FISH IN MYELOMA

BTG 2013

FISH in Myeloma

Dr Edmond S K Ma

Department of Pathology

Hong Kong Sanatorium & Hospital

BTG 2013

Agenda

• Review HKS&H experience

• CG abnormalities: East meets West

• Diagnostic applications

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)

BTG 2013

Prognostic significance

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

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)

BTG 2013

Agenda




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 %

BTG 2013

CD138 immunomagnetic bead sorting

Before After

BTG 2013

FISH probes

• 13q, t(4;14), 17p, CEP 5/9/15

• t(4;14), t(14;16), 17p, CEP 5/9/15

• Current panel– t(4;14)(p16;q32) (IGH/FGFR3 DF probe)– t(14;16)(q32;q23) (IGH/MAF DF probe)– TP53/CEP17 – CKS1B/CDKN2C(p18) amplification/deletion

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

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

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

Summary of HKS&H dataCG 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%

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

BTG 2013

Agenda




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%

BTG 2013

Agenda




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

BTG 2013


CD20 CD138

Kappa PAX-5

BTG 2013


• t(11;14) positive• MYD88 L256P negative

• Conclusion

CD20+ IgM myeloma

Cyclin D1

BTG 2013

IgG paraprotein: myeloma or WM?

• F/68 Chinese• Nephrotic syndrome,

renal impairment• IgG kappa

paraprotein – increasing trend

• First BM in 2010: not myeloma

Peripheral blood

BTG 2013

Plasma cell myeloma with t(11;14)(q13;q32)

• Around 20% of myeloma• Cyclin D1 over-expression• Lymphoplasmacytic morphology (40 – 50%)• CD138+ 19+ 20+ PAX5+ cyclin D1+• Mostly IgG, IgA and light chain myeloma• 80% IgM-myeloma associated with t(11;14)

– but CD20-ve in contrast to t(11;14) non-IgM PCM

BTG 2013


BM

BM

Trephine x 100

Trephine x 600

BTG 2013


CD20 CD138

Kappa ISH

Lambda ISH

BTG 2013


• t(11;14) -ve• MYD88 L256P +ve

• Conclusion: IgG WM

T C

BTG 2013

Disease progression

• M/72 Chinese• Light chain myeloma• Achieved CR• Sudden relapse as

plasma cell leukaemia 10 months later

• Acquired del(17p) that was negative at diagnosis

Diagnosis

Relapse

BTG 2013

Acknowledgements

• Prof. C S Chim

• Mr. Anthony Wong• Ms. Candy Wang

• Referring clinicians

FISH IN MYELOMA

Education

Transcript of FISH IN MYELOMA