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Biologically Inspired Engineering: The Next Technology Wave Don Ingber, MD,PhD Founding Director, Wyss Institute Judah Folkman Professor of Vascular Biology, Harvard Medical School & Boston Children’s Hospital Professor of Bioengineering, Harvard School of Engineering and Applied Sciences Hansjörg Wyss Institute for Biologically Inspired Engineering at Harvard University Convergence of the Sciences Life Sciences Engineering Physical Sciences …boundaries between living and non-living systems are beginning to break down THE BIG CHALLENGE: Current Drug Development Model is Broken Computer Power Doubles Every 18 Months Moore’s Law Number of Medicines Invented HALVES Every 9 Years Eroom’s Law (Moore’s law backwards) (Scannell et al., Nature Rev. Drug Discov. 2012; 11:191-200) The Drug Development Model is Broken: • Testing a single compound can cost > $2 million • Cells cultured in dishes don’t function like in our bodies • Animal studies take years to complete • Innumerable animal lives are lost • Results often don’t predict clinical responses! • Lack of new drugs reaching patients (Scannell et al., Nature Rev. Drug Discov. 2012) Need better lab models that mimic whole human organ function Biomimetic Microsystems • Engineer microchips containing living human cells that reconstitute organ-level functions for drug screening, diagnostic and therapeutic applications • ACCELERATE drug development & REPLACE animal testing Biomimetic Spleen Microchip Manufacturing offers control of features at same size scale of living cells Photolithographic Etching: UV Mask nm to mm Photoresist Silicon Cells A Human Breathing Lung-on-a-Chip (Dan Huh, Wyss Institute; Huh et al., Science 2010) Alveoli (air sacs) Measure Cell INJURY (Reactive Oxygen Species) SMOG Air Pollutants Lung cells on-chip (Nanoparticulates) Nanoparticles Epithelium Endotheliu m Cell Injury Inflammation (only + Breathing Motions) + Breathing Motions WBCs Breathing Increases Nanoparticle Absorption Epithelium Nanoparticles % Absorption Flow Flow Endothelium No Stretching Outflow from Lower Channel + Cyclic Stretching 0 hr 1 hr 4 hr Prediction Tested in Whole Mouse Lung Nanoparticles Flow Flow Epithelium Endothelium % Absorption Nebulizer Ventilation Perfusion Prediction Confirmed in Whole Lung Nanoparticles Flow Flow Epithelium Endothelium % Absorption PC AS AS AS AS AS PC AS Human Disease Model: Pulmonary Edema (‘Fluid on the Lungs’)-on-a-Chip Air Membrane IL-2 (CANCER DRUG) Liquid Day 0 Air Liquid Liquid Meniscus Liquid (Huh et al. Sci. Trans. Med., 2012) IL-2 IL-2 Day 2 Day 3 Liquid Day 4 Effects of IL-2 Cancer Drug on Lung Permeability Lung Vascular Leakage Model Flow Capillary On-Chip IL-2 FITC-inulin Prediction Confirmed In Vivo Predicting Drug Efficacy DRUGInhibitor A TRPV4 Lung DRUG A Inhibitor TRPV4 Flow Capillary IL2 + TRPV4 Inhibitor IL-2 & Drug A FITC-inulin Human Disease Model On-Chip: Pulmonary Edema (‘fluid on the lungs”) • Human Disease Model • Drug Toxicity Model • Drug Efficacy Model (Huh et al., Science 2010 & Sci. Trans. Med. 2012) Beating Heart-on-a-chip (Kit Parker Lab) (Grosberg et al., Lab Chip 2011) Structural and functional quantitation on-chip (Wang et al., Nature Medicine 2014) Modeling Barth