Second primary malignancies in multiple myeloma: an overview and IMWG consensus.

Background: Therapeutic advancements following the introduction of autologous stem cell transplantation and 'novel' agents have significantly improved clinical outcomes for patients with multiple myeloma (MM). Increased life expectancy, however, has led to renewed concerns about the long-term risk of second primary malignancies (SPMs). This review outlines the most up-to-date knowledge of possible host-, disease-, and treatment-related risk factors for the development of SPMs in patients with MM, and provides practical recommendations to assist physicians. Design: A Panel of International Myeloma Working Group members reviewed the most relevant data published in the literature as full papers, or presented at meetings of the American Society of Clinical Oncology, American Society of Hematology, European Hematology Association, or International Myeloma Workshops, up to June 2016. Here, we present the recommendations of the Panel, based on this literature review. Results: Overall, the risk of SPMs in MM is low, multifactorial, and partially related to the length of patients' survival and MM intrinsic susceptibility. Studies suggest a significantly increased incidence of SPMs when lenalidomide is administered either following, or concurrently with, oral melphalan. Increased SPM incidence has also been reported with lenalidomide maintenance following high-dose melphalan, albeit to a lesser degree. In both cases, the risk of death from MM was significantly higher than the risk of death from SPMs, with lenalidomide possibly providing a survival benefit. No increase in SPM incidence was reported with lenalidomide plus dexamethasone (without melphalan), or with bortezomib plus oral melphalan, dexamethasone, or thalidomide. Conclusion: In general, the risk of SPMs should not alter the current therapeutic decision-making process in MM. However, regimens such as lenalidomide plus dexamethasone should be preferred to prolonged exposure to lenalidomide plus oral melphalan. SPM risk should be carefully discussed with the patient in the context of benefits and risks of different treatment options.

High speed scintillation autoradiography.

Impregnation of nuclear track emulsion with liquid scintillator and exposure at -85 degrees C allows rapid autoradiographic labeling. With tritiated thymidine of high specific activity (40 to 60 curies per millimole), exposure time can be shortened to 20 to 60 minutes, allowing complete sample processing within 4 hours. In experiments requiring isotopes with low incorporation rates or low specific activity, exposure time can be shortened from months to several days.

Cystic fibrosis: decreased conjugation and excretion of [14C]spermidine.

Free and conjugated [14C]spermidine were measured in plasma samples from normal individuals and cystic fibrosis patients. Within 4 minutes, the 14C-labeled material in the plasma from normal individuals was 70 percent conjugated compared to no detectable conjugation by cystic fibrosis patients. Further, the patients excreted only 11 to 13 percent of the [14C]spermidine in their urine within 72 hours whereas normal excretion was 60 to 76 percent. In both cases, the labeled material was in a conjugated form.

Next generation flow for minimally-invasive blood characterization of MGUS and multiple myeloma at diagnosis based on circulating tumor plasma cells (CTPC).

Here, we investigated for the first time the frequency and number of circulating tumor plasma cells (CTPC) in peripheral blood (PB) of newly diagnosed patients with localized and systemic plasma cell neoplasms (PCN) using next-generation flow cytometry (NGF) and correlated our findings with the distinct diagnostic and prognostic categories of the disease. Overall, 508 samples from 264 newly diagnosed PCN patients, were studied. CTPC were detected in PB of all active multiple myeloma (MM; 100%), and smoldering MM (SMM) patients (100%), and in more than half (59%) monoclonal gammopathy of undetermined significance (MGUS) cases (p <0.0001); in contrast, CTPC were present in a small fraction of solitary plasmacytoma patients (18%). Higher numbers of CTPC in PB were associated with higher levels of BM infiltration and more adverse prognostic features, together with shorter time to progression from MGUS to MM (p <0.0001) and a shorter survival in MM patients with active disease requiring treatment (p ≤ 0.03). In summary, the presence of CTPC in PB as assessed by NGF at diagnosis, emerges as a hallmark of disseminated PCN, higher numbers of PB CTPC being strongly associated with a malignant disease behavior and a poorer outcome of both MGUS and MM.

International uniform response criteria for multiple myeloma.

