MSI and other molecular markers: how useful are...

MSI and other molecular markers:how useful are they?Daniela E. Aust, Institute for Pathology, University Hospital Dresden,

Germany

www.pathologie-universitaetsmedizin-dresden.de

Institut für PathologieInstitut für Pathologie

Disclosure slide

I Member of advisory boards for AMGEN, ROCHE, BOEHRINGER

I Speaker honoraria from FALK Pharma, Pfizer, Lilly and ROCHE

I Third party funds from MERCK for immunohistochemistry in a clinical

trial



What can (molecular) pathology offer forclinical decisions in colorectal cancer?

Better understanding of the disease

Predictive markers

Prognostic markers



Weinberg R &

Hanahan D, 2011

Therapeutic Targeting of the„Hallmarks of Cancer“


Institut für Pathologie

GOAL:



Predictive markers

I MSI for 5-FU, irinotecan, immune checkpoint therapy ?

I RAS for anti-EGFR-therapy

I TS, TP, DPD for 5-FU-therapy

I ERCC1 for oxaliplatin



MSI and 5-FU

I Several studies showed the absence of benefit for adjuvant chemotherapy

in MSI-H patients

― Ribic et al. N Engl J Med 2003

• 570 cancers, 95 (16,7 %) with MSI-H.

• Interaction chemotherapy*MSI status p=0.009

― Jover et al. Gut 2006

• 505 patients stage II-III 125 stages II (42.2%) 135 stages III (64.5%)

with adjuvant chemotherapy

• Interaction chemotherapy*MSI status p=0.007



No chemotherapy 5FU chemotherapy

1027 patients included in trials

demonstrating the effect of FU in

adjuvant settings

MSI + (dMMR) 185 pts (18%)

Dan Sargent et al.

Dan Sargent et al, JCO 2010



Stage IIStage II Stage IIIStage III

MSI

MSS

Dan Sargent et al, JCO 2010



2141 patients, 344 MSI + (16%) ; positive effect limited to the group of lynch

syndrome

No chemotherapy

Lynch syndrome Sporadic MSI

Chemotherapy 5FU



ReferencesType of

study

Number of

patients

Number of MSI

tumours (%)

Tumour

stage

Number of patients

receiving adjuvant CTStratification

Survival

analysis

criteria

Survival results for MSI patients

Elsaleh et al,

2000 [19]R 656 56 (8.5) III 272 MMR 5 yr-OS Longer survival

Hemminki et al,

2000 [20]P / NR 95 11 (12) III 95 MMR 3 yr-RFS Longer survival

Ribic et al,

2003 [30]

R from

RCT570 95 (16.7) II + III 283 MMR and CT

5 yr-DFS

5 yr-OS No benefit or detrimentala

Carethers et al,

2004 [22]R 204 36 (17.6) II + III 66 CT OS No benefit

de Vos Tot Neder-

veen Cappel et al,

2004 [23]

R 92 92 (100)b III 28 CT 5 yr-OS No benefit

Benatti et al,

2005 [24]R 1263 256 (20.3) All stages 304 CT 5 yr-OS No benefit

Westra et al,

2005 [21]

R from

RCT273 44 (16) III 273 MMR 5 yr-DFS Longer survival

c

Jover et al,

2006 [25]P / NR 754 66 (8.8) All stages 260 CT OS No benefit

Lanza et al,

2006 [26]R 718 114 (15.9) II + III 193 MMR and CT 6 yr-OS No benefit

Kim et al,

2007 [27]

R from

RCT542 98 (18) II + III 369 MMR 5 yr-RFS/OS No significant difference

d

Lamberti et al,

2007 [28]P / NR 416 52 (13) All stages 89

e MMR OS No significant difference

457 70 (15) II + III 229 No benefit

1027f 165 (16) II + III 512 No benefit or detrimental

Hutchins et al,

2011 [29]

R from

RCT1913 218 (11.4) II (90%) 924 MMR and CT 2 yr-RFS No benefit

Sargent et al,

2010 [31]MMR and CT

5 yr-DFS

5 yr-OS

R from

RCT

5-fluorouracil-based adjuvant chemotherapy

To cut the long 5 FU-story short (from A. Zaanan)



