Surgical Pathology Lab of the Future Thomas M. Grogan, M.D. Professor of Pathology, University of Arizona Founder, Ventana Medical Systems, Inc. 28 April 2010 Objective Demonstrate how the next generation of tissue-based diagnostics will inform cancer therapy. 2 2 1
Illustrative Patient Cases A 48 year old with breast carcinoma A 32 year old with breast carcinoma. A 28 year old with gastric carcinoma. A 58 year old with lung carcinoma. 3 A 63 year old with prostate cancer. A 34 year old with neuroendocrine cancer. A 59 year old with an anaplastic glioma. A 61 year old with abdominal lymphoma. 3 The Key Scientific Advances Beyond diagnosis to therapeutics. Beyond single analyte to multiplexing. Beyond protein to gene plus protein assays. 4 Beyond qualitative to quantitative assays. Beyond informatics to cellular informatics. Beyond written reports to electronic patient-centric reports. 4 2
Patient # 1 The Importance of Morphologic Context 48 year old with 2 centimeter mass left breast. Histology (H&E) = invasive ductal breast carcinoma. 5 Immunohistochemistry (IHC): Triple negative (ER -, PR -, Her2 - ) PCR: ER Postive Resolution by microscopy: ER on normal cells, not cancer 5 Invasive Breast Carcinoma (CA) NL 6 CA H&E Estrogen Receptor 6 3
Chemistry on a Glass Slide Microarrays ISH FISH 7 IHC Immuno FITC 7 Chemistry in Context 8 Intact Extracted 8 4
Utility of Slide-Based Tissue Chemistry Chemistry of an intact biopsy. Chemistry on a glass slide. Biomarkers in microanatomic content. Chemistry of lesional tissue. 9 9 Breast Cancer Heterogeneity HER-2 10 1+ 2+ 3+ 0 10 Objective 4x 5
Patient # 2 The Importance of Next- Generation Molecular Assays 32 year old female with 11 right breast mass. Surgical biopsy (H&E): invasive breast cancer. Right mastectomy 85 mm mass: DCIS and invasive Grade: 3 Margins: Free Axillary dissection: 1/23 LN positive 11 Breast Cancer Symphony H&E ER (SP1) PR (1E2) 12 HER2 (4B5) HER2 SISH Ki-67 (30-9) 12 6
Mis-Diagnosis of HER2 Status 13 Sources: GNE market research, Perez et al., 2004 and personal communication, Paik et al., 2002, Reddy et al., 2004 and data on file 13 2007 SISH HER2 Launch 14 14 7
Comparison of Automated Silver Enhanced in-situ Hybridization (SISH) and Fluorescence-ISH (FISH) for Validation of HER2 Gene Status in Breast Carcinoma According to the Guidelines of the American Society of Clinical Oncology and the College of American Pathologists 96% concordance of SISH with FISH fulfils ASCO/CAP requirement 15 Dietel, M 1, I. Ellis 2, H. Höfler 3, H. Kreipe 4, H. Moch 5, A. Dankof 1, K. Kölble 1 and G. Kristiansen 1 1 Institute of Pathology, Charité-University Medicine Berlin, Berlin, Germany 2 Division of Pathology, School of Molecular Medical Sciences, University of Nottingham, Department of Histopathology, City Hospital NHS Trust Hospital, Nottingham, United Kingdom 3 Institute of Pathology, Technische Universität München, Munich, Germany 4 Institute of Pathology, Medizinische Hochschule Hannover, Hannover, Germany 5 Institute of Surgical Pathology, Department Pathology, University Hospital Zurich, Zürich, Switzerland Virchow Arch Path 451(1):19-25, 2007 15 A Multicenter Study Comparing Silver In Situ Hybridization with FISH Overall, 94.8% of cases were successfully 16 analyzed by SISH across 6 participating labs. Concordance for diagnosis of HER2 amplification by SISH compared with FISH was high (96%) overall. Bartlett, Campbell F, Ibrahim M, Di Palma, Miller K. Am J Clin Path, Oct; 132(4): 514-20 2009. 16 8
