Circulating tumour cells: The Good, the Bad and the Ugly Bates, Mark; Mohamed, Bashir M.; Ward, Mark P. ...
Biochimica et biophysica acta. Reviews on cancer,
March 2023, 2023-03-00, 20230301, Volume:
1878, Issue:
2
Journal Article
Peer reviewed
Open access
This review is an overview of the current knowledge regarding circulating tumour cells (CTCs), which are potentially the most lethal type of cancer cell, and may be a key component of the metastatic ...cascade. The clinical utility of CTCs (the "Good"), includes their diagnostic, prognostic, and therapeutic potential. Conversely, their complex biology (the "Bad"), including the existence of CD45+/EpCAM+ CTCs, adds insult to injury regarding their isolation and identification, which in turn hampers their clinical translation. CTCs are capable of forming microemboli composed of both non-discrete phenotypic populations such as mesenchymal CTCs and homotypic and heterotypic clusters which are poised to interact with other cells in the circulation, including immune cells and platelets, which may increase their malignant potential. These microemboli (the "Ugly") represent a prognostically important CTC subset, however, phenotypic EMT/MET gradients bring additional complexities to an already challenging situation.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Circulating tumour cells (CTCs) are a critical intermediate step in the process of cancer metastasis. The reliability of CTC isolation/purification has limited both the potential to report on ...metastatic progression and the development of CTCs as targets for therapeutic intervention. Here we report a new methodology, which optimises the culture conditions for CTCs using primary cancer cells as a model system. We exploited the known biology that CTCs thrive in hypoxic conditions, with their survival and proliferation being reliant on the activation of hypoxia-inducible factor 1 alpha (HIF-1α). We isolated epithelial-like and quasi-mesenchymal CTC phenotypes from the blood of a cancer patient and successfully cultured these cells for more than 8 weeks. The presence of CTC clusters was required to establish and maintain long-term cultures. This novel methodology for the long-term culture of CTCs will aid in the development of downstream applications, including CTC theranostics.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
High-grade serous ovarian cancer (HGSOC) is the most prevalent and deadliest subtype of epithelial ovarian cancer (EOC), killing over 140,000 people annually. Morbidity and mortality are compounded ...by a lack of screening methods, and recurrence is common. Plasminogen-activator-inhibitor 1 (PAI-1, the protein product of SERPIN E1) is involved in hemostasis, extracellular matrix (ECM) remodeling, and tumor cell migration and invasion. Overexpression is associated with poor prognosis in EOC. Platelets significantly increase PAI-1 in cancer cells
, and may contribute to the hematogenous metastasis of circulating tumor cells (CTCs). CTCs are viable tumor cells that intravasate and travel through the circulation-often aided by platelets - with the potential to form secondary metastases. Here, we provide evidence that PAI-1 is central to the platelet-cancer cell interactome, and plays a role in the metastatic cascade.
SK-OV-3 cells where PAI-1 had been silenced, treated with healthy donor platelets, and treated with platelet-conditioned medium were used as an
model of metastatic EOC. Gene expression analysis was performed using RNA-Seq data from untreated cells and cells treated with PAI-1 siRNA or negative control, each with and without platelets. Four cohorts of banked patient plasma samples (n = 239) were assayed for PAI-1 by ELISA. Treatment-naïve (TN) whole blood (WB) samples were evaluated for CTCs in conjunction with PAI-1 evaluation in matched plasma.
Significant phenotypic changes occurring when PAI-1 was silenced and when platelets were added to cells were reflected by RNA-seq data, with PAI-1 observed to be central to molecular mechanisms of EOC metastasis. Increased proliferation was observed in cells treated with platelets. Plasma PAI-1 significantly correlated with advanced disease in a TN cohort, and was significantly reduced in a neoadjuvant chemotherapy (NACT) cohort. PAI-1 demonstrated a trend towards significance in overall survival (OS) in the late-stage TN cohort, and correlation between PAI-1 and neutrophils in this cohort was significant. 72.7% (16/22) of TN patients with plasma PAI-1 levels higher than OS cutoff were CTC-positive. These data support a central role for PAI-1 in EOC metastasis, and highlight PAI-1's potential as a biomarker, prognostic indicator, or gauge of treatment response in HGSOC.
•Platelet cancer cell interactions are a key factor in driving the pro-metastatic phenotype.•Platelet cancer cell interactions appear to be mediated by 5 key genes which have established roles in ...metastasis.•Targeting these mediators of metastasis could improve outcomes for cancer patients.
