Abstract 888: A novel assay for the evaluation of ADCC in 3D-culture

Abstract

Abstract In the last few years, one of the most innovative areas of cancer research has been immuno-oncology where significant advancements have been seen, particularly, in the use of monoclonal antibodies to treat cancers. The regulation of immune-mediated killing by monoclonal antibodies is an important mechanism of action, like in the case of trastuzumab, a humanized monoclonal antibody used in the treatment of HER-2 positive breast cancers. Binding of the humanized antibody to the receptor induces antibody-dependent cellular cytotoxicity (ADCC). Assays that monitor this activity are historically difficult to execute suffering from low sensitivity and reproducibility. Furthermore, experiments are typically done in suspension of cells grown as monolayers; often, showing limited clinical translatability when studying solid tumor models. Unencumbered by the limitations of 2D assays, the tumor microenvironment of 3D-cultures includes cell-cell interactions and formations of metabolic gradients, which are vital to understanding drug efficacy and resistance. To study the ADCC triggered by trastuzumab in 3D-cellular models, we developed a novel method to evaluate cellular toxicity in spheroids. In this study, we demonstrated the quantification, with specificity and precision, of the ADCC activity elicited by trastuzumab in 3D-culture. The high HER2-expressing BT474 cell line (target cells) was pre-labeled and formed into spheroids utilizing Kuraray's 384-well Elplasia's microplates. PBMCs (effector cells) isolated from healthy donors were added in the presence or absence of the antibody, trastuzumab, and incubated overnight. Cytotoxicity of the tumor-derived cells was evaluated by counting stained dead cells using a cell imager. In parallel, a 2D-platform for ADCC assay was run and the pharmacology of trastuzumab was characterized. BT474 cells grown as spheroids show an IC50 of 0.09 µg/mL, when the study was done in 2D-culture an IC50 of 0.028 µg/mL was obtained. The ability to evaluate ADCC in both 2D- and 3D- may help the development of more translatable models in the clinic. Citation Format: Macarena Irigoyen, Andrea Alms, Yoko Ejiri, Satoru Ayano, Gonzalo Castillo. A novel assay for the evaluation of ADCC in 3D-culture [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 888.