Molecular Pathogenesis of Multiple Myeloma

我们继续在多发性骨髓瘤（MM）和意义不明的单克隆γ病（MGUS）中识别和表征Ig易位。首先，我们确定了MAFA （8q24.3）是一个新的复发易位伙伴。这种易位发生在约1%的MM肿瘤中，与我们之前发现的c-MAF和MAFB易位具有相同的基因表达谱（最值得注意的是CYCLIN D2的高表达）。其次，我们克隆了MM中的第一个Ig lambda易位，并确定了失调的靶点为nfkb诱导激酶（NIK/MAP3K14）。我们正在积极研究NFKB在MM发病机制中的作用。第三，我们对46个人类MM细胞系（HMCL）进行了阵列CGH分析，并确定了复发性双等位基因缺失和可能在肿瘤发生中重要的基因扩增。这也使我们能够更精确地绘制针对c-MYC、CYCLIN D1和其他基因的易位断点。第四，我们刚刚完成了46例HMCL和50例原发性MM肿瘤的分子细胞遗传学分析。这使我们能够深入了解MM中原发性易位和继发性易位的机制和作用。最后，我们对易位和基因表达谱的研究使我们能够提出一个假设，即CYCLIN D基因的失调是MGUS和MM肿瘤的早期和统一事件。这些研究还使我们能够提出TC（易位/ CYCLIN D）分类，该分类确定了8组具有不同生物学和临床特性的肿瘤。我们继续研究MM的其他致癌事件。首先，我们确定K-和N-RAS突变在MGUS中不常见（少于5%），但在MM肿瘤中发生约30%，因此RAS突变有时可能是MGUS/MM转变的标志，如果不是因果事件。此外，我们发现K-RAS突变的流行率在所有TC组中相似，而N-RAS突变的流行率在不同TC组中有显著差异，这一发现我们尚未理解。其次，我们已经确定p16INK4a在MM肿瘤中的表达水平通常非常低，与基因是否甲基化无关，尽管一些MM肿瘤和细胞系表达该基因的水平相当高。第三，我们已经确定p18INK4c的表达，它似乎是正常浆细胞G1>S转变的关键调节因子，提供了一个重要的增殖标志物。在大约30%的HMCL和近10%的增殖性MM肿瘤中，p18的双等位基因缺失似乎是导致增殖增加的关键事件。然而，矛盾的是，高增殖的HMCL和MM肿瘤中约有60%的p18水平升高，这表明这些肿瘤细胞对增殖增加导致的p18水平升高不敏感。一小部分表达高水平p18的HMCL和增生性肿瘤使RB1失活，这可以解释对高水平p18不敏感的原因。我们正在继续尝试确定当p18水平高时导致增殖增加的其他机制。最后，我们已经确定了46例HMCL中90%以上的p53失活的机制。

We continue to identify and characterize Ig translocations in multiple myeloma (MM) and monoclonal gammopathy of undetermined significance (MGUS). First, we have identified MAFA (8q24.3) as a new recurrent translocation partner. This translocation, which occurs in about 1% of MM tumors, is associated with the same gene expression profile (most notably, very high expression of CYCLIN D2) as c-MAF and MAFB translocations that we identified previously. Second, we have cloned the first Ig lambda translocation in MM, and have identified the dysregualated target as NFKB-inducing kinase (NIK/MAP3K14). We are actively investigating the role of NFKB in the pathogenesis of MM. Third, we have done array CGH analyses on our panel of 46 human MM cell lines (HMCL), and have identified recurrent bi-allelic deltions and amplifications of genes that are likely important in oncogenesis. This also has enabled us to map more precisely translocation breakpoints that target c-MYC, CYCLIN D1, and other genes. Fourth, we have just completed molecular cytogenetic analyses of 46 HMCL and 50 primary MM tumors. This has enabled us to gain insights regarding the mechanisms causing and roles of primary vs secondary translocations in MM. Finally, our studies on translocations and gene expression profiling enabled to propose the hypothesis that dysregulation of a CYCLIN D gene is a an early and unifying event in MGUS and MM tumors. These studies also enabled us to propose a TC (translocation/ CYCLIN D) classification, which identified 8 groups of tumors with differing biolgical and clinical properties.We continue to investigate other oncogenic events in MM. First, we have determined that K- and N-RAS mutations are infrequent (less than 5%) in MGUS but occur in about 30% of MM tumors, so that RAS mutations may sometimes be a marker if not a causal event in the MGUS/MM transition. In addition, we found that the prevalence of K-RAS mutations is similar in all TC groups, whereas the prevalence of N-RAS mutations differs markedly among the different TC groups, a finding that we do not yet understand. Second, we have determine that p16INK4a is usually expressed at very low levels in MM tumors independent of whether or not the gene is methylated, although some MM tumors and cell lines express quite high levels of this gene. Third, we have determined that the expression p18INK4c, which seems to be a key regulator of the G1>S transition in normal plasma cells, provides an important marker of proliferation. In approximately 30% of HMCL and nearly 10% of proliferative MM tumors, the bi-allelic deletion of p18 appears to be a critical event that leads to increased proliferation. Paradoxically, however, about 60% of both HMCL and MM tumors that are hightly proliferative have increased levels of p18, suggesting that these tumor cells are insensitive to the increased levels of p18 that occur as a consequence of increased proliferation. A small fraction of the HMCL and proliferative tumors that express high levels of p18 have inactivated RB1, which would explain the insensitivy to high p18 levels. We are continuing to attempt to identify other mechanisms responsible for increased proliferation when there are high p18 levels. Finally, we have been determing the mechanisms by which p53 is inactivated in more than 90% of our panel of 46 HMCL.