Hypercoagulability and Thrombosis

Hypercoagulability and Thrombosis

Maria Domenica CappelliniErika Poggiali

University of Milan and Policlinico Foundation IRCCS

Milan, Italy

Clinical challenges in NTDT

• Iron overload

• Hypercoagulability

• Iron overload

• Hypercoagulability

Hypercoagulability

Why are we so concerned?

Epidemiology of thromboembolism in thalassaemia patients

Taher A, et al. Blood Rev 2008;22:283-92.

ReferenceTI

n (%)TM

n (%)

Type of thrombosis

VT PE AT Stroke

Zurlo et al., 1989 – 4/159 (2.5) N/A

Michaeli et al., 1992 – 4/100 (4) * * * *

Aessopos J et al., 1997 3/5 (60) 3/5 (60) *

Moratelli et al., 1998 12/74 (16.2) 14/421 (3.3) N/A

Borgna Pignatti et al., 1998 5/52 (9.6) 27/683 (4.0) * * * *

Cappellini et al., 2000 24/83 (29) – * * *

Borgna Pignatti et al., 2004 – 8/720 (1.1) N/A

TI = β-thalassaemia intermedia; TM = β-thalassaemia major;VT = venous thrombosis; PE = pulmonary embolism; AT = arterial thrombosis; N/A = not available.

Taher A, et al. Thromb Haemost. 2006;96:488-91.

12

8

19

12

39

9

66

30

0

11

8

23

28

48

0 20 40 60 80

Others

STP

PVT

PE

DVT

Stroke

Venous

TM (n = 61)

TI (n = 85)

• Patients (N = 8,860)– 6,670 with TM– 2,190 with TI

• 146 (1.65%) thrombotic events – 61 (0.9%) with TM – 85 (3.9%) with TI

Taher A, et al. Thromb Haemost. 2006;96:488-91.DVT = deep vein thrombosis; PE = pulmonary embolism;PVT = portal vein thrombosis; STP = superficial thrombophlebitis.

Thromboembolic events (%)

Type

of e

vent

Risk factors for developing thrombosis in TI were

age (> 20 years)previous thrombotic eventfamily history splenectomy

Thromboembolic events

Can we redefine thalassaemia as

a hypercoagulable state?

Pathophysiology of Thalassaemia

Extravascular haemolysis + Ineffective erythropoiesis

Release into the peripheral circulation of damaged RBCs and erythroid precursors

Pulmonary hypertension (PHT) and thromboembolic events (TEE)

Pathophysiology of thrombosis in NTDT

• Cellular factors– Platelet activation– Pathology and alteration in red blood cells– Endothelial cells and peripheral blood activation

(microparticles)• Nitric oxide• Splenectomy• Inherited and acquired coagulation defects• Other factors

Cappellini MD, et al. Ann N Y Acad Sci 2010;1202:231-6.

Platelets• Increased platelet aggregation• Increased expression of

activation markers • Presence of platelet

morphologic abnormalities

Nitric oxide• Hallmark of haemolysis• ↓ Levels leading to vasoconstriction

Thrombophilia• No role for prothrombotic

mutations• Decreased levels of antithrombin III,

protein C, and protein S • Anti-phospholipid antibodies

Splenectomy• High platelet counts and

hyperactivity• High levels of negatively charged

RBCs

RBCs• Formation of reactive

oxygen species • Expression of negatively

charged phospholipids• Enhanced cohesiveness

and aggregability

Hypercoagulability

Other factors• Cardiac dysfunction• Hepatic dysfunction• Endocrine dysfunction

Peripheral blood elements• Expression of endothelial

adhesion molecules and tissue factor on endothelial cells (ELAM-1, ICAM-1, vWF, VCAM-1)

• Formation of microparticles


Hypercoagulability

Platelet activation

Winichagoon P, et al. Asian J Trop Med Public Health 1981;12:556–60.Del Principe D, et al. Br J Hematol 1993;84:111–7.

