The genomic profiling of high-risk smoldering myeloma patients treated with an intensive strategy unveils potential markers of resistance and progression.

Smoldering multiple myeloma (SMM) precedes multiple myeloma (MM). The risk of progression of SMM patients is not uniform, thus different progression-risk models have been developed, although they are mainly based on clinical parameters. Recently, genomic predictors of progression have been defined for untreated SMM. However, the usefulness of such markers in the context of clinical trials evaluating upfront treatment in high-risk SMM (HR SMM) has not been explored yet, precluding the identification of baseline genomic alterations leading to drug resistance. For this reason, we carried out next-generation sequencing and fluorescent in-situ hybridization studies on 57 HR and ultra-high risk (UHR) SMM patients treated in the phase II GEM-CESAR clinical trial (NCT02415413). DIS3, FAM46C, and FGFR3 mutations, as well as t(4;14) and 1q alterations, were enriched in HR SMM. TRAF3 mutations were specifically associated with UHR SMM but identified cases with improved outcomes. Importantly, novel potential predictors of treatment resistance were identified: NRAS mutations and the co-occurrence of t(4;14) plus FGFR3 mutations were associated with an increased risk of biological progression. In conclusion, we have carried out for the first time a molecular characterization of HR SMM patients treated with an intensive regimen, identifying genomic predictors of poor outcomes in this setting.

Are we ready to look beyond plasma cells in assessing high-risk smoldering myeloma?

Sklavenitis-Pistofidis et al. report clinical and correlative results of a single-arm phase II trial of elotuzumab, lenalidomide, and dexamethasone in patients with high-risk smoldering myeloma. The authors explore the interactions between the genetics of the plasma cell clone and the immune microenvironment as potential biomarkers of treatment susceptibility and efficacy.

Genetic subtypes of smoldering multiple myeloma are associated with distinct pathogenic phenotypes and clinical outcomes.

Smoldering multiple myeloma (SMM) is a precursor condition of multiple myeloma (MM) with significant heterogeneity in disease progression. Existing clinical models of progression risk do not fully capture this heterogeneity. Here we integrate 42 genetic alterations from 214 SMM patients using unsupervised binary matrix factorization (BMF) clustering and identify six distinct genetic subtypes. These subtypes are differentially associated with established MM-related RNA signatures, oncogenic and immune transcriptional profiles, and evolving clinical biomarkers. Three genetic subtypes are associated with increased risk of progression to active MM in both the primary and validation cohorts, indicating they can be used to better predict high and low-risk patients within the currently used clinical risk stratification models.

Smoldering multiple myeloma - Past, present, and future.

Smoldering multiple myeloma (SMM) routinely precedes the development of multiple myeloma. While some patients experience aggressive disease, others have more indolent courses akin to those with monoclonal gammopathy of undetermined significance. Much effort has been made to understand the pathobiological basis of this heterogeneity. Scientific advancements have led to the emergence of various clinical and genomic markers of relevance, translating into evolution of disease definitions over time. More recently, the interest in manipulation of biological pathways has intensified in a bid to stall or halt disease progression. Studies with lenalidomide have exemplified the promise of early intervention, whereas numerous therapeutic approaches remain the subject of ongoing clinical investigation. This review summarizes the historic progress made in defining SMM as a distinct clinicopathologic entity, provides a critical appraisal of the evidence guiding risk assessment, and suggests a pragmatic approach to its modern-day management. Finally, an overview of developments on the horizon is also provided.

Plasma cells expression from smouldering myeloma to myeloma reveals the importance of the PRC2 complex, cell cycle progression, and the divergent evolutionary pathways within the different molecular subgroups.

Sequencing studies have shed some light on the pathogenesis of progression from smouldering multiple myeloma (SMM) and symptomatic multiple myeloma (MM). Given the scarcity of smouldering samples, little data are available to determine which translational programmes are dysregulated and whether the mechanisms of progression are uniform across the main molecular subgroups. In this work, we investigated 223 SMM and 1348 MM samples from the University of Arkansas for Medical Sciences (UAMS) for which we had gene expression profiling (GEP). Patients were analysed by TC-7 subgroup for gene expression changes between SMM and MM. Among the commonly dysregulated genes in each subgroup, PHF19 and EZH2 highlight the importance of the PRC2.1 complex. We show that subgroup specific differences exist even at the SMM stage of disease with different biological features driving progression within each TC molecular subgroup. These data suggest that MMSET SMM has already transformed, but that the other precursor diseases are distinct clinical entities from their symptomatic counterpart.

