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Molecular profiling of colorectal cancers
Dr Angela Silmon12th September 2014
Colorectal NSSG Audit Day
Outline
• Brief introduction to NewGene• Personalised Medicine• Molecular testing in colorectal cancer• Horizon scanning
NewGene Ltd
A pioneer in developing, validating and delivering molecular diagnostics using the
latest high throughput sequencing and genotyping technologies
Technology platforms
Genotyping
• Sequenom MassARRAY 4– MALDI TOF mass spectrometer– Low cost– Rapid turn around– Targeted mutations
F
R
5’end
3’end
[C/G]
[G/C]3’end
5’end
[C/G]
[G/C]
extension into SNP site5500DaC
5800Da
G6100Da
G
C
Allele 1
Allele 2
PCR SAP clean
Extension reaction Resin Spotting MALDI TOF Data
analysis
Laser
Detector
Time of flight
Sequenom chip
Sequenom Mass Array 4
Sequenom Mass Array 4
5500Da 7000Da 7000Da 7000Da
Wild type allele only (TT)
Mutant allele only (CC)Wild type and mutant allele present (TC)
Personalised medicine portfolio
Gene Mutations
Non-small cell lung cancer
EGFR 17 point mutations Indels exon 19 and 20Inclds T790M
Metastatic colorectal cancer
KRAS, BRAF, NRAS 23 mutations in each RAS geneInclds BRAF V600E
Melanoma BRAF
GIST cKIT, PDGFRA Two step process
FundingGene Mutations
Non-small cell lung cancer
EGFR Astra Zeneca until Oct 2010CCGs
ALK* Pfizer2015, likely change
Metastatic colorectal cancer
KRAS, BRAF, NRAS Merck Serono until May 2014 NHS England
Melanoma BRAF Roche2015, TBD
GIST cKIT, PDGFRA Referring Trust
* IHC carried out at Cellular Pathology, RVI, FISH test carried out by Cytogenetics
Referral pathway
Request CDF number
Refer for testing incld CDF number
Automatic un-registration from CDF
Test result
Therapy decisionCDF number if req
T176
Refer for testingCheck T176 box
Test result
Therapy decision
Funding: CDF
Funding: NHS England
Molecular testing in mCRC
• Mutation in the RAS genes offer prognostic value with patients with a mutation in the KRAS or NRAS gene having lower overall survival rates compared to wild type
• Mutations predict lack of benefit from EGFR targeted therapies such as Cetuximab and Panitumumab
• Mutations in codons 12 and 13 of KRAS most common, ~40% of mCRC patients
• Mutations in RAS genes found in ~ 50% of patients
• Targeted mutations– KRAS codons 12, 13, 59, 61, 117 and 146– NRAS codons 12, 13, 59 and 61
• Interpretation– Therapy license - wild type RAS genes only– BRAF prognostic indicator but guidelines may
change
Association of Clinical Pathologists Molecular Pathology and Diagnostics Group
Wong NACS, Gonzalez D, Salto-Tellez M, et al. J Clin Pathol 2014
Association of Clinical Pathologists Molecular Pathology and Diagnostics Group
• Primary or metastatic CRC tissue can be used • Biopsy or resection specimen tissue can be
used• The minimum neoplastic cell content tested
should be at least two times the assay’s LOD.• Audit– TAT ≤7 working days for >90% of samples– Incident rates – Failure rates
Wong NACS, Gonzalez D, Salto-Tellez M, et al. J Clin Pathol 2014
NewGene Audit 2014• Turn around time
– Target 5 working days– Range 2 - 11 days– Average 3.4 days– 90% of reports issued within 5d
• Incidence rate
• Failure rate– < 1%
• EQA– Participation since 2010– All genotyping maximum
marks, no poor performance
Wild type samples 42%Mutation positive samples 58%Gene NewGene Reported
KRAS 42.7% 40%
NRAS 6% 5%
BRAF 9.3% 10%
Jan – June 2014N = 500
Horizon scanning
• Require a comprehensive joined up approach to molecular testing– Funding mechanism– Testing mechanism
Gene Horizon
Non-small cell lung cancer
EGFR KRAS
Metastatic colorectal cancer
KRAS, BRAF, NRAS
Melanoma BRAF NRAS
Horizon scanning
• OncoFocus– Comprehensive test for actionable mutations
• UltraSeek– Circulating tumour DNA– 1% tumour content
Gene AAABL1 T3151AKT1 E17KALK F1174LBRAF V600E, V600K, V600REGFR G719S, T790M, L861Q, E746- A750delFLT 1836delIDH1 R132HIDH2 R140Q, R172KJAK2 V617FKRAS G12A, G12D, G12V, G12C, G12S, G13DNRAS Q61K, Q61RPIK3CA E545K, E542K, H1047R
Horizon scanning
• Pilot data generation– OncoFocus– UltraSeek– Business case for service delivery– Phase I Clinical trials– Clinical implementation
