Vaccines and Vaccine Adjuvants

Written by Maja Hunter, Ph.D.

Not too long ago, during the COVID-19 pandemic that started at the end of the year 2019, vaccination was proven once again to be an important preventive tool against infectious diseases.

The beginnings of vaccination date back to May 1796. An English physician, Dr. Edward Jenner, knowing the theory that humans infected beforehand with cowpox were protected against smallpox, inoculated an 8-year-old boy with a matter collected from a cowpox sore on the hand of the milkmaid. After several days of feeling unwell and suffering a local reaction, the boy recovered and was two months later resistant to smallpox [1]. Dr. Edward Jenner proved the concept of vaccination (lat. vacca – cow). Since then, scientists have expanded knowledge about the immune system, and how it recognizes and reacts upon detection of anything foreign.

Specific antibodies are made to recognize antigens

An antigen is a marker that notifies your immune system if your body is dealing with anything that is not recognized as a part of the body. Antigens, usually proteins or sugars (polysaccharides), are found on the surface of viruses, bacteria, tumors and normal cells of your body [2]. Unknown antigens are recognized by the immune system as a treat, and custom-made antibodies are designed against them (Figure 1). During the first encounter with a specific antigen, the immune system learns and creates a defense strategy (specific antibodies). Moreover, upon reencounter with that pathogen, presenting that specific antigen, the immune system immediately reacts and defends the body against the infection – without you even noticing it.

Figure 1. An antibody is made to respond to specific antigens [3].

Recombinant antigens for vaccine production

The main components of the vaccines are antigens. Native antigens are extracted from the original source in their natural form, whereas recombinant antigens are manufactured artificially. To produce recombinant antigens, a vector expressing the protein of interest is transformed into a suitable heterologous expression system, e.g., E. coli, with well-established culturing conditions and all the “gene tools” easily accessible.

The advantages of recombinant proteins vs. native proteins are:

larger and consistent production yield;
easier and faster purification; purer final product;
well-established production protocol and time savings because of improved purity and consistency of the product, which makes recombinant protein production cheaper;
accessibility – heterologous expression systems produce high levels of desirable antigens; scaling up is a well-established process [4].

If produced protein is expressed inside of the host cell, the cell has to be disrupted in order to access, isolate and purify the overexpressed antigen.

High-shear processing is an optimal method for cell disruption. High pressure forces cell suspension through narrow microchannels, reaching high velocities and experiencing high shear rates. Warming up of the sample is inevitable but Dyhydromatics ShearJet® processors have a heat exchanger positioned just after the Reaction Chamber® module, which efficiently cools down the cell suspension. Most importantly, temperature-sensitive proteins retain their activity.

Pneumococcal polysaccharide vaccines prepared using high-shear processing technology

The pneumococcal polysaccharide vaccines (PPSV) are important to protect the vaccinated individuals from bacteria Streptococcus pneumoniae, causing pneumonia, meningitis, otitis media and bacteremia. The vaccines are made from purified polysaccharide capsules acting as antigens [5]. A single bacterial species can exist in many variants, called serotypes, distinguished by the humoral immune response [6]. PPSV23 contains polysaccharide capsules of 23 different serotypes. Other existing versions with reduced number of serotypes, such as PPSV7, PPSV14 and others, were made for the younger population or individuals with medical condition [5].

Reduction of the polysaccharide molecular weight can be achieved by chemical or mechanical methods. The chemical method is suitable for only certain polysaccharides, e.g., pneumococcal serotypes 3, 6A, 8 and 12F, which do not contain non-saccharide substituents, such as O-acetyl, glycerol phosphate, pyruvyl, etc. [7]. However, a mechanical method for polysaccharide molecular weight reduction, such as high-shear processing, is indispensable for polysaccharides containing non-saccharide substituents [7]. Chemical treatment of such polysaccharide chains would alter the chemical structure of the molecule or its attached groups, which would result in diverse polymers. Importantly, for the preparation of pneumococcal polysaccharide vaccines, the homogenous polysaccharide mixture is required for the immune system to build a reliable defense mechanism.

Processing the serotype 24F on our pilot scale ShearJet® HP350 processor

No matter what equipment are you using for reducing the molecular weight (Mw) of polysaccharides, it will take a few passes before the desired size range will be reached. We used the opportunity and compared the capabilities of the Dyhydromatics ShearJet® HP350-30 high-shear processor with the table top homogenizer GEA Panda 2000plus. The desired molecular weight range was 100-150 kDa.

Table 1. Processing of serotype 24F on GEA Panda 2000plus – high-pressure homogenization

GEA Panda 2000plus

Table 2. Processing of serotype 24F on Dyhydromatics ShearJet® HP350 – high-shear homogenization

ShearJet® HP350

As you can see in Table 2, the desired results were achieved in fewer than 35 passes with the Dyhydromatics ShearJet® HP350 processor. GEA Panda 2000plus would need over 70 passes to achieve the goal (Table 1). High-shear homogenization with fixed-geometry proved to be a more efficient method for polysaccharide molecular weight reduction than high-pressure homogenization.

