worldwide.
We are indebted to Drs. Griffin Rodgers and Eitan Fibach for their
critical reading of the manuscript.

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