Mechanisms governing metastatic dormancy and reactivation

Abstract

Many cancer patients suffer from metastatic relapse several years after they have undergone radical surgery. Early cancer cell dissemination followed by a protracted period of dormancy potentially explains this prevalent clinical behavior. Increasing evidence suggests that the metastasis-initiating cells are cancer stem cells or revert to this functional state upon infiltrating a target organ. Their entry into dormancy and subsequent reactivation are governed by intrinsic programs and by contextual cues, which resemble those regulating the self-renewal capability of adult stem cells. In addition, metastatic cells undergoing reactivation are nursed by specialized extracellular matrix niches, which support positive signals, such as Wnt and Notch, and attenuate negative signals, such as BMP. In spite of significant remaining uncertainties, these findings provide a framework to understand the logic of metastatic dormancy and reactivation and open new avenues for therapeutic intervention.

Figure 1. The invasion-metastasis cascade

Genetic and epigenetic alterations endow cancer cells with the capabilities that enable them to negotiate the sequential steps comprising the invasion-metastasis cascade. A partial or complete EMT allows individual carcinoma cells or small groups of carcinoma cells to disassociate from adjacent epithelial cells and to invade into the underlying interstitial matrix (invasion). In a tightly linked process, cancer cells coopt a wide spectrum of host cells to create a permissive microenvironment. Upon recruiting angiogenic endothelial cells and inducing the development of a defective vasculature (angiogenesis), cancer cells enter into the circulation (intravasation) and disseminate via the bloodstream (blood-born dissemination). In a variation from the predominant sequence depicted here, cancer cells enter into lymphatic vessels and colonize loco-regional lymphnodes prior to entering into the blood stream. Upon arresting in the microcirculation, cancer cells disrupt the endothelial junctions and penetrate into the stroma of the target organ (extravasation). In the final step, colonization, they resist apoptosis (initial survival), undergo a variable period of dormancy (dormancy) and finally outgrow into macroscopic lesions (outgrowth). In order to colonize a target organ, cancer cells need to mold a permissive microenvironment. In certain cases, systemic signals retard the vascularization of micrometastases (systemic inhibition), potentially explaining why surgical resection of the primary tumor may induce rapid outgrowth of metastatic lesions (Demicheli et al., 2007). Other systemic signals are proposed to spur metastatic outgrowth via the recruitment of bone marrow-derived hematopoietic cells (systemic instigation).

Figure 2. Relationship between early dissemination, metastatic dormancy and reactivation

(A) Although carcinomas in situ can release potentially metastatic cells in the bloodstream, the number and metastatic capacity of cancer cells deposited at pre-metastatic sites presumably increases as primary tumors progress toward increasing malignancy. Experiments performed by inoculating B16F1 melanoma cells directly in the circulation of mice suggest that the efficiency of extravasation is approximately 20% and that of initial survival approximately 4% (Luzzi et al., 1998). (B) A large fraction of cancer cells, which have remained viable in the target organ, enters into solitary dormancy before surgical resection of the primary tumor interrupts dissemination. After a variable lag time, a small minority of dormant cells undergoes reactivation and gives rise to metastatic outgrowths. Experiments performed by inoculating B16F1 melanoma cells directly in the circulation of mice suggest that a large fraction of micrometastases regress because they fail to establish a permissive microenvironment, further contributing to the inefficiency of colonization (Luzzi et al., 1998). However, a small fraction of micrometastases spawns macroscopic lesions. It is debated whether micrometastatic dormancy occurs and, if so, whether it interrupts secondary tumor growth for a significant period of time. The cancer cells that comprise a macrometastasis have solved the adaptation problem and can therefore seed additional macrometastases in the same organ. Percentages of attrition are derived from the analysis of a single xenograft model and are therefore intended for illustrative purpose.

Figure 3. Survival and stress signaling in metastatic dormancy and reactivation

Adhesive and signaling interactions leading to activation of AKT support the survival of cancer cells during dormancy and reactivation (top). Stress signals initiated by p38 kinase and leading to the activation of the Unfolded Protein Response (UPR) and of TOR contribute to dormancy, whereas activation of ERK may contribute to reactivation (bottom). mΦ: macrophage.

Figure 4. Stem cell signaling pathways and transcriptional circuits implicated in metastatic reactivation

The stem cell signaling pathways and transcriptional circuits implicated in metastatic colonization or, specifically, reactivation are illustrated diagrammatically. The interactions between signaling components, transcription factors, and functional outputs are largely inferred from studies on embryonic and adult stem cells (Clevers and Nusse, 2012; Guruharsha et al., 2012; Wakefield and Hill, 2013; Young, 2011). Signaling pathways, such as Wnt/p-catenin and Notch, promote cell cycle progression via Myc and Cyclin D1. Myc also induces expression of the Polycomb Repressor Complex 1 component Bmi-1. Together with JAK/STAT3, these pathways induce expression of SOX2, OCT4, and NANOG, which constitute the core transcriptional circuit regulating self-renewal. BMP signaling opposes the upregulation of these core factors. Additional transcription factors determine progenitor identity and/or induce an EMT. PRRX1 also controls expression of SOX9 (Reichert et al., 2013). Broken lines denote indirect signaling interactions. Solid lines illustrate direct transcriptional interactions. Rectangles: transcription factors; ovals: cell cycle components; octagon: epigenetic regulator. Functional groups are color-coded.

Figure 5. Metastatic niches

Coco is retained at the cell surface presumably because it binds to cell surface proteoglycans. It thereby effectively shields outgrowing cancer cells from the inhibitory action of BMP proteins produced by host cells. Fibronectin fibrils that are decorated by tenascin-C and periostin nurse outgrowing micrometastases by promoting activation of the Notch and β-catenin/TCF signaling pathways. Tenascin-C can engage integrins as well as Syndecan 4. The latter can function as a co-receptor for Frizzled. Periostin facilitates presentation of Wnt to Frizzled and also binds to integrins. Tenascin-C promotes activation of Notch and β-catenin/TCF signaling via Musashi-1 and Lgr5, respectively (Oskarsson et al., 2011)(not shown).

Figure 6. Relationship between EMT, MET and metastatic reactivation

The existence of two types of metastatic cancer stem cells potentially explains the relationship between MET and reactivation. Mesenchymal stem cells are cycling slowly or are dormant, whereas E-cadherin-expressing cancer stem cells are proliferating vigorously, mirroring the behavior of normal adult stem cells and transit-amplifying progenitors, respectively. Experiments in mouse models suggest that, when metastasis is initiated by mesenchymal cancer stem cells, an MET may be required for reactivation. If the mesenchymal stem cell is cycling slowly, it can give rise to its immediate progeny, which expresses E-cadherin and proliferates rapidly, spurring metastatic outgrowth (1). If the mesenchymal stem cell is dormant, it may have to undergo a MET in order to become competent for reactivation (2).

Figure 7. Anti-metastatic therapies

Since dormant cancer cells are not cycling, they may be relatively resistant to anti-mitotic therapies. The studies discussed in this Review suggest that dormant cancer cells may undergo apoptosis in response to Src, PI3K/TOR or AKT inhibitors. In addition, Notch and Wnt inhibitors or BMP-R agonists may prevent the reactivation of these cells. In contrast, adjuvant chemotherapies and targeted therapies inhibit the survival and proliferation of reactivated cancer cells, interfering with the outgrowth of micrometastases.