2 G-EXJ-1030713 May 2012 TABLE OF CONTENTS Use of MRI in
evaluating liver iron loading (and monitoring therapy) Cardiac MRI
Cytogenetic Assessment in MDS Transcranial Doppler Ultrasonography
(TCD) for assessment of stroke risk in Sickle Cell Disease Glossary
of terms
Slide 3
G-EXJ-1030713 May 2012 USE OF MRI IN EVALUATING LIVER IRON
LOADING (AND MONITORING THERAPY) NOTE: These slides are for use in
educational oral presentations only. If any published
figures/tables from these slides are to be used for another purpose
(e.g. in printed materials), it is the individuals responsibility
to apply for the relevant permission. Specific local use requires
local approval
Slide 4
4 G-EXJ-1030713 May 2012 Outline Introduction to iron and liver
iron overload Key methods for assessing liver iron liver biopsy SF
SQUID liver MRI SIR method relaxometry methods (R2 and R2*)
Clinical recommendations for measuring LIC Summary LIC = liver iron
concentration; MRI = magnetic resonance imaging; SF = serum
ferritin; SIR = signal intensity ratio; SQUID = superconducting
quantum interface device.
Slide 5
G-EXJ-1030713 May 2012 Introduction to iron and iron
overload
Slide 6
6 G-EXJ-1030713 May 2012 Iron overload Iron overload is common
in patients who require intermittent or regular blood transfusions
to treat anaemia and associated conditions it may be exacerbated in
some conditions by excess gastrointestinal absorption of iron Iron
overload can lead to considerable morbidity and mortality 1 Excess
iron is deposited in major organs, resulting in organ damage the
organs that are at risk of damage due to iron overload include the
liver, heart, pancreas, thyroid, pituitary gland, and other
endocrine organs 2,3 1 Ladis V, et al. Ann NY Acad Sci.
2005;1054:445-50. 2 Gabutti V, Piga A. Acta Haematol.
1996;95:26-36. 3 Olivieri NF. N Engl J Med. 1999;341:99-100.
Slide 7
7 G-EXJ-1030713 May 2012 Importance of analysing liver iron A
patients LIC is the best measure of total body iron stores Knowing
the liver iron concentration helps to predict the risk of hepatic
and extra-hepatic complications 14 1 Batts KP. Mod Pathol.
2007;20:S31-9. 2 Jensen PD, et al. Blood. 2003;101:91-6. 3
Angelucci E, et al. Blood. 2002;100:17-21. 4 Telfer PT, et al. Br J
Haematol. 2000;110:971-7.
Slide 8
8 G-EXJ-1030713 May 2012 LIC threshold of 7 mg Fe/g dry wt 0 5
10 15 20 25 All (n = 1,744) TM (n = 937) TI (n = 84) SCD (n = 80)
Mean LIC + SD over previous year prior to enrolment in EPIC trial
(mg Fe/g dry wt) Cappellini MD, et al. Blood. 2008;112:[abstract
3880]. Importance of analysing liver iron (cont.) All
transfusion-dependent patients prior to study enrolment had
moderate-to-severe hepatic iron loading
Slide 9
9 G-EXJ-1030713 May 2012 Overview of LIC correlations with
other measurements DFS = disease-free survival. 1 Angelucci E, et
al. N Engl J Med. 2000;343:327-31. 2 Jensen PD, et al. Blood.
2003;101:91-6. 3 Angelucci E, et al. Blood. 2002;100:17-21. 4
Telfer PT, et al. Br J Haematol. 2000;110:971-7. 5 Noetzli LJ, et
al. Blood. 2008;112:2973-8. LIC Hepatocellular injury 2 and
fibrosis 3 Body iron stores 1 Cardiac iron 5 Cardiac DFS 4
Slide 10
10 G-EXJ-1030713 May 2012 LIC prediction of total body iron
stores BMT = bone marrow transplantation. 1 Olynyk JK, et al. Am J
Gastroenterol. 1998;93:346-50. 2 Angelucci E, et al. N Engl J Med.
2000;343:327-31. Sample > 1 mg dry wt (n = 25) r = 0.98
0510152025 300 250 200 150 100 50 0 Body iron stores (mg/kg) LIC
(mg Fe/g dry wt) Hereditary haemochromatosis 1 Iron removed (g) LIC
(g/g) 0510152025 50,000 40,000 30,000 20,000 10,000 0 -TM 2 LIC is
a reliable measure of total body iron stores in hereditary
haemochromatosis and -TM
Slide 11
11 G-EXJ-1030713 May 2012 Serum ferritin measurement alone
underestimates the body iron load Origa R, et al. Haematologica.
2007;92:583-8. Taher A, et al. Haematologica. 2008;93:1584-6. -TI
-TM 05101520253035 LIC (mg Fe/g dry wt) SF ( g/L) 2,000 4,000 6,000
8,000 10,000 12,000 14,000 0 SF ( g/L) 05101520253035404550 LIC (mg
Fe/g dry wt) 1,000 2,000 3,000 4,000 5,000 6,000 7,000 8,000 9,000
10,000 0 -TI -TM SF has almost no sensitivity or specificity for
iron stores in thalassaemia intermedia
Slide 12
G-EXJ-1030713 May 2012 Assessing liver iron overload
Slide 13
13 G-EXJ-1030713 May 2012 Key methods for assessing liver iron
Liver biopsy LIC advantages and disadvantages correlation of LIC
with other measurements SF concentration over time advantages and
disadvantages correlation of SF levels with other measurements
SQUID advantages and disadvantages Liver MRI advantages and
disadvantages relaxometry methods (T2 and T2*) SIR method Olivieri
NF, Brittenham GM. Blood. 1997;89:739-61. Direct method Indirect
methods
Slide 14
G-EXJ-1030713 May 2012 Liver biopsy
Slide 15
15 G-EXJ-1030713 May 2012 Technique for taking a percutaneous
liver biopsy Step 1. The patient lies on his back, or his left side
Step 4. The patient must hold breath for 5-10 seconds when the
needle is quickly pushed in and out. As the needle comes out it
brings with it a small sample of liver tissue Patient preparation:
Blood tests are done shortly before the biopsy to check blood
clotting time, to exclude risk of bleeding following the biopsy.
The biopsy is commonly preceded by an ultrasound examination of the
liver to determine the best and safest biopsy site Step 3. A
special hollow needle is inserted into the liver, usually between
the 2 lower ribs on the right hand side Step 2. The place for the
biopsy is cleaned with antiseptic and local anaesthesia is provided
(s.c. on the right hand side) Liver biopsy A tiny incision is made
between the ribs, and a needle is inserted to reach the area of the
liver where a tissue sample is taken. The procedure requires local
anaesthesia Area where a tissue sample is taken from Overall: The
procedure is carried out by a qualified physician or surgeon in an
outpatient care centre or hospital. It is fast (not longer than 5
min) and the patient is discharged shortly after adam.com
Slide 16
16 G-EXJ-1030713 May 2012 Processing the liver biopsy sample
Gross histopathological examination reveals presence of abnormal
cells or liver tissue used to determine presence and degree of
cirrhosis and fibrosis LIC measurement by iron staining by atomic
absorption spectroscopy: the current gold standard! Who does the
test? preparation of the samples might be by a trained technician
the analysis requires a qualified pathologist Angelucci E, et al.
Haematologica. 2008;93:741-52. Image from:
www.pathguy.com/lectures/cirrhosis_trichrome.jpg
Slide 17
17 G-EXJ-1030713 May 2012 Liver biopsy Liver biopsy with iron
measurement by atomic absorption spectroscopy is the gold standard
for measuring LIC 1 LIC threshold (mg Fe/g dry wt) 2 LIC threshold
( mol Fe/g dry wt) Clinical relevance 1.832Upper 95% of normal 15.0
269 Greatly increased risk of cardiac disease and early death 1
Angelucci E, et al. Haematologica. 2008;93:741-52. 2 St Pierre TG,
et al. Blood. 2005;105:855-61.
Slide 18
18 G-EXJ-1030713 May 2012 Liver biopsy: pros and cons Pros 1
Cons Direct measurement of LIC Validated reference standard
Quantitative, specific, and sensitive Allows for measurement of
non-haem storage iron Provides information on liver
histology/pathology Correlates with morbidity and mortality
Invasive and painful procedure with risk of potentially serious
complications 1 May involve sampling errors, especially in patients
with cirrhosis 1 Requires skilled physicians 1 Laboratory
techniques not standardized 1 iron measurement by atomic absorption
spectroscopy 2 or chemical determination 3 wet or dry weight quoted
iron concentration varies throughout the liver, 4 sample size often
insufficient (requires 1 mg dry weight, or > 4 mg wet weight) 1
TIF. Guidelines for the Clinical Management of Thalassemia. 2nd
rev. ed. Cyprus: TIF; 2008. Available from:
www.thalassaemia.org.cy/pdf/Guidelines_2nd_revised_edition_EN.pdf.
Accessed December 2010. 2 Angelucci E, et al. Haematologica.
2008;93:741-52. 3 Wood JC. Blood Rev. 2008;22 Suppl 2:S14-21. 4
Ambu R, et al. J Hepatol. 1995;23:544-9.
Slide 19
19 G-EXJ-1030713 May 2012 Heterogeneity of iron concentration
throughout the liver From autopsy of a patient with
beta-zero-thalassaemia. Ambu R, et al. J Hepatol. 1995;23:544-9.
020% 2040% 4060% 6080% 80100% Iron is unevenly distributed in the
liver; therefore, a small sample may not give an absolutely
representative mean LIC
Slide 20
G-EXJ-1030713 May 2012 SF Concentration
Slide 21
21 G-EXJ-1030713 May 2012 Ferritin and SF Ferritin is primarily
an intracellular protein that stores iron in a form readily
accessible to cells releases iron in a controlled fashion The
molecule is shaped like a hollow sphere and it stores ferric (Fe 3+
) iron in its central cavity the storage capacity of ferritin is
approximately 4,500 Fe 3+ ions per molecule Ferritin is found in
all tissues, though primarily in the liver, spleen, and bone marrow
A small amount is also found in the blood as serum ferritin
Harrison PM, Arosio P. Biochim Biophys Acta. 1996;1275:161-203. SF
> 1,000 g/L is a marker of excess body iron
Slide 22
22 G-EXJ-1030713 May 2012 SF: pros and cons SF levels from a
blood sample are measured ProsCons Easy to assess Inexpensive
Positive correlation with morbidity and mortality Allows
longitudinal follow-up of patients Indirect measurement of iron
burden Fluctuates in response to inflammation, abnormal liver
function, ascorbate deficiencies TIF. Guidelines for the Clinical
Management of Thalassaemia. 2nd rev. ed. Cyprus: TIF; 2008.
Available from:
www.thalassaemia.org.cy/pdf/Guidelines_2nd_revised_edition_EN.pdf.
Accessed December 2010.
Slide 23
G-EXJ-1030713 May 2012 SQUID
Slide 24
24 G-EXJ-1030713 May 2012 SQUID: superconducting quantum
interference device Carneiro AA, et al. Reson Med. 2005;43:122-8.
