DOCSLIB.ORG

Peptide Nucleic Acids, Morpholinos and Related Antisense Biomolecules

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Peptide Nucleic Acids, Morpholinos and Related Antisense Biomolecules MEDICAL INTELLIGENCE UNIT Peptide Nucleic Acids, Morpholinos and Related Antisense Biomolecules Christopher G. Janson, M.D. Departments of Neurosurgery, Neurology and Molecular Genetics Cell and Gene Therapy Center UMDNJ-Robert Wood Johnson Medical School Camden, New Jersey, U.S.A. Matthew J. During, M.D., ScD. Department of Molecular Medicine and Pathology University of Auckland Auckland, New Zealand LANDES BIOSCIENCE / EUREKAH.COM KLUWER ACADEMIC / PLENUM PUBLISHERS GEORGETOWN, TEXAS NEW YORK, NEW YORK USA U.SA PEPTIDE NUCLEIC ACIDS, MORPHOLINOS AND RELATED ANTISENSE BIOMOLECULES Medical Intelligence Unit Landes Bioscience / Eurekah.com Kluwer Academic / Plenum Publishers Copyright ©2006 Eurekah.com and Kluwer Academic / Plenum Publishers All rights reserved. No part of this book may be reproduced or transmitted in any form or by any means, electronic or mechanical, including photocopy, recording, or any information storage and retrieval system, without permission in writing from the publisher, vdth the exception of any material supplied specifically for the purpose of being entered and executed on a computer system; for exclusive use by the Purchaser of the work. Printed in the U.S.A. Kluwer Academic / Plenum PubHshers, 233 Spring Street, New York, New York, U.S.A. 10013 http ://www.wkap .nl/ Please address all inquiries to the Publishers: Landes Bioscience / Eurekah.com, 810 South Church Street, Georgetown, Texas, U.S.A. 78626 Phone: 512/ 863 7762; FAX: 512/ 863 0081 http ://v^^ww.eurekah. com http://www.landesbioscience.com Peptide Nucleic Acids, Morpholinos and Related Antisense BiomoleculeSy edited by Christopher G. Janson and Matthew J. During, Landes / Kluwer dual imprint / Landes series: Medical Intelligence Unit ISBN: 0-306-48230-4 While the authors, editors and publisher believe that drug selection and dosage and the specifications and usage of equipment and devices, as set forth in this book, are in accord with current recommendations and practice at the time of publication, they make no warranty, expressed or implied, with respect to material described in this book. In view of the ongoing research, equipment development, changes in governmental regulations and the rapid accumulation of information relating to the biomedical sciences, the reader is urged to carefully review and evaluate the information provided herein. Library of Congress Cataloging-in-Publication Data Peptide nucleic acids, morpholinos, and related antisense biomolecules / [edited by] Christopher G. Janson, Matthew J. During. p. ; cm. ~ (Medical intelligence unit) ISBN 0-306-48230-4 1. Antisense nucleic acids. 2. Antisense peptides. I. Janson, Christopher G. II. During, Matthew J. III. Series: Medical intelligence unit (Unnumbered : 2003) [DNLM: 1. Peptide Nucleic Acids. 2. Antisense Elements (Genetics) QU 58 P42394 2006] QP624.5.A57P47 2006 612'.015756-dc22 2005016425 This volume is dedicated to Dr. Linda Bartoshuk, who facilitated its planning by assisting one of the editors (Dr. Janson) during his leave from Yale University; and to Dr. Stanley Miller, who first proposed the role of PNA in prebiotic chemical evolution and the ancient history of life on Earth, thereby raising the question if the recent discovery of PNA was in fact a fortuitous rediscovery of our common pre-DNA, pre-RNA origins. CONTENTS Preface xvii Parti Research Applications 1. The Many Faces of PNA 3 Peter E. Nielsen Introduction to PNA 3 PNA Chemistry 3 Cellular Uptake of PNA 4 Antisense Applications 5 Antigene Properties 5 Antimicrobial PNAs 9 Genetic