Project Publications

High blood flow shear stress values are associated with circulating tumor cells cluster disaggregation in a multi-channel microfluidic device.
Marrella A, Fedi A, Varani G, Vaccari I, Fato M, Firpo G, Guida P, Aceto N, Scaglione S.

» Abstract

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.

PLoS One. 2021 Jan 14;16(1):e0245536. doi: 10.1371/journal.pone.0245536.
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Bring along your friends: Homotypic and heterotypic circulating tumor cell clustering to accelerate metastasis.
Aceto N.

» Abstract

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.

Biomed J. 2020 Feb;43(1):18-23. doi: 10.1016/j.bj.2019.11.002. Epub 2020 Feb 11. PMID: 32200952; PMCID: PMC7090281.
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Tracking cancer progression: from circulating tumor cells to metastasis.
Castro-Giner F, Aceto N.

» Abstract

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 make 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.

Genome Med. 2020 Mar 19;12(1):31. doi: 10.1186/s13073-020-00728-3. PMID: 32192534; PMCID: PMC7082968.
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Quantification of Protein Secretion from Circulating Tumor Cells in Microfluidic Chambers
Armbrecht L, Rutschmann O, Szczerba BM, Nikoloff J, Aceto N, Dittrich PS.

» Abstract

Cancer cells can be released from a cancerous lesion and migrate into the circulatory system, from whereon they may form metastases at distant sites. Today, it is possible to infer cancer progression and treatment efficacy by determining the number of circulating tumor cells (CTCs) in the patient’s blood at multiple time points; further valuable information about CTC phenotypes remains inaccessible. In this article, a microfluidic method for integrated capture, isolation, and analysis of membrane markers, as well as quantification of proteins secreted by single CTCs and CTC clusters, is introduced. CTCs are isolated from whole blood with extraordinary efficiencies above 95% using dedicated trapping structures that allow co-capture of functionalized magnetic beads to assess protein secretion. The patform is tested with multiple breast cancer cell lines spiked into human blood and mouse-model-derived CTCs. In addition to immunostaining, the secretion level of granulocyte growth-stimulating factor (G-CSF), which is shown to be involved in neutrophil recruitment, is quantified The bead-based assay provides a limit of detection of 1.5 ng mL-1 or less than 3700 molecules per cell. Employing barcoded magnetic beads, this platform can be adapted for multiplexed analysis and can enable comprehensive functional CTC profiling in the future.

Adv Sci (Weinh). 2020 Apr 24;7(11):1903237. doi: 10.1002/advs.201903237.
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Circulating tumor cells: Ready for translation?
Diamantopoulou Z, Castro-Giner F, Aceto N.

» Abstract

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 with 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 the prevention of CTC cluster generation.

J Exp Med. 2020 Aug 3;217(8):e20200356. doi: 10.1084/jem.20200356. PMID: 32644115; PMCID: PMC7398171.
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Hypoxia Triggers the Intravasation of Clustered Circulating Tumor Cells.
Donato C, Kunz L, Castro-Giner F, Paasinen-Sohns A, Strittmatter K, Szczerba BM, Scherrer R, Di Maggio N, Heusermann W, Biehlmaier O, Beisel C, Vetter M, Rochlitz C, Weber WP, Banfi A, Schroeder T, Aceto N.

» Abstract

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 the prevention of CTC cluster generation.

Cell Rep. 2020 Sep 8;32(10):108105. doi: 10.1016/j.celrep.2020.108105. PMID: 32905777; PMCID: PMC7487783
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Circulating Tumor Cell-Neutrophil Tango along the Metastatic Process.
Saini M, Szczerba BM, Aceto N.

» Abstract

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.

Cancer Res. 2019 Dec 15;79(24):6067-6073. doi: 10.1158/0008-5472.CAN-19-1972. Epub 2019 Sep 16. LINK

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To Be Taken in Count: Prostatic Tumor Cells Break Free upon Needle Biopsy.
Saini M, Aceto N.
Clin Chem. 2020 Jan 1;66(1):6-8. doi: 10.1373/clinchem.2019.311696. LINK

 

Therapeutic vascularization in regenerative medicine.
Gianni-Barrera R, Di Maggio N, Melly L, Burger MG, Mujagic E, Gürke L, Schaefer DJ, Banfi A.

