A virus is a small collection of proteins and fats that can penetrate specific cell types to deliver a genetic payload into that cell. A virus is therefore said to be a vector for this genetic payload.
Before we answer this question let’s look at what a virus is.
A virus is a small collection of proteins and fats that can penetrate specific cell types to deliver a genetic payload into that cell. A virus is therefore said to be a vector for this genetic payload. Viruses deliver a genetic payload that, in the case of most wild-type viruses, reprograms the cells to make many copies of the virus. This is what makes many viruses so prolific as, unchecked by the immune system, one virus can create many thousands of copies of itself, rapidly spreading through the host organism, and as in the case of COVID-19, between people.
AZD1222 uses a virus to fight a virus. It uses a strain of virus called an adenovirus, that is common among chimpanzees and similar to one of the viruses that causes the common cold in humans. The team at Oxford University developed a way to remove the genetic instructions from this adenovirus and to replace this with new genetic instructions. When this new viral vector infects the targeted cells, rather than making copies of itself, it instructs the cell to manufacture a protein that is found on the surface of the SARS-CoV-2 virus that causes COVID-19. In the case of AZD1222 this protein stimulates a strong immune response which helps to prevent future infections.
So how is AZD1222 made?
The manufacturing process for viral vectors, whether used for the manufacture of vaccines or for delivering other genetic instructions, as in gene therapy applications, follows the same basic manufacturing method (Cells + food + virus – contaminants = millions of cells and trillions of viral vectors). It is often broken into upstream, downstream, and fill-finish operations.
Upstream: the first step is to generate a large number of special cells through a process called cell culture. This starts with a small number of mammalian cells that, when kept in the right sterile conditions, fed with food and oxygen, multiply, typically doubling in number every day.
When there are enough of these cells, they can then be infected (or more accurately transfected) with a virus that is engineered to program these cells to produce the viral vector described above.
Both the cells and the virus used are stored, frozen, in a working cell bank, in much the same way as a starter culture for sourdough bread making always starts from the same starter culture. If these working banks run low, they can be refreshed from a master bank in a process that is comparable to the feeding of the sourdough starter culture.
(This part of the manufacturing process typically takes about 3 weeks).
The faster the cells grow, and the faster the viral vectors are produced, the faster the manufacturing process and the more viral vectors that can be produced per batch. In addition to this, the total number of cells, the number of viral vectors, and the number of doses can also be increased by simply increasing the volume of starting cell culture. Typically, this can be from 200 L up to 2000 L for vaccines of this type and these volumes are capable of generating sufficient viral vectors for 1 million to 10 million doses per batch. The number of doses is directly linked to the dosing regimen for each vaccine, but this is a typical number to illustrate the scale of the process. The word ‘simply’ here is a misnomer. It does not reflect the ongoing challenge of upscaling product manufacture to maintain quality and consistent vaccine characteristics as volumes increase. In the context of vaccine manufacture any change is highly undesirable, and often forbidden. This scaling depends very much on process expertise, equipment knowledge, and a host of analytical tests that demonstrate equivalency at every stage in the process.
As well as scaling up, manufacturing can also be increased by scaling out. This refers to the number of duplicate processes that can be run in parallel. For example, an existing 200 L process can be scaled up to a single 1000 L process or a similar number of doses generated using 5 × 200 L processes running simultaneously.
Downstream: when the viral vectors are produced there are also a lot of unwanted contaminants that are left over that would be undesirable in the final vaccine. It is the role of the downstream process to remove these contaminants and to leave the highly purified viral vectors.
This occurs through a series of separation techniques that exploit differences in size, charge, and chemistry between the wanted viruses and the unwanted contaminants.
(This part of the manufacturing process typically takes about 1 week).
The end of this process delivers the bulk drug substance, typically in large flexible bags or plastic bottles ready for shipping to a different location for the next specialized operation. The starting volume of 200 to 2000 L is typically now 10 to 100 L but still enough for 1 to 10 million doses. This may now be formulated into bulk drug product with additional fluids to maximize the stability and to dilute the product to a manageable dosage. At this point the drug substance may also be frozen to safeguard against degradation and to prolong the shelf life. This takes place in carefully controlled freezers that consistently achieve the same rate of freezing and are safeguarded by rigorous temperature monitoring and control during the shipment to the next location.
The entire process is designed to consistently work exactly the same way, every time, to deliver exactly the same product with every batch. It is essential that the manufacturing process remain very clean, and often sterile throughout the upstream and downstream stages. The manufacturing environment is kept clean with highly controlled air filtration systems, rigorous operator cleanliness, such as head-to-toe gowns, facemasks, gloves and head coverings, and the strictest levels of cleanliness for anything that touches the process fluid or that directly feeds into the process.
At the heart of many modern pharmaceutical manufacturing processes is the application of single-use technology. While this is an undesirable option elsewhere in life, in the context of drug manufacture, the use of single-use technology significantly accelerates the development of the manufacturing process and is an off-the-shelf way to ensure the strictest degree of processing cleanliness. Processes that utilize re-usable fluid contact components and technology take many years to commission, build, and test before being put into service and then regularly retested to ensure that they remain clean. Also, in the context of drug manufacture, single-use technology is more environmentally friendly, with a lower carbon footprint when compared to alternative methods, due to the high energy demands of steam sterilization and smaller environmental impact due to the removal of large volumes of cleaning chemicals needed. But that is a subject that can be explored separately. Without single-use technology this, and many other COVID-19 vaccines would take far longer to develop.
At its most basic level, single-use processing can be as simple as replacing a stainless-steel tank, connected by lengths of steel pipework, with flexible bags connected by flexible tubing. However, the concept extends to include ways to safely (aseptically) join different processing elements as well as each of the key technologies used in the manufacturing process. Once designed to all fit together this means that a manufacturing process can quickly be copied, anywhere in the world with minimum additional design. For AZD1222 the single-use concept supports global scale-out and scale-up to deliver the capacity to produce billions of doses in multiple manufacturing locations.
Fill/finish: This is the part of the process that takes the drug substance, in bulk form and puts it into the final containers that are ultimately distributed to the locations where the vaccination will take place.
If the bulk drug substance is frozen, this must first be thawed in a carefully controlled manner.
From this bulk liquid, automated filling lines then transfer this liquid into the desired final container. This can typically be hundreds of thousands, if not millions of glass vials but may also be in prefilled syringes or a bulk container used for multiple doses.
Each single dose contains up to about 5 × 1010 (50 billion) viral vectors in less than < 0.5 mL of fluid.
This filling takes place in an even cleaner, sterile environment with the vaccine passing through filters that ensure absolute cleanliness of the vaccine and ultimately assuring the safety of the vaccine by excluding the possibility of any environmental contamination.
From this point, all paperwork is scrutinized across all sites to ensure that everything has been manufactured in accordance with the strictest of protocols. Only when every datapoint is confirmed as being acceptable is the product released and distributed, ensuring that strict temperature controls are in place at every stage along the way to the patients’ arms.