Information Carrier 11 PNA in Diagnostics 12 Prospects 13 2. Modulation of Nucleic Acid Information Processing by PNAs: Potential Use in Anti-Viral Therapeutics 18 Lionel Bastide, Bernard Lebleu and Ian Robbins PNAs as Gene-Modulator Agents 21 Virus Specific Nucleic Acid Processing 25 PNA-Tuning 26 3. Targeted Gene Delivery: The Role of Peptide Nucleic Acid 30 Kenneth W. Liangs Feng Liu and Leaf Huang Targeted Gene Delivery through Polycation/DNA Complex 31 Targeted Delivery of Naked DNA 33 4. Imaging Gene Expression in the Brain with Peptide Nucleic Acid (PNA) Antisense Radiopharmaceuticals and Drug Targeting Technology 38 Ruben]. Boado and William M. Pardridge Mechanism of Action of Antisense Drugs 40 Medical Diagnostic and Therapeutic Applications 41 Functional Genomics 45 Overview of Antisense Molecules and Effective Delivery 45 Brain Drug Targeting Systems A7 Physiological Brain Efflux of IgG 52 Imaging of Brain Gene Expression 52 5. Receptor-Specific Targeting with Complementary Peptide Nucleic Acids Conjugated to Peptide Analogs and Radionuclides 61 Eric Wickstrom, Mathew L Thakur and Edward R, Sauter Synthesis of Probes 68 AppHcations in Cells 71 Applications in Animals 7G Applications in Patients 80 6. Morpholinos and PNAs Compared 89 James E. Summerton Classification of Antisense Structural Types 90 Preparation of Morpholinos and PNAs 91 Properties of Morpholinos and PNAs 93 Applications 106 7. Chemistry of Locked Nucleic Acids (LNA): Design, Synthesis and Bio-Physical Properties 114 ]esper WengeU Michael Petersen, Miriam Frieden and Troels Koch Synthesis of LNA Monomers 114 Solid Phase Synthesis of LNA Oligonucleotides 115 Hybridization Characteristics of LNA Oligonucleotides 118 Hybridization Kinetics of LNA Oligonucleotides 119 Structure of LNA Oligonucleotides 122 DNA and RNA Structure 122 LNA Structure 123 LNAiRNA Duplexes 123 LNArDNA Duplexes 125 LNA:LNA Duplexes 126 a-L-LNA:RNA Duplexes 126 a-L-LNA:DNA Duplexes 126 Implications for RNase H Activity of LNA:RNA and a-L-LNA:RNA Duplexes 127 Stability of LNA and a-L-LNA Modified Nucleic Acids 127 LNA Triplexes 128 8. Recent Applications of RNA Interference (RNAi) in Mammalian Systems 133 Lisa Scherer and John J. Rossi siRNA Design 133 Recent Applications of RNAi 137 Functional Genomics 142 RNAi versus Ribozymes 143 Part II Clinical Applications 9. Peptide Nucleic Acids as Epigenetic Inhibitors of HIV-1 151 Shizuko Set HIV-1 Life Cycle and Potential Molecular Targets 152 Genetic Strategies to Inhibit HIV-1 Replication 153 Peptide Nucleic Acids as Epigenetic HIV-1 Inhibitors 154 Potential Use of PNA Against Other Infectious Pathogens 161 Bio-Delivery of PNA 161 Future Perspectives 163 10. Therapeutic Uses of Peptide Nucleic Acids (PNA) in Oncology 171 Nadia Zaffaroniy Raffaella Villa and Marco Folini Potential of PNAs as Tools for Anticancer Therapeutic Interventions 172 Perspectives 178 11. PNAs as Novel Cancer Therapeutics 181 Luca Mologni and Carlo Gamhacorti-Passerini Biochemistry of Peptide Nucleic Acids 182 PNA as a Biomolecular Tool 183 Antisense and Anti-Gene Properties of PNA 184 Future Directions 189 12. Medicinal Chemistry of Plasmid DNA with Peptide Nucleic Acids: A New Strategy for Gene Therapy 195 Olivier Zelphati, Jiin Feigner^ Yan Wang, Xiaowu Liang, Xiaodong Wang and Philip Feigner Principle of PNA Dependent Gene Chemistry Technology 197 Labeling of Plasmid DNA to Study Gene Delivery Mechanism 198 Transition 199 Overcoming the Barriers to Improve Gene Delivery and Expression 200 PNA-Conjugates for Targeting DNA to Cell Surface Receptors 201 PNA-Peptide Conjugates to Overcome Cell Membrane Barriers 202 Other Potential Applications of PNA Conjugate for Gene Delivery 206 13. Locked Nucleic Acids (LNA) and Medical Applications 212 Henrik 