Lipid Nanoparticles (LNP) for mRNA Drug Delivery.pdf
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Lipid Nanoparticles (LNP) for mRNA Drug
Delivery
mRNA (messenger RNA), is a type of single-stranded RNA that is transcribed
from a strand of DNA as a template, which carries genetic information and can
guide protein synthesis. mRNA was first proposed by Jacques Monod and
François Jacob, and later discovered by Jacob, Sydney Brenner and Matthew
Meselson at Caltech in 1961. Since 2015, the mRNA technology has been
growing at an accelerated pace as delivery and modification technologies have
matured, and biotechnology companies such as CureVac, BioNTech, and
Moderna, which are focused on mRNA technology, have come to prominence.
The COVID-19 pandemic has brought mRNA vaccines and drugs to
widespread attention worldwide and has led to a boom in research in the
mRNA field. According to Nature related statistics, as of the end of July 2021,
there are 180 mRNA vaccine and drug pipelines in development worldwide,
the vast majority of which are related to infectious diseases, rare diseases and
oncology.
mRNA has high molecular weight and strong hydrophilicity, but its
single-stranded structure makes it extremely unstable and susceptible to
degradation. The limited lifetime of mRNA allows cells to rapidly change
protein synthesis in response to its changing needs, but it is difficult to meet
the requirements of druggability. In addition, mRNA molecules carry a negative
charge and have difficulty crossing the cell membrane with the same negative
charge on the surface, so special modification or wrapping delivery system is
required to achieve intracellular expression of mRNA drugs, therefore, delivery
technology is one of the core patented technologies of mRNA companies.
Lipid Nanoparticle (LNP) is currently the dominant delivery system. They are
often used in vaccines due to their relatively easy uptake by
antigen-presenting cells. The three major mRNA vaccine giants Moderna,
CureVac and BioNTech are currently using LNP delivery technology for
their COVID-19 vaccines.
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Structure of Lipid Nanoparticles
The LNP components that are widely used at this stage include the following
four major categories: cationic or ionizable lipids, cholesterol, PEGylated lipids
and phospholipids.
1. Cationic Or Ionizable Lipids
Cationic lipids (CLs) and ionizable lipids (ILs) can initiate the first step of
self-assembly through electrostatic interactions. Lipid complexes containing
cationic lipids are still widely used for nucleic acid delivery. However, they
have been largely replaced by pH-responsive ionizable lipids due to toxicity
concerns and lack of in vivo potency. ionizable lipids in LNP formulations
behave as neutral at physiological pH, while positively charged in the acidic
environment of endosomes. The pH-dependent ionization ability makes
ionizable lipids suitable materials for nucleic acid delivery due to the large
improvement in potency and toxicity characteristics.
The development of cationic lipids requires a balance between delivery
efficiency and cytotoxicity. The cytotoxicity of cationic lipids depends on the
structure of their hydrophilic head groups, e.g., amphiphilic molecules
containing quaternary ammonium head groups are more toxic than amphiphilic
molecules containing tertiary amines. Therefore, addressing the cytotoxicity of
ionizable cationic lipids is one of the key points of LNP technology and the
focus of patent protection. The proprietary cationic lipids ALC-0315
(Acuitas/Pfizer) and SM-102 (Moderna) both have hydrophilic head groups
that are tertiary amines that can be positively charged by protonation in a
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physiological low pH environment and are safely cleared after mRNA is
delivered.
2. Cholesterol
Cholesterol is often included as a helper that improves intracellular delivery as
well as LNP stability in vivo. Helper neutral lipids (e.g., various phospholipids)
are also used to construct LNP bilayers because the bilayer structure of
cationic liposomes is not stable.
3. PEGylated Lipids
LNPs modified by PEG with specific structure can control the particle size of
nanoparticles during nanoparticle synthesis. Due to the strong hydration of
polyethylene glycol ethoxy links, the PEG structure can form a hydrophilic
protective layer in the aqueous phase, which can effectively prevent the
aggregation of nanoparticles during storage, thus maintaining the spatial
stability of LNP. At the same time, PEG on the surface of LNP particles can
protect the particles from being detected by immune proteins in vivo, shield the
particles from being bound by plasma proteins and other components, and
prevent LNP particles from being cleared in vivo. The proprietary PEG lipid
structure ALC-0159 (M-DTDAM-2000) currently selected by Pfizer
and M-DMG-2000 selected by Moderna are both derivatives of PEG-2000.
4. Phospholipids
Helper lipids are represented by phospholipids such as DOPE, DSPC and
DOPC. In the preparation of cationic liposomes, helper lipids have very strong
synergistic effects, mainly including stabilizing bilayer membranes and
reducing the toxicity of cationic components, promoting the release of mRNA
when LNP is endocytosed and assisting the cell permeation of cationic
liposomes, and determining the morphology of mRNA-LNP complexes,
making the complexes well fusible and improving the transmembrane
efficiency.
Huateng Pharma has the capability to produce LNPs delivery system
excipients on a large scale and has been supplying a wide range of PEG lipids
and helper lipids, including mPEG2000-DMG and ALC-0159, to domestic and
international customers. Huateng Pharma is an industry leader in impurity
