Molecular monitoring in AML: Impact on Therapy
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Transcript of Molecular monitoring in AML: Impact on Therapy
Molecular monitoring in AML: Impact on Therapy
Hwei-Fang Tien, National Taiwan University Hospital
West part of NTUH
• Molecular markers for monitoring
• Prognostic implications
• Pre-emptive therapy on molecular relapse
• Samples and timing for monitoring
Outline
• Molecular markers for monitoring
• Prognostic implications
• Pre-emptive therapy on molecular relapse
• Samples and timing for monitoring
Outline
Molecular markers for monitoring in AML
Abnormal fusion genes: ~ 30%
t(8;21)/RUNX1-RUNX1T1
inv(16)/CBFβ-MYH11
t(15;17)/PML-RARα
t(7;11)/NUP98-HOXA9
t(11;v)/MLL-partner gene
Molecular mutations that are stable during disease progression
NPM1, IDH1/2, DNMT3A ,CEBPA, MLL-PTD
Gene overexpression
WT1
Abnormal fusion genes: ~ 30%
t(8;21)/RUNX1-RUNX1T1
inv(16)/CBFβ-MYH11
t(15;17)/PML-RARα
t(7;11)/NUP98-HOXA9
t(11;v)/MLL-partner gene
Molecular mutations that are stable during disease progression
NPM1, IDH1/2, DNMT3A ,CEBPA, MLL-PTD
Gene overexpression
WT1
Molecular markers for monitoring in AML
NPM1 mutant as a molecular marker for MRD monitoring
0
1
2
3
4
5
6
7
0 10 20 30 40 50 60 70 80 90 100
Log
Mut
ants
Blast % in BM
NTUH, Leukemia, 2007, 21998
… ...
TCTG
CCTG
CATG
TCGG
CCAG
CCGG
… ...
951 960 964
TGGAGGA
TGGAGGA
TGGAGGA
TGGAGGA
TGGAGGA
TGGAGGA
TGGAGGA
TGGAGGACGGC
…….
……
……
……
……
……
……
AGATCTCTG
AGATCTCTG
AGATCTCTG
AGATCTCTG
AGATCTCTG
AGATCTCTG
AGATCTCTG
AGATCTCTG
GCAG
GCAG
GCAG
GCAG
GCAG
GCAG
GCAG
GCAG
Wild type
Type VII
Type VI
Type V
Type IV
Type III
Type II
Type I
No. patients
26 5
1
2
1
1
2
A NPM1 mutations: 4-nucleotide insertions
The copy number of mutants correlate well with BM blast %, but frequently in BM samples with blasts <5%, NPM1 mutant levels can be high.* sensitivity: 1/105
B Quantitative real-time PCR
0
1
2
3
4
5
6
0 5 10 15 20 25 30
0
1
2
3
4
5
6
0 5 10 15 20 25 30 35
Patient 3 Patient 31
Mon. Mon.
CR CR
CR
CR
Relapse Relapse
Relapse
CR
CR
CR CR CR CR CR
NPM1 mutant monitoring during the clinical course
intensive chemotherapy
t(7;11)/NUP98-HOXA9 as a marker
More common in Asia and is associated with poor prognosis
#11
#7
fusion
NTUH, Leukemia, 2009, 23:1303
Blast percentage in bone marrow
Lo
g N
UP
98
-HO
XA
9/
10
6 H
UP
O
0
1
2
3
4
5
6
0 10 20 30 40 50 60
Correlation between marrow blast percentage and mutant load of NUP98-HOXA9
The mutant signals always remain high even when BM blasts <5% and are rarely undetectable.
NTUH, Leukemia, 2009, 23:1303
Real-time PCR for NUP98–HOXA9
0
1
2
3
4
5
6
0 1 2 3 4 5 6 7 8
MUD HSCT (myeloablative)
relapse
0
1
2
3
4
5
6
0 2 4 6 8 10 12
Sibling HSCT (reduced intensity)
0
12
34
5
0 10 20 30 40 50
Sibling HSCT (myeloablative)
DLIDLI
Relapse
Lo
g N
UP
98-H
OX
A9/
106 H
UP
O
Lo
g N
UP
98-H
OX
A9/
106 H
UP
O
Lo
g N
UP
98-H
OX
A9/
106 H
UP
O
Months from diagnosis
Months from diagnosis
Months from diagnosis
A B
C D
Patient No. 7 Patient No. 11
Patient No. 6
0 10 20 30 40 50
0
5
4
3
1
CR
CR
CR
CR
CR
CR
CR
CRCR
CR
CR
CR
CR
CR
CRCR
0
1
2
3
4
5
6
0 5 10 15 20
Months from diagnosis
Lo
g N
UP
98-H
OX
A9/
106 H
UP
OAutoBMT
Patient No. 8
MUD HSCTmyeloablative
2
PR
PR
CR Relapse
CR CR
CR
CR
CR
CR
MRD monitoring of t(7;11)/NUP98-HOXA9
NTUH, Leukemia, 2009, 23:1303
In general, chemotherapy and even HSCT can only partially reduce the mutant signals which are always detectable even at CR. Most pts died of disease.
