# Characteristics of Botulinum Neurotoxins the Determine Potency

> **NIH NIH R01** · UNIVERSITY OF WISCONSIN-MADISON · 2024 · $763,801

## Abstract

Botulinum Neurotoxins (BoNTs) are a large family of protein toxins that possess extreme potency and cause
severe disease in humans and animals. Botulism is a neuroparalytic disease of long duration, lasting up to
several months. Without proper medical care, naturally occurring botulism is lethal in up to 50% of cases, and
even with supportive care and antitoxin administration, botulism is a devastating and severe disease and
remains lethal in ~ 5 % of cases. While naturally occurring botulism is rare, BoNTs are classified as a Tier 1
Category A Select Agents due to their threat as potential bioterrorist weapons and severity and long duration of
the disease. Conversely, BoNTs are widely used human biotherapies to treat more than 200 neuromuscular
disorders, some of which are devastating without this unique treatment. BoNTs are divided immunologically
into seven BoNT serotypes (A-G), which are further subdivided into subtypes. For example, there are eight
subtypes of BoNT/A. Hundreds of BoNT and BoNT-like variants have been identified by sequencing efforts, but
only few variants have been investigated for potency and duration of action at the protein level. Note, only two
subtypes, BoNT/A1 and BoNT/B1 are currently used as therapeutics. Our recent studies have used BoNT/A
subtypes to determine, for the first time, the basis for long duration of action, stable association of LC/A1 on
the intracellular plasma membrane, and mechanisms for the high BoNT/A1 potency. This renewal will
determine detailed molecular mechanism for the specific durations of action of BoNT serotypes that elicit
natural human botulism and LC targeting of SNAP-25 on the plasma membrane. A novel mRNA-based BoNT
Light Chain expression system will be used to standardize the determination of BoNT duration of action and
potency. Translational studies will develop long duration variants of other BoNT serotypes as alternates for
BoNT/A1 as a therapeutic agent to overcome BoNT/A specific treatment resistance. The collaborative efforts of
the Pellett and Barbieri laboratories combine computational, molecular, and cellular approaches with BoNT
studies by mouse bioassays and in human and rodent cell-based assays, including human motor-neurons.
These studies will use native BoNTs, recombinant BoNTs produced in native expression hosts, and individual
subunits to assess the two most important aspects of BoNTs, duration of action and potency. A streamlined
approach will first investigate subunit domains in functional studies and select specific alterations for the more
effort- and cost-intensive construction and analyses of holotoxins. Completion of these studies will provide a
detailed understanding of the mechanisms underlying BoNT potency, where in addition to cell entry and
catalysis, intracellular LC trafficking and membrane association are contributing factors. The molecular
concepts identified in these studies can be extrapolated to other protein toxins.

## Key facts

- **NIH application ID:** 10980625
- **Project number:** 2R01AI139306-06A1
- **Recipient organization:** UNIVERSITY OF WISCONSIN-MADISON
- **Principal Investigator:** Joseph T Barbieri
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $763,801
- **Award type:** 2
- **Project period:** 2019-01-18 → 2029-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10980625

## Citation

> US National Institutes of Health, RePORTER application 10980625, Characteristics of Botulinum Neurotoxins the Determine Potency (2R01AI139306-06A1). Retrieved via AI Analytics 2026-06-11 from https://api.ai-analytics.org/grant/nih/10980625. Licensed CC0.

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