Macromolecular Drug Delivery

M E T H O D S I N M O L E C U L A R B I O L O G YTM

John M. Walker, SERIES EDITOR

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481. Hepatocyte Transplantation: Methods andProtocols, edited by Anil Dhawan and Robin D.Hughes, 2008

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479. Plant Signal Transduction: Methods andProtocols, edited by Thomas Pfannschmidt, 2008

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M E T H O D S I N M O L E C U L A R B I O L O G YTM

MacromolecularDrug DeliveryMethods and Protocols

Edited by

Mattias BeltingLund University, Lund, Sweden

EditorMattias BeltingLund UniversityLund, Swedenmattias.belting@med.lu.sebelting@scripps.edu

Series EditorJohn M. WalkerUniversity of HertfordshireHatfield, HertfordshireUK

ISSN: 1064-3745 e-ISSN: 1940-6029ISBN: 978-1-58829-999-4 e-ISBN: 978-1-59745-429-2DOI 10.1007/978-1-59745-429-2

Library of Congress Control Number: 2008937921

C© Humana Press, a part of Springer Science+Business Media, LLC 2009All rights reserved. This work may not be translated or copied in whole or in part without the written permis-sion of the publisher (Humana Press, c/o Springer Science+Business Media, LLC, 233 Spring Street, NewYork, NY 10013, USA), except for brief excerpts in connection with reviews or scholarly analysis. Use inconnection with any form of information storage and retrieval, electronic adaptation, computer software, orby similar or dissimilar methodology now known or hereafter developed is forbidden.The use in this publication of trade names, trademarks, service marks, and similar terms, even if they arenot identified as such, is not to be taken as an expression of opinion as to whether or not they are subject toproprietary rights.While the advice and information in this book are believed to be true and accurate at the date of going topress, neither the authors nor the editors nor the publisher can accept any legal responsibility for any errorsor omissions that may be made. The publisher makes no warranty, express or implied, with respect to thematerial contained herein.

Cover illustration: From Chapter 12, Figure 3D

Printed on acid-free paper

springer.com

Preface

Macromolecular drugs hold great promise as novel therapeutics of severalmajor disorders, e.g., cancer and cardiovascular disease. However, their use islimited by lack of efficient, safe, and specific delivery strategies. Successfuldevelopment of such strategies requires interdisciplinary collaborations, whichprovide opportunities for the unexpected at the interface between different dis-ciplines. Once available, macromolecular drugs will revolutionize the treatmentof various diseases as well as provide novel diagnostic tools for the benefit ofthe patient.

This volume gives an introduction to macromolecular drug delivery and acomprehensive review of the methods used in the field. In vitro and in vivomodels are described that should be of interest to a broad scientific audience,including experts in biophysics, chemistry, cell biology, and pre-clinical andclinical in vivo studies. Researchers involved in the rapidly expanding field oftargeted therapy strategies, e.g., in oncology, should also find this volume highlyinteresting.

v

ContentsPreface ............................................................................................ vContributors ..................................................................................... ixColor Plates ................................................................................... xiii

1 Developments in Macromolecular Drug Delivery.................................. 1Mattias Belting and Anders Wittrup

2 Synthetic vs. Natural/Biodegradable Polymersfor Delivery of shRNA-Based Cancer Therapies .............................. 11

John S. Vorhies and John J. Nemunaitis3 siRNA and DNA Transfer to Cultured Cells ........................................ 31

Bagavathi Gopalakrishnan and Jon Wolff4 Poly(�-amino esters): Procedures for Synthesis

and Gene Delivery .................................................................. 53Jordan J. Green, Gregory T. Zugates, Robert Langer,

and Daniel G. Anderson5 Systemic Delivery and Pre-clinical Evaluation

of Nanoparticles Containing AntisenseOligonucleotides and siRNAs..................................................... 65

Chuanbo Zhang, Joseph T. Newsome, Rajshree Mewani, Jin Pei,Prafulla C. Gokhale, and Usha N. Kasid

6 Peptide-Based Delivery of Steric-BlockPNA Oligonucleotides.............................................................. 85

Saıd Abes, Gabriela D. Ivanova, Rachida Abes, Andrey A. Arzumanov,Donna Williams, David Owen, Bernard Lebleu, and Michael J. Gait

7 Characterizing Peptide-Mediated DNA Internalizationin Human Cancer Cells ........................................................... 101

Anders Wittrup and Mattias Belting8 Selection and Characterization of Antibodies from Phage Display

Libraries Against Internalizing Membrane Antigens......................... 113Johan Fransson and Carl A.K. Borrebaeck

9 Artificial Membrane Models for the Study of Macromolecular Delivery .. 129Lena Maler and Astrid Graslund

10 Enhanced Delivery of Macromolecules into Cellsby Electroendocytosis .............................................................. 141

