Cell-Penetrating Peptide Conjugates of Steric Blocking Oligonucleotides as Therapeutics for Neuromuscular Diseases from a Historical Perspective to Current Prospects of Treatment.

Michael J Gait,Corinne Betts,Caroline Godfrey,Graham Mcclorey,Andrey A Arzumanov,Suzan Hammond,Matthew J.A Wood

doi:10.1089/nat.2018.0747

Abstract

The review starts with a historical perspective of the achievements of the Gait group in synthesis of oligonucleotides (ONs) and their peptide conjugates toward the award of the 2017 Oligonucleotide Therapeutic Society Lifetime Achievement Award. This acts as a prelude to the rewarding collaborative studies in the Gait and Wood research groups aimed toward the enhanced delivery of charge neutral ON drugs and the development of a series of Arg-rich cell-penetrating peptides called Pip (peptide nucleic acid/phosphorodiamidate morpholino oligonucleotide [PNA/PMO] internalization peptides) as conjugates of such ONs. In this review we concentrate on these developments toward the treatment of the neuromuscular diseases Duchenne muscular dystrophy and spinal muscular atrophy toward a platform technology for the enhancement of cellular and in vivo delivery suitable for widespread use as neuromuscular and neurodegenerative ON drugs.

Highlights

Therapeutic antisense oligonucleotides (ONs) of the type where the mechanism of mRNA silencing occurs through the action of the intracellular RNase H enzyme to cleave a complementary RNA strand have been reviewed extensively previously [1,2], including recently by Stanley Crooke, the recipient of the 2016 Oligonucleotide Therapeutic Society (OTS) Lifetime Achievement Award [3]
We recently reviewed the chemistries used in steric blocking ON approaches [4]. Such steric blocking ONs have emerged as approved therapeutic agents for neuromuscular diseases, such as eteplirsen for treatment of Duchenne muscular dystrophy (DMD) [5] and nusinersen for treatment of spinal muscular atrophy (SMA) [6]
In the years that followed most of the development of therapeutic ONs took place using gapmers, where a central core of deoxynucleotides containing PS linkages was flanked by a number of 2¢-modified nucleotide analogs on each arm, such analogs being insensitive to recognition by RNase H

Summary

Introduction

Therapeutic antisense oligonucleotides (ONs) of the type where the mechanism of mRNA silencing occurs through the action of the intracellular RNase H enzyme to cleave a complementary RNA strand have been reviewed extensively previously [1,2], including recently by Stanley Crooke, the recipient of the 2016 Oligonucleotide Therapeutic Society (OTS) Lifetime Achievement Award [3]. The poorer intracellular efficacy of steric blocking ONs, compared to the RNase H antisense types, is partly due to their intrinsic RNA target-blocking mode of action where a competition must occur for interaction with components of the RNA recognition machinery (eg, splicing complexes) This process must require at least a stoichiometric quantity of ON to reach and bind to its RNA target. Many attempts have been made over recent years to enhance the cellular and in vivo delivery of ONs to improve their effectiveness, both steric blocking and otherwise This may be achieved in some cases by judicious choices of the chemistry of the backbone of ONs. For example, the use of a phosphorothioate (PS) backbone, to replace a phosphodiester backbone, results in a greater stability against nuclease degradation for which it was designed and serendipitously in enhanced serum protein binding upon systemic injection, and, increases bioavailability [8,9]. This review concentrates initially on the contributions of the Gait group in ON and conjugate synthesis toward the award of the 2017 OTS Lifetime Achievement Award as a prelude to the exciting collaborative work of the Gait and Wood groups in the development of peptide-ON conjugates as therapeutics in treatment for neuromuscular diseases

RNAse H and Initial Steric Blocking Activities of Antisense ONs

These include on the left side

Findings

CPP Conjugates of Charge Neutral PNA ONs