Faculty Bibliography

PURPOSE/OBJECTIVE:Our purpose in this report was to define genes and pathways dysregulated as a consequence of the t(4;14) in myeloma, and to gain insight into the downstream functional effects that may explain the different prognosis of this subgroup. EXPERIMENTAL DESIGN/METHODS:Fibroblast growth factor receptor 3 (FGFR3) overexpression, the presence of immunoglobulin heavy chain-multiple myeloma SET domain (IgH-MMSET) fusion products and the identification of t(4;14) breakpoints were determined in a series of myeloma cases. Differentially expressed genes were identified between cases with (n = 5) and without (n = 24) a t(4;14) by using global gene expression analysis. RESULTS:Cases with a t(4;14) have a distinct expression pattern compared with other cases of myeloma. A total of 127 genes were identified as being differentially expressed including MMSET and cyclin D2, which have been previously reported as being associated with this translocation. Other important functional classes of genes include cell signaling, apoptosis and related genes, oncogenes, chromatin structure, and DNA repair genes. Interestingly, 25% of myeloma cases lacking evidence of this translocation had up-regulation of the MMSET transcript to the same level as cases with a translocation. CONCLUSIONS:t(4;14) cases form a distinct subgroup of myeloma cases with a unique gene signature that may account for their poor prognosis. A number of non-t(4;14) cases also express MMSET consistent with this gene playing a role in myeloma pathogenesis.

The ability of donor lymphocyte infusions (DLIs) to induce complete responses (CRs) in patients with relapsed myeloma after allogeneic bone marrow transplantation (BMT) provides clear evidence of an effective graft-versus-myeloma (GVM) response. To identify target antigens of the GVM response, we screened a myeloma cDNA expression library with post-DLI serum from 4 patients with myeloma who achieved CR after DLI and 1 patient who was in CR before DLI. We identified a panel of 13 gene products reactive with post-DLI serum but negative with pre-DLI and pre-BMT serum. Antibodies to these proteins were not detected in the sera of 10 patients who underwent allogeneic BMT without DLI and 5 patients with acute graft-versus-host disease (GVHD). Minimal reactivity with these proteins was detected in the sera of 20 healthy donors and 20 patients with chronic GVHD. In contrast, 5 of these proteins were recognized by more than 1 myeloma DLI responder. Testing of serial serum samples showed an association between antibody response and time of best response after DLI. The expression of these genes was evaluated in primary myeloma cells and in normal plasma cells. This study demonstrates that the GVM response is associated with antibody responses to highly expressed myeloma-associated antigens.

Much has been learned regarding the biology and clinical implications of genetic abnormalities in multiple myeloma. Because of recent advances in the field, an International Workshop was held in Paris in february of 2003. This summary describes the consensus recommendations arising from that meeting with special emphasis on novel genetic observations. For instance, it is increasingly clear that translocations involving the immunoglobin heavy-chain locus are important for the pathogenesis of one-half of patients. As a corollary, it also clear that the remaining patients, lacking IgH translocations, have hyperdiploidy as the hallmark of their disease. Several important genetic markers are associated with a shortened survival such as chromosome 13 monosomy, hypodiploidy, and others. The events leading the transformation of the monoclonal gammopathy of undetermined significance (MGUS) to myeloma are still unclear. One of the few differential genetic lesions between myeloma and MGUS is the presence of ras mutations in the latter. Gene expression platforms are capable of detecting many of the genetic aberrations found in the clonal cells of myeloma. Areas in need of further study were identified. The study of the genetic aberrations will likely form the platform for targeted therapy for the disease.

Multiple myeloma (MM) cells express certain tumour-associated antigens (TAAs) that could serve as targets for active-specific immunotherapy. The aim of the present study was to test the MM/dendritic cell (DC) fusion as a vaccination strategy. We fused MM cells with DC to generate fusion cells (FCs) and tested their antigen presenting cell (APC) function in mixed lymphocyte reactions and cytotoxicity assays. First, the HS Sultan and SK0-007 HAT sensitive human MM cell lines and DCs generated from peripheral blood of normal donors were fused in the presence of 50% polyethylene glycol to form FCs. Next, tumour cells freshly isolated from patients were similarly fused with autologous DCs to generate FCs. The FCs demonstrated a biphenotypic profile, confirmed both by flow-cytometry and dual immunofluorescence microscopy. These FCs induced MM-specific cytotoxicity. FCs, but not MM cells or DCs alone, were potent stimulators of autologous patient T cells. More importantly, FC-primed autologous peripheral blood mononuclear cells demonstrated major histocompatibility complex-restricted MM-specific cytolysis. These studies therefore demonstrated that MM/DC FC can trigger an autologous immune response to MM cells and formed the framework for a clinical trial currently underway.

We have previously shown that thalidomide and its potent immunomodulatory derivatives (IMiDs) inhibit the in vitro growth of multiple myeloma (MM) cell lines and patient MM cells that are resistant to conventional therapy. In this study, we further characterize the effect of these drugs on growth of B cell malignancies and angiogenesis. We established a beige-nude-xid (BNX) mouse model to allow for simultaneous in vivo measurement of both anti-tumor and anti-angiogenic effects of thalidomide and its analogs. Daily treatment (50 mg/kg/d) with thalidomide or IMiDs was nontoxic. The IMiDs were significantly more potent than thalidomide in vivo in suppressing tumor growth, evidenced by decreased tumor volume and prolonged survival, as well as mediating anti-angiogenic effects, as determined by decreased microvessel density. Our results therefore show that the IMiDs have more potent direct anti-tumor and anti-angiogenic effects than thalidomide in vivo, providing the framework for clinical protocols evaluating these agents in MM and other B cell neoplasms.

The introduction of microarrays offers the opportunity to examine the expression of many thousands of genes in a single experiment. Investigations in leukaemia and lymphoma have led to the identification of a number of subgroups, with a defined gene expression pattern, not previously identified by morphology, cytogenetics or molecular techniques. In many cases these expression patterns can be linked to the tumour cells normal developmental counterpart, and represent distinct disease subgroups with different clinical presentations and outcomes. The technology has also identified genes that may be important in tumour cell biology including key genes in cell proliferation, adhesion, apoptosis, and the development of drug resistance. These early studies demonstrate that genetic microarrays will be useful in classifying haematological malignancies, predicting response to treatment, predicting prognosis, and identifying novel targets for therapy.