Impact of the non-cellular tumor microenvironment on metastasis: potential therapeutic and imaging opportunities

Evidence is accumulating that the malignant phenotype of a given tumor is dependent not only on the intrinsic characteristics of tumor cells, but also on the cooperative interactions of non-neoplastic cells, soluble secreted factors and the non-cellular solid-state ECM network that comprise the tumor microenvironment. Given the ability of the tumor microenvironment to regulate the cellular phenotype, recent efforts have focused on understanding the molecular mechanisms by which cells sense, assimilate, interpret, and ultimately respond to their immediate surroundings. Exciting new studies are beginning to unravel the complex interactions between the numerous cell types and regulatory factors within the tumor microenvironment that function cooperatively to control tumor cell invasion and metastasis. Here, we will focus on studies concerning a common theme, which is the central importance of the non-cellular solid-state compartment as a master regulator of the malignant phenotype. We will highlight the non-cellular solid-state compartment as a relatively untapped source of therapeutic and imaging targets and how cellular interactions with these targets may regulate tumor metastasis.     

Conjunction of tumor cells with lymphocytes: implications for tumor invasion and metastasis

Our previous studies have led to a novel hypothesis that tumor metastasis is triggered by aberrant lymphocyte infiltration that disrupts intercellular junctions and surface adhesion molecules and causes dissociation of tumor cells from the primary tumor core, allowing lymphocytes to conjoin with dissociated tumor cells and physically 'drag' them to different tissue sites. Our hypothesis is supported by morphological and immunohistochemical data from multiple types of human cancer. This hypothesis challenges the traditional belief that the physical conjunction between tumor cells and lymphocytes would lead to degeneration of the tumor cells. To validate our hypothesis, H&E and immunostained sections were examined under high magnification to identify potential signs of degeneration-related changes. Our study revealed that >60% of isolated tumor cells overlying focal capsule disruptions, or within the stroma and vascular structures, were physically conjoined with lymphocytes to form tumor cell-lymphocyte chimeras (TLCs). Approximately 90% of the tumor cell partners of TLCs were morphologically indistinguishable from their counterparts within the tumor core. In addition, one third of the tumor cells of TLCs expressed high levels of cell proliferation specific proteins, or were undergoing mitosis. Our study also revealed that a subset of dilated lymphatic ducts or blood vessels at the site of focal capsule disruptions harbored variable numbers of tumor cells, and the wall of these structures was in direct physical continuity with the myoepithelial cell layer. Our study suggests that the onset of tumor metastasis may occur in two forms: (1) lymphocyte-mediated shuttling that allows lymphocytes to physically 'drag' tumor cells to different sites, and (2) tumor progenitor-mediated angiogenesis that allows tumor cells to directly enter the vascular structures.     

microRNAs调节肿瘤细胞转移表型的分子机制

microRNAs(miRNAs)是一类在转录后水平调控基因表达的内源性非编码小RNA分子.miRNAs具有癌基因与抑癌基因的功能,参与肿瘤细胞的增殖、粘附、侵袭、转移和肿瘤血管形成等过程.miRNAs可调节肿瘤细胞转移表型,主要通过改变肿瘤细胞黏附力、侵袭力与迁移力.本文重点介绍调节肿瘤细胞转移表型相关miRNAs及其作用的分子机制,以便为肿瘤转移的研究提供新思路.     

Immunopotentiation of NKT cells by low-protein diet and the suppressive effect on tumor metastasis

Mice were fed with a 5% low-protein diet for two weeks, at which point tumor inoculation was conducted. Following this inoculation, the 5% low-protein diet was continued. On the other hand, control mice were fed with a normal diet (25% protein) and such diet was continued after tumor inoculation. In comparison with control mice, mice fed with the 5% low-protein diet showed a prominent prolongation of survival rate when injected with both EL4 and 3LL tumors. Interestingly, CD1d(-/-) mice, which primarily lack natural killer T (NKT) cells, did not show the prolongation of survival rate even when they received a 5% low-protein diet. The most striking phenomenon seen in tumor-bearing mice fed with the 5% low-protein diet was the suppression of tumor metastasis to the liver and lung. Such suppression was not seen in CD1d(-/-) mice who were fed with a 5% low-protein diet. Phenotypic study revealed that the proportion of NKT cells after tumor inoculation decreased in the mice fed with a normal diet. However, such decrease did not occur in mice fed with the 5% low-protein diet. Reflecting the activation of NKT cells by feeding, tumor cytotoxicity and cytokine production were also augmented by the 5% low-protein diet. These results suggest that a low-protein diet has the potential to augment the innate immunity against tumors, especially mediated by the activation of NKT cells.     

