Phenotypic Profiling of Raf Inhibitors and Mitochondrial Toxicity in 3D Tissue Using Biodynamic Imaging

Abstract

The existence of phenotypic differences in the drug responses of 3D tissue relative to 2D cell culture is a concern in high-content drug screening. Biodynamic imaging is an emerging technology that probes 3D tissue using short-coherence dynamic light scattering to measure the intracellular motions inside tissues in their natural microenvironments. The information content of biodynamic imaging is displayed through tissue dynamics spectroscopy (TDS) but has not previously been correlated against morphological image analysis of 2D cell culture. In this article, a set of mitochondria-affecting compounds (FCCP, valinomycin, nicardipine, ionomycin) and Raf kinase inhibitors (PLX4032, PLX4720, GDC, and sorafenib) are applied to multicellular tumor spheroids from two colon adenocarcinoma cell lines (HT-29 and DLD-1). These were screened by TDS and then compared against conventional image-based high-content analysis (HCA). The responses to the Raf inhibitors PLX4032 and PLX4720 are grouped separately by cell line, reflecting the Braf/Kras difference in these cell lines. There is a correlation between TDS and HCA phenotypic clustering for most cases, which demonstrates the ability of dynamic measurements to capture phenotypic responses to drugs. However, there are significant 2D versus 3D phenotypic differences exhibited by several of the drugs/cell lines.

Inverse-power-law behavior of cellular motility reveals stromal–epithelial cell interactions in 3D co-culture by OCT fluctuation spectroscopy

“…While speckle fluctuation models have been well-developed for particle flow and diffusion in OCT [6,7], we are only beginning to understand the fluctuation spectra arising from intracellular motions in living tissues. Tissue fluctuation measurements are a growing area of research for assessing drug-response in cancer models in vitro [8,9], oocyte viability [10], and cortical cell recovery after stroke in vivo [3]. …”

Challenges and Opportunities Toward Enabling Phenotypic Screening of Complex and 3D Cell Models

“…One of such emerging technologies is biodynamic imaging, which takes advantage of short-coherence dynamic light scattering to measure the intracellular motions inside tissues in their natural microenvironments. This platform has already been demonstrated to be able to capture phenotypic responses to drugs in spheroids [52]. This type of technology may have the potential to enable the direct, dynamic measurement of cell viability and cell proliferation within 3D tissue, in the absence of any disturbance by a chemical or biological reporter.…”

Role of cellular adhesions in tissue dynamics spectroscopy

“…Figure 2 shows example drug response spectrograms to nocodazole, a chemotherapeutic drug, for the two cell lines. BDI techniques have been used to image the structure of mouse eye and multicellular tumor spheroids 7 , to detect mitosis in multicellular tumor spheroids 14 , and to study the phenotypic response of 3D tissue to mitochondrial toxins and other chemotherapeutic drugs 15 . This paper presents recent investigations into how the level of cellular adhesions influences tissue drug response behavior using BDI.…”