PATHOPHYSIOLOGY OF MYELOMA BONE DISEASE: THE KEY ROLE OF RANKL IN THE MYELOMA BONE MARROW MICROENVIRONMENT

Under normal physiologic conditions, osteoblasts and osteoclasts work in unison to remodel bone by bone formation and bone resorption, respectively.26,27 Immature osteoblasts secrete cytokines such as IL-6 to upregulate osteoclasts and mature osteoblasts secrete osteoprotegerin (OPG) to inhibit the activation of osteoclasts. As new bone is formed, osteoblasts become trapped and differentiate into osteocytes which contribute factors to both osteoclastogenesis and osteoblastogenesis.8 MPC cause the dysregulation and uncoupling of this bone remodeling process by interacting with the BMME to induce osteoclast-activating factors to promote osteoclastogenesis while simultaneously secreting osteoblast inhibitory factors to inhibit osteoblastogenesis (Figure 1).12 In the initial stages of the disease, both osteoblasts and osteoclasts are recruited to initiate bone resorption. MPC produce IL-1 and tumor necrosis factor (TNF) which stimulate osteoblast progenitor cells to differentiate into osteoblasts. Osteoblasts in turn secrete IL-6 which acts as MPC growth factor and promoter of osteoclastogenesis.11,28 Once myeloma bone disease (MBD) is established, osteoblasts decrease in number. MPC and bone marrow stromal cells secrete Dkk-1 while osteocytes secrete sclerosin, both of which inhibit the canonical Wnt pathway and result in a decrease in osteoblastogenesis.29,30 Dkk-1 additionally inhibits mesenchymal stromal cells from differentiating into osteoblasts which enables the maximum amount of IL-6 to be secreted thus promoting MPC growth.31 sFRP-2, a Wnt antagonist secreted by MPC, further inhibits osteoblastogenesis.27,32

Figure 1

The balance between osteoblasts and osteoclasts is maintained by the ratio of OPG:RANKL.33 The interaction between RANK and RANKL activate downstream nuclear factor kappa B (NF-kB) which in turn activates osteoclast precursors and causes their differentiation to mature osteoclasts and decreases osteoclast apoptosis.9 OPG is a soluble decoy receptor for RANKL that inhibits the RANK-RANKL interaction via molecular mimicry in order to increase osteoblast activity and promote bone formation.27,34 MPC interact with the BMME and activate molecular cascades that ultimately result in increased RANKL and decreased OPG expression.35,36 MPC secrete soluble RANKL as well as PTHrP, IL-1, IL-6, IL-11 and other cytokines which in turn stimulate RANKL expression by osteoblasts and bone marrow stromal cells.8,37,38 In addition, MPC express syndecan-1 which binds to OPG resulting in subsequent endocytosis and degradation of OPG by MPC.39


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Greater serum RANKL/OPG ratios are associated with shorter survival. At 60 months, the survival probability for patients with soluble RANKL/OPG <1 was 89% and for patients with a ratio of 1–3 was 32%. The level of soluble RANKL also correlated with the extent of bone disease as examined by radiographic imaging.40 MPC are able to tip the balance of RANKL/OPG in favor of greater levels of RANKL with subsequent suppression of osteoblastogenesis, hyperactivation of osteoclasts and the propagation of osteolytic lesions throughout the entire bone marrow.8

ANTI-MYELOMA THERAPIES AND THEIR EFFECT ON THE RANKL PATHWAY AND BONE REMODELING

Given the key role of the RANKL pathway and osteoclastogenesis in MPC survival, anti-myeloma therapies that simultaneously target MPC and osteoclast differentiation have the potential to cause deep clinical responses as well as prevent SRE.

Proteasome inhibitors (PIs) such as bortezomib, carfilzomib and ixazomib have been reported to affect bone remodeling via their ability to modulate the RANK/RANKL pathway. One of the main cytotoxic effects of proteasome inhibitors is attributed to inhibition of NF-kB activity.41 Given that binding of RANKL to RANK on the surface of osteoclast precursors activates NF-kB which promotes osteoclast maturation and bone resorption, proteasome-dependent inhibition of NF-kB by PIs lead to a reduction in RANKL-mediated osteoclast differentiation.42,43 In patients with MM, bortezomib was associated with an increase in the levels of biomarkers associated with bone formation and decreased serum levels of RANKL and markers of bone resorption.44 Carfilzomib has been shown to directly inhibit osteoclast formation and bone resorption in vitro, while enhancing osteogenic differentiation and matrix mineralization. Carfilzomib increased trabecular bone volume, decreased bone resorption and enhanced bone formation in mouse models of MM.45 Ixazomib has demonstrated the ability to inhibit in vitro osteoclastogenesis and resorption and these effects on osteoclasts were partially mediated by inhibition of RANKL-induced NF-κB signaling. Ixazomib also stimulates osteogenic differentiation of mesenchymal cells in vitro and promotes osteoblast function and matrix mineralization.46

Immunomodulatory drugs such as thalidomide, lenalidomide and pomalidomide possess anti-myeloma properties including immune-modulation, anti-angiogenic, anti-inflammatory and anti-proliferative effects. Lenalidomide has been shown to inhibit osteoclast formation and activation through inhibition of key factors during osteoclastogenesis in vitro. The combination of lenalidomide and bortezomib blocked osteoclast-derived secretion of growth and survival factors and RANKL secretion from bone marrow stromal cells. Furthermore, lenalidomide treatment decreased serum bone-remodeling markers in patients with relapsed and refractory MM.47 In patients with relapsed and refractory MM, intermediate doses of thalidomide (200mg/day) with dexamethasone led to significant reduction of the soluble RANKL/OPG ratio and markers of bone remodeling.48 Pomalidomide has been shown to inhibit osteoclastogenesis by downregulating transcription factor PU.1 and by significantly blunting RANKL upregulation normalizing the RANKL/OPG ratio in human osteoprogenitor cells when co-cultured with MM cells.49,50

Monoclonal antibodies against CD 38 are the newest group of drugs that have revolutionized anti-MM therapy. Daratumumab, an anti-CD38 monoclonal antibody, has shown in-vitro inhibition of osteoclastogenesis and bone resorption activity in bone marrow cells of MM patients by blocking the interaction of CD 38 expressing monocytes and early osteoclast progenitors.51 Furthermore, the inhibition of T-cell proliferation caused by osteoclasts is partially overcome by another anti-CD38 monoclonal antibody, isatuximab, via inhibition of multiple immune checkpoint molecules expressed on osteoclasts which in turn decrease the immune-evasive properties of MPC.52

In a study of 51 MM patients, patients who received high dose chemotherapy followed by autologous stem cell transplant (ASCT) had a significant reduction of sRANKL/OPG ratio, with a concomitant decrease in markers of bone resorption starting the second month post-ASCT. Bone formation markers started to increase after the 9th month post-ASCT while the increase of OPG preceded this. Thus, it is postulated that high dose chemotherapy followed by ASCT normalizes the abnormal bone resorption in MM patients through the decrease of the RANKL/OPG ratio.53

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Histone deacetylase inhibitors (HDACs) inhibit HDAC enzymes, curtailing the aberrant HDAC enzyme activity in MPC.54 Vorinostat has been shown to inhibit RANKL-induced osteoclast formation by suppressing the induction of the osteoclastogenic transcription factor c-Fos.55 Panobinostat has also been shown to inhibit RANKL-mediated osteoclast formation in vitro and in a mouse model of MM.56

By targeting the RANKL pathway, the most active myeloma therapies not only cause apoptosis of the MPC but also inhibit osteoclastogenesis and other key signaling events that underlie SRE.

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