National Toxicology Program

Workshop Documents

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 Workshop Report Now Available [PDF]

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 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 authorities, and the animal protection community. The workshop goals were: (a) 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 and (b) to identify priorities for research, development, and validation efforts needed to advance the use of alternative methods (see the agenda for additional details).

Five workshop sessions covered a variety of topics including:

• An overview of public health needs for botulinum toxin testing and regulatory requirements
• An overview of the current understanding and knowledge gaps for botulinum toxin
• A review of methods as potential replacements or refinements for the current botulinum toxin test
• A review of strategies to reduce the number of animals used for the current botulinum toxin test

During the workshop, panel discussions were held to (a) review and supplement the background information presented during the speakers' presentations, (b) more precisely define the state-of-the-science, knowledge gaps and identify priorities for research, development, and validation efforts needed to advance the use of alternative methods for each area discussed, and (c) develop specific guidance and recommendations to advance the use of alternative methods for each area discussed.

In general, the consensus of the panel discussions was that the reviewed methods could be used, in specific circumstances or in a tiered-testing strategy, to reduce and 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 botulinum toxin test method. The panel discussions 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.