Thalassemia majorclinical aspects and management
Prof Renzo GalanelloPediatric Clinic 2deg University of CagliariOspedale Regionale Microcitemie-ASL8
Clinical characteristics ofβ talassemia major
severe early-onset hypochromicmicrocytic anemialiver and spleen enlargementfailure to thrivesevere skeletal manifestationregular transfusions required
(transfusion dependent)
DIAGNOSIS OF HOMOZYGOUS BETA THALASSEMIA
DIAGNOSIS OF HOMOZYGOUS BETA THALASSEMIA
cbc
Hb electrophoresisHPLC
DNA globin gene analysis
globin chain synthesis
cbc
Hb electrophoresisHPLC
DNA globin gene analysis
globin chain synthesis
cellulose acteate electrophoresis (pH 86)cellulose acteate electrophoresis (pH 86)
origin
HbXHbFHbA
HbA2
-
+
Peripheral blood smear in β-thalassemia major (A) intermedia (B) and in heterozygous β-thalassemia (C)
Peripheral blood smear in β-thalassemia major (A) intermedia (B) and in heterozygous β-thalassemia (C)
AA
BB
CC
Figure 7
7
Management of Thalassemia Majorbull Conventional
ndash RBC transfusionsndash Iron chelationndash (Splenectomy)
bull Bone marrow transplantationbull Prospectives
ndash gene therapyndash Hb F induction
8
AIMS OF TRANSFUSION TREATMENTbull Normal growth and physical activitybull darr Chronic hypoxemiabull darr Compensatory marrow hyperplasia
ndash darr hypervolemiandash darr bone changes ndash darr extramedullary erythropoiesis
bull darr Splenomegaly and hypersplenismbull darr Gastro-intestinal iron absorption
Improvementof cardiacfunction
9
Guidelines For RBC TransfusionIn Thalassemia Major
lt10 mlkghRate
2-4 weeksInterval
lt20 mlKgVolume
lt140 gdLPost-transfusional hemoglobin
95-105 gdLPre-transfusional hemoglobin
10
0
50
100
150
200
250
300
350
400
0 20 40 60 80 100PATIENT WEIGHT kg
Hct 80
Hct 60
Hct 75
Hct 50
ml
AMOUNT OF BLOOD REQUIRED TO RAISE AMOUNT OF BLOOD REQUIRED TO RAISE PATIENTPATIENTrsquorsquoS HAEMOGLOBIN BY 1 gdlS HAEMOGLOBIN BY 1 gdl
TIF 2000
11
Transfusional Indices (Annual)
bull Mean hemoglobin bull Pure red cell consumptionbull Iron inputbull Mean transfusional interval
12
bull Row data fromndash transfusionndash chelationndash tests
bull Complicationsbull Therapies
Transfusional indexesIron balanceAppointmentsPrint-outsGraphsCentres Cooperation
A THALASSEMIA-ORIENTED COMPUTERIZED CLINICAL RECORDWEBTHAL
13
Transfusion - Dependent Complications
bull Iron overload
bull Infections
bull Immunization (allergic reactions alloimmunization)
1 ml pure RBC = 116 mg Fe
1 blood unit = 200 mg Fe
To maintain mean Hb = 12 g dl
100-200 mlkgy of pure RBC
Mean iron imput = 03-06 mgkgday
Includes iron absorbed by the GI tract (1 to 4 mg day)
Complications in thalassemia major
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
Clinical characteristics ofβ talassemia major
severe early-onset hypochromicmicrocytic anemialiver and spleen enlargementfailure to thrivesevere skeletal manifestationregular transfusions required
(transfusion dependent)
DIAGNOSIS OF HOMOZYGOUS BETA THALASSEMIA
DIAGNOSIS OF HOMOZYGOUS BETA THALASSEMIA
cbc
Hb electrophoresisHPLC
DNA globin gene analysis
globin chain synthesis
cbc
Hb electrophoresisHPLC
DNA globin gene analysis
globin chain synthesis
cellulose acteate electrophoresis (pH 86)cellulose acteate electrophoresis (pH 86)
origin
HbXHbFHbA
HbA2
-
+
Peripheral blood smear in β-thalassemia major (A) intermedia (B) and in heterozygous β-thalassemia (C)
Peripheral blood smear in β-thalassemia major (A) intermedia (B) and in heterozygous β-thalassemia (C)
