Impact of a Desmoplastic Tumor Microenvironment for Colon Cancer Drug Sensitivity: A Study with 3D Chimeric Tumor Spheroids

Abstract

spheroid culture provides opportunities to model tumor growth closer to its natural context. The collagen network in the extracellular matrix supports autonomic tumor cell proliferation, but its presence and role in tumor spheroids remain unclear. In this research, we developed an in vitro 3D co-culture model in a microwell 3D (μ-well 3D) cellculture array platform to mimic the tumor microenvironment (TME). The modular setup is used to characterize the paracrine signaling molecules and the role of the intraspheroidal collagen network in cancer drug resistance. The μ-well 3D platform is made up of poly(dimethylsiloxane) that contains 630 round wells for individual spheroid growth. Inside each well, the growth surface measured 500 μm in diameter and was functionalized with the amphiphilic copolymer. HCT-8 colon cancer cells and/or NIH3T3 fibroblasts were seeded in each well and incubated for up to 9 days for TME studies. It was observed that NIH3T3 cells promoted the kinetics of tumor organoid formation. The two types of cells self-organized into core−shell chimeric tumor spheroids (CTSs) with fibroblasts confined to the shell and cancer cells localized to the core. Confocal microscopy analysis indicated that a type-I collagen network developed inside the CTS along with increased TGF-β1 and α-SMA proteins. The results were correlated with a significantly increased stiffness in 3D co-cultured CTS up to 52 kPa as compared to two-dimensional (2D) co-culture. CTS was more resistant to 5-FU (IC 50 = 14.0 ± 3.9 μM) and Regorafenib (IC 50 = 49.8 ± 9.9 μM) compared to cells grown under the 2D condition 5-FU (IC 50 = 12.2 ± 3.7 μM) and Regorafenib (IC 50 = 5.9 ± 1.9 μM). Targeted collagen homeostasis with Sclerotiorin led to damaged collagen structure and disrupted the type-I collagen network within CTS. Such a treatment significantly sensitized collagen-supported CTS to 5-FU (IC 50 = 4.4 ± 1.3 μM) and to Regorafenib (IC 50 = 0.5 ± 0.2 μM). In summary, the efficient formation of colon cancer CTSs in a μ-well 3D culture platform allows exploration of the desmoplastic TME. The novel role of intratumor collagen quality as a drug sensitization target warrants further investigation.

Controlling the Trimerization of the Collagen Triple-Helix by Solvent Switching

“…Finally, we explored the stability of the hybrid triple helix formed by the CHP strand and the denatured collagen chains in different solvents. Although CHP–collagen hybridization has been reported in many studies as an established method to examine collagen damage in tissues, ,,, the direct characterization and structural understanding of the hybrid triple helix itself are extremely limited. Therefore, assessing the effect of different solutions on the stability of the hybrid triple helix can help us start appreciating the stabilizing interactions between the CHP and denatured collagen chains.…”

Glioblastoma-Derived Three-Dimensional Ex Vivo Models to Evaluate Effects and Efficacy of Tumor Treating Fields (TTFields)

“…Moreover, they can be developed to resemble the tumor microenvironment more closely, allowing functional investigations of drug responses as well as metastasis, such as combining the 3D model with biomaterials [ 17 ], or engineering them to mimic physiological environments by using tumor-on-a-chip technology [ 38 ]. Three-dimensional models have been successfully used for drug screening studies for breast, lung and colon cancer [ 20 , 21 , 22 ]. Studies in GBM using 3D systems are also not far behind, as more research favoring them emerge [ 19 , 39 , 40 ].…”

“…Organoids represent the histological characteristics, cellular diversity, gene expression, and mutational profiles of their corresponding parental tumors, as was successfully demonstrated for breast, lung and colon cancer, as well as GBM [ 18 , 20 , 21 , 22 , 40 , 45 ]. In addition, they can be generated quickly and reliably within two weeks from intraoperatively gained tissue [ 24 ].…”

“…Some of the more recent 3D model systems available for GBM include glioma cell lines, GBM stem cell derived cultures, microtubes, human-induced pluripotent stem cells, organoids, organotypic slice cultures, as well as bioprinted chip systems [ 19 ]. Nonetheless, most of the 3D models that exist, even apart from those used for GBM, are mainly engineered for the purpose of drug screening [ 20 , 21 , 22 ]. None currently exist in combination with TTFields to screen for the aforementioned modalities including patient-suitability to predict maximum beneficial outcomes.…”

Synthesis and characterization of a fluorescent polymeric nano-thermometer: dynamic monitoring of 3D temperature distribution in co-cultured tumor spheroids

“…3.6 3D distribution and dynamic monitoring of temperature in co-cultured 3D tumor spheroids HCT-8 and NIH3T3 cells were co-cultured in a µ-well 3D cellculture platform as mentioned in our previous manuscript. 56 These two types of cells are self-organized into core-shell structures, HCT-8 cells localized into the center, and NIH3T3 confined to the shell (see ESI Fig. S5(b) †).…”