0rumy Andreas Wolter and Lars Kongsbak Biochemistry of LNA 212 LNA in Diagnostics and Genomics Applications 215 LNA in Therapeutic Applications 218 Future Perspectives on LNA 221 14. Peptide Nucleic Acids as Agents to Modify Target Gene E3q)ression and Function 223 Gan Wang and Peter M. Glazer PNA Binding Affinity 224 PNA Binding Specificity 224 Detection of PNA Binding-Induced Transcription in Hela Nuclear Extract in Vitro Transcription System 224 Determination of the Initiation Sites of PNA Binding-Induced Transcription 226 PNA Binding-Generated D-Loops Lead to GFP Gene Expression in Mammalian Cells 228 PNA-Induced Endogenous y-Globin Gene Expression in Human Cells 228 The Correlation between PNA Binding-Generated D-Loops and the Natural Promoter of the Gene in Target Gene Transcription 229 The PNA Length Requirement for Inducing Transcription from the PNA Binding Sites 231 Transcription Components Involved in PNA Binding-Induced Transcription 232 The Limitation of PNA Binding-Induced Target Gene Expression 233 PNAs for Targeted Genome Modification 234 15. Peptide Nucleic Acids: Cellular Delivery and Recognition of DNA and RNA Targets 236 David K Corey Hybridization by PNA: Affinity Isn't Everything 236 Strand Invasion by PNAs 238 Intracellular Delivery of PNAs 239 Applications for PNAs Delivered within Cells 240 16. The Use of PNAs and Their Derivatives in Mitochondrial Gene Therapy 243 PaulM. Smith, Gunther F. Ross, Theresa M. Wardell, Robert W. Taylor, Douglass M. Turnhull and Robert N. Lightowlers The Antigenomic Hypothesis 244 PNA as the Choice of Antigenomic Agent 244 Cellular Uptake and the Problem of Mitochondrial Import 246 PNAs as Antigenomic Molecules - Do They Work? 247 Trouble-Shooting the Antigenomic Approach to mtDNA Disease .... 248 17. Gene Silencing through RNA Interference: Potential for Therapeutics and Functional Genomics 252 David O. Azorsa, Spyro Mousses and Natasha J. Caplen RNAi: An Historical Perspective 252 RNAi: Summary of Mechanism 253 Physiological Role of RNAi and Related Post-Transcriptional Gene Silencing Mechanisms 253 RNAi in Mammalian Cells 255 The Delivery of Triggers of RNAi 256 RNAi as a Functional Genomics Tool 257 RNAi as a Therapeutic Strategy 258 18. Transcriptional
Load more

Recommended publications
  • New Constrained Amino Acids and Peptide Nucleic Acid Building Blocks for the Construction of Bio-Polymers Alexandra Gresika
    New constrained amino acids and peptide nucleic acid building blocks for the construction of bio-polymers Alexandra Gresika To cite this version: Alexandra Gresika. New constrained amino acids and peptide nucleic acid building blocks for the construction of bio-polymers. Other. Université Côte d’Azur, 2018. English. NNT : 2018AZUR4104. tel-02073232 HAL Id: tel-02073232 https://tel.archives-ouvertes.fr/tel-02073232 Submitted on 19 Mar 2019 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. THÈSE DE DOCTORAT Nouveaux synthons contraints de type - amino acides et PNA en vue de l´élaboration de bio-foldamères New constrained amino acids and peptide nucleic acid building blocks for the construction of biopolymers Alexandra Gresika Molécules Bioactives Présentée en vue de l’obtention Devant le jury, composé de : du grade de docteur en Chimie Muriel Amblard, Directrice de Recherche d’Université Côte d’Azur CNRS, IBMM UMR 5247, Université de Dirigée par : Nadia Patino Montpellier Karine Alvarez, Chargée de Recherche Soutenue le : 16.11.2018 CNRS, AFBM UMR 7257, Université Aix- Marseille Mohamed Mehiri, Maître de Conférences, ICN UMR 7272, Université Côte d’Azur Nadia Patino, Professeur des Universités, ICN UMR 7272, Université Côte d’Azur 1 New constrained amino acids and peptide nucleic acid building blocks for the construction of bio-polymers.
