mRNA

1. Traditional biologics timeline vs. Emergency COVID-19 steps to approval

2. Factors which allowed for quicker development

Ulrich Kalinke, Dan H. Barouch, Ruben Rizzi, Eleni Lagkadinou, Özlem Türeci, Shanti Pather & Pieter Neels (2022) Clinical development and approval of COVID-19 vaccines, Expert Review of Vaccines, 21:5, 609-619, DOI: 10.1080/14760584.2022.2042257

3. Timelines for specific COVID-19 vaccines

Ulrich Kalinke, Dan H. Barouch, Ruben Rizzi, Eleni Lagkadinou, Özlem Türeci, Shanti Pather & Pieter Neels (2022) Clinical development and approval of COVID-19 vaccines, Expert Review of Vaccines, 21:5, 609-619, DOI: 10.1080/14760584.2022.2042257

4. Timeline for development of COVID-19 vaccine vs. vaccines for other diseases

Ball, Philip. “The lightning-fast quest for covid vaccines — and what it means for other diseases.” Nature, vol. 589, no. 7840, 2020, pp. 16–18, https://doi.org/10.1038/d41586-020-03626-1

5. Timeline for development of COVID-19 vaccine vs. vaccines for other diseases, including vaccine type

Kumari, M., Lu, RM., Li, MC. et al.A critical overview of current progress for COVID-19: development of vaccines, antiviral drugs, and therapeutic antibodies. J Biomed Sci 29, 68 (2022). https://doi.org/10.1186/s12929-022-00852-9

 6. Milestones during COVID-19

  Operation Warp Speed

• • >$18 Billion
  • Provided funding for development, manufacturing, and distribution of COVID-19 vaccine candidates
  • View the breakdown in this report: Operation Warp Speed Contracts

1. Function: mRNA is a transient intermediate between DNA (genetic instructions) and protein (functional units). The “m” stands for messenger. The mRNA is a transcribed subset of the DNA, meaning it represents only a portion of the genome.  This strand of mRNA is translated by ribosomes, which will then build an amino acid chain. The amino acid chain will fold into a functional protein, a SARS-CoV-2 spike protein in the case of the Pfizer/BioNTech and Moderna vaccines.

“The Central Dogma” Introduction to Genomics for Engineers, St. Jude Children’s Research Hospital, learngenomics.dev/. Accessed 21 Nov. 2023.

2. Structure: RNA differs from DNA in that it contains a ribose sugar (extra -OH group) and incorporates uracil (U) bases instead of thymine (T) bases. The structures are similar but not interchangeable.

Parker, Nina, et al. “10.3 Structure and Function of RNA.” Microbiology, OpenStax, 1 Nov. 2016, openstax.org/books/microbiology/pages/1-introduction.

3. Delivery: mRNA is an inherently unstable molecule which degrades within hours to days within your body. mRNA also does not readily cross cell membranes. mRNA is enveloped in lipids for protection and to enter your cells cytoplasm (where translation takes place).

Aldosari, B.N.; Alfagih, I.M.; Almurshedi, A.S. Lipid Nanoparticles as Delivery Systems for RNA-Based Vaccines. Pharmaceutics 2021, 13, 206. https://doi.org/10.3390/pharmaceutics13020206

4. Sequence: We’ve already mentioned mRNA encodes the sequence for just a small portion of a genome. In terms of mRNA vaccines, that sequence encodes for a spike protein (antigen) of the SARS-CoV-2 virus, not the whole viral genome! This mRNA sequence is a set of instructions for your own cells to build only the spike protein. mRNA is a good potential platform technology because we can easily change the sequence but keep the majority of the manufacturing process the same. The complicated processes of translation and post-translational modification is outsourced to your own cells!

Richter, Greg. “Why Do mRNA Vaccines Cause Strongest Immune Response in Younger Individuals? Lipid Nanoparticles Offer Some Answers.” Drexel News, Drexel University, 31 Mar. 2023, drexel.edu/news/archive/2023/March/Why-do-mRNA-Vaccines-Cause-Strongest-Immune-Response-in-Younger-Individuals.