Gene Expression Analysis of the Bone Marrow Microenvironment Reveals Distinct Immunotypes in Smoldering Multiple Myeloma Associated to Progression to Symptomatic Disease.

Background: We previously reported algorithms based on clinical parameters and plasma cell characteristics to identify patients with smoldering multiple myeloma (SMM) with higher risk of progressing who could benefit from early treatment. In this work, we analyzed differences in the immune bone marrow (BM) microenvironment in SMM to better understand the role of immune surveillance in disease progression and to identify immune biomarkers associated to higher risk of progression. Methods: Gene expression analysis of BM cells from 28 patients with SMM, 22 patients with monoclonal gammopathy of undetermined significance (MGUS) and 22 patients with symptomatic MM was performed by using Nanostring Technology. Results: BM cells in SMM compared to both MGUS and symptomatic MM showed upregulation of genes encoding for key molecules in cytotoxicity. However, some of these cytotoxic molecules positively correlated with inhibitory immune checkpoints, which may impair the effector function of BM cytotoxic cells. Analysis of 28 patients with SMM revealed 4 distinct clusters based on immune composition and activation markers. Patients in cluster 2 showed a significant increase in expression of cytotoxic molecules but also inhibitory immune checkpoints compared to cluster 3, suggesting the presence of cytotoxic cells with an exhausted phenotype. Accordingly, patients in cluster 3 had a significantly longer progression free survival. Finally, individual gene expression analysis showed that higher expression of TNF superfamily members (TNF, TNFAIP3, TNFRSF14) was associated with shorter progression free survival. Conclusions: Our results suggest that exhausted cytotoxic cells are associated to high-risk patients with SMM. Biomarkers overexpressed in patients with this immune gene profile in combination with clinical parameters and PC characterization may be useful to identify SMM patients with higher risk of progression.

Whole-genome sequencing reveals progressive versus stable myeloma precursor conditions as two distinct entities.

Multiple myeloma (MM) is consistently preceded by precursor conditions recognized clinically as monoclonal gammopathy of undetermined significance (MGUS) or smoldering myeloma (SMM). We interrogate the whole genome sequence (WGS) profile of 18 MGUS and compare them with those from 14 SMMs and 80 MMs. We show that cases with a non-progressing, clinically stable myeloma precursor condition (n = 15) are characterized by later initiation in the patient's life and by the absence of myeloma defining genomic events including: chromothripsis, templated insertions, mutations in driver genes, aneuploidy, and canonical APOBEC mutational activity. This data provides evidence that WGS can be used to recognize two biologically and clinically distinct myeloma precursor entities that are either progressive or stable.

The molecular make up of smoldering myeloma highlights the evolutionary pathways leading to multiple myeloma.

Smoldering myeloma (SMM) is associated with a high-risk of progression to myeloma (MM). We report the results of a study of 82 patients with both targeted sequencing that included a capture of the immunoglobulin and MYC regions. By comparing these results to newly diagnosed myeloma (MM) we show fewer NRAS and FAM46C mutations together with fewer adverse translocations, del(1p), del(14q), del(16q), and del(17p) in SMM consistent with their role as drivers of the transition to MM. KRAS mutations are associated with a shorter time to progression (HR 3.5 (1.5-8.1), p = 0.001). In an analysis of change in clonal structure over time we studied 53 samples from nine patients at multiple time points. Branching evolutionary patterns, novel mutations, biallelic hits in crucial tumour suppressor genes, and segmental copy number changes are key mechanisms underlying the transition to MM, which can precede progression and be used to guide early intervention strategies.

A longitudinal analysis of chromosomal abnormalities in disease progression from MGUS/SMM to newly diagnosed and relapsed multiple myeloma.

