Pharmaceutical Aspects of Oligonucleotides

Oligonucleotides with base sequences complementary (antisense) to a specific RNA or DNA offer the exciting potential of selectively modulating the expression of an individual gene. Those molecules are gaining interest of the rapid progress in molecular biology and broad scope of potential applications in treating human diseases. However, oligonucleotides diffuse poorly through the biological barriers, including cell membrane. In addition, they are rapidly degraded in vivo by nucleases. Thus, to improve the administration of these compounds is a real challenge that this book will, in part, focus on. Therefore, particular emphasis will be placed on the developments of the chemistry as well as on the formulation and the controlled delivery of these compounds. Background information concerning intracellular trafficking and tissue distribution/pharmacokinetics will also be given. In addition, the therapeutical aspects will also be given consideration.The target audience are researchers working within academia and industry as well as those in management positions in companies that are involved in the field of oligonucleotide therapy. This book is also vital for postgraduate students who need to be well informed about this currently hot topic.

Patrick Couvreur is Director of the CNRS Unit 'Physico-chimie, Pharmacotechnie, Biopharmacie', Chatyenay-Malabry, France. Claude Malvy is Research Director at CNRS, Institut G. Roussy, Villejuif, France.

Contributors

x

Preface

xii

PART ONE General Features

1

(32)

Mechanisms of Action of Antisense Oligonucleotides

3

(30)

D.M. Tidd

R.V. Giles

Introduction

3

(1)

Cell Proliferation Arrest through Release of Deoxynucleosides

4

(2)

Extracellular Aptameric Effects of Phosphorothioate Oligodeoxynucleotide Analogues

6

(1)

Aptameric Effects of G-quartet Oligonucleotides and Analogues

7

(3)

Immune Stimulation by Oligodeoxynucleotides Containing CpG Motifs

10

(1)

Other Activities of Oligodeoxynucleotides Containing CpG Motifs

11

(1)

Antisense Inhibition of Gene Expression by Steric Block

12

(2)

Antisense Inhibition of Gene Expression through Ribonuclease H-mediated Destruction of Target mRNA

14

(7)

Oligonucleotides in Vivo

21

(1)

Conclusions

21

(12)

Acknowledgements

22

(1)

References

22

(11)

PART TWO Chemical Aspects

33

(76)

Chemistry of Oligonucleotides

35

(44)

J.W. Engels

E. Uhlmann

Introduction

35

(4)

Design of Oligonucleotides

39

(3)

Oligonucleotide Modifications: Synthesis and Properties

42

(17)

Analysis of Oligonucleotides

59

(9)

Conclusion

68

(11)

Acknowledgement

68

(1)

References

68

(11)

The Oligonucleotide Prodrug Approach: The Pro-oligonucleotides

79

(19)

F. Morvan

J.-J. Vasseur

E. Vives

B. Rayner

J.-L. Imbach

Introduction

79

(1)

What Kind of Enzymolabile Group?

80

(3)

First Pro-oligonucleotide Models

83

(3)

Pro-oligonucleotides of the Second Generation

86

(6)

Conclusion

92

(6)

Acknowledgements

94

(1)

References

94

(4)

Peptide Nucleic Acids

98

(11)

P.E. Nielsen

Introduction

98

(1)

Antisense

99

(2)

Antimicrobials

101

(1)

Anti-telomerase

101

(1)

Antigene

101

(2)

Anti-HIV

103

(1)

Pharmacology

103

(1)

Further Developments

103

(6)

Acknowledgement

104

(1)

References

104

(5)

PART THREE Delivery

109

(90)

Peptide-mediated Delivery of Oligonucleotides

111

(17)

E. Vives

B. Lebleu

Delivery Vehicles for the Improved Uptake of Nucleic Acids: a Survey

111

(1)

The Potential of Peptides for Nucleic Acids Delivery

112

(2)

Strategies for the Coupling of Peptides to Oligonucleotides

114

(2)

Poly (L-lysine)-based Delivery Systems

116

(1)

Conjugation to Fusogenic Peptides Allowing Membrane Fusion or Membrane Translocation

117

(6)

Conclusions

123

(5)

Acknowledgements

123

(1)

References

123

(5)

