Molecular Characterization and Disease-Related Morbidities of β-Thalassemia Patients from the Northeastern Part of Iraq

Figures & data

Table 1 Demographic and Disease Characteristics of 242 β-Thalassemia Patients

Parameter	Total (242)	NTDT (159)	TDT (83)	P-value
Age (years)
•<18	144 (59.5)	90 (56.6)	54 (65.1)	<0.051
•18–34	82 (33.9)	54 (34.0)	28 (33.7)
•≥35	16 (6.6)	15 (9.4)	1 (1.2)
Splenomegaly	123 (50.8)	93 (58.5)	30 (36.1)	<0.001
Mean spleen size ± SD	13.8 ± 3.5	14.4 ± 3.4	12.5 ± 3.3	<0.001
Splenectomized	83 (34.3)	52 (32.7)	31 (37.3)	0.477
Hepatomegaly	87 (36.0)	60 (37.7)	27 (32.5)	0.26
Mean liver size ± SD	13.8 ± 2.5	13.9 ± 2.5	13.5 ± 2.5	0.261
Serum ferritin (μg/l)
•<1000	148 (61.2)	122 (76.7)	26 (31.3)	<0.001
•1000–2500	61 (25.2)	26 (16.4)	35 (42.2)
•>2500	33 (13.6)	11 (6.9)	22 (26.5)
Treatment
•None transfused	33 (13.6)	33 (20.7)	0 (0)	<0.001
•Occasional transfusion	77 (31.8)	75 (47.2)	2 (2.4)	<0.001
•Regular transfusion	132 (54.5)	51 (32.1)	81 (97.6)	<0.001
•Iron chelation	142 (58.7)	63 (39.6)	79 (95.2)	<0.001
•Hydroxyurea	84 (34.7)	75 (47.2)	9 (10.8)	<0.001
Complications
•Bone disease	162 (66.9)	99 (62.3)	63 (75.9)	0.031
▲ Facial deformity	162 (66.9)	99 (62.3)	63 (75.9)	0.031
▲ Osteoporosis	37 (37)	17 (28.3)	20 (50)	0.023
•Endocrinopathies	78 (32.2)	44 (27.7)	34 (41)	0.079
▲ Growth retardation	49 (34)	25 (27.8)	24 (44.4)	0.122
▲ Hypothyroidism	33 (15.5)	22 (16.8)	11 (13.4)	0.418
▲ Diabetes mellitus	5 (2.1)	0 (0)	5 (6.0)	0.002
•Hepatobiliary disease	70 (28.9)	46 (28.9)	24 (28.9)	0.988
▲ High ALT ≥50 IU/l	31 (12.9)	12 (7.5)	19 (23.2)	0.001
▲ Biliary complications	48 (19.9)	41 (25.8)	7 (8.5)	0.044
•Pulmonary hypertension	24 (9.9)	18 (11.3)	6 (7.2)	0.321
•Thrombosis	2 (0.8)	2 (1.3)	0 (0)	0.308
•EMH	1 (0.4)	1 (0.6)	0 (0)	0.469
•Leg ulcer	1 (0.4)	1 (0.6)	0 (0)	0.469
Rate of complications	191 (78.9)	122 (76.7)	69 (83.1)	0.25

Notes: Osteoporosis and hypothyroidism were evaluated in patients ≥10 years old and/or symptomatic, and growth retardation (height >2 SD below 3rd percentile for the mean age and gender) was evaluated in patients ≤18 years.

Table 2 Impact of Hydroxyurea Therapy on Disease Characteristics

Hydroxyurea Therapy (N, %)	Transfusion/Year (Mean ± SD)	s.Ferritin (Mean ± SD)	Chelation Therapy (N, %)
Yes 84 (34.7) No 158 (65.3)	3 ± 5.3 10.4 ± 6.5 P-value <0.001	695.2 ± 1039.5 1553.4 ± 1584.5 P-value <0.001	26 (18.3) 116 (81.7) P-value <0.001

Figure 1 Frequency of disease-related morbidities among 242 β-thalassemia patients at different age intervals.

Figure 2 Clinical management of 242 β-thalassemia patients.

