Observations
LD50 test on mice given a bad airing in public
Toxin manufacturers quick to show they are working for a more humane alternative
The testing of products on animals is mired in controversy in the UK. Not wanting to incur the disdain of the British public, who have long been known for their close affection for animals, botulinum toxin manufacturers responded quickly to reports from the British Union for the Abolition of Vivisection (BUAV) about the testing of their products on mice and the continued use of the LD50 test—the lethal dose at which 50% of the mice would be expected to die when injected with the toxin.
The issue was given a fresh round of publicity when celebrities Ricky Gervais and Chrissie Hynde joined the call to end the LD50 botulinum toxin test on mice. What reignited the contentious issue was a video shot undercover by the BUAV at a testing laboratory in Wickham, Hampshire. The video, The Ugly Truth about Botox, shown on the BUAV’s website, provides a grizzly account of the testing of the Dysport toxin on mice. Mice are injected with the toxin, suffer progressive paralysis, and some appear to suffer severe breathing difficulties before dying.
Sarah Kite, BUAV special projects director, said: "Our shocking findings show that crude, archaic and extremely cruel animal tests are still allowed in the UK even when an alternative test exists and animal testing is not required by official bodies."
There are a number of tests for evaluating toxicity, including the SNAP-25 assay, however, the LD50 assay is the mandatory test required by the European (EMEA), American (FDA) and UK (MHRA) regulatory authorities for a product license to be granted. Other tests can be used to determine toxicity but only to supplement, and not replace, the LD50 test.
Ipsen Biopharm, which manufactures Dysport, said it took the allegations "very seriously" and would be discussing with Wickham. "Animal welfare is of primary importance to Ipsen, and the approval of Wickham by the Home Office has been critical in Ipsen’s decision to work with this company," said a spokesperson.
Alternatives
Ipsen says its LD50 replacement research is focused on alternative testing methods such as endopeptidase activity assays (including SNAP-25), enzyme-linked immunosorbent assays (ELISA) and assays that measure the binding capability of the product to the neuromuscular junction target.
"Such research is governed by the importance of establishing the exact potency of all batches using standardised methodology because such potency will determine the precise efficacy and safety of each injection to guarantee patient safety," Ipsen says.
Allergan was quick to distance itself from the Wickham laboratory, saying it does not use it and "strives to minimise animal testing." Allergan says it shares the pharmaceutical industry’s goal of reducing, refining and, wherever possible, eliminating animal testing. A spokesperson said: "We have worked with the FDA and other worldwide regulatory agencies on revised testing methods that tighten acceptance criteria for product release, enabling Allergan to reduce the number of animals used in the test by approximately 50%."
The BUAV called for alternative tests such as the SNAP-25, which measures the activity of the toxin in a test tube.
But Ispen says the test is inadequate. "The SNAP-25 assay method developed by the UK National Institute for Biological Standards and Control for specific internal use has never been accepted for product quality testing and release by any worldwide regulatory authority, since it measures only one property of botulinum toxin."
Allergan points out that it is "investing heavily" to develop reliable alternative methods. "Over the last 10 years Allergan has invested more than $40 million in the development of alternative assays and are at the forefront of the industry in terms of finding alternative tests."
Merz says it has also been working to provide a replacement for the LD50 test, as well as refining the test with respect to animal protection. Merz says it is in regular consultation with regulatory authorities in Europe and the US to achieve this goal, and it is an active member of a European team of experts with the aim of implementing the 3R principles (reduce—refine—replace) for botulinum toxins.
Merz says it has developed and validated a method for antibody screening that has reduced the number of mice required by more than 90%. Over the last seven years this method has been deployed in all of the Xeomin clinical trials, Merz says.
