Correction: Pomalidomide, bortezomib, and low-dose dexamethasone in lenalidomide-refractory and proteasome inhibitor-exposed myeloma.

Following the publication of this article, the authors noted that the pomalidomide dose for the additional SC cohort in Fig. 1 was incorrectly listed. The correct dose for pomalidomide in the additional SC cohort should be the maximum tolerated dose of 4 mg/day, not 2 mg/day as listed in the original Fig. 1. The authors apologize for any inconvenience caused.

Integrative network analysis identifies novel drivers of pathogenesis and progression in newly diagnosed multiple myeloma.

Multiple myeloma (MM) is an incurable malignancy of bone marrow plasma cells characterized by wide clinical and molecular heterogeneity. In this study we applied an integrative network biology approach to molecular and clinical data measured from 450 patients with newly diagnosed MM from the MMRF (Multiple Myeloma Research Foundation) CoMMpass study. A novel network model of myeloma (MMNet) was constructed, revealing complex molecular disease patterns and novel associations between clinical traits and genomic markers. Genomic alterations and groups of coexpressed genes correlate with disease stage, tumor clonality and early progression. We validated CDC42BPA and CLEC11A as novel regulators and candidate therapeutic targets of MMSET-related myeloma. We then used MMNet to discover novel genes associated with high-risk myeloma and identified a novel four-gene prognostic signature. We identified new patient classes defined by network features and enriched for clinically relevant genetic events, pathways and deregulated genes. Finally, we demonstrated the ability of deep sequencing techniques to detect relevant structural rearrangements, providing evidence that encourages wider use of such technologies in clinical practice. An integrative network analysis of CoMMpass data identified new insights into multiple myeloma disease biology and provided improved molecular features for diagnosing and stratifying patients, as well as additional molecular targets for therapeutic alternatives.

14-3-3ζ binds the proteasome, limits proteolytic function and enhances sensitivity to proteasome inhibitors.

14-3-3 proteins are a family of master regulators of intracellular signaling, yet their impact on proteasome function is unknown. We demonstrate that 14-3-3ζ binds the 11S proteasome activator, limiting proteasome assembly and cellular capacity for protein degradation. To define the functional impact of 14-3-3ζ proteasomal binding in myeloma cells, silencing and overexpression experiments are performed. We find that downregulation of 14-3-3ζ impairs myeloma cell growth and confers resistance to clinically used proteasome inhibitors. In a large cohort of newly diagnosed myeloma patients, elevated expression of 14-3-3ζ is associated with high risk myeloma genetic subtypes and worse prognosis overall. Our work demonstrates the important role of 14-3-3ζ in regulating proteasome function, myeloma cell growth and sensitivity to therapeutics, and suggests regulation of 14-3-3ζ as a new approach in myeloma therapy.

High somatic mutation and neoantigen burden are correlated with decreased progression-free survival in multiple myeloma.

Tumor-specific mutations can result in immunogenic neoantigens, both of which have been correlated with responsiveness to immune checkpoint inhibitors in highly mutagenic cancers. However, early results of single-agent checkpoint inhibitors in multiple myeloma (MM) have been underwhelming. Therefore, we sought to understand the relationship between mutation and neoantigen landscape of MM patients and responsiveness to therapies. Somatic mutation burden, neoantigen load, and response to therapy were determined using interim data from the MMRF CoMMpass study (NCT01454297) on 664 MM patients. In this population, the mean somatic and missense mutation loads were 405.84(s=608.55) and 63.90(s=95.88) mutations per patient, respectively. There was a positive linear relationship between mutation and neoantigen burdens (R2=0.862). The average predicted neoantigen load was 23.52(s=52.14) neoantigens with an average of 9.40(s=26.97) expressed neoantigens. Survival analysis revealed significantly shorter progression-free survival (PFS) in patients with greater than average somatic missense mutation load (N=163, 0.493 vs 0.726 2-year PFS, P=0.0023) and predicted expressed neoantigen load (N=214, 0.555 vs 0.729 2-year PFS, P=0.0028). This pattern is maintained when stratified by disease stage and cytogenetic abnormalities. Therefore, high mutation and neoantigen load are clinically relevant risk factors that negatively impact survival of MM patients under current standards of care.

Pomalidomide, bortezomib and low-dose dexamethasone in lenalidomide-refractory and proteasome inhibitor-exposed myeloma.

