Prof. Dr. Manfred Dietel

Transcription

Fortschritte in der personalisierten Medizin
M. Dietel
Institute of Pathology
(Rudolf-Virchow-Haus)
Humboldt University, Berlin
Rudolf Virchow
1821 - 1902
1710 - 2010
e-mail: manfred.dietel@charite.de
09.10.2013
Institut für Pathologie – Charité Berlin
1
Allgemeine Definition
Es geht nicht darum, für einen Patienten ein
individuell predictive
maßgeschneidertes Medikament zu
tests
based
entwickeln
und
dann zu verabreichen.
on reliable BM
Definition in der Onkologie
Unter personalisierter Medizin in der Onkologie wird die durch
die Arzneimittelzulassungsbehörde bindend
vorgeschriebene prädiktive Biomarker-basierte
Diagnostik als Voraussetzung zur Applikation eines
zielgerichteten Arzneimittels
verstanden.
Es wird also vor Gabe eines Medikamentes der Nachweis
seiner Wirksamkeit gefordert (compagnon diagnostic).
FDA: Drug-Diagnostic Co-development Initiative
09.10.2013
3
Prediction is difficult,
especially about the future
Niels Bohr, 1885-1962
2010
Thinking back to infectious diseases and seeing the current development in cancer – HER2, KRAS, BRAF, EGFR etc. - we shouldn’t be too
pessimistic.
In oncology, the predictive power of tissue
based analyses is underestimated.
09.10.2013
4
Predictive tissue-based biomarkers for targeted therapies
FDA / EMA-approved drugs associated with eligibility tests* (selection)
•
Trastuzumab → metastatic breast cancer, overexpression/amplification of HER-2
•
Cetuximab → metastatic colorectal cancer, overexpressing EGFR/wild-type KRAS
•
Panitumumab
→ now,
colorectal
KRAS a
(mutation
excluded)
Already
incancer
35%with
of wild-type
all tumors
predictive
•
Gefitinib → non-small cell lung cancer with mutated EGFR
•
is appropriate.
Notably,
molecular
Erlotinib
→ non-smalltest
cell lung
cancer with mutated
EGFR
•
Crizotinib → non-small cell lung cancer with mutated EML4-ALK
•
Nimotuzumab → metastatic colorectal cancer (still experimental)
•
Lapatinib → metastatic breast cancer overexpression HER-2/neu (?)
•
Vemurafenib (PX4032) → malignant melanoma with mutated B-RAF
•
All→these
substances(activated
have PK),
been
Imatinib
CML, bcr/abl–positive
•
Imatinib → GIST with activated c-kit receptor tyrosine kinase/CD117, exon 9 mut
•
Rituximab (+ CHOP), Y90-Ibritumomab, I131-Tositumomab → NHLymphoma with CD20
•
Gemtuzumab-Ozogamicin
AML importance
with CD33 ( > 60 yrs.),
mal. melanoma
This underlines→ the
of biobanks.
•
Tamoxifen+/- chemo → ER+/HER2 - breast cancer, mutation pattern - multigene assays
prediction of
tumour response is exclusively tissue-based.
developed on the
basis of histologically characterised human tissue.
*Strongly suggested by FDA’s Drug-Diagnostic Co-Development Initiative
09.10.2013
5
Recommendations for sample preparation and molecular analysis
Surgery
Surgical
Pathology
tumour resection
grossing
paraffin embedding
histological evaluation
Oncological
treatment
manualmicrodissection
microdissection
manual
Molecular
Pathology
DNA isolation
amplification
detection
09.10.2013
6
Manual microdissection
Director: Prof. M. Dietel
To
Dr. ##########
#############
Charitéplatz 1
10117 Berlin
Tel. 0049 30 450 536 001
Fax 0049 30 450 536 900
E-mail: manfred.dietel@charite.de
Incoming
XXXXX
Name
Birth
Molecular pathology report
Combined Report on Anatomic and Molecular Pathology
Without manual microdissection
the error rate can be
Material: external
colon biopsy, FFPE - block no. ##### -10
up to 8%* due only to inadequate
samples,
Clinical Data: Met. ### cancer ###
independent of any technicalHistopath.:
problems
Malignant epithelial ……….
