CD44 and tumor-derived extracellular vesicles (TEVs) : possible gateway to cancer metastasis

Cancer metastasis, the final stage of tumor progression, is a complex process governed by the interplay of multiple types of cells and the tumor microenvironment. One of the aspects of this interplay involves the release of various factors by the tumor cells alone or by forcing other cells to do so. As a consequence of these actions, tumor cells are prepared in favorable conditions for their dissemination and spread to other sites/organs, which guarantees their escape from immunosurveillance and further progression. Tumor-derived extracellular vesicles (TEVs) represent a heterogeneous population of membrane-bound vesicles that are being actively released by different tumors. The array of proteins (i.e., receptors, cytokines, chemokines, etc.) and nucleic acids (i.e., mRNA, miR, etc.) that TEVs can transfer to other cells is often considered beneficial for the tumor’s survival and proliferation. One of the proteins that is associated with many different tumors as well as their TEVs is a cluster of differentiation 44 in its standard (CD44s) and variant (CD44v) form. This review covers the present information regarding the TEVs-mediated CD44s/CD44v transfer/interaction in the context of cancer metastasis. The content and the impact of the transferred cargo by this type of TEVs also are discussed with regards to tumor cell dissemination

Tumour-derived microvesicles contain interleukin-8 and modulate production of chemokines by human monocytes

Background: Tumour-derived microvesicles (TMVs) may interact with cells of the immune system. Our previous observations indicated that TMVs modulate production of cytokines and reactive oxygen species (ROS) by monocytes. This study was designed to determine the role of TMVs in stimulation of chemokine production by human monocytes. Materials and Methods: Chemokines at the mRNA and protein level were detected by real-time PCR and by Western blot, respecively. Chemokine release and chemotaxis of blood leukocytes were analysed by flow cytometry. Matrigel assay was used to determine angiogenesis in a NOD-SCID mice model. Results: TMVs induced secretion of interleukin-8 (CXCL8), monocyte chemoattractant protein-1 (CCL2), macrophage inflammatory protein-1α (CCL3) and MIP-1β (CCL4), and regulated on activation normal T-cells expressed and secreted (CCL5) chemokines and accumulation of their mRNA in monocytes. Moreover, TMVs enhanced angiogenesis in NOD-SCID mice by delivering chemokines and via stimulation of monocytes. In addition, TMVs may be storage for chemokines thus inducing chemotaxis of blood leukocytes. Conclusion: These results further support the role of TMVs in modulation of monocyte biological activity

Tumour-derived microvesicles (TMV) mimic the effect of tumour cells on monocyte subpopulations

Background: Monocytes/macrophages may be affected by tumour cells via cell-to-cell contact, soluble factors and by tumour-derived microvesicles (TMV). Previous observations indicate that TMV interact with monocytes and alter their immunophenotype and activity. This study was designed to determine interactions of TMV with subpopulations (CD14++CD16– and CD14+CD16++) of human monocytes. Methods: Engulfment of TMV by subsets of monocytes was analysed by flow cytometry. Moreover cytokine release and production of reactive oxygen intermediates (ROI) and reactive nitrogen intermediates (RNI) by CD14++CD16– and CD14+CD16++ cells after TMV stimulation was determined. Results: It was found that TMV are engulfed more efficiently by CD14++CD16– than CD14+CD16++ cells. TMV-activated CD14++CD16– cells produce more ROI and interleukin -10 (IL-10) than CD14++CD16+. CD14+CD16++ cells following TMV stimulation showed an increased release of tumour necrosis factor alpha, IL-12p40 and RNI. Conclusion: TMV significantly modulate biological activity of monocyte subsets with a pattern similar to tumour cells. Therefore, TMV mimic the activating effect of tumour cells on monocytes as assessed by release of cytokines, ROI and RNI

Colorectal cancer-derived microvesicles modulate differentiation of human monocytes to macrophages