Tumour metastasis accounts for over 90% of cancer related deaths. The platelet is a key blood component, which facilitates efficient metastasis. This study aimed to understand the molecular mechanisms involved in tumour-platelet cell interactions.
The interaction between cancer cells and platelets was examined in 15 epithelial cell lines, representing 7 cancer types. Gene expression analysis of EMT-associated and cancer stemness genes was performed by RT-PCR. Whole transcriptome analysis (WTA) was performed using Affymetrix 2.0ST arrays on a platelet co-cultured ovarian model.
Platelet adhesion and activation occurred across all tumour types. WTA identified increases in cellular movement, migration, invasion, adhesion, development, differentiation and inflammation genes and decreases in processes associated with cell death and survival following platelet interaction. Increased invasive capacity was also observed in a subset of cell lines. A cross-comparison with a platelet co-cultured mouse model identified 5 common altered genes; PAI-1, PLEK2, CD73, TNC, and SDPR.
Platelet cancer cell interactions are a key factor in driving the pro-metastatic phenotype and appear to be mediated by 5 key genes which have established roles in metastasis. Targeting these metastasis mediators could improve cancer patient outcomes.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Growth hormone (GH) promotes growth in children, but is also essential for bone strength, body composition, metabolic factors, such as lipid profile, and maintenance of quality of life. The Merck ...KGaA (Germany) funded "360° GH in Europe" meeting, held in Lisbon, Portugal, in June 2016, comprised three sessions entitled "
," "
," and "
." The scientific program covered all stages of pediatric GH treatment, and reported here are the outcomes of the third session of the meeting, which considered transition from pediatric GH treatment to teenage and young adult GH therapy. A large number of patients with chronic diseases, including GH deficiency, drop out of therapy during the transition period. Multiple factors are associated with this, such as lack of understanding of the disease process, insufficient knowledge of treatment options, the patient becoming more independent, and requirement for interaction with a new set of health-care workers. Education regarding disease management and treatment options should be provided from an early age and right through the transition period. However, endocrine specialists will view the transition period differently, depending on whether they are pediatric endocrinologists who mainly deal with congenital diseases, in which auxology is important, or adult endocrinologists who are more concerned with body composition and metabolic factors. View points of both a pediatric and an adult endocrine specialist are presented, together with a case study outlining practical aspects of transition. It was noted in the meeting discussion that having one person to guide a patient through transition from an early age is important, but may be constrained by various factors such as finances, and options will differ by country.
A new model of paediatric nursing, funded initially by a charitable organisation working in partnership with UK healthcare providers, was implemented to support children living with serious long-term ...conditions. This study explored, from the perspective of multiple stakeholders, the impact of services provided by 21 ‘Roald Dahl Specialist Nurses’ (RDSN) within 14 NHS Trust hospitals.
A Mixed Methods Exploratory design commenced with interviews with RDSNs (n = 21) and their managers (n = 15), alongside a medical clinician questionnaire (n = 17). Initial themes (constructivist grounded theory) were validated through four RDSN focus groups, and informed development of an online survey of parents (n = 159) and children (n = 32). Findings related to impact were integrated using a six-step triangulation protocol.
Zones of significant impact included: Improving quality and experience of care; Improved efficiencies and cost-effectiveness; Provision of holistic family-centred care; and Impactful leadership and innovation. The RDSNs forged networks across inter-agency boundaries to safeguard the child and enhance the family experience of care. RDSNs delivered improvements across a range of metrics, and were valued for their emotional support, care navigation and advocacy.
Children living with serious long-term conditions have complex needs. Regardless of the specialty, location, organisation or service focus, this new model of care crosses organisational and inter-agency boundaries to ensure that the healthcare delivered has maximum impact. It has a profoundly positive impact on families.
This integrated and family-centred model of care is strongly recommended for children with complex needs crossing organisational divides.
•The scope and scale of the RDSN role is wider than a ‘traditional’ nursing role.•The charitable funding model supports RDSNs to bridge organisational boundaries.•RDSNs are highly valued by families for emotional support and care navigation.•RDSNs are pivotal in preventing avoidable hospital admissions or visits to A&E.•The RDSNs enable children and families to become more involved in decision-making.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Purpose
The association between pathological complete response (pCR) in patients receiving neoadjuvant chemotherapy (NAC) for breast cancer and Circulating Tumour Cells (CTCs) is not clear. The aim ...of this study was to assess whether CTC enumeration could be used to predict pathological response to NAC in breast cancer as measured by the Miller–Payne grading system.