    [Show full text]
  • Advances in Oligonucleotide Drug Delivery
    REVIEWS Advances in oligonucleotide drug delivery Thomas C. Roberts 1,2 ✉ , Robert Langer 3 and Matthew J. A. Wood 1,2 ✉ Abstract | Oligonucleotides can be used to modulate gene expression via a range of processes including RNAi, target degradation by RNase H-mediated cleavage, splicing modulation, non-coding RNA inhibition, gene activation and programmed gene editing. As such, these molecules have potential therapeutic applications for myriad indications, with several oligonucleotide drugs recently gaining approval. However, despite recent technological advances, achieving efficient oligonucleotide delivery, particularly to extrahepatic tissues, remains a major translational limitation. Here, we provide an overview of oligonucleotide-based drug platforms, focusing on key approaches — including chemical modification, bioconjugation and the use of nanocarriers — which aim to address the delivery challenge. Oligonucleotides are nucleic acid polymers with the In addition to their ability to recognize specific tar- potential to treat or manage a wide range of diseases. get sequences via complementary base pairing, nucleic Although the majority of oligonucleotide therapeutics acids can also interact with proteins through the for- have focused on gene silencing, other strategies are being mation of three-dimensional secondary structures — a pursued, including splice modulation and gene activa- property that is also being exploited therapeutically. For tion, expanding the range of possible targets beyond example, nucleic acid aptamers are structured
    [Show full text]
  • Guide for Morpholino Users: Toward Therapeutics
    Open Access Journal of Drug Discovery, Development and Delivery Special Article - Antisense Drug Research and Development Guide for Morpholino Users: Toward Therapeutics Moulton JD* Gene Tools, LLC, USA Abstract *Corresponding author: Moulton JD, Gene Tools, Morpholino oligos are uncharged molecules for blocking sites on RNA. They LLC, 1001 Summerton Way, Philomath, Oregon 97370, are specific, soluble, non-toxic, stable, and effective antisense reagents suitable USA for development as therapeutics and currently in clinical trials. They are very versatile, targeting a wide range of RNA targets for outcomes such as blocking Received: January 28, 2016; Accepted: April 29, 2016; translation, modifying splicing of pre-mRNA, inhibiting miRNA maturation and Published: May 03, 2016 activity, as well as less common biological targets and diagnostic applications. Solutions have been developed for delivery into a range of cultured cells, embryos and adult animals; with development of a non-toxic and effective system for systemic delivery, Morpholinos have potential for broad therapeutic development targeting pathogens and genetic disorders. Keywords: Splicing; Duchenne muscular dystrophy; Phosphorodiamidate morpholino oligos; Internal ribosome entry site; Nonsense-mediated decay Morpholinos: Research Applications, the transcript from miRNA regulation; Therapeutic Promise • Block regulatory proteins from binding to RNA, shifting Morpholino oligos bind to complementary sequences of RNA alternative splicing; and get in the way of processes. Morpholino oligos are commonly • Block association of RNAs with cytoskeletal motor protein used to prevent a particular protein from being made in an organism complexes, preventing RNA translocation; or cell culture. Morpholinos are not the only tool used for this: a protein’s synthesis can be inhibited by altering DNA to make a null • Inhibit poly-A tailing of pre-mRNA; mutant (called a gene knockout) or by interrupting processes on RNA • Trigger frame shifts at slippery sequences; (called a gene knockdown).
