susanna akiki wmrgl 2009 acute promyelocytic leukemia with t(15;17) (q22;q21) developing...
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Susanna Akiki WMRGL 2009
Acute Promyelocytic Leukemia with t(15;17) (q22;q21)
developing inv(16)(p13q22) secondary AML
Susanna Akiki WMRGL 2009
• Leukemia clonal neoplasia, • consequence of an accumulation of genetic damage
•Specific leukemia’s associated with very specific genetic events
Mutation arises in a single cell
Emerges as a substantial population of cells
Susanna Akiki WMRGL 2009
Acute Myeloid Leukemia• Consequence of acquired somatic mutations in haematopietic
progenitors- myeloid lineage
• Typically involve translocations
• Target is often a transcription factor involved in haemopoiesis
• Create a novel gene fusion
• Implicated in leukemiagenesis
• Gillilands 2 hit hypothesis of co operating mutations in the pathogenesis of AML two distinct classes of mutation required to induce leukaemia
• Mutations that block cellular differentiation• Mutations that increase cell proliferation & survival
• Genetic event occur well characterised & identify specific subsets with prognostic significance
Susanna Akiki WMRGL 2009
Acute Promyelocytic Leukemia
• APL subtype of AML characterised by
• Specific recurrent translocation t(15;17)
• Chimeric PML-RARA fusion encodes a novel protein
• Acts as a transcriptional repressor
• APL particularly sensitive to treatment
• Maturation block overcome by Retinoic acid
• Highly effective in combination with chemo
• Prolonged remission achieved in ~ 80% patients
Susanna Akiki WMRGL 2009
PML-RARA Monitoring
Level of PML/RARA normalised to ABL
0.000001
0.00001
0.0001
0.001
0.01
0.1
1
01/0
1/200
6
01/0
4/200
6
01/0
7/200
6
01/1
0/200
6
01/0
1/200
7
01/0
4/200
7
01/0
7/200
7
01/1
0/200
7
01/0
1/200
8
Ratioonlog
scale
PML/RARA:ABL
Sensitivity of detection
Level of PML/RARA normalised to ABL
0.000001
0.00001
0.0001
0.001
0.01
0.1
1
01/0
8/200
3
01/1
1/200
3
01/0
2/200
4
01/0
5/200
4
01/0
8/200
4
01/1
1/200
4
01/0
2/200
5
01/0
5/200
5
01/0
8/200
5
01/1
1/200
5
01/0
2/200
6
01/0
5/200
6
01/0
8/200
6
01/1
1/200
6
01/0
2/200
7
01/0
5/200
7
01/0
8/200
7
01/1
1/200
7
Ratioonlog
scale
PML/RARA:ABL
Sensitivity of detection
t(15;17) predicts a favourable response
Susanna Akiki WMRGL 2009
PML-RARA Monitoring
Re emergence is predictive of relapse
Level of PML/RARA normalised to ABL
0.000001
0.00001
0.0001
0.001
0.01
0.1
1
10
01/0
3/200
5
01/0
6/200
5
01/0
9/200
5
01/1
2/200
5
01/0
3/200
6
01/0
6/200
6
01/0
9/200
6
01/1
2/200
6
01/0
3/200
7
Ratioonlog
scale
PML/RARA:ABL
Sensitivity of detection
Level of PML/RARA normalised to ABL
0.000001
0.00001
0.0001
0.001
0.01
0.1
1Ratioonlog
scale
PML/RARA:ABL
Sensitivity of detection
Early detection allows early
effective intervention
Failure to achieve remission
Susanna Akiki WMRGL 2009
• 50 year old female presented with APML in Nov 2003
• 47,XX, +8, t(15;17)(q22;q21)
• RT-PCR confirmed a PML/RARA (bcr3) gene fusion
• consistent with the diagnosis of APML, M3.
