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Advanced Delivery Editors
Advanced Delivery
Cheng
and Therapeutic
Mahato
Applications of RNAi and Therapeutic
TA
Applications of RNAi
Editors Kun Cheng, School of Pharmacy, University of h
d
Missouri-Kansas City, USA e
v
r
a
Ram I. Mahato, Department of Pharmaceutical Sciences, a
n
University of Tennessee Health Science Center, USA p
c
e
e Editors
u
d
t
RNA interference (RNAi) is a process in living cells whereby small double stranded i Kun Cheng
cD
RNA interferes with the expression of specifi c genes with complementary nucleotide
sequence. Like many nucleic acid-based therapies, RNAi has great potential in treating Ae Ram I. Mahato
various life-threatening diseases. However, the poor stability and cellular uptake pl
i
of RNAi molecules remain considerable barriers to their effi cient delivery which is v
p
paramount to a successful therapy. This book provides a comprehensive introduction e
l
to RNAi delivery, then goes on to discuss different delivery strategies, and concludes ir
c
with current applications in various diseases. It covers: y
a
• Mechanisms, Barriers, and Analysis of RNAi Delivery ta
• Nonclinical Safety Assessments and Clinical Pharmacokinetics: A Regulatory Perspective ion
• Bioconjugation of siRNA for Site Specifi c Delivery
nd
• Nanoscale Delivery Systems for RNAi
s
• Environmentally-Responsive Delivery Systems for RNAi
• Light-sensitive RNAi o
• Viral-Mediated Delivery of shRNA and miRNA
f
• RNAi applications in Cancer Therapy, Liver Diseases, Hepatitis B, and Ocular Disease
R
• miRNA as therapeutic agents and targets
N
Advanced Delivery and Therapeutic Applications of RNAi combines the essential
aspects of developing RNAi therapeutics from bench to clinic, and is invaluable for A
researchers working on RNAi, drug discovery and delivery, biomedical engineering,
i
biomaterials, molecular biology, and biotechnology.
Advanced Delivery
and Therapeutic
Applications of RNAi
Advanced Delivery
and Therapeutic
Applications of RNAi
Edited by
KUN CHENG
School of Pharmacy, University of
Missouri-Kansas City, USA
and
RAM I. MAHATO
Department of Pharmaceutical Sciences,
University of Tennessee Health Science
Center, USA
Thiseditionfirstpublished2013
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ISBN:9781119976868(13digits)
Setin10/12pt,TimesbyThomsonDigital,Noida,India.
Idedicatethisbooktomyparents,Mr.GuangxiongChengandMrs.PingqingXu,
mywifeLizhiSun,mychildrenDanielandJessicafortheirloveand
continuoussupport,andtomymentorswhohaveinspiredmetopursue
acareerinscience.
KunCheng
IdedicatethisbooktomywifeSubhashini,mychildrenKalikaandVivekfortheir
loveandsupport;mylatemotherSarswatiforbelievinginme;andtomystudents
andmentorswhohavealwayshelpedmeinmyquestforlearning
andinachievinghighergoals.
RamI.Mahato
Contents
Preface xvii
Contributors xix
AbouttheEditors xxiii
Part1 IntroductionandBasicsofRNAi 1
1 MechanismsandBarrierstoRNAiDelivery 3
JiehuaZhouandJohnJ.Rossi
1.1 Introduction 3
1.2 BarrierstoSystemicRNAiDelivery 5
1.3 RationalDesigntoImproveRNAiEfficacy 6
1.4 ChemicalModificationstoEnhancesiRNAStabilityandReduce
ImmuneResponse 7
1.5 CellularUptakeandIntracellularReleaseofsiRNA 7
1.6 CombinatorialTargetingforTargetedRNAiDelivery 8
1.7 Cell-SpecificAptamer-FunctionalizedNanocarriersforRNAiDelivery 9
1.8 TheClinicalDevelopmentandChallengesofsiRNAsTherapeutics 10
1.9 ConclusionandPerspectives 12
References 12
2 AnalysisofsiRNADeliveryUsingVariousMethodologies 19
YiPei
2.1 Introduction 19
2.2 CheckpointsforAnalyzingsiRNADelivery 20
2.2.1 CirculationCheckpoint 22
2.2.2 OrganorTissueCheckpoint 22
2.2.3 CellularCheckpoint 22
2.2.4 RISCCheckpoint 23
2.2.5 TargetmRNAKnockdown(IndirectCheckpoint) 24
2.2.6 ProteinandOutcome(IndirectCheckpoint) 25
2.2.7 Safety(IndirectCheckpoint) 26
2.3 MethodsforAnalysisofsiRNA 26
2.3.1 GeneralConsiderations 26
2.3.2 Hybridization-Based(Non-Imaging)Methods 28
2.3.3 Non-Hybridization-Based(Non-Imaging)Methods 34
2.3.4 Imaging-Based(Non-Hybridization)Methods 35
2.3.5 Imaging-Based(Hybridization)Methods 37
viii Contents
2.4 CaseStudyforsiRNADeliveryAnalysis 38
References 39
3 ChallengesandOpportunitiesinBringingRNAiTechnologies
fromBenchtoBed 45
SandeshSubramanyaandLanceFord
3.1 Introduction 45
3.2 RNAiMediator(siRNAorshRNA) 45
3.2.1 siRNA 45
3.2.2 Vector-derivedshRNA 47
3.2.3 miRNAs 49
3.3 SafetyIssuesofRNAiMediators 50
3.3.1 ImmuneStimulation 50
3.3.2 RNAiOverexpression 52
3.4 EfficacyofRNAiMediators 52
3.4.1 TherapeuticResponse 52
3.5 RNAiMediatorsinClinicalTrials 53
3.6 Conclusion 54
References 55
4 NonclinicalSafetyAssessmentsandClinicalPharmacokineticsfor
OligonucleotideTherapeutics:ARegulatoryPerspective 63
Shwu-LuanLee,PaulBrown,JianWangandRobertT.Dorsam
4.1 Introduction 63
4.2 UniquePropertiesofOligonucleotide-basedTherapeutics 63
4.3 RegulationofOligonucleotide-BasedTherapeutics 65
4.3.1 SubmissiontotheFDA 65
4.3.2 ReviewProcessforNon-clinicalStudies 67
4.3.3 RegulatoryIssues 74
4.3.4 ClinicalPharmacokinetics 76
4.4 Conclusion 79
Disclaimer 79
Appendix 79
References 80
5 RoleofPromotersandMicroRNABackboneforEfficientGeneSilencing 83
FengLiandRamI.Mahato
5.1 Introduction 83
5.2 PromotersforshRNAExpression 84
5.2.1 ConstitutivePromoters 84
5.2.2 InduciblePromoters 87
5.2.3 SiteSpecificPromoters 93
5.3 miRNA-basedshRNAs 96