TGFbeta primes breast tumor cells for metastasis

The microenvironment of a tumor is known to influence tumor progression and spread to distant sites (metastasis). Padua et al. (2008) now show that transient exposure of breast cancer cells to the signaling molecule transforming growth factor beta (TGFbeta) promotes their extravasation from blood vessels and entry into the lung by upregulation of the adipokine angiopoietin-like 4 (ANGPTL4). Their work shows that the later stages of metastasis can be influenced by transient signals produced in the primary tumor microenvironment.     

Flavonoids inhibit melanoma lung metastasis by impairing tumor cells endothelium interactions

Flavonoids comprise a class of low molecular weight compounds displaying a variety of biological activities including inhibition of tumor growth and metastasis. To gain insight into the mechanisms underlying metastasis inhibition, we have employed the B16-BL6 murine melanoma metastasis model. B57BL/6N mice were injected i.v. with tumor cells and Apigenin, Quercetin, or Tamoxifen, each at 50 mg/kg given i.p., and lung tumor cell colonies counted 14-6 days thereafter. Three different injection schedules were used for each drug: (a) daily injection, starting 24 h before injection of the tumor cells; (b) single dose, 24 h preceding tumor challenge; (c) daily injection, starting 24 h after the injection of the tumor cells. All three compounds significantly reduced tumor lung deposits (Apigenin = Quercetin > Tamoxifen). However, when treatment was delayed by 24 h after tumor cells (schedule c), multiple daily doses of Apigenin or Quercetin were less effective that a single dose of the same compound given 24 h before tumor challenge (schedule b). Apigenin and Quercetin, but not Tamoxifen, were found to inhibit VCAM-1 expression in a dose-dependent manner in HUVEC and in murine pulmonary endothelial cells. In ex vivo experiments, the number of tumor cells adhering to lung vessels was significantly diminished in animals treated with a single dose of Apigenin and Quercetin. These findings indicate that the inhibition of tumor cell metastasis by Apigenin or Quercetin may significantly depend on the ability of these compounds to alter the host's microenvironment, further substantiating the role of the intravascular processes in the metastatic cascade.     

Cutting edge: inhibition of experimental tumor metastasis by dendritic cells pulsed with alpha-galactosylceramide.

A unique lymphoid lineage, Valpha14 NKT cells, bearing an invariant Ag receptor encoded by Valpha14 and Jalpha281 gene segments, play crucial roles in various immune responses, including protective immunity against malignant tumors. A specific ligand of Valpha14 NKT cells is determined to be alpha-galactosylceramide (alpha-GalCer) which is presented by the CD1d molecule. Here, we report that dendritic cells (DCs) pulsed with alpha-GalCer effectively induce potent antitumor cytotoxic activity by specific activation of Valpha14 NKT cells, resulting in the inhibition of tumor metastasis in vivo. Moreover, a complete inhibition of B16 melanoma metastasis in the liver was observed when alpha-GalCer-pulsed DCs were injected even 7 days after transfer of tumor cells to syngeneic mice where small but multiple metastatic nodules were already formed. The potential utility of DCs pulsed with alpha-GalCer for tumor immunotherapy is discussed.     

Visualization of body thermoregulation by infrared imaging

This paper analyzes two mechanisms applied on the human body in order to study the thermoregulatory system according to heat generation and heat loss. Two approaches are presented. The first approach is based on plethysmography, where an armband is placed on the forearm in order to modulate the blood flow. The second approach uses a cold stimulation. The visualization is achieved using infrared imaging devices. The resulting images reveal a temperature balance between the stimulated and the non-stimulated hands. The thermal behavior and typical thermographic recordings on each subject are discussed and analyzed in response to different stimulations.     