Figure 3 Distribution of disease-related morbidities among 242 β-thalassemia patients.

Table 3 Univariate Analysis of Disease-Related Morbidities Among 242 β-Thalassemia Patients

	Hypothyroidism (n= 33)	Osteoporosis (n= 37)	Biliary Disease (n= 48)	Abnormal LFT (n= 31)	Pulmonary Hypertension (n= 24)
Age (Years)
<18 years (n= 144)	19	16	8	19	11
18–34 years (n = 82)	13	18	32	11	11
≥ 35 years (n= 16)	1	3	8	1	2
P value	0.618	0.662	<0.001	0.778	0.316
Gender
Male (n= 129)	16	23	20	19	12
Female (n= 113)	17	14	28	12	12
P value	0.589	0.113	0.082	0.311	0.762
Ferritin level (μg/l)
< 1000 (n =148)	24	23	34	2	12
1000–2500 (n = 61)  >2500 (n= 33)	5 4	8 6	11 3	10 19	8 4
P value	0.113	0.833	0.122	< 0.001	0.422
Hemoglobin level (g/dL)
<9 (n = 142)	20	27	16	21	15
≥ 9 (n = 97)	13	10	32	10	9
P value	0.755	0.055	<0.001	0.326	0.737
Splenectomy
Yes (n = 83)	6	23	38	14	13
No (n = 159)	27	14	10	17	11
P value	0.031	0.781	<0.001	0.212	0.033
Transfusion/Year
None (n = 33)	2	2	4	0	1
Occasional (n = 77)	16	9	29	4	9
Regular (n = 132)	15	26	15	27	14
P value	0.074	0.401	<0.001	<0.001	0.319
Iron chelation (n=242)
No chelation (n = 100)	15	6	18	1	4
Yes chelation (n = 142)	18	31	30	30	20
P value	0.168	0.577	0.588	<0.001	0.014
Hydroxyurea
Yes (n = 84)	15	6	22	5	10
No (n= 158)	18	31	26	26	14
P value	0.191	0.011	0.061	0.011	0.488

Table 4 Laboratory Investigations of 242 β-Thalassemia Patients in the Current Study

Laboratory Tests	Mean ± SD, Range			P-value
	Total	NTDT	TDT
Hb (g/dl)	8.8 ± 1.3 (4.1–13.8)	8.9 ± 1.4 (4.1-13.9)	8.5 ± 0.9 (5.5–9.1)	0.013
MCV (fl)	74.4 ± 9.7 (48.7–116.2)	72.0 ± 10.6 (48.7–116.2)	78.9 ± 5.1 (62.5–96.4)	<0.001
MCH (pg)	25.6 ± 3.8 (15.8–38.2)	24.4 ± 4 (15.8–38.2)	27.8 ± 1.9 (21.4–33.6)	<0.001
Hb F (%)	65.7 ± 34.8 (4.6–99.8)	65.7 ± 34.8 (4.6–99.5)	64.4 ± 35.8 (6.2–99.8)	0.917
Hb A2 (%)	3.2 ± 2.3 (0.2–11.2)	3.2 ± 2.2 (0.4–8.4)	3.2 ± 3.2 (0.2–11.2)	0.933
Serum Ferritin (μg/l)	1250.5 ± 1472.5 (27–9882)	853.3 ± 1192.7 (27–9882)	2006.2 ± 1665.3 (300–9391)	<0.001
TSH (μIU/mL)	3.5 ± 2 (0.4–15.3)	3.5 ± 1.9 (0.4–14.8)	3.5 ± 2.1 (1.2–15.3)	0.969
ALT (IU/L)	28.3 ± 28.4 (6.0–198)	23.4 ± 23.8 (6.0–158)	37.5 ± 33.8 (9.0–198)	<0.001
Free T4 (Pmol/L)	18.3 ± 2.9 (14–27.6)	18.6 ± 2.8 (14.7–27.6)	17.0 ± 3.1 (14–22.9)	0.173
HCV infection (n, %)	40 (16.5)	18 (11.3)	22 (26.8)	0.039

