Metastasis formation is a hallmark of invasive cancers and it is achieved through the shedding of circulating tumor cells (CTCs) from the primary site into the blood circulation. There, CTCs are ...found as single cells or as multicellular clusters, with clusters carrying an elevated ability to survive within the bloodstream and initiate new metastatic lesions at distant sites. Clusters of CTCs include homotypic clusters made of cancer cells only, as well as heterotypic clusters that incorporate stromal or immune cells along with cancer cells. Both homotypic and heterotypic CTC clusters are characterized by a high metastasis-forming capability, high proliferation rate and by distinct molecular features compared to single CTCs, and their presence in the peripheral circulation of cancer patients is generally associated with a poor prognosis. In this short review, we summarize the current literature that describes homotypic and heterotypic CTC clusters, both in the context of their molecular characteristics as well as their value in the clinical setting. While CTC clusters have only recently emerged as key players in the metastatic process and many aspects of their biology remain to be investigated, a detailed understanding of their vulnerabilities may pave the way towards the generation of new metastasis-suppressing agents.
The metastatic process is an extraordinarily complex step‐by‐step procedure, characterized by many analogies with migratory patterns of humans or animals across our planet. The ongoing interrogation ...of circulating tumor cells (CTCs), caught in the act of spreading from one location to another, is revealing distinct behaviors including biological, physical, and mechanical features that impact on their likelihood to form metastasis. In this viewpoint, I will discuss some of these findings and provide a perspective on the metastatic journey, open questions and opportunities to exploit some of the most recent discoveries for the development of antimetastasis medicines.
Metastasis is how most cancers kill. During the metastatic cascade, cancer cells acquire different properties that influence their likelihood to succeed. In this viewpoint, I summarize some of my views on metastasis‐relevant phenomena, challenges, open questions and opportunities for intervention.
The analysis of circulating tumor cells (CTCs) is an outstanding tool to provide insights into the biology of metastatic cancers, to monitor disease progression and with potential for use in liquid ...biopsy-based personalized cancer treatment. These goals are ambitious, yet recent studies are already allowing a sharper understanding of the strengths, challenges, and opportunities provided by liquid biopsy approaches. For instance, through single-cell-resolution genomics and transcriptomics, it is becoming increasingly clear that CTCs are heterogeneous at multiple levels and that only a fraction of them is capable of initiating metastasis. It also appears that CTCs adopt multiple ways to enhance their metastatic potential, including homotypic clustering and heterotypic interactions with immune and stromal cells. On the clinical side, both CTC enumeration and molecular analysis may provide new means to monitor cancer progression and to take individualized treatment decisions, but their use for early cancer detection appears to be challenging compared to that of other tumor derivatives such as circulating tumor DNA. In this review, we summarize current data on CTC biology and CTC-based clinical applications that are likely to impact our understanding of the metastatic process and to influence the clinical management of patients with metastatic cancer, including new prospects that may favor the implementation of precision medicine.
Metastasis is the major driver of death in patients with cancer. Invasion of surrounding tissues and metastasis have been proposed to initiate following loss of the intercellular adhesion protein, ...E-cadherin
, on the basis of inverse correlations between in vitro migration and E-cadherin levels
. However, this hypothesis is inconsistent with the observation that most breast cancers are invasive ductal carcinomas and express E-cadherin in primary tumours and metastases
. To resolve this discrepancy, we tested the genetic requirement for E-cadherin in metastasis using mouse and human models of both luminal and basal invasive ductal carcinomas. Here we show that E-cadherin promotes metastasis in diverse models of invasive ductal carcinomas. While loss of E-cadherin increased invasion, it also reduced cancer cell proliferation and survival, circulating tumour cell number, seeding of cancer cells in distant organs and metastasis outgrowth. Transcriptionally, loss of E-cadherin was associated with upregulation of genes involved in transforming growth factor-β (TGFβ), reactive oxygen species and apoptosis signalling pathways. At the cellular level, disseminating E-cadherin-negative cells exhibited nuclear enrichment of SMAD2/3, oxidative stress and increased apoptosis. Colony formation of E-cadherin-negative cells was rescued by inhibition of TGFβ-receptor signalling, reactive oxygen accumulation or apoptosis. Our results reveal that E-cadherin acts as a survival factor in invasive ductal carcinomas during the detachment, systemic dissemination and seeding phases of metastasis by limiting reactive oxygen-mediated apoptosis. Identifying molecular strategies to inhibit E-cadherin-mediated survival in metastatic breast cancer cells may have potential as a therapeutic approach for breast cancer.
The crosstalk between cancer cells and the immune system is crucial for disease progression and its therapeutic targeting is providing exciting results, in particular with newly developed immune ...checkpoint inhibitors. Current approaches primarily focus on cellular interactions occurring between tumor cells and T lymphocytes; however, recent data highlight a crucial role of neutrophils in support of tumor progression and suggest yet unexplored treatment opportunities. In this review, we summarize the current understanding of those interactions that occur between neutrophils and cancer cells, focusing on both protumor and antitumor activities of neutrophils at different stages of cancer progression. These include infiltration of neutrophils into the primary tumor, their interactions with circulating tumor cells (CTC) within the bloodstream, and their involvement in the establishment of a metastatic niche. Additionally, we discuss how further investigation of CTCs and their interacting immune cell partners may point towards novel immune checkpoint inhibition strategies and provide new insights on the efficacy of already existing immunotherapies.
