Quantification of Breast Cancer Cell Invasiveness Using a Three-dimensional (3D) Model

Cvetković, Donna; Goertzen, Cameron; Bhattacharya, Moshmi

doi:10.3791/51341

Cited by 13 publications

(10 citation statements)

References 26 publications

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“…To analyze the effect of ECC on TNBC cells in three dimensions (3D), 35 we cultured MDA-MB-231, MDA-MB-468 and UCI-082014 TNBC cells expressing VC or ECC in 1:1 collagen:Matrigel gels for 6 days. We found that TNBC cells expressing ECC had less and smaller stellate structures and invaded less into the matrix (Figure 6a), had decreased proliferation as analyzed by a mitotic marker, phospho-histone H3 staining (Figure 6b) and increased apoptosis as analyzed by cleaved caspase 3 staining (Figure 6c) when compared with TNBC cells expressing VC.…”

Section: Resultsmentioning

confidence: 99%

CDCP1 cleavage is necessary for homodimerization-induced migration of triple-negative breast cancer

et al. 2016

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Triple-negative breast cancer (TNBC) is a highly aggressive and metastatic form of breast cancer that lacks the estrogen, progesterone and HER2 receptors and is resistant to targeted and hormone therapies. TNBCs express high levels of the transmembrane glycoprotein, complement C1r/C1s, Uegf, Bmp1 (CUB)-domain containing protein 1 (CDCP1), which has been correlated with the aggressiveness and poor prognosis of multiple carcinomas. Full-length CDCP1 (flCDCP1) can be proteolytically cleaved, resulting in a cleaved membrane-bound isoform (cCDCP1). CDCP1 is phosphorylated by Src family kinases in its full-length and cleaved states, which is important for its pro-metastatic signaling. We observed that cCDCP1, compared with flCDCP1, induced a dramatic increase in phosphorylation of the migration-associated proteins: PKCδ, ERK1/2 and p38 mitogen-activated protein kinase in HEK 293T. In addition, only cCDCP1 induced migration of HEK 293T cells and rescued migration of the TNBC cell lines expressing short hairpin RNA against CDCP1. Importantly, we found that only cCDCP1 is capable of dimerization, which can be blocked by expression of the extracellular portion of cCDCP1 (ECC), indicating that dimerization occurs through CDCP1’s ectodomain. We found that ECC inhibited phosphorylation of PKCδ and migration of TNBC cells in two-dimensional culture. Furthermore, ECC decreased cell invasiveness, inhibited proliferation and stimulated apoptosis of TNBC cells in three-dimensional culture, indicating that the cCDCP1 dimer is an important contributor to TNBC aggressiveness. These studies have important implications for the development of a therapeutic to block CDCP1 activity and TNBC metastasis.

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Section: Resultsmentioning

confidence: 99%

CDCP1 cleavage is necessary for homodimerization-induced migration of triple-negative breast cancer

et al. 2016

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“…HSC3 cells were derived from poorly differentiated OSCC and provided by Riken BioResource Center, Cell Engineering Division (RCB1975) (Ibaraki, Japan). HSC3‐Venus human SCC cells (HSC3‐Venus cells) stably overexpressing green fluorescent Venus protein were prepared by the same method as SAS‐Venus cells . The GFP expression vector, pZGreen1‐N1, was obtained from Clontech (CA, USA).…”

Section: Methodsmentioning

confidence: 99%

“…HSC3-Venus human SCC cells (HSC3-Venus cells) stably overexpressing green fluorescent Venus protein were prepared by the same method as SAS-Venus cells. 3 The GFP expression vector, pZGreen1-N1, was obtained from Clontech (CA, USA). For gene transfer, plasmid DNA and the gene transfer reagent FuGENE 6 (Promega, WI, USA) were used in a ratio of 1:3, mixed with MEM without addition of FBS.…”

Section: Cell Lines and Culturementioning

confidence: 99%

“…Recently, several studies of tumor cell invasion in 3‐D models have been reported. For example, breast cancer cell invasion was quantified using a 3‐D model . A cancer cell spheroid assay was used to assess invasion in a 3‐D setting .…”

