Optimizing metastatic-cascade-dependent Rac1 targeting in breast cancer: Guidance using optical window intravital FRET imaging

Floerchinger, Alessia; Murphy, Kendelle J.; Latham, Sharissa L.; Warren, Sean; McCulloch, Andrew T.; Lee, Young-Kyung; Stoehr, Janett; Mélénec, Pauline; Guaman, Cris S.; Metcalf, Xanthe L.; Lee, Victoria; Zaratzian, Anaiis; Silva, Andrew Da; Tayao, Michael; Rolo, Sonia; Phimmachanh, Monica; Sultani, Ghazal; McDonald, Laura; Mason, Susan; Ferrari, Nicola; Ooms, Lisa M; Johnsson, Anna‐Karin; Spence, Heather J.; Olson, Michael F.; Machesky, Laura M.; Sansom, Owen J.; Morton, Jennifer P.; Mitchell, Christina A.; Samuel, Michael S.; Croucher, David R.; Welch, Heidi; Blyth, Karen; Caldon, C. Elizabeth; Herrmann, David; Anderson, Kurt I.; Timpson, Paul; Nobis, Max

doi:10.1016/j.celrep.2021.109689

Cited by 16 publications

(16 citation statements)

References 141 publications

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“…Finally, we sought to determine whether the observed changes in the 3D organotypic matrices as a result of collagen XII depletion would affect cancer cell invasion in the 3D setting. PyMT cancer cells 79 were seeded onto, and allowed to invade into the CAF-remodelled organotypic matrices following removal of CAFs (see ‘Methods’) (Fig. 6h ).…”

Section: Resultsmentioning

confidence: 99%

“…PyMT 20065 cancer cells were derived with the support of Dr Karen Blyth at the CRUK Beatson Institute in Glasgow through the SEARCHbreast initiative ( https://searchbreast.org/ ), a resource to facilitate sharing of archived material derived from in vivo breast cancer models. PyMT 20065 cancer cells were maintained in DMEM supplemented with 10% FBS, 1% penicillin/streptomycin, 5 µg/mL insulin, 10 ng/mL epidermal growth factor and 10 ng/mL Cholera Toxin A as previously described 79 . All cells were kept at 37 °C in 20% O 2 and 5% CO 2 .…”

Section: Methodsmentioning

confidence: 99%

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Temporal profiling of the breast tumour microenvironment reveals collagen XII as a driver of metastasis

et al. 2022

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The tumour stroma, and in particular the extracellular matrix (ECM), is a salient feature of solid tumours that plays a crucial role in shaping their progression. Many desmoplastic tumours including breast cancer involve the significant accumulation of type I collagen. However, recently it has become clear that the precise distribution and organisation of matrix molecules such as collagen I is equally as important in the tumour as their abundance. Cancer-associated fibroblasts (CAFs) coexist within breast cancer tissues and play both pro- and anti-tumourigenic roles through remodelling the ECM. Here, using temporal proteomic profiling of decellularized tumours, we interrogate the evolving matrisome during breast cancer progression. We identify 4 key matrisomal clusters, and pinpoint collagen type XII as a critical component that regulates collagen type I organisation. Through combining our proteomics with single-cell transcriptomics, and genetic manipulation models, we show how CAF-secreted collagen XII alters collagen I organisation to create a pro-invasive microenvironment supporting metastatic dissemination. Finally, we show in patient cohorts that collagen XII may represent an indicator of breast cancer patients at high risk of metastatic relapse.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Temporal profiling of the breast tumour microenvironment reveals collagen XII as a driver of metastasis

et al. 2022

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“…Intriguingly, the presence of CD163+ Tim4+ macrophages caused spheroid compaction much like that caused by Rac1OE (see Figure 4 ). Such Rac1-driven clustering of cells in circulation may promote the increased cell survival under sheer stress experienced in the bloodstream during metastatic dissemination as suggested by the OCMetSim-Spheroids model and supported by in vitro studies of invasive breast cancer cells ( Floerchinger et al. , 2021 ).…”

