(360b) Challenges in the Pharmaceutical Development of Lipid Nanoparticle Therapeutics | AIChE

(360b) Challenges in the Pharmaceutical Development of Lipid Nanoparticle Therapeutics


Viza, N. - Presenter, Merck & Co.
Wagner, A., The University of Texas at Austin
Liu, Y., Merck & Co.
Zhao, A., Merck & Co.
Doty, A., Merck & Co.
Smith, K., Merck & Co.
Jia, X., Merck & Co.
Bothe, J., Merck & Co.
Su, Y., Merck & Co.
Li, M., Merck & Co.
Kemp, E., Merck & Co.
Socia, A., Merck & Co.
Guidry, E., Merck & Co.
Gindy, M., Merck & Co.
The discovery of RNA therapeutics has emerged as a promising modality to treat infectious diseases, cancer [1] and undruggable targets [2]. The delivery of RNA is a challenge because these molecules quickly get cleared upon administration and their size, along with negative charge does not allow it to cross into the cellular membrane [1]. As a result, many other platforms including positively charged antibody conjugates, masked endosomolytic agent-dynamic polyconjugates, or lipid nanoparticles offer an oligonucleotide delivery. These platforms also allow for DNA and RNA gene therapy, and vaccines. A recent rise in the development of clinical therapeutic RNA has led to the approvals of siRNA drugs (Patisiran) and mRNA vaccines (Moderna, CureVac, BioNTech) using lipid nanoparticle delivery [1].

The lipid nanoparticle delivery method offers the most mature platform for RNA therapeutics and has gained significant attention over the recent years [1]. However, there remains significant challenges in order to advance to commercialization. One of these challenges, requires efficient delivery of the RNA cargo to the targeted tissue and cellular compartment. Furthermore, the control of the lipid nanoparticle assembly is critical to the understanding the influence of in vivo pharmaceutical properties. A thorough comprehension of the physio-chemical properties to its bio performance is essential to ensuring the best quality commercialized drug product. This includes the molecular size, charge and polarity. The lipid nanoparticle assembly mechanism is also largely unknown [3]. We present an experimental design study of RNA loaded lipid nanoparticle self-assembly via a rapid antisolvent precipitation process, to identify critical parameters in the formation of LNPs. This will also help us understand the mechanism of the LNP assembly driving force to increase robustness for future formulation development and improve reproducibility.

RNA loaded lipid nanoparticles are complex structures of approximately 80-100nm, composed of cationic and neutral lipids. This includes PEGylated lipids in order to improve steric stability. The cationic lipid enables nucleic acid encapsulation and endosomal escape into the cytosol. The neutral lipids include phospholipid and cholesterol, which modulate the physical properties including rigidity, stability and biological membrane interactions. The oligonucleotide cargo, or RNA, is encapsulated at a defined ratio of cationic lipid to oligonucleotide. The process of forming the LNPs utilizes a micromixing device to generate controlled nanometer size, narrow polydispersity, and high RNA encapsulation efficiency [3]. The design of the mixing device is a continuous flow operation allowing for a rapid precipitation process suitable for commercia scale-up [3].

The goal of our developmental strategy is to control the LNP assembly process and reproducibility and correlate physicochemical properties with efficacy and/or toxicity. We use several analytical tools to enable robust and sensitive drug product characterization. For example, small-angle X-ray scattering (SAXS) quantitatively characterizes the structural features of RNA loaded LNPs [3]. Additionally, cryogenic transmission electron microscopy (cryo-TEM) micrographs can be used to identify LNP morphology, either hexagonal, cubic, unilamellar, or bilamellar macromolecular structures. Furthermore, differential scanning calorimetry (DSC) is a thermal analysis tool used to identify phase transitions [4] for LNPs. Both the SAXS and cryo-tem are powerful tools for characterization of LNP structure as a function of formulation manufacture.

In combination of our analytical tools and formulation composition we can generate LNPs of distinct lipid phases. A shift from lamellar to hexagonal shows to correspond to an improvement in potency. A difference in compositional heterogeneity in LNPs is observed to have significant difference in bioperformance. These overall changes in formulation or process are required frequently throughout preclinical, clinical and even sometime post market approval. Henceforth, understanding the key product attributes to in-vivo performance is essential. The analytical tools guide the formulation composition selection, manufacturing process development and ensure manufacturing process consistency.

  1. Roces et al., Pharmeceutics, 2020, 12, 1095
  2. Gindy et al., Expert Opin. 2012, 9, 171
  3. Gindy et al., Langmuir 2014, 30, 16, 4613
  4. Eygeris et al., Nanoletters, pending