Biomarcadores en cáncer colorectal€¦ · Biomarcadores en cáncer colorectal Perspectiva...
Transcript of Biomarcadores en cáncer colorectal€¦ · Biomarcadores en cáncer colorectal Perspectiva...
Biomarcadores
en cáncer colorectal
Perspectiva Clínica
Hacia una medicina molecular
Ramon Salazar MD, PhD Catalan Institute of Oncology
Terapia anti-EGFR
1. Van Cutsem E, et al. J Clin Oncol 2011; 29:2011-9;
2. Bokemeyer C, et al. Ann Oncol 2011; 22:1535-46;
3. Douillard JY, et al. J Clin Oncol 2011; 29(Suppl):3510 (oral presentation).
*Significant P-value; **Non-significant P-value;
pmab, panitumumab; cmab, cetuximab
• KRAS exon 2 mutational status is a key predictor of anti-EGFR mAbs efficacy and a detrimental effect has been observed for anti-EGFRs + FOLFOX in patients with KRAS MT tumours
• mCRC patients with WT KRAS exon 2 may benefit from anti-EGFR mAbs in combination with chemotherapy in all lines of treatment and as single agent in later lines
WT KRAS exon 2
CRYSTAL1 OPUS2 PRIME3
FOLFIRI FOLFIRI + cmab
FOLFOX FOLFOX + cmab
FOLFOX FOLFOX + pmab
RR, % 43 57 p=0.0001 34 57 p=0.002 48 57*
PFS, months 8.4 9.9 HR 0.70* 7.2 8.3 HR 0.56* 8.6 10 HR 0.80*
OS, months 20 23.5 HR 0.80* 18.5 22.8 HR 0.85** 19.7 23.9 HR 0.88**
MT KRAS
RR, % 36 31 52 34 0.02 41 40
PFS, months 7.7 7.4 HR 1.17** 8.6 5.5 HR 1.72* 9.2 7.4 HR 1.27*
OS, months 16.7 16.2 HR 1.03** 17.5 13.4 HR 1.29** 19.2 15.5 HR 1.17**
KRAS testing predicts efficacy to cetuximab and panitumumab in major
registration trials
DNA mutation in XXXX gene
DNA: normal
Ala Ala Ser Asn Leu Gln Trp Arg Protein: normal
DNA: mutated
Ala Ala Ser Asn Leu STOP Protein. truncated
MUTATION 9604C>T = Gln3126Stop in XXXX
N
Control
Patient
Sanger
Sensitivity of mutation detection 10-20%
ARMS Sensitivity of detection~1%
Mutations in codons 12, 13 and 61
(20 mutations panel)
Mutations in codons 12 and 13
(7 mutations panel)
TaqMelt
Sensitivity of mutation detection ~1-5%
Mutation detection kits in KRAS
Douillard J-Y, et al. N Engl J Med 2013;369:102334.
Percentages have been rounded. 7 patients harboured either
KRAS or NRAS codon 59 mutations. RAS and BRAF ascertainment rate: 89%.
KRAS, NRAS extended panel and BRAF mutation hotspots in
the PRIME trial (& others)
NRAS
KRAS
EXON 2 EXON 3 EXON 4 EXON 1
12 13 61 117 146
EXON 2 EXON 3 EXON 4 EXON 1
12 13 61 117 146
BRAF
EXON 15 EXON 16… EXON 1…
600
40% 4% 6%
3% 4% 0%
59
59
8%
Among WT KRASexon 2 patients, an additional
17% of tumours with RAS mutations were found
12 13 61 117 146 59
12 13 61 117 146 59
600
Overall RAS
ascertainment
rate: 90%
Other RAS mutations: CRYSTAL & OPUS
Ciardiello F et al. Ann Oncol 2014; Tejpar S et al. Ann Oncol 2014
Study N Method Sensitivity Other RAS
mt, %
CRYSTAL 430 BEAMing† 0.01% 14.7
OPUS 118 BEAMing† 0.01% 26.3
FIRE-3 407 Pyrosequencing 1-5% 16.0
PRIME 620 Dideoxy sequencing/WAVE 5% 17.4
PEAK 221 Dideoxy sequencing/WAVE 5% 23.1
WT KRAS ex 2 WT All RAS
Study PRIME (+/- Panitumumab)
N 656 512
PFS HR / p 0.8 / 0.02 0.72 / 0.04
OS HR / p 0.83 / 0.072 0.78 / 0.043
Study CRYSTAL (+/- Cetuximab)
N 666 367
PFS HR / p 0.7 / 0.001 0.56 / 0.0002
OS HR / p 0.8 / 0.009 0.69 / 0.002
Study FIRE-3 (Cetuximab vs Bevacizumab)
N 592 342
PFS HR / p 1.06 / 0.54 0.93 / 0.54
OS HR / p 0.77 / 0.017 0.7 / 0.011
Douillard JY, NEJM 2013; Ciardiello F, Ann Oncol 2014; Schwartzberg L, JCO 2013 (A 3631); Stintzing S, EJC 2013 (Proc ECCO)
All RAS mutations: additional efficacy effect
RAS mutations: Which is the right
sensitivity cut-off?
Ciardiello F et al. Ann Oncol 2014
RAS mutations: Which is the right
sensitivity cut-off?
Ciardiello F et al. Ann Oncol 2014
?
