Disseminated Tumor Cells Persist in the Bone Marrow of Breast Cancer Patients through Sustained Activation of the Unfolded Protein Response

Abstract

Disseminated tumor cells (DTC), which share mesenchymal and epithelial properties, are considered to be metastasis-initiating cells in breast cancer. However, the mechanisms supporting DTC survival are poorly understood. DTC extravasation into the bone marrow may be encouraged by low oxygen concentrations that trigger metabolic and molecular alterations contributing to DTC survival. Here, we investigated how the unfolded protein response (UPR), an important cytoprotective program induced by hypoxia, affects the behavior of stressed cancer cells. DTC cell lines established from the bone marrow of patients with breast cancer (BC-M1), lung cancer, (LC-M1), and prostate cancer (PC-E1) were subjected to hypoxic and hypoglycemic conditions. BC-M1 and LC-M1 exhibiting mesenchymal and epithelial properties adapted readily to hypoxia and glucose starvation. Upregulation of UPR proteins, such as the glucose-regulated protein Grp78, induced the formation of filamentous networks, resulting in proliferative advantages and sustained survival under total glucose deprivation. High Grp78 expression correlated with mesenchymal attributes of breast and lung cancer cells and with poor differentiation in clinical samples of primary breast and lung carcinomas. In DTCs isolated from bone marrow specimens from breast cancer patients, Grp78-positive stress granules were observed, consistent with the likelihood these cells were exposed to acute cell stress. Overall, our findings provide the first evidence that the UPR is activated in DTC in the bone marrow from cancer patients, warranting further study of this cell stress pathway as a predictive biomarker for recurrent metastatic disease. Cancer Res; 75(24); 5367-77. Ó2015 AACR.

Tumorigenic and Immunosuppressive Effects of Endoplasmic Reticulum Stress in Cancer

“…Dormant cells are often quiescent and exhibit reduced metabolic rates, which can insulate them from many anticancer drugs that rely on active proliferation. Disseminated tumor cells in the bone marrow of breast cancer patients exhibit high expression of multiple ER chaperones, including BiP, which insulates these cells from hypoxia and glucose deprivation (Bartkowiak et al, 2010; Bartkowiak et al, 2015). Furthermore, comparative proteomic analyses of highly proliferative T-HEp3 human squamous carcinoma cell line and the D-HEp3 subclone, which becomes dormant in vivo despite growing in vitro , revealed a p38-dependent program sustaining high BiP expression and constitutive PERK phosphorylation.…”

Adaptive Stress Responses During Tumor Metastasis and Dormancy

“…Several UPR-regulated genes (e.g., Grp78, Grp94, PDI, HSP47, and cyclophilin B) are upregulated in dormant cells from experimental models, patient-derived dormant DTCs, and DTC-derived cell lines [84–87], supporting a role for the UPR in metastatic dormancy. Mechanistic analysis revealed that increased PERK–eIF2α phosphorylation, XBP-1 splicing [85], and ATF6 activation [88] contribute to the survival and increased stress tolerance of dormant DTCs.…”

“…PERK signaling was also suggested to contribute to the quiescence phenotype, and inhibition of PERK signaling reverses the dormant phenotype and renders cells more susceptible to therapy [85]. Activation of the UPR in the BM possibly occurs in response to varying oxygen and nutrient availability, and is thought to confer a survival advantage to DTC within this niche [87]. Indeed, in vitro analysis of cell lines derived from BM DTCs revealed a heightened resistance to glucose and oxygen deprivation, which was reversed by Grp78 knockdown [87].…”

“…Activation of the UPR in the BM possibly occurs in response to varying oxygen and nutrient availability, and is thought to confer a survival advantage to DTC within this niche [87]. Indeed, in vitro analysis of cell lines derived from BM DTCs revealed a heightened resistance to glucose and oxygen deprivation, which was reversed by Grp78 knockdown [87]. This suggests that DTCs in the hypoxic BM environment may be especially likely to upregulate the cellular stress response.…”

Functional studies on circulating and disseminated tumor cells in carcinoma patients

“…The UPR grants cytoprotection under hostile microenvironmental conditions like hypoxia, suggesting that such a DTC phenotype is well protected from microenvironmental stress. Detailed analyses of the DTC cell lines BC‐M1, LC‐M1 and PC‐E1 revealed a dynamic network of interconnected cellular programs like induction of the UPR chaperones and oxidoreductases in dependence of EGFR/ErbB2 expression under hypoxia (Bartkowiak et al., 2015). These findings support the view that DTCs in cancer patients are able to cope with a variety of different cell stress factors like the hypoxic conditions of the hematopoietic stem cell niche.…”