Section: Introductionmentioning

confidence: 99%

“…For example, breast cancer cell invasion was quantified using a 3-D model. 3 A cancer cell spheroid assay was used to assess invasion in a 3-D setting. 4 Cell spheroids have been employed as 3-D culture models, especially 3-D cancer spheroids for modeling of cancer cell metastasis.…”

Section: Introductionmentioning

confidence: 99%

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Three‐dimensional cultured tissue constructs that imitate human living tissue organization for analysis of tumor cell invasion

Iwai

Kishimoto

Amano

et al. 2018

J Biomedical Materials Res

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Preventing cancer metastasis requires a thorough understanding of cancer cell invasion. These phenomena occur in human 3-D living tissues. To this end, we developed a human cell-based three-dimensional (3-D) cultured tissue constructs that imitate in vivo human tissue organization. We investigated whether our 3-D cell culture system can be used to analyze the invasion of human oral squamous cell carcinoma (OSCC) cells. The 3-D tissue structure consisted of five layers of normal human dermal fibroblasts along with human dermal lymphatic endothelial cell tubes and was generated by the cell accumulation technique and layer-by-layer assembly using fibronectin and gelatin. OSCC cells with different lymph metastatic capacity were inoculated on the 3-D tissues and their invasion through the 3-D tissue structure was observed. Conventional methods of analyzing cell migration and invasion, that is, 2-D culture-based transwell and Matrigel assays were also used for comparison. The results using the 3-D cultured tissue constructs were comparable to those obtained using conventional assays; moreover, use of the 3-D system enabled visualization of differential invasion capacities of cancer cells. These results indicate that our 3-D cultured tissue constructs can be a useful tool for analysis of cancer cell invasion in a setting that reflects the in vivo tissue organization. © 2018 The Authors. journal Of Biomedical Materials Research Part A Published By Wiley Periodicals, Inc. J. Biomed. Mater. Res. Part A: 00A: 000-000, 2018.

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Inhibition of MT1-MMP proteolytic function and ERK1/2 signalling influences cell migration and invasion through changes in MMP-2 and MMP-9 levels

Cepeda

Evered

Pelling

et al. 2017

J. Cell Commun. Signal.

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Membrane type-1 matrix metalloproteinase (MT1-MMP, MMP-14) is a unique protease that cleaves extracellular proteins, activates proMMPs, and initiates intracellular signalling. MCF-7 cells are non-invasive and deficient in MT1-MMP, MMP-2, and MMP-9 expression. We created an MCF-7 cell line (C2) that stably produces active MT1-MMP and demonstrated increased ERK1/2 phosphorylation. MAPK inhibition in this cell line showed an inverse relationship in MMP-2 and MMP-9 transcripts where levels of these genes increased and decreased, respectively. Using invasive MDA-MB 231 cells that endogenously produce MT1-MMP and have naturally high pERK levels, we demonstrated the identical inverse relationship between MMP-2 and -9 transcript and protein levels, suggesting that this novel relationship is conserved amongst MT1-MMP positive breast cancer cells. To further analyze the relationship between MMP-2 and -9 levels, we chemically inhibited activation and catalytic activity of MT1-MMP using a furin and MMP inhibitor, respectively, to show that interference with the functions of MT1-MMP induced changes in MMP-2 and 9 transcript levels that were always inverse of each other, and likely mediated by differential transcriptional activity of the NF-κB transcription factor. Furthermore, we analyzed the functional consequences of these expression changes to show MMP, and in particular ERK, inhibition decreased migration and invasion using 2D culture, and inhibits the formation of an invasive phenotype in Matrigel 3D culture. This study demonstrated a novel inverse transcriptional relationship between MMP-2 and -9 levels and MT1-MMP activity that have functional consequences, and also showed that increases in the levels of MMPs does not necessarily correlate with an invasive phenotype.

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Quantification of Breast Cancer Cell Invasiveness Using a Three-dimensional (3D) Model

Cited by 13 publications

References 26 publications

CDCP1 cleavage is necessary for homodimerization-induced migration of triple-negative breast cancer

CDCP1 cleavage is necessary for homodimerization-induced migration of triple-negative breast cancer

Three‐dimensional cultured tissue constructs that imitate human living tissue organization for analysis of tumor cell invasion

Inhibition of MT1-MMP proteolytic function and ERK1/2 signalling influences cell migration and invasion through changes in MMP-2 and MMP-9 levels

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