The ability of circulating tumor cells (CTCs) to form clusters has been linked to increased metastatic potential. Yet biological features and vulnerabilities of CTC clusters remain largely unknown. ...Here, we profile the DNA methylation landscape of single CTCs and CTC clusters from breast cancer patients and mouse models on a genome-wide scale. We find that binding sites for stemness- and proliferation-associated transcription factors are specifically hypomethylated in CTC clusters, including binding sites for OCT4, NANOG, SOX2, and SIN3A, paralleling embryonic stem cell biology. Among 2,486 FDA-approved compounds, we identify Na+/K+ ATPase inhibitors that enable the dissociation of CTC clusters into single cells, leading to DNA methylation remodeling at critical sites and metastasis suppression. Thus, our results link CTC clustering to specific changes in DNA methylation that promote stemness and metastasis and point to cluster-targeting compounds to suppress the spread of cancer.
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•Binding sites for OCT4, SOX2, NANOG, and SIN3A are hypomethylated in CTC clusters•CTC cluster hypomethylation profile correlates with a poor prognosis in breast cancer•Treatment with FDA-approved Na+/K+-ATPase inhibitors dissociates CTC clusters•Dissociation reverts the methylation profile of CTC clusters and suppresses metastasis
A comparative analysis of the methylation landscape of single and clusters of circulating tumor cells reveals patterns of similarity to embryonic stem cells and identifies pharmacological agents that can target clustering, suppress stemness, and blunt metastatic spreading.
Circulating tumor cell clusters (CTC clusters) are present in the blood of patients with cancer but their contribution to metastasis is not well defined. Using mouse models with tagged mammary ...tumors, we demonstrate that CTC clusters arise from oligoclonal tumor cell groupings and not from intravascular aggregation events. Although rare in the circulation compared with single CTCs, CTC clusters have 23- to 50-fold increased metastatic potential. In patients with breast cancer, single-cell resolution RNA sequencing of CTC clusters and single CTCs, matched within individual blood samples, identifies the cell junction component plakoglobin as highly differentially expressed. In mouse models, knockdown of plakoglobin abrogates CTC cluster formation and suppresses lung metastases. In breast cancer patients, both abundance of CTC clusters and high tumor plakoglobin levels denote adverse outcomes. Thus, CTC clusters are derived from multicellular groupings of primary tumor cells held together through plakoglobin-dependent intercellular adhesion, and though rare, they greatly contribute to the metastatic spread of cancer.
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•CTC clusters originate as oligoclonal groups of cells from the primary tumor•CTC clusters exhibit increased metastatic propensity compared to single CTCs•Abundance of CTC clusters in patients denotes adverse outcome•Plakoglobin mediates CTC cluster formation, enhancing metastatic spread
Circulating tumor cell (CTC) clusters originating from oligoclonal primary tumor cells exhibit increased metastatic potential compared to single CTCs and indicate adverse outcome in cancer patients.
Circulating tumor cells (CTCs) are shed into the bloodstream from primary tumors, but only a small subset of these cells generates metastases. We conducted an in vivo genome-wide CRISPR activation ...screen in CTCs from breast cancer patients to identify genes that promote distant metastasis in mice. Genes coding for ribosomal proteins and regulators of translation were enriched in this screen. Overexpression of
, which encodes a component of the large ribosomal subunit, increased metastatic growth in multiple organs and selectively enhanced translation of other ribosomal proteins and cell cycle regulators. RNA sequencing of freshly isolated CTCs from breast cancer patients revealed a subset with strong ribosome and protein synthesis signatures; these CTCs expressed proliferation and epithelial markers and correlated with poor clinical outcome. Therapies targeting this aggressive subset of CTCs may merit exploration as potential suppressors of metastatic progression.
Circulating tumor cells (CTCs) are shed from solid cancers in the form of single or clustered cells, and the latter display an extraordinary ability to initiate metastasis. Yet, the biological ...phenomena that trigger the shedding of CTC clusters from a primary cancerous lesion are poorly understood. Here, when dynamically labeling breast cancer cells along cancer progression, we observe that the majority of CTC clusters are undergoing hypoxia, while single CTCs are largely normoxic. Strikingly, we find that vascular endothelial growth factor (VEGF) targeting leads to primary tumor shrinkage, but it increases intra-tumor hypoxia, resulting in a higher CTC cluster shedding rate and metastasis formation. Conversely, pro-angiogenic treatment increases primary tumor size, yet it dramatically suppresses the formation of CTC clusters and metastasis. Thus, intra-tumor hypoxia leads to the formation of clustered CTCs with high metastatic ability, and a pro-angiogenic therapy suppresses metastasis formation through prevention of CTC cluster generation.
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•Hypoxia leads to cell-cell junction upregulation and intravasation of CTC clusters•Hypoxic CTC clusters are highly metastatic compared to normoxic CTCs•Increase in intra-tumor hypoxia leads to accelerated metastasis•Treatment with EpB2 reduces hypoxia and prevents CTC cluster formation
Donato et al. show that intra-tumor hypoxia leads to cell-cell junction upregulation and formation of hypoxic CTC clusters with high metastatic ability. Treatment with EphrinB2 improves tumor vascularization and decreases hypoxia, leading to a reduced CTC cluster shedding rate and suppression of metastasis.
The epithelial to mesenchymal transition (EMT) has been proposed to contribute to the metastatic spread of breast cancer cells. EMT-promoting transcription factors determine a continuum of different ...EMT states. In contrast, estrogen receptor α (ERα) helps to maintain the epithelial phenotype of breast cancer cells and its expression is crucial for effective endocrine therapies. Determining whether and how EMT-associated transcription factors such as ZEB1 modulate ERα signaling during early stages of EMT could promote the discovery of therapeutic approaches to suppress metastasis. Here we show that, shortly after induction of EMT and while cells are still epithelial, ZEB1 modulates ERα-mediated transcription induced by estrogen or cAMP signaling in breast cancer cells. Based on these findings and our ex vivo and xenograft results, we suggest that the functional interaction between ZEB1 and ERα may alter the tissue tropism of metastatic breast cancer cells towards bone.
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