Magnetizing coil Dewar Liquid helium SQUID Pick to coil Water bag
Patient Mattress Bed Piston H2OH2O Patient preparation: No special
patient preparation is required. Ultrasound is used to evaluate the
depth and size of the liver. The patient lies on their back with
their torso surrounded by a 5-L water bag to minimize contributions
from other tissues Step 2. LIC corresponds to the variation of
magnetization detected and is calculated using custom-made Matlab
6.5 software Principle of the technique: Normal tissue is
diamagnetic and has a magnetic susceptibility similar to that of
water. In the presence of iron, tissue susceptibility is changed
proportional to the amount of iron present. This alteration is
detected, allowing non-invasive measurement of LIC Step 1. The
susceptometer applies a low-power (114 T and 7.7 Hz) homogeneous
magnetizing field in the hepatic region. Sensitive detectors
measure the interference of tissue iron vs the water reference
medium within the field Overall: The procedure is carried out by a
qualified radiologist in a hospital. It is fast (not longer than 5
min) and the patient is discharged immediately after. Processing
could be done on the spot and is faster then LIC histopathological
examination
Slide 25
25 G-EXJ-1030713 May 2012 SQUID: pros and cons ProsCons
Non-invasive 1 Wide linear range 1 Good correlation with LIC by
biopsy 2 Requires expensive, specialized equipment and expertise 1
Not widely available 1 Each machine should be individually
calibrated 1 SQUID can underestimate LIC 3 1 TIF. Guidelines for
the Clinical Management of Thalassaemia. 2nd rev. ed. Cyprus: TIF;
2008. Available from:
www.thalassaemia.org.cy/pdf/Guidelines_2nd_revised_edition_EN.pdf.
Accessed December 2010. 2 Sheth S. Pediatr Radiol. 2003;33:373-7. 3
Piga A, et al. Blood. 2005;106:[abstract 2689]. 250 200 150 100 50
0 0 100150200250 Hepatic iron (magnetic) ( mol Fe/g wet wt) Hepatic
iron (biopsy) ( mol Fe/g wet wt) R = 0.99 p < 0.001 SQUID is a
non-invasive method that has been calibrated, validated, and used
in clinical studies, but the complexity, cost and technical demands
limit its use
Slide 26
G-EXJ-1030713 May 2012 Liver MRI
Slide 27
27 G-EXJ-1030713 May 2012 MRI ROI = region of interest; SI =
signal intensity; TE = echo time. Brittenham GM, Badman DG. Blood.
2003;101:15-9. Ridgway JP. J Cardiovasc Magn Reson. 2010;12:71.
Integral radiofrequency transmitter (body) coil Main magnet coils
x,y,z gradient coils Patient table Patient preparation: All
infusion and medication pumps should be removed. The scan does not
require contrast agent, and so no peripheral vein access is needed
Step 2. Post-processing: As TE increases, the images SI decreases.
The relationship between TE and SI in a selected part of the image
(i.e. ROI) is analysed with specialized software or manually. Data
are reported as relaxation times (T2 or T2*), depending on the
acquisition method Principle of the technique: A strong magnetic
field is used to organize the protons in the tissue in 1 direction.
Then radiofrequency is used to knock them off. The time for them to
re-align with the magnetic field and the energy they release during
the process depend on the interactions of the proton with other
ions, notably iron ions. These events could be measured at various
TEs and then analysed to reveal the iron content in the tissue Step
1. Image acquisition: Images are taken at various TEs Overall: The
procedure is carried out by a qualified radiologist in a hospital.
Acquisition is fast (approx. 5 min), and the patient is discharged
immediately after. Processing may require specialized software and
is done afterwards
Slide 28
28 G-EXJ-1030713 May 2012 MRI is increasingly being used as a
non-invasive method to measure LIC ProsCons Non-invasive 1,2
Assesses iron content throughout the liver 2 Increasingly and
widely available worldwide 2 Pathological status of liver and heart
can be assessed in parallel 2 Validated relationship with biopsy
LIC 3 6 Indirect measurement of LIC 2 Requires MRI with dedicated
imaging method 2 Sensitivity depends on type of scanner, degree of
iron overload, presence of fibrosis, and inflammation 7 1 Chavhan
GB, et al. Radiographics. 2009;29:1433-49. 2 TIF. Guidelines for
the Clinical Management of Thalassaemia. 2nd rev. ed. Cyprus: TIF;
2008. Available from: www.thalassaemia.org.cy/pdf/Guidelines_2nd_
revised_edition_EN.pdf. Accessed December 2010. 3 Christoforidis A,
et al. Eur J Haematol. 2009;82:388-92. 4 St Pierre TG, et al.
Blood. 2005;105:855-61. 5 Wood JC, et al. Blood. 2005;106:1460-5. 6
Hankins JS, et al. Blood. 2009;113:4853-5. 7 Sirlin CB, Reeder SB.
Magn Reson Imaging Clin N Am. 2010;18:359-81.
Slide 29
29 G-EXJ-1030713 May 2012 MRI scanners Manufacturers Siemens
Healthcare (Erlangen, Germany; www.siemensmedical.com) GE
Healthcare (Milwaukee, WI, USA; www.gemedicalsystems.com) Philips
Healthcare (Best, the Netherlands; www.medical.philips.com)
Magnetic field strength most imaging is done on 1.5 T machines 3 T
machines give better signal:noise ratio 1 worse susceptibility
artefacts 1 The upper detection limit is halved, therefore it is
too low for many patients 1 lower T2 and T2* values than 1.5 T
machines 2 Liver package (including standard sequences and analysis
of the data) is included in the software provided together with the
MRI machine specialized LIC analysis software can be bought
separately 1 Wood JC, Ghugre N. Hemoglobin. 2008;32:85-96. 2 Storey
P, et al. J Magn Reson Imaging. 2007;25:540-7.
Slide 30
30 G-EXJ-1030713 May 2012 Overview of MRI techniques used to
measure LIC DATA ACQUISITION DATA ANALYSIS MAJOR PROS AND CONS A
combination of gradient and spin echos Free website + Fast
acquisition Simple data analysis Limited sensitivity
Reproducibility Gradient echo (same technique as cardiac iron
measurement) (1 min) Manually (free xls sheet) or with dedicated
software (e.g CMR tool 3,000 GBP per year) + Fast acquisition
Correlates well with LIC Susceptible to artefacts Training needs
Spin echo (15min) Done centrally by Resonance Health (300 USD per
scan) + Gold Standard Little training need Longer data acquisition
time Cost of analysis Signal Intensity Ratio (SIR) method
(Gandon/Ernst) Relaxometry method R2*(T2*) R2(T2) (Ferriscan )
Liver MRI Technique
Slide 31
31 G-EXJ-1030713 May 2012 MRI measurement of LIC: techniques
There are 2 broad groups of techniques SIR methods (Gandon et al.
methods) relaxometry methods (FerriScan and T2* (R2*) methods)
ProsCons SIR method Fast data acquisition Relatively simple
algorithms and data analysis Can be used in scanners with different
magnetic strengths (0.5, 1.0, 1.5 T) Limited range of sensitivity
(upper limit is 21 mg Fe/g dry wt [380 mol/L]) Assumptions on
reference tissue Not reliable in cirrhosis Smaller reproducibility
Relaxometry method Greater range of sensitivity Does not rely on
reference tissue assumptions T2* (or R2*) is very quick (requires a
single breath-hold) Has only been calibrated at 1.5 T Takes longer
to acquire data, when done as T2 (or R2) Argyropoulou MI, Astrakas
L. Pediatr Radiol. 2007;37:1191-200. Gandon Y, et al. Lancet.
2004;363:357-62. St Pierre TG, et al. Ann N Y Acad Sci.
2005;1054:379-85. Wood JC. Curr Opin Hematol. 2007;14:183-90. Wood
JC, et al. Blood. 2005;106:1460-5.
Slide 32
32 G-EXJ-1030713 May 2012 SIR methods Most common protocol
includes 4-gradient echo sequences with different TEs 1 spin-echo
sequence 0 100 300 400 200 0100200400300 Study group Validation
group Biopsy LIC ( mol Fe/g dry wt) MRI LIC ( mol Fe/g dry wt)
Gandon Y, et al. Lancet. 2004;363:357-62. 1. Patient preparation (5
min) 2. Image acquisition (approx. 5-20 min) 3. Data analysis
(depends on experience)
Slide 33
33 G-EXJ-1030713 May 2012 SIR methods (cont.) The ROI is
selected in the liver and the reference tissue (muscle or fat), in
each image The SI of the liver region is divided by that of the
reference tissue A calculation algorithm to assist has been
developed for 0.5, 1.0, and 1.5 T MRI machines 1 1 Gandon Y.
Available from:
http://www.radio.univ-rennes1.fr/Sources/EN/HemoResult.html.
Accessed December 2010. 1. Patient preparation (5 min) 2. Image
acquisition (approx. 5-20 min) 3. Data analysis (relatively
fast)
Slide 34
34 G-EXJ-1030713 May 2012 Relaxometry methods: T2, T2*, T2, R2,
and R2* If a spin-echo sequence is used, the relaxation time is T2
If a gradient-echo sequence is used, it is T2* These are related by
the equation 1 1/T2* = 1/T2 + 1/T2 T2 is the magnetic field
inhomogeneity of the tissue To attain a positive linear
relationship with HIC T2* can be transformed into reciprocal R2*:
R2* [Hz] = 1,000/T2* [ms] T2 can be transformed into reciprocal R2:
R2 [Hz] = 1,000/T2 [ms] 1 Anderson LJ, et al. Eur Heart J.
2001;22:2171-9. 2 Wood JC, Ghugre N. Hemoglobin.
2008;32:85-96.
Slide 35
35 G-EXJ-1030713 May 2012 Relaxometry methods: R2 and R2*
Several pulse sequences are included in the MRI software package
Parameters R2 (for FerriScan ) spin echo sequence T2* (and R2*)
gradient echo sequence FOV (mm)300 x 225350 x 300 Matrix (lines)256
x 176128 x 80 Resolution (mm)1.17 x 1.28 x 5.02.73 x 3.75 x 10.0 TR
(ms)2500200 TE (ms)6, 9, 12, 15, 18Minimum possible (ideally <
2.0 ms) NEX (n)11 Flip angle ()9020 BW (Hz/px)3001,950 Segments
(n)8 FatSatOn Wood JC, Ghugre N. Hemoglobin. 2008;32:85-96.
Slide 36
36 G-EXJ-1030713 May 2012 1 Wood JC, et al. Blood.
2005;106:1460-5. 2 St Pierre TG, et al. Blood. 2005;105:855-61.
Biopsy LIC (mg Fe/g dry wt) Mean R2 (Hz) 300 250 200 150 100 50 0
Hepatitis Hereditary haemochromatosis -thalassaemia/Hb E
-thalassaemia 010203040 Biopsy LIC (mg Fe/g dry wt) 350 300 250 200
150 100 50 0 0102030405060 R2 (Hz) LIC by biopsy, R = 0.98 Linear
fit using biopsy data Controls, LIC by norms alone Correlation
between R2-estimated LIC and LIC by biopsy R2-LIC calibration curve
by St Pierre et al. 2005 2 R2-LIC calibration curve by Wood et al.
2005 1
Slide 37
37 G-EXJ-1030713 May 2012 Correlation between R2*-estimated LIC
and LIC by biopsy 1 Wood JC, et al. Blood. 2005;106:1460-5. 2
Hankins JS, et al. Blood. 2009;113:4853-5. R2* (Hz) Biopsy LIC (mg
Fe/g dry wt) R = 0.97 Patients Controls Fit 0 200 400 600 800 1,000
1,200 1,400 1,600 1,800 2,000 0102030405060 30 25 20 15 10 5 0
02004006008001000 R2*MRI (Hz) LIC (mg Fe/g dry wt) Correlation
coefficient = 0.98 p < 0.001 R2*-LIC calibration curve by
Hankins et al. 2 R2*-LIC calibration curve by Wood et al. 1
Slide 38
38 G-EXJ-1030713 May 2012 LIC estimated with R2 and R2* MRI
correlate well with each other Wood JC, et al. Blood.