Table 5 The Reported β-Globin Gene Mutations in the Current Study

β-Thalassemia Mutations	HGVS Nomenclature	Total	NTDT	TDT
β^+
1. IVS I.6 (T> C)	HBB:c.92+6T> C	87 (18.0)	74 (23.3)	13 (7.8)
2. IVS I.110 (G> A)	HBB:c.93–21G> A	29 (6.0)	16 (5)	13 (7.8)
3. IVS I.5 (G> C)	HBB: c.92+5G>C	27 (5.6)	1 (0.3)	26 (15.7)
4. IVS I.128 (T> G)	HBB: c.93–3T>G	10 (2.0)	8 (2.5)	2 (1.2)
5. IVS II.745 (C> G)	HBB:c.316–106C> G	8 (1.7)	2 (0.6)	6 (3.6)
6. CAP +1 (A-> C)	HBB:c.-50A> C	2 (0.4)	2 (0.6)	0 (0)
7. −101 (C>T)	HBB:c.-151C> T	1 (0.2)	1 (0.3)	0 (0)
8. −30 (T>A)	HBB:c.-80T> A	1 (0.2)	0 (0)	1 (0.6)
9. +20 (C>T)	HBB:c.-31C> T	1 (0.2)	1 (0.3)	0 (0)
Total		166 (34.3)	105 (33)	61 (36.7)
β^0
1. IVS II.1 (G> A)	HBB: c.315+1G>A	173 (35.7)	150 (47.2)	23 (13.9)
2. Cod 8/9 (+G)	HBB:c.27–28insG	41 (8.5)	11 (3.5)	30 (18.1)
3. IVS I.1 (G> A)	HBB:c.92+1G> A	27 (5.6)	15 (4.7)	12 (7.2)
4. Codon 8 (-AA)	HBB:c.25–26del/AA	26 (5.4)	15 (4.7)	11 (6.6)
5. Cod 5 (-CT)	HBB:c.17–18del/CT	19 (3.9)	6 (1.9)	13 (7.8)
6. Cod 39 (C> T)	HBB: c.118C>T	8 (1.7)	3 (0.9)	5 (3.0)
7. Cod 44 (-C)	HBB: c.135delC	8 (1.7)	2 (0.6)	6 (3.6)
8. Cod 36/37 (-T)	HBB: c.112delT	4 (0.8)	2 (0.6)	2 (1.2)
9. Cod 25/26 (+T)	HBB:c.78–79insT	2 (0.4)	0(0)	2 (1.2)
10. Cod 82–83 (-G)	HBB: c.251delG	2 (0.4)	1 (0.3)	1 (0.6)
11. Cod 15 (TGG>TGA)	HBB:c.48G> A	1 (0.2)	1 (0.3)	0 (0)
12. IVS II.850 (G> T)	HBB:c.316–1G> T	1 (0.2)	1 (0.3)	0 (0)
Total		312 (64.5)	207 (65.1)	105 (63.3)
Wild		5 (1.0)	5 (1.6)	0 (0)
Dominant like β-thalassemia Cod 127 (A> G)	HBB:c.383A> G	1 (0.2)	1 (0.3)	0 (0)
Total 22 alleles		484	318	166

