Clinical utility of cancer biomarkers assessed by virtual microscopy
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1 Clinical utility of cancer biomarkers assessed by virtual microscopy Lina Seiz Clinical Research Unit Head: Prof. Dr. Manfred Schmitt Department of Obstetrics and Gynecology Klinikum rechts der Isar
2 Established prognostic/predictive markers for breast cancer Steroid hormone receptors - estrogen receptor (ER) - progesterone receptor (PR) HER2 upa (urokinase-type plasminogen activator) PAI-1 (plasminogen activator inhibitor-type1) - secreted serine protease and its inhibitor - degradation and remodeling of ECM - chemotaxis, migration/invasion, adhesion, proliferation, angiogenesis
3 upa and PAI-1 are key factors in tumor invasion and metastasis Adhesion Migration PAI-1 Degradation of ECM Invasion Metastasis Vitronectin upa Plasmin Plasminogen Integrins α β upar Internalization / upar-recycling Signal Transduction Stimulation of Proliferation
4 upa and PAI-1 low Clinical impact of upa and PAI-1 in breast cancer Disease-free survival (DFS) Overall survival (OS) Enhanced benefit from adjuvant chemotherapy Chemo N0 Trial Stratification node-negative breast cancer upa/pai-1 upa and/or PAI-1 high Advanced tumor stage Poor clinical outcome (Duffy et al., 1988) Poor prognosis (Jänicke et al., 1989) Randomization Observation adj CTX Observation Prognostic and predictive impact of upa and PAI-1 in node-negative breast cancer validated at the highest level of evidence (LOE-1)
5 Clinical impact of upa and PAI-1 in breast cancer Look et al.; JNCI 2002 (meta-analysis of 8377 primary breast cancer patients) upa and PAI-1 are strong prognostic factors for DFS and OS High upa and/or PAI-1 double risk of disease recurrence or dying from breast cancer
6 Techniques for the determination of upa and PAI-1 in tumor tissues ELISA - commercially available (e.g.femtelle ; American Diagnostica Inc., USA) - international quality assurance - fresh-frozen breast cancer tissue specimens ( mg) - cut-off values: 3 ng/mg (upa), 14 ng/mg (PAI-1) - gold standard Immunohistochemistry (IHC) - fixed, paraffin-embedded tissue sections - increase in upa/pai-1 values in ELISA leads to increasing staining intensity (Jänicke et al., 1990; Reilly et al., 1992) - automated scanning of tumor tissue specimens
7 Immunohistochemical evaluation of upa and PAI-1 protein levels 418 primary breast carcinomas full-face sections tissue micro arrays automated staining Ventana BenchMark XT (Peroxidase-based labeled streptavidin biotin (LSAB) method) monoclonal antibodies directed to upa and PAI-1 digitization Hamamatsu NanoZoomer virtual microscope image analysis quantitative scoring
8 upa protein expression in ductal invasive breast cancer (mouse monoclonal antibody directed to upa, Ventana BenchMark ) scanned with a Hamamatsu NanoZoomer virtual microscope (Hamamatsu Instruments, Herrsching, Germany)
9 upa protein expression in ductal invasive breast cancer (mouse monoclonal antibody directed to upa, Ventana BenchMark ) scanned with a Hamamatsu NanoZoomer virtual microscope (Hamamatsu Instruments, Herrsching, Germany)
10 PAI-1 protein expression in ductal invasive breast cancer (mouse monoclonal antibody directed to PAI-1, Ventana BenchMark ) scanned with a Hamamatsu NanoZoomer virtual microscope (Hamamatsu Instruments, Herrsching, Germany)
11 PAI-1 protein expression in ductal invasive breast cancer (mouse monoclonal antibody directed to PAI-1, Ventana BenchMark ) scanned with a Hamamatsu NanoZoomer virtual microscope (Hamamatsu Instruments, Herrsching, Germany)
12 Automated image analysis SlidePath, Dublin tumor cells quantification ECM histoscore non-tumor cells intensity extent
13 Correlation of upa and PAI-1 values obtained by IHC score with ELISA values Clinical data - patient age - tumor size - nodal status - hormone receptors - HER2 Prognosis
14 People involved Department of Obstetrics and Gynecology Clinical Research Unit, TUM Department of Pathology TUM Prof. Dr. Manfred Schmitt Karin Mengele Kerstin Regner Apostolos Gkazepis Department of Obstetrics and Gynecology TUM PD Dr. Marcus Kremer Dr. Julia Slotta-Huspenina Prof. Dr. Nadia Harbeck Hamamatsu Instruments, Herrsching, Germany Frank Osenberg Dr. Erk Mennenga-Klopp Dr. Catherine Conway Dr. Donal O Shea
upa: From Pilot Studies to Recommendation for Clinical Use Professor Joe Duffy St Vincent s University Hospital,
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