Intrinsic signalling factors associated with cancer cell-cell fusion.

Cellular fusion e.g. between cancer cells and normal cells represents a stepwise process that is tightly regulated. During a pre-hybrid preparation program somatic cells and/or cancer cells are promoted to a pro-fusogenic state as a prerequisite to prepare a fusion process. A pro-fusogenic state requires significant changes including restructure of the cytoskeleton, e.g., by the formation of F-actin. Moreover, distinct plasma membrane lipids such as phosphatidylserine play an important role during cell fusion. In addition, the expression of distinct fusogenic factors such as syncytins and corresponding receptors are of fundamental importance to enable cellular mergers. Subsequent hybrid formation and fusion are followed by a post-hybrid selection process. Fusion among normal cells is important and often required during organismal development. Cancer cells fusion appears more rarely and is associated with the generation of new cancer hybrid cell populations. These cancer hybrid cells contribute to an elevated tumour plasticity by altered metastatic behaviour, changes in therapeutic and apoptotic responses, and even in the formation of cancer stem/ initiating cells. While many parts within this multi-step cascade are still poorly understood, this review article predominantly focusses on the intracellular necessities for fusion among cancer cells or with other cell populations of the tumour microenvironment. Video Abstract.

Extracellular Events Involved in Cancer Cell-Cell Fusion.

Fusion among different cell populations represents a rare process that is mediated by both intrinsic and extracellular events. Cellular hybrid formation is relayed by orchestrating tightly regulated signaling pathways that can involve both normal and neoplastic cells. Certain important cell merger processes are often required during distinct organismal and tissue development, including placenta and skeletal muscle. In a neoplastic environment, however, cancer cell fusion can generate new cancer hybrid cells. Following survival during a subsequent post-hybrid selection process (PHSP), the new cancer hybrid cells express different tumorigenic properties. These can include elevated proliferative capacity, increased metastatic potential, resistance to certain therapeutic compounds, and formation of cancer stem-like cells, all of which characterize significantly enhanced tumor plasticity. However, many parts within this multi-step cascade are still poorly understood. Aside from intrinsic factors, cell fusion is particularly affected by extracellular conditions, including an inflammatory microenvironment, viruses, pH and ionic stress, hypoxia, and exosome signaling. Accordingly, the present review article will primarily highlight the influence of extracellular events that contribute to cell fusion in normal and tumorigenic tissues.

Spontaneous Fusion of MSC with Breast Cancer Cells Can Generate Tumor Dormancy.

Direct cellular interactions of MDA-MB-231cherry breast cancer cells with GFP-transduced human mesenchymal stroma/stem-like cells (MSCGFP) in a co-culture model resulted in spontaneous cell fusion by the generation of MDA-MSC-hyb5cherry GFP breast cancer hybrid cells. The proliferative capacity of MDA-MSC-hyb5 cells was enhanced about 1.8-fold when compared to the parental MDA-MB-231cherry breast cancer cells. In contrast to a spontaneous MDA-MB-231cherry induced tumor development in vivo within 18.8 days, the MDA-MSC-hyb5 cells initially remained quiescent in a dormancy-like state. At distinct time points after injection, NODscid mice started to develop MDA-MSC-hyb5 cell-induced tumors up to about a half year later. Following tumor initiation, however, tumor growth and formation of metastases in various different organs occurred rapidly within about 10.5 days. Changes in gene expression levels were evaluated by RNA-microarray analysis and revealed certain increase in dormancy-associated transcripts in MDA-MSC-hyb5. Chemotherapeutic responsiveness of MDA-MSC-hyb5 cells was partially enhanced when compared to MDA-MB-231 cells. However, some resistance, e.g., for taxol was detectable in cancer hybrid cells. Moreover, drug response partially changed during the tumor development of MDA-MSC-hyb5 cells; this suggests the presence of unstable in vivo phenotypes of MDA-hyb5 cells with increased tumor heterogeneity.

Altered Tumor Plasticity after Different Cancer Cell Fusions with MSC.