New uniform response criteria are required to adequately assess clinical outcomes in myeloma. The European Group for Blood and Bone Marrow Transplant/International Bone Marrow Transplant Registry criteria have been expanded, clarified and updated to provide a new comprehensive evaluation system. Categories for stringent complete response and very good partial response are added. The serum free light-chain assay is included to allow evaluation of patients with oligo-secretory disease. Inconsistencies in prior criteria are clarified making confirmation of response and disease progression easier to perform. Emphasis is placed upon time to event and duration of response as critical end points. The requirements necessary to use overall survival duration as the ultimate end point are discussed. It is anticipated that the International Response Criteria for multiple myeloma will be widely used in future clinical trials of myeloma.

Circulating tumour DNA analysis demonstrates spatial mutational heterogeneity that coincides with disease relapse in myeloma.

Mutational characterisation in multiple myeloma (MM) currently relies on bone marrow (BM) biopsy, which fails to capture the putative spatial and genetic heterogeneity of this multifocal disease. Analysis of plasma (PL)-derived circulating free tumour DNA (ctDNA) as an adjunct to BM biopsy, for mutational characterisation and tracking disease progression, was evaluated. Paired BM MM cell DNA and ctDNA from 33 relapsed/refractory (RR) and 15 newly diagnosed (ND) patients were analysed for KRAS, NRAS, BRAF and TP53 mutations using the OnTarget Mutation Detection (OMD) platform. OMD detected 128 mutations (PL=31, BM=59, both=38) indicating the presence of PL mutations (54%). A higher frequency of PL-only mutations was detected in RR patients than ND (27.2% vs 6.6%, respectively), authenticating the existence of spatial and genetic heterogeneity in advanced disease. Activating RAS mutations were more highly prevalent than previously described with 69% harboring at least one RAS mutation. Sequential ctDNA quantitation with droplet digital PCR through longitudinal PL tracking of specific clones in seven patients demonstrated changes in fractional abundance of certain clones reflective of the disease status. We conclude that ctDNA analysis as an adjunct to BM biopsy represents a noninvasive and holistic strategy for improved mutational characterisation and therapeutic monitoring of MM.

Whole-body computed tomography versus conventional skeletal survey in patients with multiple myeloma: a study of the International Myeloma Working Group.

For decades, conventional skeletal survey (CSS) has been the standard imaging technique for multiple myeloma (MM). However, recently whole-body computed tomography (WBCT) has been implemented into the diagnostic criteria of MM. This analysis compares sensitivity and prognostic significance of WBCT and CSS in patients with smoldering MM (SMM) and MM. Fifty-four of 212 patients (25.5%) had a negative CSS and a positive WBCT for osteolytic lesions (P<0.0001). Of 66 patients with SMM based on CSS, 12 (22.2%) had osteolytic lesions on WBCT. In comparison, WBCT failed to detect some bone destructions in the appendicular skeleton possibly due to limitations of the field of view. Presence of lytic bone lesions in WBCT was of borderline prognostic significance (P=0.051) for SMM patients, with a median time to progression of 38 versus 82 months for those without bone destructions. In conclusion, WBCT identifies significantly more sites of bone destruction than CSS. More than 20% of patients with SMM according to CSS have in fact active MM detectable with WBCT. On the basis of this and other studies, WBCT (either computed tomography (CT) alone or as part of a positron emission tomography-CT protocol) should be considered the current standard for the detection of osteolytic lesions in MM.

Next Generation Flow for highly sensitive and standardized detection of minimal residual disease in multiple myeloma.

Flow cytometry has become a highly valuable method to monitor minimal residual disease (MRD) and evaluate the depth of complete response (CR) in bone marrow (BM) of multiple myeloma (MM) after therapy. However, current flow-MRD has lower sensitivity than molecular methods and lacks standardization. Here we report on a novel next generation flow (NGF) approach for highly sensitive and standardized MRD detection in MM. An optimized 2-tube 8-color antibody panel was constructed in five cycles of design-evaluation-redesign. In addition, a bulk-lysis procedure was established for acquisition of ⩾107 cells/sample, and novel software tools were constructed for automatic plasma cell gating. Multicenter evaluation of 110 follow-up BM from MM patients in very good partial response (VGPR) or CR showed a higher sensitivity for NGF-MRD vs conventional 8-color flow-MRD -MRD-positive rate of 47 vs 34% (P=0.003)-. Thus, 25% of patients classified as MRD-negative by conventional 8-color flow were MRD-positive by NGF, translating into a significantly longer progression-free survival for MRD-negative vs MRD-positive CR patients by NGF (75% progression-free survival not reached vs 7 months; P=0.02). This study establishes EuroFlow-based NGF as a highly sensitive, fully standardized approach for MRD detection in MM which overcomes the major limitations of conventional flow-MRD methods and is ready for implementation in routine diagnostics.

