Elsevier

Toxicon

Volume 42, Issue 5, October 2003, Pages 461-469
Toxicon

Intramuscular injection of 125I-botulinum neurotoxin-complex versus 125I-botulinum-free neurotoxin: time course of tissue distribution

https://doi.org/10.1016/S0041-0101(03)00196-XGet rights and content

Abstract

The diffusion from the site of intramuscular injection of 900 kDa botulinum neurotoxin–hemagglutinin complex (BoNT/A-complex) and 150 kDa free-botulinum neurotoxin (free-BoNT/A) was compared. Radioiodinated compounds were injected into the gastrocnemius muscle of rats (70 Units (U) 125I-BoNT/A-complex, 67 or 344 U free-125I-BoNT/A, or free-125I-iodide) and the eyelids of rabbits (24 U 125I-BoNT/A-complex or 108 U free-125I-BoNT/A), and measured in various tissues at different time points. There were no detectable systemic effects or generalized botulinum neurotoxin toxicity in either rats or rabbits, indicating that most of the toxin, whether as 125I-BoNT/A-complex or free-125I-BoNT/A, remained at the injection site. In rats, 125I-BoNT/A-complex and free-125I-BoNT/A diffused in a pattern that was grossly similar. Almost no radioactivity was recovered from the brain. Radioactivity recovered from distant tissues (thyroid, skin, and contralateral muscle) was primarily attributable to either low molecular weight 125I-containing peptides or 125I-iodide. After injection into rabbit eyelids, neither 125I-BoNT/A-complex nor free-125I-BoNT/A spread to distant structures, including the eye. The results indicate that most of the neurotoxin does not diffuse from the injection site, whether in free or complexed form, and this may reduce the potential for systemic effects.

Introduction

Botulinum neurotoxin is one of the most potent neuroparalytic agents known (Coffield et al., 1994). Subtoxic doses have been used successfully in a wide range of clinical applications. Approved for clinical use by the FDA in 1989, localized intramuscular injections of botulinum neurotoxin type A have been used to treat intractable muscle hyperactivity associated with strabismus, blepharospasm, dysphonia, spasticity, cerebral palsy, and different types of dystonia (Coffield et al., 1994, Osako and Keltner, 1991, Brashear et al., 2002, Koman et al., 2000). Several professional societies have endorsed the use of botulinum neurotoxin therapy (American Academies of Ophthalmology and Neurology, and the National Institutes of Health).

Purified botulinum neurotoxin type A is a 150 kDa protein. In cultures, the toxin is found complexed with nontoxin proteins to form 300, 500, or 900 kDa complexes. BOTOX® (botulinum toxin type A; Allergan, Inc.), a botulinum neurotoxin type A complex (BoNT/A-complex), contains the 900 kDa complex made up of the type A toxin molecule and associated proteins (including a hemagglutinin). In clinical use, this BoNT/A-complex is injected directly into those muscles where the clinical response is desired. Using the lowest effective drug dosage minimizes the spread of neurotoxin to surrounding tissues (Shaari and Sanders, 1993). However, it is not known whether the free-neurotoxin (free-BoNT/A in which the associated nontoxin proteins have been removed) would show significantly different pharmacokinetic and distribution properties after intramuscular injection than the BoNT/A-complex. A comparison of the distribution and pharmacokinetics of the BoNT/A-complex with free-BoNT/A is of interest both to determine the time course of diffusion away from the injection site and to identify organs and tissues into which the neurotoxin might deposit.

The purpose of the present study was to compare the diffusion of BoNT/A-complex from the site of intramuscular injection with that of free-BoNT/A.

Section snippets

Animals

All animal experimentation was conducted in accordance with the Animals (Scientific Procedures) Act of 1986. Male Sprague–Dawley CD rats (age ≈10 weeks) were purchased from Charles River (Margate, UK) and were used at body weights ranging from 284 to 393 g. Rats were maintained in glass metabolism cages to permit separate collection of urine and feces. Female New Zealand White rabbits were purchased from Interfauna Ltd (Wyton, UK) and were used at body weights ranging from 2.5 to 3.0 kg.

Test substances

Results

None of the animals developed signs of acute toxicity, and there was no mortality prior to scheduled euthanasia.

Discussion

At all time points after injection with neurotoxin-complex, the proportion of TCA-precipitable material remaining at the injection site was consistently high (>90%), indicating that most of the radioactive material was in the form of large macromolecules. The percentage of material that was TCA-precipitable was greater with neurotoxin-complex than free-neurotoxin. In contrast, less of the radiolabeled material remaining at the injection site of high-dose free-neurotoxin appeared to be in the

Acknowledgements

Financial support was provided by a grant from Allergan, Inc.

References (14)

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