Methods
Twenty-six patients were recruited, and blood samples were taken pre- and post-NAC. CTCs were isolated using the ScreenCell device and stained using a modified Giemsa stain. CTCs were enumerated by 2 pathologists and classified as single CTCs, doublets, clusters/microemboli and correlated with the pathological response as measured by the Miller–Payne grading system.
χ
2
or ANOVA was performed in SPSS 24.0 statistics software for associations.
Results
89% of patients had invasive ductal carcinoma (IDC) and 11% invasive lobular carcinoma (ILC). At baseline 85% of patients had CTCs present, median 7 (0–161) CTCs per 3 ml of whole blood. Post-chemotherapy, 58% had an increase in CTCs. This did not correlate with the Miller–Payne grade of response. No significant association was identified between the number of CTCs and clinical characteristics; however, we did observe a correlation between pre-treatment CTC counts and body mass index,
p
< 0.05.
Conclusions
Patients with a complete response to NAC still had CTCs present, suggesting enumeration is not sufficient to aid surgery stratification. Additional characterisation and larger studies are needed to further characterise CTCs isolated pre- and post-chemotherapy. Long-term follow-up of these patients will determine the significance of CTCs in NAC breast cancer patients.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OBVAL, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Abstract
Background: Circulating tumour cells (CTCs) are silent precursors of metastatic disease that utilise various mechanisms to survive in circulation and metastasise to distal sites. Classical ...CTC detection relies on EpCam affinity-based technologies; however, CTCs are highly heterogeneous and often undergo EMT. Recent research highlights the ability of platelets and neutrophils to ‘cloak' CTCs and crosstalk to aid in their proliferation and survival in circulation.
Aims: Assess the role of platelets and neutrophils in the characterisation of CTCs.
Methods: Cell lines: Breast cancer (MCF-7) and ovarian cancer (SKOV-3, OVCAR3, OAW42) cells were exposed to healthy donor isolated platelets and neutrophils for 24h. EMT and immune evasion gene assays, flow cytometry analysis and cell proliferation assays were performed following co-culture. Cell line-blood spike-in experiments were performed for gating strategy optimisation.Patient samples: Blood specimens were prospectively collected from breast and ovarian cancer patients (central venous and peripheral blood). CTCs and immune cells were then isolated using a ClearCell FX microfluidic device. FX isolated cells were immunophenotyped by either immunofluorescence (IF) or flow cytometry. Single cells were index sorted using BD FACS Melody for subsequent scRNAseq.
Results: Platelet cloaking altered EpCam, PLEK2, CCL2 and TWIST1 mRNA expression in MCF-7 cells. Platelet and neutrophil co-culture altered EpCam, PAI-1 and PD-L1 expression in ovarian cancer cell lines. Healthy neutrophils co-cultured with ovarian cancer cells increased cell viability, while induction of NETosis increased cancer cell proliferation. CTCs isolated from peripheral blood in breast and ovarian patients were EpCam/panCK+/CD45- by IF. Immunophenotyped CTCs were identified as EpCam+/E-Cadherin+/CD45-, N-Cadherin+/CD45- and as PD-L1+/EpCam+/E-Cadherin+/CD45- cells by flow cytometry. We also identified EpCam+/CD45+/CD66b+ cells in ovarian central blood. Isolated classical and non-classical CTCs were sorted based on their different immunophenotypes for downstream scRNAseq using BD Precise Whole Transcriptome Assays.
Conclusion: Platelets and neutrophils alter the expression of markers used in the identification of CTCs. Neutrophils were found to increase the proliferation of cancer cells as well as increase PD-L1 expression. We identified a heterogeneous population of CTCs across different patients and found that CTCs from central venous blood have increased numbers of non-classical CTCs (CD66b+/EpCam+ cells). Molecular scRNAseq signatures of CTC subsets form the basis for identifying the most clinically relevant CTCs in circulation. Our approach may improve diagnostic accuracy, allow for immunophenotyping of cloaked CTCs, and provide prognostic information.
Citation Format: Mark P. Ward, Bashir M. Mohamed, Laura Kane, Mark Bates, Janina Berghoff, Cathy L. Spillane, Tanya Kelly, John Kennedy, Feras A. Saadeh, Karsten Hokamp, Noreen Gleeson, Orla Sheils, Cara Martin, Michael Gallagher, Sean Hannify, Eric P. Dixon, Sharon A. O'Toole, John J. O'Leary. Influence of platelets and neutrophils on circulating tumour cells abstract. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 764.
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