Identification of novel mutational drivers reveals oncogene dependencies in multiple myeloma.

Understanding the profile of oncogene and tumor suppressor gene mutations with their interactions and impact on the prognosis of multiple myeloma (MM) can improve the definition of disease subsets and identify pathways important in disease pathobiology. Using integrated genomics of 1273 newly diagnosed patients with MM, we identified 63 driver genes, some of which are novel, including IDH1, IDH2, HUWE1, KLHL6, and PTPN11 Oncogene mutations are significantly more clonal than tumor suppressor mutations, indicating they may exert a bigger selective pressure. Patients with more driver gene abnormalities are associated with worse outcomes, as are identified mechanisms of genomic instability. Oncogenic dependencies were identified between mutations in driver genes, common regions of copy number change, and primary translocation and hyperdiploidy events. These dependencies included associations with t(4;14) and mutations in FGFR3, DIS3, and PRKD2; t(11;14) with mutations in CCND1 and IRF4; t(14;16) with mutations in MAF, BRAF, DIS3, and ATM; and hyperdiploidy with gain 11q, mutations in FAM46C, and MYC rearrangements. These associations indicate that the genomic landscape of myeloma is predetermined by the primary events upon which further dependencies are built, giving rise to a nonrandom accumulation of genetic hits. Understanding these dependencies may elucidate potential evolutionary patterns and lead to better treatment regimens.

Is molecular remission the goal of multiple myeloma therapy?

The increased number of effective therapies and the wider use of combinations that give deeper remissions have resulted in a reassessment of the goals of myeloma therapy. With the advent of new therapeutic strategies and diagnostic tools, achievement of minimal residual disease (MRD)-negative status has become increasingly important, with some even considering it as the primary endpoint for therapy. The level of MRD that is aimed for is a continuous, rather than an absolute variable, with studies in both transplant-eligible and -noneligible patients showing that the level of MRD achieved is predictive of progression-free survival and overall survival, with an improvement in survival of approximately 1 year for each log-depletion in MRD level. The most widely used methods to assess MRD status include flow cytometry and clonality detection, using next-generation sequencing technologies with sensitivity limits of 1:10-3 to 1:10-6 The timing of when to assess MRD depends on the treatment used, as well as the molecular and cytogenetic subgroup of the myeloma itself. It is also becoming clear that the level of MRD negativity, as well as microenvironmental factors, are important prognostically, including the regeneration of normal plasma cells, and the normalization of the immune repertoire. With advances in antibody-based therapy and immunotherapy, the achievement of stable MRD states is now possible for a significant proportion of patients, and is a prerequisite for myeloma cure.

Anti-myeloma effects of ruxolitinib combined with bortezomib and lenalidomide: A rationale for JAK/STAT pathway inhibition in myeloma patients.

JAK proteins have been linked with survival and proliferation of multiple myeloma (MM) cells; therefore, JAK inhibition could be a therapeutic strategy for MM. We evaluated JAK1 and JAK2 expression in MM patients and the effects of JAK/STAT pathway inhibition on apoptosis, cell cycle, gene and protein expression in RPMI-8226 and U266 MM cell lines. 57% of patients presented overexpression of JAK2 and 27%, of JAK1. After treatment with ruxolitinib and bortezomib, RPMI-8226 and U266 presented 50% of cells in late apoptosis, reduction of anti-apoptotic genes expression and higher number of cells in SubG0 phase. Co-culture with stromal cells protected RPMI-8226 cells from apoptosis, which was reversed by lenalidomide addition. Combination of ruxolitinib, bortezomib and lenalidomide induced 72% of cell death, equivalent to bortezomib, lenalidomide and dexamethasone, combination used in clinical practice. Many JAK/STAT pathway genes, after treatment, had their expression reduced, mainly in RPMI-8226, with insignificant changes in U266. In this scenario, JAK/STAT pathway could pose as a new therapeutic target to be exploited, since it is constitutively active and contributes to survival of MM tumor cells.

MAF protein mediates innate resistance to proteasome inhibition therapy in multiple myeloma.

Multiple myeloma (MM) patients with the t(14;16) translocation have a poor prognosis, and unlike other molecular subgroups, their outcome has not improved with the introduction of bortezomib (Bzb). The mechanism underlying innate resistance of MM to Bzb is unknown. In the present study, we have investigated how MAF overexpression impacts resistance to proteasome inhibitor (PI) therapy (Bzb and carfilzomib). High levels of MAF protein were found in t(14;16) cell lines; cell lines from the t(4;14) subgroup had intermediate levels, whereas cell lines from the other subgroups had low levels. High expression of MAF protein in t(14;16) was associated with significantly higher PI half-maximum inhibitory concentration values compared with other molecular subgroups. PI exposure abrogated glycogen synthase kinase 3ß (GSK3ß)-mediated degradation of MAF protein, resulting in increased MAF protein stability and PI resistance. Subsequent studies using loss-of-function and gain-of-function models showed that silencing MAF led to increased sensitivity to PIs, enhanced apoptosis, and activation of caspase-3, -7, -8, -9, poly (ADP-ribose) polymerase, and lamin A/C. In contrast, overexpression of MAF resulted in increased resistance to PIs and reduced apoptosis. These results define the role of MAF and GSK3 in the resistance of t(14;16) MM to PIs and identifies a novel mechanism by which MAF protein levels are regulated by PIs, which in turn confers resistance to PIs.

Evidence of an epigenetic origin for high-risk 1q21 copy number aberrations in multiple myeloma.

Multiple myeloma is a B-cell malignancy stratified in part by cytogenetic abnormalities, including the high-risk copy number aberrations (CNAs) of +1q21 and 17p(-). To investigate the relationship between 1q21 CNAs and DNA hypomethylation of the 1q12 pericentromeric heterochromatin, we treated in vitro peripheral blood cultures of 5 patients with balanced constitutional rearrangements of 1q12 and 5 controls with the hypomethylating agent 5-azacytidine. Using G-banding, fluorescence in situ hybridization, and spectral karyotyping, we identified structural aberrations and copy number gains of 1q21 in the treated cells similar to those found in patients with cytogenetically defined high-risk disease. Aberrations included 1q12 triradials, amplifications of regions juxtaposed to 1q12, and jumping translocations 1q12. Strikingly, all 5 patients with constitutional 1q12 rearrangements showed amplifications on the derivative chromosomes distal to the inverted or translocated 1q12 region, including MYCN in 1 case. At the same time, no amplification of the 1q21 region was found when the 1q12 region was inverted or absent. These findings provide evidence that the hypomethylation of the 1q12 region can potentially amplify any genomic region juxtaposed to it and mimic CNAs found in the bone marrow of patients with high-risk disease.