Weichert W et al. J Mol Diagn;12:35-42
BRAFV600mut
Nerv
Forward
Automated
DNA extraction
Reverse
Sequencing
Mol. analysis:
KRAS mutation GGTGGC to GATGGC
(ca. 40%)
XXXXX
Combined patho-report: Metastasized adenocarcinoma of
the ##### with ### mutation as indicated.
Pathologist…………………..
09.10.2013
7
Die aktuell immer wieder gestellte Frage …
Sind die Ergebnisse so überzeugend, dass der enorme
Aufwand und die exorbitanten Kosten der onkologsch
orientierten PM gerechtfertigt sind?
•Kosten der F&E ca. 2 Mrd.€ pro neue Substanz
•Kosten pro Therapiezyklus ca. 30-40 T€
Die aktuell immer wieder gestellte Frage
Was sind überzeugende Ergebnisse?
Einige Beispiele ………..
Ulzeriertes Dickdarmkarzinom
Dieser Tumor bzw. seine Metastasen können mit
einem zielgerichteten Medikament (therapeutischer
AK) behandelt werden, wenn mittels molekularer
Analysen ein KRAS-Wildtyp nachgewiesen wurde.
Warum?
KRAS-MAPK signalling pathway
Y Y
Ligands
Panitumumab and cetuximab inhibit ligand binding, dimerisation, activation of the receptor and the signalling pathway
EGFR
KRAS
mutant KRAS
(Wild-type)
RAF
MEK
Mutant KRAS
constitutively
active –
40% of patients*
ERK
ELK
Activation of
• Proliferation
• Angiogenesis
• Malignant phenotype
09.10.2013
Nucleus
Schubbert S et al. Nat Rev Cancer 2007;7:295-308;
für Pathologie
Charité
Berlin
*Friday BB, Adjei Institut
AA. Biochim.
Biophys.–Acta.
2005;
1756:127-144.
11
Metastasierte Colon-Ca: Wild-type/mut KRAS/BSC
1.0
Proportion Event Free (%)
0.9
Pmab + BSC WT
BSC alleine
Pmab + BSC Mut
0.8
0.7
0.6
0.5
115/124 (93)
114/119 (96)
12,3
7,3
76/84 (90)
7,4
HR = 0,45 (95% CI: 0,34–0,59)
Stratified log-rank, P < 0,0001
0.4
0.3
Events N (%)
Median
(weeks)
16 weeks
0.2
0.1
0.0
0 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30 32 34 36 38 40 42 44 46 48 50 52
Weeks
Patients at Risk
Pmab + BSC 124 119 112 106 80 69 63 58 50 45 44 44 33 25 21 20 17 13 13 13 10 7 7 6 5 5
BSC alleine 119 109 91 81 38 20 15 15 14 11 10 9 9 6 6 6 6 5 4 3 3 2 2 2 2 1
Amado R, et al. ESMO 2007;a0007; JCO, 26 (2008 1626-1634
Institut für Pathologie – Charité Berlin, Campus Mitte
Treatment efficacy - according to mut status
In CRC ca. 55% are KRAS wild type.
Out of these only ca. 50% (i.e. 25% of all CRC) respond
to EGFR antibodies.
Why?
How can they be detected or stratified?
09.10.2013
13
Mutations in KRAS and NRAS genes in colorectal cancer
KRAS
1
2
3
4
12/13
59/61
117/146
40%
4%
6%
2
3
4
12/13
59/61
117/146
3.5%
4%
0%
5
6
exon
mutation position
frequency
NRAS
1
non-coding exon
coding exon
09.10.2013
5
6
7
exon
mutation position
frequency
When the rare mutations are added
they represent 17.5 % of all CRC and
they are associated with resistance!
