The Evolution of Vaccine Manufacturing – Past, Current, and Future Trends
The vaccine industry is a robust industry that is growing substantially. According to a 2013 report (1) from the World Health Organization (WHO), the market has quadrupled from $5B in 2000 to almost $24B in 2013 and is expected to rise to $100B by 2025. The influenza vaccine market alone was $2.9B in 2011 and is expected to rise to $3.8B by 2018. The vaccine industry is also a driver of growth in the global pharmaceutical market with 10-15% growth per year vs. 5-7% for pharmaceuticals.
One reason for the projected growth in vaccines is due to the more than 120 new vaccine products in the development pipeline. Some of these pipeline vaccines are poised to have a major impact on global public health by targeting very deadly or widespread viruses. Examples include the vaccines in development for the Ebola virus. These vaccines have been receiving extensive coverage and have been fast tracked as a result of last year’s outbreak in West Africa. Other significant vaccines in development include vaccines for dengue fever, H5N1 (bird flu), HIV, Herpes Simplex Virus, Epstein Barr (mononucleosis), chikungunya virus, and the Marburg virus.
With so many vaccines in the pipeline there is a strong focus on vaccine manufacturing, specifically increasing manufacturing efficiency, reducing cost, and maintaining safety. For the purposes of this blog, we will focus on the most prevalent vaccine type and manufacturing strategy – viral vaccines manufactured using mammalian cell culture. Currently there are 20 vaccines approved for the United States market that are produced by mammalian cell culture or in part by mammalian cell culture. (See Table 1)
|Vaccine||Type||Manufacturer||Cell Type||Target Population||Initial Approval|
|Adenovirus||Live||Barr Labs/Teva||WI-38||Military||March 2011|
|DTaP-IPV (Kenrix)||Inactivated (Polio)||GSK||Vero (microcarrier)||Children 4-6||June 2008|
|DTaP-HepB-IPV (Pediarix)||Inactivated (Polio)||GSK||Vero (microcarrier)||Infants, children 6 wks-6yr||Dec 2002|
|DTaP-IPV/Hib (Pentacel)||Inactivated (Polio)||Sanofi||MRC-5 (microcarrier)||Infants, children 6 wks-4yr||June 2008|
|Hep A (Havrix)||Inactivated||GSK||MRC-5||1yr+||Feb 1995|
|Hep A (Vaqta)||Inactivated||Merck||MRC-5||1yr+||March 1996|
|Hep A/Hep B (Twinrix)||Inactivated (Hep A)||GSK||MRC-5||18+||May 2001|
|Influenza (Flucelvax)||Inactivated, split||Novartis||MDCK (suspension)||18+||Nov 2012|
|Japanese Encephalitis (Ixiaro)||Inactivated||Intercell Biomedical/ Novartis||Vero||2 mo+||March 2009|
|MMR (MMR-II)||Attenuated||Merck||Chick Embryo (Measles, Mumps) WI-38 (Rubella)||1yr+||1971|
|MMRV (ProQuad)||Attenuated||Merck||Chick Embryo (Measles, Mumps) WI-38 (Rubella), MRC-5 (Varicella)||1-12 yr||Sept 2005|
|Polio (IPV-Ipol)||Inactivated||Sanofi||Vero (microcarrier)||6 wks+||Updated Production Process 1988|
|Rabies (Imovax)||Inactivated||Sanofi||MRC-5||All ages||Dec 2005|
|Rabies (RabAvert)||Inactivated||Novartis||Chicken fibroblast||All ages||Oct. 2007|
|Rabies Vaccine Adsorbed (RVA)*||Inactivated||BioPort Corp||Unknown||Unknown||1988|
|Rotavirus (RotaTeq)||Attenuated, reassortments||Merck||Vero||6-32 wks||Feb 2006|
|Rotavirus (Rotarix)||Attenuated||GSK||Vero||6-24 wks||Apr 2008|
|SmallPox (Vaccinia, ACAM2000)||Live||Sanofi||Vero||Military||Aug 2007|
|Varicella (Varivax)||Attenuated||Merck||MRC-5||1yr+||March 1995|
Mammalian Cell Culture Based Vaccine Manufacturing Trends
When using mammalian cell culture to manufacture vaccines, cell line selection is a critical first step. Researchers look for lines that do well in culture and have a good safety profile, i.e. lack of tumorigenic potential and limited oncogenic potential, and a history of safety in vaccine manufacturing. Initially, diploid cells were favored for production because of safety due to their non-tumorigenic potential in animals. This includes MRC-5 and WI-38 human fibroblast cell lines.
The Vero cell line is a newer technology. The cell line gained approval in the 1980’s with the production of Polio vaccines and now has an established history of safety and is widely accepted for vaccine manufacturing. The WHO has even established a certified source of Vero cells for distribution to vaccine manufacturers to encourage their use. Vero cells have the advantage of being continuous, without a limited cell doubling capacity, which provides a significant advantage in cell culture. They are also non-tumorigenic and more readily adapt to modern microcarrier based production. FDA documents show that the Polio Vaccine uses Vero cells in a microcarrier process. Vero cell based manufacturing currently represents 7 of the 20 mammalian cell culture based vaccines and many other vaccines in the pipeline.
The latest mammalian cell line to be used in an approved vaccine is the Madin Darby Canine Kidney (MDCK) line that has been adapted to suspension culture, used in the manufacture of Flucelvax by Novartis. In addition, the PER.C6 human cell line is also being used in some pipeline vaccines and has been documented as being used in at least one of the Ebola vaccines in development.
