Although the classic definition of cell plasticity taken from stem cell biology implies the ability of stem cells to differentiate into various cell lineages, the term is also currently applied to the ability of a given cell type to reciprocally dedifferentiate, redifferentiate, and/or transdifferentiate in response to specific stimulation (1
). The plasticity potential of malignant cells has been extensively studied in epithelial tumors as a mechanism that allows the epithelial cells to transdifferentiate into mesenchymal cells and vice versa in a process that is highly controlled by the microenvironment (3
In contrast to solid tumors, the plasticity of hematopoietic malignancies, such as multiple myeloma, has not been extensively studied. Progression of multiple myeloma is considered a multistage and dynamic process of cell differentiation, survival, proliferation, and dissemination (4
). Although the majority of multiple myeloma patients initially respond to therapy and achieve clinical remission, they subsequently relapse (7
). The low proliferation rate of mature multiple myeloma plasma cells and the inability to attain a cure in multiple myeloma patients led to the notion that the proliferative, self-renewal compartment subsists within the immature clonal B-cell/lymphoblastoid population. Although previous studies support this notion, the identity of the multiple myeloma ‘‘stem cell’’ remains elusive.
Myeloma cells from the majority of patients are phenotypically CD45low
and have very low proliferative activity (<1%), as determined by the standard bromodeoxyuridine labeling index (BrdUrd LI) assay. In 1991, Caligaris-Capio et al. showed the ability of stromal cells to induce differentiation of patients’ B cells into monoclonal immunoglobulin-expressing multiple myeloma plasma cells (9
), suggesting that myeloma stem cells emerge from immature clonal cells. Indeed, reports from several independent laboratories have identified the myeloma clone throughout the B-cell differentiation ladder, using complementary determining region 3 (CDR3)–based PCR analysis (10
). Pliarski et al. suggested that the myeloma clone arises from a preswitched B cell that gives rise to malignant, drug-resistant B cells with characteristics resembling stem cells, and that this clone is responsible for patient relapse (13
). Other groups suggested that the highly proliferative myeloma cells are CD45-expressing lymphoblasts (16
) rather than B cells, and upon cytokine stimulation, these cells differentiate into mature CD45−
cells capable of long-term survival (6
Opposing these concepts, studies from our laboratory using the severe combined immunodeficient (SCID)-hu and the SCID-rab models for primary myeloma (18
) and other studies with the 5T murine model for myeloma (20
) showed that purified, mature CD45−/low
and CD138-selected multiple myeloma plasma cells have proliferative potential and the ability to produce myeloma in vivo
. Furthermore, the clonal, immature CD45high
cells in those studies were incapable of engraftment (18
) or had a slower rate of engraftment (20
) compared with their mature counterparts. Overall, these studies show a fundamental confusion and controversy over the origin of the proliferative, self-renewing myeloma cells.
Based on their similar properties and behaviors, tumorigenic cells are often compared with normal stem cells, suggesting a theory that cancer stem cells exist in the form of a rare, primitive, subpopulation of cells capable of self-renewal, pluripotency, and longevity. In studies of hematologic malignancies, such as acute myeloid leukemia, only the rare CD34+
cells were capable of producing acute myeloid leukemia in SCID mice (22
). Recent work on multiple myeloma suggested that a small fraction of clonal cells expressing CD138−
are the myeloma stem cells and that CD138-expressing multiple myeloma plasma cells are not clonogenic (23
). Further evidence in support of the cancer stem cell theory came from breast cancer research. Clark et al. showed that only a small fraction of CD44+
breast cancer cells formed tumors in nonobese diabetic/SCID mice (24
Recent studies using global microarray profiling to identify common stem cell markers and signaling pathways in tumor cells further fuel the cancer stem cell theory and have provided new insights into our understanding of tumorigenesis. Several stem cell–associated signaling pathways, including Wnt
/ β-catenin, Notch, and HoxB, are activated in tumor cells (2
). Other genes involved in cell proliferation (Nucleostemin
; ref. 25
) and prevention of senescence (BMI-1
; ref. 26
) were highly expressed by both tumor cells and undifferentiated stem cells. Although these findings may help identify potential targets for cancer stem cells, they also indicate that each tumor cell expresses, to a varying degree, a range of stem cell–like genes and therefore could potentially acquire the characteristics of a stem cell.
Myeloma cells from the majority of patients typically reside in the bone marrow and alter this microenvironment to their advantage. By inhibiting osteoblastogenesis and inducing osteoclastogenesis, angiogenesis, and immunosuppression, multiple myeloma cells protect themselves from spontaneous and drug/immune-induced apoptosis, thereby ensuring their continued growth. In the majority of patients with myeloma, growth of multiple myeloma cells in the bone marrow is associated with induction of osteolytic bone disease (7
). Myeloma plasma cells contribute to bone destruction through their interactions with bone marrow stromal cells, directly (29
) and indirectly (33
) stimulating differentiation of bone-destroying osteoclasts. Multiple myeloma cells also prevent differentiation of bone-building osteoblasts through secretion of the Wnt
-signaling antagonist DKK1 (36
). This indicates that multiple myeloma cells uncouple the processes of osteoclastic bone resorption and osteoblastic bone formation, resulting in disturbed bone remodeling in multiple myeloma patients.
Clinical observations and experimental studies using osteoclast inhibitors highlighted the interdependence between myeloma bone disease and tumor progression. It seems that bisphosphonates, in addition to preserving bone, are also effective antitumor agents (37
). Studies using a murine model for myeloma and our studies with the myelomatous SCID-hu mice showed that inhibition of myeloma-associated bone disease by bisphosphonates or by inactivation of the receptor activator of nuclear factor-κB ligand (RANKL) halts progression of bone resorption and also has a profound antimyeloma effect. These findings suggest that osteoclasts facilitate survival and growth of myeloma cells in bone marrow (33
). This notion is further supported by our findings that osteoclasts alone support survival and continued proliferation of purified primary multiple myeloma cells ex vivo
). Together, these studies strongly suggest that osteoclasts play a critical role in regulation of myelomagenesis.
The aim of this study was to determine the osteoclast-induced phenotypic changes associated with survival of multiple myeloma cells in long-term coculture. We showed that purified, mature multiple myeloma plasma cells, cocultured with osteoclasts for up to 20 weeks, gradually lost their mature phenotype and dedifferentiated to an immature, resilient, apoptosis-resistant phenotype. This indicates that multiple myeloma cells have a plasticity that allows them to be reprogrammed and acquire autonomous survival properties upon long-term, direct contact with the osteoclasts they promote.