Chemoprotection Across the Tumor Border: Cancer Cell Response to Doxorubicin Depends on Stromal Fibroblast Ratios and Interstitial Therapeutic Transport

Abstract

Introduction-Increasing evidence suggests that the tumor microenvironment reduces therapeutic delivery and may lead to chemotherapeutic resistance. At tumor borders, drug is convectively transported across a unique microenvironment composed of inverse gradients of stromal and tumor cells. These regions are particularly important to overall survival, as they are often missed through surgical intervention and contain many invading cells, often responsible for metastatic spread. An understanding of how cells in this tumor-border region respond to chemotherapy could begin to elucidate the role of transport and intercellular interactions in relation to chemoresistance. Here we examine the contribution of drug transport and stromal fibroblasts to breast cancer response to doxorubicin using in silico and in vitro models of the tumorstroma interface. Methods-2D culture systems were utilized to determine the effects of modulated ratios of fibroblasts and cancer cells on overall cancer cell viability. A homogenous breast mimetic in vitro 3D collagen I-based hydrogel system, with drug delivered via pressure driven flow (0.5 lm/s), was developed to determine the effects of transport and fibroblasts on doxorubicin treatment efficacy. Using a novel layered tumor bulk-to-stroma transition in vitro 3D hydrogel model, ratios of MDA-MB-231s and fibroblasts were seeded in successive layers creating cellular gradients, yielding insight into region specific cancer cell viability at the tumor border. In silico models, utilizing concentration profiles developed in COMSOL Multiphysics, were optimized for time dependent viability prediction and confirmation of in vitro findings. Results-In general, the addition of fibroblasts increased viability of cancer cells exposed to doxorubicin, indicating a protective effect of co-culture. More specifically, however, modulating ratios of cancer cells (MDA-MB-231):fibroblasts in 2D co-cultures, to mimic the tumor-stroma transition, resulted in a linear decrease in cancer cell viability from 77% (4:1) to 44% (1:4). Similar trends were seen in the

Applicability of drug response metrics for cancer studies using biomaterials

“…The in vivo tumor microenvironment is inherently spatially heterogeneous, and researchers are starting to examine how this cellular and extracellular heterogeneity may influence treatment response. Logsdon et al, found that MDA-MB-231 cells in mixed, 3D culture with fibroblasts were more resistant to 10 µM doxorubicin at low ratios of tumor to stromal cells (4:1) but equally affected by the drug at higher ratios (1:4) [39]. Shen at al., found similar results using a micro-patterned interface of tumor to stromal cells wherein MCF-7 cell proliferation was inhibited by reversine at the interface but not in the bulk [46].…”

“…The presence of stromal cells (cancer-associated fibroblasts, pericytes, or adipocytes) has been shown to drastically alter drug response, ranging from promoting drug resistance to increasing drug sensitivity [39][40][41][42][43][44]. Using multicellular cultures can account for tissue level interactions and therefore may be more physiologically relevant than monocultures.…”

“…However, because dead cells may remain within 3D cultures, total fluorescence readings may be less accurate. It is often more appropriate to stain the cells using a viability marker like propidium iodide or JC-1, then quantify cell viability and/or number using either confocal/multiphoton microscopy or flow cytometry [39,40,50]. This method can be used to track survival of one cell type of interest while ignoring the other cells or examine survival of each of multiple cell types via multiplexing of different fluorophores.…”

Three‐dimensional culture models to study drug resistance in breast cancer

“…This observation in synergistic treatment was also made in a similar model with MDA‐MB‐231 and stromal HTB‐125 cells, where without Tranilast, DOX therapy lead to increased fibrosis and thus drug resistance (Saini et al, 2018). In a similar investigation of CAF influence, the ratio of MDA‐MB‐231:human dermal fibroblast was tested for possible effects on DOX efficacy while incorporating delivery via interstitial flow (Logsdon, Beeghly, & Munson, 2017). The fibroblasts were more chemoprotective at low density (4:1) than at 1:1 or 1:4 (Logsdon et al, 2017).…”

“…In a similar investigation of CAF influence, the ratio of MDA‐MB‐231:human dermal fibroblast was tested for possible effects on DOX efficacy while incorporating delivery via interstitial flow (Logsdon, Beeghly, & Munson, 2017). The fibroblasts were more chemoprotective at low density (4:1) than at 1:1 or 1:4 (Logsdon et al, 2017).…”