We analyzed variations in terms of chromosomal abnormalities (CA) by fluorescence in situ hybridization (FISH) analysis on purified bone marrow plasma cells throughout the progression from monoclonal gammopathy of undetermined significance/smoldering multiple myeloma (MGUS/SMM) to newly diagnosed MM/plasma cell leukemia (NDMM/PCL) at diagnosis and from diagnostic samples to progressive disease. High risk was defined by the presence of at least del(17p), t(4;14), and/or t(14;16). 1p/1q detection (in the standard FISH panel from 2012 onward) was not available for all patients. We analyzed 139 MM/PCL diagnostic samples from 144 patients, with a median follow-up of 71 months: high-risk CA at diagnosis (MGUS/SMM or NDMM) was present in 28% of samples, whereas 37-39% showed high-risk CA at relapse. In 115 patients with NDMM who evolved to relapsed/refractory MM, we identified 3 different populations: (1) 31/115 patients (27%) with gain of new CA (del13, del17p, t(4;14), t(14;16) or 1q CA when available); (2) 10/115 (9%) patients with loss of a previously identified CA; and (3) 74 patients with no changes. The CA gain group showed a median overall survival of 66 months vs. 84 months in the third group (HR 0.56, 95% CI 0.34-0.92, p = 0.023). Clonal evolution occurs as disease progresses after different chemotherapy lines. Patients who acquired high-risk CA had the poorest prognosis. Our findings highlight the importance of performing FISH analysis both at diagnosis and at relapse.

Monitoring treatment response and disease progression in myeloma with circulating cell-free DNA.

Circulating cell-free DNA (cfDNA) has the potential to capture spatial genetic heterogeneity in myeloma (MM) patients. We assessed whether cfDNA levels vary according to risk status defined by the 70 gene expression profile (GEP70). cfDNA levels in 77 patients were significantly higher in the GEP70 high-risk (HR) group compared to the low-risk (LR) group and correlated weakly with clinical markers including lactate dehydrogenase, ß2 -microglobulin, and ISS. Patients with high cfDNA levels were associated with a worse PFS (hazard ratio 6.4; 95% CI of ratio 1.9-22) and OS (hazard ratio 4.4; 95% CI of ratio 1.2-15.7). Circulating tumor DNA (ctDNA) was elevated in the HR group and ctDNA correlated strongly with GEP70 risk score (Spearman r = .69, P = .0027). cfDNA concentrations were significantly elevated between days 3-5 after chemotherapy before falling back to baseline levels. ctDNA in two patients showed a similar spike in levels between days 3 and 5 after chemotherapy with a concomitant increase in allele fraction of KRAS mutations. We assessed cfDNA levels in 25 patients with smoldering myeloma with serial samples and showed increased allele fraction of mutated KRAS at progression in cfDNA. Our study shows that cfDNA is a dynamic tool to capture genetic events in myeloma.

Risk Stratification and Treatment in Smoldering Multiple Myeloma.

Smoldering multiple myeloma is a heterogeneous asymptomatic precursor to multiple myeloma. Since its identification in 1980, risk stratification models have been developed using two main stratification methods: clinical measurement-based and genetics-based. Clinical measurement models can be subdivided in three types: baseline measurements (performed at diagnosis), evolving measurements (performed over time during follow-up appointments), and imaging (for example, magnetic resonance imaging). Genetic approaches include gene expression profiling, DNA/RNA sequencing, and cytogenetics. It is important to accurately distinguish patients with indolent disease from those with aggressive disease, as clinical trials have shown that patients designated as "high-risk of progression" have improved outcomes when treated early. The risk stratification models, and clinical trials are discussed in this review.

Genomic Profiling of Smoldering Multiple Myeloma Identifies Patients at a High Risk of Disease Progression.

PURPOSE: Smoldering multiple myeloma (SMM) is a precursor condition of multiple myeloma (MM) with a 10% annual risk of progression. Various prognostic models exist for risk stratification; however, those are based on solely clinical metrics. The discovery of genomic alterations that underlie disease progression to MM could improve current risk models. METHODS: We used next-generation sequencing to study 214 patients with SMM. We performed whole-exome sequencing on 166 tumors, including 5 with serial samples, and deep targeted sequencing on 48 tumors. RESULTS: We observed that most of the genetic alterations necessary for progression have already been acquired by the diagnosis of SMM. Particularly, we found that alterations of the mitogen-activated protein kinase pathway (KRAS and NRAS single nucleotide variants [SNVs]), the DNA repair pathway (deletion 17p, TP53, and ATM SNVs), and MYC (translocations or copy number variations) were all independent risk factors of progression after accounting for clinical risk staging. We validated these findings in an external SMM cohort by showing that patients who have any of these three features have a higher risk of progressing to MM. Moreover, APOBEC associated mutations were enriched in patients who progressed and were associated with a shorter time to progression in our cohort. CONCLUSION: SMM is a genetically mature entity whereby most driver genetic alterations have already occurred, which suggests the existence of a right-skewed model of genetic evolution from monoclonal gammopathy of undetermined significance to MM. We identified and externally validated genomic predictors of progression that could distinguish patients at high risk of progression to MM and, thus, improve on the precision of current clinical models.