This phase 1 dose-escalation study evaluated pomalidomide, bortezomib (subcutaneous (SC) or intravenous (IV)) and low-dose dexamethasone (LoDEX) in lenalidomide-refractory and proteasome inhibitor-exposed relapsed or relapsed and refractory multiple myeloma (RRMM). In 21-day cycles, patients received pomalidomide (1-4 mg days 1-14), bortezomib (1-1.3 mg/m2 days 1, 4, 8 and 11 for cycles 1-8; days 1 and 8 for cycle ⩾9) and LoDEX. Primary endpoint was to determine the maximum tolerated dose (MTD). Thirty-four patients enrolled: 12 during escalation, 10 in the MTD IV bortezomib cohort and 12 in the MTD SC bortezomib cohort. Patients received a median of 2 prior lines of therapy; 97% bortezomib exposed. With no dose-limiting toxicities, MTD was defined as the maximum planned dose: pomalidomide 4 mg, bortezomib 1.3 mg/m2 and LoDEX. All patients discontinued treatment by data cutoff (2 April 2015). The most common grade 3/4 treatment-emergent adverse events were neutropenia (44%) and thrombocytopenia (26%), which occurred more frequently with IV than SC bortezomib. No grade 3/4 peripheral neuropathy or deep vein thrombosis was reported. Overall response rate was 65%. Median duration of response was 7.4 months. Pomalidomide, bortezomib and LoDEX was well tolerated and effective in lenalidomide-refractory and bortezomib-exposed patients with RRMM.

Clinical Implications of Complex Pharmacokinetics for Daratumumab Dose Regimen in Patients With Relapsed/Refractory Multiple Myeloma.

New therapeutic strategies are urgently needed to improve clinical outcomes in patients with multiple myeloma (MM). Daratumumab is a first-in-class, CD38 human immunoglobulin G1κ monoclonal antibody approved for treatment of relapsed or refractory MM. Identification of an appropriate dose regimen for daratumumab is challenging due to its target-mediated drug disposition, leading to time- and concentration-dependent pharmacokinetics. We describe a thorough evaluation of the recommended dose regimen for daratumumab in patients with relapsed or refractory MM.

Targeting 6-phosphogluconate dehydrogenase in the oxidative PPP sensitizes leukemia cells to antimalarial agent dihydroartemisinin.

The oxidative pentose phosphate pathway (PPP) is crucial for cancer cell metabolism and tumor growth. We recently reported that targeting a key oxidative PPP enzyme, 6-phosphogluconate dehydrogenase (6PGD), using our novel small-molecule 6PGD inhibitors Physcion and its derivative S3, shows anticancer effects. Notably, humans with genetic deficiency of either 6PGD or another oxidative PPP enzyme, glucose-6-phosphate dehydrogenase, exhibit non-immune hemolytic anemia upon exposure to aspirin and various antimalarial drugs. Inspired by these clinical observations, we examined the anticancer potential of combined treatment with 6PGD inhibitors and antimalarial drugs. We found that stable knockdown of 6PGD sensitizes leukemia cells to antimalarial agent dihydroartemisinin (DHA). Combined treatment with DHA and Physcion activates AMP-activated protein kinase, leading to synergistic inhibition of human leukemia cell viability. Moreover, our combined therapy synergistically attenuates tumor growth in xenograft nude mice injected with human K562 leukemia cells and cell viability of primary leukemia cells from human patients, but shows minimal toxicity to normal hematopoietic cells in mice as well as red blood cells and mononucleocytes from healthy human donors. Our findings reveal the potential for combined therapy using optimized doses of Physcion and DHA as a novel antileukemia treatment without inducing hemolysis.

Daratumumab: monoclonal antibody therapy to treat multiple myeloma.

Daratumumab (Darzalex[TM]) is a human monoclonal antibody (MAb) that targets CD38; a surface protein highly expressed across multiple myeloma (MM) cells. Preclinical studies have shown daratumumab induces MM cell death through several mechanisms, including complement-dependent cytotoxicity (CDC) antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP), apoptosis upon secondary crosslinking and immunomodulatory effects via a decrease in immune suppressive cells. Daratumumab has a favorable toxicity profile and encouraging clinical activity as a single agent and in combination with lenalidomide in heavily pretreated, relapsed patients in whom other novel agents (such as bortezomib, thalidomide and lenalidomide) and stem cell transplant have already failed. Given the encouraging efficacy and acceptable safety profile, daratumumab has emerged as a novel treatment option for MM both as a monotherapy and in combination with conventional and novel anti-MM agents. This review will focus on preclinical pharmacology, pharmacokinetics, safety and clinical development of daratumumab in MM.

Targeting glutamine metabolism in multiple myeloma enhances BIM binding to BCL-2 eliciting synthetic lethality to venetoclax.