Ultra-sensitive Detection of Nucleic Acid Targets by Silver In Situ Hybridization (SISH) Silver, Hydroquinone, & H 2 O 2 HRP-conjugated Goat Anti-Rabbit Antibody 17 Rabbit Anti-DNP Antibody DNP-labeled DNA Probe Gene Target Enzyme catalyzes reduction of silver ions to metallic silver: metal nanoparticles deposit at the site of the target hybridized to the probe. 17 Breast Tissues Stained for HER2 Gene and Chromosome 17 Centromere (CEN17) 18 Normal HER2 Gene Amplified HER2 Gene Breast Cancer HER2 Gene Chromosome 17 Centromere (CEN17) Objective 100X 18 9
ER: SP1 vs 6F11 SP1 6F11 19 19 20 20 10
Patient # 3 The Importance of Automation 28 year old woman with metastatic gastric carcinoma overexpression Her2neu. Treated post-surgery with 21 trastuzumab (anti-her2) with oxaliplatin. 4 year survival with metastatic disease controlled. Gastric Cancer 8:249-252, 2005 21 Gastric Carcinoma HER2 Expression 22 H&E PROTEIN GENE 22 11
Incidence of Gastric Carcinoma 23 below 14 15 to 20 21 to 30 above 31 Incidence per 200,000 people 23 Gastric and Breast Cancer Incidence by Country 24 Impact per 200,000 people 24 U 12
Standardization of Molecular Pathology 25 Automated platform for portability. 25 Patient # 4 The Importance of Combining Testing Modalities 58 year old WM presented with Hemoptysis. 26 Chest x-ray revealed a 3 cm left lower lung mass. Resection revealed well differentiated adeno carcinoma. 26 13
Personalized Medicine Concept 27 One Drug Fits All = will benefit from therapy = will not benefit from therapy Companion Diagnostic Predictive Biomarker Theranostic The Right Drug for the Right Patient 27 Personalized Medicine in the Making: Lung Cancer and EGFR Tyrosine Kinase Inhibitors EGFR Mutations, Copy#, IHC Primary Resistance: KRAS Mutation NSCLC Patients Erlotinib/Gefitinib 28 Responder Initial Response 1. EGFR T790M Mutation: Irreversible EGFR TKIs (e.g. HKI-272) Non-responder Acquired Resistance 2. MET Amplification: EGFR TKI + MET TKI 3. IGF1R Activation: EGFR TKI + IGF1R TKI Mechanism-Specific Treatment of Resistance: Rational Drug Combinations Disease Recurrence (1-2 years) 28 14
EGFR and KRAS Mutations Clinically relevant for response to EGFR inhibitors, eg Tarceva 29 EGFR mutations observed in ~77% responders vs. 7% refractory cases ~45% deletions in exon 19 ~40 45% L858R mutation in exon 21 ~10% of mutations in exons 18, 20 PLoS Med 2: e13 (2005); Nature Rev Cancer 7:169 (2007); Carcinogenesis 28:1851 (2007); Biochim Biophys Acta 1756:97 (2005) KRAS mutations correlate with non-response Codons 12, 13, and 61 Up to 20-30% of lung, 30-50% colorectal, 90% of pancreatic cancers 29 Lung Carcinoma EGFR 30 HER2 IGF1R Ki-67 Survivin 30 15
The Surgical Pathology Lab of the Future 31 31 Heterogeneity Predicts Partial Response 32 H&E EGFR Gene & Protein EGFR Gene EGFR Protein Objective 40X 32 16
Companion Diagnostics Identifies targets of therapy and personalizes treatment. Aids extension of treatments to other diseases. Has an expanding menu of markers. 33 Defines heterogeneity and informs add-on therapy. 33 Patient # 5 The Importance of Multiplexing 63 year old with prostate cancer of intermediate risk. Gleason score = 6. PSA level 5. 34 34 17
Prostate Carcinoma by H&E 35 35 Prostate Carcinoma by IHC (PIN4) 36 36 18
