Könitzer Jennifer D, Sieron Annette, Wacker Angelika, Enenkel Barbara
Boehringer Ingelheim, Division Research Germany, Immune Modulation and Biotherapeutics Discovery, Biberach/Riß, Germany.
Boehringer Ingelheim, Biopharma Operations Germany, Biberach/Riß, Germany.
PLoS One. 2015 Dec 29;10(12):e0145633. doi: 10.1371/journal.pone.0145633. eCollection 2015.
The direct induction of cell death, or apoptosis, in target cells is one of the effector mechanisms for the anti CD20 antibody Rituximab. Here we provide evidence that Rituximab's apoptotic ability is linked to the antibody IgG isotype. Reformatting Rituximab from the standard human IgG1 heavy chain into IgG2 or IgG4 boosted in vitro apoptosis induction in the Burkitt's lymphoma B cell line Ramos five and four-fold respectively. The determinants for this behavior are located in the hinge region and CH1 domain of the heavy chain. By transplanting individual IgG2 or IgG4 specific amino acid residues onto otherwise IgG1 like backbones, thereby creating hybrid antibodies, the same enhancement of apoptosis induction could be achieved. The cysteines at position 131 of the CH1 domain and 219 in the hinge region, involved in IgG2 and IgG4 disulfide formation, were found to be of particular structural importance. Our data indicates that the hybrid antibodies possess a different CD20 binding mode than standard Rituximab, which appears to be key in enhancing apoptotic ability. The presented work opens up an interesting engineering route for enhancing the direct cytotoxic ability of therapeutic antibodies.
诱导靶细胞发生直接细胞死亡或凋亡是抗CD20抗体利妥昔单抗的效应机制之一。在此，我们提供证据表明利妥昔单抗的凋亡能力与抗体IgG亚型有关。将利妥昔单抗从标准的人IgG1重链改造成IgG2或IgG4后，分别使伯基特淋巴瘤B细胞系拉莫斯细胞的体外凋亡诱导能力提高了5倍和4倍。这种行为的决定因素位于重链的铰链区和CH1结构域。通过将单个IgG2或IgG4特异性氨基酸残基移植到类似IgG1的骨架上，从而产生杂交抗体，也能实现相同程度的凋亡诱导增强。发现参与IgG2和IgG4二硫键形成的CH1结构域第131位的半胱氨酸和铰链区第219位的半胱氨酸具有特别重要的结构意义。我们的数据表明，杂交抗体与标准利妥昔单抗具有不同的CD20结合模式，这似乎是增强凋亡能力的关键。所展示的工作为增强治疗性抗体的直接细胞毒性能力开辟了一条有趣的工程途径。
