Delivery of therapeutic oligonucleotides with cell penetrating peptides

Abstract

Oligonucleotide-based drugs have received considerable attention for their capacity to modulate gene expression very specifically and as a consequence they have found applications in the treatment of many human acquired or genetic diseases. Clinical translation has been often hampered by poor biodistribution, however. Cell-penetrating peptides (CPPs) appear as a possibility to increase the cellular delivery of non-permeant biomolecules such as nucleic acids. This review focuses on CPP-delivery of several classes of oligonucleotides (ONs), namely antisense oligonucleotides, splice switching oligonucleotides (SSOs) and siRNAs. Two main strategies have been used to transport ONs with CPPs: covalent conjugation (which is more appropriate for charge-neutral ON analogues) and non-covalent complexation (which has been used for siRNA delivery essentially). Chemical synthesis, mechanisms of cellular internalization and various applications will be reviewed. A comprehensive coverage of the enormous amount of published data was not possible. Instead, emphasis has been put on strategies that have proven to be effective in animal models of important human diseases and on examples taken from the authors' own expertise.

Peptides for nucleic acid delivery

“…PMOs are chemically modified ONs that harbor a morpholino moiety instead of the ribose moiety and phosphorodiamidate linkages instead of the phosphodiester linkages. CPP conjugation to PMO has been performed via a variety of methods including: maleimide linkage, disulfide linkage, click chemistry, or amide linkage between the free PMO secondary amine and an activated carboxylic group at the c-terminus of the peptide, which is the most popular [21]. Initial reports showing promising results using this chemistry were based on using arginine-rich peptides together with other chemical modifications such as aminohexanoic acid and beta alanine [44,45].…”

“…Importantly, they demonstrate excellent potential for the delivery of nucleic acids, ranging from natural or synthetic short RNA or DNA ONs to much larger plasmid molecules, both in vitro and in vivo [19][20][21][22]. CPPs can have very different origin and are in many ways ambiguous to define.…”

Current Understanding of Direct Translocation of Arginine-Rich Cell-Penetrating Peptides and Its Internalization Mechanisms

“…Among them, arginine-rich CPPs such as Tat peptide from a human immunodeficiency virus (HIV)-1 trans-activator protein TAT, [1][2][3] Penetratin peptide from a Drosophila homeodomain Antennapedia 4,5) and oligoarginines 6,7) show high efficiency of internalization, facilitating intracellular delivery of a wide range of hydrophilic molecules including small compounds, peptides, proteins, nucleic acids, nanoparticles, liposomes, and others that would be otherwise difficult to enter cells. [8][9][10] Arginine-rich CPPs have, therefore, attracted much attention as one of the promising carriers for intracellular delivery of therapeutic molecules including nucleic acids and drug candidates. [8][9][10] So far, a number of peptide sequences have been reported as CPPs, which are classified by their physicochemical characteristics, such as basic/amphiphilic CPPs that contain several basic amino acids including arginine and lysine, and hydrophobic CPPs (Table 1): interestingly, polyhistidine peptide such as histidine 16-mer (H16) is recently reported as CPPs.…”

“…[8][9][10] Arginine-rich CPPs have, therefore, attracted much attention as one of the promising carriers for intracellular delivery of therapeutic molecules including nucleic acids and drug candidates. [8][9][10] So far, a number of peptide sequences have been reported as CPPs, which are classified by their physicochemical characteristics, such as basic/amphiphilic CPPs that contain several basic amino acids including arginine and lysine, and hydrophobic CPPs (Table 1): interestingly, polyhistidine peptide such as histidine 16-mer (H16) is recently reported as CPPs. 11) Although the molecular mechanisms as to how argininerich CPPs enter cells have not been fully understood and still remain debated, it is commonly accepted that internalization of these peptides involves endocytosis.…”

Antisense Oligonucleotides as Therapeutics and their Delivery

“…However, clinical use of these agents would require circumvention of the challenges relating to their appropriate design, higher biological stability and targeted delivery. Various in vitro studies have revealed increase in cellular uptake of these agents by employing a delivery system [32][33][34] . However, delivery strategies need to be more refined and in vivo studies needs to be employed to evaluate the safety and toxicity associated with the delivery system 33 .…”