Towards Cruelty-free Botulinum Toxin Testing: Current Situation and Roadblocks

Abstract

Botulinum neurotoxin (BoNT), more commonly known as Botox, is a biological toxin produced by bacteria and is characterized by its paralytic effect on nerve cells. BoNT is used for medical applications such as muscle spasms but is also known for its use in the cosmetic treatment of facial wrinkles. Since BoNT is a biological product, the characteristics and potency of the toxin can differ between production batches. For this reason, potency tests need to be performed on each batch of BoNT product to ensure its efficacy and safety. For these potency tests, European guidelines require the use of a mouse LD50 test. In this mouse model, a dilution series of BoNT product is injected into the abdomen of the mice, and the concentration at which 50% of injected mice die is used to determine the potency of this particular batch of BoNT. Over the course of the test, mice will become unable to walk and eventually suffocate due to the failure of respiratory muscles. However, during the test mice often die of starvation or dehydration because they are unable to move to their food bowls and water taps because of muscle paralysis, making the test unreliable. Both outcomes inflict great suffering on the mice. It has been estimated that over 400,000 mice have died annually across the EU for BoNT potency testing during manufacturing. In 2011 Allergan was the first major BoNT manufacturer to develop and receive approval for a cell-based model for BoNT potency testing. Unlike other available alternatives to the mouse model, this cell-based model incorporates all crucial steps of the neurotoxin’s mechanism of toxicity. Different BoNT products use different ingredients which can cause problems if the assay is not adapted for these specific products, resulting in a time-consuming validation process. This contributed to the other two major BoNT manufacturers, Merz and Ipsen, taking years to validate a cell-based assay for their own products. Even though all three major BoNT manufacturers in Europe now use a cell-based assay to replace some amount of their animal testing, the number of mice used for BoNT potency testing had actually increased in the years leading up to 2019. This is likely the result of new manufacturers bringing their BoNT products to the EU market, EU regulatory guidelines stating that some quality control tests still need to be performed using the mouse model, and the total amount of produced BoNT increasing to keep up with rising consumer demand. Animal rights organizations are disappointed with this development, especially considering BoNT is commonly used for cosmetic treatments. Even though animal testing for cosmetics has been banned in the EU since 2013, BoNT products avoid this ban due to having genuine medical applications and are thus classified as medicines. However, in both the Netherlands and the rest of the EU, off-label prescription of BoNT for vanity reasons is common. Even though LD50 testing is prohibited in the Netherlands, BoNT products which were tested on mice are still marketed and sold there for cosmetic purposes. Since 2019, the EU has done little to help finalize the replacement of LD50 testing for BoNT. EU regulatory guidelines need to be updated to fully remove LD50 testing as part of the quality control process and until all BoNT products are ‘cruelty-free’, off-label prescription of these products for vanity reasons should be prohibited.