5. Immune Reaction: After the spike protein has been created in your cells, it is sent to the cell membrane where it is displayed for recognition by antibodies and immune cells. Your immune system will “remember” the spike protein and is able to quickly respond to infections with the SARS-CoV-2 virus. Think of the vaccination like a practice run for your body.

Chavda, V.P.; Soni, S.; Vora, L.K.; Soni, S.; Khadela, A.; Ajabiya, J. mRNA-Based Vaccines and Therapeutics for COVID-19 and Future Pandemics. Vaccines 2022, 10, 2150. https://doi.org/10.3390/vaccines10122150

It should be noted that there are several ways to manufacture mRNA vaccines. Other processes typically include the same steps in a slightly different order.

1. Identify Target Antigen Sequence – performed by pathogen surveillance teams via Next-Generation Sequencing (NGS).
2. Add Sequence to Plasmid DNA (pDNA) – the plasmid will carry the gene for the antigen, also called the “gene of interest” (GOI).
3. Multiply pDNA by Bacterial Fermentation – produces many copies of the plasmid (and many copies of the GOI).
4. Purify pDNA – isolate plasmid from cells, toxins, and other fermentation wastes.
5. Linearize pDNA – cutting the circular pDNA to form a linear DNA sequence. This will promote template uniformity and increase transcription efficiency.
6. IVT Reaction – transcribes gene for antigen (DNA) into mRNA drug substance.
7. Chromatography – isolate mRNA from pDNA, nucleotides, and other IVT wastes.
8. Encapsulation – surround mRNA with special lipid mix to produce lipid nanoparticles (LNPs), allows transport to cells.
9. Ultrafiltration/Diafiltration (UF/DF) Formulation – concentrating LNPs to achieve practical doses, removing wastes, and exchanging process buffer for a storage buffer with human compatibility. Achieved by Tangential Flow Filtration (TFF)
10. Final Filling – sterile filtration of drug product and loading into sterilized containers that will be sent to pharmacies.

The National Center for Therapeutics Manufacturing (NCTM) specializes in biomanufacturing training. If you’re interested in an mRNA training program, please email Programs@NCTMmail.tamu.edu.

The QC department performs tests on the mRNA vaccine products to gather multiple measurements related to the quality of the drug substance and the drug product. Drug substance represents the molecule with the main therapeutic action (also known as the active ingredient). Drug product is the final dosage form (i.e. it is the drug substance plus other molecules added for storage, stability, administration, etc.) that will be sent to pharmacies. The measurements from these tests will be recorded and sent to the QA department.

The QA department will review the results from the QC department and determine if the measured values meet specifications for the approved product. These specifications are known as Critical Quality Attributes (CQAs). CQAs are the measurable physical, chemical, or biological properties of the drug substance and drug product which must fall within acceptable ranges, distributions, or limits to be considered acceptable. If accepted, QA will release the product for pharmacies.

The graphic below lists relevant attributes and the methods used to measure them.

The National Center for Therapeutics Manufacturing (NCTM) specializes in biomanufacturing training. If you’re interested in an mRNA training program, please email Programs@NCTMmail.tamu.edu.

In the music video, we included a graph which compared the Phase III population sizes of several vaccines based on their clinical review documents. The clinical review documents are submitted to the FDA upon the completion of Phase III clinical trials and the submission of a Biologics Licensing Application (BLA). The clinical reviews can be downloaded via the FDA website or with the links below.

• COMIRNATY^®– Pfizer-BioNTech (COVID-19)
• SHINGRIX – GSK (Zoster)
• Fluzone^® High-Dose – Sanofi Pasteur (Influenza)
• SPIKEVAX – Moderna (COVID-19)
• Gardasil^® – Merck (HPV)
• PRIORIX – GSK (Measles, Mumps, Rubella)
• TENIVAC^® – Sanofi Pasteur (Tetanus, Diptheria)