Engineered Breast Cancer Cell Spheroids Reproduce Biologic Properties of Solid Tumors

Abstract

Solid tumors develop as three-dimensional tissue constructs. As tumors grow larger, spatial gradients of nutrients and oxygen and inadequate diffusive supply to cells distant from vasculature develops. Hypoxia initiates signaling and transcriptional alterations to promote survival of cancer cells and generation of cancer stem cells (CSCs) that have self-renewal and tumor-initiation capabilities. Both hypoxia and CSCs are associated with resistance to therapies and tumor relapse. Here, we conduct a systematic study to demonstrate that 3D cancer cell models, known as tumor spheroids, generated with a polymeric aqueous two-phase system (ATPS) technology capture these important biological processes. Similar to solid tumors, spheroids of triple negative breast cancer (TNBC) cells deposit major extracellular matrix proteins. Our molecular analysis establishes presence of hypoxic cells in the core region and expression of CSC gene and protein markers including CD24, CD133, and Nanog. Importantly, these spheroids resist treatment with chemotherapy drugs. We successfully target drug resistant spheroids through a combination treatment approach using a hypoxia-activated prodrug, TH-302, and a chemotherapy drug, doxorubicin. This study demonstrates that ATPS spheroids recapitulate important biological and functional properties of solid tumors and provide a unique model for studies in cancer research.

Cell‐Inspired All‐Aqueous Microfluidics: From Intracellular Liquid–Liquid Phase Separation toward Advanced Biomaterials

“…The cultivated spheroids show spherical morphology in high reproducibility (Figure 26b) and excellent viability (Figure 26c) to proliferate into larger spheroids (Figure 26d). The patterned tumor spheroids partially addressed challenges in the heterogeneity of spheroid culture in vitro, which is therefore used as promising platforms for executing high‐throughput anticancer compounds screening. The ability to scale‐up 3D spheroid culture in microfluidic devices offers new opportunity for fast testing of multiple anticancer drugs and analyze their half‐maximum (IC 50 ) and maximum ( E max ) inhibitory concentrations, which facilitates the multiparametric analysis of cellular responses to drug compounds and identify novel chemical probes with chemotherapeutic benefits, as demonstrated in Figure 26e …”

Tumour-on-a-chip: microfluidic models of tumour morphology, growth and microenvironment

“…The advantage of the liquid overlay method is its simplicity of operation, but the disadvantage lies in its poor control over spheroid size. Similarly, to produce tumour spheroids by avoiding cell adhesion to cultureware and inducing aggregation, an aqueous two-phase system can also compartmentalize cell suspension and produce spheroids without the concern of drying and possible inefficiency in chemical transport and toxicity of an oil phase [51][52][53]. Three-dimensional spheroids can also be formed by assembly of cells using bio-orthogonal chemistry [54] or incubation of cells with magnetic particles [55,56] (figure 2g,h).…”

Biomaterials‐Based Approaches to Tumor Spheroid and Organoid Modeling

“…[6] Spheroids of triple negative breast cancer cells generated with the ATPS technology reproduce major biological properties of solid breast tumors. [124] This included growth of spheroids over time, secretion and deposition of major ECM proteins such as collagen I, fibronectin, and laminin by cancer cells [Figure 8C], gradients of proliferative cells [Figure 8D], size- and density-dependent hypoxia [Figure 8E], expression of cancer stem cell markers [Figure 8F], and hypoxia-mediated resistance to doxorubicin and cisplatin [Figure 8G]. Hypoxia was modelled by varying cellular density.…”