14
20020408 Trial RAS (Exon 4) Analysis
PFS in Patients with WT RAS* mCRC
Events
n/N (%)
Median weeks
(95% CI)
Panitumumab + BSC
(n=73)
70 (96)
14.1 (10.3–23.3)
BSC (n=63)
61 (97)
7.0 (6.0–7.4)
100
90
80
70
HR=0.36 (95% Cl: 0.25–0.52)
p<0.001
Proportion Event-Free %
60
50
40
30
20
22 weeks
10
0
0
4
8
12 16 20 24
28 32 36 40 44 48 52 56 60 64 68 72 76 80 84 88 92 96 100
Weeks
*WT KRAS and NRAS exons 2, 3, and 4
Patterson SD, et al. J Clin Oncol 31, 2013 (suppl; abstr 3617).
NSCLC - Macroscopy
aeno carcinoma
broncho-alveolar type
Für diese Tumoren kommt eine Therapie mit
Tyrosinkinase-Inhibitoren in Betracht, wenn im
Einzelfall bestimmte Mutationen nachgewiesen
werden können.
central
squamous cell carcinoma
peripheral
adenocarcinoma
NSCLC: Past and Current Landscape
1999
Histology-driven
selection1
2012
Targeting oncogenic
drivers
NO MUTATION
DETECTED
Adenocarcinoma
Squamous-cell carcinoma
Large cell carcinoma
AKT1
NRAS
MEK1
MET AMP
HER2
PIK3CA
BRAF 2%
DOUBLE
MUTANTS 3%
EML4-ALK
7%
KRAS
22%
EGFR
17%
Actionable driver mutations identified in 54% of lung adenocarcinoma tumours
LCMC, Lung Cancer Mutation Consortium
09.10.2013
Kris MG, et al. Presented at ASCO 2011; Abstract CRA7506
17
Rapid Responses Seen In Some Patients
Ou et al. J Thoracic Oncol 2010;5:2044–2046 Camidge RD et al.: ASCO 2011
09.10.2013
18
Tumour responses to crizotinib by patient
Decrease or increase from baseline (%)
100
Study A8081001
N=1161
PROFILE 1005
N=2402*
100
80
80
60
60
40
40
20
20
0
0
–20
–20
–40
–40
–60
–60
–80
–80
–100
–100
Best objective response according to RECIST:
PD
SD
PR
1. Camidge DR, et al. Lancet Oncol 2012;10:1011−9;
2. Kim DW, et al. Presented at ASCO 2012; Abstract 7533
09.10.2013
CR
*Mature population, excluding those with early death,
indeterminate response and non-measurable disease
19
EML4-ALK Fusion in NSCLC
Chromosom 2
ALK
EML4
Anaplastic
lymphoma kinase
Exon20
Exon13
constitutive
activation
Alk
11 variants
EML4
20
13
PF02341066
20
20
20
6a, b
EML4-Alk
EGFRwt
KRASwt
Modified according to
Soda et al. nature 448:561 (2007).
Institut für Pathologie – Charité Campus Mitte
ALK
EML4
Anaplastic
lymphoma kinase
Exon20
Alk
09.10.2013
Exon13
EML4
21
E23002-11_ALK-BA
Malignant Melanoma
50% of all malignant melanomas exhibit a BRAF-Mutation
*Total V600 mutation rate for BRIM-3 (cobas® 4800 BRAF V600 Mutation
Test); 9.9% of the cobas-positive cases subjected to retrospective Sanger sequencing had V600K mutations
09.10.2013
22
Overall survival (%)
Vemurafenib phase I overall survival: Updated KM estimates (08. 2011)
100
90
80
70
60
50
40
30
20
10
0
1 year
2 years
BRAF-mutated
— V600E dose escalation
(n=16)
— Extension (n=32)
— WT or sub-therapeutic
exposure (n=33)
wild-type
0
3
6
9 12 15 18 21 24 27 30 33 36
Median OS (month)
Time since first dose (months)
Extension cohort landmark
Estimated survival: 1 year = 50%, 2 years = 38%
09.10.2013
Dose escalation
25.2
Extension
13.8
WT or sub-therapeutic.
4.18
23
Vemurafenib inhibits V600 mutated BRAF kinase
Response to
BRAF-inhibitors is
given only if a
BRAF mutation is
present
This has to be
tested prior to the
therapy.