Syndrome (with human iPS cells) Physiologial Coupling of Heart & Lung Chips (with Kit Parker at Wyss Institute & SEAS, Harvard) Beating Heart Chip Breathing Lung Chip Physiological Coupling Between Lung & Heart Chips Before addition After addition of lung effluent Doxorubicin Delivery to Lung Chip Suppresses Contractility in Linked Heart Chip Liquid Doxurubicin 20 nM 200 nM 2000 nM Small Airway-On-A-Chip (unpublished work of Kambez Benam & Remi Villenave) ‘Classic’ Lung (Alveolus)-On-a-Chip Top channel Bottom Channel 400um Small Airway-On-a-Chip Small Airway: Diameter < 2 mm Regeneration of Airway Epithelium On-Chip Epithelial cells DAPI Endothelial cells Differentiation of Specialized Human Bronchiolar Epithelial Cells On-Chip Cilia (β-tubulin IV) Mucus (Muc5AC) Beating Cilia in the Airway-on-a-Chip (time-lapse at 1/7th normal rate) Top View Visualization of Mucociliary Transport (Real-Time Imaging) Comparison of Airway Chip to Human Airway ‘Flu-like’ Inflammatory Response Induced On-Chip (Induced using Viral Mimic poly I:C) Chemokine Production Monocyte Recruitment Control + Poly I:C Endothelium Influences Cytokine Response to Viral Mimic Human COPD Patient Lung Responses & Exacerbations are Recapitulated On-Chip TLR4 IL8/CXCL8 Cytokine secretion (pg/ml) % mRNA fold change 400 * 300 200 100 0 Healthy 1.5×104 100 50 0 Healthy COPD Con LPS 4×103 2×103 0 0 Healthy Cytokine secretion (pg/ml) % mRNA fold change 150 M-CSF/CSF1 * 8×103 5.0×103 TLR3 * 1×104 6×103 1.0×104 COPD 200 * 2.0×104 1×104 COPD IP10/CXCL10 ** ** 1×103 Healthy 50 40 30 1×102 20 1×101 10 1×100 0 Healthy COPD COPD RANTES/CCL5 * Con Poly I:C ** Healthy COPD Peristaltic Human Gut-on-a-Chip (Kim et al., Lab Chip 2012 & Integrative Biology 2013) Human Intestine Microfluidic Platform Lumen Intestinal Villi Capillary PNAS, 2007, 104:10295 Human Gut Epithelium (Exposed to Flow + Cyclic Deformation) Gut Chip 24 hr after seeding + Peristaltic-like motions Induction of Intestinal Villi Formation Gut Differentiation Requires Correct Mechanical Cues Restoration of Proliferative Crypts Barrier Function Differentiation Differentiation of all Intestinal Cell Lineages Drug Metabolism Mucus Production Intestinal Barrier Co-Culture with Intestinal Microbiome Probiotic Bacteria (Lactobacillus GG) (Kim et al., Lab Chip 2012) Bar, 50 μm Gut-on-a-chip with Normal Microbiome Top view Side view (8 different probiotic strains) GFP-E. coli, F-actin, Nuclei Mechanically Active Environment + Microbiome Induces Small Intestine Differentiation On-Chip Gene Expression Profile Similarities: (Across >22,000 human genes) Gut Chip Gut Chip + VSL#3 Gut Chip + VSL#3 Gut Chip Transwell Transwell Transwell Gut Chip Gut Chip +VSL#3 0.97 0.98 0.99 1 Ileum Duodenum Jejunum Organ-on-Chip Technology Pipeline • Ongoing projects – – – – – – – – – – – – – – Lung Alveolus Lung Small Airway Heart Liver Small Intestine Large Intestine Kidney Proximal Tubule Kidney Glomerulus Bone marrow Skin Blood-Brain Barrier Cancer Eye …… Integrated Human Body-on-Chips DARPA Multiphysiological Systems Grant (D. Ingber, K. Parker, J. Wikswo & CFDRC) Organ-On-Chip INTERROGATOR Instrument Universal Chip Holder – Automated instrument integrates multiple organs – Designed for ease of use “plug-and-play” approach – Generate