2005;106:1460-5. 0 10 20 30 40 50 01020304050 Estimated HIC
(mg/dry) by R2-SP Estimated HIC (mg/dry) by R2* Patient data Linear
fit, R=0.94
Slide 39
39 G-EXJ-1030713 May 2012 30 25 20 15 10 5 0 02004006008001000
Liver R2* (Hz) HIC (mg Fe/g of dry weight liver) Hankins, et al.
Wood, et al. Anderson, et al. [Fe] (mg/g dry wt) Cardiac R2* (Hz) 0
2 4 6 8 10 12 14 0100200300400 R 2 = 0.82540 Liver MRICardiac MRI
Gradient relaxometry (T2*, R2*) can conveniently measure cardiac
and liver iron HIC = hepatic iron concentration Carpenter JP, et
al. J Cardiovasc Magn Reson. 2009;11 Suppl 1:P224. Hankins et al
Blood. 2009;113:4853-4855. Cardiac and liver iron can be assessed
together conveniently by gradient echo during a single MRI
measurement.
Slide 40
40 G-EXJ-1030713 May 2012 Relaxometry methods: pros and cons
ProsCons R2* Correlate well to biopsy LIC 14 Greater sensitivity to
iron deposits 5 Faster (images can be obtained in a single
breath-hold) and easier 6 Can perform cardiac and liver iron
assessment at the same time More susceptible to artefacts Requires
expert training of a technician/ radiologist for data acquisition
and data analysis R2 (Ferriscan ) Correlate well to biopsy LIC 14
Less affected by susceptibility artefacts 6 Highly sensitive and
specific over a large range of LIC, including patients with severe
haemosiderosis 7 The gold standard method in clinical trials
Requires no training for data analysis (done centralized by
Resonance Health) Multiple breath-holds required which increases
MRI time Cost of analysis (300 USD per scan) 1 Christoforidis A, et
al. Eur J Haematol. 2009;82:388-92. 2 St Pierre TG, et al. Blood.
2005;105:855-61. 3 Wood JC, et al. Blood. 2005;106:1460-5. 4
Hankins JS, et al. Blood. 2009;113:4853-5. 5 Anderson LJ, et al.
Eur Heart J. 2001;22:2171-9. 6 Wood JC, Ghugre N. Hemoglobin.
2008;32:85-96. 7 Papakonstantinou, O, et al. J Magn Reson Imaging.
2009;29:853-9.
Slide 41
41 G-EXJ-1030713 May 2012 Relaxometry methods: R2 and R2*
(cont.) Correct position is important so that the LIC across the
whole liver can be measured Images are taken at various TEs Red
line indicates correct position of the slice 1. Patient preparation
(5 min) 2. Image acquisition (approx. 5-20 min) 3. Data analysis
(depends on experience)
Slide 42
42 G-EXJ-1030713 May 2012 Liver R2* MRI Liver with normal iron
levels Liver with severe iron overload Images courtesy of Dr J. de
Lara Fernandes. T2* = 15.7 ms or R2* = 63.7 Hz or LIC = 1.3mg/g T2*
= 1.1 ms or R2* = 909 Hz or LIC = 25.0 mg/g TE=1.3msTE=3.6ms
TE=7.1ms TE=1.3msTE=3.6ms TE=7.1ms
Slide 43
43 G-EXJ-1030713 May 2012 How can I avoid artefacts when
assessing LIC by MRI? When assessing LIC, one thing that is really
important is to use fat saturation (usually automatically included
in all the sequences). This is especially important if a patient
has steatosis (e.g. adults with haemochromatosis) How frequent are
artefacts in liver MRI? In contrast to cardiac MRI, the risk for
motion artefacts (e.g. due to breathing) or susceptibility
artefacts is much lower when performing liver MRI. As in cardiac
MRI, if artefacts are present and too severe, scans may have to be
repeated FAQ: artefacts Questions and answers were prepared under
the review of Dr J. de Lara Fernandes, University of Campinas,
Brazil.
Slide 44
44 G-EXJ-1030713 May 2012 Relaxometry methods: R2 and R2*
(cont.) Determine ROI entire liver boundary, excluding obvious
hilar vessels 1 Slice thickness varies, generally 515 mm 14 Number
of slices anything from about 1 to 20 slices can be studied 14 Red
outline shows position of ROI 1 Wood JC, et al. Blood.
2005;106:1460-5. 2 St Pierre TG, et al. Blood. 2005;105:855-61. 3
Papakonstantinou O, et al. J Magn Reson Imaging. 2009;29:853-9. 4
Hankins JS, et al. Blood. 2009;113:4853-5. 1. Patient preparation
(5 min) 2. Image acquisition (approx. 5-20 min) 3. Data analysis
(depends on experience)
Slide 45
45 G-EXJ-1030713 May 2012 Relaxometry methods: R2 and R2*
(cont.) As TE increases, SI should decrease When plotted on a graph
as iron load increases, the curve gets steeper T2 or T2* can be
calculated from the curve R2 and R2* can also be calculated
Calculations are done manually, or by specific licensed software
(e.g. CMRtools ), or images could be directly sent to a validated
centre performing FerriScan for analysis 100 80 60 40 20 0 1520 0 5
10 SI TE (ms) Typical non-iron-loaded tissue Increasing iron
loading 1. Patient preparation (5 min) 2. Image acquisition
(approx. 5-20 min) 3. Data analysis (depends on experience)
Slide 46
46 G-EXJ-1030713 May 2012 Analysis of the data The data can be
analysed manually or using post-processing software
ManuallyPost-processing software Excel spreadsheet
ThalassaemiaTools (CMRtools) cmr 42 FerriScan MRmap MATLAB
Slide 47
47 G-EXJ-1030713 May 2012 Analysis of the data (cont.)
MethodProsCons Excel spreadsheetLow cost Time-consuming Tedious
ThalassaemiaTools (CMRtools) 1 Fast (1 min) 2 Easy to use FDA
approved GBP 3,000 per year cmr 42(3) Easy to use FDA approved 3
Can generate T2*/R2* and T2/R2 maps with same software Allows
different forms of analysis Generates pixel-wise fitting with
colour maps 40,000 USD first year costs 12,000 USD per year after
FDA = Food and Drug Administration. 1
www.cmrtools.com/cmrweb/ThalassaemiaToolsIntroduction.htm. Accessed
Dec 2010. 2 Pennell DJ. JACC Cardiovasc Imaging. 2008;1:579-81. 3
www.circlecvi.com. Accessed Dec 2010.
Slide 48
48 G-EXJ-1030713 May 2012 MethodProsCons FerriScan 1
Centralized analysis of locally acquired data (206 active sites
across 25 countries) Easy set-up on most MRI machines EU approved
Validated on GE, Philips, and Siemens scanners USD 300 per scan
Patients data are sent to reference centre MRmap 2 Uses IDL
runtime, which is a commercial software (less expensive than cmr 42
/CMRtools) Can quantify T1 and T2 map with the same software Purely
a research tool Not intended for diagnostic or clinical use MATLAB
3 Low costAvailable only locally Physicists or engineers need to
write a MATLAB program for display and T2* measurement 1
www.resonancehealth.com/resonance/ferriscan. Accessed Dec 2010. 2
www.cmr-berlin.org/forschung/mrmapengl/index.html. Accessed Dec
2010. 3 Wood JC, Noetzli L. Ann N Y Acad Sci. 2010;1202:173-9.
Analysis of the data (cont.)
Slide 49
49 G-EXJ-1030713 May 2012 What is truncation? After the
selection of the ROI, the signal decay can be fitted using
different models. In the truncation model, the late points in the
curve (the plateau) are subjectively discarded to obtain a curve
with an R 2 > 0.995. A new single exponential curve is made by
fitting the remaining signals. What is the most frequent mistake
made when interpreting the data from an MRI scan? Interpreting a
liver MRI is more challenging than for a cardiac MRI, especially in
patients with severe liver iron overload. Correcting the data using
truncation analysis is very important (done automatically by some
software). The example (see following slide) clearly shows what
happens, if the truncation is not done correctly FAQ: mistakes in
manual analysis of liver MRI data Questions and answers were
prepared under the review of Dr J. de Lara Fernandes, University of
Campinas, Brazil.
Slide 50
50 G-EXJ-1030713 May 2012 Non-truncated analysis with results
with a poor R 2 (< 0.995). The apparent LIC of 4.65 suggests
mild LICs. Observe the flat plateau of the data points after a TE
of 3.62 ms The same patient, but analysing the data with only the 3
first data points results in a better (although not perfect) R 2.
The LIC results in severe iron overload, reflecting the real
concentrations of iron Analysis without truncation of the data
Analysis with truncation of the data FAQ: mistakes in manual
analysis of liver MRI data (cont.)
Slide 51
51 G-EXJ-1030713 May 2012 How to start measuring liver iron
loading in a hospital? What steps need to be taken? To start
assessing liver iron loading by MRI, these steps can be followed
1.Check MRI machine requirements 0.51.5 T (1.5 T is highly
recommended for T2* and T2 calculations; 0.5 T only for SIR)
calibrated includes a liver package 2.Optional: buy software for
analysing the data (otherwise, Excel spreadsheet can be used)
3.Optional: training of personnel for acquiring MRI images
4.Optional: training of personnel on how to analyse the data FAQ:
how to start measuring liver iron loading? Questions and answers
were prepared under the review of Dr J. de Lara Fernandes,
University of Campinas, Brazil.
Slide 52
52 G-EXJ-1030713 May 2012 LIC: interpretation of results LIC
threshold values for classification of iron overload Iron levels
LIC (mg Fe per g dry weight) LIC (mol Fe per g dry wt) R2 (s 1 )
R2* (s 1 )T2* (ms) Normal< 2< 35.6< 50< 88> 11.4
Mild overload 27 35.6 125.0 50 100 88 263> 3.8 11.4 Moderate
overload 715 125 269 100 155 263 555> 1.8 3.8 Severe overload 15
269 155 555 1.8 Values estimated based on R2 LIC calibration curve;
R2, R2* and T2* values valid for MRI machines with 1.5T only. St
Pierre TG, et al. Blood 2005;105:855861; Wood JC, et al. Blood
2005;106:14601465.
Slide 53
53 G-EXJ-1030713 May 2012 Implementation of liver and cardiac
MRI Slide presented at Global Iron Summit 2011 - With the
permission of Juliano de Lara Fernandes 1.5T MRI Scanner
Experienced radiologist Cardiac acquisition package Routine cardiac
MR exams Post-processing analysis US$1.000.000 US$50.000 US$40.000
or US$4.000/y or in-house or outsource Yes No day training 1 day
training Yes No 1-2 day training 4 day training Liver Analysis
Liver Analysis Heart Analysis Heart Analysis
Slide 54
G-EXJ-1030713 May 2012 Summary
Slide 55
55 G-EXJ-1030713 May 2012 Summary Iron overload is a serious
problem among patients who require blood transfusions to treat
anaemia and associated conditions Analysing liver iron overload is
important to predict risk of hepatic and extra-hepatic
complications The extent of iron accumulation in the liver is a key
prognostic indicator for morbidity and mortality MRI has the added
advantage that iron levels throughout the liver can be analysed,
rather than just the biopsied section (iron levels throughout the
liver can vary) R2 is the most commonly used technique in clinical
practice, although R2* is a comparable alternative across most
ranges of iron overload and is faster
Slide 56
G-EXJ-1030713 May 2012 CARDIAC MRI Diagnostic Backgrounder
NOTE: These slides are for use in educational oral presentations
only. If any published figures/tables from these slides are to be
used for another purpose (e.g. in printed materials), it is the
individuals responsibility to apply for the relevant permission.