Polymeric Nanoparticles and Microparticles as Carriers for Antisense Oligonucleotides

128

(18)

E. Fattal

P. Couvreur

Introduction

128

(1)

Nanoparticles

129

(8)

In Vitro Stability of ONs Adsorbed onto Nanoparticles

137

(1)

Cell Interactions with ON Loaded Nanoparticles

138

(1)

In Vitro Pharmacological Activity of Oligonucleotide-loaded Nanoparticles

138

(1)

In Vivo Studies with Oligonucleotide Nanoparticles

139

(1)

Microparticles

140

(1)

Conclusion

140

(6)

References

141

(5)

Liposomes for the Delivery of Oligonucleotides

146

(26)

P. Couvreur

C. Malvy

C. Dubernet

E. Fattal

Introduction

146

(1)

Anionic Liposomes

147

(3)

Cationic Liposomes

150

(7)

pH-sensitive Liposomes

157

(3)

Immunoliposomes and Other Molecularly Targeted Liposomes

160

(3)

Fusogenic Liposomes and Proteoliposomes

163

(1)

Conclusions

164

(8)

References

165

(7)

Comb-type Polycation Copolymer for Antigene Strategy and DNA Carrier

172

(27)

A. Maruyama

Introduction

172

(1)

Comb-type Polycations as a Stabilizer for DNA Duplex and Triplex

172

(12)

Comb-type Polycation Copolymers with Cell-specific Polysaccharide Side-chains as cell-specific DNA Carrier

184

(15)

Acknowledgements

192

(1)

References

192

(7)

PART FOUR Biopharmaceutics

199

(42)

Delivery of Antisense Oligonucleotides in Vitro: Experimental Points

201

(12)

G.B. Takle

C.A. Stein

Introduction

201

(2)

Oligonucleotide-Binding Proteins on the Cell Surface

203

(2)

Intracellular Compartmentalization

205

(1)

Oligonucleotide Delivery Reagents -- Practical Considerations

206

(1)

Experimental Antisense: Points to Consider

207

(6)

References

210

(3)

Mechanisms of Transmembrane Transport of Oligonucleotides

213

(13)

R.L. Juliano

Overview of Cellular Uptake of Antisense Oligonucleotides

213

(2)

Permeation of Oligonucleotides across Membranes

215

(2)

Mechanisms of Enhancement of Oligonucleotide Permeation across Membranes

217

(3)

Summary

220

(6)

References

221

(5)

Pharmacokinetics of Oligodeoxynucleotides

226

(15)

A. Gouyette

Introduction

226

(1)

Pharmacokinetics

227

(2)

Chemistry of Oligonucleotides and Formulations

229

(1)

Cellular Pharmacokinetics

229

(1)

Preclinical Pharmacokinetics

230

(4)

Clinical Pharmacokinetics

234

(2)

Conclusions

236

(5)

References

236

(5)

PART FIVE Pharmacological Activity

241

(68)

Antisense as a Novel Therapy for Cancer

243

(20)

B.P. Monia

Novel Approaches for Anticancer Therapy

243

(2)

Antisense Approaches for Cancer

245

(5)

Antisense as a Novel Anticancer Approach against ras

250

(9)

Conclusions and Future Prospects

259

(4)

Acknowledgements

259

(1)

References

260

(3)

Modulation of Inflammatory Processes with Antisense Oligonucleotides

263

(23)

C.F. Bennett

Introduction

263

(1)

ICAM-1, a Case Study

264

(11)

Other Examples

275

(3)

Regulation of Immune Response by Non-antisense Mechanisms

278

(1)

Conclusions

279

(7)

References

280

(6)

Oligonucleotides as Antiparasite Compounds

286

(23)

J.-J. Toulme

Introduction

286

(1)

Design of Antisense Oligonucleotides for Antiparasite Use

287

(3)

Antiparasite Effects of Antisense Oligonucleotides

290

(7)

RNA Structures are Valid Targets for Regulatory Oligonucleotides

297

(5)

Conclusion

302

(7)

Acknowledgements

303

(1)

References

304

(5)

Index

309

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Pharmaceutical Aspects of Oligonucleotides

Summary

Author Biography

Table of Contents

Supplemental Materials