MammaPrint, the story of the 70-gene profile René Bernards Professor of Molecular Carcinogenesis The Netherlands Cancer Institute Amsterdam Chief Scientific Officer Agendia Amsterdam

The breast cancer treatment dilemma

Of 100 women with breast cancer 3

Only 25% will develop distant metastases 4

But we treat over 75% of all patients with chemotherapy 5

Which means that 50% of all breast cancer patients get a toxic chemotherapy that they did not need! 6

MammaPrint: improved breast cancer diagnosis Low risk High risk High risk Low risk

Identification of the MammaPrint breast cancer prognosis profile 78 breast tumors (‘83-’94) patients < 55 years lymph node negative no adjuvant therapy Unbiased full genome gene expression analysis Prognosis Reporter Genes distant metastases < 5 years (n=34) no distant metastases in at least 5 years (n=44)

MammaPrint Discovery: Van ‘t Veer et al. (2002) Nature 415, 530-536.

MammaPrint prognosis Profile “the 70 gene profile” van´t Veer et al., Nature 415, p. 530-536, 2002 70 significant prognosis genes Tumor samples threshold set at 10% false negatives 91 % sensitivity, 73% specificity

MammaPrint 70 genes are involved in all aspects of tumor cell biology proliferation angiogenesis adhesion to extracellular matrix local invasion intravasation, survival, extravasation proliferation angiogenesis adhesion to extracellular matrix Genes of unknown function (25)

First validation: Van de Vijver et al. (2002) New England J. Med. 347, 1999-2009. 295 patients

MammaPrint:Improved Clinical Management Profiling vs St Gallen selection MammaPrint St Gallen MammaPrint improved prediction and more accurate metastases-free metastases-free MammaPrint: 40% in good profile 60 % in poor profile St Gallen: 15% in low risk 85% in high risk NEJM 347, p1999-2009, 2002

MammaPrint vs St Gallen guidelines Gene profiling: Reduction adjuvant chemotherapy selection Avoiding both over- and undertreatment Improved prognosis prediction NEJM 347, p1999-2009, 2002

Second validation: Buyse et al. (2006) JNCI. 98, 1183-1192. 302 patients

>30% Discordant cases! Independent External Validation: Microarray outperforms all clinical risk assessment High clinical risk Adjuvant on line! N=222 73% Low clinical risk Adjuvant on line! N=80 27% 27% microarray Low risk 35% microarray High risk Under- treatment! Over- Buyse et al JNCI 2006 >30% Discordant cases!

MammaPrint predicts early metastases Time to distant metastasis Hazard Ratios highest in first 5 years JNCI 98, p1183-1192, 2006

Chemotherapy only reduces the early metastases No effect of Chemotherapy Effect of Chemotherapy Courtesy: Peter Ravdin

High reproducibility of microarray experiments (99%) Reproducibility; repeat of the experiment Reproducibility; 2 samples, 7 month period Glas et al, BMC Genomics 2007

MammaPrint: improved breast cancer diagnosis First FDA approved molecular Diagnostic test for cancer 2007 Time Magazine: Invention of the year 2007 20

MINDACT study design ASSESS clinical RISK AND MammaPrint RISK 6000 patients, <70 YRS, 1-3 POS NODES ASSESS clinical RISK AND MammaPrint RISK (adjuvant!online & MammaPrint) 55% 10% 35% BOTH HIGH RISK DISCORDANT RISK BOTH LOW RISK RANDOMIZE decision-making Use clinical risk Use MammaPrint high low high low Chemotherapy No chemotherapy

Recent additional features of the MammaPrint test: Expanded indications: all ages

MammaPrint prognosis in postmenopausal patients 5 10 15 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 KM survival curve: Breast cancer > 55 year old Time(year) Survival Probability Chi2 = 15.38 P = 8.79e-005(wt power = 0) Good prognosis profile Metastases free probability Poor prognosis profile Poor Good 150 patients Overall survival HR 2.3, 0.95 CI [ 1.3-4.1], p=0.0049

Recent additional features of the MammaPrint test: Expanded indications: patients with 1-3 positive lymph nodes

Validation of MammaPrint in patients with 1-3 positive lymph nodes Milan and NKI series 241 patients Milan EIO (B Viale): 1994-1998 NKI series: 1984-1995 80% treated adjuvant chemo and/or hormonal therapy Median Follow Up: Milan: 8.97 years (0-10.9) NKI : 10.4 years (1.6-21.2)

MammaPrint performance in patients with 1-3 positive node(s) N=241breast cancer patients with 1-3 positive lymph node(s) Milan & NKI Multivariate HR 6.59 (95% CI 1.71 to 25.45; p = 0.006)

Breast cancer with 1-3 positive node(s) MammaPrint Distant metastases as first event Multivariate analysis

TargetPrint: Quantification of ER, PR and HER2

MammaPrint: extensive regulatory approvals MammaPrint is the first and only IVDMIA cleared for market by FDA ISO 17025 accredited and CE mark for European market CLIA registered to be able to test US patients CAP Accredited The College of American Pathologists

MammaPrint is only the “tip of the iceberg” of personalized medicine Who needs treatment? Which therapy will work best? 30

Personalized medicine: multiple answers on a single microarray chip Prognosis? Is there a hereditary component? Will tumor respond to Herceptin? Will tumor respond to DNA damaging agents?

Poor quality biomolecules: poor quality biomarkers! RNA integrity Protein integrity FFPE Fresh

Thank you!