Specific local use requires local approval
Slide 57
57 G-EXJ-1030713 May 2012 Outline Introduction to iron overload
Assessing cardiac iron loading echocardiography cardiac MRI Cardiac
MRI in practice preparation of the patient acquisition of the image
analysis of the data Excel spreadsheet ThalassaemiaTools (CMRtools)
cmr4 2 FerriScan MRmap MATLAB Summary MRI = magnetic resonance
imaging.
Slide 58
G-EXJ-1030713 May 2012 Introduction to iron overload
Slide 59
59 G-EXJ-1030713 May 2012 Introduction to iron overload Iron
overload is common in patients who require intermittent or regular
blood transfusions to treat anaemia and associated conditions it
may be exacerbated in some conditions by excess gastrointestinal
absorption of iron Iron overload can lead to considerable morbidity
and mortality 1 Excess iron is deposited in major organs, resulting
in organ damage the organs that are at risk of damage due to iron
overload include the liver, heart, pancreas, thyroid, pituitary
gland, and other endocrine organs 2,3 1 Ladis V, et al. Ann NY Acad
Sci. 2005;1054:445-50. 2 Gabutti V, Piga A. Acta Haematol.
1996;95:26-36. 3 Olivieri NF. N Engl J Med. 1999;341:99-109.
Slide 60
60 G-EXJ-1030713 May 2012 Importance of analysing cardiac iron
In -thalassaemia major, cardiac failure and arrhythmia are risk
factors for mortality 1 signs of myocardial damage due to iron
overload: arrhythmia, cardiomegaly, heart failure, and pericarditis
2 heart failure has been a major cause of death in -thalassaemia
patients in the past (5070%) 1,3 In MDS, the results of studies are
less comprehensible the reported proportion of MDS patients with
cardiac iron overload is inconsistent; from high to only a small
proportion of MDS patients 47 cardiac iron overload occurs later
than does liver iron overload 4,7,8 however, cardiac iron overload
can have serious clinical consequences in MDS patients 1
Borgna-Pignatti C, et al. Haematologica. 2004;89:1187-93. 2 Gabutti
V, Piga A. Acta Haematol. 1996;95:26-36. 3. Modell B, et al.
Lancet. 2000;355:2051-2. 4 Jensen PD, et al. Blood.
2003;101:4632-9. 5 Chacko J, et al. Br J Haematol. 2007;138:587-93.
6 Konen E, et al. Am J Hematol. 2007;82:1013-6. 7 Di Tucci AA, et
al. Haematologica. 2008;93:1385-8. 8 Buja LM, Roberts WC. Am J Med.
1971;51:209-21.
Slide 61
61 G-EXJ-1030713 May 2012 Baseline Latest follow-up p <
0.001 cT2* 20 ms cT2* < 10 ms Patients (%) cT2* = cardiac T2*. 1
Thomas AS, et al. Blood. 2010;116:[abstract 1011]. 2 Modell B, et
al. Lancet. 2000;355:2051-2. Importance of analysing cardiac iron
(cont.) In 2010, the overall mortality rate of -thalassaemia major
patients in the UK was substantially lower than a decade ago (1.65
vs 4.3 per 1,000 patient years) 1,2 due to improved monitoring and
management of iron overload over the last decade, 77% of patients
have normal cardiac T2* 1 cardiac iron overload is no longer the
leading cause of death in this population 1 60 17 23 7 0 10 20 30
40 50 60 70
Slide 62
62 G-EXJ-1030713 May 2012 Cardiac T2*: Overview of correlations
with other measurements 1 Wood JC, et al. Blood. 2004;103:1934-6. 2
Anderson LJ, et al. Eur Heart J. 2001;22:2171-9. 3 Tanner MA, et
al. J Cardiovasc Magn Reson. 2006;8:543-7. 4 Kirk P, et al.
Circulation. 2009;120:1961-8. 5 Westwood MA, et al. J Magn Reson
Imaging. 2005;22:229-33. For thalassaemia, but not sickle cell.
APFR = atrial peak filling rate; EPFR = early peak filling rate;
LIC = liver iron concentration; SF = serum ferritin. Weak or no
correlation Transfusion duration 1 Ventricular dysfunction 1-3
Arrhythmia and heart failure 4 APFR EPFR:APFR 5 Need for cardiac
medication 1-2 T2* SF and LIC 1-3
Slide 63
63 G-EXJ-1030713 May 2012 LVEF (%) 0 50 70 40 30 20 10 60 80 90
0 204060908010010305070 Cardiac T2* (ms) Cardiac T2* value of 37 ms
in a normal heart Cardiac T2* value of 4 ms in a significantly
iron-overloaded heart LVEF = left-ventricular ejection fraction.
Anderson LJ, et al. Eur Heart J. 2001;22:2171-9. Normal T2* range
Normal LVEF range Cardiac T2*: Relationship with LVEF Myocardial
T2* values < 20 ms are associated with a progressive and
significant decline in LVEF
Slide 64
64 G-EXJ-1030713 May 2012 0.1 Cardiac T2*: Relationship with
cardiac failure and arrhythmia Kirk P, et al. Circulation.
2009;120:1961-8. T2* < 10 ms: relative risk 159 (p < 0.001)
T2* < 6 ms: relative risk 268 (p < 0.001) Cardiac failure
Proportion of patients developing cardiac failure Follow-up time
(days) 600120180240300360 0.3 0.2 0 0.4 0.5 0.6 < 6 ms 68 ms 810
ms > 10 ms Arrhythmia 600120180240300360 0.15 0.10 0.05 0 0.20
0.25 0.30 < 10 ms 1020 ms > 20 ms T2* < 20 ms: relative
risk 4.6 (p < 0.001) T2* < 6 ms: relative risk 8.65 (p <
0.001) Follow-up time (days) Proportion of patients with arrhythmia
Low myocardial T2* predicts a high risk of developing cardiac
failure and arrhythmia
Slide 65
G-EXJ-1030713 May 2012 Assessing cardiac iron overload
Slide 66
66 G-EXJ-1030713 May 2012 Assessing cardiac iron loading:
Agenda Echocardiography Cardiac MRI advantages and disadvantages of
cardiac MRI MRI: a non-invasive diagnostic tool T2* is the standard
method for analysing cardiac iron
Slide 67
G-EXJ-1030713 May 2012 Echocardiography
Slide 68
68 G-EXJ-1030713 May 2012 Assessing cardiac iron loading:
Echocardiography EF = ejection fraction. 1 Leonardi B, et al. JACC
Cardiovasc Imaging. 2008;1:572-8. 2 Hoffbrand AV. Eur Heart J.
2001;22:2140-1. ProsCons Readily available 1 Relatively inexpensive
1 Does not detect early damage 2 Echocardiographic diastolic
function parameters correlate poorly with LVEF and T2* 1 Cannot
directly or indirectly quantify cardiac iron levels
Slide 69
G-EXJ-1030713 May 2012 Cardiac MRI
Slide 70
70 G-EXJ-1030713 May 2012 MRI: A non-invasive diagnostic tool
Indirectly measures levels of iron in the heart MRI measures
longitudinal (T1) and transverse (T2) relaxation times of the
protons iron deposition disrupts the homogeneous magnetic field and
shortens T1 and T2 times in a concentration-dependent manner RF =
radio-frequency. 1 Wood JC, Ghugre N. Hemoglobin. 2008;32:85-96. 2
Wood JC, et al. Circulation. 2005;112:535-43. 3 Wang ZJ, et al.
Radiology. 2005;234:749-55. 4 Ghugre NR, et al. Magn Reson Med.
2006;56:681-6. Protons Magnetic field RF/spin echo/gradient echo
Echo signal T1, T2 Signal processing Iron
Slide 71
71 G-EXJ-1030713 May 2012 MRI: A non-invasive diagnostic tool
(cont.) If a spin-echo sequence is used, the relaxation time is T2
If a gradient-echo sequence is used, it is T2* Cardiac MRI methods
gradient-echo T2* MRI: most used in clinical practice spin-echo T2
MRI: less useful (motion artefacts common due to characteristics of
the heart) TE = echo time. Adapted from Wood JC, Ghugre N.
Hemoglobin. 2008;32:85-96. Protons Magnetic field Most used in
clinical practice: Gradient echo Image acquired at different TEs
Excel or software T2* [ms} R2* [Hz]= 1,000/T2* Spin echo Image
acquired at different TEs Excel or software T2* [ms} R2* [Hz]=
1,000/T2*
Slide 72
72 G-EXJ-1030713 May 2012 Assessing cardiac iron loading:
Cardiac MRI Advantages of MRIDisadvantages of MRI Non- invasive
Rapidly assesses iron content in the septum of the heart Relative
iron burden can be reproducibly estimated Functional parameters can
be examined concurrently (e.g. LVEF) Iron status of liver and heart
can be assessed in parallel Allows longitudinal follow-up Good
correlation with morbidity and mortality outcomes Indirect
measurement of cardiac iron Requires MRI imager with dedicated
imaging method Relatively expensive and varied availability
Slide 73
73 G-EXJ-1030713 May 2012 What are sequences? Sequences are a
set of radio-frequency and gradient pulses (slight tilts in the
magnetization curves of the scanner) generated repeatedly during
the scan, which produce echoes with varied amplitudes and shapes
that will define the MR image What is gradient echo? A
gradient-echo sequence is obtained after 2 gradient impulses are
applied to the body, resulting in a signal echo that is read by the
coils. In these sequences, the spins are not refocused and,
therefore, are subject to local inhomogeneities, with a more rapid
decay curve. For gradient-echo pulse sequences, the T2* relaxation
times (which reflect these inhomogeneities) on the signal are more
significant 1 Image from Ridgway JP. J Cardiovasc Magn Reson.
2010;12:71. FAQ: Cardiac MRI
Slide 74
74 G-EXJ-1030713 May 2012 Gradient relaxometry (T2*, R2*) is
the method for analysing cardiac iron levels 1 Guo H, et al. J Magn
Reson Imaging. 2009;30:394-400. 2 Anderson LJ, et al. Eur Heart J.
2001;22:2171-9. 3 Wood JC, Noetzli L. Ann N Y Acad Sci.
2010;1202:173-9. 4 Wood JC, Ghugre N. Hemoglobin. 2008;32:85-96. 5
Westwood M, et al. J Magn Reson Imaging. 2003;18:33-9. 6 Hoffbrand
AV. Eur Heart J. 2001;22:2140-1. 7 He T, et al. Magn Reson Med.
2008;60:1082-9. T2* (gradient echo)T2 (spin echo) ProsGreater
sensitivity to iron deposition 2 Shorter acquisition time 1 Less
affected by motion artefacts 3 More readily available 3 Easier to
perform 4 Good reproducibility 5 Less affected by susceptibility
artefacts 1, due to metal implants, airtissue interfaces, proximity
to cardiac veins ConsMore sensitive to static magnetic field
inhomogeneity 1 Noise, motion, and blood artefacts can complicate
analysis (particularly in heavily iron-loaded hearts) 7 Lack of
sensitivity 6 Motion artefacts 6 Poor signal-to-background noise
ratios at longer TEs 6 Longer acquisition time 1
Slide 75
75 G-EXJ-1030713 May 2012 HIC = hepatic iron concentration
Carpenter JP, et al. J Cardiovasc Magn Reson. 2009;11 Suppl 1:P224.