Natural history of relapsed myeloma, refractory to immunomodulatory drugs and proteasome inhibitors: a multicenter IMWG study.

Introduction of new myeloma therapies offers new options for patients refractory to immunomodulatory drugs (IMiDs) and proteasome inhibitors (PIs). In this multicenter study, patients with relapsed multiple myeloma, who have received at least three prior lines of therapy, are refractory to both an IMiD (lenalidomide or pomalidomide) and a PI (bortezomib or carfilzomib), and have been exposed to an alkylating agent were identified. The time patients met the above criteria was defined as time zero (T0). Five hundred and forty-three patients diagnosed between 2006 and 2014 were enrolled in this study. Median age at T0 was 62 years (range 31-87); 61% were males. The median duration between diagnosis and T0 was 3.1 years. The median number of lines of therapy before T0 was 4 (range 3-13). The median overall survival (OS) from T0 for the entire cohort was 13 (95% confidence interval (CI) 11, 15) months. At least one regimen recorded after T0 in 462 (85%) patients, with a median (95% CI) progression-free survival and OS from T0 of 5 (4, 6), and 15.2 (13, 17) months, respectively. The study provides the expected outcome of relapsed multiple myeloma that is refractory to a PI and an IMiD, a benchmark for comparison of new therapies being evaluated.

Polyamines as markers of response and disease activity in cancer chemotherapy.

One hundred twenty-four patients with hematological and solid neoplasms had pretreatment urinary polyamine determinations. Putrescine, spermidine, and spermine were all significantly increased as compared to normals (p less than 0.001). Polyamine levels were directly related to disease activity and tumor burden. In patients with multiple myeloma, putrescine levels were significantly correlated with clinical disease activity as well as the in vitro labeling index of marrow plasma cells. Spermidine values reflected tumor cell burden. Serial studies in 56 patients indicated that greater than twofold rise in urinary spermidine during treatment was highly correlated with cell kill and subsequent clinical response (p less than 0.001). Serum polyamine levels in 17 patients were found to be comparable to urinary values. Our data indicate that polyamine determinations can potentially be clinically useful, i.e., baseline values as indicators of tumor cell mass and growth fraction, and increases in spermidine during treatment as an excellent marker of tumor cell kill.

Metabolism of [14C]spermidine and [14C]putrescine in normal volunteers and in cancer patients.

The administration of [14C]putrescine or [14C]spermidine (i.v., 100 muCi) to normal volunteers or patients with advanced cancer resulted in alpha-phase half-lives of 40 and 30 sec and beta-phase half-lives of 30 and 60 min, respectively. No significant differences were found between the plasma decay curves of normals and those of cancer patients. Urinary excretion was similar with both groups excreting approximately 45% of [ 14C]putrescine within 24 hr and 60 to 76% of [14C]spermidine within 48 hr. Dowex chromatography indicated that more than 90% of the radiolabel in the urine was in a conjugated form. Plasma conjugation studies of [14C]putrescine and [14C]spermidine in both groups indicated near-total conjugation of the radiolabel within 4 to 5 min of i.v. injection. Since putrescine and spermidine are markers of disease activity, characterization of the conjugates will be important for the development of rapid, specific tests of altered disease activity in response to multimodality therapy.

Characterization of a new drug-resistant human myeloma cell line that expresses P-glycoprotein.