AA
BB
CC
Figure 7
7
Management of Thalassemia Majorbull Conventional
ndash RBC transfusionsndash Iron chelationndash (Splenectomy)
bull Bone marrow transplantationbull Prospectives
ndash gene therapyndash Hb F induction
8
AIMS OF TRANSFUSION TREATMENTbull Normal growth and physical activitybull darr Chronic hypoxemiabull darr Compensatory marrow hyperplasia
ndash darr hypervolemiandash darr bone changes ndash darr extramedullary erythropoiesis
bull darr Splenomegaly and hypersplenismbull darr Gastro-intestinal iron absorption
Improvementof cardiacfunction
9
Guidelines For RBC TransfusionIn Thalassemia Major
lt10 mlkghRate
2-4 weeksInterval
lt20 mlKgVolume
lt140 gdLPost-transfusional hemoglobin
95-105 gdLPre-transfusional hemoglobin
10
0
50
100
150
200
250
300
350
400
0 20 40 60 80 100PATIENT WEIGHT kg
Hct 80
Hct 60
Hct 75
Hct 50
ml
AMOUNT OF BLOOD REQUIRED TO RAISE AMOUNT OF BLOOD REQUIRED TO RAISE PATIENTPATIENTrsquorsquoS HAEMOGLOBIN BY 1 gdlS HAEMOGLOBIN BY 1 gdl
TIF 2000
11
Transfusional Indices (Annual)
bull Mean hemoglobin bull Pure red cell consumptionbull Iron inputbull Mean transfusional interval
12
bull Row data fromndash transfusionndash chelationndash tests
bull Complicationsbull Therapies
Transfusional indexesIron balanceAppointmentsPrint-outsGraphsCentres Cooperation
A THALASSEMIA-ORIENTED COMPUTERIZED CLINICAL RECORDWEBTHAL
13
Transfusion - Dependent Complications
bull Iron overload
bull Infections
bull Immunization (allergic reactions alloimmunization)
1 ml pure RBC = 116 mg Fe
1 blood unit = 200 mg Fe
To maintain mean Hb = 12 g dl
100-200 mlkgy of pure RBC
Mean iron imput = 03-06 mgkgday
Includes iron absorbed by the GI tract (1 to 4 mg day)
Complications in thalassemia major
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
DIAGNOSIS OF HOMOZYGOUS BETA THALASSEMIA
DIAGNOSIS OF HOMOZYGOUS BETA THALASSEMIA
cbc
Hb electrophoresisHPLC
DNA globin gene analysis
globin chain synthesis
cbc
Hb electrophoresisHPLC
DNA globin gene analysis
globin chain synthesis
cellulose acteate electrophoresis (pH 86)cellulose acteate electrophoresis (pH 86)
origin
HbXHbFHbA
HbA2
-
+
Peripheral blood smear in β-thalassemia major (A) intermedia (B) and in heterozygous β-thalassemia (C)
Peripheral blood smear in β-thalassemia major (A) intermedia (B) and in heterozygous β-thalassemia (C)
AA
BB
CC
Figure 7
7
Management of Thalassemia Majorbull Conventional
ndash RBC transfusionsndash Iron chelationndash (Splenectomy)
bull Bone marrow transplantationbull Prospectives
ndash gene therapyndash Hb F induction
8
AIMS OF TRANSFUSION TREATMENTbull Normal growth and physical activitybull darr Chronic hypoxemiabull darr Compensatory marrow hyperplasia
ndash darr hypervolemiandash darr bone changes ndash darr extramedullary erythropoiesis
bull darr Splenomegaly and hypersplenismbull darr Gastro-intestinal iron absorption
Improvementof cardiacfunction
9
Guidelines For RBC TransfusionIn Thalassemia Major
lt10 mlkghRate
2-4 weeksInterval
lt20 mlKgVolume
lt140 gdLPost-transfusional hemoglobin
95-105 gdLPre-transfusional hemoglobin
10
0
50
100
150
200
250
300
350
400
0 20 40 60 80 100PATIENT WEIGHT kg
Hct 80
Hct 60
Hct 75
Hct 50
ml
AMOUNT OF BLOOD REQUIRED TO RAISE AMOUNT OF BLOOD REQUIRED TO RAISE PATIENTPATIENTrsquorsquoS HAEMOGLOBIN BY 1 gdlS HAEMOGLOBIN BY 1 gdl
TIF 2000
11
Transfusional Indices (Annual)
