ABSTRACT
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.
Subject(s)
Biomarkers, Tumor , Disease Progression , Drug Resistance, Neoplasm , Mutation , Smoldering Multiple Myeloma , Humans , Male , Drug Resistance, Neoplasm/genetics , Female , Smoldering Multiple Myeloma/genetics , Biomarkers, Tumor/genetics , Middle Aged , Aged , High-Throughput Nucleotide Sequencing , Antineoplastic Combined Chemotherapy Protocols/therapeutic useABSTRACT
Although the use of games as an educational strategy is an important current trend, there is practically no option available for training people on the Drug Discovery and Development (DDD) process. To fill this gap, we designed "SCREENER", a science game that is intended to be educational, but also challenging and interesting enough to ensure player engagement. Our main target audience is students of postgraduate programs in pharmacology, medicinal chemistry, pharmacy, and medicine. This game could also be of interest to the pharmaceutical industry and regulatory and patent agencies for training new employees. We discuss the creation of SCREENER, a hybrid of board and card games, and present its components with some examples of cards and resources, as well as the dynamics of the game. SCREENER mimics the process of drug discovery and development from validating a target to registering the new drug with the regulatory agency, and can be played individually (self-learning) or with the help of a monitor who assists up to six players/teams. Briefly, 29 task cards categorized in four major areas (efficacy, safety, pharmacokinetics, and pharmaceutical development) must be purchased sequentially. Classic characteristics of games such as decision making and challenge have been incorporated. More in-depth information on the tasks and technical terms is available through QR codes. The vagaries of the DDD process are mimicked by the bonus/setback cards. The evaluation of our first test with students is presented and supports the usefulness of this new tool.
Subject(s)
Learning , Students , Drug Discovery , Educational Status , HumansABSTRACT
Although the use of games as an educational strategy is an important current trend, there is practically no option available for training people on the Drug Discovery and Development (DDD) process. To fill this gap, we designed "SCREENER", a science game that is intended to be educational, but also challenging and interesting enough to ensure player engagement. Our main target audience is students of postgraduate programs in pharmacology, medicinal chemistry, pharmacy, and medicine. This game could also be of interest to the pharmaceutical industry and regulatory and patent agencies for training new employees. We discuss the creation of SCREENER, a hybrid of board and card games, and present its components with some examples of cards and resources, as well as the dynamics of the game. SCREENER mimics the process of drug discovery and development from validating a target to registering the new drug with the regulatory agency, and can be played individually (self-learning) or with the help of a monitor who assists up to six players/teams. Briefly, 29 task cards categorized in four major areas (efficacy, safety, pharmacokinetics, and pharmaceutical development) must be purchased sequentially. Classic characteristics of games such as decision making and challenge have been incorporated. More in-depth information on the tasks and technical terms is available through QR codes. The vagaries of the DDD process are mimicked by the bonus/setback cards. The evaluation of our first test with students is presented and supports the usefulness of this new tool.
ABSTRACT
Long-term survival after liver transplantation is affected by de novo neoplasia. The incidence of this type of malignancy is increased in the setting of immunosuppressive therapy. The aim of this study was to characterize the immunologic pattern of liver transplant recipients with de novo malignancies. Fifty-one liver recipients were studied, 19 of whom had a history of de novo neoplasia. Immunophenotypic patterns among patients with/without tumors were compared. The subpopulations of CD4+ T lymphocytes and CD8+ T lymphocytes differed between the 2 types of patients studied. In patients with tumor, activation membrane markers in CD4+ T lymphocytes and CD8+ T-lymphocytes, such as CD56 or CD25, were expressed in a greater proportion, whereas activation markers CD314 and CD16 were reduced in CD56bright natural killer (NK) cells. We concluded that cytotoxic response seems to be more activated in de novo neoplasia patients, which highlights the still unknown malignancy risk effect on these immune cells.
Subject(s)
Immunocompromised Host/immunology , Liver Transplantation , Neoplasms/immunology , CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , Cross-Sectional Studies , Female , Humans , Immunophenotyping , Killer Cells, Natural/immunology , Male , Middle AgedABSTRACT
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.