Section: Discussionmentioning

confidence: 84%

“…Model simulations of tumor cell crowding showed Rac1 OE cells to be resistant to the effects of cell crowding while higher rates of death were observed for Rac1 CRISPR-Cas9 KD cells. These findings are interesting in view of recent live animal imaging studies of the invasive MMTV-PyMT breast cancer model showing that Rac1 activity (measured using a Rac1 fluorescence resonance energy transfer sensor) is spatially regulated at distinct perivascular sites within locally invasive tumors ( Floerchinger et al. , 2021 ).…”

Section: Discussionmentioning

confidence: 90%

Agent-based modeling predicts RAC1 is critical for ovarian cancer metastasis

Rivera

Toledo-Jacobo

Romero

et al. 2022

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Experimental and computational studies pinpoint rate limiting step(s) in metastasis governed by Rac1. Using ovarian cancer cell and animal models, Rac1 expression was manipulated and quantitative measurements of cell-cell and cell-substrate adhesion, cell invasion, mesothelial clearance and peritoneal tumor growth discriminated the behaviors most highly influenced by Rac1. The experimental data were used to parameterize an agent-based computational model simulating peritoneal niche colonization, intravasation and hematogenous metastasis to distant organs. Increased ovarian cancer cell survival afforded by the more rapid adhesion and intravasation upon Rac1 overexpression is predicted to increase the numbers and the rates at which tumor cells disseminate to distant sites. Surprisingly, crowding of cancer cells along the blood vessel was found to decrease the numbers of cells reaching a distant niche irrespective of Rac1 overexpression or knockdown, suggesting sites for tumor cell intravasation are rate limiting and become accessible if cells intravasate rapidly or are displaced due to diminished viability. Modeling predictions were confirmed through animal studies of Rac1 dependent metastasis to the lung. Collectively, the experimental and modeling approaches identify cell adhesion, rapid intravasation, and survival in the blood as parameters in the ovarian metastatic cascade that are most critically dependent on Rac1.

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“…This was coupled with high-throughput assessment of primary Akt-FRET cancer cells in three-dimensional (3D) organotypic invasion assays to validate the anti-invasive capacity of AKT-PI3K pathway inhibition. Furthermore, using optical window imaging ( 18 – 21 ) in breast cancer GEMMs, we mapped drug targeting in an AKT hyperactivated setting, demonstrating the utility of the Akt-FRET biosensor mouse in preclinical imaging of targeted therapies in a longitudinal manner ( 22 – 24 ).…”

Section: Introductionmentioning

confidence: 99%

Monitoring AKT activity and targeting in live tissue and disease contexts using a real-time Akt-FRET biosensor mouse

et al. 2023

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Aberrant AKT activation occurs in a number of cancers, metabolic syndrome, and immune disorders, making it an important target for the treatment of many diseases. To monitor spatial and temporal AKT activity in a live setting, we generated an Akt-FRET biosensor mouse that allows longitudinal assessment of AKT activity using intravital imaging in conjunction with image stabilization and optical window technology. We demonstrate the sensitivity of the Akt-FRET biosensor mouse using various cancer models and verify its suitability to monitor response to drug targeting in spheroid and organotypic models. We also show that the dynamics of AKT activation can be monitored in real time in diverse tissues, including in individual islets of the pancreas, in the brown and white adipose tissue, and in the skeletal muscle. Thus, the Akt-FRET biosensor mouse provides an important tool to study AKT dynamics in live tissue contexts and has broad preclinical applications.

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Optimizing metastatic-cascade-dependent Rac1 targeting in breast cancer: Guidance using optical window intravital FRET imaging

Cited by 16 publications

References 141 publications

Temporal profiling of the breast tumour microenvironment reveals collagen XII as a driver of metastasis

Temporal profiling of the breast tumour microenvironment reveals collagen XII as a driver of metastasis

Agent-based modeling predicts RAC1 is critical for ovarian cancer metastasis

Monitoring AKT activity and targeting in live tissue and disease contexts using a real-time Akt-FRET biosensor mouse

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