National Toxicology Program

ICCVAM-NICEATM/ECVAM Scientific Workshop on Alternative Methods to Refine, Reduce and Replace the Mouse LD50 Assay For Botulinum Toxin Testing Nov. 13-14, 2006      Silver Spring, MD Read the Workshop Report NIH Publication No. 08-6416 View detailed workshop agenda View presentations and other workshop materials

In response to a test method nomination from the Humane Society of the United States (HSUS), ICCVAM, in collaboration with NICEATM and ECVAM, co-sponsored a Scientific Workshop on Alternative Methods to Refine, Reduce, and Replace the Mouse LD₅₀ Assay for Botulinum Toxin Testing. The 116 attendees from 9 countries included scientists from government and academic institutions, regulatory agencies, and the animal protection community.

The goals of the workshop were:

• To review the state-of-the-science and current knowledge of alternatives that may reduce, replace, and refine (less pain and distress) the use of mice for botulinum toxin testing;
• To identify priorities for research, development, and validation efforts needed to advance the use of alternative methods.

Specific objectives of the workshop included:

• Review the public health needs for botulinum toxin testing, including the necessity to determine the safety and efficacy of products containing botulinum toxin;
• Review the current state-of-the-science and identify knowledge gaps regarding botulinum toxin structural aspects, mechanisms, and modes of action that are important to the development of alternative methods for in vivo botulinum toxin tests, and prioritize future research initiatives that would address these knowledge gaps;
• Review current development and/or validation status of alternative test methods for in vivo botulinum toxin tests and their potential to reduce, refine (less pain and distress), or replace the use of the mouse LD₅₀ assay;
• Identify alternative methods that should have the highest priority for future development and validation studies to assess potency/toxicity of botulinum toxin.

Workshop Outcomes

In general, the panel's consensus was that the reviewed methods could be used, in specific circumstances or in a tiered-testing strategy, to reduce or refine the use of mice in the current botulinum toxin test method. However, none of the reviewed methods could currently be used as a complete replacement for the current botulinum toxin test method. The panel noted that with additional development and validation efforts, one or more of the reviewed methods might be useful as a replacement for the current botulinum toxin test methods in the future. It was noted that additional validation studies were needed for most methods (e.g., comparison of results to in vivo outcomes, validation of methods for the specific applications). Finally, some best practices discussed to decrease the number of animals tested for studies included (a) use of reference standards to minimize the number of replicate animals needed, (b) use of standardized methodology, and (c) reduction in the number of doses tested for assays where confirmation of potency is being evaluated.

Background

In October 2005, the Humane Society of the United States (HSUS) submitted a test method nomination entitled Alternative Methods to Replace the Mouse LD₅₀ Assay for Botulinum Toxin Potency Testing. In the nomination, the HSUS proposed a review of the validation status of available in vitro, ex vivo, and non-lethal in vivo test methods as potential replacements for the current botulinum toxin potency test method. The nomination also requested that ICCVAM work with partners and stakeholders to validate one or more of the alternative test methods available. The HSUS nomination was reviewed by both ICCVAM and SACATM in November to December 2005. Both committees endorsed the sponsorship of a workshop on alternative botulinum toxin test methods with high priority.

Related Information

HSUS Nomination : Nomination of Alternative Methods to Replace the Mouse LD₅₀ Assay for Botulinum Toxin Potency Testing (October 2005)
View relevant HSUS letter to Allergan, Inc. (Dec. 15, 2005)