Gene therapy represents an innovative approach to treating diseases by targeting the underlying genetic causes. By modulating gene expression, gene therapies aim to provide root-cause treatment for genetic disorders, focusing on aberrant genes or gene networks. At Cytiva, we have developed the genetic medicine toolkit, a set of essential technologies designed to accelerate the development of gene therapies that tackle some of the most challenging diseases.
The promise of non-viral gene therapy
Non-viral gene therapy is a promising strategy for diseases caused by loss-of-function mutations, where the body is unable to produce a crucial protein. This approach enables protein replacement therapy using messenger RNA (mRNA) to deliver the instructions for protein production. Key advantages of non-viral gene therapy:
- Decreased complexity: no cell culture is required to produce the drug product, streamlining both R&D and manufacturing.
- Increased modularity: mRNA can be designed to express virtually any protein, while optimized LNP formulations simplify the development of future gene therapies.
For instance, Cytiva's genetic medicine toolkit demonstrated proof-of-concept (PoC) for protein replacement therapy in an anemia model using mRNA that encodes erythropoietin (Epo). This mRNA was encapsulated in GenVoy-ILM™ lipid nanoparticles (LNP) and manufactured using our NanoAssemblr™ platform.
Case study: non-viral gene therapy for anemia
In a model of anemia, mRNA encoding human erythropoietin (Epo) was encapsulated in GenVoy-ILM™ LNP and administered intravenously. Once delivered to the liver, hepatocytes expressed and secreted EPO into the bloodstream, leading to increased red blood cell production. As the therapy was scaled up using the NanoAssemblr™ platform, the production remained consistent, showcasing the reproducibility and scalability of this non-viral gene therapy solution.
Fig 1. Non-viral gene therapy mediates erythropoietin production reversing disease phenotype in an anemia model
Genetic medicine framework and toolkit
Developing gene therapies follows a framework similar to other therapeutic modalities but with unique considerations. Our genetic medicine framework guides gene therapy development from conception to commercialization, with technologies designed to accelerate every stage.
Fig 2. Genetic medicine framework
Defining the target product profile (TPP)
The target product profile (TPP) outlines the ideal characteristics of a drug product for treating a specific disease. For gene therapy, the TPP varies based on the disease, the therapeutic target, and market considerations. A simplified profile for chronic gene therapy includes:
- Regimen: administered every 2 to 4 weeks
- Route of administration: intravenous
- Stability: long-term storage at -20°C or higher
- Presentation: single-dose liquid suspension with a maximum dose volume of 20 mL
1. Target Selection: precision at the core
The first step in developing gene therapy is target selection, often using bioinformatics to identify gene targets associated with the disease. Gene therapy approaches can either knock down faulty genes using siRNA or express functional genes using mRNA. Emerging technologies like CRISPR/Cas9 offer the potential for long-term treatment by editing genes directly. Both in vitro and in vivo models validate the selected targets before progressing to human studies.
2. Vector selection: choosing the right carrier
For gene knockdown therapies, siRNA mimics microRNA to degrade specific mRNA transcripts through RNA interference (RNAi) pathways (1). Meanwhile, mRNA allows for transient gene expression without integrating into the genome, reducing long-term risks. For protein replacement therapies, mRNA featuring a cap1 structure and mammalian base modifications are preferred to minimize immune reaction and maximize half-life (2,3).
Both mRNA and siRNA therapies can be manufactured through cell-free enzymatic synthesis, simplifying production and accelerating the development of RNA therapeutics (4).
3. Delivery and formulation technology: optimizing efficacy
Lipid nanoparticles (LNP) have emerged as the most advanced non-viral delivery system for gene therapies. Notably, the FDA-approved ONPATTRO for treating polyneuropathy caused by hATTR amyloidosis employs an LNP-siRNA approach. More recently, mRNA-LNP COVID-19 vaccines have been authorized for clinical use in several countries.
Our LNP are engineered with ionizable cationic lipids, offering a versatile platform for non-viral gene delivery across multiple applications, including protein replacement therapies.
The benefits of LNP delivery include:
- Cell-free production: streamlines the manufacturing process
- Scalability: continuous-flow manufacturing accelerates drug development
4. Scalable manufacturing: from discovery to commercialization
Our NanoAssemblr™ platform uses microfluidic mixing to produce high-quality RNA-LNP. The proprietary NxGen™ microfluidic architecture enables precision control of LNP self-assembly, ensuring that gene therapies can be manufactured consistently at every stage of development.
NanoAssemblr™ Spark™, Ignite™/Ignite+™, Blaze™/Blaze+™ systems, and the GMP and NanoAssemblr™ commercial formulation system (CFS) systems share the same NxGen™ technology, allowing seamless scaling from early-stage research to commercial production.
End-to-end gene therapy development solutions – We offer comprehensive, flexible solutions to support gene therapy development from concept to clinical implementation. With advanced LNP technology and development framework, we are confident in accelerating the timeline for advancing genetic medicines from the lab to commercial use.
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REFERENCES
- Samaridou E, Walgrave H, Salta E, et al. Nose-to-brain delivery of enveloped RNA - cell permeating peptide nanocomplexes for the treatment of neurodegenerative diseases. Biomaterials. 2020;230:119657. doi:10.1016/j.biomaterials.2019.119657
- Karikó K, Buckstein M, Ni H, Weissman D. Suppression of RNA Recognition by Toll-like Receptors: The Impact of Nucleoside Modification and the Evolutionary Origin of RNA.
- Robinson E, MacDonald KD, Slaughter K, et al. Lipid Nanoparticle-Delivered Chemically Modified mRNA Restores Chloride Secretion in Cystic Fibrosis. Mol Ther. 2018;26(8):2034-2046. doi:10.1016/j.ymthe.2018.05.014
- Mollocana-Lara EC, Ni M, Agathos SN, Gonzales-Zubiate FA. The infinite possibilities of RNA therapeutics. J Ind Microbiol Biotechnol. 2021;48(9-10):kuab063. doi:10.1093/jimb/kuab063