Abstract Multiple myeloma (MM) cells are the malignant counterpart of post-germinal center (GC) long-lived PCs, characterized by strong BM dependence, somatic hypermutation (SHM) of immunoglobulin (Ig) genes, and isotype class switch recombination (CSR) resulting in absence of IgM expression in all but 1% of tumors. Virtually every case of MM is preceded by a pre-malignant PC tumor called monoclonal gammopathy of undetermined significance (MGUS) that, like MM, produces a typical M-spike by SPEP or free light chain in the urine. Translocations involving the immunoglobulin heavy chain (IgH) locus are present in at least half of MM cases and are thought to result from errors during the physiological process of CSR or SHM. These translocations result in dysregulated expression of an oncogene that is juxtaposed to the strong Ig enhancers. There are three recurrent primary IgH translocation groups in MM: CYCLIN D (11q13-CYCLIN D1; 12p13-CYCLIN D2; 6p25-CYCLIN D3) MAF (16q23-MAF; 20q12-MAFB; 8q24.3-MAFA; MMSET/(FGFR3)-4p16-(MMSET in all but also FGFR3 in 80% of these tumors. Multiple trisomies represent an alternative pathogenetic pathway. There is a consensus that chromosome content reflects at least two pathways of pathogenesis. Nearly half of MGUS and MM tumors are hyperdiploid (HRD), usually with extra copies of three or more specific chromosomes (3,5,7,9,11,15,19, 21). Strikingly, HRD tumors rarely (~10%) have a primary IgH translocation, whereas Non-hyperdiploid (NHRD) tumors usually (~70%) have an IgH translocation. Almost all cases of plasma cell neoplasm starting from the MGUS stage and independently on the chromosome content aberrantly express one or more of the CYCLIN D genes and it has been proposed that dysregulation of a CYCLIN D gene provides a unifying, early oncogenic event in MGUS and MM. Consistently, the dysregulation of CYCLIN D1 in mouse B cells is sufficient to induce a monoclonal gammopathy that does not progress to MM. Secondary oncogenic events drive MGUS and MM progression MYC dysregulation. There is increased expression of c-MYC in most newly diagnosed MM tumors compared to MGUS tumor. Recently, it was shown that sporadic activation of a MYC transgene (Vk*MYC) in GC B cells in an MGUS prone mouse strain led to the universal development of MM tumor. Hence, increased MYC expression may be responsible for progression from MGUS to MM. Complex translocations involving MYC appear to be secondary progression events that often do not involve Ig loci, but rather other super enhancers that are very active in plasma cells. They are rare or absent in MGUS, but recently identified in over 40% of untreated MM, and 90% of MM cell lines. Activating mutations of RAS and BRAF. The prevalence of activating NRAS or KRAS mutations is about 15-18% each in newly diagnosed and relapsed MM tumors, and recently, BRAF mutations were described in 4% of MM tumors. Expression of an activated of KRAS in mouse B cells is sufficient to induce a monoclonal gammopathy but it does not progress to MM. Not surprisingly, dysregulating pairs of CYCLIN D1, KRAS and/or MYC in mouse B cells accelerated the progression of MM. Activating mutations of NFkappaB pathway. Extrinsic ligands (APRIL and BAFF) produced by BM stromal cells provide critical survival signals to long-lived PCs by stimulating TACI, BCMA, and BAFF receptors to activate the NFKB pathways. Most MGUS and MM tumors highly express NFKB target genes, suggesting a continued role of extrinsic signaling in PC tumors and at least in part explaining the constant dependency of MM cells on the BM microenvironment. Activating mutations in positive regulators and inactivating mutations in negative regulators of the NFKB pathway have been identified in at least 20% of untreated MM tumors and ~50% of HMCLs, rendering the cells less dependent on ligand-mediated NFKB activation and most likely contributing to extra-medullary spread of the disease. Additional changes include monosomy 13 (DIS3, mir15/16, RB1), del17p (TP53), add1q (MCL1) and del1p (CDKN2C, FAM46C). While the role of these putative tumor suppressor/oncogene remain to be defined, we found that loss of one copy of miR15/16 cooperated with MYC to cause the progression of MM in mice, while loss of one copy of RB1 did not, implicating loss of miR15/16 as one driving force behind selection for monosomy 13 in human MM. High-risk MM is associated with intra-clonal tumor heterogeneity Recent evidence indicates that tumor heterogeneity is prevalent in MM, as in many other cancers, and that different subclones are present within the tumor population, characterized by distinct genetic mutations that contributed independently to the tumor progression. The degree of clonal heterogeneity is presumably related to the degree of genomic instability, and appears to correlate with the presence of high-risk genetics. Three patterns of relapse are seen following successful therapy. A third of the time the dominant clone at diagnosis predominates, unchanged, at relapse. A third of the time the diagnostic clone has acquired additional genetic changes at relapse. And a third of the time an ancestral clone to the one dominant at diagnosis emerges, lacking some of the genetic changes present in the diagnostic clone, and having acquired additional genetic changes. Activating the immune system to overcome tumor heterogeneity MM cells develop a malignant relationship with components of the innate immune system in the BM. Initially inflammatory macrophages support MM tumor development, but over time convert to an immunosuppressive M2 phenotype. This relationship can be disrupted by activating signaling downstream of TLR and TNFR receptors in macrophages through the use of IAP antagonists which target for destruction negative regulators of these pathways (cIAP1/2). In MM bearing mice with a mature (but not those with an immature) immune system this results in a complete response, with no evidence of a direct anti-tumor effect of IAP antagonists on the tumor cells at clinically relevant doses. This has led to a phase II clinical trial of IAP antagonists to activate the innate immune system in patients with MM. The treatment is associated with dramatic activation and mobilization of macrophages, with cytokine release and early evidence of anti-tumor effects. Conclusion Significant progress has been made is understanding the molecular pathogenesis and biology of MM. Oncogenic pathways can be activated through cell intrinsic or extrinsic mechanisms. Similar to other cancers, MM is characterized by multi-stage accumulation of genetic abnormalities deregulating different pathways. Much of this knowledge is already being utilized for diagnosis, prognosis and risk-stratification of patients. Importantly, from a clinical standpoint, this knowledge has led to development of novel therapeutic strategies, some of which are already in clinical use, and many others showing promise in pre-clinical and early clinical studies. Citation Format: Marta Chesi, P. Leif Bergsagel. Targeting genetic heterogeneity in multiple myeloma through immune activation. [abstract]. In: Proceedings of the AACR Special Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; Sep 20-23, 2014; Philadelphia, PA. Philadelphia (PA): AACR; Clin Cancer Res 2015;21(17 Suppl):Abstract nr IA25.