#8. Scaling of COVID vaccine manufacturing: It’s all about the lipids

by Öner Tulum, William Lazonick, and Ken Jacobson                          

This is the first installment of a three-part series on the scalability of mRNA vaccine production. The current article focuses on the decades-long research in various technology domains that enabled drug companies to develop mRNA-based vaccines and to bring the first two COVID vaccines onto the market within a year. 

In September 2020, the BioNTech/Pfizer partnership was scaling production of its messenger ribonucleic acid (mRNA) vaccine. With phase III clinical trials still in progress, Pfizer CEO Albert Bourla and BioNTech CEO Ugur Sahin met each other in person for the first time. The occasion was a flight in a Pfizer company jet to visit a small family-owned firm, Polymun, based in Klosterneuburg, Austria. 

Polymun is one of only a handful of companies in the world that can produce lipid nanoparticles (LNPs), the substance most critical in the mass manufacture of an mRNA COVID vaccine. The two CEOs made this unusual trip because their COVID vaccine effort depended not only on Polymun’s capability to meet its production target for LNPs but also on its willingness to share its knowhow to enable scaling of production of LNPs at Pfizer facilities in Europe and the United States.

An LNP is a miniscule bubble of fat used to encapsulate mRNA that is encoded with the genetic information of a disease-specific antigen–in this case, the spike protein of the SARS-CoV-2 virus. LNPs constitute the packaging that protects the mRNA molecule as the vaccine travels through the human body to reach its target cells and trigger an immune response.

Messenger RNA technology has enabled the remarkably rapid design and development of what appear to be the safest and most effective COVID vaccines yet approved. The BioNTech and Moderna vaccines represented the very first attempts to produce mRNA-based vaccines. Scaling production of these novel vaccines was a complex manufacturing puzzle that had to be solved “on the job.” Whereas, as we have previously explained, the greatest complications in producing viral-vector vaccines occur in the drug-substance (DS) stage of the manufacturing process, the major challenges in the mass production of mRNA vaccines take place in the subsequent drug-product (DP) stage, in particular during the formulation process.

Pharmaceutical applications of RNAs

The fight against the COVID pandemic marks the first time that mRNA-based vaccines have ever been deployed. But the constituent technologies such as the chemical compounds, the excipients used in the vaccines, and the RNA and non-viral drug delivery platforms were well known to scientists and regulators prior to the pandemic. The ready availability of these technologies made it possible for Moderna and BioNTech/Pfizer to develop approved COVID vaccines in such short periods of time.

Since the 1990s, scientists have been exploring potential therapeutic applications of various RNA molecules, such as small interfering RNA (siRNA) and  messenger RNA (mRNA). Through an animal study in 1990, scientists demonstrated the expression of a protein in cells by injecting a naked RNA encoded with the genetic information of the target protein. In the decade leading up to the start of the pandemic in 2019, scientists had already developed a significant understanding of the spike protein of the SARS-CoV-2 virus, building on knowledge acquired in researching the SARS-CoV-1 (or SARS) and MERS-CoV (or MERS) outbreaks. Based on previous scientific learning, both Moderna and BioNTech were well-positioned to quickly sequence a single-stranded RNA molecule encoded with the genetic information that could instigate the production of these spike proteins within cells, triggering an immune response that would produce antibodies to fight future viral attacks. 

Lipid-based drug-delivery technology for mRNA vaccines

Without previous advances in drug-delivery technologies such as adeno-associated virus vectors (used in the Oxford, Janssen, and Sputnik V vaccines) and LNPs (used in the BioNTech, Moderna, and CureVac mRNA vaccines), it would have taken many more years to develop vaccines against COVID-19. But while scientists have known about mRNA’s potential as a therapeutic tool since the 1990s, it was not until 2018 that LNPs were used as an effective vehicle for the delivery of an mRNA-based therapeutic–one for a rare genetic disease.

Messenger RNA is a highly fragile molecule, but this fragility has both advantages and disadvantages in preparing the immune system to fight a disease-causing agent such as the SARS-COV-2 virus. The advantage is that mRNA degrades quickly after performing the job for which it was programmed, thus eliminating excessive protein replication that could be harmful to the human body. The rapidity of mRNA’s biodegradability occurs because the human body has an enzyme called Ribonuclease (or RNase) that breaks mRNA molecules into pieces small enough so that they are no longer a threat to healthy cells. But the disadvantage is that the naked mRNA can degrade so rapidly upon entering the human body that it does not have time to perform the task of carrying a viral pathogen’s genetic information to an immune system that has no antibodies readily available.   

The term lipid nanoparticle (LNP) refers to the assembly of lipids and mRNA for the safe and effective delivery of the mRNA molecule packed with the genetic sequence of the protein. The formulations of the three existing mRNA vaccines contain four types of lipids, each performing a specific task. 

The image below shows two structural lipids (also referred to as “helper lipids”): distearoylphosphatidylcholine (DSPC) and cholesterol. These structural lipids, which have been widely used in both the pharmaceutical and cosmetics industries, are critical for the stability and delivery efficiency of LNPs. In addition, there are two functional lipids used in the LNP assembly, poly(ethylene glycol)-conjugated phospholipids (PEGylated lipids or PPLs) and ionizable cationic lipids (ICLs). By mimicking the membrane of organic cells, PPLs play a critical role in the survival of the LNPs while they are circulating in the body by preventing LNPs from getting tagged by certain proteins as foreign microorganisms. PEGylated lipids also prevent LNPs from shrinking in size as they come into contact with water.

mRNA lipid nanoparticle structure

[Click image to enlarge]
The molecular structure of the ICLs is manipulated to acquire a positive charge, thus ensuring that the negatively charged mRNA is encapsulated within a liposome (a spherical vesicle around the mRNA molecule). Furthermore, ICLs instigate the process of endocytosis, through which the negatively charged cell membrane (outer layer of cells) swallows an LNP, of which positively charged ICLs account for more than one-half of the lipid content. After accessing the cytoplasm (that part of a cell between its membrane and nucleus), ICLs begin to dissolve due to the low acid/water (pH) ratio in the cell and to release mRNA into the cytoplasm. 

Now that we know what LNPs are and what they do, our upcoming posts will explain the process of lipid procurement and the subsequent assembly of mRNA into LNPs.