Multiple myeloma is a disease with a high initial chemotherapeutic response but virtually no cures due to emergence of drug resistance. A doxorubicin-resistant human myeloma cell line (8226/Dox) has been selected from the myeloma cell line RPMI8226 by continuously exposing cells to gradually increasing doses of doxorubicin. The resistant phenotype has been retained for over 2 months despite growth in drug-free medium. The resistant subline was cross-resistant to mitoxantrone, acronycine, etoposide, and vincristine. The 8226/Dox cell line remained sensitive to melphalan but acquired collateral sensitivity to dexamethasone. Intracellular doxorubicin accumulation, as measured by [14C]doxorubicin and high-performance liquid chromatography, was decreased by 54% at 1 h for 8226/Dox compared to the sensitive line. Efflux of doxorubicin was significantly greater in the resistant subline as compared to the sensitive parent cell line. Membrane analysis using immunoblotting techniques detected increased expression of the integral membrane protein P-glycoprotein (Mr 170,000) in the resistant subline. Cytogenetic analysis of 8226/Dox revealed a 7q-anomaly not seen in the parent cell line. No double minutes or homogeneously staining regions were observed. The drug sensitivity/resistance pattern of the resistant cell line correlates well with clinical observations indicating the potential of this cell line as a model for resistance in multiple myeloma.

In vitro chemosensitivities of human tumor stem cells to the Phase II drug 4'-(9-acridinylamino)methanesulfon-m-anisidide and prospective in vivo correlations.

A potential application of the human tumor stem cell colony assay is to guide Phase II clinical investigations by identifying classes of tumors (or individual patients) which are sensitive in vitro to a new antitumor compound. We have tested human tumor stem cells from 140 tumor biopsies representing 20 different tumor types for chemosensitivity to the Phase II drug 4'-(9-acridinylamino)methanesulfon-m-anisidide. In vitro sensitivity was defined as a reduction in the number of tumor colony-forming cells to 30% of the control or less after a 1-hr exposure to one-tenth of the pharmacologically achievable plasma concentration of 4'-(9-acridinylamino)methanesulfon-m-anisidide. In vitro sensitivity was found in 29 cases: non-Hodgkin's lymphoma (2 of 2); cervical carcinoma (1 of 1); sarcoma (3 of 6); neuroblastoma (1 of 2); acute myelogenous leukemia (6 of 16); chronic myelogenous leukemia (1 of 3); melanoma (8 of 34); uterine carcinoma (1 of 5); lung carcinoma (1 of 9); ovarian carcinoma (4 of 36); and breast carcinoma (1 of 11). Prospective in vitro-in vivo correlations in eight patients with various tumor types showed that three of three patients sensitive in vitro to 4'-(9-acridinylamino)methanesulfon-m-anisidide responded in vivo, while five of five patients resistant in vitro had no clinical response. The results provide support for further evaluation of the utility of the human tumor stem cell colony assay for targeting Phase II clinical trials.

Phenotypic and functional analysis of 1,25-dihydroxyvitamin D3 receptor mediated modulation of the human myeloma cell line RPMI 8226.

Several recent studies have demonstrated the presence of specific receptors for the 1,25-dihydroxyvitamin D3 (calcitriol) in activated normal lymphocytes. By DNA cellulose chromatography, we show evidence of such specific receptors in the human myeloma cell line RPMI 8226. Nanomolar concentrations of 1,25-dihydroxyvitamin D3 reduce the proliferation of RPMI 8226 cells significantly and simultaneously induce the appearance of both new properties and phenotype expression, such as butyrate esterase, enhanced expression of CD20 (B1), CD15 (Leu-M1) antigens and lambda chains, and decreased expression of the PC1 antigen using microfluorometric analysis. But such an increased expression of membrane lambda chains was not associated with an enhanced secretion of lambda chains. Furthermore, the bone resorbing activity produced normally by RPMI 8226 cells was reduced significantly after 1,25-dihydroxyvitamin D3 treatment. The possible mechanisms and significance of these new functional and phenotypic properties are discussed with respect to the B-cell lineage.

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.

Gene signature combinations improve prognostic stratification of multiple myeloma patients.

