Establishing a Pediatric CNS Biorepository Javad Nazarian, PhD, MS Associate Professor, Integrative Systems Biology George Washington University School of Medicine and Health Sciences Principal Investigator, Children's National Medical Center Center for Genetic Medicine Research (CGMR) Center for Cancer and Immunology Research (CCIR) May 25, 2016

High-quality, data-rich samples are essential for future research High-quality, data-rich samples are essential for future research. But obtaining and storing these samples is not as straightforward as many researchers think. 7 JUNE 2012 | VOL 486 | NATURE | 141

The Need To Biobank at CNHS Stems From Active Research Areas Some of the active areas of research: Medulloblastoma DIPG Immunology / Immunotherapy, Clinical Trials Low grade glioma NF, AML, etc.

Prerequisites and Necessities Recognition of challenges Understanding disease biology is hindered by: Lack of biological specimen Tissue type (frozen, matched FFPE, matched blood, etc.) Clinical annotation Molecular annotation Hypothesis testing and translational research is hindered by Lack of robust preclinical models of the disease Clinical annotation

Rapid Translational Application of Targeted Molecules Collaborative & Translational Vision Neurooncology Neurosurgery Neurology Research Specimen Specimen Specimens: Tumor & Non-tumor, CSF, Blood, Urine In Vivo/Vitro Models Rapid preclinical testing Molecular Testing Genome, mRNA, Proteome Rapid Translational Application of Targeted Molecules

Rapid Translational Application of Targeted Molecules Collaborative & Translational Vision at CNHS Neurooncology Neurosurgery Neurology Research Specimen Specimen Specimens: Tumor & Non-tumor, CSF, Blood, Urine In Vivo/Vitro Models Rapid preclinical testing Molecular Testing Genome, mRNA, Proteome Rapid Translational Application of Targeted Molecules

Rapid Translational Application of Targeted Molecules Collaborative & Translational Vision at CNHS Neurooncology Neurosurgery Neurology Research Specimen Specimen Specimens: Tumor & Non-tumor, CSF, Blood, Urine In Vivo/Vitro Models Rapid preclinical testing Molecular Testing Genome, mRNA, Proteome Rapid Translational Application of Targeted Molecules

Rapid Translational Application of Targeted Molecules Collaborative & Translational Vision at CNHS Neurooncology Neurosurgery Neurology Research Specimen Specimen Specimens: Tumor & Non-tumor, CSF, Blood, Urine In Vivo/Vitro Models Preclinical testing Molecular Testing Genome, mRNA, Proteome Rapid Translational Application of Targeted Molecules

Collaborative & Translational Vision at CNHS Neurooncology Neurosurgery Neurology Research Specimen Specimen Specimens: Tumor & Non-tumor, CSF, Blood, Urine In Vivo/Vitro Models Preclinical testing Molecular Testing Genome, mRNA, Proteome Rapid Translational Application of Targeted Molecules: responders vs non-responders, etc

Collaborative & Translational Vision at CNHS National Collaborative Efforts Postmortem Tumor Banking (DIPG) Clinical Specimen (e.g. PNOC-003)

Coordinating Postmortem tissue collection Physician Study Coordinator Hospice Nurse/Social Worker Neuro Pathologist Funeral home Availability Hours of operation Vested in detailed procedure Alternatives : Diener service Regional path Point of contact with the family Update on patient’s health status Funeral home information Paper work Timing Autopsy site

Non-CNMC Coordinated Autopsy Cases

Outcome: Biorepository Total Number of Subjects In the Biorepository

Biorepository Breakdown: Age and Gender

Biorepository Breakdown: Tumor Type OPG

Biorepository Breakdown: Tissue type Frozen FFPE 20 2.6% 101 13.2% 113 14.4% 24 3.1% 34 4.4% 2 0.3% 474 61.7% CSF, Blood, Urine

Prerequisites and Necessities Recognition of challenges STUDY THE DISEASE Lack of biological specimen Tissue type (frozen, matched FFPE, matched blood, etc.) Clinical annotation Molecular annotation √ TEST HYPOTHESIS  TRANSLATE Lack of robust preclinical models of the disease Clinical annotation

Characterization of DIPG primary cells: DAPI H3K27M Merge Neurosphere DAPI GFAP Olig2 Merge Nestin Sox2 Adherent

Models Using Autopsied Specimen DIPG Xenograft Models Using Autopsied Specimen Murine Pontine tumor Murine Cortical tumor a b c H&E H&E Ki67 MAB1273 Ki67 H3K27me3 H3K27M Kambhampati et. al Sridevi Yadavilli

Pre- clinical models of DIPG ID Tissue Source Mutations (major) Primary Cell Xenograft CNMC 762 Biopsy (PNOC-003) H3F3A K27M/ TP53 A159P/ KDR/ KIT/ PDGFRA Yes SF 10423 H3F3A K27M / PPM1D / TP53 R282W / PIK3R1 K567E N/A CNMC 787 TP53 R110L / ATRX E2279* CNMC 586 Autopsy H3F3A K27M / TP53 / PBRM1 CNMC 739 H3F3A K27M CNMC 675 H3F3A K27M/ PPM1D E525X/ ATRX V1514D

Lessons Learned: DIPG Whole Brain Procurement 1 2 7 3 4 6 5

Lessons Learned: Detailed processing is a essential! Kambhampati et al. 2015

Complexity of Biobanking

FreezerPro database to annotate the biobank

Current Challenges Logistics (e. g. pathology, shipping, etc) Funding Awareness (clinicians & families) Collaborative efforts Uniformity of consent form/process The established biobank has resulted in: Collaborations Better understanding of DIPG biology (>12 peer reviewed publications) Robust preclinical resources

ACKNOWLEDGEMENTS PATIENTS & THEIR FAMILIES CHILDREN’S NATIONAL HEALTH SYSTEM Cheng-Ying Ho, Brian Rood, Lindsay Kilburn Eugene Hwang, Suresh Magge, Roger Packer Pacific Pediatric Neuro-Oncology Consortium Sabine Mueller, Michael Prados, CHOP Mariarita Santi, Adam Resnick, Angela Waanders, Michael Fisher Isabella Kerr Molina Foundation Zickler Family Foundation