Targeting the unfolded protein response in multiple myeloma with monoclonal antibodies to overcome drug resistance

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Multiple myeloma (MM) is a rare hematological cancer originating from transformed plasma cells in the bone marrow. Excessive synthesis and secretion of monoclonal immunoglobulins (Ig) or incomplete molecules, the Ig light chains, is one hallmark of MM. This secretory activity leads to a significant increased level of stress in the endoplasmatic reticulum (ER) and results frequently in the accumulation of incorrect or missfolded proteins. To circumvent unnecessary or even harmful protein biosynthesis this accumulation can trigger a highly conserved mechanism called the Unfolded Protein Response (UPR), which finally results in the apoptosis of the cell. A novel class of therapeutics uses this mechanism by inhibiting the cellular degradation system- the proteasome- which causes severe ER stress. Proteasome inhibitors such as bortezomib represent the strongest plasma cell depleting agents of today and are used very successfully in the treatment of MM. Accumulation of missfolded Ig on the other hand increases the number of chaperone molecules which are involved in protein assembly as well as in the regulation of the UPR. A key sensor of the UPR is chaperone molecule Glucose regulated protein (GRP) 78 which is mainly located in the ER but was also found on the surface of tumor cells or as secreted variant. The latter variant seems to contribute to PI resistance in vitro. However, the connection of GRP78 and the ubiquitin-proteasome pathway is barely understood. We have developed a therapeutic anti-GRP78 antibody and tested the product in clinical trials. Single-agent activity was moderate, but in combination with PI objective responses were seen even in a patient with PI refractory disease. In this proposal I aim to go back from bedside to the bench to elucidate GRP78 induced PI resistance in vitro and to investigate whether treatment with anti-GRP78 antibodies can overcome PI resistance. I will explore the frequency and functional role of secreted GRP78 in myeloma patients at different stages of their disease using ELISA and Western blot techniques. In addition, I will compare GRP78 serum levels from PI sensitive to PI resistant MM patients. Furthermore, intracellular overexpression as well as knock down of GRP78 will be performed to study its impact on susceptibility to PI. Recombinant expressed GRP78 will be co-incubated with PI in several cytotoxicity assays and finally different anti-GRP78 antibodies will be evaluated in combination with PI in sensitive and refractory cell lines and patients material. Positive combinations will be validated in an experimental animal model of MM. The aim of the study is to obtain insights in the molecular pathogenesis of GRP78 mediated resistance and on the other hand to develop an antibody based treatment strategy to overcome PI resistance in refractory MM patients.