BACKGROUND: Tumour-derived microvesicles (TMVs) are important players in tumour progression, modulating biological activity of immune cells e.g. lymphocytes, monocytes and macrophages. This phenomenon is particularly interesting in the progression of colon cancer, as macrophages in this type of tumour are relevant for the recovery processes. In the present study, the role of colon cancer cell-derived microvesicles in monocyte differentiation and activity profile (polarization) was investigated. METHODS: Monocyte-derived macrophages (MDM) were differentiated in vitro in the presence of TMVs obtained from colon cancer: Caco-2, SW620, LoVo or SW480 cell lines and analysed according to their morphology and biological functions, as defined by cytokine secretion, reactive oxygen intermediate (ROI) production and cytotoxic activity against respective colon cancer cells. RESULTS: Monocytes differentiated with TMVs exhibited morphological and phenotypical characteristics of macrophages. An early contact (beginning with the first day of the in vitro culture) of monocytes with TMVs resulted in increased IL-10 secretion and only slightly elevated TNF release. Early, or prolonged contact resulted in low ROI production and low cytotoxicity against tumour cells. On the other hand, late contact of MDM with TMVs, stimulated MDM to significant TNF and IL-12 secretion, ROI production and enhanced cytotoxicity against tumour cells in vitro. In addition, differences in MDM response to TMVs from different cell lines were observed (according to cytokine secretion, ROI production and cytotoxicity against tumour cells in vitro). Biological activity, STATs phosphorylation and microRNA profiling of MDMs indicated differences in their polarization/activation status which may suggest mixed polarization type M1/M2 with the predominance of proinflammatory cells after late contact with TMVs. CONCLUSIONS: Macrophage activity (polarization status) may be regulated by contact with not only tumour cells but also with TMVs. Their final polarization status depends on the contact time, and probably on the vesicle “cargo”, as signified by the distinct impact of TMVs which enabled the switching of MDM maturation to regulatory macrophages

Similarities in the general chemical composition of colon cancer cells and their microvesicles investigated by spectroscopic methods-potential clinical relevance

Colon cancer constitutes 33% of all cancer cases in humans and the majority of patients with metastatic colon cancer still have poor prognosis. An important role in cancer development is the communication between cancer and normal cells. This may occur, among others, through extracellular vesicles (including microvesicles) (MVs), which are being released by both types of cells. MVs may regulate a diverse range of biological processes and are considered as useful cancer biomarkers. Herein, we show that similarity in the general chemical composition between colon cancer cells and their corresponding tumor-derived microvesicles (TMVs) does exist. These results have been confirmed by spectroscopic methods for four colon cancer cell lines: HCT116, LoVo, SW480, and SW620 differing in their aggressiveness/metastatic potential. Our results show that Raman and Fourier Transform InfraRed (FTIR) analysis of the cell lines and their corresponding TMVs did not differ significantly in the characterization of their chemical composition. However, hierarchical cluster analysis of the data obtained by both of the methods revealed that only Raman spectroscopy provides results that are in line with the molecular classification of colon cancer, thus having potential clinical relevance

Properties of monocytes generated from haematopoietic CD34+ stem cells from bone marrow of colon cancer patients

Monocytes exhibit direct and indirect antitumour activities and may be potentially useful for various forms of adoptive cellular immunotherapy of cancer. However, blood is a limited source of them. This study explored whether monocytes can be obtained from bone marrow haematopoietic CD34(+) stem cells of colon cancer patients, using previously described protocol of expansion and differentiation to monocytes of cord blood-derived CD34(+) haematopoietic progenitors. Data show that in two-step cultures, the yield of cells was increased approximately 200-fold, and among these cells, up to 60 % of CD14(+) monocytes were found. They consisted of two subpopulations: CD14(++)CD16(+) and CD14(+)CD16(−), at approximately 1:1 ratio, that differed in HLA-DR expression, being higher on the former. No differences in expression of costimulatory molecules were observed, as CD80 was not detected, while CD86 expression was comparable. These CD14(+) monocytes showed the ability to present recall antigens (PPD, Candida albicans) and neoantigens expressed on tumour cells and tumour-derived microvesicles (TMV) to autologous CD3(+) T cells isolated from the peripheral blood. Monocytes also efficiently presented the immunodominant HER-2/neu(369–377) peptide (KIFGSLAFL), resulting in the generation of specific cytotoxic CD8(+) T lymphocytes (CTL). The CD14(++)CD16(+) subset exhibited enhanced cytotoxicity, though nonsignificant, towards tumour cells in vitro. These observations indicate that generation of monocytes from CD34(+) stem cells of cancer patients is feasible. To our knowledge, it is the first demonstration of such approach that may open a way to obtain autologous monocytes for alternative forms of adaptive and adoptive cellular immunotherapy of cancer