Hankins et al Blood. 2009;113:4853-4855. 30 25 20 15 10 5 0
02004006008001000 Liver R2* (Hz) HIC (mg Fe/g of dry weight liver)
Hankins, et al. Wood, et al. Anderson, et al. [Fe] (mg/g dry wt)
Cardiac R2* (Hz) 0 2 4 6 8 10 12 14 0100200300400 R 2 = 0.82540
Liver MRI Cardiac MRI Gradient relaxometry (T2*, R2*) can
conveniently measure cardiac and liver iron Cardiac and liver iron
can be assessed together conveniently by gradient echo during the a
single MRI measurement.
Slide 76
76 G-EXJ-1030713 May 2012 Cardiac T2* MRI is usually measured
in the septum of the heart Heart with normal iron levels Heart with
severe iron overload Images courtesy of Dr J. de Lara Fernandes.
T2* = 22.8 ms or R2* = 43.9 Hz T2* = 5.2 ms or R2* = 192 Hz
Slide 77
77 G-EXJ-1030713 May 2012 Conversion from T2* to R2* is a
simple mathematical calculation: R2* = 1,000/T2* Level of cardiac
iron overloadT2*, msR2*, Hz Normal 20 1 < 50 Mild, moderate1020
1 50100 Severe< 10 2 > 100 1 Anderson LJ, et al. Eur Heart J.
2001;22:2171-9. 2 Kirk P, et al. Circulation. 2009;120:1961-8.
These values are only applicable to 1.5 T scanners 1 What is
R2*?
Slide 78
78 G-EXJ-1030713 May 2012 Why should the data be presented as
R2* and not T2*? Seven whole hearts from patients with
transfusion-dependent anaemias were assessed by histology and
cardiac MRI [Fe] (mg/g dry wt) Cardiac T2* (ms) [Fe] (mg/g dry wt)
0 2 4 6 8 10 12 14 010203040506070 R 2 = 0.949 Cardiac R2* (Hz) 0 2
4 6 8 10 12 14 0100200300400 R 2 = 0.82540 Carpenter JP, et al. J
Cardiovasc Magn Reson. 2009;11 Suppl 1:P224. R2* has a linear
relationship with tissue iron concentration, which simplifies the
interpretation of data and allows comparison of changes over
time
Slide 79
79 G-EXJ-1030713 May 2012 Anderson LJ, et al. Eur Heart J.
2001;22:2171-9. Hockey stick effect?Or a more gradual relationship?
The relationship between cardiac T2*/R2* and LVEF Heart T2* (ms)
LVEF (%) R2* (s 1 ) LVEF (%) 90 80 70 60 50 40 30 20 10 0
2030405060708090100 0 80 60 40 20 0 050100150200250 Why should the
data be presented as R2* and not T2*? (cont.) R2* allows
demonstration of cardiac risk in a more gradual way
Slide 80
80 G-EXJ-1030713 May 2012 Transform to R2* Standard errors on a
single measurement are approximately constant with R2*, but are
non-uniform with T2* Westwood M, et al. J Magn Reson Imaging.
2003;18:33-9. 60 50 40 30 20 10 0 0 2030405060 T2* second
measurement (ms) T2* first measurement (ms) 120 100 80 60 40 20 0 0
406080100120 R2* second measurement (s 1 ) R2* first measurement (s
1 ) Why should the data be presented as R2* and not T2*? (cont.)
R2* has a constant standard error that makes assessment of the
significance of changes easier
Slide 81
G-EXJ-1030713 May 2012 Cardiac T2* MRI in practice
Slide 82
82 G-EXJ-1030713 May 2012 MRI scanners Manufacturers Siemens
Healthcare (Erlangen, Germany; www.siemensmedical.com) GE
Healthcare (Milwaukee, WI, USA; www.gemedicalsystems.com) Philips
Healthcare (Best, the Netherlands; www.medical.philips.com)
Magnetic field T2* varies with magnetic field strength 1 need 1.5 T
for cutoff levels of 20 ms (iron overload) and 10 ms (severe iron
overload) 1,2 Cardiac package needs to be acquired separately from
the manufacturers. The cost is about USD 40,000. However, in most
centres, this is available since MRI is frequently used in
non-iron-related cardiovascular imaging includes all necessary for
acquisition of the image sequences are included in Siemens and
Philips Healthcare cardiac packages, but for GE Healthcare they
need to be acquired separately (note: variations may exist between
countries) 1 Anderson LJ, et al. Eur Heart J. 2001;22:2171-9. 2
Kirk P, et al. Circulation. 2009;120:1961-8.
Slide 83
83 G-EXJ-1030713 May 2012 Cardiac T2* MRI in practice: The
process T2*, R2* *Time to manually calculate T2*/R2* values in an
Excel spreadsheet depends on the experience of the physician. 1.
Patient preparation (5 min) 2. Acquisition of the MRI image
(approx. 5-20 min) 3. Analysis of MRI data (time depends on
experience*)
Slide 84
84 G-EXJ-1030713 May 2012 Cardiac T2* MRI in practice: The
process (cont.) Preparation of the patient Acquisition of the image
Analysis of the data (post-processing) Excel spreadsheet
ThalassaemiaTools, CMRtools cmr 42 FerriScan MRmap MATLAB
Slide 85
G-EXJ-1030713 May 2012 Preparation of the patient
Slide 86
86 G-EXJ-1030713 May 2012 Preparation of the patient Standard
precautions need to be taken There is no need for peripheral vein
access since no contrast agent is required Special care remove all
infusion/medication pumps (e.g. with insulin, pain-relieving drugs)
stop continuous i.v. application of ICT during the measurement ECG
signal should be positioned according to scanner specifications ECG
= electrocardiography.
Slide 87
87 G-EXJ-1030713 May 2012 Cardiac T2* MRI in practice: The
process (cont.) Preparation of the patient Acquisition of the image
Analysis of the data (post-processing) Excel spreadsheet
ThalassaemiaTools, CMRtools cmr 42 FerriScan MRmap MATLAB
Slide 88
G-EXJ-1030713 May 2012 Acquisition of the image
Slide 89
89 G-EXJ-1030713 May 2012 Acquisition of the image: MRI pulse
sequences Pulse sequences are a preselected set of defined
radio-frequency and gradient pulses are computer programs that
control all hardware aspects of the scan determine the order,
spacing, and type of radio-frequency pulses that produce magnetic
resonance images according to changes in the gradients of the
magnetic field Several different pulse sequences exist 1 a
gradient-echo sequence generates T2* 1 Wood JC, Ghugre N.
Hemoglobin. 2008;32:85-96.
Slide 90
90 G-EXJ-1030713 May 2012 1 Anderson LJ, et al. Eur Heart J.
2001;22:2171-9. 2 Westwood M, et al. J Magn Reson Imaging.
2003;18:33-9. 3 He T, et al. J Magn Reson Imaging. 2007;25:1205-9.
4 He T, et al. Magn Reson Med. 2008;60:1082-9. 5 Pepe A, et al. J
Magn Reson Imaging. 2006;23:662-8. SequenceGroup Number of echoes
per breath-hold Heart regions Pre- pulse RR intervals TR Bright
blood (Anderson et al.) 1 London (Pennell) 1 (but multiple
breath-holds) 1 (septum)No1Variable Novel bright blood (Westwood et
al) 2 London (Pennell) Multiple1 (septum)No1Fixed Black blood (He
et al) 3-4 London (Pennell) Multiple1 (septum)Yes2Fixed Multi-slice
(Pepe et al) 5 Pisa (Pepe) Multiple Multi- region No1Fixed The most
common commercially available T2* acquisition techniques The
various techniques give clinically comparable results. 2-3, 5
Slide 91
91 G-EXJ-1030713 May 2012 Acquisition of the image: TEs Images
are taken at a minimum of 5 different TEs, normally 8 12 1 The
choice of minimum TE determines the smallest measurable T2 1
ideally, min TE 2 ms, max TE 17 20 ms Different T2* acquisition
techniques according to TE multiple breath-hold: acquire an image
for each TE in separate breath-holds 2 single breath-hold
multi-echo acquisition: acquire images for all TE during 1
breath-hold 3 Mean R2* compared with true value in the case of
synthetic images for different minimum TEs, but same echo duration
(18 ms) 4 1 Wood JC, Noetzli L. Ann N Y Acad Sci. 2010;1202:173-9.
2 Anderson LJ, et al. Eur Heart J. 2001;22:2171-9. 3 Westwood M, et
al. J Magn Reson Imaging. 2003;18:33-9. 4 Ghugre NR, et al. J Magn
Reson Imaging. 2006;23:9-16. 500 450 400 350 300 250 200 150 100 50
0 0100200300400500 True R2* (Hz) Mean R2*: ramp, dualtone, &
uniform (Hz) Shortest TE = 2 ms Shortest TE = 1 ms Shortest TE = 4
ms Shortest TE = 5.5 ms True
Slide 92
92 G-EXJ-1030713 May 2012 How does the MRI data output looks
like? Data visualizationMRI data output 1 Wood JC, Ghugre N.
Hemoglobin. 2008;32:85-96. During a single breath hold the pulse
sequence run several times at increasing echo time (TE), generating
data points corresponding to decreased signal intensity 1 FrameTE
(ms)Mean ST 01.989.5 13.683.6 25.376.8 37.070.6 48.764.5 510.459.2
612.254.9 713.950.2 815.645.8 917.342.4
Slide 93
93 G-EXJ-1030713 May 2012 Which is recommended: single or
multiple breath-hold technique? Comparison of the 2 methods, single
and multiple breath-hold, showed no significant skewing between T2*
values in all patients with -thalassaemia major, regardless of
their T2* value (see Bland- Altman plots) 1 However, in cardiac MRI
the most recommended technique is single breath-hold, because it
allows quick acquisition of the information. This is especially
important to avoid movement artefacts (heart beating, breathing)
and assure the good quality of the MRI image 1 Westwood M, et al. J
Magn Reson Imaging. 2003;18:33-9. Patients with T2* < 20 ms 1
Patients with T2* 20 ms 1 FAQ: Acquisition technique
Slide 94
94 G-EXJ-1030713 May 2012 Acquisition of the image Single
breath-hold multi-echo acquisition take a short-axis slice of the
ventricle (halfway between the base and the apex): orange line
image acquisition should occur immediately after the R wave do not
alter any settings that could alter TE (e.g. FOV) Image courtesy of
Dr J. de Lara Fernandes.
Slide 95
95 G-EXJ-1030713 May 2012 Cardiac T2* MRI in practice: The
process (cont.) Preparation of the patient Acquisition of the image
Analysis of the data (post-processing) Excel spreadsheet
ThalassaemiaTools, CMRtools cmr 42 FerriScan MRmap MATLAB
Slide 96
G-EXJ-1030713 May 2012 Analysis of the data
(post-processing)
Slide 97
97 G-EXJ-1030713 May 2012 How T2* is calculated from the MRI
output? Data visualization 1 Wood JC, Ghugre N. Hemoglobin.
2008;32:85-96. Curve Fitting T2* Noise level T2* calculation is
fitting a curve on the data points and calculating at what echo
time no signal is left from iron (only noise) 1
Slide 98
98 G-EXJ-1030713 May 2012 Analysis of the data The data can be
analysed manually or using post-processing software
ManuallyPost-processing software Excel spreadsheet
ThalassaemiaTools (CMRtools) cmr 42 FerriScan MRmap MATLAB
Slide 99
99 G-EXJ-1030713 May 2012 Analysis of the data (cont.)
MethodProsCons Excel spreadsheetLow costTime-consuming Tedious
ThalassaemiaTools (CMRtools) 1 Fast (1 min) 2 Easy to use FDA
approved GBP 3,000 per year cmr 42(3) Easy to use FDA approved 3
Can generate T2*/R2* and T2/R2 maps with same software Allows
different forms of analysis Generates pixel-wise fitting with
colour maps 40,000 USD first year costs 12,000 USD per year after
FDA = Food and Drug Administration. 1
www.cmrtools.com/cmrweb/ThalassaemiaToolsIntroduction.htm. Accessed
Dec 2010. 2 Pennell DJ. JACC Cardiovasc Imaging. 2008;1:579-81. 3
www.circlecvi.com. Accessed Dec 2010.