Multiple myeloma (MM) is a plasma cell malignancy that is largely incurable due to development of resistance to therapy-elicited cell death. Nutrients are intricately connected to maintenance of cellular viability in part by inhibition of apoptosis. We were interested to determine if examination of metabolic regulation of BCL-2 proteins may provide insight on alternative routes to engage apoptosis. MM cells are reliant on glucose and glutamine and withdrawal of either nutrient is associated with varying levels of apoptosis. We and others have demonstrated that glucose maintains levels of key resistance-promoting BCL-2 family member, myeloid cell leukemic factor 1 (MCL-1). Cells continuing to survive in the absence of glucose or glutamine were found to maintain expression of MCL-1 but importantly induce pro-apoptotic BIM expression. One potential mechanism for continued survival despite induction of BIM could be due to binding and sequestration of BIM to alternate pro-survival BCL-2 members. Our investigation revealed that cells surviving glutamine withdrawal in particular, enhance expression and binding of BIM to BCL-2, consequently sensitizing these cells to the BH3 mimetic venetoclax. Glutamine deprivation-driven sensitization to venetoclax can be reversed by metabolic supplementation with TCA cycle intermediate α-ketoglutarate. Inhibition of glucose metabolism with the GLUT4 inhibitor ritonavir elicits variable cytotoxicity in MM that is marginally enhanced with venetoclax treatment, however, targeting glutamine metabolism with 6-diazo-5-oxo-l-norleucine uniformly sensitized MM cell lines and relapse/refractory patient samples to venetoclax. Our studies reveal a potent therapeutic strategy of metabolically driven synthetic lethality involving targeting glutamine metabolism for sensitization to venetoclax in MM.

Monoclonal antibodies in the treatment of multiple myeloma: current status and future perspectives.

The treatment landscape for patients with multiple myeloma (MM) is constantly evolving. Over the past decade, the introduction of novel agents such as proteasome inhibitors and immunomodulatory drugs has led to notable changes in therapeutic strategy, and improvements in survival, yet MM remains incurable in the vast majority of cases. More recently, a targeted approach to MM treatment has emerged, using monoclonal antibodies (mAbs) to target antigens expressed on the surface of MM cells. MAbs tested to date kill MM cells via the host's immune system and/or by promoting apoptosis, and appear to have generally improved tolerability compared with currently available treatments. Due to their distinct mode of action, mAbs are promising both for patients who have exhausted current regimens, and as part of first-line treatments in newly diagnosed patients. This review examines the recent developments in mAb-based therapy for MM, primarily focused on those agents in ongoing clinical testing.

Dexamethasone treatment promotes Bcl-2 dependence in multiple myeloma resulting in sensitivity to venetoclax.

Venetoclax (ABT-199), a specific inhibitor of the anti-apoptotic protein Bcl-2, is currently in phase I clinical trials for multiple myeloma. The results suggest that venetoclax is only active in a small cohort of patients therefore we wanted to determine its efficacy when used in combination. Combining venetoclax with melphalan or carfilzomib produced additive or better cell death in four of the five cell lines tested. The most striking results were seen with dexamethasone (Dex). Co-treatment of human myeloma cell lines and primary patient samples, with Dex and venetoclax, significantly increased cell death over venetoclax alone in four of the five cell lines, and in all patient samples tested. The mechanism by which this occurs is an increase in the expression of both Bcl-2 and Bim upon addition of Dex. This results in alterations in Bim binding to anti-apoptotic proteins. Dex shifts Bim binding towards Bcl-2 resulting in increased sensitivity to venetoclax. These data suggest that knowledge of drug-induced alterations of Bim-binding patterns may help inform better combination drug regimens. Furthermore, the data indicate combining this novel therapeutic with Dex could be an effective therapy for a broader range of patients than would be predicted by single-agent activity.

Low expression of pro-apoptotic Bcl-2 family proteins sets the apoptotic threshold in Waldenström macroglobulinemia.

Waldenström macroglobulinemia (WM) is a proliferative disorder of IgM-secreting, lymphoplasmacytoid cells that inhabit the lymph nodes and bone marrow. The disease carries a high prevalence of activating mutations in MyD88 (91%) and CXCR4 (28%). Because signaling through these pathways leads to Bcl-xL induction, we examined Bcl-2 family expression in WM patients and cell lines. Unlike other B-lymphocyte-derived malignancies, which become dependent on expression of anti-apoptotic proteins to counter expression of pro-apoptotic proteins, WM samples expressed both pro- and anti-apoptotic Bcl-2 proteins at low levels similar to their normal B-cell and plasma cell counterparts. Three WM cell lines expressed pro-apoptotic Bcl-2 family members Bim or Bax and Bak at low levels, which determined their sensitivity to inducers of intrinsic apoptosis. In two cell lines, miR-155 upregulation, which is common in WM, was responsible for the inhibition of FOXO3a and Bim expression. Both antagonizing miR-155 to induce Bim and proteasome inhibition increased the sensitivity to ABT-737 in these lines indicating a lowering of the apoptotic threshold. In this manner, treatments that increase pro-apoptotic protein expression increase the efficacy of agents treated in combination in addition to direct killing.