data to predict human response X 10 Organ Chips Organ ‘Interrogator’ Prototype Custom Enclosure Interrogator Integrated Inside (12 cartridge capacity) Final Prototype Personalized Organs on Chips (from individuals to populations) Emulate Inc. was recently founded to commercialized the Automated Instruments and Vascularized Microfluidic Organ Chips that have emerged from our research effort Disclosure Statement of Financial Interest: I hold equity in Emulate Inc. & chair it’s Scientific Advisory Board Programmable Nanomaterials Platform • Create ‘smart’ medical nanotechnologies • From Implantable Devices to INJECTABLE MEDICAL DEVICES Bioinspired Drug Delivery for Cardiovascular Medicine Vascular Blockage is Leading Cause of Death Platelets ‘Activated’ by Shear Stress •Heart Attack •Stroke •Pulmonary Embolism •Atherosclerosis •Coronary Spasm •Peripheral Vascular Disease •… Shear-Targeted Drug Delivery (Korin et al., Science 2012) Shear-induced platelet activation Shear Stress Synthetic ‘Platelet Mimetics’ Shear Stress Vascular Blockage-Targeted Drug Delivery Shear-Targeted Drug Delivery (Korin et al., Science 2012) Shear-induced platelet activation Shear Stress Synthetic ‘Platelet Mimetics’ Shear Stress Vascular Blockage-Targeted Drug Delivery Targeting of a Clot-Busting Drug (tPA: tissue plasminogen activator) Removal of Artificial Clots using Shear-Targeted tPA Nanotherapeutics 0 min 1 min 60 min In Vitro Study Clinical Responses in an Animal Model In Vivo (effective at 1/100th injected clinical dose of free tPA) Targeting Pulmonary Embolism Pulmonary Embolism Model Significant Increase in Survival SA-NT (80% Survived) Control (100% Dead) Embolus Released Nanoparticles Pathogen-Targeted Technology SEPSIS: • A major killer worldwide with few treatment options • >30% mortality even with best antibiotic therapy and ICU care • Bloodstream infections occur in ~10 % percent of hospital patients, but cause of ~50% of all U.S. hospital deaths (May 18, 2014/Drugs.com) • More than $19B spent in U.S. on treatment Increasing Resistant Microbes New Antibiotic Approvals # New Antibacterial Agents Rapid Disease Progression 20 15 10 5 0 a b ‘Biospleen’ Dialysis-Like Sepsis Therapeutic Device MBL CRD MBL Magnetic nanobead neck Collagen helix IgG1 Fc Artificial Spleen Micro-architecture Blood FcMBL Cleansed (90 kDa) Blood MBL (650Contaminated kDa) Magnetic Opsonins arterial Magnetic bead Magnetic Beads Opsonized Pathogens Vasculature E.coli sinusoid ΔB ΔB Sinusoid ΔB External Magnets S.aureus c venous Collection Fluid Magnetically Separated Pathogens & Beads Venous system Sinusoid slits Lymphoid Discard'Out' Discard out Saline'In' in Saline Magnets c MAGNET' Venous Arterial red-pulp cord Stress fiber Waste ΔB! ΔB' ΔB! Sinusoid Arterial Magne+c' Magnetic Opsonins' Endothelial cells d Sep+c'Blood'In' Septic blood in opsonin in Cleansed'Blood'Out' Cleansed blood out Peristaltic pump Saline in Magnetic opsonins Anesthesia Saline out for ANALYSIS TESTJugular SAMPLE IN catheters Magnets SAMPLE OUT Static mixer 51 Incubation a b ‘Biospleen’ Dialysis-Like Sepsis Therapeutic Device MBL CRD MBL Magnetic nanobead neck Collagen helix IgG1 Fc Artificial Spleen Micro-architecture Blood FcMBL Cleansed (90 kDa) Blood MBL (650Contaminated kDa) Magnetic