Subject(s)
Flow Cytometry , Monoclonal Gammopathy of Undetermined Significance/diagnosis , Multiple Myeloma/diagnosis , Neoplastic Cells, Circulating/metabolism , Plasma Cells/metabolism , Adult , Aged , Aged, 80 and over , Biomarkers , Diagnosis, Differential , Female , Flow Cytometry/methods , Humans , Immunophenotyping , Male , Middle Aged , Monoclonal Gammopathy of Undetermined Significance/metabolism , Multiple Myeloma/metabolism , Neoplasm Staging , Neoplastic Cells, Circulating/pathology , Plasma Cells/pathology , Prognosis , Sensitivity and SpecificityABSTRACT
Solitary plasmacytoma is an infrequent form of plasma cell dyscrasia that presents as a single mass of monoclonal plasma cells, located either extramedullary or intraosseous. In some patients, a bone marrow aspiration can detect a low monoclonal plasma cell infiltration which indicates a high risk of early progression to an overt myeloma disease. Before treatment initiation, whole body positron emission tomography-computed tomography or magnetic resonance imaging should be performed to exclude the presence of additional malignant lesions. For decades, treatment has been based on high-dose radiation, but studies exploring the potential benefit of systemic therapies for high-risk patients are urgently needed. In this review, a panel of expert European hematologists updates the recommendations on the diagnosis and management of patients with solitary plasmacytoma.
Subject(s)
Plasmacytoma/diagnosis , Plasmacytoma/therapy , Disease Management , Europe/epidemiology , Humans , Magnetic Resonance Imaging/methods , Plasmacytoma/epidemiology , Positron Emission Tomography Computed Tomography/methods , Prognosis , Treatment OutcomeABSTRACT
This corrects the article DOI: 10.1038/bcj.2017.90.
ABSTRACT
Persistence of minimal residual disease (MRD) after treatment for myeloma predicts inferior outcomes, but within MRD-positive patients there is great heterogeneity with both early and very late relapses. Among different MRD techniques, flow cytometry provides additional information about antigen expression on tumor cells, which could potentially contribute to stratify MRD-positive patients. We investigated the prognostic value of those antigens required to monitor MRD in 1265 newly diagnosed patients enrolled in the GEM2000, GEM2005MENOS65, GEM2005MAS65 and GEM2010MAS65 protocols. Overall, CD19pos, CD27neg, CD38lo, CD45pos, CD81pos, CD117neg and CD138lo expression predicted inferior outcomes. Through principal component analysis, we found that simultaneous CD38lowCD81posCD117neg expression emerged as the most powerful combination with independent prognostic value for progression-free survival (HR:1.69; P=0.002). This unique phenotypic profile retained prognostic value among MRD-positive patients. We then used next-generation flow to determine antigen stability throughout the course of the disease, and found that the expression of antigens required to monitor MRD is mostly stable from diagnosis to MRD stages, except for CD81 whose expression progressively increased from baseline to chemoresistant tumor cells (14 vs 28%). Altogether, we showed that the phenotypic profile of tumor cells provides additional prognostic information, and could be used to further predict risk of relapse among MRD-positive patients.
Subject(s)
Antigens, CD/metabolism , Multiple Myeloma/metabolism , Multiple Myeloma/pathology , Adult , Aged , Aged, 80 and over , Female , Humans , Male , Middle Aged , Neoplasm Recurrence, Local/metabolism , Neoplasm Recurrence, Local/pathology , Neoplasm, Residual/metabolism , Neoplasm, Residual/pathology , PrognosisABSTRACT
Precise classification of acute leukemia (AL) is crucial for adequate treatment. EuroFlow has previously designed an AL orientation tube (ALOT) to guide towards the relevant classification panel (T-cell acute lymphoblastic leukemia (T-ALL), B-cell precursor (BCP)-ALL and/or acute myeloid leukemia (AML)) and final diagnosis. Now we built a reference database with 656 typical AL samples (145 T-ALL, 377 BCP-ALL, 134 AML), processed and analyzed via standardized protocols. Using principal component analysis (PCA)-based plots and automated classification algorithms for direct comparison of single-cells from individual patients against the database, another 783 cases were subsequently evaluated. Depending on the database-guided results, patients were categorized as: (i) typical T, B or Myeloid without or; (ii) with a transitional component to another lineage; (iii) atypical; or (iv) mixed-lineage. Using this automated algorithm, in 781/783 cases (99.7%) the right panel was selected, and data comparable to the final WHO-diagnosis was already provided in >93% of cases (85% T-ALL, 97% BCP-ALL, 95% AML and 87% mixed-phenotype AL patients), even without data on the full-characterization panels. Our results show that database-guided analysis facilitates standardized interpretation of ALOT results and allows accurate selection of the relevant classification panels, hence providing a solid basis for designing future WHO AL classifications.