The leukemic cells are very resistant to the present treatment and novel therapy is needed for these patients.
WT1 overexpression as a marker to monitor MRD
European LeukemiaNet, JCO, 2009, 27:5195
Overexpression of WT1 can be found in >80% of AML patients
European LeukemiaNet (ELN) has standardized the assay method for WT1 expression
• Molecular markers for monitoring
• Prognostic implications
• Pre-emptive therapy on molecular relapse
• Samples and timing for monitoring
Outline
Prognostic implication of MRD in patients with NPM1 mutation
More aggressive treatment may be needed for this group of patients.
NPM1-mutated patients with less than 2 logs reduction of NPM1 mutant after consolidation C/T had shorter OS and RFS
OS RFS
Reduction <2 logs, n=6
Reduction <2 logs, n=6
P=0.010 P=0.001
Month Month
Others, n=16 Others, n=16
NTUH 2006
NTUH: Leukemia 21:998, 2007
After consolidation
P=0.002
Any > 1.5%, n=18
OS
Month
n=13
RFS
Any > 1.5%, n=18
Month
n=13
P<0.001
Prognostic Significance of MRD Levels of NPM1 mutants during Follow-ups
NTUH, : Leukemia 21:998, 2007
Early intervention at this moment may be needed for these patients.
During sequential follow-ups, patients who had >1.5% of NPM1 mutant in any sample had poorer prognosis
Gene mutations as biomarkers for MRD monitoring
CR patients
after doubleinduction C/T
after completion of therapy
JCO, 2011, 29:2709
NPM1 mutation: German-Austrian AML Study Group
MRD Monitoring in patients with CBF-AML
Jourdan et al, Blood , prepublished online January 15, 2013; DOI 10.1182
French AML Intergroup198 patients , aged 18 to 60 yearold and with newly diagnosed CBF-AML with t(8;21)/RUNX1-RUNX1T1 or inv(16)/CBFB-MYH11
A more than 3-log MRD reduction after the second consolidation C/T was associated with better prognosis
Relapse rate
reduction<3 log
reduction>=3 log
P<0.001
Overall survival
reduction>=3 log
reduction<3 log
P=0.066
Same for PFS, p<0.001
After 2nd consolidation C/T
Detection of MRD by ELN Standardized WT1 Assay
91 AML patients: with significant high WT1 expression (>2x 104 WT1 copies/104 ABL copies)
Cilloni et al, a European LeukemiaNet Study, JCO, 2009, 27:5195
After induction chemotherapy
• Molecular markers for monitoring
• Prognostic implications
• Pre-emptive therapy on molecular relapse
• Samples and timing for monitoring
Outline
MRC, UK 1. Detection of MRD at the end of consolidation in APL patients could predict RFS.
MRD Monitoring of PML/RARα Fusion Transcript by Real-Time PCR in APL patients
AML15: with pre-emptive therapy
Grimwade et al, JCO 2009 27:3650
AML12: without pre-emptive therapy
2. Pre-emptive therapy with As2O3 on molecular relapse reduced rate of clinical relapse.
DLI Improve the Outcome of Patients with MRD after allo-HSCT
MRD: WT1 expression for AML, IgH and TCR for ALL
Dominietto et al, Blood, 2007, 109:5063
• Molecular markers for monitoring
• Prognostic implications
• Pre-emptive therapy on molecular relapse
• Samples and timing for monitoring
Outline
Sampling interval (mo)
CBFB-MYH11 PB 6* BM AvoidRUNX1-RUNX1T1 PB 3 BM 4PML-RARA PB 1 BM 2NPM1c/FLT3-ITD-
PB 4 BM 6NPM1c/FLT3-ITD+
PB 3 BM 4WT1 expression PB 2 BM 4
Proposed guidelines for MRD
Modified from Hokland & Ommen, Blood 2011, 117:2577
*One additional MRD sampling recommended 3 months after end of C/T.# incidence in NTUH **In normal karyotype
During follow-ups: according to the relapse kinetics of different molecular alterations During treatment:
according to the prognostic significance of MRD at different time points
PML-RARA, CBFB-MYH11, RUNX1-RUNX1T1: after consolidation
NPM1 mutation: after double induction and consolidation
WT1 expression: after induction
Summary
• Molecular monitoring after treatment is helpful for
risk-stratification.
• The optimal timing for MRD assessment varies with
molecular groups.
• Pre-emptive therapy at the time of molecular relapse during follow-ups may improve the clinical outcome.
• Molecular monitoring by quantitative real-time PCR
is sensitive and specific for MRD detection in AML.
Future Prospective
Standardize the method and timing of molecular
monitoring.
Incorporate MRD detection into pretreatment risk factors in clinical trials to evaluate more clearly the prognostic significance of MRD and to design preemptive measurement on a molecular relapse.
Apply next generation deep sequencing of targeted genes for MRD detection.
台灣藍鵲( Formosan Blue Magpie )