Synthetic Nucleic Acid Delivery Systems in Gene Therapy


Synthetic nucleic acid delivery systems are prepared primarily by means of synthetic organic chemistry for the functional delivery of therapeutic nucleic acids to cells in vitro, ex vivo and in vivo. From the inception of this field, cationic liposomes/micelles and cationic polymers have been prepared and combined with therapeutic nucleic acids to form lipid‐based nanoparticles (LNPs) or polymer‐based nanoparticles (PNPs), respectively, which were found to mediate functional delivery of therapeutic nucleic acids to target cells in vitro, ex vivo and in vivo. Regrettably, these early‐version PNPs progressed with difficulty into clinical studies frequently owing to issues of cytotoxicity. On the other hand, even though early‐version LNPs were extensively studied in clinic, they too proved poorly biocompatible and too unstable in terms of formulation and storage properties for routine clinical use. More fundamentally, these early‐version LNPs did not mediate the hoped for therapeutic benefits in clinic. Given this, the past decade of research into synthetic nucleic acid delivery systems has focused on how to upgrade both the chemistry and capabilities of lipids to derive significantly more sophisticated LNPs with enhanced levels of delivery efficacy and biocompatibility. There is still no marketed gene therapy using synthetic nucleic acid delivery systems, but prospects remain bright.

Key Concepts

  • Gene therapy has been described as the use of genes as medicines to treat disease or as the delivery of nucleic acids by means of a ‘vector’ to patients for some therapeutic purpose.
  • Gene therapy should now be defined more broadly as the use of nucleic acids as medicines to treat disease by exploiting genetic level mechanism(s) involving gene modification, replacement, replication and/or transcription. In truth, practical realisation of most, if not all, gene therapies in clinic will require delivery of therapeutic nucleic acids by means of ‘vectors’.
  • ‘Vectors’ are viral or else are nonviral including those whose origin is synthetic.
  • Synthetic nonviral vectors are prepared using synthetic chemistry and formulated with therapeutic nucleic acids of choice to give synthetic nucleic acid delivery systems.
  • Lipid‐based nanoparticles (LNPs) prepared from lipids and nucleic acids or polymer‐based nanoparticles (PNPs) prepared from polymers and nucleic acids are the main classes of synthetic nucleic acid delivery systems.
  • Early‐version LNPs have entered clinic in the past but are poorly efficient and not particularly biocompatible.
  • LNPs that conform to the ABC or ABCD nanoparticle concept appear most appropriate for in vivo and clinical use.
  • ABC‐type LNPs that deliver ncRNA therapeutic nucleic acids have proven the most efficacious in clinic to date.
  • Recent research is beginning to show how the four Ss of nanotechnology (size, shape, surface and structure) can be understood to impact on the four Ts of delivery (trafficking, triggerability, targeting and timing).
  • Such a biophysical framework for understanding delivery outcomes coupled with technical improvements in lipid synthesis can be expected to help significantly in efforts to improve synthetic nucleic acid delivery systems for routine clinical gene therapy applications.

Keywords: liposomes; polymers; lipid‐based nanoparticles; polymer‐based nanoparticles; synthetic nonviral vectors; nucleic acids; gene therapy; therapeutic nucleic acids; pDNA; RNAi effectors