KRAS und BRAF

beim Kolorektalkarzinom

The prognostic impact of MMR depended on tumor site . The interaction is highly

significant p=0.009. Validated on CALGB 88903

Right colon Left colon

NO-147



Defective mismatch repair as a prognostic marker in UICC stage III colon cancer with adjuvant FOLFOX

Zaanan et al., Clin Cancer Res. 2011



ReferencesType of

study

Number of

patients

Number of MSI

tumours (%)

Tumour

stage

Number of patients

receiving adjuvant CTStratification

Survival

analysis

criteria

Survival results for MSI patients

2011 [29] RCT

Kim et al,

2010 [38]R 135 12 (8.8) All stages FOLFOX, n=121 MMR

3 yr-DFS

3 yr-OS No significant difference

Des Guetz et al,

2010 [39]R 105 19 (18) II + III FOLFOX, n=105 MMR DFS Longer survival

Zaanan et al,

2010 [40]R 233 32 (14) III

5FU, n=124

FOLFOX, n=109CT 3 yr-DFS Longer survival with FOLFOX

Zaanan et al,

2011 [41]R 303 34 (11.2) III FOLFOX, n=303 MMR 3 yr-DFS Longer survival

Bertagnolli et al,

2009 [44]

R from

RCT702 96 (13) III

5FU, n=348

5FU + IRI, n=354MMR and CT 5 yr-DFS Longer survival with IRI

Tejpar et al, 2009

[45]

R from

RCT1254 188 (15) II + III

5FU, n=633

5FU + IRI, n=621MMR and CT

RFS

OS No significant difference

Oxaliplatin-containing adjuvant chemotherapy

Irinotecan-containing adjuvant chemotherapy

From A. Zaanan

To cut the long oxaliplatin/irinotecan-story short ….



Immune checkpoint inhibitors

15

Sunshine et al, Current Opinion in Pharmacology 2015



Tumormicroenvironment in colorectal cancer

16

Llosa Cancer Discovery 2015



Tumormicroenvironment in colorectal cancer

17

Llosa Cancer Discovery 2015



Mismatch repair deficiency predicts response topembrolizumab (Le et al, NEJM 2015)

I Phase II-study with 41 patients:

― 11 mismatch repair deficient CRC

― 10 mismatch repair deficient non-CRC

― 21 mismatch repair proficient CRC

I Primary endpoints:

― Immune-related response

― Immune related progression free survival

I Significant difference in immune-related response and immune-

related progression-free survival between mismatch repair deficient

and mismatch repair proficient CRC

I Whole exome sequencing revealed 1782 somatic mutations in

mismatch repair deficient tumors and 73 in mismatch repair proficient

tumors

18



Immune-checkpoint-inhibitors for mismatch-repair-deficient colorectal cancer

19

Kelderman, Cancer Cell 2015

But: only 4% of mCRC are mismatch repair deficient!



RAS-/RAF-pathway



KRAS-mutation as a negative predictor foranti-EGFR-treatment

BRAF + PIK3CA mutation 2%NRAS mutation 3%

BRAF mutation5%

KRAS + PIK3CA mutation8%

PIK3CA mutation / PTEN loss12%

Molecular aberration not yet identified

23%

KRAS mutation32%

Non-Response85%

Response KRAS, BRAF,

NRAS,

PIK3CA wild

type, no

PTEN loss



KRAS

NRAS

1 2 3 4 5 6 exon

12/13 59/61 117/146 mutation position

40% 4% 6% frequency (Amgen)

43-49% TCGA

1 2 3 4 5 6 7 exon

12/13 59/61 117/146 mutation position

3.5% 4% 0% frequency (Amgen)

