Chemoprotection Across the Tumor Border: Cancer Cell Response to Doxorubicin Depends on Stromal Fibroblast Ratios and Interstitial Therapeutic Transport
- 372 Downloads
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 tumor-stroma interface.
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 µm/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.
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 breast-mimetic in vitro 3D collagen I-based homogenous hydrogel system. Our in vitro and in silico tumor border models indicate that MDA-MB-231s at the top of the gel, indicative of the tumor bulk, receive the greatest concentration of drug for the longest time, yet cellular death is lowest in this region. This trend is reversed for MDA-MB-231s alone.
Together, our data indicate that fibroblasts are chemoprotective at lower density, resulting in less tumor death in regions of higher chemotherapy concentration. Additionally, chemotherapeutic agent transport properties can modulate this effect.
KeywordsTumor microenvironment Drug delivery Doxorubicin Fibroblasts Breast cancer Interstitial flow 3D cell culture
Human breast triple negative adenocarcinoma cell line (luminal)
Human breast triple negative invasive ductal carcinoma cell line (luminal)
Human breast ER+/PR+ adenocarcinoma cell line (basal)
Human dermal fibroblast
Tumor to stroma transition model
Interstitial fluid pressure
The researchers would like to acknowledge Lynette Sequeira for her technical laboratory assistance, Charles Calderwood for statistical assistance. We also thank RC Cornelison, RP Pompano, SM Peirce-Cottler, and SS Blemker for helpful discussion. We would also like to acknowledge the Janes Lab at UVa for initial contribution of cell lines, the Advanced Microscopy Facility and the Biorepository and Tissue Research Facility. This research was funded in part through funding to JM Munson from the UVa Cancer Center through the NCI Cancer Center Support Grant P30 CA44579 and support from the Snell Endowment Fund and Commonwealth of VA, the School of Medicine, and funding to GF Beeghly from the Harrison Undergraduate Research Awards Center at UVa.
Conflict of interest
The authors declare that they have no conflicts of interest.
All human samples were acquired according to the ethical standards. No animal studies were performed in this work.
- 5.Conze, D., et al. Autocrine production of interleukin 6 causes multidrug resistance in breast cancer cells. Cancer Res. 61(24):8851–8858, 2001.Google Scholar
- 15.Gattazzo, F., A. Urciuolo, and P. Bonaldo. Extracellular matrix: a dynamic microenvironment for stem cell niche. Biochim. Biophys. Acta Gen. Subj. 2506–2519:2014, 1840.Google Scholar
- 24.Jain, R. K. Transport of molecules in the tumor interstitium: a review. Cancer Res. 47:3039–3051, 1987.Google Scholar
- 26.Jain, R. K., and L. T. Baxter. Mechanisms of heterogeneous distribution of monoclonal antibodies and other macromolecules in tumors: significance of elevated interstitial pressure. Cancer Res. 48:7022–7032, 1988.Google Scholar
- 32.Lankelma, J., et al. Doxorubicin gradients in human breast cancer. Clin. Cancer Res. 5:1703–1707, 1999.Google Scholar
- 35.Ljungkvist, A. S. E., J. Bussink, P. F. J. W. Rijken, J. H. A. M. Kaanders, A. J. Van der Kogel, and J. Denekamp. Vascular architecture, hypoxia, and proliferation in first-generation xenografts of human head-and-neck squamous cell carcinomas. Int. J. Radiat. Oncol. Biol. Phys. 54:215–228, 2002.CrossRefGoogle Scholar
- 47.Netti, P. A., D. A. Berk, M. A. Swartz, A. J. Grodzinsky, and R. K. Jain. Role of extracellular matrix assembly in interstitial transport in solid tumors. Cancer Res. 60:2497–2503, 2000.Google Scholar
- 56.Shain, K. H., and W. S. Dalton. Cell adhesion is a key determinant in de novo multidrug resistance (MDR): new targets for the prevention of acquired MDR. Mol. Cancer Ther. 1:69–78, 2001.Google Scholar
- 64.Subik, K., et al. The expression patterns of ER, PR, HER2, CK5/6, EGFR, Ki-67 and AR by immunohistochemical analysis in breast cancer cell lines. Breast Cancer (Auckl.) 4:35–41, 2010.Google Scholar
- 70.Weinstein, R. S., et al. Relationship of the expression of the multidrug resistance gene product (P-glycoprotein) in human colon carcinoma to local tumor aggressiveness and lymph node metastasis. Cancer Res. 51:2720–2726, 1991.Google Scholar