Clinical, genomic, and imaging predictors of myeloma progression from asymptomatic monoclonal gammopathies (SWOG S0120).

All cases of clinical myeloma (CMM) are preceded by an asymptomatic monoclonal gammopathy (AMG), classified as either monoclonal gammopathy of undetermined significance (MGUS) or asymptomatic multiple myeloma (AMM). We analyzed data from AMG patients (n = 331) enrolled in a prospective, observational clinical trial (S0120). Baseline data from clinical variables, gene expression profiles (GEP) of purified tumor cells, and findings of magnetic resonance imaging (MRI) were correlated with the risk of progression to CMM requiring therapy. GEP of purified tumor cells revealed that all molecular subtypes of CMM are also represented in the AMG phase. An increased risk score (>-0.26) (based on a 70-gene signature, GEP70) was an independent predictor of the risk of progression to CMM. Combination of elevated serum free light chain, M-spike, and GEP70 risk score identified a subset with high risk (67% at 2 years) of progression to CMM requiring therapy. Importantly, absence of these factors in AMM patients predicted low risk similar to MGUS. Detection of multiple (>1) focal lesions by MRI also conferred an increased risk of progression. These data demonstrate that signatures associated with high-risk CMM impact disease risk and support inclusion of genomic analysis in the clinical management of AMGs.

Jumping translocations of 1q12 in multiple myeloma: a novel mechanism for deletion of 17p in cytogenetically defined high-risk disease.

Multiple myeloma (MM) is a B-cell malignancy driven in part by increasing copy number alterations (CNAs) during disease progression. Prognostically significant CNAs accumulate during clonal evolution and include gains of 1q21 and deletions of 17p, among others. Unfortunately, the mechanisms underlying the accumulation of CNAs and resulting subclonal heterogeneity in high-risk MM are poorly understood. To investigate the impact of jumping translocations of 1q12 (JT1q12) on receptor chromosomes (RCs) and subsequent clonal evolution, we analyzed specimens from 86 patients selected for unbalanced 1q12 aberrations by G-banding. Utilizing spectral karyotyping and locus-specific fluorescence in situ hybridization, we identified 10 patients with unexpected focal amplifications of an RC that subsequently translocated as part of a sequential JT1q12 to one or more additional RCs. Four patients exhibited amplification and translocation of 8q24 (MYC), 3 showed amplification of 16q11, and 1 each displayed amplification of 18q21.3 (BCL2), 18q23, or 4p16 (FGFR3). Unexpectedly, in 6 of 14 patients with the combination of the t(4;14) and deletion of 17p, we identified the loss of 17p as resulting from a JT1q12. Here, we provide evidence that the JT1q12 is a mechanism for the simultaneous gain of 1q21 and deletion of 17p in cytogenetically defined high-risk disease.

Prognostic implications of serial 18-fluoro-deoxyglucose emission tomography in multiple myeloma treated with total therapy 3.

Prognostic implications of 3 imaging tools, metastatic bone survey, magnetic resonance imaging, and positron emission tomography (PET), were evaluated in 2 consecutive Total Therapy 3 trials for newly diagnosed myeloma. Data including PET at baseline and on day 7 of induction as well as standard prognostic factors were available in 302 patients of whom 277 also had gene expression profiling (GEP)-derived risk information. According to multivariate analysis, more than 3 focal lesions on day 7 imparted inferior overall survival and progression-free survival, overall and in the subset with GEP-risk data. GEP high-risk designation retained independent significance for all 3 end points examined. Thus, the presence of > 3 focal lesions on day 7 PET follow-up may be exploited toward early therapy change, especially for the 15% of patients with GEP-defined high-risk disease with a median overall survival expectation of 2 years. This trial was registered at www.clinicaltrials.gov as #NCT00081939 and # NCT00572169.

Risk factors for MDS and acute leukemia following total therapy 2 and 3 for multiple myeloma.

