The Influence of Matrix-Induced Dormancy on Metastatic Breast Cancer Chemoresistance

Metastasis remains the leading cause of cancer-associated death worldwide. Disseminated tumor cells can undergo dormancy upon infiltration of secondary organs, and chemotherapeutics fail to effectively eliminate dormant populations. Mechanistic understanding of dormancy-associated chemoresistance could lead to development of targeted therapeutic strategies. Toward this goal, we implemented three poly(ethylene glycol) (PEG)-based hydrogel formulations fabricated from proteolytically degradable PEG (PEG-PQ), integrin ligating PEG-RGDS, and the non-degradable cross-linker N-vinylpyrrolidone (NVP) to induce three distinct phenotypes in triple negative MDA-MB-231 breast cancer cells. With constant 5% w/v PEG-PQ, PEG-RGDS and NVP concentrations were tuned to induce (i) a growth state characterized by high proliferation, high metabolic activity, significant temporally increased cell density, and an invasive morphology; (ii) a balanced dormancy state characterized by a temporal balance (~1:1 ratio) in new live and dead cell density and a non-invasive morphology; and (iii) a cellular dormancy state characterized by rounded, solitary quiescent cells with low viability, proliferation, and metabolic activity. The cellular responses to doxorubicin (DOX), paclitaxel (PAC), and 5-fluorouracil (5-FU) in the three phenotypic states were quantified. Under DOX treatment, cells in dormant states demonstrated increased chemoresistance with a 1.4- to 1.8-fold increase in half maximal effective concentration (EC₅₀) and 1.3- to 1.8-fold increase in half maximal inhibitory concentration (IC₅₀) compared to cells in the growth state. PAC and 5-FU treatment led to similar results. To mechanistically investigate the role of dormancy in conferring DOX resistance, cytoplasmic and nuclear accumulation of DOX was measured. The results indicated comparable DOX accumulation between all three phenotypic states; however, the intracellular to intranuclear distribution indicated a ~1.5 fold increase in DOX nuclear accumulation in cells in the growth state compared to the two dormant states. These results further validate the utility of implementing engineered hydrogels as in vitro platforms of breast cancer dormancy for the development of anti-dormancy therapeutic strategies.