Identification of the early footprint of myeloma bone disease: spatio-temporal mapping and quantifying of extracellular matrix biomarkers in murine model and human bone biopsies

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Multiple myeloma (MM) causes massive destruction of the extracellular matrix (ECM) in bone, and is consequently one of the most serious bone diseases. Damage is induced by stimulating extensive osteolytic activity of osteoclasts while blocking bone regeneration. Osteocytes are the orchestrators of adaptive bone remodeling, stimulate osteolysis, and at the same time are potent modulators of key bone formation signaling pathways such as Wnt and Notch signaling. With their dendrite-like interconnectivity, they are at the center of the architectural design of the microstructure of mature bone. To detect damage to ECM and microstructure at the initial stage and to characterize decisive starter signals, we require new molecular and ultrastructural biomarkers for bone ECM and cutting edge methods for microstructure imaging. In preliminary work, we found decreased expression of structural proteins of the ECM in murine osteocytes within fourteen days after inoculation of MM cells into bone. By comparing expression data sets of human MM cells from clinical cohorts, we demonstrated that decreased expression of the same ECM proteins is associated with poor patient survival. While osteocytes interact directly with MM cells through the Notch signaling pathway, we have shown that Jagged1-mediated Notch activation drives MM cell proliferation and that Notch inhibition diminishes MM bone disease. It is not known which signaling pathways in osteocytes control ECM destruction in MM. We hypothesize that the Notch signaling pathway in osteocytes contributes early on to the altered remodeling of the ECM after MM cell spread to and expansion in the bone marrow. We use our murine model of MM bone disease as a platform to compare molecular changes in osteocytes and ultrastructural degradation of ECM. We will quantify in space and time, early ECM damage evolution at local sites after MM cell inoculation and throughout the skeleton as the MM cells spread. We will apply three-dimensional high-resolution material-science techniques suited for bone structural characterization. Especially three-dimensional, sub-millimeter and sub-micrometer synchrotron methods will quantify ECM structural damage in terms of voids, mineral deposits, and fiber orientation, in which we will correlate with the progression and severity of MM bone disease. Using transcriptomic profiling, we will track the components of our candidate pathway and its molecular targets over time and identify yet unknown very early molecular changes in osteocyte gene expression and pathway activity. Results in the mouse model will be validated with co-cultures of MM cells and osteocytes in-vitro and by comparison with human bone biopsies and databases in cases of monoclonal gammopathy of undetermined significance, smoldering and symptomatic MM. We are convinced that this will allow us to identify new biomarkers and innovative therapeutic targets in host and tumor.