Current and potential applications of positron emission tomography for multiple myeloma and plasma cell disorders.

Fluorine-18 (18F)-fluorodeoxyglucose (FDG) positron emission tomography (PET) allows evaluation of elevated glucose metabolism in malignancies. There has been increasing interest in FDG PET/CT for plasma cell disorders since the International Myeloma Working Group outlined multiple applications of this imaging modality, including distinguishing smoldering myeloma from active multiple myeloma, confirmation of solitary plasmacytoma, and multiple indications in patients with known multiple myeloma, including determining extent of initial disease, monitoring therapy response, and detection of residual disease following therapy. The field of molecular imaging is now shifting focus from evaluation of metabolism to targeted evaluation of specific tumor markers. Targeted PET imaging targeted of CXCR4 and CD38 has advanced into translational clinical trials, bringing us closer to powerful imaging options for myeloma. In this review we discuss the current applications of FDG PET/CT in plasma cell disorders, as well as advances in targeted PET imaging.

Moving From Cancer Burden to Cancer Genomics for Smoldering Myeloma: A Review.

Importance: All patients who develop multiple myeloma have a preceding asymptomatic expansion of clonal plasma cells, clinically recognized as monoclonal gammopathy of undetermined significance or smoldering multiple myeloma (SMM). During the past decade, significant progress has been made in the classification and risk stratification of SMM. Observations: This review summarizes current clinical challenges and discusses available models for risk stratification in the context of SMM. Owing to several novel, more effective, and less toxic drugs, these aspects are becoming increasingly important to identify patients eligible for early treatment. However, all proposed criteria were built around indirect markers of disease burden and therefore are generally able to identify a fraction of patients with SMM in whom transformation to multiple myeloma and genomic subclonal diversification are already happening. In contrast, next-generation sequencing approaches have the potential to identify myeloma precursor disease that will progress even before the major clonal expansion and progression, providing a potential base for more effective treatment and better precision regarding the optimal timing of treatment initiation. Conclusions and Relevance: Owing to modern technologies, in the near future, prognostic models derived from genomic signatures independent of the disease burden will allow better identification of the optimal timing to treat a plasma cell clonal disorder at the very early stages, when the chances of eradication are higher.

Therapeutic Advances in the Management of Smoldering Myeloma.

BACKGROUND: The International Myeloma Working Group has defined smoldering multiple myeloma (SMM) as the presence of 10%-60% plasma cells in the bone marrow and M-protein (IgG, IgA) ≥3 g/dL without end-organ damage (an increased calcium level, renal failure, anemia, and destructive bone lesions). AREAS OF UNCERTAINTY: Patients considered to have SMM should not have any myeloma-defining events or amyloidosis. Different risks factors classify SMM into low-, intermediate-, or high-risk categories. The rate of progression from SMM to symptomatic myeloma is ∼10% per year during the first 5 years of diagnosis. SMM requires frequent follow-up ∼every 3 months during the first 5 years as compared to monoclonal gammopathy of undermined significance, which usually requires follow-up every 6-12 months after the first year of diagnosis. DATA SOURCES: A literature search was performed from electronic bibliographic databases: MEDLINE (Ovid SP/PubMed), EMBASE, the Cochrane Library (Cochrane Database of Systematic Reviews), and Cochrane Central Register of Controlled Trials and from annual meeting abstracts from inception to May 2017. THERAPEUTIC ADVANCES: This review presents the literature and available data that support or do not support early treatment of high-risk SMM (HR-SMM) and provides evidence-based recommendations for management of SMM patients. Despite emerging data recommending early treatment of HR-SMM, we predict the SMM category may disappear in the near future and patients will be diagnosed with either multiple myeloma or monoclonal gammopathy of undermined significance. CONCLUSIONS: Success with early therapy trials for HR-SMM is largely due to patients meeting current criteria for multiple myeloma that may have been classified as SMM and, therefore, benefitted from therapy. Based on current practices and the literature, SMM should be managed with close follow-up. Based on available data, we suggest SMM to only be treated in clinical trial settings.

Eruptive ulcerative follicular spicules heralding progression of smoldering multiple myeloma.