Ruf A, et al. Br J Hematol 1997;98:51–6.Eldor A, et al. Am J Hematol 1989;32:94–9.

Pathology and alteration in red blood cells

Endothelial cells and peripheral blood activation

Haemolysis

Butthep P, et al. Thromb Hemost 1995;74:1045–9.Butthep P, et al. Southeast Asian J Trop Med Public Health

1997;28(Suppl. 3):141A–8A.Hovav T, et al. Br J Hematol 1999;106:178–81.

Platelets• Increased platelet aggregation• Increased expression of

activation markers • Presence of platelet

morphologic abnormalities

Nitric oxide• Hallmark of haemolysis• ↓ levels leading to vasoconstriction

Thrombophilia• No role for prothrombotic

mutations• Decreased levels of antithrombin III,

protein C, and protein S • Anti-phospholipid antibodies

Splenectomy• High platelet counts and

hyperactivity• High levels of negatively charged

RBCs

RBCs• Formation of reactive

oxygen species • Expression of negatively

charged phospholipids• Enhanced cohesiveness

and aggregability

Hypercoagulability

Other factors• Cardiac dysfunction• Hepatic dysfunction• Endocrine dysfunction

Peripheral blood elements• Expression of endothelial

adhesion molecules and tissue factor on endothelial cells (ELAM-1, ICAM-1, vWF, VCAM-1)

• Formation of microparticles


Hypercoagulability

The epidemiological data

and the clinical experience

Complication Parameter RR* 95% CI p value

Thrombosis Age > 35 years 2.60 1.39-4.87 0.003

Hb ≥ 9 g/dL 0.41 0.23-0.71 0.001

Serum ferritin ≥ 1,000 µg/L

1.86 1.09-3.16 0.023

Splenectomy 6.59 3.09-14.05 < 0.001

Transfusion 0.28 0.16-0.48 < 0.001

OPTIMAL CARE study: incidence and risk factors for thrombosis

Parameter Frequencyn (%)

Age (years)< 18 172 (29.5 )18–35 288 (49.3) > 35 124 (21.2)

Male : Female 291 (49.8) : 293 (50.2)Splenectomized 325 (55.7)Serum ferritin (µg/L)

< 1,000 376 (64.4)1,000–2,500 179 (30.6)> 2,500 29 (5)

ComplicationsOsteoporosisEMHHypogonadism CholelithiasisThrombosis Pulmonary hypertensionAbnormal liver functionLeg ulcers HypothyroidisimHeart failureDiabetes mellitus

134 (22.9)124 (21.2)101 (17.3)100 (17.1)

82 (14)64 (11)57 (9.8)46 (7.9)33 (5.7)25 (4.3)10 (1.7)

Taher AT, et al. Blood. 2010;115:1886-92.EMH = extramedullary haematopoiesis; CI = confidence interval.

*RR indicates adjusted relative risk.

Frequency of thrombosis increases with age in NTDT patients

EMH

Leg ulce

rs

Thrombosis PHT HF

Cholelith

iasis

ALF DM

Hypothyro

idism

Osteoporo

sis

Hypogonad

ism0

5

10

15

20

25

30

35

40

45

6.73.3 3.3 3.3

0

13.3

3.30 0

6.7

0

13.3

6.7

13.3

6.73.3

10.06.7

03.3

16.7 16.716.7 16.720.0

10.06.7

20.0

10.0

3.3

10.0

23.3 23.3

40.0

26.7 26.7

20.0

10.0

33.3

13.3 13.316.7

30.0

20.0

< 10 years 11-20 years 21-32 years >32 years

Freq

uenc

y (%

)

*

* ** *

** = statistically significant trend

Taher A, et al. Br J Haematol 2010;150:486-9.

< 10 years 11–20 years 21–32 years > 32 years

● N = 120 treatment-naive* TI patients

*never received any treatment intervention (splenectomy, transfusion,iron chelation therapy, or HbF-inducing agents).

