Screening of anti-cancer drug candidates follows a step-by-step testing procedure on different models, involving in vitro cell cultures, in vivo animal models and humans. Traditionally, in vitro 2D cell cultures are frequently used to evaluate cytotoxic effects of drug candidates on tumors. Although simple and efficient, 2D tumor cultures do not accurately reflect drug responses in a 3D environment, which hampers the use of these models for in vivo studies. Multicellular tumor spheroids (MCTS), a 3D model, have attracted much attention in cancer research due to their physiological similarities with in vivo tumors, such as their structures, oxygen and nutrient gradients, and drug resistances. Therefore, they can serve as a more predictive model for the cancer drug discovery.

High-throughput screening (HTS) systems employing various imaging approaches, e.g. bright-field microscopy, fluorescence microscopy or confocal microscopy, have been employed to perform routine imaging and analyses of structures and functions of MCTSs. One drawback of the current HTS systems is that they do not readily provide a 3D view of the entire MCTS, especially for large spheroids (>500μm), due to limited imaging penetration, which hampers accurate characterization of the spheroid volumetric changes. In this work, we developed a new screening system to visualize the entire 3D structure of MCTS in 96-well ultra-low attachment (ULA) round bottom microplates using optical coherence tomography (OCT), a high resolution and 3D imaging modality. We showed that this imaging system was able to perform longitudinal tracking of 3D structural changes, e.g. sizes, shapes and volumes, for individual MCTS. In particular, we demonstrated that spheroid volume measured based on 3D OCT imaging data provided a more accurate and robust result as compared to volume calculated based barely on the spheroid size (diameter). Furthermore, we demonstrated that this system could perform non-invasive, label-free monitoring of pathophysiological status of MCTS over time. The 3D distribution of a spheroid’s

necrotic core could be assessed based on intrinsic optical contrast. This OCT-based screening system would open up new opportunities for accurate 3D characterization of morphological and pathophysiological features of MCTS, and benefit the development of the state-of-the-art 3D HTS system for cancer drug discovery.

Citation Format: Yongyang Huang, Shunqiang Wang, Sarah Kessel, Ian Rubinoff, Leo Li-Ying Chan, Peter Li, Yaling Liu, Jean Qiu, Chao Zhou. Noninvasive characterization of 3D morphology and pathophysiology of multicellular tumor spheroids using optical coherence tomography [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5765. doi:10.1158/1538-7445.AM2017-5765