Alexander Barbul, Yulia Antov, Yosef Rosenberg,and Rafi Korenstein

vii

viii Contents

11 In Vitro Systems for Studying Epithelial Transport of Macromolecules ..... 151Nicole Daum, Andrea Neumeyer, Birgit Wahl, Michael Bur,

and Claus-Michael Lehr12 Preparation of Macromolecule-Containing Dry Powders for Pulmonary

Delivery............................................................................... 165Kelly S. Kraft and Marshall Grant

13 Macromolecular Delivery Across the Blood–Brain Barrier ................... 175Kullervo Hynynen

14 Positron Emission Tomography (PET)and Macromolecular Delivery In Vivo ......................................... 187

Ludwig G. Strauss and Antonia Dimitrakopoulou-Strauss

Index .......................................................................................... 199

Contributors

RACHIDA ABES • UMR 5235 CNRS, Universite Montpellier 2, Place EugeneBataillon, 34095 Montpellier cedex 5, France

SAıD ABES • UMR 5235 CNRS, Universite Montpellier 2, Place EugeneBataillon, 34095 Montpellier cedex 5, France

DANIEL G. ANDERSON • Department of Biological Engineering, Department ofChemical Engineering, and Center for Cancer Research, MassachusettsInstitute of Technology, Cambridge, MA 02139, USA

YULIA ANTOV • Department of Physiology and Pharmacology, Sackler Facultyof Medicine, Tel-Aviv University, Ramat Aviv, 69978 Tel-Aviv, Israel

ANDREY A. ARZUMANOV • Medical Research Council, Laboratory ofMolecular Biology, Hills Road, Cambridge CB2 2QH, UK

ALEXANDER BARBUL • Department of Physiology and Pharmacology, SacklerFaculty of Medicine, Tel-Aviv University, Ramat Aviv, 69978 Tel-Aviv, Israel

MATTIAS BELTING • Department of Clinical Sciences, Section of Oncology,Lund University, Sweden

CARL A.K. BORREBAECK • Department of Immunotechnology, Lund University,CREATE Health – Center for Translational Cancer Research, Lund, Sweden

MICHAEL BUR • Saarland University, Biopharmaceutics and PharmaceuticalTechnology, Saarbruecken, Germany

NICOLE DAUM • Saarland University, Biopharmaceutics and PharmaceuticalTechnology, Saarbruecken, Germany

ANTONIA DIMITRAKOPOULOU-STRAUSS • Medical PET Group, ClinicalCooperation Unit Nuclear Medicine, German Cancer Research Center,Heidelberg, Germany

JOHAN FRANSSON • Centocor Discovery Research – San Diego, 3210Merryfield Row, San Diego, CA 92121, USA

MICHAEL J. GAIT • Medical Research Council, Laboratory of MolecularBiology, Hills Road, Cambridge CB2 2QH, UK

PRAFULLA C. GOKHALE • Departments of Radiation Medicine andBiochemistry, Molecular and Cellular Biology, Georgetown UniversityMedical Center, Washington, DC 20057, USA

BAGAVATHI GOPALAKRISHNAN • Mirus Bio Corporation, 505 S. Rosa Road,Madison, WI 53719, USA

MARSHALL GRANT • MannKind Corporation, 1 Casper Street, Danbury, CT06810, USA

ix

x Contributors

JORDAN J. GREEN • Department of Biological Engineering, Department ofChemical Engineering, and Center for Cancer Research, MassachusettsInstitute of Technology, Cambridge, MA 02139, USA

ASTRID GRÄSLUND • Center for Biomembrane Research, Department ofBiochemistry and Biophysics, Stockholm University, SE-106 91 Stockholm,Sweden

KULLERVO HYNYNEN • Department of Medical Biophysics, University ofToronto and Sunnybrook Health Sciences Centre, 2075 Bayview Ave,Toronto, ON, Canada

GABRIELA D. IVANOVA • Medical Research Council, Laboratory of MolecularBiology, Hills Road, Cambridge CB2 2QH, UK

USHA N. KASID • Departments of Radiation Medicine and Biochemistry,Molecular and Cellular Biology, Georgetown University Medical Center,Washington, DC 20057, USA

RAFI KORENSTEIN • Department of Physiology and Pharmacology, SacklerFaculty of Medicine, Tel-Aviv University, Ramat Aviv, 69978 Tel-Aviv, Israel

KELLY S. KRAFT • MannKind Corporation, 1 Casper Street, Danbury, CT06810, USA

ROBERT LANGER • Department of Biological Engineering, Department ofChemical Engineering, and Center for Cancer Research, MassachusettsInstitute of Technology, Cambridge, MA 02139, USA

BERNARD LEBLEU • UMR 5235 CNRS, Universite Montpellier 2, Place EugeneBataillon, 34095 Montpellier cedex 5, France