Slide 100
100 G-EXJ-1030713 May 2012 MethodProsCons FerriScan 1
Centralized analysis of locally acquired data (206 active sites
across 25 countries) Easy set-up on most MRI machines EU approved
Validated on GE, Philips, and Siemens scanners USD 100 per scan
Patients data are sent to reference centre MRmap 2 Uses IDL
runtime, which is a commercial software (less expensive than cmr 42
/CMRtools) Can quantify T1 and T2 map with the same software Purely
a research tool Not intended for diagnostic or clinical use MATLAB
3 Low costAvailable only locally Physicists or engineers need to
write a MATLAB program for display and T2* measurement 1
www.resonancehealth.com/resonance/ferriscan. Accessed Dec 2010. 2
www.cmr-berlin.org/forschung/ mrmapengl/index.html. Accessed Dec
2010. 3 Wood JC, Noetzli L. Ann N Y Acad Sci. 2010;1202:173-9.
Analysis of the data (cont.)
Slide 101
101 G-EXJ-1030713 May 2012 What are the most common mistakes in
analysing the data that could lead to a wrong interpretation of the
T2* value? Interpreting the data from cardiac MRI is usually quite
straightforward; problems may arise when analysing data from
patients with severe cardiac iron overload. In this case, the
signal from heavily iron-loaded muscle will decay quickly and a
single exponential decay curve does not fit the data well. 1 Models
exist that can help to solve this issue (see next slide): 1. the
offset model (Prof Wood and colleagues) 2. truncation of the data
(Prof Pennell and colleagues) Both models should give comparable
results; the differences should not be clinically relevant 1 Wood
JC, Noetzli L. Ann N Y Acad Sci. 2010;1202:173-9. 2 Ghugre NR, et
al. J Magn Reson Imaging. 2006;23:9-16. Signal decay curve from a
patient with T2* 5 ms, showing that the data do not fit well 2 FAQ:
Mistakes in analysing the data
Slide 102
102 G-EXJ-1030713 May 2012 What is truncation? After the
selection of the ROI, the signal decay can be fitted using
different models. In the truncation model, the late points in the
curve that form a plateau are subjectively discarded; the objective
is to have a curve with an R 2 > 0.995. A new single exponential
curve is made by fitting the remaining signals. 1 Generally, a
truncation model should be used with the bright-blood technique to
obtain more reproducible and more accurate T2* measurements 1 What
is an offset model? The offset model consists of a single
exponential with a constant offset. Using only the exponential
model can underestimate the real T2* values (at quick signal loss
at short TE, there is a plateau), while inclusion of the offset
model into the fitting equation can improve this. 2 Generally, the
offset model is recommended to be used with the black-blood
technique 1 He T, et al. Magn Reson Med. 2008;60:1082-9. 2 Ghugre
NR, et al. J Magn Reson Imaging. 2006;23:9-16. FAQ: Mistakes in
analysing the data (cont.)
Slide 103
103 G-EXJ-1030713 May 2012 How to start measuring cardiac iron
loading in a hospital? What steps need to be taken? To start
assessing cardiac iron loading by MRI, these steps can be followed:
1.Check MRI machine requirements 1.5 T calibrated 2.Buy cardiac
package from the manufacturer. It must include all that is
necessary for acquisition of the data (the sequences are included
with Siemens and Philips Healthcare cardiac packages, but for GE
Healthcare they need to be acquired separately) 3.Optional: buy
software for analysing the data (if not, Excel spreadsheet can be
used) 4.Highly recommended: training of personnel for acquisition
of cardiac MR images (e.g. functional analyses) 5.Highly
recommended: training of personnel on how to analyse the data with
the chosen software FAQ: How to start measuring cardiac iron
loading?
Slide 104
104 G-EXJ-1030713 May 2012 Implementation of liver and cardiac
MRI 1.5T MRI Scanner Experienced radiologist Cardiac acquisition
package Routine cardiac MR exams Post-processing analysis
US$1.000.000 US$50.000 US$40.000 or US$4.000/y or in-house or
outsource Yes No day training 1 day training Yes No 1-2 day
training 4 day training Liver Analysis Liver Analysis Heart
Analysis Heart Analysis Slide presented at Global Iron Summit 2011
- With the permission of Juliano de Lara Fernandes
Slide 105
G-EXJ-1030713 May 2012 Summary
Slide 106
106 G-EXJ-1030713 May 2012 Summary Iron overload is common in
patients who require intermittent or regular blood transfusions to
treat anaemia and associated conditions Analysing cardiac iron
levels is important in -thalassaemia major, cardiac failure and
arrhythmia are risk factors for mortality in MDS, cardiac iron
overload can have serious clinical consequences due to improved
monitoring and management of iron overload over the last decade,
77% of patients have normal cardiac T2* 1 MRI: the method to
rapidly and effectively assess cardiac iron loading T2* allows
specific assessment of cardiac iron levels. The use of this
convenient, non-invasive procedure has had a significant impact on
outcomes in patients with cardiac iron overload 1 R2* is a simple
calculation from T2* and has a linear relationship with cardiac
iron concentration 1 Thomas AS, et al. Blood. 2010;116:[abstract
1011]. 2 Modell B, et al. J Cardiovasc Magn Reson.
2008;10:42-9.
Slide 107
G-EXJ-1030713 May 2012 CYTOGENERIC ASSESSMENT IN MDS NOTE:
These slides are for use in educational oral presentations only. If
any published figures/tables from these slides are to be used for
another purpose (e.g. in printed materials), it is the individuals
responsibility to apply for the relevant permission. Specific local
use requires local approval.
Slide 108
108 G-EXJ-1030713 May 2012 Outline MDS Classification and
prognosis scoring Practical guide to bone marrow aspirate analysis
in MDS Cytogenetics of MDS in bone marrow aspirates Summary LIC =
liver iron concentration; MRI = magnetic resonance imaging; SF =
serum ferritin; SIR = signal intensity ratio; SQUID =
superconducting quantum interface device.
Slide 109
109 G-EXJ-1030713 May 2012 What are the myelodysplastic
syndromes (MDS)? MDS are a spectrum of heterogeneous myeloid clonal
disorders Occurrence: De novo (primary MDS) Secondary or
treatment-related MDS MDS are associated with significant morbidity
and mortality due to: Risk of transformation to acute myeloid
leukemia (AML) Cytopenias Impaired quality of life (frequent
transfusions, iron overload, etc)
Slide 110
110 G-EXJ-1030713 May 2012 Common features of MDS MDS are
marked by ineffective haematopoiesis and defective development of
blood cells, e.g.: dyserythropoiesis (affects red blood cells
production) dysgranulopoiesis (affects granulocytes production)
dysmegakaryopoiesis (affects platelet production) Ineffective
haematopoiesis and maturation of the blood cells results in one or
more cytopenias, e.g.: anaemia (reduced red blood cell count)
neutropenia (reduced absolute neutrophil count) thrombocytopenia
(reduced number of platelets)
Slide 111
111 G-EXJ-1030713 May 2012 Minimal diagnostic criteria in MDS
consensus, Vienna 2006 Prerequisite criteria constant cytopenia in
one or more cell lineages complete blood count exclusion of all
other causes of cytopenia / dysplasia MDS-related (decisive)
criteria dysplasia in > 10% of all bone marrow cells in one or
more of the lineages, or > 15% ringed sideroblasts complete
blood count iron staining of bone marrow smears 519% blast cells
bone marrow smears typical chromosomal abnormality karyotyping or
FISH FISH = fluorescence in situ hybridization. Loken MR, et al.
Leuk Res. 2008;32:5-17. Nimer SD. Blood. 2008;111:4841-51. Valent
P, et al. Leuk Res. 2007;31:727-36.
Slide 112
112 G-EXJ-1030713 May 2012 Minimal diagnostic criteria in MDS
consensus, Vienna 2006 (cont.) Additional criteria (for patients
not fulfilling the decisive MDS criteria) abnormal phenotype of
bone marrow cells flow cytometry molecular signs of a monoclonal
cell population HUMARA assay, gene chip profiling, point mutation
or SNP analysis markedly and persistently reduced colony formation
CFU assay CFU = colony-forming unit; SNP-a = single-nucleotide
polymorphism. Loken MR, et al. Leuk Res. 2008;32:5-17. Van de
Loosdrecht AA. Leuk Res. 2008;32:205-7. Van de Loosdrecht AA, et
al. Blood. 2008;111:1067-77. Van de Loosdrecht AA, et al.
Haematologica. 2009;94:1124-1134. Nimer SD. Blood.
2008;111:4841-51. Valent P, et al. Leuk Res. 2007;31:727-36.
Slide 113
G-EXJ-1030713 May 2012 MDS classification and prognostic
scoring
Slide 114
114 G-EXJ-1030713 May 2012 Diagnostic and prognostic of MDS
Risk-stratification is necessary for clinical decision-making:
predicts treatment outcomes predicts survival predicts risk of
progression to AML Classification systemBasis of evaluation Derived
prognostic score system FAB (1982)Cellular morphologyIPSS (1997)
WHO (2002) Cellular morphology, cytogenetics WPSS (2007) FAB,
French-American-British; IPSS, International Prognostic Scoring
System; WPSS, WHO-based Prognostic Scoring System. Bennett JM, et
al. Br J Haematol 1982;51:189199; Jaffe, et al, eds. Lyon: IARC
Press; 2001; Greenberg P, et al. Blood 1997;89:20792088; Malcovati
L, et al. J Clin Oncol 2007;23:35033510.
Slide 115
115 G-EXJ-1030713 May 2012 French-American-British (FAB)
classification Blast percentage MDS Subtype Peripheral blasts (%)
Bone marrow blasts (%) Additional features AML transformation (%)
RARefractory anaemia140 Bennett JM, et al. Br J Haematol
1982;51:189199.
117 G-EXJ-1030713 May 2012 Risk stratification and prognosis
scoring for MDS: IPSS Influence of karyotype according to IPSS Good
= normal, Y, del(5q), del(20q) Poor = complex ( 3 abnormalities) or
chromosome 7 anomalies Int. = all other abnormalities
Variable00.51.01.52.0 BM blasts (%)< 551011202130
KaryotypeGoodInt.Poor Cytopenia(s)0/12/3 Greenberg P, et al. Blood.