The size of the equipment – lab vs. pilot scale, does not play any role in achieving the results. The Dyhydromatics Reaction Chamber® technology ensures linearly scalable processing of any sample. The lab scale ShearJet® HL60 utilizes the Reaction Chamber® module with one microchannel, whereas for pilot or production scale, the Reaction Chamber® modules utilize multiple microchannels in parallel to compensate for higher flow rate demand of the sample being processed.

Vaccine adjuvants

Frequently, antigens are not the only component of the vaccines. Vaccine adjuvants are used to stimulate and enhance the magnitude and durability of the immune system [8]. The advantages of using vaccine adjuvant are the reduction of antigen amount per vaccine dose, and the number of vaccination sessions. Additionally, adjuvants may increase the stability of an antigen, extend its half-life and indirectly improve its immunogenic power [9].

Most common adjuvants included in vaccines are aluminum salts but recent research introduced a few more with effective adjuvant properties and improved safety. Nanotechnology and molecular biology entered into the production processes of both antigen and adjuvant components, which resulted in improved vaccine efficacy. Currently, microparticles, emulsions, liposomes [10] and immune stimulators are seen as the most promising adjuvants in vaccine production [9].

How do Dyhydromatics high-shear processors fit into vaccine and adjuvant production?

High-shear processors are versatile and indispensable for production of certain types of vaccines and their adjuvants. The high-pressure pump generates high pressure that forces the sample through the microchannels of the Reaction Chamber® module. Due to the shear and impact forces, the temperature of the sample raises 1.7 °C per every 1,000 psi (70 bar). As mentioned above, our ShearJet® processors boast efficient cooling of the sample. Just after the sample exits the Reaction Chamber® module, it enters the heat exchanger to quickly cool down. Efficient cooling is important to prevent coalescence of nanoemulsion droplets, as well as preserving the protein activity.

Fixed-geometry technology ensures constant pressure for the complete sample volume and therefore reproducible and consistent results. Depending on selected processing conditions, such as pressure, number of passes and Reaction Chamber® module type, you can obtain the desired particle size with narrow particle size distribution. Importantly, you can use the same Dyhydromatics ShearJet® processor for cell disruption to access the antigen and facilitate its purification, for polysaccharide molecular weight reduction and nanoemulsion droplet reduction. You only have to exchange the Reaction Chamber® module and processing can start. For polysaccharide molecular weight reduction and for cell disruption the Reaction Chamber® 87.1 L-type is required, whereas Reaction Chamber® T-type is best for droplet reduction of emulsions and liposomes.

We offer proof-of-concept testing to ensure your application is suitable for our equipment and desired particle sizes are achievable. If you want to learn more about our ShearJet® processors and our technology, do not hesitate to reach out to our friendly customer service. We look forward to answering your questions. Visit our website at www.dyhydromatics.com.

Literature

[1] World Health Organization. A Brief History of Vaccination. [Retrieved July 13th, 2023]. Website: https://www.who.int/news-room/spotlight/history-of-vaccination/a-brief-history-of-vaccination

[2] "Antigen". Cleveland Clinic. 2023. Retrieved July 26th, 2023. Website: <https://my.clevelandclinic.org/health/diseases/24067-antigen>.

[3] “Antibody”. National Human Genome Research institute. 2024. Retrieved January, 8th, 2024. <https://www.genome.gov/genetics-glossary/Antibody>.

[4] Why choose a recombinant antigen? Ana Camacho. Retrieved July 26th, 2023. Website: <https://www.rekombiotech.com/en/blog/why-recombinant-antigen>.

[5] Daniels CC, Rogers PD, Shelton CM. A Review of Pneumococcal Vaccines: Current Polysaccharide Vaccine Recommendations and Future Protein Antigens. J Pediatr Pharmacol Ther. 2016 Jan-Feb;21(1):27-35. doi: 10.5863/1551-6776-21.1.27.

[6] Simon-Loriere, E., Schwartz, O. Towards SARS-CoV-2 serotypes?. Nat Rev Microbiol 20, 187–188 (2022). Doi: 10.1038/s41579-022-00708-x

[7] Prasad, A. Krishna, Kim, JJ. and Gu, J. Design and Development of Glycoconjugate Vaccines. Carbohydrate-Based Vaccines: From Concept to Clinic. ACS Symposium Series Vol. 1290, Chapter 4, p. 75-100 (2018). Doi: 10.1021/bk-2018-1290.ch004

[8] Pulendran, B., S. Arunachalam, P. & O’Hagan, D.T. Emerging concepts in the science of vaccine adjuvants. Nat Rev Drug Discov 20, 454–475 (2021). Doi: 10.1038/s41573-021-00163-y

[9] Facciolà A, Visalli G, Laganà A, Di Pietro A. An Overview of Vaccine Adjuvants: Current Evidence and Future Perspectives. Vaccines (Basel). 2022 May 22;10(5):819. doi: 10.3390/vaccines10050819.

[10] Li M, Kaminskas LM, Marasini N. Recent advances in nano/microparticle-based oral vaccines. J Pharm Investig. 2021;51(4):425-438. doi: 10.1007/s40005-021-00537-9.

Posted on January 25, 2024