CLAUS-MICHAEL LEHR • Saarland University, Biopharmaceutics andPharmaceutical Technology, Saarbruecken, Germany

RAJSHREE MEWANI • Departments of Radiation Medicine and Biochemistry,Molecular and Cellular Biology, Georgetown University Medical Center,Washington, DC 20057, USA

LENA MÄLER • Center for Biomembrane Research, Department ofBiochemistry and Biophysics, Stockholm University, SE-106 91 Stockholm,Sweden

JOHN J. NEMUNAITIS • Mary Crowley Medical Research Center, Dallas, TX,USA

ANDREA NEUMEYER • Saarland University, Biopharmaceutics andPharmaceutical Technology, Saarbruecken, Germany

JOSEPH T. NEWSOME • Lombardi Comprehensive Cancer Center, and Divisionof Comparative Medicine, Georgetown University Medical Center,Washington, DC 20057, USA

DAVID OWEN • Medical Research Council, Laboratory of Molecular Biology,Hills Road, Cambridge CB2 2QH, UK

Contributors xi

JIN PEI • Departments of Radiation Medicine and Biochemistry, Molecularand Cellular Biology, Georgetown University Medical Center, Washington,DC 20057, USA

YOSEF ROSENBERG • Department of Physiology and Pharmacology, SacklerFaculty of Medicine, Tel-Aviv University, Ramat Aviv, 69978 Tel-Aviv, Israel

LUDWIG G. STRAUSS • Medical PET Group, Clinical Cooperation UnitNuclear Medicine, German Cancer Research Center, Heidelberg, Germany

BIRGIT WAHL • Saarland University, Biopharmaceutics and PharmaceuticalTechnology, Saarbruecken, Germany

DONNA WILLIAMS • Medical Research Council, Laboratory of MolecularBiology, Hills Road, Cambridge CB2 2QH, UK

ANDERS WITTRUP • Department of Clinical Sciences, Section of Oncology,Lund University, Sweden

JON WOLFF • Departments of Pediatrics and Medical Genetics, WaismanCenter, University of Wisconsin-Madison, Madison, WI 53705, USA

JOHN S. VORHIES • Mary Crowley Medical Research Center, Dallas, TX, USACHUANBO ZHANG • Departments of Radiation Medicine and Biochemistry,

Molecular and Cellular Biology, Georgetown University Medical Center,Washington, D.C. 20057, USA

GREGORY T. ZUGATES • Department of Biological Engineering, Department ofChemical Engineering, and Center for Cancer Research, MassachusettsInstitute of Technology, Cambridge, MA 02139, USA

Color Plates

Color Plate 1 Effects of systemically delivered LErafsiRNA on Raf-1expression in normal and tumor tissues. (See discussionand complete caption on p. 66.)

Color Plate 2 Low electric field-stimulated adsorption and uptake ofBSA-FITC by COS 5–7 cells. Cell suspensions of COS5–7 cells in DMEM-H supplemented with 6.8 �M BSA-FITC are subjected to LEF treatment of 20 V/cm (180 �spulse duration at frequency of 500 Hz) for 1 min, fol-lowed by two successive washings with DMEM-H solu-tion. Cells are observed by confocal microscopy. a and bare confocal images of FITC fluorescence taken at cen-tral optical section through the X–Y plane of COS 5–7cells, in b it is superimposed also with phase contrastimage (blue channel). Bar = 5 �m. (a) Imaging immedi-ately after washing with DMEM-H. It can be seen thata large amount of BSA-FITC is adsorbed and some isalready internalized. (b) Imaging after 25 min of incuba-tion at 24 C followed by trypsinization. It resulted in aneffective uptake of BSA-FITC and its removal from theplasma membrane. (c) Distribution histograms of BSA-FITC fluorescence intensity in COS 5–7 cells measuredby flow cytometry 1 h after exposure to LEF and incu-bation at 24◦C: (1) control cells in the absence of BSA-FITC in the medium; (2) control cells in the presence ofBSA-FITC – constitutive uptake; (3) cells exposed to LEFin the presence of BSA-FITC – enhanced adsorption anduptake; (4) same exposed cells as in (3), but further sub-jected to trypsinization before measurement – enhanceduptake only (See discussion on p. 145.)

xiii

xiv Contributors

Color Plate 3 Particle deposition in the lungs. Particles largerthan 10 �m are deposited in the mouth and throat(yellow/orange area) and are swallowed. Particlesbetween 6 and 10 �m are deposited in the upperairways (blue area). Particles between 0.5 and 6 �mare within the respirable range and are depositedin the alveolar region (pink area). Image from:http://www.filterair.info/articles/article.cfm/ArticleID/36856F0C-747B-4E08-B730798D614269E9/Page/1 (Seediscussion on p. 166.)