1. Diagnostic Capabilities Beyond the Tumor Cell Genome Gregory J. Tsongalis, PhD, HCLD Professor of Pathology Director, Molecular Pathology Geisel School of Medicine at Dartmouth Dartmouth Hitchcock Medical Center Norris Cotton Cancer Center Lebanon, NH gregory.j.tsongalis@hitchcock.org

2. Outline Technology and Personalized Medicine miRNAs The liquid biopsy Beyond that…….

3. Technology and Therapeutics as Drivers of Medical Care Basic Science Clinical Medicine Translational Research Clinical Laboratory Medicine TherapeuticsTechnology

4. The Genome Or The Human Karyotype

5. Massively Parallel or Next Generation Sequencing at DHMC MiSeq PGM Sequence by Synthesis

6. Personalized Medicine Shaw A T and Engelman J A JCO 2013;31:1105-1111 (A) Before crizotinib; (B) after 7 weeks of crizotinib.

7. Nucleus Cytoplasm Pri-miRNA Pre-miRNA miRNA:miR NA* duplex Unwinding Drosha DGCR8 Exportin 5 Dicer RISC Ribosome Translational Repression mRNA Cleavage C. L. Bartels and G.J. Tsongalis, Clin Chem, 2009;55: 623-631 Specificity (miRNAs as Biomarkers)

8. miRNA Profiling from Pancreatic FNAs Szafranska, Doleshal, Edmunds, Gordon, Luttges, Mundingv Barth Jr, Gutmann, Siriawinata, Pipas, Tannapfel, Korc, Hahn, Labourier, Tsongalis. Clin Chem 54(10):1716-1724, 2008. 5 16 4 5 * ** Hanoun, Delpu, Suriawinata, Bournet, Bureau, Selves, Tsongalis, Dufresne, Buscail, Cordelier, Torrisani. Clin Chem 56(7):1107-1118, 2010. 24 FFPE matched tumor and normal HNSCC (6 laryngeal, 6 oral tongue) miRs: 21, 205, 12746 upregulated miRs:451, 125b downregulated

9. Issues With The Tissues –Tumor tissue has inherent limitations –Tumor tissue is a single snapshot in time –Selection bias due to tumor heterogeneity –Invasive procedure

10. Resected SpecimenSmall Needle Core Biopsy Exhausted Block A Huge Problem

11. The Liquid Biopsy Cell Free DNA (cfDNA) –Cells undergoing apoptosis or necrosis –Amount of cfDNA correlates with tumor staging and prognosis

13. Xenograft Model Kerrington Smith, MD

14. Promega PlexorHY

15. J. E. Hill, PhD Thayer School of Engineering at Dartmouth Breath-based Diagnostics

17. Conclusions Circulating nucleic acids become future biomarkers of choice More advanced analysis of simpler specimens

18. Molecular Pathology Laboratory and Translational Research Program Samantha Allen Paul Burchard Francine de Abreu, Ph.D. Leanne Cook Betty Dokus Amber Erskine Torrey Gallagher Arnold Hawk Joel Lefferts, Ph.D. Emmeline Liu Rebecca O’Meara Jason Peterson Elizabeth Reader Barbara Soares Heather Steinmetz Laura Tafe, M.D. Scott Turner, PhD Wendy Wells, M.D. Terri Wilson Eric York