09.10.2013
24
Vemurafenib inhibits V600 mutated BRAF kinase
RTK
RAS
50-60%* of melanomas
RAFV600mut
BRAF
ATP
• Constitutive
activation is
independent of
extracellular factors
VEMURAFENIB
(PLX4032, RG7204, RO5185426)
MEK-IB
MEK
ATP
• Not responsive to
normal regulatory
signals
ERK
Cellular
Proliferation
Cellular
Survival
*Total V600 mutation rate for BRIM-3 (cobas® 4800 BRAF V600 Mutation
Test); 9.9% of the cobas-positive cases subjected to retrospective Sanger sequencing had V600K mutations
Presented in Vienna at ESMO 09/2012:
Flaherty (NEJM, 2012):
OS from 5.8 months with monotherapy to 9.9 months
with combinational targeted therapy.
Institut für Pathologie – Charité Campus Mitte
Next Steps in Molecular Pathology –
Multigene Assays in Breast Cancer
• Multi-gene analyses, predictive molecular pathology and
response to chemotherapy in breast cancer
• Which patient with ER+ and Her2 neg. breast carcinoma
will show a good prognosis when treated by endocrine
therapy only?“
09.10.2013
27
EPclin: Validation
ABCSG-6 – mol. + clin. EP
1
96%
0.8
78%
Martin Filipits, Margaretha Rudas,
Raimund Jakesz, et al.
0.6
Following the EPclin-based
predictive data 96% of the
A new molecular predictor of distant recurrence in
low-risk patients do not show-up
with metastases
P < 0.001
ER-positiveHER2-negative breast cancer adds
0.4
independent information to conventional
after 10 years.HR 7.97 (3.56-17.83)
clinical risk factors.
Clin Cancer Res Published OnlineFirst August 1, 2011.
0.2
EPclin low
EPclin high
0
0
20
40
numbers at risk:
09.10.2013
60
80
months
100
120
208
200
194
129
120
114
91
170
156
141
87
76
71
57
28
Stratification by EndoPredictclin ®
1702 Patientinnen in
ABCSG 6 & 8
nach S3-Leitlinien
248 Pat.
„low risk“*
5,3 %
Metastasen
w/o Endopredict
these patients may
have recieved CTx
09.10.2013
1.371 Pat.
„intermed. risk“*
840 EPclin
„low risk“
4,5 %
Metastasen
531 EPclin
„high risk“
83 Pat.
„high risk“*
44,5 %
Metastasen
19,7 % Metastasen
*nach S3-Leitlinien
29
A look into the future, exemplified by a current case
09.10.2013
30
Up-coming Molecular Diagnostic
histological
diagnosis
metastasized
neuro-endocrine
carcinoma, grade 3
standard sequential
molecular diagnostics
KRAS
BRAF
EGFR exons 18,19, 21
cKIT
usw.
parallel molecular
diagnostics
IonAmpliseq* Cancer
Panel in 46 gene
(total 604 loci).
other relevant
mutations
no mutations
????
*Ion Torrent
09.10.2013
31
Proprietary Semiconductor Sequencing Technology
Ion Torrent®, the chip is the mashine
If a nucleotide, for example a C, is
added to a DNA template and is then
incorporated into a strand of DNA, a
hydrogen ion will be released. The
charge from that ion will change the pH
of the solution, which can be detected
by our proprietary ion sensor.
09.10.2013
32
Up-coming Molecular Diagnostic
histological
diagnosis
metastasized
neuro-endocrine
carcinoma, grade 3
standard sequential
molecular diagnostics
parallel molecular
diagnostics
KRAS
BRAF
EGFR exons 18,19, 21
cKIT
usw.
IonAmpliseq* Cancer
Panel in 46 gene
(total 604 loci).
ABL
APC
ALK
KRAS
BRAF
no mutations Iressa ⇒ EGFR mut exon 20
ERBB2
FGFR2 mut
FGFR-inhibitor ⇒
FGFR3
cKIT
sorafinib/sufitinib ⇒
KDL mut
604 further loci…….
