The OncoScan microarray offers high-quality copy number, genotype, and somatic mutation data with whole-genome coverage and high resolution in cancer genes for use with challenging FFPE samples.

1. The OncoScan FFPE Assay Kit for analysis of copy
number and somatic mutations in cancer
Larry Greenfield M.D., Ph.D

2. Affymetrix’ clinical product portfolio
Cancer
Hematological malignancies
Solid tumors
Drug Metabolism
Reproductive
Health
Prenatal
Postnatal
Mendelian diseases

3. For Research Use Only
Challenges for laboratories
 Growing need for genome wide copy number profiles
 Large and growing list of actionable copy number
aberrations
 Detection of clonal populations enables longitudinal patient
assessments
 Number of copy number aberrations indicative of prognosis
 For solid tumor analysis, FFPE is a critical specimen type but is
often highly degraded, and generates limited DNA
 Molecular profiles need to be generated rapidly and at an
affordable price
 Multiplex multiple markers with minimal data analysis
burden

4. For Research Use Only
Pan Indication Copy Number Actionable Genes
AKT2 CTNNB1 JAK2 MPL PTPN11
ALK DNMT3A JUN MPM1 RAB25
APC EGFR KIT MYC RB1
AR ERBB2 KRAS MYCN REL
ASXL1 ESR1 MAP2K1 MYD88 RET
ATM FGFR1 Map2K2 NOTCH1 RUNX1
BRAF FGFR4 MDM2 NRAS TET2
CCND1 FLT3 MET PDGFRA Tp53
CDK4 IDH1 MITF PIK3CA VHL
CEBPA IDH2 MLL PTEN WT1
 Leading cancer centers* are
developing a growing panel of
actionable pan indication copy
number aberrations
 Examples include MYC
amplification (prognostic in
colorectal cancer, prostate and
breast), Her2 amplification
(predictive in breast, ovarian and
endometrial cancers)
Large and growing list of actionable
copy number changes
*Test panels from Washington University, MD Anderson, MSKCC

5. For Research Use Only
 Chromosomal copy
number changes
contribute significantly to
tumor development and
progression in
gastrointestinal stromal
tumors (GIST)
 Analysis evidencing
“…genomic complexity as
the best predictor of
disease relapse”
 In addition, pattern of
genomic loss may
provide differential
diagnostic information
Genomic complexity indicative of prognosis
GIST as an example
Chromosome copy number changes carry prognostic information independent of KIT/PDGFRA point mutations in
gastrointestinal stromal tumors
Mara Silva1, Isabel Veiga1, Franclim R Ribeiro1, Joana Vieira1, Carla Pinto1, Manuela Pinheiro1, Bárbara
Mesquita1,Catarina Santos1, Marta Soares2, José Dinis2, Lúcio Santos3, Paula Lopes4, Mariana Afonso4, Carlos
Lopes4 and Manuel R Teixeira*1,5
1q gain 12q gain
1p loss 14q loss
15q loss 22q loss

6. OncoScan enables previously
inaccessible solid tumor studies
 “There are currently few studies to have
realized this potential for FFPE samples,
owing to the inherent problems of
working with chemically modified and
damaged DNA, the requirements for
larger amounts of genomic DNA and/or
high depth of sequencing coverage to
minimize ambiguous read mapping and
the subsequent added bioinformatics
challenges that this brings”
 “The recently developed OncoScan, a
high-resolution SNP array based on
molecular inversion probes, appears to
perform well in comparison with aCGH
platforms and so might prove to be a
successful method for studying FFPE
samples for CNAs”

7. For Research Use Only
 “…Our results suggest that
this FISH panel would
have missed 6
melanomas in our cohort
that OncoScan detected,
one of which proved to be
lethal to the patient
 This implies a single
OncoScan test could be
used to confirm the
diagnosis of malignant
melanoma, stratify the
risk of recurrence, and
direct pharmacologic
interventions”
With enormous implications
FISH-based misdiagnosis
OncoScan can also be used
to multiplex the commonly
tested druggable somatic
mutation BRAFv600E along
with copy number
information

8. OncoScan™ FFPE Assay Kit
Key features
• Input DNA of 80ng
• Will work with archived & highly degraded FFPE samples
• 48 hrs from DNA to results
• Off the shelf product with kitted reagents including OncoScan™
Nexus Express software
• Works with existing Affymetrix instruments
• Universal reference to establish normal baseline
• Content
– Genome wide copy number determination
– High resolution on ~900 cancer genes
– Genome wide LOH resolution
– Detection of frequently tested somatic mutations
• Clonal population detection
• High copy number dynamic range