Lenalidomide has been linked to myelodysplastic syndrome (MDS) after autotransplants for myeloma. Total therapy trials (TT; TT2(-/+) thalidomide) and TT3 (TT3a with bortezomib, thalidomide; TT3b with additional lenalidomide) offered the opportunity to examine the contribution of these immune-modulatory agents to MDS-associated cytogenetic abnormalities (MDS-CA) and clinical MDS or acute leukemia ("clinical MDS/AL"). Of 1080 patients with serial cytogenetic studies, MDS-CA occurred in 11% and clinical MDS/AL in 3%. Risk features of MDS-CA included TT3b, age ≥65 years, male gender, levels of ß-2-microglobulin >5.5 mg/L, and multiple myeloma relapse. Clinical MDS/AL occurred less frequently in the control arm of TT2 and more often with TT3a and TT3b. Since MDS-CA often antedated clinical disease, periodic cytogenetic monitoring is recommended. Larger CD34 quantities should be collected upfront as the risk of MDS could be reduced by applying higher CD34 doses with transplant. Thus, treatment, host, and myeloma features could be linked to MDS development after therapy for this malignancy. This trial was registered at www.clinicaltrials.gov: TT3A: NCT00081939, TT3B: NCT00572169.

Second malignancies in total therapy 2 and 3 for newly diagnosed multiple myeloma: influence of thalidomide and lenalidomide during maintenance.

Thalidomide and lenalidomide constitute an important part of effective myeloma therapy. Recent data from the Intergroup Francophone du Myélome, Cancer and Leukemia Group B, and Gruppo Italiano Malattie Ematologiche dell Adulto MM-015 trials suggest that lenalidomide maintenance therapy is associated with a higher incidence of second primary malignancies (SPMs), including both hematologic and solid malignancies. In the present study, we analyzed data from the Total Therapy 2 (TT2) trial, along with the 2 Total Therapy 3 (TT3) trials. TT2 patients were assigned randomly to either a control group (no thalidomide) or to the experimental group (thalidomide during induction, between transplantations, and during consolidation and maintenance). The 2 TT3 trials used thalidomide and bortezomib during induction, before and in consolidation after tandem melphalan-based transplantation; TT3A applied VTD (bortezomib, thalidomide, dexamethasone) in the first year of maintenance and TD for 2 more years, whereas TT3B used VRD (bortezomib, lenalidomide, dexamethasone) maintenance for 3 years. The cumulative incidence of SPMs did not differ significantly among the TT trial components when measured from enrollment (P = .78) or from initiation of maintenance (P = .82). However, a pairwise comparison of the TT2 arms suggested a lower incidence of hematologic SPMs in the thalidomide maintenance arm (hazard ratio = 0.38; P = .09). These trials are registered at www.clinicaltrials.gov as NCT00573391 (TT2), NCT00081939 (TT3A), and NCT00572169 (TT3B).

Hepatitis B reactivation in patients with multiple myeloma and isolated positive hepatitis B core antibody: a call for greater cognizance.

Hepatitis B virus (HBV) reactivation is a well-established complication of severe immunosuppression in patients with hematologic malignancy and positive hepatitis B surface antigen (HBsAg). Patients who receive high-dose chemotherapy, corticosteroids, rituximab, or have a bone marrow transplant are particularly at increased risk for HBV reactivation. However, limited information is available in the literature regarding HBV reactivation in patients with isolated anti-HBc, particularly in the setting of multiple myeloma (MM). We report two cases of HBV reactivation in MM patients with isolated anti-HBc positive with a rather atypical presentation. In conclusion, our cases highlight that clinicians need to be cognizant about this potentially fatal but preventable complication of chemotherapy and immunosuppression.

Pharmacogenomics of bortezomib test-dosing identifies hyperexpression of proteasome genes, especially PSMD4, as novel high-risk feature in myeloma treated with Total Therapy 3.

