Determining tamoxifen sensitivity using primary breast cancer tissue in collagen-based three-dimensional culture

Leeper, Alexander; Farrell, Joanne; Williams, Linda; Thomas, Jeremy; Dixon, JM; Wedden, Sarah; Harrison, David J.; Katz, Elad

doi:10.1016/j.biomaterials.2011.10.028

Cited by 20 publications

(23 citation statements)

References 39 publications

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“…We have recently succeeded in establishing a robust system for long-term culture of human breast cancer explants, successfully growing >90% tumours of all major sub-types [4, 5]. In these cultures, primary breast cancer biopsies from invasive breast cancers are explanted into the centre of a type I collagen matrix.…”

Section: Resultsmentioning

confidence: 99%

“…Furthermore, there are serious doubts regarding the relevance of drug testing in established cancer cell lines and transfer of results to the clinic [3]. We have developed a three-dimensional culture system that allows maintenance of complex breast cancer specimens explanted ex vivo for up to four weeks after surgical resection in a supporting matrix of exogenous stromal type I collagen [4, 5]. These ex vivo cultures offer a relatively rapid and quantitative avenue to explore mechanisms of tumour spread and evaluate new treatments.…”

Section: Introductionmentioning

confidence: 99%

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Targeting of Rac GTPases blocks the spread of intact human breast cancer

et al. 2012

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High expression of Rac small GTPases in invasive breast ductal carcinoma is associated with poor prognosis, but its therapeutic value in human cancers is not clear. The aim of the current study was to determine the response of human primary breast cancers to Rac-based drug treatments ex vivo.Three-dimensional organotypic cultures were used to assess candidate therapeutic avenues in invasive breast cancers. Uniquely, in these primary cultures, the tumour is not disaggregated, with both epithelial and mesenchymal components maintained within a three-dimensional matrix of type I collagen. EHT 1864, a small molecule inhibitor of Rac GTPases, prevents spread of breast cancers in this setting, and also reduces proliferation at the invading edge. Rac1+ epithelial cells in breast tumours also contain high levels of the phosphorylated form of the transcription factor STAT3. The small molecule Stattic inhibits activation of STAT3 and induces effects similar to those seen with EHT 1864. Pan-Rac inhibition of proliferation precedes down-regulation of STAT3 activity, defining it as the last step in Rac activation during human breast cancer invasion.Our data highlights the potential use of Rac and STAT3 inhibition in treatment of invasive human breast cancer and the benefit of studying novel cancer treatments using three-dimensional primary tumour tissue explant cultures.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Targeting of Rac GTPases blocks the spread of intact human breast cancer

et al. 2012

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“…Potentially this is a more flexible model as the authors cite a success rate of >95%. Along with conventional immunohistochemistry, optical projection tomography, which enables 3D visualisation of the explants in situ, allowed tamoxifen-sensitivity to be determined 6. With the complexity and heterogeneity of breast cancer now firmly recognised, together these models provide humanised alternatives for the in vitro study of breast cancer and could be applied to other solid cancer types.…”

Section: Discussionmentioning

confidence: 99%

The practicalities of using tissue slices as preclinical organotypic breast cancer models

et al. 2012

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Models considering breast cancer complexity cannot be easily or accurately replicated in routine cell line or animal models. We aimed to evaluate the practicality of organotypic tissue slice culture in breast cancer. Following ethical approval, 250 µm thick sections from surplus breast tumours (n=10) were prepared using a vibrating blade microtome. Triplicate tissue slices were placed in 6-well plates and cultured for up to 7 days ± tamoxifen (1 nM) or doxorubicin (1 µM). Tissue slices were fixed and embedded before sectioning for morphological evaluation and immunohistochemistry. H&E showed good preservation of tissue morphology. Collagen production was evident. Biomarkers of proliferation and apoptosis could be evaluated using immunohistochemistry and used as surrogates to quantify drug effects. In summary, breast cancer tissue slices can be cultured in vitro as organotypic models. Nevertheless, although simple in concept, the delicacy of the model with regard to handling makes subsequent analytical processes challenging.

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“…In contrast, approaches that utilize re-aggregation of primary tissue cultures into functional 3D matrices or scaffolds can lead to the formation of complex, functional organoids or microtissues that naturally include stromal and ECM components [20]. The direct embedding of cell lines, primary cells [8], [9] or primary explants [21], [22] into biological relevant ECM preparations remains the most promising and practical method to recapitulate morphologic aspects such as tissue formation, differentiation and homeostasis; also including tumor progression and invasion (reviewed in [4]). In addition, it is critical to assess the physical force, pressure and local stiffness or rigidity of the matrix, which promotes tumor progression, cell motility and impacts on the modes of cell invasion used by cancer cells [23], [24].…”

Section: Introductionmentioning

confidence: 99%

“…These various spheroid or acinus phenotypes correlate with incremental activation of oncogenic signalling pathways and re-arrangement of the cytoskeleton in tumor progression [34], [35]. Imaging-based analyses of 3D morphology can therefore be highly informative for in vitro tumour biology, based on cancer cell lines [36], [37] or primary, patient-derived tissue cultures [21], [38]. This approach can be further assisted by mathematical modelling [39]–[41], machine learning and Bayesian networks [42].…”

Section: Introductionmentioning

confidence: 99%

Quantification of Dynamic Morphological Drug Responses in 3D Organotypic Cell Cultures by Automated Image Analysis

et al. 2014

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Glandular epithelial cells differentiate into complex multicellular or acinar structures, when embedded in three-dimensional (3D) extracellular matrix. The spectrum of different multicellular morphologies formed in 3D is a sensitive indicator for the differentiation potential of normal, non-transformed cells compared to different stages of malignant progression. In addition, single cells or cell aggregates may actively invade the matrix, utilizing epithelial, mesenchymal or mixed modes of motility. Dynamic phenotypic changes involved in 3D tumor cell invasion are sensitive to specific small-molecule inhibitors that target the actin cytoskeleton. We have used a panel of inhibitors to demonstrate the power of automated image analysis as a phenotypic or morphometric readout in cell-based assays. We introduce a streamlined stand-alone software solution that supports large-scale high-content screens, based on complex and organotypic cultures. AMIDA (Automated Morphometric Image Data Analysis) allows quantitative measurements of large numbers of images and structures, with a multitude of different spheroid shapes, sizes, and textures. AMIDA supports an automated workflow, and can be combined with quality control and statistical tools for data interpretation and visualization. We have used a representative panel of 12 prostate and breast cancer lines that display a broad spectrum of different spheroid morphologies and modes of invasion, challenged by a library of 19 direct or indirect modulators of the actin cytoskeleton which induce systematic changes in spheroid morphology and differentiation versus invasion. These results were independently validated by 2D proliferation, apoptosis and cell motility assays. We identified three drugs that primarily attenuated the invasion and formation of invasive processes in 3D, without affecting proliferation or apoptosis. Two of these compounds block Rac signalling, one affects cellular cAMP/cGMP accumulation. Our approach supports the growing needs for user-friendly, straightforward solutions that facilitate large-scale, cell-based 3D assays in basic research, drug discovery, and target validation.

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Determining tamoxifen sensitivity using primary breast cancer tissue in collagen-based three-dimensional culture

Cited by 20 publications

References 39 publications

Targeting of Rac GTPases blocks the spread of intact human breast cancer

Targeting of Rac GTPases blocks the spread of intact human breast cancer

The practicalities of using tissue slices as preclinical organotypic breast cancer models

Quantification of Dynamic Morphological Drug Responses in 3D Organotypic Cell Cultures by Automated Image Analysis

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