Genomic

Heterogeneity of

Multiple Myeloma and

Implications

for Targeted Therapy

and Treatment

Resistance

Niels Weinhold, PhD

September 13, 2016

2

Introduction to myeloma genetics

Double-hit myeloma

Private clones

Overview

3

Introduction to myeloma genetics

Double-hit myeloma

Private clones

Overview

4

Primary (initiating) & secondary events

+3

+5

+7

+9

+11

+15

+19

+21

Manier et al., Nat. Rev. Clin. Oncol., 2016

5

Primary (initiating) & secondary events

+3

+5

+7

+9

+11

+15

+19

+21

t(MYC)

del(17p)/TP53

del1p32, del1p12 &

gain/amp 1q21

Manier et al., Nat. Rev. Clin. Oncol., 2016

6

GEP70: 70 genes associated with outcome

7

Primary (initiating) & secondary events

+3

+5

+7

+9

+11

+15

+19

+21

del(17p)/TP53

Presence

vs

Type

Manier et al., Nat. Rev. Clin. Oncol. 2016

8

Introduction to myeloma genetics

Double-hit myeloma

Private clones

Overview

9

Longitudinal exome sequencing study: overview

Weinhold et al., Blood 2016

17p12 (TP53)

???

10

Frequence and prognostic value of TP53 aberrations

Presentation

del(17p) in 5 patients

TP53mut in 4 patients

Relapse

del(17p) in 10 patients

TP53mut in 10 patients

11 patients with one

affected allele

5 patients with double

event

Weinhold et al., Blood 2016

11

Acquired double-hits at relapse

Presentation: 18 bi-allelic events in 12 patients

Relapse: 26 bi-allelic events in 17 patients

FAM46C bi-allelic events in 6 patients

Risk at presentation: 4/19 vs. 7/10, P=0.02

Risk at relapse: 3/15 vs. 12/14, P=0.001

Weinhold et al., Blood 2016

12

Variant allele frequency: presentation

Va

ria

nt

all

ele

fre

qu

en

cy:

rela

pse

TP53: Competing subclones

NRAS

DIS3

del(17p)

KRAS

DIS3

TP53 Subclonal vs. double hit

Frequency information

for both aberrations

important!

Clonal competition

Pre-existing?

Shared

mutations

Clone with del(17p)

Weinhold et al., Blood 2016

13

Introduction to myeloma genetics

Double-hit myeloma

Private clones

Overview

14

“Focal lesion” project

Aim: determine the extent of heterogeneity in space

42 newly diagnosed myeloma patients

Whole exome sequencing & high resolution arrays

Paired samples: aspirates from iliac crest & focal lesion(s)

Focal lesions

Accumulations of malignant plasma cells in

restricted areas within the bone marrow

74% of patients with >=1 according to MRI*

Mean number: 10*

Sample 1

“Random” aspirate

from iliac crest

Paired sample(s)

Focal lesion(s)

*Walker et al. JCO 2007

15

Heterogeneity: “primary” event hyperdiploidy

shared unshared Not present

16

Heterogeneity: ploidy

Focal lesion (L4)

Iliac crest

Trisomies: hyperdiploid karyotype

Initiating event???

17

Heterogeneity: “primary” t(IgH) and progression events

shared unshared Not present

18

Focal lesion (L4)

del(1p) del(17p)

Heterogeneity in space: high risk & bi-allelic event

TP53 Pre-existing ultra-high-risk event!!!

19

Mutations: types of heterogeneity

Shared

minor>major

Unshared

Variant allele frequency: random aspirate (iliac crest)

Va

ria

nt

all

ele

fre

qu

en

cy:

foca

l le

sio

n

Shared

20

Total number of mutations and heterogeneity

Median total:64

Min->major clone

Unshared >=20% VAF

Unshared <20% VAF 0% (0-26%)

11% (0-62%)

5% (0-42%)

21

Actionable and driver mutations: distribution

“Driver” genes (Other) actionable mutations*

*Walker et al. JCO 2015

Nr of patients

with mutation

Blue = unshared

Mutations in “driver” genes are not

necessarily shared

22

BRAF V600E as target: example

BRAF mutat ion in 1/ 3 of reads

Trisomy 7

Majority of cells with mutat ion

“Good target !”

Expectation*

*Andrulis et al. Cancer Discovery 2013

BRAF

???

23

Same patient: clonality analysis based on seq data

Clone in

focal

lesion

Clone at

iliac crest

Variant allele frequency: random aspirate (iliac crest)

Va

ria

nt

all

ele

fre

qu

en

cy:

foca

l le

sio

n

BRAF

High

Risk

Low risk

24

KRAS

FGF12

BRAF STAT3

Site-specific dominance

Allopatric speciation?

Impact on standard risk stratification?

25

GEP70: random aspirate vs. focal lesion

42

(16%)

GEP70 focal lesion

221

(84%)

Regionally restricted dominance

of high risk clones!

26

Outcome according to GEP70 of paired samples

27

Spatial analyses: conclusions

Heterogeneity in space in majority of cases on the nucleotide level

Full spectrum of mutations not detectable with single sample

“Driver” and other actionable mutations are frequently non-

ubiquitous

High risk sub-clones frequently unequally distributed

Targeted treatment may even support high risk clones

Possible explanation for low sensitivity of high risk markers

“Simple” Darwinian model (clonal sweeps) does not explain

patterns in myeloma

28

Acknowledgements

Thank you for your

attention!