bull Mean hemoglobin bull Pure red cell consumptionbull Iron inputbull Mean transfusional interval
12
bull Row data fromndash transfusionndash chelationndash tests
bull Complicationsbull Therapies
Transfusional indexesIron balanceAppointmentsPrint-outsGraphsCentres Cooperation
A THALASSEMIA-ORIENTED COMPUTERIZED CLINICAL RECORDWEBTHAL
13
Transfusion - Dependent Complications
bull Iron overload
bull Infections
bull Immunization (allergic reactions alloimmunization)
1 ml pure RBC = 116 mg Fe
1 blood unit = 200 mg Fe
To maintain mean Hb = 12 g dl
100-200 mlkgy of pure RBC
Mean iron imput = 03-06 mgkgday
Includes iron absorbed by the GI tract (1 to 4 mg day)
Complications in thalassemia major
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
cellulose acteate electrophoresis (pH 86)cellulose acteate electrophoresis (pH 86)
origin
HbXHbFHbA
HbA2
-
+
Peripheral blood smear in β-thalassemia major (A) intermedia (B) and in heterozygous β-thalassemia (C)
Peripheral blood smear in β-thalassemia major (A) intermedia (B) and in heterozygous β-thalassemia (C)
AA
BB
CC
Figure 7
7
Management of Thalassemia Majorbull Conventional
ndash RBC transfusionsndash Iron chelationndash (Splenectomy)
bull Bone marrow transplantationbull Prospectives
ndash gene therapyndash Hb F induction
8
AIMS OF TRANSFUSION TREATMENTbull Normal growth and physical activitybull darr Chronic hypoxemiabull darr Compensatory marrow hyperplasia
ndash darr hypervolemiandash darr bone changes ndash darr extramedullary erythropoiesis
bull darr Splenomegaly and hypersplenismbull darr Gastro-intestinal iron absorption
Improvementof cardiacfunction
9
Guidelines For RBC TransfusionIn Thalassemia Major
lt10 mlkghRate
2-4 weeksInterval
lt20 mlKgVolume
lt140 gdLPost-transfusional hemoglobin
95-105 gdLPre-transfusional hemoglobin
10
0
50
100
150
200
250
300
350
400
0 20 40 60 80 100PATIENT WEIGHT kg
Hct 80
Hct 60
Hct 75
Hct 50
ml
AMOUNT OF BLOOD REQUIRED TO RAISE AMOUNT OF BLOOD REQUIRED TO RAISE PATIENTPATIENTrsquorsquoS HAEMOGLOBIN BY 1 gdlS HAEMOGLOBIN BY 1 gdl
TIF 2000
11
Transfusional Indices (Annual)
bull Mean hemoglobin bull Pure red cell consumptionbull Iron inputbull Mean transfusional interval
12
bull Row data fromndash transfusionndash chelationndash tests
bull Complicationsbull Therapies
Transfusional indexesIron balanceAppointmentsPrint-outsGraphsCentres Cooperation
A THALASSEMIA-ORIENTED COMPUTERIZED CLINICAL RECORDWEBTHAL
13
Transfusion - Dependent Complications
bull Iron overload
bull Infections
bull Immunization (allergic reactions alloimmunization)
1 ml pure RBC = 116 mg Fe
1 blood unit = 200 mg Fe
To maintain mean Hb = 12 g dl
100-200 mlkgy of pure RBC
Mean iron imput = 03-06 mgkgday
Includes iron absorbed by the GI tract (1 to 4 mg day)
Complications in thalassemia major
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
Peripheral blood smear in β-thalassemia major (A) intermedia (B) and in heterozygous β-thalassemia (C)
Peripheral blood smear in β-thalassemia major (A) intermedia (B) and in heterozygous β-thalassemia (C)
AA
BB
CC
Figure 7
7
Management of Thalassemia Majorbull Conventional
ndash RBC transfusionsndash Iron chelationndash (Splenectomy)
bull Bone marrow transplantationbull Prospectives
ndash gene therapyndash Hb F induction
8
AIMS OF TRANSFUSION TREATMENTbull Normal growth and physical activitybull darr Chronic hypoxemiabull darr Compensatory marrow hyperplasia