Subject(s)
Leukemia, Myeloid, Acute/pathology , Acute Disease , Adolescent , Adult , Aged , Aged, 80 and over , Child , Child, Preschool , Female , Humans , Immunophenotyping/methods , Infant , Infant, Newborn , Male , Middle Aged , Precursor Cell Lymphoblastic Leukemia-Lymphoma/pathology , Young AdultABSTRACT
A six-month-old female cat suffered aspiration of an abundant amount of barium sulfate during a radiographic procedure for the diagnosis of megaesophagus. Latero-lateral contrast radiography revealed severe dilation of the thoracic esophagus cranial to the base of the heart. Persistence of the right aortic arch was suspected and later confirmed during corrective surgery. Accumulation of barium sulfate, used as a contrast agent, was clearly observed in the lumen of the bronchi, bronchioles, and alveoli in the radiographic image. Days after the surgery, the animal developed severe respiratory distress, which resulted in death. Cytology results and histology analysis using polarized light demonstrated that the lumen of bronchi, bronchioles, and alveoli exhibited evident histiocytic infiltration with cytoplasm filled by abundant amorphous refractive granular material consistent with barium sulfate. In this report, we describe the anatomical, cytological, histopathological (using polarized light), and x-ray findings of a case of barium sulfate aspiration pneumonia in a cat resulting from the use of this contrast medium for the diagnosis of megaesophagus secondary to persistent right aortic arch.(AU)
Uma gata de seis meses aspirou grande quantidade de sulfato de bário durante procedimento radiográfico para diagnóstico de megaesôfago. Contraste radiográfico látero-lateral revelou dilatação de esôfago torácico até a base do coração. Persistência do arco aórtico direito foi confirmada durante cirurgia corretiva. Acúmulo de sulfato de bário, utilizado como agente de contraste, foi observado no lúmen de brônquios, bronquíolos e alvéolos à imagem radiográfica. Dias após a cirurgia o animal desenvolveu insuficiência respiratória grave e evoluiu para óbito. Resultado de citologia e histologia utilizado luz polarizada demonstrou que lúmen de brônquios, bronquíolos e alvéolos mostraram infiltração histológica com citoplasma cheio de material refratário granular amorfo compatível com sulfato de bário. Nesse relatório, descrevemos achados anatômicos, citológicos, histopatológicos e radiográficos de um caso de pneumonia aspirativa em gata resultante do uso desse meio de contraste para diagnóstico de megaesôfago secundário a persistência do arco aórtico direito.(AU)
Subject(s)
Animals , Cats , Barium Sulfate/analysis , Esophageal Achalasia/veterinary , Pneumonia, Aspiration/veterinary , Aorta, ThoracicABSTRACT
Multiparameter flow cytometry (MFC) has become standard in the management of patients with plasma cell (PC) dyscrasias, and could be considered mandatory in specific areas of routine clinical practice. It plays a significant role during the differential diagnostic work-up because of its fast and conclusive readout of PC clonality, and simultaneously provides prognostic information in most monoclonal gammopathies. Recent advances in the treatment and outcomes of multiple myeloma led to the implementation of new response criteria, including minimal residual disease (MRD) status as one of the most relevant clinical endpoints with the potential to act as surrogate for survival. Recent technical progress led to the development of next-generation flow (NGF) cytometry that represents a validated, highly sensitive, cost-effective and widely available technique for standardized MRD evaluation, which also could be used for the detection of circulating tumor cells. Here we review current applications of MFC and NGF in most PC disorders including the less frequent solitary plasmocytoma, light-chain amyloidosis or Waldenström macroglobulinemia.
Subject(s)
Flow Cytometry/methods , Paraproteinemias/diagnostic imaging , Humans , Paraproteinemias/pathologySubject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , High-Throughput Nucleotide Sequencing/methods , Multiple Myeloma/drug therapy , Neoplasm, Residual/physiopathology , Aged , Female , Humans , Male , Monitoring, Physiologic , Multiple Myeloma/genetics , Multiple Myeloma/pathology , Survival AnalysisABSTRACT
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.