    [Show full text]
  • De Novo Nucleic Acids: a Review of Synthetic Alternatives to DNA and RNA That Could Act As † Bio-Information Storage Molecules
    life Review De Novo Nucleic Acids: A Review of Synthetic Alternatives to DNA and RNA That Could Act as y Bio-Information Storage Molecules Kevin G Devine 1 and Sohan Jheeta 2,* 1 School of Human Sciences, London Metropolitan University, 166-220 Holloway Rd, London N7 8BD, UK; [email protected] 2 Network of Researchers on the Chemical Evolution of Life (NoR CEL), Leeds LS7 3RB, UK * Correspondence: [email protected] This paper is dedicated to Professor Colin B Reese, Daniell Professor of Chemistry, Kings College London, y on the occasion of his 90th Birthday. Received: 17 November 2020; Accepted: 9 December 2020; Published: 11 December 2020 Abstract: Modern terran life uses several essential biopolymers like nucleic acids, proteins and polysaccharides. The nucleic acids, DNA and RNA are arguably life’s most important, acting as the stores and translators of genetic information contained in their base sequences, which ultimately manifest themselves in the amino acid sequences of proteins. But just what is it about their structures; an aromatic heterocyclic base appended to a (five-atom ring) sugar-phosphate backbone that enables them to carry out these functions with such high fidelity? In the past three decades, leading chemists have created in their laboratories synthetic analogues of nucleic acids which differ from their natural counterparts in three key areas as follows: (a) replacement of the phosphate moiety with an uncharged analogue, (b) replacement of the pentose sugars ribose and deoxyribose with alternative acyclic, pentose and hexose derivatives and, finally, (c) replacement of the two heterocyclic base pairs adenine/thymine and guanine/cytosine with non-standard analogues that obey the Watson–Crick pairing rules.
    [Show full text]
  • Conformation and Polymorphism in PNA-DNA and PNA-RNA Hybrids (Chimera/Duplex/Triple Helix/Molecular Mechanics)
    Proc. Natl. Acad. Sci. USA Vol. 90, pp. 9542-9546, October 1993 Biochemistry Peptide nucleic acid (PNA) conformation and polymorphism in PNA-DNA and PNA-RNA hybrids (chimera/duplex/triple helix/molecular mechanics) ORN ALMARSSON AND THOMAS C. BRUICE Department of Chemistry, University of California at Santa Barbara, Santa Barbara, CA 93106 Contributed by Thomas C. Bruice, July 19, 1993 ABSTRACT Two hydrogen-bonding motifs have been pro- nucleic acids, is their stability toward nucleases. Although the posed to account for the extraordinary stability of polyamide designing of the PNA analogues of DNA has been described "peptide" nucleic acid (PNA) hybrids with nucleic acids. These by the inventors (4), the design rationale does not explain the interresidue- and intraresidue-hydrogen-bond motifs were in- great stability of the PNA hybrids with nucleic acids, as vestigated by molecular mechanics calculations. Energy- exemplified by PNA-strand invasion of double-helical DNA. iinimized structures of Watson-Crick base-paired decameric Our recent molecular mechanics study (5) with the B-helix duplexes of PNA with A-, B-, and Z-DNA and A-RNA poly- structure of the DNA*PNA duplex (dAp)lo'(pnaT)lo [where morphs indicate that the inherent stability of the complemen- (dAp)10 is the decanucleotide of 2'-deoxyadenosine 5'- tary PNA helical structures is derived from interresidue, rather phosphate and (pnaT)lo is the decamer of thyminyl PNA with than from intraresidue, hydrogen bonds in all hybrids studied. lysine amide attached to the C terminus] strongly suggests that Intraresidue-hydrogen-bond lengths are consistently longer the stability of DNA*PNA duplexes is due to interresidue than interresidue hydrogen bonds.
    [Show full text]
  • Dna Enzymes for Peptide-Nucleic Acid Conjugation and for Lysine Methylation
    DNA ENZYMES FOR PEPTIDE-NUCLEIC ACID CONJUGATION AND FOR LYSINE METHYLATION BY CHIH-CHI CHU DISSERTATION Submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Chemistry in the Graduate College of the University of Illinois at Urbana-Champaign, 2017 Urbana, Illinois Doctoral Committee: Professor Scott K. Silverman, Chair Professor Paul J. Hergenrother Associate Professor Douglas A. Mitchell Professor Steven C. Zimmerman Abstract Proteins and RNA are known to be enzymes in nature. These biopolymers have complex secondary and tertiary structures that can enable substrate binding and catalysis. DNA is primarily double-stranded and is not known to be catalytic in nature. However, given the similarities in chemical structure between DNA and RNA, it is reasonable to think that single- stranded DNA can also form complex structures. In fact, artificial DNA enzymes have been identified in laboratories by in vitro selection. The identification of new enzymes favors the use of nucleic acids over proteins for several reasons. First, nucleic acids can be amplified by natural enzymes whereas proteins cannot be amplified in any way. Second, the number of possible sequences is smaller for nucleic acids (4n, where n is the length of the biopolymer) than for proteins (20n). Therefore, selection experiments for identifying nucleic acid enzymes will cover a larger fraction of total sequence space. Furthermore, of the sequence space that is covered, a large portion of nucleic acid sequences can fold into secondary or tertiary structures, whereas most random sequences of proteins often will not fold and thus will aggregate. Between the two nucleic acid polymers, DNA offers additional advantages over RNA because DNA can be directly amplified by polymerases whereas RNA requires an extra reverse transcription step.