PHT = pulmonary hypertension; HF = heart failure; ALF = abnormal liver function; DM = diabetes mellitus.

High prevalence of thromboembolic events, particularly in splenectomized patients

Thromboembolic events occurred in 24/83 (29%) transfusion-independent patients with TI who had undergone splenectomy

Cappellini MD, et al. Br J Haematol. 2000;111:467-73.

Thro

mbi

n-ge

nera

ted

(nM

)

0 10 30 60 90 120 1500

30

60

90

120

150Splenectomized patient with TINon-splenectomized patient with TINon-splenectomized controlsSplenectomized controls

Time (s)

OPTIMAL CARE study: patient stratification according to splenectomy and TEE status

• Three groups of patients identified – Group I, splenectomized patients with a documented TEE (n = 73)– Group II, age- and sex-matched splenectomized patients without TEE (n = 73)– Group III, age- and sex-matched non-splenectomized patients without TEE (n = 73)

Taher A, et al. J Thromb Haemost. 2010;8:2152-8.

Type of thromboembolic event in splenectomized TI patients (Group I) n (%)

DVT 46 (63.0)

PE* 13 (17.8)

STP 12 (16.4)

PVT 11 (15.1)

Stroke 4 (5.5)

TEE = thromboembolic events; DVT = deep vein thrombosis; PE = pulmonary embolism; STP = superficial thrombophlebitis; PVT = portal vein thrombosis.

*All patients who had PE had confirmed DVT.

Parameter

Group ISplenectomized with TEE(n = 73)

Group IISplenectomized without TEE(n = 73)

Group IIINon-splenectomized(n = 73)

p value

Mean age ± SD, years 33.1 ± 11.7 33.3 ± 11.9 33.4 ± 13.1 0.991Male : female 33 : 40 35 : 38 34 : 39 0.946Mean Hb ± SD, g/dL 9.0 ± 1.3 8.8 ± 1.2 8.7 ± 1.3 0.174Mean HbF ± SD, % 45.9 ± 28.0 54.4 ± 32.8 44.2 ± 27.2 0.429Mean NRBC count ± SD, x106/L 436.5 ± 205.5 279.0 ± 105.2 239.5 ± 128.7 < 0.001Mean platelet count ± SD, x109/L 712.6 ± 192.5 506.3 ± 142.1 319.2 ± 122.0 < 0.001PHT, n (%) 25 (34.2) 17 (23.3) 3 (4.1) < 0.001HF, n (%) 7 (9.6) 5 (6.8) 1 (1.4) 0.101DM, n (%) 4 (5.5) 5 (6.8) 1 (1.4) 0.256Abnormal liver function, n (%) 2 (2.7) 2 (2.7) 3 (4.1) 0.863Family history of TEE 3 (4.7) 1 (1.4) 3 (4.7) 0.554Thrombophilia, n (%) 3 (4.7) 2 (2.7) 2 (2.7) 0.863Malignancy, n (%) 1 (1.4) 2 (2.7) 0 (0) 0.363Transfused, n (%) 32 (43.8) 48 (65.8) 54 (74.0) 0.001Anti-platelet or anticoagulant use, n (%) 1 (1.4) 3 (4.1) 2 (2.7) 0.598Hydroxyurea use, n (%) 13 (17.8) 17 (23.3) 29 (27.4) 0.383

OPTIMAL CARE study: patient stratification according to splenectomy and TEE status (cont.)

Hb = total haemoglobin; HbF = fetal haemoglobin; NRBC = nucleated red blood cell; PHT = pulmonary hypertension; HF = heart failure; DM = diabetes mellitus. Taher A, et al. J Thromb Haemost. 2010;8:2152-8.

Time-to-thrombosis (TTT) since splenectomy

• The median TTT following splenectomy was 8 years (range 1–33 years)


Time-to-thrombosis (TTT) since splenectomy (cont.)


