Shark-Sourced Vaccine Ingredient May Be on Its Way Out

Adjuvants are substances that can be added to vaccines to enhance the body’s immune response, essentially helping the vaccine to “work better”. The immune “boost” provided by adjuvants is useful for immunizing pregnant, infant or elderly populations that have unique immune system characteristics.

You may be familiar with aluminum salts – such as aluminum hydroxide, aluminum phosphate and aluminum potassium sulfate – as being the most well-known adjuvants, due to their safe use in vaccines for over 70 years. A perhaps lesser-known adjuvant product is squalene, a natural terpene that can be obtained from botanical sources but is of the highest purity when derived from shark liver.

“The history of adjuvants has been quite an empirical endeavor and squalene was one of those things, along with other substances that were identified several decades ago, as enhancing vaccine responses,” says Dr. Christopher Fox, affiliate associate professor in Global Health and vice president of Formulations at the University of Washington, and a synthetic biologist at the Seattle-based biotechnology nonprofit Access to Advanced Health Institute.

Fox says that the discovery of squalene as an adjuvant was “serendipitous”. He offers a fun fact: “A little bit about squalene – you put six isoprene units together, so it’s a linear triterpene. It’s interesting in that when it is in an emulsified form, formulated appropriately, it’s known to enhance immune responses to vaccines. Whereas other oils, like triglyceride oils that you might find commonly in vegetable-based sources, do not enhance immune responses to vaccines.”

Since the 1990s, it has been used in hundreds of millions of vaccine doses, including seasonal influenza vaccines and pediatric vaccines. “In more recent times, there are squalene-based adjuvants in licensed COVID-19 vaccines – not in the United States, but in other countries. It is a widely used ingredient,” Fox describes.

Beyond vaccine adjuvants, squalene has swiftly become a highly sought after ingredient for cosmetic products due to its hydrating effects on skin and its anti-inflammatory properties. Whether the squalene found in a cosmetic product is shark-derived can be unclear as some companies may not specify where the squalene is sourced from on their labels.

Protection achieved in the short-term, but a long-term pain persists

In recent years, calls for an alternative to shark-derived squalene use in vaccine and cosmetic products have grown louder. “Developing vaccines with shark liver oil may help protect people in the short-term but it comes with a long-term pain for shark sustainability,” says Glenn Sant, senior advisor on Fisheries Trade and Traceability at TRAFFIC, a global non-governmental organization that monitors the trading of wild animals and plants. “It is a no-win situation for the wild species involved, nor the long-term availability of the ingredients for something as important as lifesaving vaccinations, unless sustainability is at the fore front of our thinking.”

Fox says that there has been some encouraging science in recent years, particularly in the cosmetic industry, where substitutes for squalene-like molecules that are not derived from sharks are making headway. “We hope that carries over into the pharmaceutical industry,” he expresses.

Beyond its impact on shark welfare, exactly how shark-derived squalene works so effectively as an adjuvant is not totally clear according to Fox. This, coupled with its complex chemical structure, has made it challenging to synthesize using traditional organic chemistry techniques. Fox is interested in sourcing sustainable alternatives and the structure–function relationship of squalene analogues. “Part of the value of generating new analogues and evaluating their biological activity is that it helps inform how the structure of this class of molecules relates to their function. That information can then be used to optimize adjuvant activity,” he describes.

Searching for alternatives to shark-derived squalene

Fox and colleagues recently published a study in npj Vaccines, where over 20 squalene analogues were chemically generated from β-farnesene, a compound produced via the fermentation of yeast. The work was conducted in collaboration with Amyris, a synthetic biotechnology and renewable chemical company based in California. β-farnesene is also a terpene that is naturally produced in many different plants, but at small concentrations. That’s where Amyris offers a helping hand, with its “precision fermentation” approach.

Taken from Technology Networks.

“You can think about it as an advanced version of the way beer has been made for ages. We program yeast – like you would program a computer – and feed it sugarcane-derived sugar, which results in molecules – like β-farnesene – that are bioidentical to those traditionally sourced from nature,” says Chris Paddon, technical liaison director at Amyris.

The researchers took advantage of the availability of β-farnesene, which contains 15 carbon atoms, to create analogues of squalene, which contains 30 carbon atoms. By switching up the compound’s chemistry, the team were able to generate over 20 different analogues, all of which carry similarities to naturally-derived squalene, and some differences.

“Four or five molecules showed enhanced activity compared to shark-derived squalene, another handful showed comparable activity and a further handful showed decreased activity. These were not aligned with the stability of the emulsion, which was interesting. The length of the chain matters. So, you can have linear molecules that go too short, and you begin to lose biological activity, whereas longer chains may increase activity. The saturation of the chain is also important. Stronger structural changes in the center of the molecule may also affect the biological activity,” Fox explains.

From sharks to yeast

The initial data is promising, but the hunt for a sustainable alternative isn’t over just yet. All adjuvant products approved for human use undergo the exact same clinical testing protocols as the vaccines in which they are added to – and it is (rightly so) a thorough process. “The safety profile needs to be assessed in well-established preclinical models. That is an important next step for the research,” says Fox. In addition to safety evaluations, the analogues will need to be assessed for how well they protect against diseases in well-established preclinical models.

Alongside preclinical testing, manufacturing process development and characterization will have to be mapped out and tested in order for the analogues to reach human clinical trials.

Fox sounds enthusiastic when asked about these challenges: “The β-farnesene precursor is available at industrial scale, and then the synthetic chemistry steps to arrive at the molecules described in the manuscript are just a handful of steps – 3 to 4 – not 30+,” he says. “It’s also very standard, so a pharmaceutical chemistry facility could perform the steps and then there would need to be a final purification to ensure that they are pure enough for pharmaceutical use.”

Dr. Christopher Fox and Chris Paddon were speaking to Molly Campbell, Senior Science Writer for Technology Networks.

Reference: Fisher KJ, Kinsey R, Mohamath R, et al. Semi-synthetic terpenoids with differential adjuvant properties as sustainable replacements for shark squalene in vaccine emulsions. npj Vaccines. 2023;8(1):14. doi:10.1038/s41541-023-00608-y.