    [Show full text]
  • 12.12.2017 1 CHAPTER 1 OLIGONUCLEOTIDES Part 2
    12.12.2017 Canonical nucleobase pairing and alternative base pairs CHAPTER 1 OLIGONUCLEOTIDES Part 2 – noncanonical nucleobases Natural and non-natural base pairs that function in polymerase reactions Self-assembly of whole genes and DNA nanostructures The technology tested by assembly of the kanamycin-resistance gene and growing the bacteria in the environment containing kanamycin after assembly and conversion of that gene. S. Benner et al., Beilstein J. Org. Chem. 2014, 10, 2348–2360. doi:10.3762/bjoc.10.245 1 12.12.2017 AEGIS – Artificially Expanded Genetic Information System AEGIS – Artificially Expanded Genetic Information System ZP CG S. Benner et al., Beilstein J. Org. Chem. 2014, 10, 2348–2360. doi:10.3762/bjoc.10.245 S. Benner et al., J. Am. Chem. Soc., 2011 , 133 (38), pp 15105–15112 AEGIS – Permanent orthogonal nucleobases surviving PCR ACGTZP-DNA crystal structures Electron density presented to the minor groove recognition site by polymerases „ minor groove scanning hypothesis” Error rate 0,2% per a PCR cycle – both removal and incorporation of Z and P the artificial genetic system capable to evolve. Pol: Deep Vent – 2 Z/P, Taq/Phu – 3-4 Z/P 18-mers: 2+2 Z:P pairs B-DNA dZTP ( deprotonated ) at higher pH pairs slightly with G 6 consecutive Z:P A-DNA loss of some Z, but also gain of some new Z mutants. 0,1 nm wider, but otherwise alike G:C pairs S. Benner et al., J. Am. Chem. Soc., 2011 , 133 (38), pp 15105–15112 S. Benner et al., J. Am. Chem. Soc., 2015, 137, pp 6947–6955 2 12.12.2017 Unnatural aminoacid incorporation using a noncanonical base pair (A) The coupled transcription–translation system using the nonstandard codon– anticodon interaction for the site-specific incorporation of 3-chlorotyrosine into the Ras protein.
    [Show full text]
  • Synthesis of Phosphorodiamidate Morpholino Oligonucleotides Using Trityl and Fmoc Chemistry-A New Method Amenable to Automated Synthesizer
    Synthesis of Phosphorodiamidate Morpholino Oligonucleotides Using Trityl and Fmoc Chemistry-A New Method Amenable to Automated Synthesizer Ujjwal Ghosh†a, Jayanta Kundu†a, Atanu Ghosh†, Arnab Das†, Dhriti Nagar¶, Aurnab Ghose¶ and Surajit Sinha†* †School of Applied and Interdisciplinary Sciences, Indian Association for the Cultivation of Science, 2A and 2B Raja S.C. Mullick Road, Jadavpur, Kolkata 700032, India ¶Indian Institute of Science Education and Research, Pune, Maharashtra 411008, India aAuthors have equal contribution *Corresponding author: [email protected] KEYWORDS: Morpholino oligonucleotides, Trityl and Fmoc chemistry, DNA Synthesizer, Activators ETT and Iodine, Antisense reagents. Abstract: Phosphorodiamidatemorpholino oligo- nucleotides (PMO) are routinely used for gene silencing and the recently developed PMO-based drug “Exondys51” has highlighted the importance of PMO as excellent antisense reagents. Howev- er, the synthesis of PMO has remained challeng- ing. Here a method for the synthesis of PMO us- ing either trityl or Fmoc-protected active morpho- lino monomers using chlorophosphoramidate chemistry in the presence of a suitable coupling agent on a solid support has been reported. After screening several coupling agents (tetrazole, 1,2,4-triazole, ETT, iodine, LiBr and dicyanoimidazole), ETT and iodine were found to be suitable for efficient coupling. Fmoc chemistry was not known for PMO synthesis because the preparation of Fmoc-protected chloro- phosphoramidate monomers was not trivial. Synthesis of Fmoc-protected activated monomers and their use in PMO synthesis is reported for the first time. 25-mer PMO has been synthesized using both the methods and vali- dated in vivo in the zebrafish model by targeting the no tail gene.