Opsonins arterial Magnetic bead Magnetic Beads Opsonized Pathogens Vasculature E.coli sinusoid ΔB ΔB Sinusoid ΔB External Magnets S.aureus c venous Collection Fluid Magnetically Separated Pathogens & Beads Venous system Sinusoid slits Lymphoid Discard'Out' Discard out Saline'In' in Saline Magnets c MAGNET' Venous Arterial red-pulp cord Stress fiber Waste ΔB! ΔB' ΔB! Sinusoid Arterial Magne+c' Magnetic Opsonins' Endothelial cells d Sep+c'Blood'In' Septic blood in opsonin in Cleansed'Blood'Out' Cleansed blood out Peristaltic pump Saline in Magnetic opsonins Anesthesia Saline out for ANALYSIS TESTJugular SAMPLE IN catheters Magnets SAMPLE OUT Static mixer 52 Incubation Generic Pathogen Capture Technology Biological Inspiration: HUMAN OPSONINS • Natural blood proteins • Molecular components of the Innate Immune system • Many are LECTINS that bind to surface carbohydrates on pathogens that are not commonly found on human cells Engineered Human Opsonin (FcMBL) Human Mannose Binding Lectin (MBL): Binds diverse pathogens, targets them for phagocytosis, stimulates immune response MBL Carbohydrate Recognition Domain (CRD) neck Engineered FcMBL: •Binds diverse pathogens •Lacks complement activation & coagulation domains • Easy purification MBL CRD MBL neck Fc IgG Complement & coagulation activation Collagen helices FcMBL on magnetic nanobead Broad Pathogen-Binding Spectrum S. aureus 1 mm 1 mm (128 nm beads) C. albicans E. coli 1 mm FcMBL Opsonin: Bind > 90 Pathogens + Toxins Fungi Gram negative Gram positive Viruses Parasites Aspergillus spp Acinetobacter baumanii* Burkholderia cepacia, Bacterioides fragilis Bacillus subtilis “Dengue” Cryptosporidium Blastomyces Chlamydia trachomatis, Clostridium neoformans, Clostridium difficile, perfringens “Ebola” Leishmania Candida, albicans, glabrata, guilliermondii, krusei, parapsilosis, tropicalis Escherichia coli* Enterobacter aerogenes* Enterobacter cloacae (abx)* EBV Malaria Cryptococcus Haemophilus inf b Listeria monocytogenes* Hep B, C Schistosoma Fusarium spp. Helicobacter pylori Mycobactrium avium HIV Trypanosoma Mucor spp. Klebsiella oxytoca, K. pneumonia (abx)* Mycobacterium tuberculosis, leprae HSV 1, 2, CMV Saccharomyces Legionella pneumophila Nocardia farcinica, P. acnes “ Influenza A” Pneumocystis jirovecii (carinii) Neisseria meningitidis B,C, Neisseria gonorrhoeae Staphylococcus aureus MSSA*, MRSA* S. epidermidis*, Marburg LPS Pseudomonas aeruginosa*, S. marcescens (abx)* S. maltophilia* Streptococcus pyogenes Group A, RSV LTA SARS-CoV Ricin Salmonella typhi, paratyphi, typhimurium (abx)* Shigella flexneri Mycoplasma Yersinia pseudotuberculosis M. pneumoniae, M. hominis, M. orale West Nile Toxins Rat Sepsis Model Peristaltic pump Catheterized Rat Microfluidic Separator Syringe pump injecting MBL1 beads & Heparin Static mixer Spleen-on-a-Chip Cleansing of Blood Pathogens in Rats Spleen chip + no beads Control Spleen Chip • Rats treated with mf-DLT device had consistently lower levels of S. aureus pathogens at all time points (n=5) (p<0.001) • 90% of pathogens removed from human whole blood within 1 hr (n=4) Cytokine Levels (pg/mL) Reduction of Cytokine Levels In Vivo 120 SAIP No treatment 100 80 SAIP Biospleen 60 40 20 0 GM-CSF IFN-G IL-1A IL-4 IL-6 Blood Cleansing Increases