Time to thrombosis

Duration since splenectomy (years)

Cum

ulat

ive

thro

mbo

sis-

free

sur

viva

l

0

0.2

0.4

0.6

0.8

1

0 5 10 15 20 25 30 35 40

NRBC count< 300 x 106/L≥ 300 x 106/L


Cum

ulat

ive

thro

mbo

sis-

free

sur

viva

l

0

0.2

0.4

0.6

0.8

1

0 5 10 15 20 25 30 35 40

TransfusedYesNo


Cum

ulat

ive

thro

mbo

sis-

free

sur

viva

l

0

0.2

0.4

0.6

0.8

1

0 5 10 15 20 25 30 35 40

Platelet count< 500 x 109/L≥ 500 x 109/L


Cum

ulat

ive

thro

mbo

sis-

free

sur

viva

l

0

0.2

0.4

0.6

0.8

1

0 5 10 15 20 25 30 35 40

Pulmonary hypertensionYesNo

Clinical recommendations for the prevention of thromboembolic events

• A guarded approach to splenectomy in β-thalassaemia patients is recommended unless strongly indicated

• In already-splenectomized NTDT patients, those at high risk of thrombosis may be identified early by their high NRBC and platelet counts, evidence of PHT, and transfusion naivety– attention should also be paid to the aging NTDT patients

• Prospective clinical trials that evaluate the efficacy, safety, and cost-effectiveness of transfusions and anti-platelet and anticoagulant therapy in preventing thromboembolism are necessary– aspirin for stroke prevention and lifelong anticoagulation treatment in patients

with a history of thrombotic events

Taher AT, et al. Br J Haematol. 2011;152:512-23.

Complication Parameter RR 95% CI p value

EMH Splenectomy 0.44 0.26–0.73 0.001Transfusion 0.06 0.03–0.09 < 0.001Hydroxyurea 0.52 0.30–0.91 0.022

Pulmonary hypertension Age > 35 years 2.59 1.08–6.19 0.032Splenectomy 4.11 1.99–8.47 < 0.001Transfusion 0.33 0.18–0.58 < 0.001Hydroxyurea 0.42 0.20–0.90 0.025Iron chelation 0.53 0.29–0.95 0.032

Heart failure Transfusion 0.06 0.02–0.17 < 0.001Thrombosis Age > 35 years 2.60 1.39–4.87 0.003

Hb ≥ 9 g/dL 0.41 0.23–0.71 0.001Serum ferritin ≥ 1,000 µg/L 1.86 1.09–3.16 0.023Splenectomy 6.59 3.09–14.05 < 0.001Transfusion 0.28 0.16–0.48 < 0.001

Cholelithiasis Age > 35 years 2.76 1.56–4.87 < 0.001Female 1.96 1.18–3.25 0.010Splenectomy 5.19 2.72–9.90 < 0.001Transfusion 0.36 0.21–0.62 < 0.001Iron chelation 0.30 0.18–0.51 < 0.001

Abnormal liver function Serum ferritin ≥ 1,000 µg/L 1.74 1.00–3.02 0.049

OPTIMAL CARE study: transfusion therapy reduces the risk of complications

Taher AT, et al. Blood. 2010;115:1886-92.

n = 445 occasionally/regularly transfused patients (N = 584)

Complication Parameter RR 95% CI p value

Leg ulcers Age > 35 years 2.09 1.05–4.16 0.036Splenectomy 3.98 1.68–9.39 0.002Transfusion 0.39 0.20–0.76 0.006Hydroxyurea 0.10 0.02–0.43 0.002

Hypothyroidism Splenectomy 6.04 2.03–17.92 0.001Hydroxyurea 0.05 0.01–0.45 0.003

Osteoporosis Age > 35 years 3.51 2.06–5.99 < 0.001Female 1.97 1.19–3.27 0.009Splenectomy 4.73 2.72–8.24 < 0.001Transfusion 3.10 1.64–5.85 < 0.001Hydroxyurea 0.02 0.01–0.09 < 0.001Iron chelation 0.40 0.24–0.68 0.001

Hypogonadism Female 2.98 1.79–4.96 < 0.001Serum ferritin ≥ 1,000 µg/L 2.63 1.59–4.36 < 0.001Transfusion 16.13 4.85–52.63 < 0.001Hydroxyurea 4.32 2.49–7.49 < 0.001Iron chelation 2.51 1.48–4.26 0.001

Only significant associations presented.