Figure 1. Structures of some notable neutral colipids (B‐layer) used in the preparation of LNP (lipid‐based nanoparticle) type synthetic nucleic acid delivery systems: Chol, cholesterol; DOPE, 1,2‐dioleoyl‐sn‐glycero‐3‐phosphoethanolamine or dioleoyl l‐α‐phosphatidylethanolamine; DOPC, 1,2‐dioleoyl‐sn‐glycero‐3‐phosphocholine or dioleoyl l‐α‐phosphatidylcholine; DSPE, 1,2‐distearoyl‐sn‐glycero‐3‐phosphoethanolamine or distearoyl l‐α‐phosphatidylethanolamine and μ (mu), mature adenovirus core peptide.
Figure 2. Structures of some notable cytofectins (B‐layer) used in the preparation of LNP type synthetic nucleic acid delivery systems: DC‐Chol, 3β‐[N‐(N′,N′‐dimethylaminoethane)‐carbamoyl]cholesterol; CDAN, N1‐cholesteryloxycarbonyl‐3,7‐diazanonane‐1, 9‐diamine; GL‐67, lipid 67; DODAG, N′,N′‐dioctadecyl‐N‐4,8‐diaza‐10‐aminodecanoylglycine amide and DS(14‐yne)TAP, 1,2‐(distear‐14‐ynoyloxy)‐3‐(trimethylammonium) propane.
Figure 3. Structures of some notable cationic polymers (B‐layer) used in the preparation of early version PNP type synthetic nucleic acid delivery systems: pLL, poly‐l‐lysine; PEI, polyethylenimine and PAMAM, polyamidoamine dendrimer.
Figure 4. Graphic illustration of synthetic, self‐assembly ABCD nanoparticles to show how APIs (active pharmaceutical ingredients) – such as pDNA (plasmid deoxyribonucleic acid), siRNA (small interfering ribonucleic acid) (or other drug agents) (A‐components) are condensed within functional concentric layers of chemical components to facilitate complementary delivery functions; B‐components (typically lipids or polymers) are introduced for the compaction/association of APIs, and in order to facilitate their cellular delivery followed by intracellular trafficking; C‐component(s) (typically stealth/biocompatibility polymers such as PEG) are required to engender biological and physical stability; D‐components (bona fide biological receptor‐specific targeting ligands) may be employed to target specific cell populations. By definition, in lipid‐based nanoparticles, also known in text as LNPs, B‐components are primarily lipids; in polymer‐based nanoparticles, also known in text as PNPs, B‐components are primarily polymers. Reproduced by kind permission from KP Therapeutics Ltd.
Figure 5. Structures of some notable cytofectins and colipids (B‐layer) and PEG‐lipids (BC‐layer) used in the preparation of some more advanced LNP ABC‐type synthetic nucleic acid delivery systems: DODMA, N‐[1‐(2,3‐dioleyloxy)propyl]‐N,N‐dimethyl ammonium chloride; DSPC, 1,2‐distearoyl‐sn‐glycero‐3‐phosphocholine or distearoyl l‐α‐phosphatidylcholine; PEG2000‐DSG, (ω‐methoxy‐polyethylene glycol 2000)‐distearoyl‐sn‐glycerol; DLinDMA, 1,2‐dilinoleyloxy‐3‐dimethyl‐aminopropane; DLin‐KC2‐DMA, 2,2‐dilinoleyl‐4‐(2‐dimethyl‐aminoethyl)‐[1,3]‐dioxolane; DLin‐MC3‐DMA (MC3), (6Z, 9Z, 28Z, 31Z)‐heptatriaconta‐6,9,28,31‐tetra‐en‐19‐yl 4‐(dimethylamino)‐butanoate; DPPC, 1,2‐dipalmitoyl‐sn‐glycero‐3‐phosphocholine or dipalmitoyl l‐α‐phosphatidylcholine; PEG2000‐C‐DMA, 3‐N‐(ω‐methoxy‐polyethylene glycol 2000) carbamoyl‐1,2‐dimyristyl‐oxypropylamine; AtuFECT01, l‐arginyl‐2,3‐l‐diaminopropionic acid‐N‐palmityl‐N‐oleylamide trihydrochloride; DPhyPE, l,2‐diphytanoyl‐sn‐glycero‐3‐phosphoethanolamine or l,2‐diphytanoyl l‐α‐phosphatidylethanolamine and PEG2000‐DSPE, (ω‐methoxy‐polyethylene glycol 2000)‐N‐carboxy‐1,2‐distearoyl‐sn‐glycero‐3‐phosphoethanolamine or (ω‐methoxy‐polyethylene glycol 2000)‐N‐carboxy‐distearoyl l‐α‐phosphatidylethanolamine.
Figure 6. Structures of well‐known labelling lipids (B‐layer) and targeting lipids (BCD‐layer) used in the preparation of some more advanced LNP ABC and ABCD type synthetic nucleic acid delivery systems: Gd.DOTA.Chol, gadolinium (III) 2‐(4,7‐bis‐carboxymethyl‐10‐[2‐N′‐(cholesteryloxycarbonyl)‐2‐aminoethylamidomethyl]‐1,4,7,10‐ tetraazacyclododec‐1‐yl) acetic acid; Gd.DOTA.DSA, gadolinium (III) 2‐(4,7‐bis‐carboxymethyl‐10‐[(N,N‐distearyl‐amidomethyl)‐N′‐amidomethyl]‐1,4,7,10‐tetraazacyclododec‐1‐yl) acetic acid; DOPE‐Rhoda, 1,2‐dioleoyl‐sn‐glycero‐3‐phosphoethanolamine‐N‐(lissamine rhodamine B sulphonyl) or dioleoyl l‐α‐phosphatidylethanolamine‐N‐(lissamine rhodamine B sulphonyl); folate‐PEG2000‐DSPE, (folate‐N‐ω‐polyethylene glycol 2000)‐N‐carboxy‐1,2‐distearoyl‐sn‐glycero‐3‐phosphoethanolamine.


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Further Reading

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Miller, Andrew D(Apr 2017) Synthetic Nucleic Acid Delivery Systems in Gene Therapy. In: eLS. John Wiley & Sons Ltd, Chichester. [doi: 10.1002/9780470015902.a0005745.pub3]