Haematologica 2003 - Supplements

4.4 Evolving from MGUS/ SMOLDERING MM to
symptomatic MM.
091
Characterisation of the Transition of MGUS to Multiple
Myeloma using Expression Microarrays.
Ann M. Dring, Faith E. Davies, Cheng Li, Andrew C.
Rawstron, Masood A. Shammas, James A. Fenton, Teru
Hideshima, Dharminder Chauhan, Karen Rees, David
Gonzalez-Castro, Daniel Auclair, Wing H. Wong, Nikhil C.
Munshi, Gareth J. Morgan, Kenneth C. Anderson.
Academic Department of Haematology and Oncology, University of
Leeds, Leeds, West Yorkshire, United Kingdom; Department of
Adult Oncology, Dana-Farber Cancer Institute, Boston, MA, USA;
Department of Biostatistics, Harvard School of Public Health,
Boston, MA, USA
MGUS can be considered a premalignant phase of myeloma
(MM) and MGUS plasma cells (PCs) although distinct in their
clinical behavior are clearly related to MM PCs. Understanding
the molecular basis of the transition from MGUS to MM can
provide considerable insight into the multi-step pathogenesis of
MM. This transition has been studied using cytogenetics and
mutational analysis however consistent changes suitable for the
further investigation have not been identified. Global expression
based analysis can highlight genes and gene families important in
this transition which may be potential future therapeutic targets.
We have analyzed PCs from 5 normals (N), 5 MGUS and 31 MM
following CD138+ selection and SMART PCR-based
amplification using the Affymetrix U95Av2 gene chip comparing
12,000 known expressed sequences (10,000 genes).
Unsupervised analyses were performed to identify genes which
had most variation across all samples and supervised analyses,
using the ‘compare samples’ function in DCHIP, looked for
genes which varied significantly between specific sample groups.
Hierarchical clustering was then used to study the results. 380
genes separated N PCs from MM PCs, whereas 263 genes
separated N PCs from MGUS PCs. Interestingly the majority of
genes were downregulated (252 downregulated, 128
upregulated). The transition of MGUS to MM was more closely
examined and using the same strict analysis criteria the number of
genes separating MGUS and MM was considerably fewer (74
genes), than those separating N and malignant PCs suggesting
that MGUS and MM PCs are more similar to each other than to
PCs from normal donors. The underlying basis of these changes
may be understood more clearly by looking at the functional
classes of genes that are altered. Important genes in the N vs
Malignant comparison include: oncogenes and tumor suppressor
genes (c-myb, LAF4, DOC1, Rb1); cell signaling genes (CD163,
small inducible cytokine subfamily C and CDK2 associated
protein 2); death genes (MAD3 and beclin 1); DNA binding and
transcription factors (XBP1, YY1, CBF and seven in absentia);
and developmental genes (WNT and sonic pathways). In
comparison the genes differentiating MGUS from MM are more
limited but include a number of potentially important genes
affecting cell growth and maintenance, signal transduction,
structural proteins, and developmental processes. Interestingly
no genes involved in apoptosis were highlighted as being
differentially expressed between MGUS and MM. The data sets
were validated by comparing the expression of genes
differentiating N and malignant PCs using flow cytometry and
RT-PCR. Using these techniques there was good correlation with
gene expression data identified on the arrays, and also good
correlation with previously published data. In conclusion gene
array analysis highlights the differences in gene expression levels
between N, MGUS, and MM PCs and supports the multi-step
pathogenesis of MM. Genes involved in the control of
transcription and developmental pathways are important in this
transition, with the critical differences relating to the transition
from normal to MGUS. To fully define this transition further
analysis of different sub-populations of CD138+ PCs in MGUS
in comparison with N PCs populations is required. The
amplification technique used in this study will be critical for this
analysis.
092
ROLE OF ANTI-TUMOR IMMUNE EFFECTORS IN THE
CONTROL OF “MALIGNANT TRANSFORMATION” IN
GAMMOPATHIES.
M. V. Dhodapkar, M. Geller, J. Krasovsky, D. Chang
Laboratory of Tumor Immunology and Immunotherapy, Rockefeller
University, and Memorial Sloan Kettering Cancer Center, New
York, NY
Clonal expansion of a transformed cell is essential, but not
sufficient for the development of clinical cancer. This is
particularly puzzling in the case of monoclonal gammopathies,
because many of the cytogenetic and genomic changes initially
described in myeloma have now also been detected in premalignant
monoclonal gammopathy of undetermined significance
(MGUS). Immune system has been postulated to play a role in
surveillance of tumors in mice. The role of immune effectors in
the control of transformed cells in patients with gammopathies
remains to be fully defined. We have studied tumor reactive
immune effectors in the blood and tumor bed of patients with
MGUS and myeloma. We have recently shown that freshly
isolated T cells from blood or tumor bed of patients with
progressive myeloma lack tumor reactive rapid effector function.
Targeting tumor antigens to Fc receptors of dendritic cells
(DCs) leads to enhancement of cross presentation and generation
of tumor reactive effector T cells (Dhodapkar et al, J Exp Med
2002). Using this approach, T cells from the tumor bed of even
patients with progressive tumors can be activated to yield tumor
reactive killer T cells (Dhodapkar et al, PNAS 2002). Anti-tumor
reactivity in these cultures was specific for autologous tumor, and
mostly not directed against Ig derived determinants. Natural killer
T cells are distinct lymphocytes that recognize glycolipid
antigens in the context of CD1 family of antigen presenting
molecules. We find that clinical progression in myeloma is
associated with a loss of ligand reactive rapid effector function in
NKT cells, which can be restored ex vivo using dendritic cells.
Direct analysis of tumor specific effector T cell function in
MGUS now indicates the presence of an active anti-tumor
effector T cell response. This response is enriched in the marrow
and is specific for antigens expressed by tumor cells in each
patient. These data, to our knowledge, provide the first direct
evidence for active and tumor specific immune recognition in the
bed of a human non-viral “preneoplastic state”, and strongly
suggest that the development of “clinical myeloma” is regulated
at least in part at the level of the host immune response consisting
of both innate and adaptive immune effectors. The finding that
reversible host factors might determine clinical malignancy may
also have important implications for therapeutic goals in these
diseases.
S129