Identification of glycan structure alterations on cell membrane proteins in desoxyepothilone B resistant leukemia cells

Resistance to tubulin-binding agents used in cancer is often multifactorial and can include changes in drug accumulation and modified expression of tubulin isotypes. Glycans on cell membrane proteins play important roles in many cellular processes such as recognition and apoptosis, and this study investigated whether changes to the glycan structures on cell membrane proteins occur when cells become resistant to drugs. Specifically, we investigated the alteration of glycan structures on the cell membrane proteins of human T-cell acute lymphoblastic leukemia (CEM) cells that were selected for resistance to desoxyepothilone B (CEM/dEpoB). The glycan profile of the cell membrane glycoproteins was obtained by sequential release of N- and O-glycans from cell membrane fraction dotted onto polyvinylidene difluoride membrane with PNGase F and β-elimination respectively. The released glycan alditols were analyzed by liquid chromatography (graphitized carbon)-electrospray ionization tandem MS. The major N-glycan on CEM cell was the core fucosylated α2-6 monosialo-biantennary structure. Resistant CEM/dEpoB cells had a significant decrease of α2-6 linked sialic acid on N-glycans. The lower α2-6 sialylation was caused by a decrease in activity of β-galactoside α2-6 sialyltransferase (ST6Gal), and decreased expression of the mRNA. It is clear that the membrane glycosylation of leukemia cells changes during acquired resistance to dEpoB drugs and that this change occurs globally on all cell membrane glycoproteins. This is the first identification of a specific glycan modification on the surface of drug resistant cells and the mechanism of this downstream effect on microtubule targeting drugs may offer a route to new interventions to overcome drug resistance.     

Involvement of tumor necrosis factor-related apoptosis-inducing ligand in surveillance of tumor metastasis by liver natural killer cells

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis in various tumor cells in vitro, but its physiological role in tumor surveillance remains unknown. Here, we report that TRAIL is constitutively expressed on murine natural killer (NK) cells in the liver and plays a substantial role in suppressing tumor metastasis. Freshly isolated NK cells, but not natural killer T cells or ordinary T cells, from the liver expressed cell surface TRAIL, which was responsible for spontaneous cytotoxicity against TRAIL-sensitive tumor cells in vitro along with perforin and Fas ligand (FasL). Administration of neutralizing monoclonal antibody against TRAIL significantly increased experimental liver metastases of several TRAIL-sensitive tumor cell lines. Such an anti-metastatic effect of TRAIL was not observed in NK cell-depleted mice or interferon-gamma-deficient mice, the latter of which lacked TRAIL on liver NK cells. These findings provide the first evidence for the physiological function of TRAIL as a tumor suppressor.     

Intravascular origin of metastasis from the proliferation of endothelium-attached tumor cells: a new model for metastasis

Metastasis is a frequent complication of cancer, yet the process through which circulating tumor cells form distant colonies is poorly understood. We have been able to observe the steps in early hematogenous metastasis by epifluorescence microscopy of tumor cells expressing green fluorescent protein in subpleural microvessels in intact, perfused mouse and rat lungs. Metastatic tumor cells attached to the endothelia of pulmonary pre-capillary arterioles and capillaries. Extravasation of tumor cells was rare, and it seemed that the transmigrated cells were cleared quickly by the lung, leaving only the endothelium-attached cells as the seeds of secondary tumors. Early colonies were entirely within the blood vessels. Although most models of metastasis include an extravasation step early in the process, here we show that in the lung, metastasis is initiated by attachment of tumor cells to the vascular endothelium and that hematogenous metastasis originates from the proliferation of attached intravascular tumor cells rather than from extravasated ones. Intravascular metastasis formation would make early colonies especially vulnerable to intravascular drugs, and this possibility has potential for the prevention of tumor cell attachment to the endothelium.     

Cancer stem cells and tumor metastasis: first steps into uncharted territory

In several forms of human cancer, only a phenotypic subset of cancer cells, usually termed "cancer stem cells" (CSC), can initiate tumor growth when transplanted. In this issue of Cell Stem Cell, Hermann et al. (2007) analyze the relationship between CSC and tumor metastasis.     

