Measurement of Botulinum Toxin Activity: Evaluation of the Lethality Assay

Pearce, L. Bruce; Borodic, G E; First, E R; MacCallum, R D

doi:10.1006/taap.1994.1181

Cited by 114 publications

(69 citation statements)

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“…9 This independent work by NIBSC confirmed that the unit ratios are similar to those obtained for the marketed products as described by other authors, 7,8,10 but the interlaboratory variation remained substantial even when the same assay protocol was employed.…”

supporting

confidence: 83%

Current Evidence on the Unit Equivalence of Different Botulinum Neurotoxin A Formulations and Recommendations for Clinical Practice in Dermatology

Karsai

Raulin

2009

Dermatologic Surgery

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BACKGROUND The unit equivalence between the two main Botulinum neurotoxin A (BoNTA) preparations, Dysport (Ipsen Ltd., Slough, Berkshire, UK) and BOTOX (Allergan Inc., Irvine, CA), is a matter of discussion. The UK assay used to test Dysport is more sensitive than the U.S. assay used for BOTOX, resulting in a different efficacy per unit in both formulations. Ratios ranging from 6:1 to 1:1 can be found in the literature, but the more recently published literature suggests that 1 unit of BOTOX is equivalent to approximately 2 to 4 units of Dysport (ratio 2:1–4:1). OBJECTIVE Because the number of BoNTA treatments is constantly increasing, these differences warrant a systematic review of published evidence about the unit equivalence of UK and U.S. formulations. METHODS The review is based on a detailed literature research in all relevant databases (MEDLINE, PubMed, Cochrane Library, specialist textbooks). RESULTS The present review supports the recent assumption that dose ratios of less than 3:1 (e.g., 2.5:1 or even 2:1) between Dysport and BOTOX are probably more suitable. CONCLUSIONS The current evidence is still insufficient, and further investigation of lower dose ratios is recommended.

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supporting

confidence: 83%

Current Evidence on the Unit Equivalence of Different Botulinum Neurotoxin A Formulations and Recommendations for Clinical Practice in Dermatology

Karsai

Raulin

2009

Dermatologic Surgery

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“…intraperitoneal (ip) LD 50 method [18]. The mouse ip LD 50 was determined using a seven-serial dose with a dilution interval of 1.25 and 20 mice per dose.…”

Section: Methodsmentioning

confidence: 99%

Comparison of Systemic Toxicity between Botulinum Toxin Subtypes A1 and A2 in Mice and Rats

Torii

Goto

Nakahira

et al. 2014

Basic Clin Pharma Tox

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The adverse events caused by botulinum toxin type A (subtype A1) product, thought to be after-effects of toxin diffusion after high-dose administration, have become serious issues. A preparation showing less diffusion in the body than existing drugs has been sought. We have attempted to produce neurotoxin derived from subtype A2 (A2NTX) with an amino acid sequence different from that of neurotoxin derived from subtype A1 (A1NTX). In this study, to investigate whether A2NTX has the potential to resolve these issues, we compared the safety of A2NTX, a progenitor toxin derived from subtype A1 (A1 progenitor toxin) and A1NTX employing the intramuscular lethal dose 50% (im LD 50 ) in mice and rats and the compound muscle action potential (CMAP) in rats. Mouse im LD 50 values for A1 progenitor toxin and A2NTX were 93 and 166 U/kg, respectively, and the rat im LD 50 values were 117 and 153 U/kg, respectively. In the rat CMAP test, the dose on the contralateral side, which caused a 50% reduction in the CMAP amplitude, that is, CMAP-TD 50 , was calculated as 19.0, 16.6 and 28.7 U/kg for A1 progenitor toxin, A1NTX and A2NTX, respectively. The results indicate that A2NTX is safer than A1 progenitor toxin and A1NTX.Botulinum toxins act on neuromuscular junctions and inhibit the release of acetylcholine from pre-junctional membranes, inducing muscle relaxation [1][2][3][4]. The toxins have been used as important therapeutic agents for neurological disorders [5][6][7]. There are seven subtypes of type A organism, A1-A7, based on variations in the amino acid sequence of the neurotoxin produced [8][9][10], and all botulinum toxin type A products are derived from subtype A1 organisms (e.g. BOTOX â ;Allergan Inc., Irvine, CA, USA; Dysport â ; Ipsen Ltd, Slough, UK and Xeomin â ; Merz Pharmaceuticals GmbH, Frankfurt am Main, Germany) [11]. These toxin products have an upper dosage limit, and their adverse events have become serious issues, speculated as after-effects of diffusion after high-dose administration [12][13][14][15]. Therefore, a preparation with greater therapeutic efficacy at lower dosages, and showing less diffusion in the body than existing drugs, has been sought. We have attempted to produce botulinum neurotoxin derived from subtype A2 (A2NTX, 150 kDa), with an amino acid sequence different from that of botulinum neurotoxin derived from subtype A1 (A1NTX, 150 kDa). Our aim is to create a product more effective and safer than the existing toxin products derived from subtype A1, and we demonstrated that A2NTX was more effective than complex of botulinum neurotoxin, hemagglutinin, and nontoxic-nonhemagglutinin derived from subtype A1 (A1 progenitor toxin, 900 kDa) and A1NTX in previous reports [16]. In this study, to investigate whether A2NTX is safer than existing botulinum toxin type A products, we compared the systemic toxicities of A1 progenitor toxin, A1NTX and A2NTX. We compared A1 progenitor toxin and A2NTX on administering the intramuscular lethal dose 50% (im LD 50 ) to mice and rats. In addition, w...