Survival in Rats with Endotoxemia LPS intensity (a.u.) (Kang et al., Nature Medicine 2014) Survival (%) 100 50 0 0 1 2 3 4 5 0.8 0.6 0.4 0.2 0.0 Lung Time (h) • • LPS (1.4 x107 EU/mL) injected IV into rats (n=6) 100% of control animals die by 4.5 hrs vs. 85% survival LPSDLT Device with therapy − + + − + Biospleen + Inspiration from the Non-Medical World Non-Stick Slippery Liquid-Infused Porous Surfaces (SLIPS) (Joanna Aizenberg Lab) Winner of 2012 R&D Technology Award Medical SLIPS Tethered Liquid Perfluorocarbon (TLP) Fig. 1 A Liquid Perfluorocarbon (LP) Blood Blood Substrate Substrate q Sliding Angle Tethered Perfluorocarbon (TP) t=5 s are silanized with Tethered Perfluorocarbon (TP), then B FDA approved indwelling devices 0 sec 2 sec 5 sec 1. sterilized and stored > 1 year 1. Prior to use, an FDA-approved Liquid Perfluorocarbon (e.g. Perfluorodecalin; PFD) is added 1. Control The TP retains the LP and repels blood TLP Coating for Medical Devices (Leslie et al., Nature Biotechnology 2014) • TLP works on SMOOTH FDA-approved Medical Materials • TLP-treated arteriovenous (AV) shunt functioned for 8 hrs in a pig without heparin Arterial PU Venous PC PVC It’s Just the Tip of the Wave… Injectable Engineered Heart Valve Kit Parker (Wyss Institute Harvard) and Simon Hoerstrup (U. Zurich/Wyss Translation Center) - 290,000 heart valve replacements annually worldwide Bioprostheses Mechanical Valves ( 3x by 2050; Yacoub et al., 2005) - Current replacements materials are suboptimal lack of growth / regeneration surface thrombogenicity / Immunogenicity Engineer scalable nanofiber based, seamless semilunar cardiac valves using automated RJS Fast, inexpensive and regenerative Easily manufacturable / off-the-shelf Biomimetic mechanics in vitro / in vivo in situ remodeling & regeneration Rotary Jet Spinning (RJS) Robbins et al., 2009 Robbins et al., 2009 Biomimetic Mechanics In Vitro Injectable Engineered Heart Valve (JetValve) Kit Parker (Wyss Institute Harvard) and Simon Hoerstrup (U. Zurich/Wyss Translation Center) After implantation Before implantation Follow up 1 MONTH in vivo: aSMA JetValve 3 2 2 2 1 1 MG Von Kossa 3 H&E The thin and fully loaded heart leaflet shows cellular infiltration, remodeling, and endothelialization after 1 month in vivo implantation at the pulmonary valve position Bioinspired Robotics • Develop biologically inspired robots that move and adapt like living organisms • Create AUTONOMOUS SELF-ASSEMBLING MACHINES Normal Embryogenesis (Karlstrom & Kane; http://zfin.org) Bioinspired Programming Algorithms Kasper Stoy (U. Southern Denmark & Wyss Visiting Scholar) & Radhika Nagpal (Wyss Institute) Bioinspired Robotics Rob Wood Radhika Nagpal L. Mahadevan Kit Parker George Whitesides Conor Walsh Autonomous Construction Autonomous Control Automated “POP-UP” Manufacturing Wyss Core Faculty Donald Ingber Kevin Parker Joanna Aizenberg George Church 2008 L. Mahadevan Pamela Silver James Collins Peng Yin Radhika Nagpal David Mooney David Edwards William Shih Conor Walsh Ary Goldberger Robert Wood George Whitesides Jennifer Lewis Neel Joshi 2009 2012 2013 ATT Staff Add Broad Industrial Experience: CELL BIOLOGY ENGINEERING PHARMACOLOGY VIROLOGY MODELING MEDICINE PHYSIOLOGY IMMUNOLOGY MATERIALS SOFTWARE wyss.harvard.edu