Gene expression profiling (GEP) of purified plasma cells 48 hours after thalidomide and dexamethasone test doses showed these agents' mechanisms of action and provided prognostic information for untreated myeloma patients on Total Therapy 2 (TT2). Bortezomib was added in Total Therapy 3 (TT3), and 48 hours after bortezomib GEP analysis identified 80 highly survival-discriminatory genes in a training set of 142 TT3A patients that were validated in 128 patients receiving TT3B. The 80-gene GEP model (GEP80) also distinguished outcomes when applied at baseline in both TT3 and TT2 protocols. In context of our validated 70-gene model (GEP70), the GEP80 model identified 9% of patients with a grave prognosis among those with GEP70-defined low-risk disease and 41% of patients with favorable prognosis among those with GEP70-defined high-risk disease. PMSD4 was 1 of 3 genes common to both models. Residing on chromosome 1q21, PSMD4 expression is highly sensitive to copy number. Both higher PSMD4 expression levels and higher 1q21 copy numbers affected clinical outcome adversely. GEP80 baseline-defined high risk, high lactate dehydrogenase, and low albumin were the only independent adverse variables surviving multivariate survival model. We are investigating whether second-generation proteasome inhibitors (eg, carfilzomib) can overcome resistance associated with high PSMD4 levels.

Standard and novel imaging methods for multiple myeloma: correlates with prognostic laboratory variables including gene expression profiling data.

Multiple myeloma causes major morbidity resulting from osteolytic lesions that can be detected by metastatic bone surveys. Magnetic resonance imaging and positron emission tomography can detect bone marrow focal lesions long before development of osteolytic lesions. Using data from patients enrolled in Total Therapy 3 for newly diagnosed myeloma (n=303), we analyzed associations of these imaging techniques with baseline standard laboratory variables assessed before initiating treatment. Of 270 patients with complete imaging data, 245 also had gene expression profiling data. Osteolytic lesions detected on metastatic bone surveys correlated with focal lesions detected by magnetic resonance imaging and positron emission tomography, although, in two-way comparisons, focal lesion counts based on both magnetic resonance imaging and positron emission tomography tended to be greater than those based on metastatic bone survey. Higher numbers of focal lesions detected by magnetic resonance imaging and positron emission tomography were positively linked to high serum concentrations of C-reactive protein, gene-expression-profiling-defined high risk, and the proliferation molecular subgroup. Positron emission tomography focal lesion maximum standardized unit values were significantly correlated with gene-expression-profiling-defined high risk and higher numbers of focal lesions detected by positron emission tomography. Interestingly, four genes associated with high-risk disease (related to cell cycle and metabolism) were linked to counts of focal lesions detected by magnetic resonance imaging and positron emission tomography. Collectively, our results demonstrate significant associations of all three imaging techniques with tumor burden and, especially, disease aggressiveness captured by gene-expression-profiling-risk designation. (Clinicaltrials.gov identifier: NCT00081939).

Metronomic therapy is an effective salvage treatment for heavily pre-treated relapsed/refractory multiple myeloma.

Relapsed/refractory multiple myeloma represents a major challenge in multiple myeloma therapy. For patients with relapsed/refractory multiple myeloma, we developed a treatment schema of metronomically scheduled drug therapy. We identified 186 patients who had been treated with metronomic therapy between March 2004 and January 2012 with a median follow up of 24.2 months. Median age was 61 years (range 36-83). Median number of prior therapies was 14 (range 1-51). Median number of completed metronomic therapy cycles was 1 (range 1-5), while 45 of 186 (25%) received 2 or more cycles. Responses included complete remission in 11 of 186 patients (6%), very good partial remission in 12 of 186 (7%), partial remission in 65 of 179 (36%), and minimal response in 29 of 186 (16%), for an overall response rate of 63% (117 of 186). Median overall survival and progression-free survival were 11.2 and 3.6 months, respectively. Hematologic toxicity grading was problematic as 146 of 186 (78%) of patients presented with at least grade 2 thrombocytopenia within 90 days prior to starting metronomic therapy. Grade 4 leukopenia, anemia, and/or thrombocytopenia following metronomic therapy occurred in 108 of 186 (58%), 12 of 186 (6%), and 147 of 186 (79%) patients, respectively. Incidence of grade 3-4 neutropenic fever was 4 of 186 (2%). Most patients (177 of 186, 95%) were treated in an outpatient unit and secondary admissions due to regimen-related toxicity occurred in 37 of 186 (20%). Treatment-related mortality was evident in 2 of 186 (1%). In conclusion, metronomic therapy is an effective late salvage treatment in relapsed/refractory multiple myeloma, with a high overall response rate and a favorable toxicity profile.