Second transplants for multiple myeloma relapsing after a previous autotransplant-reduced-intensity allogeneic vs autologous transplantation.

There is no standard therapy for multiple myeloma relapsing after an autotransplant. We compared the outcomes of a second autotransplant (N=137) with those of an allotransplant (N=152) after non-myeloablative or reduced-intensity conditioning (NST/RIC) in 289 subjects reported to the CIBMTR from 1995 to 2008. NST/RIC recipients were younger (median age 53 vs 56 years; P<0.001) and had a shorter time to progression after their first autotransplant. Non-relapse mortality at 1-year post transplant was higher in the NST/RIC cohort, 13% (95% confidence interval (CI), 8-19) vs 2% (95% CI, 1-5, P0.001). Three-year PFS and OS for the NST/RIC cohort were 6% (95% CI, 3-10%) and 20% (95% CI, 14-27%). Similar outcomes for the autotransplant cohort were 12% (95% CI, 7-19%, P=0.038) and 46% (95% CI, 37-55%, P=0.001). In multivariate analyses, risk of death was higher in NST/RIC recipients (hazard ratio (HR) 2.38 (95% CI, 1.79-3.16), P<0.001), those with Karnofsky performance score<90 (HR 1.96 (95% CI, 1.47-2.62), P<0.001) and transplant before 2004 (HR 1.77 (95% CI, 1.34-2.35) P0.001). In conclusion, NST/RIC was associated with higher TRM and lower survival than an autotransplant. As disease status was not available for most allotransplant recipients, it is not possible to determine which type of transplant is superior after autotransplant failure.

New drugs and novel mechanisms of action in multiple myeloma in 2013: a report from the International Myeloma Working Group (IMWG).

Treatment in medical oncology is gradually shifting from the use of nonspecific chemotherapeutic agents toward an era of novel targeted therapy in which drugs and their combinations target specific aspects of the biology of tumor cells. Multiple myeloma (MM) has become one of the best examples in this regard, reflected in the identification of new pathogenic mechanisms, together with the development of novel drugs that are being explored from the preclinical setting to the early phases of clinical development. We review the biological rationale for the use of the most important new agents for treating MM and summarize their clinical activity in an increasingly busy field. First, we discuss data from already approved and active agents (including second- and third-generation proteasome inhibitors (PIs), immunomodulatory agents and alkylators). Next, we focus on agents with novel mechanisms of action, such as monoclonal antibodies (MoAbs), cell cycle-specific drugs, deacetylase inhibitors, agents acting on the unfolded protein response, signaling transduction pathway inhibitors and kinase inhibitors. Among this plethora of new agents or mechanisms, some are specially promising: anti-CD38 MoAb, such as daratumumab, are the first antibodies with clinical activity as single agents in MM. Moreover, the kinesin spindle protein inhibitor Arry-520 is effective in monotherapy as well as in combination with dexamethasone in heavily pretreated patients. Immunotherapy against MM is also being explored, and probably the most attractive example of this approach is the combination of the anti-CS1 MoAb elotuzumab with lenalidomide and dexamethasone, which has produced exciting results in the relapsed/refractory setting.

Favorable impact of pre-transplant ATG on outcomes of reduced-intensity hematopoietic cell transplants from partially mismatched unrelated donors.

Reduced-intensity conditioning (RIC) permits allogeneic hematopoietic progenitor cell transplantation in patients who would not be considered candidates for transplantation using a myeloablative preparative regimen because of age, comorbidities or prior therapy. In the setting of myeloablative transplantation, use of antithymocyte globulin (ATG) can reduce the risk of GVHD without negatively affecting transplant outcomes; however, limited data exist on the impact of ATG in the setting of RIC, particularly when there is HLA-mismatch. We performed a retrospective analysis of 85 patients who received unrelated donor transplants at our institution for hematologic malignancies following conditioning with fludarabine and melphalan (FluMel), with or without rabbit ATG (6 mg/kg). ATG was targeted to patients receiving HLA-mismatched grafts. With a median follow-up of 36 months, those receiving ATG and a mismatched graft had similar rates of acute and chronic GVHD, relapse, and similar OS compared with those receiving HLA-matched grafts without ATG. In a multivariate analysis, HLA-mismatched donor was not associated with a decrement in OS. We conclude that this intermediate dose of ATG is effective in preventing severe GVHD in the setting of HLA-mismatch, without undue compromise of the graft versus tumor effects on which RIC transplants depend.