While cell fusion demonstrates an important pathway during tissue development and regeneration of distinct organs, this process can also contribute to pathophysiological phenotypes during tumor progression. Hybrid cell formation after heterofusion between cancer cells and various other cell types within the tumor microenvironment is observed in vitro and in vivo. In particular, mesenchymal stroma/stem-like cells (MSC) perform diverse levels of communication with cancer cells by exhibiting anti- and pro-tumorigenic effects. During these cellular interactions, MSC can eventually fuse with cancer cells. Thereby, the newly generated disparate hybrid populations display aneuploidy associated with chromosomal instability. Based upon a subsequent post-hybrid selection process (PHSP), fused cancer cells can undergo apoptosis/necroptosis, senescence, dormancy, or a proliferative state by acquisition of new properties. Consequently, PHSP-surviving hybrid cancer cells demonstrate altered functionalities within the tumor tissue. This is accompanied by changes in therapeutic responsiveness and a different metastatic behavior. Accordingly, enhanced tumor plasticity interferes with successful therapeutic interventions and aggravates patient prognoses. The present review article focusses on fusion of MSC with different human cancer cells, in particular breast cancer populations and resulting characteristics of various cancer hybrid cells. Moreover, some mechanisms of cancer cell fusion are discussed together with multiple PHSP pathways.

Mechanisms of Cell Fusion in Cancer.

Cell-cell fusion is a normal physiological mechanism that requires a well-orchestrated regulation of intracellular and extracellular factors. Dysregulation of this process could lead to diseases such as osteoporosis, malformation of muscles, difficulties in pregnancy, and cancer. Extensive literature demonstrates that fusion occurs between cancer cells and other cell types to potentially promote cancer progression and metastasis. However, the mechanisms governing this process in cancer initiation, promotion, and progression are less well-studied. Fusogens involved in normal physiological processes such as syncytins and associated factors such as phosphatidylserine and annexins have been observed to be critical in cancer cell fusion as well. Some of the extracellular factors associated with cancer cell fusion include chronic inflammation and inflammatory cytokines, hypoxia, and viral infection. The interaction between these extracellular factors and cell's intrinsic factors potentially modulates actin dynamics to drive the fusion of cancer cells. In this review, we have discussed the different mechanisms that have been identified or postulated to drive cancer cell fusion.

A melanoma patient with macrophage-cancer cell hybrids in the primary tumor, a lymph node metastasis and a brain metastasis.

In 1911 it was proposed that cancer might result from fusion and hybridization between macrophages and cancer cells. Using immunohistochemistry it was determined that essentially all solid tumors expressed macrophage-like molecules on their cell surface. More recently we have used forensic (STR) genetics that allows one to detect DNA from more than one individual in the same sample. By studying biopsies from individuals receiving allogeneic stem cell transplants and later developed solid tumor metastases, we were able to detect both donor and patient DNA sequences suggesting that hybrids were present. Previously we found hybrids in biopsies of a renal cell carcinoma, a melanoma in a brain metastasis and a melanoma in a primary tumor with lymph node metastases. Here we have traced hybrids from a primary melanoma to an axillary lymph node to a brain metastasis. This is the first time that the entire metastatic process has been documented.

Impact of the Tumor Microenvironment on Tumor Heterogeneity and Consequences for Cancer Cell Plasticity and Stemness.

Tumor heterogeneity is considered the major cause of treatment failure in current cancer therapies. This feature of solid tumors is not only the result of clonal outgrowth of cells with genetic mutations, but also of epigenetic alterations induced by physical and chemical signals from the tumor microenvironment (TME). Besides fibroblasts, endothelial and immune cells, mesenchymal stroma/stem-like cells (MSCs) and tumor-associated macrophages (TAMs) intimately crosstalk with cancer cells and can exhibit both anti- and pro-tumorigenic effects. MSCs can alter cancer cellular phenotypes to increase cancer cell plasticity, eventually resulting in the generation of cancer stem cells (CSCs). The shift between different phenotypic states (phenotype switching) of CSCs is controlled via both genetic programs, such as epithelial-mesenchymal transdifferentiation or retrodifferentiation, and epigenetic alterations triggered by signals from the TME, like hypoxia, spatial heterogeneity or stromal cell-derived chemokines. Finally, we highlight the role of spontaneous cancer cell fusion with various types of stromal cells. i.e., MSCs in shaping CSC plasticity. A better understanding of cell plasticity and phenotype shifting in CSCs is a prerequisite for exploiting this phenomenon to reduce tumor heterogeneity, thereby improving the chance for therapy success.

The Intimate Relationship Among EMT, MET and TME: A T(ransdifferentiation) E(nhancing) M(ix) to Be Exploited for Therapeutic Purposes.

