Dilchert Janine, Hofmann Martin, Unverdorben Felix, Kontermann Roland, Bunk Sebastian
Immatics Biotechnologies GmbH, Paul-Ehrlich-Str. 15, 72076 Tuebingen, Germany.
Institute of Cell Biology and Immunology, University of Stuttgart, Allmandring 31, 70569 Stuttgart, Germany.
Antibodies (Basel). 2022 May 10;11(2):34. doi: 10.3390/antib11020034.
Bispecific T cell receptor (TCR)-based molecules capable of redirecting and activating T cells towards tumor cells represent a novel and promising class of biotherapeutics for the treatment of cancer. Usage of TCRs allows for targeting of intracellularly expressed and highly selective cancer antigens, but also requires a complex maturation process to increase the naturally low affinity and stability of TCRs. Even though TCR domains can be matured via phage and yeast display, these techniques share the disadvantages of non-human glycosylation patterns and the need for a later reformatting into the final bispecific format. Here, we describe the development and application of a Chinese Hamster Ovary (CHO) display for affinity engineering of TCRs in the context of the final bispecific TCR format. The recombinase-mediated cassette exchange (RCME)-based system allows for stable, single-copy integration of bispecific TCR molecules with high efficiency into a defined genetic locus of CHO cells. We used the system to isolate affinity-increased variants of bispecific T cell engaging receptor (TCER) molecules from a library encoding different CDR variants of a model TCR targeting preferentially expressed antigen in melanoma (PRAME). When expressed as a soluble protein, the selected TCER molecules exhibited strong reactivity against PRAME-positive tumor cells associated with a pronounced cytokine release from activated T cells. The obtained data support the usage of the CHO display-based maturation system for TCR affinity maturation in the context of the final bispecific TCER format.
能够将T细胞重定向并激活使其靶向肿瘤细胞的双特异性T细胞受体（TCR）分子代表了一类用于癌症治疗的新型且有前景的生物疗法。TCR的使用能够靶向细胞内表达的高选择性癌症抗原，但也需要一个复杂的成熟过程来提高TCR天然较低的亲和力和稳定性。尽管TCR结构域可以通过噬菌体和酵母展示进行成熟，但这些技术存在非人糖基化模式的缺点，并且需要在后期重新构建成最终的双特异性形式。在此，我们描述了一种中国仓鼠卵巢（CHO）展示技术的开发及应用，用于在最终双特异性TCR形式的背景下对TCR进行亲和力工程改造。基于重组酶介导的盒式交换（RCME）系统能够将双特异性TCR分子高效、稳定地单拷贝整合到CHO细胞的一个特定基因位点。我们使用该系统从一个编码优先靶向黑色素瘤中优先表达抗原（PRAME）的模型TCR不同互补决定区（CDR）变体的文库中分离出亲和力增强的双特异性T细胞结合受体（TCER）分子变体。当作为可溶性蛋白表达时，所选的TCER分子对PRAME阳性肿瘤细胞表现出强烈反应性，并伴有活化T细胞显著的细胞因子释放。所获得的数据支持在最终双特异性TCER形式的背景下使用基于CHO展示的成熟系统进行TCR亲和力成熟。