ndash darr hypervolemiandash darr bone changes ndash darr extramedullary erythropoiesis
bull darr Splenomegaly and hypersplenismbull darr Gastro-intestinal iron absorption
Improvementof cardiacfunction
9
Guidelines For RBC TransfusionIn Thalassemia Major
lt10 mlkghRate
2-4 weeksInterval
lt20 mlKgVolume
lt140 gdLPost-transfusional hemoglobin
95-105 gdLPre-transfusional hemoglobin
10
0
50
100
150
200
250
300
350
400
0 20 40 60 80 100PATIENT WEIGHT kg
Hct 80
Hct 60
Hct 75
Hct 50
ml
AMOUNT OF BLOOD REQUIRED TO RAISE AMOUNT OF BLOOD REQUIRED TO RAISE PATIENTPATIENTrsquorsquoS HAEMOGLOBIN BY 1 gdlS HAEMOGLOBIN BY 1 gdl
TIF 2000
11
Transfusional Indices (Annual)
bull Mean hemoglobin bull Pure red cell consumptionbull Iron inputbull Mean transfusional interval
12
bull Row data fromndash transfusionndash chelationndash tests
bull Complicationsbull Therapies
Transfusional indexesIron balanceAppointmentsPrint-outsGraphsCentres Cooperation
A THALASSEMIA-ORIENTED COMPUTERIZED CLINICAL RECORDWEBTHAL
13
Transfusion - Dependent Complications
bull Iron overload
bull Infections
bull Immunization (allergic reactions alloimmunization)
1 ml pure RBC = 116 mg Fe
1 blood unit = 200 mg Fe
To maintain mean Hb = 12 g dl
100-200 mlkgy of pure RBC
Mean iron imput = 03-06 mgkgday
Includes iron absorbed by the GI tract (1 to 4 mg day)
Complications in thalassemia major
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
7
Management of Thalassemia Majorbull Conventional
ndash RBC transfusionsndash Iron chelationndash (Splenectomy)
bull Bone marrow transplantationbull Prospectives
ndash gene therapyndash Hb F induction
8
AIMS OF TRANSFUSION TREATMENTbull Normal growth and physical activitybull darr Chronic hypoxemiabull darr Compensatory marrow hyperplasia
ndash darr hypervolemiandash darr bone changes ndash darr extramedullary erythropoiesis
bull darr Splenomegaly and hypersplenismbull darr Gastro-intestinal iron absorption
Improvementof cardiacfunction
9
Guidelines For RBC TransfusionIn Thalassemia Major
lt10 mlkghRate
2-4 weeksInterval
lt20 mlKgVolume
lt140 gdLPost-transfusional hemoglobin
95-105 gdLPre-transfusional hemoglobin
10
0
50
100
150
200
250
300
350
400
0 20 40 60 80 100PATIENT WEIGHT kg
Hct 80
Hct 60
Hct 75
Hct 50
ml
AMOUNT OF BLOOD REQUIRED TO RAISE AMOUNT OF BLOOD REQUIRED TO RAISE PATIENTPATIENTrsquorsquoS HAEMOGLOBIN BY 1 gdlS HAEMOGLOBIN BY 1 gdl
TIF 2000
11
Transfusional Indices (Annual)
bull Mean hemoglobin bull Pure red cell consumptionbull Iron inputbull Mean transfusional interval
12
bull Row data fromndash transfusionndash chelationndash tests
bull Complicationsbull Therapies
Transfusional indexesIron balanceAppointmentsPrint-outsGraphsCentres Cooperation
A THALASSEMIA-ORIENTED COMPUTERIZED CLINICAL RECORDWEBTHAL
13
Transfusion - Dependent Complications
bull Iron overload
bull Infections
bull Immunization (allergic reactions alloimmunization)
1 ml pure RBC = 116 mg Fe
1 blood unit = 200 mg Fe
To maintain mean Hb = 12 g dl
100-200 mlkgy of pure RBC
Mean iron imput = 03-06 mgkgday
Includes iron absorbed by the GI tract (1 to 4 mg day)
Complications in thalassemia major
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
8
AIMS OF TRANSFUSION TREATMENTbull Normal growth and physical activitybull darr Chronic hypoxemiabull darr Compensatory marrow hyperplasia
ndash darr hypervolemiandash darr bone changes ndash darr extramedullary erythropoiesis
