Department of Neurology, Hannover Medical School, Hannover, Germany.
Institute of Toxicology, Hannover Medical School, Hannover, Germany.
Toxicon. 2019 Oct;168:158-163. doi: 10.1016/j.toxicon.2019.07.005. Epub 2019 Jul 16.
Botulinum neurotoxin (BoNT) is synthesized as a progenitor toxin complex (PTC) by Clostridium botulinum. This PTC comprises, in addition to the neurotoxin itself, neurotoxin associated proteins (NAPs) which are composed of three hemagglutinins and one non-toxic, non-hemagglutinin protein. After oral ingestion, these NAPs protect the neurotoxin from the low pH and proteases in the gastrointestinal tract and play a role in the entry via the intestinal barrier. Two of the three therapeutically used botulinum neurotoxin serotype A (BoNT/A) products (onabotulinumtoxinA and abobotulinumtoxinA) contain different amounts of NAPs, while incobotulinumtoxinA, lacks these proteins. In addition, human serum albumin (HSA) that is supposed to stabilize BoNT/A is added at different concentrations. Up to now, the function of the NAPs and HSA after parenteral therapeutic application is not completely understood. To investigate the influence of NAPs and HSA on potency of BoNT/A, we used the ex vivo mouse phrenic nerve hemidiaphragm assay. Increasing doses of HSA resulted dose-dependently in a more pronounced effect of BoNT/A. Though, a plateau was reached with concentrations of 0.8 mg/ml HSA and higher, the accessory addition of NAPs in a relevant amount (4 ng/ml) did not further enhance the effect of BoNT/A. In conclusion, in our ex vivo assay an adequate concentration of HSA prevented BoNT/A from loss of effect and supplementary NAPs did not alter this effect. A confirmation of these data in an in vivo assay is still lacking. However, it might be supposed that even in clinically applied BoNT/A products an increase of HSA accompanied by the avoidance of NAPs could potentially reduce the injected dose and, thus, the risk of unwanted side effects, the treatment costs as well as the risk of a secondary therapy failure due to BoNT/A neutralizing antibodies.
肉毒杆菌神经毒素（BoNT）由肉毒梭菌合成，作为前体毒素复合物（PTC）存在。该 PTC 除了神经毒素本身外，还包含神经毒素相关蛋白（NAPs），NAPs 由三种血凝素和一种无毒、非血凝素蛋白组成。口服摄入后，这些 NAPs 可保护神经毒素免受胃肠道内低 pH 值和蛋白酶的影响，并在通过肠道屏障进入机体的过程中发挥作用。目前临床上使用的三种肉毒梭菌神经毒素血清型 A（BoNT/A）产品中的两种（onabotulinumtoxinA 和 abobotulinumtoxinA）含有不同量的 NAPs，而 incobotulinumtoxinA 则不含这些蛋白。此外，还添加了不同浓度的人血白蛋白（HSA），以稳定 BoNT/A。到目前为止，对于这些 NAPs 和 HSA 在经皮治疗应用后的作用还不完全清楚。为了研究 NAPs 和 HSA 对 BoNT/A 效力的影响，我们使用了离体小鼠膈神经半膈肌试验。结果发现，随着 HSA 剂量的增加，BoNT/A 的作用呈剂量依赖性增强。尽管当 HSA 浓度达到 0.8mg/ml 及更高时达到了一个平台期，但以相关浓度（4ng/ml）添加辅助 NAPs 并没有进一步增强 BoNT/A 的作用。总之，在我们的离体试验中，适当浓度的 HSA 可防止 BoNT/A 失去作用，而补充 NAPs 不会改变这种作用。这些数据在体内试验中的验证仍然缺乏。然而，可以推测，即使在临床上应用的 BoNT/A 产品中，增加 HSA 并避免 NAPs 的使用，有可能降低注射剂量，从而降低因 BoNT/A 中和抗体而产生的不良反应、治疗成本以及二次治疗失败的风险。