1998;89:2079-88. IPSS score
Slide 118
118 G-EXJ-1030713 May 2012 Years Cumulative survival of MDS
patients by IPSS Low Int-1 Int-2 High 0246810141618 0 10 20 30 40
50 60 70 80 90 100 Survival Years Percent 0246810141618 0 10 20 30
40 50 60 70 80 90 100 Percent AML Evolution Low Int-1 Int-2 High
Greenberg P, et al. Blood 1997;89:20792088. 12
Slide 119
119 G-EXJ-1030713 May 2012 MDS: WHO classification 2008 Blast
percentage MDS Subtype Dysplasia Peripheral blasts (%) Bone marrow
blasts (%) Ringed sideroblasts (%) Cytogenetics 5q syndromeMostly
DysE< 1%< 5%< 15%5q sole RA, RN, RT, RCUDDysE, N, T<
1%< 5%< 15%Various RARSMostly DysE0< 5%> 15%Various
RCMD23 lineagesrare< 5%< 15%Various RAEB-113 lineages<
5%59%< 15%Various RAEB-213 lineages519% Auer rods +/- 1019% Auer
rods +/- < 15%Various MDS-U1 lineage< 1%< 5%<
15%Various BM = bone marrow; DysE = dyserythropoiesis; MDS-U =
myelodysplastic syndrome, unclassified; N = neutropenia; pB =
peripheral blood; RCMD = refractory cytopenia with multilineage
dysplasia; RCUD = refractory cytopenia with unilineage dysplasia;
RN = refractory neutropenia; RT = refractory thrombocytopenia; T =
thrombocytopenia. Swerdlow SH, et al., eds. WHO Classification of
Tumours of Haematopoietic and Lymphoid Tissues. Lyon: IARC Press;
2008:109-38.
Slide 120
120 G-EXJ-1030713 May 2012 Survival of MDS patients according
to transfusion dependency Transfusion-dependent patients
Transfusion-independent patients Time (months) Proportion surviving
0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 020406080100120140
Cazzola M, Malcovati L. N Engl J Med. 2005;352:536-8.
Slide 121
121 G-EXJ-1030713 May 2012 The WHO classification-based
prognostic scoring system (WPSS) for MDS Points 0123 WHO subtype
RA, RARS, del(5q) RCMD, RCMD-RS RAEB-1RAEB-2 Transfusion
requirementNoneRegular Cytogenetic categoryGoodInt.Poor Risk
groupsScoreMedian survival (months) Very low0103 Low172
Intermediate240 High3421 Very high5612 Malcovati L, et al. J Clin
Oncol. 2007;25:3503-10. Transfusion dependence is an independent
indicator of severity of disease and has a significant effect on
survival
Slide 122
122 G-EXJ-1030713 May 2012 Overall survival and AML risk
assessments in MDS by WPSS Malcovati L et al. J Clin Oncol
2007;25:35033510.
Slide 123
123 G-EXJ-1030713 May 2012 WHO prognosis scoring system allows
time dependent prognosis scoring IPSSWPSS ProsMore widely used and
recognized Allows time-dependent prognosis scoring and risk
stratification Takes into account individual patient transfusion
need ConsApplies only at the time of diagnosis Underestimates the
impact of transfusion requirement and cytogenetics Underestimates
the impact of poor cytogenetics Greenberg P, et al. Blood
1997;89:20792088. Malcovati L, et al. J Clin Oncol.
2007;25:35033510. Schanz J, et al. Blood. 2007;110:[abstract 248].
Kantarjian H, et al. Cancer. 2008;113:1351-61. Garcia-Manero G, et
al. Leukemia. 2008;22:538-43.
Slide 124
G-EXJ-1030713 May 2012 Practical guide to bone marrow aspirate
analysis in MDS
Slide 125
125 G-EXJ-1030713 May 2012 Bone marrow biopsy How is it done:
Marrow aspirate and bone (trephine) biopsy are removed by physician
in an outpatient procedure under local anesthesia heparinized
sample can be stored at room temperature for 24 hours What tests
are done? assessment of cellularity, architecture and focal
collection of blasts for hematopoietic dysplasia smear staining
examination cytogenetic analysis karyotype FISH What other tests
might be done? flow cytometry cell counts cell sorting
immunophenotyping Hellstrm-Lindberg E. Myelodysplastic Syndromes.
London: Remedica; 2008. Van de Loosdrecht, et al,
Haematologica.2009; 94:1124-34. Image from: Medline Plus. NIH/NLM.
http://www.nlm.nih.gov/medlineplus/ency/imagepages/1129.htm.
Accessed Jan, 2011.
Slide 126
126 G-EXJ-1030713 May 2012 Analysis of bone marrow smears Bone
marrow aspirate smears are prepared on a slide by a medical
technician using: Wrights stain Perls staining (for ringed
sideroblasts) May-Grnwald-Giemsa staining Immunohistochemical
stainings, i.e. CD34 A pathologist or hematologist then examines
the slides for cell abnormalities under microscope: at least 400
nucleated cells and 20 megakaryocytes should be examined for
morphology Hellstrm-Lindberg E. Myelodysplastic Syndromes. London:
Remedica; 2008.
Slide 127
127 G-EXJ-1030713 May 2012 Abnormal large megakaryocyte (double
arrow), abnormal hypo-granular and Pelger neutrophils (single
arrows). Refractory anaemia (RA) Analysis of bone marrow smears
(cont.) Refractory anaemia with excess blasts (RAEB) Erythroid
cells with peri-nuclear iron accumulation Prussian blue staining
(left); Perls stain showing ringed sideroblasts with peri-nuclear
(mitochondrial) deposition of iron (right) ASH Image Bank, used
with permission, all rights reserved. Courtesy of J. Goasguen,
Universit de Rennes, France.
Slide 128
128 G-EXJ-1030713 May 2012 Immunophenotyping by flow cytometry
Various lineages are labeled with antibodies that recognize
specific haematopoeitic identifiers Combination of no more than
four labels recommended Important for identification of blasts -
CD34+/abnormal granularity/CD45dim Well-correlated with other
diagnostic techniques and prognostic systems Van de Loosdrecht, et
al. Haematologica.2009; 94:1124-34. Image from:
http://www.bio.davidson.edu/courses/genomics/method/FACS.html
Slide 129
G-EXJ-1030713 May 2012 Cytogenetics of MDS in bone marrow
aspirates
Slide 130
130 G-EXJ-1030713 May 2012 Importance of cytogenetic analysis
in MDS Nearly half of the patients with MDS present with
cytogenetic abnormalities 1 changes have a pathogenetic relevance
(i.e. loss or gain of gene function) Cytogenetic analysis is
essential for the diagnosis and classification of MDS according to
IPSS and WPSS 2 Chromosomal aberrations have prognostic relevance
for OS and for the time to leukaemic transformation, independent of
other factors Cytogenetic analysis forms the basis for therapeutic
decisions cytogenetics is indicative of response to therapy, i.e.
lenalidomide in del(5q) 3 and azacitidine in 7/del(7q) 4 IPSS =
International Prognostic Scoring System; OS = overall survival;
WPSS = WHO classification-based Prognostic Scoring System. 1 Haase
D, et al. Blood. 2007;110:4385-95. 2 Greenberg P, et al. Blood.
1998;89:2079-88. 3 List AF, et al. N Engl J Med. 2006;355:1456-65.
4 Fenaux P, et al. Lancet Oncol. 2009;10:223-32.
Slide 131
131 G-EXJ-1030713 May 2012 Cytogenetic aberrations are frequent
in patients with MDS 9% 14% 29% 48% Normal karyotype One
abnormality Two abnormalities Complex karyotype N = 2,072 patients
with MDS Haase D, et al. Blood. 2007;110:4385-95.
Slide 132
132 G-EXJ-1030713 May 2012 WHO 2008: Incidence of the most
common cytogenetic aberrations in MDS (over 5%) Unbalanced
aberrations De novo MDS (%) Secondary MDS (%) +810 7/del(7q)1050
5/del(5q)1040 del(20q)5858 Y5 iso(17q)/t(17p)/del(17p)3535 Swerdlow
SH, et al. In: WHO Classification of Tumours of Haematopoietic and
Lymphoid Tissues. 4th ed. Lyon: IARC; 2008:441. Unbalanced
aberrations with loss of genetic information (deletions or
monosomies) are most common in MDS; balanced aberrations are
rare
Slide 133
133 G-EXJ-1030713 May 2012 When should cytogenetic testing be
performed in patients with MDS: Diagnosis WHO 2008 guidelines
recommend a complete cytogenetic analysis of BM at initial
diagnosis in all patients with MDS Cytogenetic analysis is
mandatory for diagnosis of MDS associated with del(5q) patients
with refractory cytopenia(s) who lack MDS diagnostic features;
these patients may be considered as having presumptive evidence of
MDS if they have MDS-related cytogenetic abnormalities (slides 4
and 5) rather than indicating abnormality, isolated loss of Y
chromosome might be an age-related phenomenon and mosaicism with
trisomy 8 might be a constitutional change. Therefore, they might
not be sufficient to prove MDS BM = bone marrow. Swerdlow SH, et
al. In: WHO Classification of Tumours of Haematopoietic and
Lymphoid Tissues. 4th ed. Lyon: IARC; 2008:441. Valent P, Horny HP.
Eur J Clin Invest. 2009;39:548-53. Vardiman JW, et al. Blood.
2009;114:937-51.
Slide 134
134 G-EXJ-1030713 May 2012 Is cytogenetic testing required for
all patients? Yes, cytogenetic testing should be performed whenever
possible at initial diagnosis and every 612 months during follow-up
since karyotype and prognosis might change during the course of the
disease. There are a few exceptions that will have no therapeutic
consequences, e.g. very frail and multi-morbid patients. Are there
disease presentations that correlate with specific cytogenetic
abnormalities? There is no correlation between specific
abnormalities and disease presentation (with exception of the
association between isolated del(5q) syndrome and RA, with typical
dysplasia of megakaryocytes). FAQs: Cytogenetic testing for
diagnosis of patients with MDS Questions and answers were prepared
under the review of Dr. Haase, University of Gttingen,
Germany.
Slide 135
135 G-EXJ-1030713 May 2012 FAQs: Cytogenetic testing during
treatment in patients with MDS Why is it important to perform
cytogenetic testing during the treatment course? Cytogenetic
remissions represent a better quality of remission and are more
reliable than those determined by blood films or cytomorphology
from the BM. Furthermore, karyotype might change during the course
of the disease affecting the prognosis and the treatment of the
patient. First data show that a cytogenetic progression might be
detectable several weeks before clinical manifestation. Is
cytogenetic testing important for all patients or is it recommended
for specific focus groups? All patients with clonal abnormalities
identified before start of therapy should be followed up during
therapy. Also, patients with initially normal karyotype might
develop abnormalities during the course of disease and, therefore,
require regular monitoring. Questions and answers were prepared
under the review of Dr. Haase, University of Gttingen,
Germany.
Slide 136
136 G-EXJ-1030713 May 2012 Cytogenetics from BM is recommended
when possible at least once every 6 months during therapy. When
FISH is performed using peripheral blood (i.e. CD34-FISH), testing
should be conducted every 3rd month. How often should cytogenetic
testing be performed? When should treatment be altered as a result
of changes in the cytogenetic profile? When a clear cytogenetic
progression is seen, or when an abnormal clone is completely
eliminated and karyotype turns normal and stays normal over time
(at least 3 months); this depends on the type of therapy. What is
the definition of cytogenetic progression in MDS? 1) Occurrence of
new cytogenetic abnormalities (first in patients with normal
karyotype or additional for patients with abnormalities). 2)
Significant increase (> 50%) of the size of the clone with
certain cytogenetic abnormalities. FAQs: Cytogenetic testing during
treatment in patients with MDS (cont.) Questions and answers were
prepared under the review of Dr. Haase, University of Gttingen,
Germany.