Subject(s)
Flow Cytometry/methods , Immunophenotyping/methods , Multiple Myeloma/diagnosis , Plasma Cells/pathology , Adult , Aged , Aged, 80 and over , Antibody Specificity , Cell Count , Equipment Design , Female , Flow Cytometry/instrumentation , Humans , Immunophenotyping/instrumentation , Male , Middle Aged , Multiple Myeloma/drug therapy , Multiple Myeloma/pathology , Neoplasm, Residual , Sensitivity and Specificity , Software , Specimen Handling , Treatment OutcomeABSTRACT
The notion that plasma cells (PCs) are terminally differentiated has prevented intensive research in multiple myeloma (MM) about their phenotypic plasticity and differentiation. Here, we demonstrated in healthy individuals (n=20) that the CD19-CD81 expression axis identifies three bone marrow (BM)PC subsets with distinct age-prevalence, proliferation, replication-history, immunoglobulin-production, and phenotype, consistent with progressively increased differentiation from CD19+CD81+ into CD19-CD81+ and CD19-CD81- BMPCs. Afterwards, we demonstrated in 225 newly diagnosed MM patients that, comparing to normal BMPC counterparts, 59% had fully differentiated (CD19-CD81-) clones, 38% intermediate-differentiated (CD19-CD81+) and 3% less-differentiated (CD19+CD81+) clones. The latter patients had dismal outcome, and PC differentiation emerged as an independent prognostic marker for progression-free (HR: 1.7; P=0.005) and overall survival (HR: 2.1; P=0.006). Longitudinal comparison of diagnostic vs minimal-residual-disease samples (n=40) unraveled that in 20% of patients, less-differentiated PCs subclones become enriched after therapy-induced pressure. We also revealed that CD81 expression is epigenetically regulated, that less-differentiated clonal PCs retain high expression of genes related to preceding B-cell stages (for example: PAX5), and show distinct mutation profile vs fully differentiated PC clones within individual patients. Together, we shed new light into PC plasticity and demonstrated that MM patients harbouring less-differentiated PCs have dismal survival, which might be related to higher chemoresistant potential plus different molecular and genomic profiles.
Subject(s)
Multiple Myeloma/diagnosis , Multiple Myeloma/metabolism , Plasma Cells/metabolism , Plasma Cells/pathology , Adult , Antigens, CD/metabolism , Biomarkers , Bone Marrow/metabolism , Bone Marrow/pathology , Bone Marrow Cells/metabolism , Bone Marrow Cells/pathology , Case-Control Studies , Cell Cycle , DNA Methylation , Female , Gene Expression Profiling , Genetic Heterogeneity , High-Throughput Nucleotide Sequencing , Humans , Immunophenotyping , In Situ Hybridization, Fluorescence , Male , Middle Aged , Multiple Myeloma/genetics , Multiple Myeloma/mortality , Mutation , Neoplasm Grading , Phenotype , Prognosis , Single-Cell Analysis , Young AdultABSTRACT
The role of endothelial progenitor cell (EPC)-mediated vasculogenesis in hematological malignancies is not well explored. Here, we showed that EPCs are mobilized from the bone marrow (BM) to the peripheral blood at early stages of multiple myeloma (MM); and recruited to MM cell-colonized BM niches. Using EPC-defective ID1+/- ID3-/- mice, we found that MM tumor progression is dependent on EPC trafficking. By performing RNA-sequencing studies, we confirmed that endothelial cells can enhance proliferation and favor cell-cycle progression only in MM clones that are smoldering-like and have dependency on endothelial cells for tumor growth. We further confirmed that angiogenic dependency occurs early and not late during tumor progression in MM. By using a VEGFR2 antibody with anti-vasculogenic activity, we demonstrated that early targeting of EPCs delays tumor progression, while using the same agent at late stages of tumor progression is ineffective. Thus, although there is significant angiogenesis in myeloma, the dependency of the tumor cells on EPCs and vasculogenesis may actually precede this step. Manipulating vasculogenesis at an early stage of disease may be examined in clinical trials in patients with smoldering MM, and other hematological malignancies with precursor conditions.