Cell Culture Media
While biopharmaceutical CHO cell culture based manufacturing has made continual improvements over the years, including advances in creating defined media formulations; vaccine cell culture remained relatively unchanged with several media formulations utilizing a combination of classic media and serum. The optimization of cell culture media is one area where improvements can be made in vaccine manufacturing. The removal of animal products and creation of a robust, defined and customizable media, which maximizes output per cell and earlier harvest times, would be ideal.
One area that has been under development is to remove serum from vaccine manufacturing. The latest cell culture based influenza vaccines to be approved, Flucelvax and Flublok, both use a serum free process. New technologies including animal-free media ingredients that enhance the growth of vaccine cell lines could be employed to either reduce or eliminate the need for serum in new vaccines or in reformulations of approved vaccines. Some of these technologies include, recombinant human serum albumin, recombinant human transferrin, recombinant growth factors and specialty products such as InVitria’s Zap-SR serum reducer.
Another area where vaccine manufacturing is evolving is in the use of newer vessels, including bioreacators. Bioreactors are currently being used in vaccine manufacturing and their use will likely continue to expand. Bioreactors can provide cells the optimum environment, which can lead to an increase in productivity and reduced overall cost. While many vaccines are still manufactured using static culture or roller bottles, several studies have shown that manufacturing can be greatly improved by employing bioreactors.
Bioreactor technologies for vaccine manufacturing include:
Microcarriers and stainless steel or single-use stirred tank bioreactors
Bioreactors can be used when cells are in suspension or adherent by employing another technology – microcarriers. Microcarriers enable more cells per milliliter of culture by expanding the surface area for cell proliferation. More cells in culture increases overall vaccine titer (yield). With an increase in the use of stirred-tank or single-use bioreactors, microcarrier innovations have been necessary to allow adherent cells to be cultured in these conditions. In addition, the use of microcarriers allow manufacturers to increase the number of cells that can be cultured in one tank enabling more efficient large-scale production and permitting the use of greater than 1,000 liter bioreactors.
Fixed bed bioreactors
Another type of bioreactor being utilized in vaccine manufacturing is a disposable fixed-bed bioreactor. These bioreactors contain multiple individual, hydrophilized microcarriers. Each of these has a surface area and is compacted into a fixed bed. These fixed-bed matrices are then incorporated within a bioreactor. An internal centrifuge pump within the bioreactor ensures a homogeneous distribution of media and cells throughout the fixed bed.
Hollow fiber bioreactors
Hollow fiber bioreactors have also been used to manufacture viruses. Hollow fiber bioreactors employ perfusion culture with semipermeable hollow fibers in a cartridge that contains inlet and outlet ports. The media flows through these fibers providing nutrients to cells. Spent media can then be filtered and replaced with fresh media or oxygenated and returned.
Single use technologies and flexible facilities
In addition to expanding the use of bioreactors, vaccine manufacturing is also looking for improvements by employing single-use technologies, where possible, to reduce capital investment and speed up the cleaning and validation time. The introduction of single-use technologies has enabled manufacturing facilities that are more “flexible” thereby improving a facility’s utilization. While flexible facility layouts are dependent on specific manufacturing needs, the principle of flexible design integrates the ability to move equipment around as needed to provide the type of manufacturing necessary for each product. If more product is required, more single-use bioreactors or larger ones can be plugged into the manufacturing design. Because this equipment is portable it enables one size to be moved in while another is moved out or another is added. Mixers, single-use bioreactors, tanks, totes and other equipment are simply moved around to meet the manufacturing needs.
Some companies opt to employ two adjustable suites in manufacturing, each with similar equipment that can be moved in or out based on need. At a recent conference, one company described their two adjacent suites; one fitted for viral vaccines the other for antibodies. They utilized a similar layout, equipment and process design for each suite which allowed employees to easily move from one manufacturing suite to the other as demand dictated with little to no training or adjustment on their part. This company used a variety of single-use bioreactors in the 50 liter to 2,000-liter scale that they could employ based on demand.
Transitioning to new manufacturing technologies – the Influenza Vaccine as an example
On November 20, 2012, the Food and Drug Administration (FDA) announced that it approved the use of Flucelvax manufactured by Novartis to prevent seasonal flu in people over age 18. This approval was the first for a cell culture based seasonal influenza vaccine in the United States. Instead of using the traditional chicken egg production method, Novartis propagates the virus in Madin Darby Canine Kidney (MDCK) cells. They also employ a serum-free process. There are many advantages to using a cell culture-based manufacturing system over egg-based production including that it is quicker to manufacture, efficiently scalable and offers more control over the manufacturing environment.
In 2013, another cell culture based influenza vaccine was approved. On January 16, 2013, the FDA approved Flublok, manufactured by Protein Sciences Corporation. Flublok utilizes recombinant DNA technology and a baculovirus in insect cell expression system, also serum free. The insect cells are used to manufacture recombinant hemagglutinin, a flu virus vaccine antigen. This technology offers similar advantages in that it can be used to manufacture influenza vaccines much faster than traditional egg based production. In addition, it doesn’t require availability of the influenza virus, just the genetic code.
These two influenza vaccines are good examples of the direction of vaccine manufacturing as a whole. Increased manufacturing speed, scalability, and defined production.
Viral outbreaks like the West Africa Ebola outbreaks or the Disneyland measles outbreak are reminders of the importance of vaccines to public health. Vaccines have been responsible for the reduction and eradication of some of the worst diseases in human history and innovation in vaccines is critical as concerns grow about potential public health threats such as the Ebola virus, H5N1 (avian flu), pandemic strains of influenza and others. Just as it is important to continue to develop new vaccines, it is also critical that we continue to improve the way we manufacture these vaccines. Reducing cost and improving safety are always key drivers in vaccine manufacturing and will continue to drive manufacturing innovations now and in the future.