抗体-药物偶联物（ADCs）相对于纯抗体（mAbs）可以提高治疗指数。然而，ADC的合成很复杂，因为其组成部分是在CHO细胞（mAb）和化学合成（药物）中分别制备的。它们分别经过纯化、偶联，然后纯化ADC，与常规mAbs相比增加了生产成本。相比之下，合成的亲和素融合蛋白更容易生产，它由抗体衍生物、连接物和蛋白毒素组成，即重组免疫毒素（RIT）。植物能够进行功能性抗体所需的翻译后修饰，并且还能表达活性的蛋白毒素，例如重组槲寄生凝集素粘凝素，这在原核生物和哺乳动物细胞中均不可能。本文中，我们使用了Nicotiana benthamiana和N. tabacum植物以及基于烟草BY-2细胞的植物细胞包装（PCPs）来生产用于治疗骨髓单核细胞白血病所需的有效RIT。我们比较了具有不同亚细胞定位信号、连接物和蛋白毒素的RIT。通过设计实验方法，选取的候选物的积累量提高到每千克湿质量约40mg，使用优化的亲和层析方法分离的相应蛋白质纯度约为80%，总收率约为84%。我们对一种基于抗CD64靶向的毒素粘凝素的药物候选物进行了贮存稳定性和体外细胞毒性测试，在人骨髓单核细胞白血病细胞系中进行了表征。我们确定了植物基因表达平台中需要进一步改进的瓶颈，并评估了在植物制造RIT的过程开发中应考虑的关键工艺参数。

Antibody-drug conjugates (ADCs) can improve therapeutic indices compared to plain monoclonal antibodies (mAbs). However, ADC synthesis is complex because the components are produced separately in CHO cells (mAb) and often by chemical synthesis (drug). They are individually purified, coupled, and then the ADC is purified, increasing production costs compared to regular mAbs. In contrast, it is easier to produce recombinant fusion proteins consisting of an antibody derivative, linker and proteinaceous toxin, i.e. a recombinant immunotoxin (RIT). Plants are capable of the post-translational modifications needed for functional antibodies and can also express active protein toxins such as the recombinant mistletoe lectin viscumin, which is not possible in prokaryotes and mammalian cells respectively. Here, we used Nicotiana benthamiana and N. tabacum plants as well as tobacco BY-2 cell-based plant cell packs (PCPs) to produce effective RITs targeting CD64 as required for the treatment of myelomonocytic leukemia. We compared RITs with different subcellular targeting signals, linkers, and proteinaceous toxins. The accumulation of selected candidates was improved to ~ 40 mg kg-1 wet biomass using a design of experiments approach, and corresponding proteins were isolated with a purity of ~ 80% using an optimized affinity chromatography method with an overall yield of ~ 84%. One anti-CD64 targeted viscumin-based drug candidate was characterized in terms of storage stability and cytotoxicity test in vitro using human myelomonocytic leukemia cell lines. We identified bottlenecks in the plant-based expression platform that require further improvement and assessed critical process parameters that should be considered during process development for plant-made RITs.