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“…Because of the small therapeutic index and potential lethal complication of an overdose, a reliable determination of the biological activity of the BoNT preparation is mandatory (Hakami et al, 2010). The gold standard method to quantify the activity of BoNT is the mouse lethality assay (Pearce et al, 1994(Pearce et al, , 1997Singh et al, 2013). This assay determines the dose of the drug that causes asphyxic death in 50% of treated mice.…”

Section: Introductionmentioning

confidence: 99%

Botulinum neurotoxin dose-dependently inhibits release of neurosecretory vesicle-targeted luciferase from neuronal cells

Pathe-Neuschäfer-Rube

Neuschäfer-Rube

Genz

et al. 2015

ALTEX

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SummaryBotulinum toxin is a bacterial toxin that inhibits neurotransmitter release from neurons and thereby causes a flaccid paralysis. It is used as drug to treat a number of serious ailments and, more frequently, for aesthetic medical interventions. Botulinum toxin for pharmacological applications is isolated from bacterial cultures. Due to partial denaturation of the protein, the specific activity of these preparations shows large variations. Because of its extreme potential toxicity, pharmacological preparations must be carefully tested for their activity. For the current gold standard, the mouse lethality assay, several hundred thousand mice are killed per year. Alternative methods have been developed that suffer from one or more of the following deficits: In vitro enzyme assays test only the activity of the catalytic subunit of the toxin. Enzymatic and cell based immunological assays are specific for just one of the different serotypes. The current study takes a completely different approach that overcomes these limitations: Neuronal cell lines were stably transfected with plasmids coding for luciferases of different species, which were N-terminally tagged with leader sequences that redirect the luciferase into neuro-secretory vesicles. From these vesicles, luciferases were released upon depolarization of the cells. The depolarization-dependent release was efficiently inhibited by botulinum toxin in a concentration range (1 to 100 pM) that is used in pharmacological preparations. The new assay might thus be an alternative to the mouse lethality assay and the immunological assays already in use.

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Measurement of Botulinum Toxin Activity: Evaluation of the Lethality Assay

Cited by 114 publications

References 0 publications

Current Evidence on the Unit Equivalence of Different Botulinum Neurotoxin A Formulations and Recommendations for Clinical Practice in Dermatology

Current Evidence on the Unit Equivalence of Different Botulinum Neurotoxin A Formulations and Recommendations for Clinical Practice in Dermatology

Comparison of Systemic Toxicity between Botulinum Toxin Subtypes A1 and A2 in Mice and Rats

Botulinum neurotoxin dose-dependently inhibits release of neurosecretory vesicle-targeted luciferase from neuronal cells

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