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Background: Effective in vivo T-cell depletion is a critical component of many transplantation tolerance protocols. We have previously demonstrated T-cell depletion in miniature swine using a CRM9-based CD3-immunotoxin, pCD3-CRM9. CRM9 is a mutant form of diphtheria toxin (DT) that binds less efficiently than wild-type DT to the DT receptor (proHB-EGF) of primates. In this report, we describe and characterize the dose-dependent neurotoxicity associated with CRM9-based immunotoxin administration in swine., Methods: Miniature swine were treated with varying doses of pCD3-CRM9 followed by daily monitoring for symptoms of neuropathy, including limb weakness, paresis, sluggishness, and/or respiratory distress. Animals demonstrating severe respiratory distress were euthanized and peripheral nerve, spinal cord, and skeletal muscle tissue samples were obtained at autopsy for microscopic examination. Unconjugated CRM9 was administered to one animal to define its toxicity independent of the effects of T-cell depletion., Results: Excellent T-cell depletion was obtained using doses of pCD3-CRM9 greater than 0.1 mg/kg. However, neurotoxicity was observed at these doses, as manifested by transient muscle weakness or paresis, which in some cases progressed to respiratory failure and death. Dorsal root ganglia samples revealed pathological changes typical of diphtheritic polyneuropathy. The animal receiving unconjugated CRM9 exhibited the same neurotoxic side effects as those receiving the pCD3-CRM9 conjugate., Conclusions: Administration of pCD3-CRM9 immunotoxin provides excellent T-cell depletion in miniature swine but is associated with significant dose-dependent neurotoxicity. A possible reason for CRM9-associated neurotoxicity in swine, but not primates, is suggested on the basis of a known amino acid difference in the exodomain of the DT receptor (proHB-EGF) of swine compared with that of primates.