Reconstituting botulinum toxin drugs: shaking, stirring or what?

Abstract

Most botulinum toxin (BT) drugs are stored as powders which need to be reconstituted with normal saline before clinical use. As botulinum neurotoxin (BNT), the therapeutically active ingredient, is a large double-stranded protein the process of reconstitution should be performed with special attention to mechanical stress applied. We wanted to test the mechanical stability of BNT during the reconstitution process. For this, 100MU onabotulinumtoxinA (Botox(®), Irvine, CA, USA) was reconstituted with 2.0ml of NaCl/H2O. Gentle reconstitution (GR) was performed with a 5ml syringe, a 0.90×70mm injection needle, one cycle of injection-aspiration-injection and two gentle shakes of the vial. Aggressive reconstitution (AR) was performed with a 5ml syringe, a 0.40×40mm injection needle, ten injection-aspiration-injection cycles and 30s of continuous shaking of the vial. AR increased the time to paralysis in the mouse hemidiaphragm assay (HDA) from 72.0±4.6 to 106.0±16.0min (*p=0.002, two-tailed t test after Kolmogorov-Smirnova test with Lilliefors correction for normal distribution). Construction of a calibration curve revealed that the increase in the time to paralysis was correlated with a loss of potency of from 100 to 58MU (-42%). BT users should use large diameter injection needles for reconstitution, apply two or three injection-aspiration-injection cycles and, maybe, shake the vials a few times to rinse the entire glass wall. Aggressive reconstitution with small diameter needles, prolonged injection-aspiration-injection and violent shaking should be avoided.