Table 6 Genotypes of 242 Thalassemia Patients in the Current Study

Genotypes	Total	NTDT	TDT
β^0/β^0
1. IVS II.1 (G> A)/IVS II.1 (G> A)	60 (24.8)	57 (35.9)	3 (3.6)
2. IVS II.1 (G> A)/Cod 8/9 (+G)	10 (4.1)	5 (3.2)	5 (6.0)
3. Cod 8/9 (+G)/Cod 8/9 (+G)	6 (2.5)	0 (0)	6 (7.2)
4. IVS I.1 (G> A)/IVS I.1 (G> A)	6 (2.5)	2 (1.3)	4 (4.8)
5. Cod 5 (-CT)/Cod 5 (-CT)	5 (2.1)	1 (0.6)	4 (4.8)
6. Cod 8 (-AA)/Cod 8 (-AA)	5 (2.1)	2 (1.3)	3 (3.6)
7. Cod 8 (-AA)/IVS II.1 (G> A)	5 (2.1)	4 (2.5)	1 (1.2)
8. IVS I.1 (G> A)/IVS II.1 (G> A)	5 (2.1)	4 (2.5)	1 (1.2)
9. Cod 44 (-C)/Cod 44 (-C)	4 (1.7)	1 (0.6)	3 (3.6)
10. Cod 8 (-AA)/Cod 8/9 (+G)	4 (1.7)	3 (1.9)	1 (1.2)
11. IVSI.1 (G>A)/Cod 8/9 (+G)	3 (1.2)	0 (0)	3 (3.6)
12. Cod 39 (C> T)/Cod 39 (C> T)	2 (0.8)	0 (0)	2 (2.4)
13. Cod 39 (C> T)/IVS II.1 (G> A)	2 (0.8)	2 (1.3)	0 (0)
14. Cod 5 (-CT)/IVS II.1 (G> A)	2 (0.8)	2 (1.3)	0 (0)
15. Cod 8 (-AA)/IVS I.1 (G> A)	2 (0.8)	2 (1.3)	0 (0)
16. IVS II.1 (G> A)/Cod 36/37 (-T)	2 (0.8)	2 (1.3)	0 (0)
17. Cod 36/37 (-T)/Cod 36/37 (-T)	1 (0.4)	0 (0)	1 (1.2)
18. Cod 5 (-CT)/Cod 82–82 (-G)	1 (0.4)	0 (0)	1 (1.2)
19. Cod 5 (-CT)/Cod 8/9 (+G)	1 (0.4)	0 (0)	1 (1.2)
20. IVS II.1 (G> A)/Cod 15 (G> A)	1 (0.4)	1 (0.6)	0 (0)
21. IVS II.1 (G> A)/Cod 82–83 (-G)	1 (0.4)	1 (0.6)	0 (0)
Total	128 (52.9)	89 (56)	39 (47)
β^+/β^+
IVS I.6 (T> C)/IVS I.6 (T> C) IVS I.110 (G> A)/IVS I.110 (G> A) IVS I.5 (G> C)/IVS I.5 (G> C) IVS I.5 (G> C)/IVS I.6 (T> C) IVS II.745 (C> G)/IVS II.745 (C> G) IVS I.128 (T> G)/IVS I.128 (T> G) IVS I.128 (T> G)/IVS I.110 (G> A) IVS I.6 (T> C)/IVS II.745 (C> G)	32 (13.2) 6 (2.5) 5 (2.1) 4 (1.7) 3 (1.2) 2 (0.8) 2 (0.8) 1 (0.4)	30 (18.9) 4 (2.5) 0 (0) 0 (0) 0 (0) 2 (1.3) 1 (0.6) 1 (0.6)	2 (2.4) 2 (2.4) 5 (6.0) 4 (4.8) 3 (3.6) 0 (0) 1 (1.2) 0 (0)
Total	55 (22.7)	38 (23.9)	17 (20.5)
β^0/β^+
1. IVS II.1 (G> A)/IVS I.6 (T> C)	9 (3.7)	7 (4.4)	2 (2.4)
2. IVS II.1 (G> A)/IVS I.110 (G> A)	7 (2.9)	4 (2.5)	3 (3.6)
3. IVS II.1 (G> A)/IVS I.5 (G> C)	6 (2.5)	1 (0.6)	5 (6.0)
4. Cod 8/9 (+G)/IVS I.110 (G> A)	5 (2.1)	2 (1.3)	3 (3.6)
5. Cod 8/9 (+G)/IVS I.5 (G> C)	3 (1.2)	0 (0)	3 (3.6)
6. Cod 5 (-CT)/IVS I.128 (T> G)	2 (0.8)	2 (1.3)	0 (0)
7. Cod 5 (-CT)/IVS I.6 (T> C)	2 (0.8)	0 (0)	2 (2.4)
8. Cod 8 (-AA)/IVS I.110 (G> A)	2 (0.8)	1 (0.6)	1 (1.2)
9. IVS I.1 (G> A)/CAP +1 (A> C)	2 (0.8)	2 (1.3)	0 (0)
10. IVS I.1 (G> A)/IVS I.6 (T> C)	2 (0.8)	2 (1.3)	0 (0)
11. Cod 39 (C> T)/IVS I.6 (T> C)	2 (0.8)	1 (0.6)	1 (1.2)
12. Cod 8 (-AA)/IVS I.5 (G> C)	2 (0.8)	0 (0)	2 (2.4)
13. Cod 25/26 (+T)/IVS I.5 (G> C)	2 (0.8)	0 (0)	2 (2.4)
14. IVS I.1 (G> A)/-101 (C>T)	1 (0.4)	1 (0.6)	0 (0)
15. Cod 5 (-CT)/IVS I.110 (G> A)	1 (0.4)	0 (0)	1 (1.2)
16. Cod 8 (-AA)/IVS I.6 (T> C)	1 (0.4)	1 (0.6)	0 (0)
17. Cod 8/9 (+G)/- 30 (T>A)	1 (0.4)	0 (0)	1 (1.2)
18. Cod 8/9 (+G)/IVS I.128 (T> G)	1 (0.4)	0 (0)	1 (1.2)
19. Cod 8/9 (+G)/IVS I.6 (T> C)	1 (0.4)	1 (0.6)	0 (0)
20. IVS II.850 (G> T)/IVS I.6 (T> C)	1 (0.4)	1 (0.6)	0 (0)
21. IVS II.1 (G> A)/IVS I.128 (T> G)	1 (0.4)	1 (0.6)	0 (0)
Total	54 (22.3)	27 (17)	27 (32.5)
β+/wt +20 (C> T), IVS II.745 (C> G)/wt	2 (0.8)	2 (1.25)	0 (0)
β^0/wt IVS II.1 (G> A)/wt (ααα ^anti3,7)	2(0.8)	2 (1.25)	0(0)
Dominant like β-thalassemia Cod 127 (A> G)/wt	1(0.4)	1 (0.6)	0(0)
Total 53 genotypes	242	159	83

