A 3D adrenocortical carcinoma tumor platform for preclinical modeling of drug response and matrix metalloproteinase activity

Abstract

Adrenocortical carcinoma (ACC) has a poor prognosis, and no new drugs have been identified in decades. The absence of drug development can partly be attributed to a lack of preclinical models. Both animal models and 2D cell cultures of ACC fail to accurately mimic the disease, as animal physiology is inherently different than humans, and 2D cultures fail to represent the crucial 3D architecture. Organoids and other small 3D in vitro models of tissues or tumors can model certain complexities of human in vivo biology; however, this technology has largely yet to be applied to ACC. In this study, we describe the generation of 3D tumor constructs from an established ACC cell line, NCI-H295R. NCI-H295R cells were encapsulated to generate 3D ACC constructs. Tumor constructs were assessed for biomarker expression, viability, proliferation, and cortisol production. In addition, matrix metalloproteinase (MMP) functionality was assessed directly using fluorogenic MMP-sensitive biosensors and through infusion of NCI-H295R cells into a metastasis-on-a-chip microfluidic device platform. ACC tumor constructs showed expression of biomarkers associated with ACC, including SF-1, Melan A, and inhibin α. Treatment of ACC tumor constructs with chemotherapeutics demonstrated decreased drug sensitivity compared to 2D cell culture. Since most tumor cells migrate through tissue using MMPs to break down extracellular matrix, we validated the utility of ACC tumor constructs by integrating fluorogenic MMP-sensitive peptide biosensors within the tumor constructs. Lastly, in our metastasis-on-a-chip device, NCI-H295R cells successfully engrafted in a downstream lung cell line-based construct, but invasion distance into the lung construct was decreased by MMP inhibition. These studies, which would not be possible using 2D cell cultures, demonstrated that NCI-H295R cells secreted active MMPs that are used for invasion in 3D. This work represents the first evidence of a 3D tumor constructs platform for ACC that can be deployed for future mechanistic studies as well as development of new targets for intervention and therapies.

In vitrodifferentiation of mouse pluripotent stem cells into glucocorticoid-producing adrenocortical cells

“…In other systems, 3D cultures have been shown to be more conducive to differentiation by providing molecular cues from neighbouring cells as well as mechanical stimulation 31,32 . We therefore tested the aggregation of aIM/LPM cells in ultra-low attachment plates, or in microwell plates, the latter allowing the formation of hundreds of small spheroids in a single well.…”

Grand challenges in organoid and organ-on-a-chip technologies

“…Simply moving a common cancer cell line from a 2D tissue culture plastic to a 3D ECM hydrogel matrix can do wonders. A cell line thought to be somewhat artificial can take on highly relevant disease-specific characteristics and serve as a useful research tool ( Aleman and Skardal, 2018 ; Dedhia et al, 2023 ; Nairon and Skardal, 2023 ). This is particularly true in rare cancers where patient-derived biospecimen availability for research might be limited.…”