bull darr Splenomegaly and hypersplenismbull darr Gastro-intestinal iron absorption
Improvementof cardiacfunction
9
Guidelines For RBC TransfusionIn Thalassemia Major
lt10 mlkghRate
2-4 weeksInterval
lt20 mlKgVolume
lt140 gdLPost-transfusional hemoglobin
95-105 gdLPre-transfusional hemoglobin
10
0
50
100
150
200
250
300
350
400
0 20 40 60 80 100PATIENT WEIGHT kg
Hct 80
Hct 60
Hct 75
Hct 50
ml
AMOUNT OF BLOOD REQUIRED TO RAISE AMOUNT OF BLOOD REQUIRED TO RAISE PATIENTPATIENTrsquorsquoS HAEMOGLOBIN BY 1 gdlS HAEMOGLOBIN BY 1 gdl
TIF 2000
11
Transfusional Indices (Annual)
bull Mean hemoglobin bull Pure red cell consumptionbull Iron inputbull Mean transfusional interval
12
bull Row data fromndash transfusionndash chelationndash tests
bull Complicationsbull Therapies
Transfusional indexesIron balanceAppointmentsPrint-outsGraphsCentres Cooperation
A THALASSEMIA-ORIENTED COMPUTERIZED CLINICAL RECORDWEBTHAL
13
Transfusion - Dependent Complications
bull Iron overload
bull Infections
bull Immunization (allergic reactions alloimmunization)
1 ml pure RBC = 116 mg Fe
1 blood unit = 200 mg Fe
To maintain mean Hb = 12 g dl
100-200 mlkgy of pure RBC
Mean iron imput = 03-06 mgkgday
Includes iron absorbed by the GI tract (1 to 4 mg day)
Complications in thalassemia major
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
9
Guidelines For RBC TransfusionIn Thalassemia Major
lt10 mlkghRate
2-4 weeksInterval
lt20 mlKgVolume
lt140 gdLPost-transfusional hemoglobin
95-105 gdLPre-transfusional hemoglobin
10
0
50
100
150
200
250
300
350
400
0 20 40 60 80 100PATIENT WEIGHT kg
Hct 80
Hct 60
Hct 75
Hct 50
ml
AMOUNT OF BLOOD REQUIRED TO RAISE AMOUNT OF BLOOD REQUIRED TO RAISE PATIENTPATIENTrsquorsquoS HAEMOGLOBIN BY 1 gdlS HAEMOGLOBIN BY 1 gdl
TIF 2000
11
Transfusional Indices (Annual)
bull Mean hemoglobin bull Pure red cell consumptionbull Iron inputbull Mean transfusional interval
12
bull Row data fromndash transfusionndash chelationndash tests
bull Complicationsbull Therapies
Transfusional indexesIron balanceAppointmentsPrint-outsGraphsCentres Cooperation
A THALASSEMIA-ORIENTED COMPUTERIZED CLINICAL RECORDWEBTHAL
13
Transfusion - Dependent Complications
bull Iron overload
bull Infections
bull Immunization (allergic reactions alloimmunization)
1 ml pure RBC = 116 mg Fe
1 blood unit = 200 mg Fe
To maintain mean Hb = 12 g dl
100-200 mlkgy of pure RBC
Mean iron imput = 03-06 mgkgday
Includes iron absorbed by the GI tract (1 to 4 mg day)
Complications in thalassemia major
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
10
0
50
100
150
200
250
300
350
400
0 20 40 60 80 100PATIENT WEIGHT kg
Hct 80
Hct 60
Hct 75
Hct 50
ml
AMOUNT OF BLOOD REQUIRED TO RAISE AMOUNT OF BLOOD REQUIRED TO RAISE PATIENTPATIENTrsquorsquoS HAEMOGLOBIN BY 1 gdlS HAEMOGLOBIN BY 1 gdl
TIF 2000
11
Transfusional Indices (Annual)
bull Mean hemoglobin bull Pure red cell consumptionbull Iron inputbull Mean transfusional interval
12
bull Row data fromndash transfusionndash chelationndash tests
bull Complicationsbull Therapies
Transfusional indexesIron balanceAppointmentsPrint-outsGraphsCentres Cooperation
A THALASSEMIA-ORIENTED COMPUTERIZED CLINICAL RECORDWEBTHAL
13
Transfusion - Dependent Complications
bull Iron overload
bull Infections
bull Immunization (allergic reactions alloimmunization)
1 ml pure RBC = 116 mg Fe
1 blood unit = 200 mg Fe
To maintain mean Hb = 12 g dl
100-200 mlkgy of pure RBC
Mean iron imput = 03-06 mgkgday
