Through several research projects led by Dr. Bergsagel, the Cancer Genetics Laboratory is investigating the development of multiple myeloma and related conditions. The lab's research on oncogene activation, genetic mutations and gene-expression profiling is helping better characterize disease progression in multiple myeloma and other bone marrow and blood disorders, which ultimately could lead to better treatment options.

Oncogene activation

Based on a novel mouse model that activates oncogene-expression using somatic hypermutation (occurring in 10 percent of the translocations in multiple myeloma), the Cancer Genetics Lab has developed a faithful model of multiple myeloma. The mouse model uses the c-Myc oncogene, which is critically important in human multiple myeloma — translocated in 15 percent of newly diagnosed multiple myeloma, 40 percent of advanced refractory multiple myeloma and more than 90 percent of human myeloma cell lines. A major clinical implication of these studies is that both the timing of oncogene activation and the nature of the oncogene are critical determinants of the resulting disease phenotype.

AID-dependent MYC activation

The enzymatic activity of activation-induced deaminase (AID) during the physiological processes of somatic hypermutation and class switch recombination occurring in germinal center B lymphocytes has been implicated in oncogene activation in mature B cell malignancies. By misdirecting the activity of AID to a conditional MYC transgene, Dr. Bergsagel's research team has achieved sporadic, AID-dependent MYC activation in germinal center B cells of Vk*MYC mice.

Whereas control C57Bl/6 mice develop benign monoclonal gammopathy with age, in contrast, all Vk*MYC mice progress to an indolent multiple myeloma associated with the biological and clinical features highly characteristic of the human disease. In one-third of mice, a more aggressive extramedullary form of myeloma develops and could be accelerated by immunization or through secondary signals including deliberate co-expression of Bcl2.

Consistent with these findings in mice, more-frequent MYC rearrangements, elevated levels of MYC mRNA and MYC target genes distinguish people with multiple myeloma from people with monoclonal gammopathy, implicating a causal role for MYC in the progression of monoclonal gammopathy to multiple myeloma.

The NF-KB pathway

While activation of NF-KB has been noted in many tumor types, only rarely has it been linked to an underlying genetic mutation. An integrated analysis of high-density oligonucleotide array CGH and gene-expression profiling data from 155 multiple myeloma samples identified a promiscuous array of abnormalities contributing to the dysregulation of NF-KB in about 20 percent of patients.

The Cancer Genetics Lab has reported mutations in 10 genes causing the inactivation of TRAF2, TRAF3, CYLD, cIAP1/cIAP2, and activation of NFKB1, NFKB2, CD40, LTBR, TACI and NIK that result primarily in constitutive activation of the noncanonical NF-KB pathway, with the single most common abnormality being inactivation of TRAF3. These results highlight the critical importance of the NF-KB pathway in the pathogenesis of multiple myeloma.

Gene-expression profiling

In a gene-expression study, Dr. Bergsagel's research team compared the transcription profiles of Waldenstrom macroglobulinemia with those of other malignant B cells, including chronic lymphocytic leukemia and multiple myeloma, as well as normal cells (peripheral blood B cells and bone marrow plasma cells). The Cancer Genetics Lab found that Waldenstrom macroglobulinemia has a homogenous gene-expression regardless of 6q deletion status and clusters with chronic lymphocytic leukemia and normal B cells on unsupervised clustering with very similar expression profiles. Only a small gene set has expression profiles unique to Waldenstrom macroglobulinemia compared with chronic lymphocytic leukemia and multiple myeloma.

The most significantly up-regulated gene is IL-6 and the most significantly associated pathway for this set of genes is MAPK signaling. Thus, IL-6 and its downstream signaling may be of biological importance in Waldenstrom macroglobulinemia. Further elucidation of the role of IL-6 in Waldenstrom macroglobulinemia is warranted as this may offer a potential therapeutic avenue.

Cyclin D dysregulation

Two oncogenic pathways have been hypothesized for multiple myeloma and premalignant monoclonal gammopathy of undetermined significance (MGUS) tumors: a nonhyperdiploid pathway associated with a high prevalence of IgH translocations and a hyperdiploid pathway associated with multiple trisomies of eight chromosomes. Cyclin D1, D2 or D3 expression seems to be increased or dysregulated, or both, in virtually all multiple myeloma tumors despite their low proliferative capacity. Translocations can directly dysregulate CCND1 (11q13) or CCND3 (6p21), or MAF (16q23) or MAFB (20q11) transcription factors that target CCND2. Biallelic dysregulation of CCND1 occurs in nearly 40 percent of tumors, most of which are hyperdiploid. Other tumors express increased CCND2, either with or without a t(4;14) translocation.

Using gene-expression profiling to identify five recurrent translocations, specific trisomies and expression of cyclin D genes, multiple myeloma tumors can be divided into eight translocation/cyclin D groups (11q13, 6p21, 4p16, maf, D1, D1+D2, D2 and none) that seem to be defined by early, and perhaps initiating, oncogenic events. However, despite subsequent progression events, these groups have differing gene-expression profiles and significant differences in the prevalence of bone disease, frequency at relapse and progression to extramedullary tumor.

Overexpression of c-Maf

The oncogene c-Maf is translocated in about 5 to 10 percent of multiple myelomas. Unexpectedly, Dr. Bergsagel's research team observed c-Maf expression in myeloma cell lines lacking c-Maf translocations and in 50 percent of multiple myeloma bone marrow samples. By gene-expression profiling, the lab identified three c-Maf target genes: cyclin D2, integrin beta7 and CCR1. C-Maf transactivated the cyclin D2 promoter and enhanced myeloma proliferation, whereas dominant inhibition of c-Maf blocked tumor formation in immunodeficient mice. C-Maf-driven expression of integrin beta7 enhanced myeloma adhesion to bone marrow stroma and increased production of VEGF.

The Cancer Genetics Lab research team proposes that c-Maf transforms plasma cells by stimulating cell cycle progression and by altering bone marrow stromal interactions. The frequent overexpression of c-Maf in myeloma makes it an attractive target for therapeutic intervention.