    [Show full text]
  • An in Vitro Translation, Selection and Amplification System for Peptide Nucleic Acids
    An in vitro translation, selection and amplification system for peptide nucleic acids The Harvard community has made this article openly available. Please share how this access benefits you. Your story matters Citation Brudno, Yevgeny, Michael E Birnbaum, Ralph E Kleiner, and David R Liu. 2009. “An in Vitro Translation, Selection and Amplification System for Peptide Nucleic Acids.” Nat Chem Biol 6 (2) (December 27): 148–155. doi:10.1038/nchembio.280. Published Version doi:10.1038/nchembio.280 Citable link http://nrs.harvard.edu/urn-3:HUL.InstRepos:25275370 Terms of Use This article was downloaded from Harvard University’s DASH repository, and is made available under the terms and conditions applicable to Other Posted Material, as set forth at http:// nrs.harvard.edu/urn-3:HUL.InstRepos:dash.current.terms-of- use#LAA NIH Public Access Author Manuscript Nat Chem Biol. Author manuscript; available in PMC 2010 August 1. NIH-PA Author ManuscriptPublished NIH-PA Author Manuscript in final edited NIH-PA Author Manuscript form as: Nat Chem Biol. 2010 February ; 6(2): 148–155. doi:10.1038/nchembio.280. An In Vitro Translation, Selection, and Amplification System for Peptide Nucleic Acids Yevgeny Brudno, Michael E. Birnbaum, Ralph E. Kleiner, and David R. Liu* Department of Chemistry and Chemical Biology and the Howard Hughes Medical Institute, Harvard University, 12 Oxford Street, Cambridge, MA 02138 Abstract Methods to evolve synthetic, rather than biological, polymers could significantly expand the functional potential of polymers that emerge from in vitro evolution. Requirements for synthetic polymer evolution include: (i) sequence-specific polymerization of synthetic building blocks on an amplifiable template; (ii) display of the newly translated polymer strand in a manner that allows it to adopt folded structures; (iii) selection of synthetic polymer libraries for desired binding or catalytic properties; and (iv) amplification of template sequences surviving selection in a manner that allows subsequent translation.
    [Show full text]
  • Multifunctional Delivery Systems for Peptide Nucleic Acids
    pharmaceuticals Review Multifunctional Delivery Systems for Peptide Nucleic Acids Stefano Volpi , Umberto Cancelli, Martina Neri and Roberto Corradini * Department of Chemistry, Life Sciences and Environmental Sustainability, University of Parma, 43124 Parma, Italy; [email protected] (S.V.); [email protected] (U.C.); [email protected] (M.N.) * Correspondence: [email protected]; Tel.: +39-0521-905410 Abstract: The number of applications of peptide nucleic acids (PNAs)—oligonucleotide analogs with a polyamide backbone—is continuously increasing in both in vitro and cellular systems and, parallel to this, delivery systems able to bring PNAs to their targets have been developed. This review is intended to give to the readers an overview on the available carriers for these oligonucleotide mimics, with a particular emphasis on newly developed multi-component- and multifunctional vehicles which boosted PNA research in recent years. The following approaches will be discussed: (a) conjugation with carrier molecules and peptides; (b) liposome formulations; (c) polymer nanopar- ticles; (d) inorganic porous nanoparticles; (e) carbon based nanocarriers; and (f) self-assembled and supramolecular systems. New therapeutic strategies enabled by the combination of PNA and proper delivery systems are discussed. Keywords: peptide nucleic acids; delivery; nanoparticles; conjugates; multifunctional systems 1. Introduction 1.1. Peptide Nucleic Acids and Their Uses Peptide nucleic acids (PNAs, Figure1) are DNA analogues in which the sugar- Citation: Volpi, S.; Cancelli, U.; phosphate units connecting the nucleobases are replaced by N-(2-aminoethyl)glycine Neri, M.; Corradini, R. moieties [1]. These molecules are excellent binding partners for cognate DNA and RNA Multifunctional Delivery Systems for strands, being able to exploit both the canonical Watson–Crick (WC) base pairing to form Peptide Nucleic Acids.