High-risk myeloma is associated with global elevation of miRNAs and overexpression of EIF2C2/AGO2.

MicroRNAs (miRNAs) are noncoding RNAs that regulate global gene expression. miRNAs often act synergistically to repress target genes, and their dysregulation can contribute to the initiation and progression of a variety of cancers. The clinical relationship between global expression of miRNA and mRNA in cancer has not been studied in detail. We used whole-genome microarray analyses of CD138-enriched plasma cells from 52 newly diagnosed cases of multiple myeloma to correlate miRNA expression profiles with a validated mRNA-based risk stratification score, proliferation index, and predefined gene sets. In stark contrast to mRNAs, we discovered that all tested miRNAs were significantly up-regulated in high-risk disease as defined by a validated 70-gene risk score (P < 0.01) and proliferation index (P < 0.05). Increased expression of EIF2C2/AGO2, a master regulator of the maturation and function of miRNAs and a component of the 70-gene mRNA risk model, is driven by DNA copy number gains in MM. Silencing of AGO2 dramatically decreased viability in MM cell lines. Genome-wide elevated expression of miRNAs in high-risk MM may be secondary to deregulation of AGO2 and the enzyme complexes that regulate miRNA maturation and function.

Superior results of Total Therapy 3 (2003-33) in gene expression profiling-defined low-risk multiple myeloma confirmed in subsequent trial 2006-66 with VRD maintenance.

The Total Therapy 3 trial 2003-33 enrolled 303 newly diagnosed multiple myeloma patients and was noted to provide superior clinical outcomes compared with predecessor trial Total Therapy 2, especially in gene expression profiling (GEP)-defined low-risk disease. We report here on the results of successor trial 2006-66 with 177 patients, using bortezomib, lenalidomide, and dexamethasone maintenance for 3 years versus bortezomib, thalidomide, and dexamethasone in year 1 and thalidomide/dexamethasone in years 2 and 3 in the 2003-33 protocol. Overall survival (OS) and event-free survival (EFS) plots were super-imposable for the 2 trials, as were onset of complete response and complete response duration (CRD), regardless of GEP risk. GEP-defined high-risk designation, pertinent to 17% of patients, imparted inferior OS, EFS, and CRD in both protocols and, on multivariate analysis, was the sole adverse feature affecting OS, EFS, and CRD. Mathematical modeling of CRD in low-risk myeloma predicted a 55% cure fraction (P < .001). Despite more rapid onset and higher rate of CR than in other molecular subgroups, CRD was inferior in CCND1 without CD20 myeloma, resembling outcomes in MAF/MAFB and proliferation entities. The robustness of the GEP risk model should be exploited in clinical trials aimed at improving the notoriously poor outcome in high-risk disease.

Total Therapy 3 for multiple myeloma: prognostic implications of cumulative dosing and premature discontinuation of VTD maintenance components, bortezomib, thalidomide, and dexamethasone, relevant to all phases of therapy.

The impact of cumulative dosing and premature drug discontinuation (PMDD) of bortezomib (V), thalidomide (T), and dexamethasone (D) on overall survival (OS), event-free survival (EFS), time to next therapy, and post-relapse survival in Total Therapy 3 were examined, using time-dependent methodology, relevant to induction, peritransplantation, consolidation, and maintenance phases. Univariately, OS and EFS were longer in case higher doses were used of all agents during induction, consolidation (except T), and maintenance (except V and T). The favorable OS and EFS impact of D induction dosing provided the rationale for examining the expression of glucocorticoid receptor NR3C1, top-tertile levels of which significantly prolonged OS and EFS and rendered outcomes independent of D and T dosing, whereas T and D, but not V, dosing was critical to outcome improvement in the bottom-tertile NR3C1 setting. PMDD of V was an independent highly adverse feature for OS (hazard ratio = 6.44; P < .001), whereas PMDD of both T and D independently imparted shorter time to next therapy. The absence of adverse effects on postrelapse survival of dosing of any VTD components and indeed a benefit from V supports the use up-front of all active agents in a dose-dense and dose-intense fashion, as practiced in Total Therapy 3, toward maximizing myeloma survival.