Disseminated tumor cells and dormancy in prostate cancer metastasis

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Optimized mouse model for the imaging of tumor metastasis upon experimental therapy

Development of new cancer treatments focuses increasingly on the relation of cancer tissue with its microenvironment. A major obstacle for the development of new anti-cancer therapies has been the lack of relevant animal models that would reproduce all the events involved in disease progression from the early-stage primary tumor until the development of mature metastatic tissue. To this end, we have developed a readily imageable mouse model of colorectal cancer featuring highly reproducible formation of spontaneous liver metastases derived from intrasplenic primary tumors. We optimized several experimental variables, and found that the correct choice of cell line and the genetic background, as well as the age of the recipient mice, were critical for establishing a useful model system. Among a panel of colorectal cancer cell lines tested, the epithelial carcinoma HT29 line was found to be the most suitable in terms of producing homogeneous tumor growth and metastases. In our hands, SCID mice at the age of 125 days or older were the most suitable in supporting consistent HT29 tumor growth after splenic implantation followed by reproducible metastasis to the liver. A magnetic resonance imaging (MRI) protocol was optimized for use with this mouse model, and demonstrated to be a powerful method for analyzing the antitumor effects of an experimental therapy. Specifically, we used this system to with success to verify by MRI monitoring the efficacy of an intrasplenically administered oncolytic adenovirus therapy in reducing visceral tumor load and development of liver metastases. In summary, we have developed a highly optimized mouse model for liver metastasis of colorectal cancer, which allows detection of the tumor load at the whole body level and enables an accurate timing of therapeutic interventions to target different stages of cancer progression and metastatic development.     

肿瘤细胞和骨微环境在骨转移中的作用

随着肿瘤治疗水平的提高,肿瘤患者的生存期显著延长,转移性骨肿瘤的发生率呈增长趋势.骨转移引起的剧烈的临床症状和其较长的潜伏期,以及缺乏有效的治疗方法,极大降低了患者的生活质量.本文主要综述了骨转移相关的细胞特征及骨微环境在骨转移中的作用,并分析了影响骨转移形成的相关分子因素,为骨转移的定向分子治疗提供进一步的理论依据.     

Functional glycosylation of human podoplanin: glycan structure of platelet aggregation-inducing factor

Podoplanin (Aggrus) is a mucin-type sialoglycoprotein that plays a key role in tumor cell-induced platelet aggregation. Podoplanin possesses a platelet aggregation-stimulating (PLAG) domain, and Thr52 in the PLAG domain of human podoplanin is important for its activity. Endogenous or recombinant human podoplanin were purified, and total glycosylation profiles were surveyed by lectin microarray. Analyses of glycopeptides produced by Edman degradation and mass spectrometry revealed that the disialyl-corel (NeuAc alpha2-3Gal beta l-3(NeuAc alpha2-6)GalNAc alpha l-O-Thr) structure was primarily attached to a glycosylation site at residue Thr52. Sialic acid-deficient podoplanin recovered its activity after additional sialylation. These results indicated that the sialylated Corel at Thr52 is critical for podoplanin-induced platelet aggregation.     

Epithelial-mesenchymal transition: a hallmark in metastasis formation linking circulating tumor cells and cancer stem cells

The occurrence of either regional or distant metastases is an indicator of poor prognosis for cancer patients. The mechanism of their formation has not yet been fully uncovered, which limits the possibility of developing new therapeutic strategies. Nevertheless, the discovery of circulating tumor cells (CTCs), which are responsible for tumor dissemination, and cancer stem cells (CSCs), required for tumor growth maintenance, shed light on the metastatic cascade. It seems that CTCs and CSCs are not necessarily separate populations of cancer cells, as CTCs generated in the process of epithelial-mesenchymal transition (EMT) can bear features characteristic of CSCs. This article describes the mechanisms of CTC and CSC formation and characterizes their molecular hallmarks. Moreover, we present different types of EMT occurring in physiological and pathological conditions, and we demonstrate its crucial role in providing CTCs with a CSC phenotype. The article delineates molecular changes acquired by cancer cells undergoing EMT that facilitate metastasis formation. Deeper understanding of those processes is of fundamental importance for the development of new strategies of early cancer detection and effective cancer treatment approaches that will be translated into clinical practice.