9. OncoScan™ FFPE Assay
enabled by MIP technology
 High specificity – low false positives and assay noise: unique approach to interrogating the
genome where the probe is amplified rather than the genomic DNA
 Ideal for FFPE samples that often suffer from severe degradation and chemical impurities: assay
requires only a small footprint (150bp) of intact DNA sequence per region
 Genetic content extension possible by the addition of new probes with new tag sequences that
are already captured by complementary sequences on the array
~40 base pairs

10. OncoScan™ FFPE Assay
Advances in algorithm and data visualization
Advances in data analysis:
 internal bioinformatics advances in algorithm development
 data visualization partnership with BioDiscovery (Nexus
software)
Enabling the following novel features:
 Incorporation of the Affymetrix’ TuScan algorithm that
supports the determination of tumor burden and ploidy to
establish a baseline for copy number determination
 Corrected for systematic variability (e.g. local and probe GC
correction)
 Ability to correlate copy number and clinical outcomes data
across multiple samples in a clinical cohort

11. Benefit and creation of a universal reference
 Inclusion of a universal reference eliminates need to run
normal controls in each batch and allows for single sample
analysis
 Universal reference will be initially derived from FFPE
samples encompassing:
– 11 countries
– 11 states within the United States
– 23 different tissue types
– Block ages ranging from <1 year - 21 years
 The universal reference will be further evaluated over
time and is intended to represent the diversity of samples
analyzed using the platform

12. From array data to copy number, LOH and
somatic mutation results
Array signal data are:
 Normalized against the universal reference
 Corrected for systematic variability (e.g. GC correction)
Next semi “raw data” are provided:
 log2 ratio
 B-allele frequency (BAF)
Data conclusion layer:
 Copy number summary and somatic mutation data
 Uses Affymetrix’ TuScan algorithm to compute the integer CN of the
cancer component (when possible)
 Reports Copy Neutral (and other) LOH segments
 Presence/absence of Somatic Mutation

13. 1
2
Algorithm Output:
% Tumor and ploidy
Integer CN Track for Tumor component
Normal Adjacent
Tissue:
Colorectal Cancer
Site: Liver
% Tumor: Normal
Ploidy = 2
Algorithm Input:
: B-allele frequency
(~signal(B)/(signal(A) + signal(B))
: log2 ratio
1
2
3
3
CN conclusion: A normal male sample

14. For Research Use Only
Algorithm Output:
% Tumor: ~80%
Ploidy: 2
Adenocarcioma
Site: Prostate
An example of a tumor with few aberrations
detection of genome wide copy number aberrations
2 31 4 5 6 7 8 9 10 X11 2 31 4 5 6 7 8 9 10 X11
2 31 4 5 6 7 8 9 10 X11

15. For Research Use Only
Algorithm Output:
% Tumor: ~80%
Ploidy: 2
Adenocarcioma
Site: Prostate
CN=2
CN=2
2 31 4 5 6 7 8 9 10 X11 2 31 4 5 6 7 8 9 10 X11
2 31 4 5 6 7 8 9 10 X11
CN=2
An example of a tumor with few aberrations
detection of genome wide copy number aberrations

16. For Research Use Only
Algorithm Output:
% Tumor: ~80%
Ploidy: 2
Adenocarcioma
Site: Prostate
CN=3
CN=3
CN=3
2 31 4 5 6 7 8 9 10 X11 2 31 4 5 6 7 8 9 10 X11
2 31 4 5 6 7 8 9 10 X11
CN=3
An example of a tumor with few aberrations
detection of genome wide copy number aberrations

17. For Research Use Only
Algorithm Output:
% Tumor: ~80%
Ploidy: 2
Adenocarcioma
Site: Prostate
CN=1
CN=1
CN=1
2 31 4 5 6 7 8 9 10 X11 2 31 4 5 6 7 8 9 10 X11
2 31 4 5 6 7 8 9 10 X11
CN=1
An example of a tumor with few aberrations
detection of genome wide copy number aberrations

18. For Research Use Only
Algorithm Output:
% Tumor: ~90%
Ploidy: 2
Stromal Carcinoma
Site: Stomach
2 31 4 5 6 7 8 9 10 X11 2 31 4 5 6 7 8 9 10 X11
2 31 4 5 6 7 8 9 10 X11
CN=2
CN=2
CN=1
CN=1
CN=1
CN=3
CN=3
CN=3
A slightly more complex tumor
detection of genome wide copy number aberrations
CN=3
CN=2
CN=1