    [Show full text]
  • Peptide Nucleic Acids Targeted to the Neurotensin Receptor and Administered I.P
    Proc. Natl. Acad. Sci. USA Vol. 96, pp. 7053–7058, June 1999 Neurobiology Peptide nucleic acids targeted to the neurotensin receptor and administered i.p. cross the blood–brain barrier and specifically reduce gene expression BETH M. TYLER*†,KAREN JANSEN*, DANIEL J. MCCORMICK‡,CHRISTOPHER L. DOUGLAS*, MONA BOULES*, JENNIFER A. STEWART*, LIHONG ZHAO*, BENJAMIN LACY*, BERNADETTE CUSACK*, ABDUL FAUQ§, AND ELLIOTT RICHELSON* Laboratories of *Neuropsychopharmacology and §Neurochemistry, Mayo Foundation for Medical and Educational Research, Jacksonville, FL 32224; and ‡Department of Biochemistry and Molecular Biology, Mayo Foundation for Medical and Educational Research, Rochester, MN 55905 Communicated by Susan E. Leeman, Boston University School of Medicine, Boston, MA, April 14, 1999 (received for review January 7, 1999) ABSTRACT Intraperitoneal injection of an unmodified mRNA levels. A sensitive assay developed to detect the antisense peptide nucleic acid (PNA) complementary to amount of PNAs in tissue (gel shift assay) confirmed the mRNA of the rat neurotensin (NT) receptor (NTR1) was presence of PNA in brain after i.p. injection. Therefore, these demonstrated by a gel shift assay to be present in brain, thus results provided evidence that any antisense strategy targeted indicating that the PNA had in fact crossed the blood–brain to brain proteins can work by i.p. delivery and by crossing the barrier. An i.p. injection of this antisense PNA specifically normal (i.e., not compromised by malignancy) BBB. Also, of inhibited the hypothermic and antinociceptive activities of NT great interest was the fact that a sense-NTR1 PNA, targeted microinjected into brain. These results were associated with a to the DNA sequence (in this case injected directly into brain) reduction in binding sites for NT both in brain and the small caused the same blockade of the responses to NT and signif- intestine.
    [Show full text]
  • Theranostics Molecular Beacons of Xeno-Nucleic Acid for Detecting
    Theranostics 2013, Vol. 3, Issue 6 395 Ivyspring International Publisher Theranostics 2013; 3(6):395-408. doi: 10.7150/thno.5935 Review Molecular Beacons of Xeno-Nucleic Acid for Detecting Nucleic Acid Qi Wang2, Lei Chen1, Yitao Long1, He Tian1 and Junchen Wu1, 1. Key Lab for Advanced Materials and Institute of Fine Chemicals, East China University of Science and Technology, China. 2. College of Public Health, Nantong University, China. Corresponding author: Junchen Wu, Meilong 130, Shanghai 200237, China. Telephone: (+86) 21-6425-3674, fax: (+86) 21-6425-2258, E-mail: [email protected]. © Ivyspring International Publisher. This is an open-access article distributed under the terms of the Creative Commons License (http://creativecommons.org/ licenses/by-nc-nd/3.0/). Reproduction is permitted for personal, noncommercial use, provided that the article is in whole, unmodified, and properly cited. Received: 2013.01.23; Accepted: 2013.04.10; Published: 2013.05.05 Abstract Molecular beacons (MBs) of DNA and RNA have aroused increasing interest because they allow a continuous readout, excellent spatial and temporal resolution to observe in real time. This kind of dual-labeled oligonucleotide probes can differentiate between bound and unbound DNA/RNA in homogenous hybridization with a high signal-to-background ratio in living cells. This review briefly summarizes the different unnatural sugar backbones of oligonucleotides combined with fluoro- phores that have been employed to sense DNA/RNA. With different probes, we epitomize the fundamental understanding of driving forces and these recognition processes. Moreover, we will introduce a few novel and attractive emerging applications and discuss their advantages and dis- advantages.
    [Show full text]