Extramedullary disease portends poor prognosis in multiple myeloma and is over-represented in high-risk disease even in the era of novel agents.

BACKGROUND: Extramedullary disease is an uncommon manifestation in multiple myeloma and can either accompany newly diagnosed disease or develop with disease progression or relapse. We evaluated the impact of this disease feature on patients' outcome in the context of novel agents. DESIGN AND METHODS: We analyzed clinical and biological features of extramedullary disease in 936 patients with multiple myeloma enrolled in Total Therapy protocols, 240 patients in non-Total Therapy protocols, and 789 non-protocol patients, all of whom had baseline positron emission tomography scans to document extramedullary disease at diagnosis and its subsequent development at the time of disease progression or relapse. RESULTS: The most common sites for extramedullary disease at diagnosis were skin and soft tissue whereas liver involvement was the striking feature in extramedullary disease at disease relapse or progression. Regardless of therapy, extramedullary disease was associated with shorter progression-free and overall survival, as well as the presence of anemia, thrombocytopenia, elevated serum lactate dehydrogenase, cytogenetic abnormalities, and high-risk features in 70-and 80-gene risk models in univariate analysis. Multivariate analysis with logistic regression revealed that this disease feature was more prevalent in patients with an elevated centrosome index, as determined by gene expression profiling, as well as in myeloma molecular subtypes that are more prone to relapse. These include the MF subtype (also called the "MAF" subtype, associated with over-expression of the MAF gene seen with chromosome translocation 14;16 or 14;20) and the PR subtype (also called the "Proliferation" subtype, associated with overexpression of pro-proliferative genes). CONCLUSIONS: These data show that extramedullary disease is more prevalent in genomically defined high-risk multiple myeloma and is associated with shorter progression-free survival and overall survival, even in the era of novel agents. All clinical trials included in the analyses were registered with www.clinicaltrials.gov (NCT00083551, NCT00083876, NCT00081939, NCT00572169, NCT00644228,NCT00002548,NCT00734877).

Gene expression profiling of plasma cells at myeloma relapse from tandem transplantation trial Total Therapy 2 predicts subsequent survival.

We report on prognostic implications for post-relapse survival (PRS) of a gene expression profiling (GEP)-defined risk score at relapse available in 120 myeloma patients previously enrolled in tandem transplantation trial Total Therapy 2. Among the 71 patients with additional GEP baseline information, 3-year PRS was 71% in 40 patients with low risk present both at baseline and relapse contrasting with only 17% in 28 patients with high risk at relapse, 12 of whom with baseline low-risk status fared better than the remainder (P = .08). On multivariate analysis of relapse parameters available in 104 patients, high risk conferred short PRS (hazard ratio = 4.00, P < .001, R(2) = 33%), whereas relapse hyperdiploidy predicted long PRS (hazard ratio = 0.37, P = .022, cumulative R(2) = 41%). In case the initial partial response lasted less than 2 years, relapse low-risk identified 26 patients with superior 3-year PRS of 61% versus 9% among 32 with relapse high-risk (P < .001). Based on its PRS predictive power, GEP analysis should be an integral part of new agent trials in search of better therapy for high-risk myeloma.

Complete remission in multiple myeloma examined as time-dependent variable in terms of both onset and duration in Total Therapy protocols.

Landmark analyses are used to investigate the importance for survival of achieving complete response (CR), an important initial goal of myeloma therapy. With median times to CR in Total Therapy (TT) trials of approximately 1 year, this approach excludes a sizeable fraction of patients dying before such a landmark. To permit inclusion of all trial participants, we investigated the prognostic implications of both onset and duration of CR as time-dependent variables. Superseding the adverse effects of cytogenetic abnormalities and other standard prognostic parameters, both failure to achieve CR (non-CR) and, especially, loss of CR (los-CR) were independently associated with inferior survival in TT1, TT2, and TT3 protocols. In the context of gene array-defined risk, available in TT2 and TT3 subsets, both los-CR and non-CR terms were retained in the survival model as dominant adverse variables, stressing the prognostic importance of sustaining CR status, especially in high-risk disease.