Includes iron absorbed by the GI tract (1 to 4 mg day)
Complications in thalassemia major
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
11
Transfusional Indices (Annual)
bull Mean hemoglobin bull Pure red cell consumptionbull Iron inputbull Mean transfusional interval
12
bull Row data fromndash transfusionndash chelationndash tests
bull Complicationsbull Therapies
Transfusional indexesIron balanceAppointmentsPrint-outsGraphsCentres Cooperation
A THALASSEMIA-ORIENTED COMPUTERIZED CLINICAL RECORDWEBTHAL
13
Transfusion - Dependent Complications
bull Iron overload
bull Infections
bull Immunization (allergic reactions alloimmunization)
1 ml pure RBC = 116 mg Fe
1 blood unit = 200 mg Fe
To maintain mean Hb = 12 g dl
100-200 mlkgy of pure RBC
Mean iron imput = 03-06 mgkgday
Includes iron absorbed by the GI tract (1 to 4 mg day)
Complications in thalassemia major
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
12
bull Row data fromndash transfusionndash chelationndash tests
bull Complicationsbull Therapies
Transfusional indexesIron balanceAppointmentsPrint-outsGraphsCentres Cooperation
A THALASSEMIA-ORIENTED COMPUTERIZED CLINICAL RECORDWEBTHAL
13
Transfusion - Dependent Complications
bull Iron overload
bull Infections
bull Immunization (allergic reactions alloimmunization)
1 ml pure RBC = 116 mg Fe
1 blood unit = 200 mg Fe
To maintain mean Hb = 12 g dl
100-200 mlkgy of pure RBC
Mean iron imput = 03-06 mgkgday
Includes iron absorbed by the GI tract (1 to 4 mg day)
Complications in thalassemia major
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
13
Transfusion - Dependent Complications
bull Iron overload
bull Infections
bull Immunization (allergic reactions alloimmunization)
1 ml pure RBC = 116 mg Fe
1 blood unit = 200 mg Fe
To maintain mean Hb = 12 g dl
100-200 mlkgy of pure RBC
Mean iron imput = 03-06 mgkgday
Includes iron absorbed by the GI tract (1 to 4 mg day)
Complications in thalassemia major
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
1 ml pure RBC = 116 mg Fe
1 blood unit = 200 mg Fe
To maintain mean Hb = 12 g dl
100-200 mlkgy of pure RBC
Mean iron imput = 03-06 mgkgday
Includes iron absorbed by the GI tract (1 to 4 mg day)
Complications in thalassemia major
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
Complications in thalassemia major
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
Associated with iron overloadbull Cardiopathybull Liver diseasebull EndocrinopathiesOthersbull Gallbladderbull Osteoporosisbull Hypersplenismbull Nephrolithiasisbull Thrombosisbull Pulmonary hypertension
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
LiverLiver biopsybiopsy
SQUIDSQUID--magneticmagneticsusceptometrysusceptometry((liverspleenliverspleen))
Serum Serum ferritinferritin
UrinaryUrinary Iron Iron ExcretionExcretion(UIE)(UIE)
MagneticMagnetic ResonanceResonanceImagingImaging ((liverheartotherliverheartotherorgansorgans) )
FreeFree ironironNTBILPI LIPNTBILPI LIP
Quantitative Quantitative phlebotomyphlebotomy
BODYIRON STORES
DETERMINATIONDirect Indirect
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
Royal Bromptonamp Harefield
NHLIImperial College
5ms 6ms 7ms 8ms 9ms 11ms 13ms 15ms 17ms
Deriving Myocardial T2Deriving Myocardial T2Deriving Myocardial T2
020406080
100
0 4 8 12 16 20
TE
Sign
al
Signal = Ke-
TET2
Anderson LJ Eur Heart J 2001 22 2171-9
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