RAS mutations: Which is the right
sensitivity cut-off?
Ciardiello F et al. Ann Oncol 2014
?
?
RAS mt clone RAS wt clone
Sensitive tumor
RAS mutations: Which is the right
sensitivity cut-off?
Resistant tumor
Sensitivity cut-off?
Cobas 4800 (ROCHE) – Cobas Kits Sensitivity 1-5%
PyroMark (QIAGEN) – TherascreenPyro Kits Sensitivity 1%
BioMark Digital PCR Arrays (Fluidigm) Sensitivity 0.1%
LightCycler 480 Sensitivity 1-5 %
Sensitivity 5-10%
The puzzle of RAS testing
Slide provided by R. Salazar
BEAMing Sysmex Inostics Sensitivity 0.01%
ICORAS Panel – LightCycler and Fluidigm –
38 mutations (RAS+BRAF+PI3KCA)
Common mutations detected by the KRAS, NRAS, RAS Extension and BRAF Pyro Kit
81 mutations panel (RAS+BRAF) Sensitivity 5%
ICO-HUB National Referral Diagnostic Lab
Clinical Trial “ULTRA” (TTD)
N=80+
N=28+
N=82+
LightCycler Roche (ICO) Wave Surveyor (Others)
PyroMark Qiagen
Clinical Practice (routine)
Analysis Platforms Comparative Study (TTD-RTICC)
Cobas 4800 (ROCHE) – Cobas Kits Sensitivity 1-5%, extended panel
PyroMark (QIAGEN) – TherascreenPyro Kits Sensitivity 1%, 81 mut panel
BioMark Digital PCR Arrays (Fluidigm) Sensitivity 0.1%
LightCycler 480 Sensitivity 1-5 %
38 mut panel (sizeable up to 48)
Plataformas Next Generation Sequencing
Illumina MiSeq
Illumina HiSeq
Life Technologies Solid & Ion Torrent
Junior 454-Roche
Construcción librería para capturar genes a analizar
Access Array Target-specific Primers
Multiplicom Somatic 1MASTR
Illumina Trusight Tumor
Diseño personalizado, se ha de validar. Más flexibilidad, más económico
26 genes Está validado y en uso en varios laboratorios Menos flexibilidad, más caro
3 genes NRAS, KRAS, BRAF Está validado y en uso en varios laboratorios Menos flexibildad, máscaro
Agilent Sure Select
CONCORDANCE: High sensitivity methodologies for mutational analysis in
circulating tumor DNA
BEAMing technology Sensitivity 0.01
Fludigm technology Sensitivity 0.1
PlasmaSeq technology Sensitivity 0.1?
Vilar E & Tabernero J, Nature 2012
“Under pressure” clonal selection vs evolution
28
Treatment of Resistance in present
or future clinical trials
RAS mutants:
Cetuximab + Irinotecan in KRAS G13D mutant CRC….
MEK inhibitors + pan HER inhibitors
BRAF mutants:
BRAF + EGFR + PIK3Ca
S492R mutants:
Panitumumab
C-MET amplifications
C-MET inhibitors ?
Tumor debulking in unresectable metastasic disease
proof of principle cfDNA load as surrogate endpoint of
effective debulking of clones with resistance potential
Acknowledgements & Collaborations Agendia
Iris Simon
Paul Roepman
Annuska Glas
Sun Tian
Laura V´Veer
IDIBELL CRC Group, HUB & ICO CRTeam
Víctor Moreno
Adriana López-Doriga
Rebeca Sanz-Pamplona
UMC Leiden
Rob Tollenaar
Wilma Mesker
H. Del Mar BCN
Clara Montagut
NKI R. Bernards Lab
Anirudh Prahallad
Ian Majewski
Magali Michaut Slotervaart Hospital
Johan Westerga
Sjoerd Bruin
Rechts der Isar Munich Robert Rosenberg Ulricht Nitsche
Hospital Gasthuisberg Leuven Sabine Tejpar
Swiss Inst Bioinformat
Vlad Popovici Mauro Delorenzi
VHIO BCN
Josep Tabernero
J. Capdevila
Rodrigo Dienstmann
Gabriel Capellà
Alberto Villanueva
David García-Molleví
Dani Azuara
Marga Nadal
E. Kreisler, S.Biondo
Xavier Sanjuan
Antonio Soriano
Cristina Santos
Gemma Soler
Merche Martínez V.
Sonia Abenia
Alex Teulé
Julieta Grasselli
Maria Cambray
Xavi Pérez
JR Germà
UBB
LRT UGR
HUB UFCCR
ICO CRT
SOM ORT UIC
IRB
E Batlle E.Sancho
PEBC
M Esteller Ana Martínez
Idibell G. Thomas MDACC S. Kopetz
www.iconcologia.net
Institut Català d’Oncologia
ICO l’Hospitalet Hospital Duran i Reynals Gran Via de l’Hospitalet, 199-203 08907 l’Hospitalet de Llobregat
ICO Badalona Hospital Germans Trias i Pujol Ctra. del Canyet s/n 08916 Badalona
ICO Girona Hospital Doctor Trueta Av. França s/n 17007 Girona
PCR Digital
770 nanopocillos: se realiza una PCR en tiempo real en cada uno de ellos
PCR Digital
770 nanopocillos: se realiza una PCR en tiempo real en cada uno de ellos