Thalidomide arm of Total Therapy 2 improves complete remission duration and survival in myeloma patients with metaphase cytogenetic abnormalities.

Total Therapy 2 examined the clinical benefit of adding thalidomide up-front to a tandem transplant regimen for newly diagnosed patients with multiple myeloma. When initially reported with a median follow-up of 42 months, complete response rate and event-free survival were superior among the 323 patients randomized to thalidomide, whereas overall survival was indistinguishable from that of the 345 patients treated on the control arm. With further follow-up currently at a median of 72 months, survival plots segregated 5 years after initiation of therapy in favor of thalidomide (P = .09), reaching statistical significance for the one third of patients exhibiting cytogenetic abnormalities (CAs; P = .02), a well-recognized adverse prognostic feature. The duration of complete remission was also superior in the cohort presenting with CAs such that, at 7 years from onset of complete remission, 45% remained relapse-free as opposed to 20% on the control arm (P = .05). These observations were confirmed when examined by multivariate analysis demonstrating that thalidomide reduced the hazard of death by 41% among patients with CA-positive disease (P = .008). Because two thirds of patients without CAs have remained alive at 7 years, the presently emerging separation in favor of thalidomide may eventually reach statistical significance as well.

TP53 deletion is not an adverse feature in multiple myeloma treated with total therapy 3.

Contrary to Total Therapy (TT) 2 for multiple myeloma patients, FGFR3- translocation bore no adverse effects on outcome in TT3 with added bortezomib. Del TP53, another poor-risk feature in TT2 and present in 10% of 441 patients treated, was examined for its prognostic consequences in TT3. Not affecting rate or duration of complete response, TP53 haplo-insufficiency also did not compromise, in the 83% with genomically defined low-risk myeloma, survival or event-free survival. FGFR3+ and FGFR3- molecular subgroups fared worse in the presence of del TP53 when applying TT2 but not TT3. Thus, the prognostic implications of del TP53 were protocol-, genome-defined risk- and molecular subgroup-dependent.

Correction: Spectrum of Cerebrovascular Disease in Patients with Multiple Myeloma Undergoing Chemotherapy-Results of a Case Control Study.

[This corrects the article DOI: 10.1371/journal.pone.0166627.].

Assessment of Total Lesion Glycolysis by 18F FDG PET/CT Significantly Improves Prognostic Value of GEP and ISS in Myeloma.

Purpose: Fluorine-18 fluorodeoxyglucose positron emission tomography with CT attenuation correction (18F-FDG PET/CT) is useful in the detection and enumeration of focal lesions and in semiquantitative characterization of metabolic activity (glycolytic phenotype) by calculation of glucose uptake. Total lesion glycolysis (TLG) and metabolic tumor volume (MTV) have the potential to improve the value of this approach and enhance the prognostic value of disease burden measures. This study aims to determine whether TLG and MTV are associated with progression-free survival (PFS) and overall survival (OS), and whether they improve risk assessments such as International Staging System (ISS) stage and GEP70 risk.Experimental Design: 192 patients underwent whole body PET/CT in the Total Therapy 3A (TT3A) trial and were evaluated using three-dimensional region-of-interest analysis with TLG, MTV, and standard measurement parameters derived for all focal lesions with peak SUV above the background red marrow signal.Results: In multivariate analysis, baseline TLG > 620 g and MTV > 210 cm3 remained a significant factor of poor PFS and OS after adjusting for baseline myeloma variables. Combined with the GEP70 risk score, TLG > 205 g identifies a high-risk-behaving subgroup with poor expected survival. In addition, TLG > 205 g accurately divides ISS stage II patients into two subgroups with similar outcomes to ISS stage I and ISS stage III, respectively.Conclusions: TLG and MTV have significant survival implications at baseline and offer a more precise quantitation of the glycolytic phenotype of active disease. These measures can be assessed more readily than before using FDA-approved software and should be standardized and incorporated into clinical trials moving forward. Clin Cancer Res; 23(8); 1981-7. ©2016 AACR.