Slide 137
137 G-EXJ-1030713 May 2012 Chromosome analysis in MDS:
Karyotyping Each cell in the body contains 46 chromosomes
representing the normal human karyotype MDS patients frequently
have a clonal abnormality in the hematopoietic progenitor cells,
where a proportion of these have an abnormal karyotype with altered
number of chromosomes and/or large alterations in their structure
Changes in the number of the chromosomes is due to monosomies (loss
of a chromosome) and/or polysomies (more then 2 copies of a
chromosome) Changes in the structure of the chromosomes are
commonly noted as: deletions, when part or entire chromatid is
missing insertions, when additional material is included in a
chromosome translocations, when genetic material is exchanged
between chromosomes
Slide 138
138 G-EXJ-1030713 May 2012 How to perform cytogenetic testing
in patients with MDS: Karyotyping Sample Collect at least 1020 mL
of heparinized BM aspirates Storage Samples should reach the lab
within 24 hours after biopsy Number of metaphases needed Analyse 25
metaphases if possible, especially if the karyotype is normal, to
exclude a small aberrant cell clone 678910 Metaphase quality To
detect structural abnormalities, adequate chromosome banding is
required ( 150250 bands per karyogram) See presenter for
references.
Slide 139
139 G-EXJ-1030713 May 2012 How to perform cytogenetic testing
in patients with MDS: Karyotyping (cont.) Medical technician
prepares metaphase slides for karyotyping, by: culturing bone
marrow aspirate for 24-72 hours, and then exposing the cells to
slightly hypotonic solution synchronizing them in metaphase (e.g.
using colchicine) finally, staining them (e.g. DAPI staining) and
fixing them on slides A pathologist or hematologist then examines
the slides for chromosomal abnormalities under microscope: Samples
are examined under microscope manually or with aid of analysis
software (separating, enhancing banding pattern, chromosome
pairing) At least 25 metaphases should be examined, especially if
the karyotype is normal, to exclude a small aberrant cell clone
Holland and Frei. Cancer Medicine 6. American Cancer Society; 2003.
Lucia Cytogenetics. http://www.lucia.cz/en/products/lucia-karyo.
Accessed Feb.2011.
Slide 140
140 G-EXJ-1030713 May 2012 Can peripheral blood be used?
Peripheral blood is usually not an alternative. No BM available:
attempt banding analysis from peripheral blood (CD34-FISH),
especially when blasts are present. Metaphase yield from peripheral
blood is generally worse than that from BM aspirates. CD34-FISH can
be an option. 1 What sample-related factors could influence the
accuracy of the test? Ex-vivo time over 24 hours: clotting;
bacterial/fungal contamination; and low cellular content (e.g.
hypocellular BM, or if several syringes are filled during biopsy
and the last syringe is used for banding analysis). Need for
dividing cells; cell clones < 10% of abnormal cells can be
overlooked; submicroscopic abnormalities cannot be detected. What
are the limitations of this method? FISH = fluorescent in situ
hybridization. 1 Braulke F, et al. Leuk Res. 2010;34:1296-301.
Questions and answers were prepared under the review of Dr. Haase,
University of Gttingen, Germany. FAQs: Karyotyping
Slide 141
141 G-EXJ-1030713 May 2012 Chromosome analysis in MDS:
Fluorescence in situ hybridization (FISH) Fluorescently labeled DNA
probes recognize complementary sequences on the chromosomes probes
that recognize centromeres detect changes in the number of
chromosomes probes that recognize specific genes can detect changes
in the chromosome sequence Fluorescently labeled probes for bcr and
abl genes show exchange of DNA between Chromosomes in chronic
myelogenous leukemia Images from:
http://www.wikidoc.org/index.php/Chronic_myelogenous_leukemia.
Accessed Feb.2011.
Slide 142
142 G-EXJ-1030713 May 2012 Preparation of the FISH sample
Metaphase slides for karyotyping could also use bone marrow smears
and paraffin-embedded bone marrow biopsies) Denature DNA (heat up
the sample) Hybridize with denatured (pre-heated) labeled probes
Wash to remove excess probe DAPI stain for DNA View with
fluorescence microscope and take photographs Image from:
http://www.creative-biolabs.com/fish/tissuearray5.htm. Accessed
Feb. 2011 Protocol from:
labs.mmg.pitt.edu/gjoerup/FISH%20protocol%20Vysis.doc. Accessed
Feb. 2011. Preparation of the FISH could be done by trained medical
technician or clinical geneticist. The analysis of the samples is
carried by clinical geneticists and/or hematologists.
Slide 143
143 G-EXJ-1030713 May 2012 FISH: Selection of probes MDS FISH
panel for initial diagnosis detects the most common aberrations,
and typically includes probes for 5q, 7q, #8, 11q, 12p, 13q, 17p,
and 20q Probes*ManufacturerWeb site Multiprobe MDS/AML
panelCytocellwww.cytocell.com 5/5q, 7/7q, 8cen, 20q Genzyme
Genetics www.genzymegenetics.com 5p/q, 7q, 17p13,
20q13Kreatechwww.kreatech.com 5p/q, 7cen/q, 8cen, 17p13, 20q13,
Ycen Abbott Molecular www.abbottmolecular.com 5/5q, 7/7q,
20qMetasystems www.metasystems- international.com Selection of the
FISH probes is essential part of the analysis and is typically done
by clinical geneticists in cooperation with hematologists *Examples
are not representative of the complete spectrum of probes available
from each provider. 1 Cherry AM, et al. Blood. 2010;116:[abstract
2922].
Slide 144
144 G-EXJ-1030713 May 2012 How is a probe selected? Use the
standard MDS FISH panel for initial diagnosis. When the aberration
is known/suspected, use additional probes in the region. If disease
morphology is suggestive of a certain genotype, one could directly
use the respective probe e.g. if RA with typical dysplasia of
megakaryocytes occurs, then use probes for del(5q). Note, however,
that one can miss other genetic changes with this approach. How
many probes are typically used in a panel? A standard panel
consists of 78 probes. 1 An extended panel can include up to 12
probes. FAQs: FISH 1 Cherry AM, et al. Blood. 2010;116:[abstract
2922]. Questions and answers were prepared under the review of Dr.
Haase, University of Gttingen, Germany.
Slide 145
145 G-EXJ-1030713 May 2012 The available survival data are
based on karyotyping (banding) studies. However, in addition to
karyotyping, one should use a standard FISH panel: 5p/q, 7cen/q,
#8cen, 17p13, 20q13, and Ycen (in males). What probes should be
used for survival prognosis? What are the limitations of FISH? One
sees/finds what one is looking for; therefore, one could overlook
complex or rare abnormalities (e.g. finding an isolated del(5q) by
FISH might give a false good prognosis if it is a part of a complex
genotype that has been misidentified). FAQs: FISH (cont.) Questions
and answers were prepared under the review of Dr. Haase, University
of Gttingen, Germany.
Slide 146
G-EXJ-1030713 May 2012 Summary
Slide 147
147 G-EXJ-1030713 May 2012 Summary WHO 2008 guidelines
recommend complete cytogenetic analysis of BM at initial diagnosis
of MDS Chromosomal aberrations, among other factors, have
prognostic relevance on overall survival and time to leukaemic
transformation IPSS was first to define cytogenetic risk groups and
to show an association with the survival prognosis of the patient
improved understanding of the cytogenetic risk factors provides a
better prognosis scoring for the patients with MDS Approximately
50% of MDS patients have abnormal cytogenetics in addition, it is
supposed that many patients with normal cytogenetics actually have
clonal abnormalities that remain undetected by metaphase
cytogenetics
Slide 148
148 G-EXJ-1030713 May 2012 Summary (cont.) Cytogenetic analysis
is essential for making therapeutic decisions with regard to
patients with MDS as it has shown associations with the response to
hypomethylating and immunomodulating agents Sequential cytogenetic
analysis is recommended to improve clinical management in MDS
Slide 149
G-EXJ-1030713 May 2012 TRANSCRANIAL DOPPLER ULTRASONOGRAPHY
(TCD) FOR ASSESSMENT OF STROKE RISK IN SICKLE CELL DISEASE NOTE:
These slides are for use in educational oral presentations only. If
any published figures/tables from these slides are to be used for
another purpose (e.g. in printed materials), it is the individuals
responsibility to apply for the relevant permission. Specific local
use requires local approval.
Slide 150
150 G-EXJ-1030713 May 2012 Outline Sickle cell disease (SCD)
Transcranial Doppler (TCD) TCD in SCD TCD Equipment Guidelines for
TCD in SCD Summary LIC = liver iron concentration; MRI = magnetic
resonance imaging; SF = serum ferritin; SIR = signal intensity
ratio; SQUID = superconducting quantum interface device.
Slide 151
G-EXJ-1030713 May 2012 Sickle Cell Disease (SCD)
Slide 152
152 G-EXJ-1030713 May 2012 What is SCD? An inherited disorder
affecting haemoglobin (Hb) synthesis Sickle cell erythrocytes have
a mutant form of Hb and HbS resulting from GluVal mutation in 6th
codon of -globin chain HbS turns normally pliable erythrocytes into
rigid, sickle-shaped cells The irregular erythrocyte morphology
leads to episodes of vascular occlusion and acute pain progressive
organ damage Children have increased risk of infection and stroke
Life expectancy may be shortened HbS = sickle cell haemoglobin.
Schnog JB, et al. Neth J Med. 2004;62:364-74. Image from
www2.med.umich.edu/prmc/media/newsroom/downloadImages.cfm?ID=656.
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153 G-EXJ-1030713 May 2012 Clinical manifestations of SCD
AnaemiaRed cell survival ~ 17 days (120 days in healthy people) 1
PainAcute and chronic 1 Central nervous systemOvert stroke, silent
stroke, and neurocognitive impairment 13 Pulmonary Recurrent acute
chest syndrome, pulmonary hypertension, and chronic sickle lung
disease 1,2 SkinChronic ulcers, typically around the ankles 1
Joints Osteonecrosis (avascular necrosis) of femoral and humeral
heads 1,2 EyesRetinal ischaemia, detachments sickle retinopathy 1,2
Kidneys Inability to concentrate urine; proteinuria progressing to
nephrotic syndrome; end-stage renal failure 4 CardiovascularCardiac
decompensation and cardiomyopathy 1 1 Schnog JB, et al. Neth J Med.
2004;62:364-74. 2 Claster S, Vichinsky EP. Br Med J.
2003;327:1151-5. 3 Prengler M, et al. Ann Neurol. 2002;51:543-52. 4
Ataga KI, Orringer EP. Am J Hematol. 2000;63:205-11.
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154 G-EXJ-1030713 May 2012 Prevalence of SCD The frequency of
the HbS gene is highest in populations in which malaria is (or was)
endemic 1,2 Approximately 200,000 new cases of SCD occur in Africa
every year 1 Recent population migrations have led to an increase
in disease frequency in other areas 1 Weatherall DJ, Clegg JB. Bull
World Health Organ. 2001;79:704-12. 2 Modell B, Darlison M. Bull
World Health Organ. 2008;86:480-7. 3 Sickle cell disease:
screening, diagnosis, management, and counseling in newborns and
infants clinical practice guideline number 6 AHCPR 1993;Publication
93-0562. 4 Sickle Cell Society. Sickle Cell Society Publication SC4
2005: www.sicklecellsociety.org/pdf/SC4.pdf. Image from
Christianson A, et al. March of Dimes Global Report on Birth
Defects: the hidden toll of dying and disabled children. 2006
(www.marchofdimes.com/MOD-Report- PF.pdf). 1 out of 2,400 live
births (all ethnic groups) are affected in England, where 12,500
individuals live with the disease 4 Births with a pathological Hb
disorder per 1,000 live births
165 G-EXJ-1030713 May 2012 Correlation between TAMMV on TCD and
stenoses on MRA in SCD Study rationale relationship between
neuroimaging abnormalities and TCD is unclear in adult patients
with SCD imaging abnormalities reported in up to 44% of children
with SCD; prevalence in adults unknown Differences: adults vs
children frequency of imaging abnormalities in a