Case history
Susanna Akiki WMRGL 2009
APML Diagnosed Nov2003
Good induction response
(4.7 log reduction, lab mean= 2.54)
? Relapse 31 months post consolidation
Level of PML/RARA normalised to ABL
0.0000010.000010.00010.0010.010.1
110Ratio
onlog
scale
PML/RARA:ABL
Sensitivity of detectionLevel of PML/RARA normalised to ABL
0.0000010.000010.00010.0010.010.1
110
10/1
1/200
3
10/0
2/200
4
10/0
5/200
4
10/0
8/200
4
10/1
1/200
4
10/0
2/200
5
10/0
5/200
5
10/0
8/200
5
10/1
1/200
5
10/0
2/200
6
10/0
5/200
6
10/0
8/200
6
10/1
1/200
6
10/0
2/200
7
10/0
5/200
7
10/0
8/200
7
10/1
1/200
7
Ratioonlog
scale
PML/RARA:ABL
Sensitivity of detection
Remained PML-RARA negative
Susanna Akiki WMRGL 2009
Diagnosis of secondary AML rather than relapsed APML considered
• 46,XX,inv(16)(p13q22)[15]
• CBFB/MYH11 type D fusion
900bp
203bp
Susanna Akiki WMRGL 2009
Kinetics of emergence
• cDNA banked as a consequence of APL monitoring
• Retrospective analysis for CBFB/MYH11
• First appearance of inv(16) clone
• Study kinetics of disease emergence
Susanna Akiki WMRGL 2009
Inv(16) retrospective analysis
? relapse
Level of CBFB/MYH11 normalised to ABL
0.000001
0.00001
0.0001
0.001
0.01
0.1
1
10
10/1
1/200
3
10/0
2/200
4
10/0
5/200
4
10/0
8/200
4
10/1
1/200
4
10/0
2/200
5
10/0
5/200
5
10/0
8/200
5
10/1
1/200
5
10/0
2/200
6
10/0
5/200
6
10/0
8/200
6
10/1
1/200
6
10/0
2/200
7
10/0
5/200
7
10/0
8/200
7
10/1
1/200
7
Ratioonlog
scale
CBFB/MYH11:ABL
Sensitivity of detection
15 month plateau phase Exponential phase
APL diagnosis Inv (16)
9 months 27 months
Susanna Akiki WMRGL 2009
Gilliland’s 2 hit hypothesisGilliland & Griffin Blood 2002
these mutations ‘cooperate’: the ‘2-hit’ model of leukaemogenesis
Susanna Akiki WMRGL 2009
Model 1: Inv (16) pre existing clone ?Level of CBFB/MYH11 normalised to ABL
1E-12
1E-11
1E-10
1E-09
1E-08
1E-07
0.000001
0.00001
0.0001
0.001
0.01
0.1
1
10
10/1
1/200
3
10/0
2/200
4
10/0
5/200
4
10/0
8/200
4
10/1
1/200
4
10/0
2/200
5
10/0
5/200
5
10/0
8/200
5
10/1
1/200
5
10/0
2/200
6
10/0
5/200
6
10/0
8/200
6
10/1
1/200
6
10/0
2/200
7
10/0
5/200
7
10/0
8/200
7
10/1
1/200
7
Ratioonlog
scale
CBFB/MYH11:ABL
Sensitivity of detection
treatment
Susanna Akiki WMRGL 2009
Model 2: True secondary leukaemia?Level of CBFB/MYH11 normalised to ABL
1E-12
1E-11
1E-10
1E-09
1E-08
1E-07
0.000001
0.00001
0.0001
0.001
0.01
0.1
1
10
10/1
1/200
3
10/0
2/200
4
10/0
5/200
4
10/0
8/200
4
10/1
1/200
4
10/0
2/200
5
10/0
5/200
5
10/0
8/200
5
10/1
1/200
5
10/0
2/200
6
10/0
5/200
6
10/0
8/200
6
10/1
1/200
6
10/0
2/200
7
10/0
5/200
7
10/0
8/200
7
10/1
1/200
7
Ratioonlog
scale
CBFB/MYH11:ABL
Sensitivity of detection
treatment
Susanna Akiki WMRGL 2009
FLAG
Level of CBFB/MYH11 normalised to ABL
0.000001
0.00001
0.0001
0.001
0.01
0.1
1
10
10/11
/200
3
10/02
/200
4
10/05
/200
4
10/08
/200
4
10/11
/200
4
10/02
/200
5
10/05
/200
5
10/08
/200
5
10/11
/200
5
10/02
/200
6
10/05
/200
6
10/08
/200
6
10/11
/200
6
10/02
/200
7
10/05
/200
7
10/08
/200
7
10/11
/200
7
10/02
/200
8
10/05
/200
8
10/08
/200
8
10/11
/200
8
10/02
/200
9
Ratioonlog
scale
CBFB/MYH11:ABL
Sensitivity of detection
Patient Outcome
? APL Relapse – As0
2ndry AML diagnosed
100% mets <5% IF 0% IF
Flag-Ida
Susanna Akiki WMRGL 2009
Summary• Novel opportunity to study the biology of AML
• It is possible the inv(16) clone was a pre existing clone, present at the time of diagnosis with APL with a second mutation arising following APL therapy
• Alternatively this case represents a true secondary AML arising as a consequence of APL therapy with both the CBFB-MYH11 and presumed tyrosine kinase hit induced during different treatment cycles
• Retrospective analysis suggests biphasic kinetics of inv(16) clone supporting Gilliland's 2 hit hypothesis of the pathogenesis of acute leukaemia
Susanna Akiki WMRGL 2009
Acknowledgments
Mike GriffithsVal Davison
Fiona MacDonald
Molecular Oncology TeamJoanne Mason
Jane BryonMax Rindl
Rachel DoakSarah Whelton
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