Table 7 Genotype Correlation with Different Parameters

	β^+/β^+ (n=55)	β^+/β ^0 (n=54)	β ^0/β ^0 (n=128)	P value
Age at diagnosis (Years)
Mean ± SD	6.14 ± 8.9	3.02 ± 2.9	4.88 ± 5.4	0.028
Age at first transfusion
Mean ± SD	4.94 ± 9.2	2.61 ± 2.8	3.67 ± 4.2	0.124
Transfusion/year
No	16.4	9.3	14.8	0.510
Yes	83.6	90.7	85.2
Iron chelation
Yes	49.1	64.8	60.2	0.219
No	50.9	35.2	39.8
HbA2%
<3.5	10%	72.7	93.4	<0.001
≥ 3.5	90%	27.3	6.6
Hb F %
<50.0	90%	13.6	9.5	<0.001
≥ 50	10%	86.4	90.5
Splenectomy
Yes	35.2	33.3	33.9	0.977
No	64.8	66.7	66.1
PHT
Yes	3.7	14.8	10.4	0.152
No	96.3	85.2	89.6
Hepatomegaly
Yes	31.5	31.5	43.2	0.181
No	68.5	68.5	56.8
Hepatitis C infection
Yes	16.7	11.1	19.2	0.410
No	83.3	88.9	80.8
Osteoporosis
Yes	38.1	32	39.2	0.816
No	61.9	68	60.8
Biliary complications
Yes	24.1	9.3	20	0.113
No	75.9	90.7	80
Hypothyroidism
No	84.9	83.0	86.1	0.870
Yes	15.1	17.0	13.9
Growth retardation
Yes	45.2	22.5	43.9	0.052
No	54.8	77.5	56.1
ALT
ALT ≥50	7.5	11.1	16.9	0.211
ALT <50	92.5	88.9	83.1
Ferritin level
< 1000	68.5	57.4	60.8	0.561
1000–2500 >2500	24.1 7.4	27.8 14.8	23.2 16.0
Facial deformity
Yes	54.5	64.8	73.4	0.041
No	45.5	35.2	26.6

Notes: Osteoporosis and hypothyroidism were evaluated in patients ≥10 years old and/or symptomatic, and growth retardation (height>2 SD below 3rd percentile for the mean age and gender) was evaluated in patients ≤18 years.