T2 - Cardiac Risk Ranging
High Intermediate Low
Anderson et al 2002
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
Iron balance in thalassemia
input excretion4transfusions
05 mgKgd
4 absorption
1 - 3 mgd
4chelators
urine stool
4 loss 1 mgd
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
OBJECTIVES OF CHELATIONOBJECTIVES OF CHELATION
Reduceprevent iron overload
Detoxify iron
Organ targeting
Reduceprevent iron overload Reduceprevent iron overload
Detoxify ironDetoxify iron
Organ targetingOrgan targeting
Labile Labile IronIron PoolPoolFeFe2+2+ + H+ H22OO22 rarrrarr FeFe3+3+ + + OHOHmacr + OH+ OH
excessexcess ironiron
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
Available iron chelators
Deferoxamine
Deferiprone
Deferasirox
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
Possible Chelation Treatments
Monotherapy
Alternate therapy
Combination therapy simultaneoussequential
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
bullSafety is the issue ofmajor concern for a drugthat patients take longlife
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
YESYESYESYESRareRareGI GI symptomssymptomsrarerareYESYESRareRareNeutropenia Neutropenia
YESYESnonoHigh High dosesdosesRenalRenal changeschanges
RareRarenonononoLensLens opacityopacityYESYESnonononoSkinSkin rashesrashes
nonoYESYESnonoAgranulocytosisAgranulocytosis
nonoYESYESnonoWeightWeight gaingainYESYESYESYESnonoLiverLiver enzymesenzymes changeschanges
nono
YESYES
nono
nono
nono
nono
nono
no no
DEFERIPRONEDEFERIPRONE
RareRareRareRare
YESYESYESYESYESYESYESYESYESYESYESYESYESYESDEFEROXAMINEDEFEROXAMINE ICL670ICL670
nonoLensLens opacityopacity
nonoArthropathyArthropathy
nonoOsteoporosisOsteoporosisRareRareHearingHearing lossloss
nonoRetinalRetinal toxicitytoxicity
nonoYersiniaYersinia infectionsinfections
nonoBoneBone changeschanges
nonoGrowthGrowth arrestarrest
nonoLocalLocal side side effecteffect
SIDE EFFECTS FROM IRON CHELATORSSIDE EFFECTS FROM IRON CHELATORS
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
Consequences of improved conventionaltreatment
Prolonged life expectancyImproved endocrine functionBetter clinical conditionBetter quality of lifeImproved psychosocial status
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
THALASSEMIA MAJOR
Bone marrow transplantation
Gene therapy
HbF induction
Non conventional treatment
1
2
3
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
Thalassemia comprehensive control program
Thalassemia comprehensive control program
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
ndash Preventionraquo community educationraquo population screeningraquo genetic counselingraquo prenatal diagnosisraquo program evaluation
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
MCV flMCV fl
MCH pgMCH pg
Hb patternHb pattern
gt 78
gt 27
A + A2 lt 32
lt 78
lt 27
A + F (01-7) + A2 gt 36
lt 78
lt 27
A + A2 lt 36
ZnPP or iron studies
NORMAL β-THALcarrier uarr ZnPP
darr ironNormal ZnPP
IRON DEFICIENCY
Globin chainsynthesis or
α-globin gene analysis
α-THALcarrier
δ and β genesanalysis
δ + β THAL
γδβ THAL
NORMAL HbA2 β THAL
le 78
le 27
A + F (3-16) + A2 lt 32
HbF quantitαβ ratio
DNA analysis
δ β THAL
HPFH
A + abnormalband + A2
Sickling test
Negative Positive
Furtherinvestigation
HbS
Screening for common mutations by
DNA analysis
Characterization of undefined mutationsby DGGE and direct
sequencing
or
or
or
Flow chart for thalassemia screeningFlow chart for thalassemia screening