Multiple myeloma (MM) is a plasma cell neoplasm with significant molecular heterogeneity. Gene expression profiling (GEP) has contributed significantly to our understanding of the underlying biology and has led to several prognostic gene signatures. However, the best way to apply these GEP signatures in clinical practice is unclear. In this study, we investigated the integration of proven prognostic signatures for improved patient risk stratification. Three publicly available MM GEP data sets that encompass newly diagnosed as well as relapsed patients were analyzed using standardized estimation of nine prognostic MM signature indices and simulations of signature index combinations. Cox regression analysis was used to assess the performance of simulated combination indices. Taking the average of multiple GEP signature indices was a simple but highly effective way of integrating multiple GEP signatures. Furthermore, although adding more signatures in general improved performance substantially, we identified a core signature combination, EMC92+HZDCD, as the top-performing prognostic signature combination across all data sets. In this study, we provided a rationale for gene signature integration and a practical strategy to choose an optimal risk score estimation in the presence of multiple prognostic signatures.

Management of relapsed multiple myeloma: recommendations of the International Myeloma Working Group.

The prognosis for patients multiple myeloma (MM) has improved substantially over the past decade with the development of new, more effective chemotherapeutic agents and regimens that possess a high level of anti-tumor activity. In spite of this important progress, however, nearly all MM patients ultimately relapse, even those who experience a complete response to initial therapy. Management of relapsed MM thus represents a vital aspect of the overall care for patients with MM and a critical area of ongoing scientific and clinical research. This comprehensive manuscript from the International Myeloma Working Group provides detailed recommendations on management of relapsed disease, with sections dedicated to diagnostic evaluation, determinants of therapy, and general approach to patients with specific disease characteristics. In addition, the manuscript provides a summary of evidence from clinical trials that have significantly impacted the field, including those evaluating conventional dose therapies, as well as both autologous and allogeneic stem cell transplantation. Specific recommendations are offered for management of first and second relapse, relapsed and refractory disease, and both autologous and allogeneic transplant. Finally, perspective is provided regarding new agents and promising directions in management of relapsed MM.

Management of refractory myeloma: a review.

Management of refractory myeloma represents a common and challenging clinical problem. Approximately 30% to 50% of patients with multiple myeloma do not respond to first-line therapy, and those who initially achieve a remission will eventually relapse. Surprisingly, there is no routinely accepted approach to patients with refractory disease. Therefore, we review the literature in an attempt to provide an overview of the published results and outline our treatment recommendations for such patients. We suggest the following: (1) for truly resistant patients (ie, those who clearly progress with initial therapy), administration of high-dose or pulsed glucocorticosteroids is the best treatment, with an expected response of 40% (defined as a greater than or equal to 50% reduction in monoclonal [M]-protein concentration); (2) for patients who relapse during therapy or relapse within 6 months of stopping the initial treatment, the VAD regimen (vincristine, doxorubicin, and dexamethasone) is one of the most effective salvage therapies, resulting in an approximately 75% response rate (greater than or equal to 50% reduction in M-protein concentration); (3) for patients who relapse within more than 6 months of stopping therapy (unmaintained remission), reinitiation of the initial therapy represents an excellent alternative, leading to recontrol in 60% to 70% of patients (greater than or equal to 50% reduction in M-protein concentration). If progression is observed or if there is response and then relapse in this setting, VAD chemotherapy can be administered again. (4) Patients who fail second-line salvage therapies should enter well-designed clinical trials to evaluate new treatment modalities. If this is not feasible, alpha-interferon or "systemic" radiotherapy are recommended in selected cases.

Prognostic factors and staging in multiple myeloma: a reappraisal.

To assess the important factors in the prognosis and staging of multiple myeloma (MM), we have correlated the presenting clinical features of 147 previously untreated patients with MM with the survival duration using multiple regression analyses. We have included the three major available myeloma-staging systems (MSS), ie, Durie-Salmon (DS), Medical Research Council (MRC), and Merlini-Waldenström-Jayakar (MWJ), plus two new variables related to disease activity: the serum beta 2-microglobulin level (S beta 2M) and the instantaneous rate of bone resorption. Our study confirms the validity of the three MSS in the prediction of survival duration, with a slight but significant advantage for the DS MSS. Among single variables, S beta 2M was the most powerful indicator of prognosis (P less than .0001), serum albumin level being the only variable adding to this significantly (P = .02). Of major interest, S beta 2M alone was a better indicator than MRC and MWJMSS. Finally, S beta 2M and the serum albumin level, variables not included in the three MSS, were better indicators than the classical DS MSS and could be combined simply to give a very powerful system of stratification.