OPTIMAL CARE study: transfusion therapy reduces the risk of complications (cont.)

Taher AT, et al. Blood. 2010;115:1886-92.

n = 445 occasionally/regularly transfused patients (N = 584).

• Transfusion therapy was protective for thrombosis, EMH, PHT, HF, cholelithiasis and leg ulcers

• Transfusion therapy was associated with an increased risk of iron overload-related endocrinopathy

Indications of RBC transfusion in TI• Hb < 5 g/dL• Declining Hb level with progressive splenic enlargement (> 3 cm/year)*

• Poor growth and/or development• Evidence of

– bone deformities– clinically relevant tendency to thrombosis– leg ulcers– EMH– PHT– infections

• Prior to surgical procedures• Pregnancy

Taher A. et al. Blood Reviews 26S (2012); S24-S27.Hb = heamoglobin.* At least in periods of maximal growth and development.

Initiating transfusion therapy in NTDT patients

• The decision to initiate transfusion in these patients is very difficult due to the heterogeneity of the disease

• There is no benefit in limiting the quantity or frequency of transfusions once they have begun

• Starting transfusions after the third year of life has been shown to increase the risk of alloimmunization

• Transfused patients with TI experience fewer TEE, PHT and silent brain infarcts compared to transfusion-naïve patients, due to the correction of the ineffective erythropoiesis and resulting damaged RBCs with thrombogenic potential

Taher A. et al. Blood Reviews 26S (2012); S24-S27.

Summary

• Thromboembolic events are frequent in β-thalassaemia patients– oxidative damage to RBCs, impacting their membrane properties, resulting in increased

aggregation and risk of thromboembolism– risk of thromboembolism increases with age, and is influenced strongly by splenectomy and

transfusion navïety

• Splenectomy is associated with a high risk of thrombosis, particularly in patients with high NRBC or platelet counts, who are transfusion-naïve

• Transfusion therapy reduces the risk of thrombosis in NTDT patients– transfusion iron intake inevitably increases the risk of iron overload, but the

benefit of transfusion therapy may greatly outweigh the cost and inconvenience of iron chelation therapy

Summary• Despite various treatment options are available, no clear

guidelines exist: each patient must be assessed individually and assigned a personalized thrombotic risk based on intrinsic and extrinsic factors

• Several studies are highlighting the roles of transfusion, iron chelation therapy, and fetal haemoglobin induction (hydroxycarbamide, HU) in the management of NTDT; thus these approaches merit large prospective evaluation

• Another approach would be to correct the reactive oxygen species-induced RBC membrane damage using antioxidants, although this approach has not yet been verified in clinical trials

Acknowledgments I would like to thank you for the attention,

and all the Congenital Anemia Center Staff for their support

Prof. M.D. CappelliniGiovanna Graziadei Irene MottaAlessia Marcon Ilaria GandolfiLaura Zanaboni Marianna GiudittaElena Cassinerio Marta MazzoleniClaudia Cesaretti Silvio De Fazio

All the nursing staff

Alteration of the phospholipid “Flip-Flop” mechanism:

RBCs with negatively charged phospholipids

Adherence of RBCs to endothelial cells is increased

Courtesy of Dr A. Taher.

Thrombus formation

Fibrin/ platelets

Thrombin generationPhosphatidylserine on damaged or senescent RBCs leads to

● Recognition by phagocytes

● Removal from circulation

● Apoptosis

Splenectomy favours persistence of these RBCs in the circulation

Documents

Hypercoagulability and Thrombosis