*Ion Torrent
09.10.2013
33
Clinical
data
tissue
Digital pathology
wild type
normal + mutated
Personalisierte (Onko-)Medizin basiert auf einer
All test are
done on
formalin fixed
paraffin
embedded
tissue
komplexen
diagnostischen Prozedur, die Pathologie und Klinik
gleichermaßen zu erfüllen haben.
Multigene
Weitere Fortschritte, z.B. neue Substanzen, neue
SubstanzAssays
Kombinationen, weitere Tumorentitäten etc., werden im
Verbund von Akademia und forschender Pharma-Industrie in
den kommenden Jahren erreicht werden.
09.10.2013
34
Institute
of Pathology,
Institut
für Pathologie,
Rudolph-Virchow-Haus, Charité
Rudolf-Virchow-Haus, Charité
Humboldt-Universität zu Berlin
Humboldt-Universität zu Berlin
Berliner
Medizinhistorisches
Museum
Berlin Wall
The
Berliner
Mauer
Alexander Ufer
Institut für Pathologie – Charité, Berlin, Campus Mitte
35
Multidisciplinary cooperation
enables personalised oncology
Cohorts:
Clinical Trials
Tumour Registries
Radiology
Sampling
Tumour
board
Endoscopy
Surgery
Patient
Oncologist
Targeted
therapies
pre-analytic
Predictive
biomarkers
Results
Diagnosis
IHC
Pathology
QA/QC
I+D+i
In situ hybridization
PC
R
Publications
Consensus
Teaching
Conde E, et al. Clin Transl Oncol 2013;15:503–8; AMP Whole Genome Analysis WG. J Mol Diagn 2011;13:249–51
09.10.2013
36
Three Cellular RAS Genes Encode
Four Highly Homologous 21 kD Proteins
Hypervariable
region
G domain
P loop
12
Switch I
13
Switch II
61
1
189
KRAS4A
C CIIM
12
13
61
1
188
KRAS4B
KKKKKK
12
13
CVIM
61
1
189
NRAS
C
1
12
10
13
16
32
38
61
59
67
85
165
CVVM
189
HRAS
C C
09.10.2013
CVLS
37
*Boxes at the bottom of each isoform representation show the
conserved
residues in magenta and the variable residues in pink.
Adapted from Schubbert S, et al. Nat Rev Cancer 2007; 7:295-308.
PFS in Patients with WT KRAS Exon 2 mCRC
Events
n (%)
Median weeks
(95% CI)
Panitumumab + BSC
(n=124)
115 (93)
12.3 (8.3–16.1)
BSC (n=119)
114 (96)
7.3 (7.0–7.7)
100
90
80
70
HR=0.45 (95% Cl: 0.34–0.59)
p<0.001
60
50
40
30
20
10
0
0
4
8
12 16 20 24
28 32 36 40 44 48 52 56 60 64 68 72 76 80 84 88 92 96 100
Weeks
Patterson SD, et al. J Clin Oncol 31, 2013 (suppl; abstr 3617); 1. Amado RG, et al. J Clin Oncol 2008; 26:1626-34.
Overall Survival Kaplan-Meier Plot in Patients
with Non-Mutated RAS
Overall
PFS in Patients with MT RAS* Exon mCRC
Events
n/N (%)
Median weeks
(95% CI)
Panitumumab + BSC
(n=99)
90 (91)
7.4 (7.3–7.7)
BSC (n=114)
108 (95)
7.3 (6.4–7.9)
100
90
80
70
HR=0.97 (95% Cl: 0.73–1.29)
p=0.729
60
50
40
30
20
10
0
0
4
8
12
16
20
24
28
32
36
40
44
48
52
56
60
64
68
72
76
80
84
86
Weeks
*MT in any KRAS and NRAS exons 2, 3, and 4
Patterson SD, et al. J Clin Oncol 31, 2013 (suppl; abstr 3617).
Predictive Molecular Pathology and
Personalized Medicine
A prerequisite of personalized medicine is the
capability to predict pre-therapeutically the
response of individual tumors to certain (targeted)
drug.
For this prediction one needs reliable and
reproducible biomarker and predictive assays.
This is the current challenge of predictive molecular
pathology.

Prof. Dr. Manfred Dietel

Transcription

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