5-9% TCGA

non-coding exon

coding exon

KRAS and NRAS Mutations in Colorectal Cancer



Douillard JY & Oliner KS et al, NEJM 2013



From Tissue to biomarker



25

Hausfälle generell /K-Fälle

nach Anforderung

Anforderung KRAS

KRAS 12/13

KRAS WT KRAS Mut

KRAS 59/61/117/146 Ende

KRAS Mut KRAS WT

Ende NRAS 12/13/59/61

NRAS Mut NRAS WT

NRAS 117/146

Ende

Ende

Workflow

„erweiterte RAS-Testung“


Institut für PathologieExtended RAS-Analyses

Wild type: 2.166 Analyses =

60%

Mutationen: 1.423 Analyses

= 40%

KRAS-Mutational analyses codon 12/13n=5000

WT Mut

Wildtyp: 50%

KRAS Mutation

12/13: 40%

KRAS Mutation 59/61: 3%

KRAS Mutation 117/146:

4%

NRAS Mutation

12/13: 2%

NRAS Mutation 59/61: 1%

NRAS Mutation 117/146:

0%

Extended RAS -Analyses n=1000

0

200

400

600

800

1000

1200

1400

1600

125 RAS cases

1.179 RAS Analyses

Extended RAS: cases versus analyses

KRAS Fälle 01.07.2013-01.09.2013

KRAS Analysen 01.07.2013-01.09.2013



Summary predictive markers

IMSI-H tumors (UICC II) do not benefit from 5-FU

treatment, but do benefit from FOLFOX or FOLFIRI

I Impact of MSI-H is dependent on tumor location and

hereditary background

I KRAS- and NRAS-mutations are negative predictors of

response to anti-EGFR-therapy, but

IOther members of the pathway may also contribute to

non-response: PI3K, PTEN, EGFR, etc.

I Biomarker testing needs rigorous quality control



Molecular signatures in CRC – do we need them?

YES

• To select stage II patients who are at risk of

recurrence (~15%)

• To select stage III patients who are at low risk of

recurrence (~50%)

• To select stage II and III patients who will benefit

from adjuvant chemotherapy



UICC Stage II and Stage III prognosis

I Inside each tumour stage the risk

of recurrence is depending of

various risk factors

I For UICC stage II :

obstruction/perforation, emergent

admission, T4 stage, high-grade,

less than 12 LN are indicative of

poor prognosis

I For stage III the number of

positive lymph-nodes are

associated with the risk of

recurrence

I The only validated prognostic

biomarker is the MSI status in

stage II patients

O’Connor J Clin Oncol 2011;29:3381-88 Weisser J Clin Oncol 2011; 29:4796-802Roth J Cin Oncol 2009; 28:466-74



International Colorectal Cancer Subtyping Consortium ICCSC

Bionetwork Sage initiative (Justin Guinney Steven Friend)

+TCGA COLON DATA + AGENDIA Unpublished


Institut für PathologieTCGA MSI/CIMP CIN Invasive

Mesenchymal

Group DSwiss

Goblet Inflammatory

Enterocyte Transit Amplifying

Group CAgendia

Group BFrench

Group EAMCAJCCII

Group APetacc3

A-type B-type C-type

CIN immune down dMMR CSC CIN Wnt up CIN normal

KRASm

CCS1 CCS2 CCS3

Surface crypt Lower crypt Mixed

Group FNetherlands

1.1. 1.2 1.3 2.1. 2.2.

Rodrigo Dienstmann (version 3, Jan 3rd 2013)

Stem-like

CIMP+



Consensus molecularsubtypes of CRC

32

Guinney et al., Nature Medicine 2015



Consensus molecular subtypes of CRC

33




Concensus molecular subtypes of CRC

34




Conclusion

ICRC is a comlex disease with several subentities derived

through different pathways

I dMMR colon tumors are clearly a subgroup of colon cancer

with specific behavior

I The division of CRC in various subentities generates the

necessity of multicenter trials, since subgroups will be small

I FFPE-material should be collected in these trials and

investigated for potential prognostic and predictive markers

I The addition of molecular data will – hopefully – allow the

development of a more personalized treatment of CRC



FFPE tumor block

HE I: label tumor area

gDNA

MSI

blo

ck a

deq

uate

on e

ntr

y?

(sam

ple

s)

mic

rosta

llelit

ein

sta

bili

ty(M

SI)

adeq

uate

for

analy

sis

(FF

PE

Q)

tum

or

cell

conte

ntetc

.(D

NA

)

am

ountD

NA

HE II: check for tumor



37

Thank you for your attention.

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