Herein, we describe a patient with immunoglobulin G (IgG)-lambda smoldering multiple myeloma with translocation t(4:14) who developed widespread ulcerative horny-like spicules, heralding rapid progression to overt myeloma requiring aggressive chemotherapy and autologous stem cell transplantation. The serum abnormal immunoglobulin in the blood was cryoglobulin, which typically precipitates in the tissues at low temperatures causing inflammation and tissue damage. Histopathological changes, observed in lesions at different evolutionary stages, evidenced columns of horny-like eosinophilic homogeneous material, immunoreactive for IgG lambda, protruding from the dilated and/or distorted follicular openings or acrosyringia and small vessel thrombotic vasculopathy and vasculitis in concert with an inflammatory neutrophilic and lymphocytic reaction. Biochemical investigations on material from a spicule and ulcero-necrotic lesion revealed cryoprecipitates containing IgG-lambda with electrophoretic characteristics identical to those of the serum dysprotein. Our findings suggest that the formation of spicules and development of ulcerative lesions are a part of the same clinical spectrum where the cold-dependent precipitation of the immunogenic dysprotein, both in the skin vessels and hair follicle infundibula, play a major pathogenetic role. Whether this highly characteristic paraneoplastic dermatosis can identify patients with high-risk cytogenetic abnormalities and be incorporated into prognostic models, applicable early on in the course of myeloma, warrants further investigation.

Single cell dissection of plasma cell heterogeneity in symptomatic and asymptomatic myeloma.

Multiple myeloma, a plasma cell malignancy, is the second most common blood cancer. Despite extensive research, disease heterogeneity is poorly characterized, hampering efforts for early diagnosis and improved treatments. Here, we apply single cell RNA sequencing to study the heterogeneity of 40 individuals along the multiple myeloma progression spectrum, including 11 healthy controls, demonstrating high interindividual variability that can be explained by expression of known multiple myeloma drivers and additional putative factors. We identify extensive subclonal structures for 10 of 29 individuals with multiple myeloma. In asymptomatic individuals with early disease and in those with minimal residual disease post-treatment, we detect rare tumor plasma cells with molecular characteristics similar to those of active myeloma, with possible implications for personalized therapies. Single cell analysis of rare circulating tumor cells allows for accurate liquid biopsy and detection of malignant plasma cells, which reflect bone marrow disease. Our work establishes single cell RNA sequencing for dissecting blood malignancies and devising detailed molecular characterization of tumor cells in symptomatic and asymptomatic patients.

Genomic patterns of progression in smoldering multiple myeloma.

We analyzed whole genomes of unique paired samples from smoldering multiple myeloma (SMM) patients progressing to multiple myeloma (MM). We report that the genomic landscape, including mutational profile and structural rearrangements at the smoldering stage is very similar to MM. Paired sample analysis shows two different patterns of progression: a "static progression model", where the subclonal architecture is retained as the disease progressed to MM suggesting that progression solely reflects the time needed to accumulate a sufficient disease burden; and a "spontaneous evolution model", where a change in the subclonal composition is observed. We also observe that activation-induced cytidine deaminase plays a major role in shaping the mutational landscape of early subclinical phases, while progression is driven by APOBEC cytidine deaminases. These results provide a unique insight into myelomagenesis with potential implications for the definition of smoldering disease and timing of treatment initiation.

Bone marrow eosinophils in plasma cell disorders.

In experimental studies, eosinophils have been shown to promote the survival, proliferation, and retention of plasma cells in the bone marrow (BM). The clinical significance of eosinophils in plasma cell disorders (PCDs) in humans is largely unknown. This study focuses on the frequency and phenotype of eosinophils in the BM and peripheral blood (PB) in patients with untreated PCD compared with healthy controls. The number of eosinophils per se did not correlate with the number of BM plasma cells or disease stage. The expression of chemokine receptor 4, which is important in the homing capacity to bone marrow stromal cells, was significantly higher in patient eosinophils and increased with disease stage. BM eosinophils from patients, especially from those with manifest disease, were more activated. Another finding in this study was that eosinophils in PB and BM from both patients and healthy controls expressed CD80 (B7-1). We discuss probable immunomodulatory consequences of surface expression of CD80 by eosinophils in conditions with marked T-cell exhaustion (e.g., multiple myeloma). Finally, we found that patients treated with corticosteroids had low levels of circulating eosinophils but preserved levels of eosinophils in the BM.