Subject(s)
Multiple Myeloma/pathology , Neovascularization, Pathologic/drug therapy , Animals , Antibodies/therapeutic use , Bone Marrow , Cell Movement , Clone Cells/pathology , Disease Progression , Endothelial Cells/pathology , Mice , Multiple Myeloma/blood supply , Multiple Myeloma/drug therapy , Neovascularization, Pathologic/prevention & control , Secondary Prevention , Vascular Endothelial Growth Factor Receptor-2/immunologySubject(s)
B-Lymphocytes/immunology , Palatine Tonsil/immunology , Palatine Tonsil/microbiology , Tonsillitis/immunology , Tonsillitis/microbiology , Adenoidectomy , Adolescent , Child , Child, Preschool , Female , Flow Cytometry , Humans , Immunity, Mucosal , Immunophenotyping , Male , Palatine Tonsil/surgery , Principal Component Analysis , Tonsillectomy , Tonsillitis/surgery , Young AdultSubject(s)
B7-H1 Antigen/metabolism , Bone Marrow/metabolism , Multiple Myeloma/pathology , Neoplasm Recurrence, Local/pathology , Neoplasm, Residual/pathology , Programmed Cell Death 1 Receptor/antagonists & inhibitors , Tumor Microenvironment , Animals , Case-Control Studies , Cells, Cultured , Female , Follow-Up Studies , Humans , Immunophenotyping , Killer Cells, Natural/metabolism , Killer Cells, Natural/pathology , Mice , Mice, Inbred C57BL , Multiple Myeloma/metabolism , Multiple Myeloma/mortality , Neoplasm Recurrence, Local/metabolism , Neoplasm Recurrence, Local/mortality , Neoplasm Staging , Neoplasm, Residual/metabolism , Neoplasm, Residual/mortality , Prognosis , Programmed Cell Death 1 Receptor/metabolism , Prospective Studies , Stem Cells/metabolism , Stem Cells/pathology , Survival Rate , T-Lymphocytes/metabolism , T-Lymphocytes/pathologyABSTRACT
Knowledge about clonal diversity and selection is critical to understand multiple myeloma (MM) pathogenesis, chemoresistance and progression. If targeted therapy becomes reality, identification and monitoring of intraclonal plasma cell (PC) heterogeneity would become increasingly demanded. Here we investigated the kinetics of intraclonal heterogeneity among 116 MM patients using 23-marker multidimensional flow cytometry (MFC) and principal component analysis, at diagnosis and during minimal residual disease (MRD) monitoring. Distinct phenotypic subclones were observed in 35/116 (30%) newly diagnosed MM patients. In 10/35 patients, persistent MRD was detected after 9 induction cycles, and longitudinal comparison of patient-paired diagnostic vs MRD samples unraveled phenotypic clonal tiding after therapy in half (5/10) of the patients. After demonstrating selection of distinct phenotypic subsets by therapeutic pressure, we investigated whether distinct fluorescence-activated cell-sorted PC subclones had different clonogenic and cytogenetic profiles. In half (5/10) of the patients analyzed, distinct phenotypic subclones showed different clonogenic potential when co-cultured with stromal cells, and in 6/11 cases distinct phenotypic subclones displayed unique cytogenetic profiles by interphase fluorescence in situ hybridization, including selective del(17p13). Collectively, we unravel potential therapeutic selection of preexisting diagnostic phenotypic subclones during MRD monitoring; because phenotypically distinct PCs may show different clonogenic and cytogenetic profiles, identification and follow-up of unique phenotypic-genetic myeloma PC subclones may become relevant for tailored therapy.
Subject(s)
Multiple Myeloma/genetics , Cell Separation , Coculture Techniques , Disease Progression , Drug Resistance, Neoplasm , Flow Cytometry , Humans , Immunophenotyping , In Situ Hybridization, Fluorescence , Multiple Myeloma/classification , Phenotype , Plasma Cells/cytology , Principal Component Analysis , Prognosis , Stromal Cells/cytologyABSTRACT
We have analyzed the applicability, sensitivity and prognostic value of allele-specific oligonucleotide real-time quantitative PCR (ASO RQ-PCR) as a method for minimal residual disease (MRD) assessment in patients with multiple myeloma (MM), comparing the results with those of multiparameter flow cytometry (MFC). A total of 170 patients enrolled in three consecutive Spanish trials achieving at least partial response after treatment were included. Lack of clonality detection (n=31), unsuccessful sequencing (n=17) and suboptimal ASO performance (n=51) limited the applicability of PCR to 42% of cases. MRD was finally investigated in 103 patients (including 32 previously studied) with persistent disease identified by PCR and MFC in 54% and 46% of cases, respectively. A significant correlation in MRD quantitation by both the techniques was noted (r=0.881, P<0.001), being reflective of treatment intensity. Patients with <10(-4) residual tumor cells showed longer progression-free survival (PFS) compared with the rest (not reached (NR) vs 31 months, P=0.002), with similar results observed with MFC. Among complete responders (n=62), PCR discriminated two risk groups with different PFS (49 vs 26 months, P=0.001) and overall survival (NR vs 60 months, P=0.008). Thus, although less applicable than MFC, ASO RQ-PCR is a powerful technique to assess treatment efficacy and risk stratification in MM.