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FISH IN MYELOMA
1. BTG 2013
FISH in Myeloma
Dr Edmond S K Ma
Department of Pathology
Hong Kong Sanatorium & Hospital
2. BTG 2013
Agenda
• Review HKS&H experience
• CG abnormalities: East meets West
• Diagnostic applications
3. BTG 2013
CG abnormalities in myeloma
• Conventional cytogenetics 30%
• Interphase FISH ~90%
– Drach J et al Cancer Res 1995; 55: 3854 – 9
• Reasons:
– Low plasma cell % in BM
– Low proliferative index of plasma cells
– Translocations may be morphologically cryptic
e.g. t(4;14)(p16;q32)
5. Prognostic significance
Black: No 13q-, t(4;14)
and 17p- and low B2M
Blue: Positive for
13q-, No t(4;14) and
17p- and low B2M
Green: No 13q-, t(4;14)
and 17p- and high B2M
Red: Positive for 13q-,
No t(4;14) and 17p-
and high B2M
Gray: Either t(4;14) of
17p-, and low B2M
Pink: Either t(4;14) of
17p-, and high B2M
Avet-Loiseau et al, Blood 2007; 109: 3489 - 95
6. BTG 2013
International Myeloma Workshop Consensus
• Risk stratification (panel 2)
Blood 2011; 117: 4696 – 4700
– Cytogenetically detected chromosomal 13 or 13q
deletion, t(4;14) and del(17p), and detection by
FISH of t(4;14), t(14;16) and del(17p)
– Repeat at relapse
• Standard investigative workup (panel 3)
Blood 2011; 117: 4701 – 5
– Standard metaphase cytogenetics and FISH for
t(4;14), t(14;16) and del(17p)
7. BTG 2013
Agenda
• Review HKS&H experience
• CG abnormalities: East meets West
• Diagnostic applications
8. BTG 2013
Myeloma FISH
• Aim at same day processing
• Examination of bone marrow smears
• Reject if < 5% plasma cells
• Cell sorting by CD138 immunomagnetic
beads (Miltenyi Biotec)
– For samples with < 50% plasma cells
– Aim at > 90% purity
• Pre-analytical variables: sample
adequacy, freshness, plasma cell %
12. BTG 2013
Gain of 1q: new high-risk CG marker in myeloma
From Cytocell website Red (1q) = CKS1B
Green (1p) = CDKN2C (p18)
Avet-Loiseau H et al, J Clin Oncol 30: 1949 – 52, 2012
13. BTG 2013
FISH analysis
• Analyze at least 100 cells
• Two observers
• Laboratory threshold
– Mean + 3SD of normal BM controls
• Positive cutoff levels
Ross FM et al 2012; Avet-Loiseau H et al 2012
– 10% for fusion or break-apart probes
– 20% for numerical abnormalities
– 30% for chromosome 1q gain
– 40% for del(17p)/p53
14. BTG 2013
Summary of HKS&H data
• n = 105 cases
– From April 2011 – Dec 2012
• Male: female = 73 : 32
• Median age = 62 years (range: 42 – 88)
• Median % plasma cells = 53%
– Range 5 – 95%
• Cell sorting performed in 44/105 (42%)
– i.e. < 50% plasma cells
15. Summary of HKS&H data
CG abnormality Frequency (%) Number Range of +ve cells
13q- 24% 5/21 60 – 90%
t(4;14) 16% 17/105 10 – 90%
t(14;16) 3% 2/80 50 – 80%
17p- 6% 6/105 20 – 80%
1q gain 42% 5/12 40 – 90%
Polysomy 5/9/15 36% 31/87 20 – 95%
16. BTG 2013
Summary of HKS&H data
• Secondary FISH abnormalities
– 6/17 t(4;14)+ve showed atypical (1Y1O1G)
signal pattern
– Polysomy / amplification 4, 14 or 16
• Common
– Monosomy / deletion 17, 4, 14 or 16
• Occasional
17. BTG 2013
Agenda
• Review HKS&H experience
• CG abnormalities: East meets West
• Diagnostic applications
18. Frequency of FISH abnormalities
across studies
Reference t(4;14)
t(14;16
)
t(11;14
)
del(17p) del(13q)
Hyper-
diploidy
MYC 1q gain
Fonseca 2003
(ECOG)
12.7% 4.6% 15.8% 10.7% 54.2% NA NA NA
Avet-Loiseau
2007 (IFM)
14% NA 21% 11% 48% 39% 13% NA
Huang S-Y 2005
(Taiwan)
NA NA NA 6.8% 34.1% ~20% NA NA
Tan 2010
(Singapore)
14.9% 5.4% 18.9% 20.3% 29.7% NA NA NA
Lai Y-Y 2012
(China)
IgH translocation 57.6% 34.7% 60.4% NA NA 49%
Yu H 2011
(China)
NA NA NA 22.2% ~50% NA NA 45.8%
HKS&H
experience
16% 3% NA 6% 24% 36% NA 42%
19. Frequency of FISH abnormalities
across studies
Reference t(4;14)
t(14;16
)
t(11;14
)
del(17p) del(13q)
Hyper-
diploidy
MYC 1q gain
Fonseca 2003
(ECOG)
12.7% 4.6% 15.8% 10.7% 54.2% NA NA NA
Avet-Loiseau
2007 (IFM)
14% NA 21% 11% 48% 39% 13% NA
Huang S-Y 2005
(Taiwan)
NA NA NA 6.8% 34.1% ~20% NA NA
Tan 2010
(Singapore)
14.9% 5.4% 18.9% 20.3% 29.7% NA NA NA
Lai Y-Y 2012
(China)
IgH translocation 57.6% 34.7% 60.4% NA NA 49%
Yu H 2011
(China)
NA NA NA 22.2% ~50% NA NA 45.8%
HKS&H
experience
16% 3% NA 6% 24% 36% NA 42%
20. Frequency of FISH abnormalities
across studies
Reference t(4;14)
t(14;16
)
t(11;14
)
del(17p) del(13q)
Hyper-
diploidy
MYC 1q gain
Fonseca 2003
(ECOG)
12.7% 4.6% 15.8% 10.7% 54.2% NA NA NA
Avet-Loiseau
2007 (IFM)
14% NA 21% 11% 48% 39% 13% NA
Huang S-Y 2005
(Taiwan)
NA NA NA 6.8% 34.1% ~20% NA NA
Tan 2010
(Singapore)
14.9% 5.4% 18.9% 20.3% 29.7% NA NA NA
Lai Y-Y 2012
(China)
IgH translocation 57.6% 34.7% 60.4% NA NA 49%
Yu H 2011
(China)
NA NA NA 22.2% ~50% NA NA 45.8%
HKS&H
experience
16% 3% NA 6% 24% 36% NA 42%
21. BTG 2013
Agenda
• Review HKS&H experience
• CG abnormalities: East meets West
• Diagnostic applications
22. IgM paraprotein: WM or myeloma?
• F/57 Caucasian
• Bone lesion
• Hb 11.3 g/dL
• WBC 6.7 x 109
/L
• Platelets 285 x 109
/L
• RFT normal
• Paraprotein
– IgM kappa at 10.4 g/L