Consolidation and maintenance therapy with lenalidomide, bortezomib and dexamethasone (RVD) in high-risk myeloma patients.

Prior studies have shown that myeloma patients exhibiting either genetically defined high-risk disease or plasma cell leukemia have a poor outcome with a median overall survival (OS) of ≤3 years. Results of IFM 2005-01 and 02 suggest that relatively limited bortezomib-containing induction regimens did not produce a major survival benefit among these patients. However, results of recent studies suggest that combination therapy may benefit these patients when given early and again later in the treatment. We evaluated a combination maintenance/consolidation regimen (RVD) following autologous stem cell transplant (ASCT) for high-risk patients to evaluate the impact of this approach on outcome. Following initiation of RVD maintenance, 51% of patients achieved stringent complete response (sCR), with 96% achieving at least VGPR as best response. Median progression free survival (PFS) for all patients is 32 months with a 3-year OS of 93%. The regimen was well tolerated with no grade 3/4 neuropathy. Early ASCT followed by RVD maintenance is a promising strategy for high-risk myeloma patients and delivered excellent response rates, and promising PFS and OS.

Association of response endpoints with survival outcomes in multiple myeloma.

Since the introduction of the proteasome inhibitor bortezomib and the immunomodulatory drugs (IMiDs) thalidomide and lenalidomide, more patients with multiple myeloma are achieving deep, durable responses and disease control, and are living longer. These improvements have afforded more robust analyses of the relationship between response and survival. Generally, these studies have demonstrated that improvements in the quality of response across all stages of treatment are associated with better disease control and longer survival. Thus, achievement of maximal response should be strongly considered, particularly in the frontline setting, but must also be balanced with tolerability, quality of life and patient preferences. In select patients, achievement of a lesser response may be adequate to prolong survival, and attempts to treat these patients to a deeper response may place them at unnecessary risk without significant benefit. Maintenance therapy has been shown to improve the quality of response and disease control and, in some studies, survival. Studies support maintenance therapy for high-risk patients as a standard of care, and there are emerging data supporting maintenance therapy in standard-risk patients to improve progression-free and possibly overall survival. Multidrug regimens combining a proteasome inhibitor and an IMiD have shown exceptional response outcomes with acceptable increases in toxicity in both the frontline and salvage settings, and are becoming a standard treatment approach. Moving forward, the use of immunophenotypic and molecular response criteria will be essential in better understanding the impact of highly active and continuous treatment regimens across myeloma patient populations. Future translational studies will help to develop antimyeloma agents to their fullest potential. The introduction of novel targeted therapies, including the IMiD pomalidomide and the proteasome inhibitors carfilzomib and ixazomib (MLN9708), will provide greater options to individualize treatment and help patients achieve a clinically meaningful response.

Treatment outcomes in patients with relapsed and refractory multiple myeloma and high-risk cytogenetics receiving single-agent carfilzomib in the PX-171-003-A1 study.

Several cytogenetic abnormalities are associated with poor outcomes in multiple myeloma (MM). We prospectively analyzed the impact of cytogenetic abnormalities on outcomes during the phase 2 PX-171-003-A1 study of single-agent carfilzomib for relapsed and refractory MM. In the response-evaluable population (257/266), fluorescence in situ hybridization (FISH)/conventional cytogenetic profiles were available for 229 patients; 62 (27.1%) had high-risk cytogenetics--del 17p13, t(4;14) or t(14;16) by interphase FISH or deletion 13 or hypodiploidy by metaphase cytogenetics--and 167 (72.9%) had standard-risk profiles. Generally, baseline characteristics were similar between the subgroups, but International Staging System stage III disease was more common in high- vs standard-risk patients (41.9% vs 27.5%) as was Eastern Cooperative Oncology Group performance status 1/2 (85.5% vs 68.3%). Overall response was comparable between the subgroups (25.8% vs 24.6%, respectively; P=0.85), while time-to-event end points showed a trend of shorter duration in high-risk patients, including median duration of response (5.6 months (95% confidence interval (CI) 3.7-7.8) vs 8.3 months (95% CI 5.6-12.3)) and overall survival (9.3 (95% CI 6.5-13.0) vs 19.0 months (95% CI 15.4-NE); P=0.0003). Taken together, these findings demonstrate that single-agent carfilzomib is efficacious and has the potential to at least partially overcome the impact of high-risk cytogenetics in heavily pre-treated patients with MM.