Intratumoral heterogeneity is considered the major cause of drug unresponsiveness in cancer and accumulating evidence implicates non-mutational resistance mechanisms rather than genetic mutations in its development. These non-mutational processes are largely driven by phenotypic plasticity, which is defined as the ability of a cell to reprogram and change its identity (phenotype switching). Tumor cell plasticity is characterized by the reactivation of developmental programs that are closely correlated with the acquisition of cancer stem cell properties and an enhanced potential for retrodifferentiation or transdifferentiation. A well-studied mechanism of phenotypic plasticity is the epithelial-mesenchymal transition (EMT). Current evidence suggests a complex interplay between EMT, genetic and epigenetic alterations, and clues from the tumor microenvironment in cell reprogramming. A deeper understanding of the connections between stem cell, epithelial-mesenchymal, and tumor-associated reprogramming events is crucial to develop novel therapies that mitigate cell plasticity and minimize the evolution of tumor heterogeneity, and hence drug resistance. Alternatively, vulnerabilities exposed by tumor cells when residing in a plastic or stem-like state may be exploited therapeutically, i.e., by converting them into less aggressive or even postmitotic cells. Tumor cell plasticity thus presents a new paradigm for understanding a cancer's resistance to therapy and deciphering its underlying mechanisms.

Leukocyte⁻Cancer Cell Fusion-Genesis of a Deadly Journey.

According to estimates from the International Agency for Research on Cancer, by the year 2030 there will be 22 million new cancer cases and 13 million deaths per year. The main cause of cancer mortality is not the primary tumor itself but metastasis to distant organs and tissues, yet the mechanisms of this process remain poorly understood. Leukocyte⁻cancer cell fusion and hybrid formation as an initiator of metastasis was proposed more than a century ago by the German pathologist Prof. Otto Aichel. This proposal has since been confirmed in more than 50 animal models and more recently in one patient with renal cell carcinoma and two patients with malignant melanoma. Leukocyte⁻tumor cell fusion provides a unifying explanation for metastasis. While primary tumors arise in a wide variety of tissues representing not a single disease but many different diseases, metastatic cancer may be only one disease arising from a common, nonmutational event: Fusion of primary tumor cells with leukocytes. From the findings to date, it would appear that such hybrid formation is a major pathway for metastasis. Studies on the mechanisms involved could uncover new targets for therapeutic intervention.

Potential Role of MSC/Cancer Cell Fusion and EMT for Breast Cancer Stem Cell Formation.

Solid tumors comprise of maturated cancer cells and self-renewing cancer stem-like cells (CSCs), which are associated with various other nontumorigenic cell populations in the tumor microenvironment. In addition to immune cells, endothelial cells, fibroblasts, and further cell types, mesenchymal stroma/stem-like cells (MSC) represent an important cell population recruited to tumor sites and predominantly interacting with the different cancer cells. Breast cancer models were among the first to reveal distinct properties of CSCs, however, the cellular process(es) through which these cells are generated, maintained, and expanded within neoplastic tissues remains incompletely understood. Here, we discuss several possible scenarios that are not mutually exclusive but may even act synergistically: fusion of cancer cells with MSC to yield hybrid cells and/or the induction of epithelial-mesenchymal transition (EMT) in breast cancer cells by MSC, which can relay signals for retrodifferentiation and eventually, the generation of breast CSCs (BCSCs). In either case, the consequences may be promotion of self-renewal capacity, tumor cell plasticity and heterogeneity, an increase in the cancer cells' invasive and metastatic potential, and the acquisition of resistance mechanisms towards chemo- or radiotherapy. While specific signaling mechanisms involved in each of these properties remain to be elucidated, the present review article focusses on a potential involvement of cancer cell fusion and EMT in the development of breast cancer stem cells.

Pro-tumor activities of macrophages in the progression of melanoma.

Macrophages are located in essentially all tissues due to their "janitor" function. Macrophages can exert either anti- or pro-tumor activities depending upon the specific tumor microenvironment they inhabit. Substantial evidence indicates that macrophages, owing to their plasticity, can be reeducated to adopt a protumoral phenotype within a tumor microenvironment through the help of growth factors in the microenvironment and intercellular interactions. As the lethality of malignant melanoma is due to its aggressive capacity for metastasis and resistance to therapy, considerable effort has gone toward treatment of metastatic melanoma. In the present review, we focus on the pro-tumor activities of macrophages in melanoma. Based upon the information presented in this review it is anticipated that new therapies will soon be developed that target pro-tumor activities of macrophages for use in the treatment of melanoma.