19. An example of a complex tumor
2 31 4 5 6 7 8 9 10 X
2 31 4 5 6 7 8 9 10 X 2 31 4 5 6 7 8 9 10 X
Algorithm Output:
% Tumor: ~80%
Ploidy: 2
Endometrioid Carcinoma
Site: Ovary

20. 2 31 4 5 6 7 8 9 10 X
2 31 4 5 6 7 8 9 10 X 2 31 4 5 6 7 8 9 10 X
Algorithm Output:
% Tumor: ~80%
Ploidy: 2
Endometrioid Carcinoma
Site: Ovary
CN=2
CN = 2
CN = 2
An example of a complex tumor

21. 2 31 4 5 6 7 8 9 10 X
2 31 4 5 6 7 8 9 10 X 2 31 4 5 6 7 8 9 10 X
Algorithm Output:
% Tumor: ~80%
Ploidy: 2
Endometrioid Carcinoma
Site: Ovary
CN = 1
CN=1
CN = 1
CN=1
An example of a complex tumor

22. 2 31 4 5 6 7 8 9 10 X
2 31 4 5 6 7 8 9 10 X 2 31 4 5 6 7 8 9 10 X
Algorithm Output:
% Tumor: ~80%
Ploidy: 2
Endometrioid Carcinoma
Site: Ovary
CN = 3
CN = 3
CN = 3
CN = 3
An example of a complex tumor

23. 2 31 4 5 6 7 8 9 10 X
2 31 4 5 6 7 8 9 10 X 2 31 4 5 6 7 8 9 10 X
Algorithm Output:
% Tumor: ~80%
Ploidy: 2
Endometrioid Carcinoma
Site: Ovary
CN = 2
Copy
Neutral
LOH
CN = 2
Copy
Neutral
LOH
An example of a complex tumor

24. 2 31 4 5 6 7 8 9 10 X
2 31 4 5 6 7 8 9 10 X 2 31 4 5 6 7 8 9 10 X
Algorithm Output:
% Tumor: ~80%
Ploidy: 2
Endometrioid Carcinoma
Site: Ovary
CN = 4
CN=4
+LOH
CN = 4
CN=4
+LOH
An example of a complex tumor

25. AGENDA
For Research Use Only
Introduction of the OncoScan FFPE Assay Kit
in 2H 2013 represents a very significant new
capability
Why?
Only solution for solid tumor genome wide copy
number profiling of a typical clinical sample
offering results in 48 hours

26. For Research Use Only
Large And Growing List Of Actionable
Copy Number Changes!
MD Anderson 2012– 200 genes actionable for SM
MD Anderson 2013- Same 200 genes now actionable for CN as well!
ABL1 BAP1 CEBPA EP300 FGFR1 GNAS KDM6A MPL NOTCH4 PIK3CG RB1 TGFB1
AKT1 BRAF CHEK2 EPHA3 FGFR2 HNF1B KIT MSH2 NPM1 PIK3R1 RET TGFBR2
ALK BRCA1 CPAMD8 ERBB2 FGFR3 HRAS KRAS MSH6 NRAS PKHD1 RUNX1 TNFAIP3
APC BRCA2 CREBBP ERBB3 FGFR4 IDH1 LAMA1 NAV3 NSD1 PPP1R3A RUNX1T1 TOP1
ARAF CARD11 CSMD3 ERCC3 FLT3 IDH2 LPHN3 NF1 PALB2 PPP2R1A SETD2 TOP2A
ARID1A CASP8 CTNNB1 ERCC4 FOXL2 IKZF1 LRP1B NF2 PAX5 PRDM1 SMARCB1 TP53
ASXL1 CBL CYLD ERCC5 GABRB3 IRS1 MAP2K4 NFKB2 PBRM1 PTCH1 SMO TSC1
ATM CDH1 DNMT3A ETV5 GATA1 JAK1 MEN1 NOTCH1 PDGFRA PTEN SOS1 TSC2
ATR CDH11 ELN EZH2 GATA3 JAK2 MLH1 NOTCH2 PDGFRB PTPN11 SYK TSHR
AURKA CDK6 EML4 FBXW7 GNAQ JAK3 MLL3 NOTCH3 PIK3CA RAD51 TET2 VHL
Partial List Of MD Anderson 200 genes

27. For Research Use Only
Large and growing List of actionable
copy number changes
Genes Amplification Deletion
1 ALK
2 EGFR
3 FGFR1
4 FGFR2
5 KRAS
6 MET
7 PTEN
8 AKT1
9 AXL
10 FGFR3
11 ERBB2
12 MAP2K1
13 PIK3CA
14 PDGFRB
15 TP53
CRUK 1st List – Only SM , no CN
CRUK 2nd List- 15 CN Genes