The Influence of Ligand Density and Degradability on Hydrogel Induced Breast Cancer Dormancy and Reactivation

Abstract

The role of hydrogel properties in regulating the phenotype of triple negative metastatic breast cancer is investigated using four cell lines: the MDA-MB-231 parental line and three organotropic sublines BoM-1833 (bone-tropic), LM2-4175 (lung-tropic), and BrM2a-831 (brain-tropic). Each line is encapsulated and cultured for 15 days in three poly(ethylene glycol) (PEG)-based hydrogel formulations composed of proteolytically degradable PEG, integrin-ligating RGDS, and the non-degradable crosslinker N-vinyl pyrrolidone. Dormancy-associated metrics including viable cell density, proliferation, metabolism, apoptosis, chemoresistance, phosphorylated-ERK and -p38, and morphological characteristics are quantified. A multimetric classification approach is implemented to categorize each hydrogel-induced phenotype as: 1) growth, 2) balanced tumor dormancy, 3) balanced cellular dormancy, or 4) restricted survival, cellular dormancy. Hydrogels with high adhesivity and degradability promote growth. Hydrogels with no adhesivity, but high degradability, induce restricted survival, cellular dormancy in the parental line and balanced cellular dormancy in the organotropic lines. Hydrogels with reduced adhesivity and degradability induce balanced cellular dormancy in the parental and lung-tropic lines and balanced tumor mass dormancy in bone-and brain-tropic lines. The ability to induce escape from dormancy via dynamic incorporation of RGDS is also presented. These results demonstrate that ECM properties and organ-tropism synergistically regulate cancer cell phenotype and dormancy.

A Biomimetic Hyaluronic Acid Hydrogel Models Mass Dormancy in Brain Metastatic Breast Cancer Spheroids

“…This protocol has been widely used to detect proliferating cells. [ 16,22,24 ] Similar trend was noticed as observed in Ki67 staining data. The percentage of EdU‐positive cells were significantly higher in spheroids in suspension culture compared to the HA hydrogel (Figure S3, Supporting Information).…”

“…However, biomimetic biomaterial scaffolds provide an important tool to study tumor cell–matrix interactions in vitro and thus have been employed as an experimental model to investigate the role of ECM derived cues on cellular dormancy in both the primary [ 18–23 ] and metastatic tumor setting. [ 15,16,24,25 ] In the context of BMBC, very limited experimental models have been reported to investigate the role of ECM derived cues on dormancy. [ 15,16,24 ] Moreover, to the best of our knowledge, none of the studies till date have reported the impact of an ECM mimetic scaffold on the regulation of mass dormancy in both TNBC and HER2+ BMBC spheroids.…”

“…[ 15,16,24,25 ] In the context of BMBC, very limited experimental models have been reported to investigate the role of ECM derived cues on dormancy. [ 15,16,24 ] Moreover, to the best of our knowledge, none of the studies till date have reported the impact of an ECM mimetic scaffold on the regulation of mass dormancy in both TNBC and HER2+ BMBC spheroids.…”

Low dose chemotherapy induces a dormant state in brain metastatic breast cancer spheroids

“…Previous studies have reported that in multiple cancers, tumor cells tend to achieve chemotherapeutic resistance by upregulating p‐p38 expression 41–44 . In the context of dormancy, p‐ERK/p‐p38 activity ratio has been studied extensively, where lower activity ratio (<1) is indicative of the dormant state and high activity ratio is indicative of the proliferative state 27,45,46 . To identify if p‐ERK and p‐p38 are involved in attaining the dormant state in 80 nM drug treated spheroids, we performed p‐ERK and p‐p38 immunofluorescence staining.…”

“…Immunofluorescence staining was performed to detect Ki67, p‐ERK, p‐p38, and Cleaved caspase 3, which were differentially expressed between proliferating versus dormant spheroids as described in prior studies 23,26,27 . Briefly, on day 7, spheroids were dissociated into single cells and added to a 96 well plate.…”