» Abstract

Therapeutic angiogenesis, that is, the generation of new vessels by delivery of specific factors, is required both for rapid vascularization of tissue‐engineered constructs and to treat ischemic conditions. Vascular endothelial growth factor (VEGF) is the master regulator of angiogenesis. However, the uncontrolled expression can lead to aberrant vascular growth and vascular tumors (angiomas). Major challenges to fully exploit VEGF potency for therapy include the need to precisely control in vivo distribution of growth factor dose and duration of expression. In fact, the therapeutic window of VEGF delivery depends on its amount in the microenvironment around each producing cell rather than on the total dose, since VEGF remains tightly bound to extracellular matrix (ECM). On the other hand, short‐term expression of less than about 4 weeks leads to unstable vessels, which promptly regress following cessation of the angiogenic stimulus. Here, we will briefly overview some key aspects of the biology of VEGF and angiogenesis and discuss their therapeutic implications with a particular focus on approaches using gene therapy, genetically modified progenitors, and ECM engineering with recombinant factors. Lastly, we will present recent insights into the mechanisms that regulate vessel stabilization and the switch between normal and aberrant vascular growth after VEGF delivery, to identify novel molecular targets that may improve both safety and efficacy of therapeutic angiogenesis.

Stem Cells Transl Med. 2020 Apr;9(4):433-444. doi: 10.1002/sctm.19-0319. Epub 2020 Jan 10. LINK

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3D Perfusable Hydrogel Recapitulating the Cancer Dynamic Environment to in Vitro Investigate Metastatic Colonization.
Vitale C, Fedi A, Marrella A, Varani G, Fato M, Scaglione S.

» Abstract

Despite the great advances that the tissue engineering field has experienced over the last two decades, the amount of in vitro engineered tissues that have reached a stage of a clinical trial is limited. While many challenges are still to be overcome, the lack of vascularization represents a major milestone if tissues bigger than approximately 200 µm are to be transplanted. Cell survival and homeostasis are to a large extent conditioned by the oxygen and nutrient transport (as well as waste removal) by blood vessels on their proximity and spontaneous vascularization in vivo is a relatively slow process, leading all together to necrosis of implanted tissues. Thus, in vitro vascularization appears to be a requirement for the advancement of the field. One of the main approaches to this end is the formation of vascular templates that will develop in vitro together with the targeted engineered tissue. Bioprinting, a fast and reliable method for the deposition of cells and materials in a precise manner, appears like an excellent fabrication technique. In this review, we provide a comprehensive background to the fields of vascularization and bioprinting, providing details on the current strategies, cell sources, materials and outcomes of these studies.

Polymers (Basel). 2020 Oct 24;12(11):E2467. doi: 10.3390/polym12112467. LINK

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Bioprinting Vasculature: Materials, Cells and Emergent Techniques.
Tomasina C, Bodet T, Mota C, Moroni L, Camarero-Espinosa S.

» Abstract


Despite the great advances that the tissue engineering field has experienced over the last two decades, the amount of in vitro engineered tissues that have reached a stage of a clinical trial is limited. While many challenges are still to be overcome, the lack of vascularization represents a major milestone if tissues bigger than approximately 200 µm are to be transplanted. Cell survival and homeostasis are to a large extent conditioned by the oxygen and nutrient transport (as well as waste removal) by blood vessels on their proximity and spontaneous vascularization in vivo is a relatively slow process, leading all together to necrosis of implanted tissues. Thus, in vitro vascularization appears to be a requirement for the advancement of the field. One of the main approaches to this end is the formation of vascular templates that will develop in vitro together with the targeted engineered tissue. Bioprinting, a fast and reliable method for the deposition of cells and materials in a precise manner, appears like an excellent fabrication technique. In this review, we provide a comprehensive background to the fields of vascularization and bioprinting, providing details on the current strategies, cell sources, materials, and outcomes of these studies.

Materials (Basel). 2019 Aug 23;12(17):2701. doi: 10.3390/ma12172701. LINK

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Cancer Diagnosis Using a Liquid Biopsy: Challenges and Expectations.
Castro-Giner F, Gkountela S, Donato C, Alborelli I, Quagliata L, Ng CKY, Piscuoglio S, Aceto N

» Abstract


The field of cancer diagnostics has recently been impacted by new and exciting developments in the area of liquid biopsy. A liquid biopsy is a minimally invasive alternative to surgical biopsies of solid tissues, typically achieved through the withdrawal of a blood sample or other body fluids, allowing the interrogation of tumor-derived material including circulating tumor cells (CTCs) and circulating tumor DNA (ctDNA) fragments that are present at a given time point. In this short review, we discuss a few studies that summarize the state-of-the-art in the liquid biopsy field from a diagnostic perspective and speculate on current challenges and expectations of implementing liquid biopsy testing for cancer diagnosis and monitoring in the clinical setting.

Diagnostics (Basel). 2018 May 9;8(2):31. doi: 10.3390/diagnostics8020031LINK

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