The absolute number of circulating nonclassical (CD14+CD16++CD14^{+}CD16^{++}) monocytes negatively correlates with DAS28 and swollen joint count in patients with peripheral spondyloarthritis

A different clinical course and pattern of skeletal involvement in peripheral and axial spondyloarthritis (SpA) suggests a distinct pathophysiology of these 2 phenotypic manifestations of SpA. Monocytes, as part of the innate immune system, seem to play an important role in the pathogenesis of SpA, but the exact inflammatory pathways remain to be elucidated. Regulatory T lymphocytes ($T_{reg}$) and Th17 lymphocytes are also known to influence proinflammatory and anti‑inflammatory reactions. The aim of our study was to compare the absolute numbers of monocyte subpopulations, $T_{reg}$, and Th17 lymphocytes with clinical measures of disease activity in patients with peripheral and axial SpA. We enrolled 21 patients with peripheral SpA and 27 patients with axial SpA diagnosed according to the Assessment of SpondyloArthritis International Society classification criteria, as well as 23 healthy controls. Patients were under 45 years, naïve to synthetic and biological disease‑modifying antirheumatic drugs and without the administration of systemic glucocorticoids. The absolute numbers of classical, intermediate, nonclassical monocytes, $T_{reg}$, and Th17 in peripheral blood were analyzed. Disease activity was assessed using the Ankylosing Spondylitis Disease Activity Score (ASDAS-CRP), Bath Ankylosing Spondylitis Disease Activity Index (BASDAI), and Disease Activity Score 28 (DAS28). In patients with SpA, the number of circulating nonclassical monocytes was decreased in comparison with controls. Only in the peripheral SpA group, a significant negative correlation was found between the concentration of nonclassical monocytes and DAS28 and the number of swollen joints. The 3 groups did not differ in terms of the concentrations of classical or intermediate monocytes and $T_{reg}$ or Th17 lymphocytes. Nonclassical monocytes may play a role in induction and perpetuation of peripheral joint inflammation, at least in peripheral SpA, as cells infiltrating the synovium

Interactions of tumour-derived micro(nano)vesicles with human gastric cancer cells

BACKGROUND: Tumour cells release membrane micro(nano)fragments called tumour-derived microvesicles (TMV) that are believed to play an important role in cancer progression. TMV suppress/modify antitumour response of the host, but there is also some evidence for their direct interaction with cancer cells. In cancer patients TMV are present in body fluid and tumour microenvironment. The present study aimed at characterization of whole types/subpopulations, but not only exosomes, of TMV from newly established gastric cancer cell line (called GC1415) and to define their interactions with autologous cells. METHODS: TMV were isolated from cell cultures supernatants by centrifugation at 50,000×g and their phenotype was determined by flow cytometry. The size of TMV was analysed by dynamic light scattering and nanoparticle tracking analysis, while morphology by transmission electron microscopy and atomic force microscopy. Interactions of TMV with cancer cells were visualized using fluorescence-activated cell sorter, confocal and atomic force microscopy, biological effects by xenografts in NOD SCID mice. RESULTS: Isolated TMV showed expression of CD44H, CD44v6 (hyaluronian receptors), CCR6 (chemokine receptor) and HER-2/neu molecules, exhibited different shapes and sizes (range 60–900 nm, highest frequency of particles with size range of 80–120 nm). TMV attached to autologous cancer cells within 2 h and then were internalized by them at 24 h. CD44H, CD44v6 and CCR6 molecules may play a role in attachment of TMV to cancer cells, while HER-2 associated with CD24 be involved in promoting cancer cells growth. Pre-exposure of cancer cells to TMV resulted in enhancement of tumour growth and cancer cell-induced angiogenesis in NOD SCID mice model. CONCLUSIONS: TMV interact directly with cancer cells serving as macro-messengers and molecular cargo transfer between gastric cancer cells resulting in enhancement of tumour growth. TMV should be considered in future as target of anticancer therapy