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
0
20
40
60
80
100
120
75 77 79 81 83 85 87 89 91 93 95 97 99 01 03 05 07
Year
Ndeg
of p
atie
nts
RefusalRefusal of of prenatalprenatal diagnosisdiagnosisor or refusalrefusal of of pregnancypregnancy interruptioninterruption
ReductionReduction of of homozygoushomozygous ββ--thalassaemiathalassaemia newbornsnewbornsin in SardiniaSardinia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
β-thal alleles (47)β-thal alleles (47)-α(37) initiation cd ACCATGrarr - - CATGα2 ATGrarrACGα1 ATGrarrGTGα2 cd 29 TrarrC (Agrinio)α2 IVS1 (-5nt)α2α1 cd 110 CrarrA (Petah Tikva)α2 cd 142 TrarrA (Icaria)α2 polyA ATAArarrATGA
α-thal alleles (18)α-thal alleles (18) δ-thal allelesδ-thal alleles-65 ArarrG-55 TrarrCcd 4 CrarrTcd 27 GrarrTCorfugrave ndash72 Kb
Hb LeporeHb Lepore
δβ-thal allelesδβ-thal allelesSicilianSpanishTurkishMacedonian
HPFHHPFH
Common structuralvariants
Common structuralvariants
HbSHbCHbD PunjabHbO-Arab
HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN HETEROGENEITY OF HEMOGLOBINOPATHIES IN MEDITERRANEAN
-α37
-α42
-α37 Hb Hasharon-(α)205
--MED-I--MED-II-(α)52
--(MA)--(CL)--(CAL)
HPFH ndash4HPFH ndash5
-196 Aγ C rarrT-117 Aγ G rarrA
Sardinian
5 - CT89 + G30 C1-1 A1-2 A1-2 G1-5 C1-5 T1-110 A1-116 G1-130 C39 T37 A44-C54-T59-A7-C2-705 C2-745 G2-843 G2-844 A2-850-G2-850 C
-90 T-88 A-88 T-87 G-87 T-87 A-86 A-31 C-30 A-30 C6 - A8 - AA27 T1-5 A1-6 C2-1 AAACAAAAATGAAA
-101 T-92 TUTR +10-T+ 33 G2-844 GAATAAG
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
31
Severity ofclinical picture
Diseasecomplications
Genetic Factors Affecting The Severity Of β-thalassemia Phenotype
bull Primary Modifiersndash Variability at globin gene loci
bull Secondary Modifiersndash Variability in other genes
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
32
a globin
β + γglobin
Mild Phenotype
α thalassemia Mildsilent β allelesIncreased γ
Severe Phenotype
a globin
β + γglobin
Mechanisms Of β-Thalassemia Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
33
ComplicationsComplications of betaof beta--thalassemia and thalassemia and geneticgenetic modifiersmodifiers
ComplicationComplication
Hyperbilirubinemia and Hyperbilirubinemia and
gallstonesgallstones
IronIron loadingloading
OsteoporosisOsteoporosisosteopeniaosteopenia
CardiacCardiac diseasedisease
ThrombosisThrombosis
HepatitisHepatitis and and liverliver cirrhosiscirrhosis
Modifier gene(s)
UGT1A1
HFE
ERGVDR Col1A1 TGFβ1
APOE4 HLAGSTM1
Factor V Leiden Prothrombin
HLA
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
PREVALENCE OF CHOLELITHIASIS IN BETA THALASSEMIA
5712035THALASSEMIA INTERMEDIA
20353261THALASSEMIA MAJOR
PREVALENCE
CHOLELITHIASISTOTAL
PATIENTS
Galanello Galanello etet alBJH 2001alBJH 2001
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
5rsquo 3rsquo
A(TA)nTAA
- 53 - 39
TATA box
Exon 1 2 3 4 5
Normal subjects A(TA)6TAA
Subjects with Gilbert syndrome A(TA)7TAA
Normal enzyme activity
Enzyme activity 30 reduction
Mutation of UGT1A1 gene promoter
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
36
0102030405060708090
(TA)6(TA)6 (TA)6(TA)7 (TA)7(TA)7
UGT1-A1 promoter genotype
o
f pat
ient
s w
ith
chol
elith
iasi
sMajorIntermedia
Plt005Plt005
Galanello et al BJH 2001
Prevalence of Cholelithiasis And UGT1-A1 Promoter Genotype In Thalassemia Major
And Intermedia
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