Predicting Outcome in Prostate Cancer Gene Fusion in Prostate Cancer 37 37 38 38 19
ERG Break-apart FISH Assay ERG Rearrangement Assay - Interpretation Translocation through Deletion 39 Translocation through Insertion 3 ERG/5 ERG No Rearrangement No re-arrangement Rubin, LaFargue 2008 Translocation through 5 ERG Deletion and 3 ERG Duplication 39 Quantum Dots Highly fluorescent, nanometer-size, single crystals of semiconductor materials. 25nm 40 655 605 585 565 525 nm Size of the nanocrystal determines the color Size is tunable from ~2-10 nm (±3%) Size distribution determines the spectral width 40 20
Patient # 6: The Importance of Companion Diagnostics 59 year old patient with episodic difficulty talking. 41 41 Magnetic Resonance Image (MRI) 42 42 21
Anaplastic Glioma, Grade III by Hematoxylin and Eosin (H&E) 43 43 Epithelial Growth Factor Receptor (EGFR) Gene Status by silver in situ hybridization (SISH) 44 44 22
45 45 46 46 23
Infiltrating Ductal Breast Carcinoma IHC Multiplexing with Haptens ER - 585 Ki67-605 PR 625 P53-655 HER2-705 47 3.50E-01 Spectral Analysis of Breast Carcinoma HER2 Relative Fluorescence Intensity 3.00E-01 2.50E-01 p53 2.00E-01 ER PR 1.50E-01 Ki67 1.00E-01 5.00E-02 47 0.00E+00 520 570 620 670 72 Wavelength (nm) Patient # 7 The Importance of patient safety 48 48 24
Potential for Cross Contamination 49 49 Individual Slide Staining Protects Against H&E Cross Contamination 50 50 25
Cross Contamination Study Tissue Floaters and Contaminants in the Histology Laboratory 51 Platt E, Sommer P, McDonald L, Bennett A, Hunt J. Arch Pathol Lab Med. Vol 133, June 2009. 51 Cross Contamination Survey % of contaminated slides 9 Hospitals 1 Days Run 100 Blank Detector Slides # of tissue frags in bath 1 2 3 4 5 6 7 8 952 Customer Site # of different tissue types in baths Conclude: 1 2 3 4 5 6 7 8 9 Customer Site 1 2 3 4 5 6 7 8 9 Customer Site Dip-dunk Baths typically contain multiple tissue types. Cross contamination was variable (0-12%). Cross contamination included malignant tissues. 52 26
Patient # 8 The Importance of Communication 60 year old female with persistent nausea and vomiting. 53 Prior history: Recurrent urinary tract infections x 3 Clinical Impression: Chronic pyelonephritis Cholecystitis 53 CT SCAN: PRE TREATMENT * 54 54 27
CT GUIDED BIOPSY 55 55 PAN B DLBCL PAN T 56 CD20 CD3 56 28
Role of the Pathologist Interpreter of biomarkers in microanatomic context Integrator of information - Multiparameter data in context 57 Great Communicator - Deliverer of the integrated results - Provider of the patient-centric report = 57 58 58 29
59 59 Conclusion Surgical pathology will be substantially improved based on: - Full automation of tissue 60 based chemistry - Multiparameter protein and molecular assays - An expanding menu of assays linked to etiology and therapeutics - Generation of an integrated report which incorporates multiparameter, multiplexed assays, morphology, image analysis ancillary labs and literature = 60 30
The Ideal Future Multiparameter, multiplex results with morphology, phenotype and genotype with full definition of all therapeutic targets communicated in a same day patient report integrating pathology, radiology, surgery and oncology findings. 61 Results communicated by a multitasking pathologist who serves as diagnostician, lab director and information integrator. 61 31