Kurzfassung: Introduction. Ibrutinib is a first-in-class, oral, once-a-day Bruton's tyrosine kinase inhibitor that achieves high overall response rates and durable remissions in patients with chronic lymphocytic leukemia (CLL) including those with high-risk features (unmutated IGHV, TP53 abnormalities, 11q deletion). Survival with continuous single-agent ibrutinib in previously-untreated CLL patients is comparable to an age-matched general population (Figure 1). IBRORS is an observational, retrospective, multicentre study to describe the characteristics and clinical outcomes of patients with CLL treated with single-agent ibrutinib in routine clinical practice in Spain. This present analysis reviews the subset of patients in IBRORS who received ibrutinib as the first-line of treatment. This series includes a significant number of patients with high risk cytogenetic/molecular alterations (del17p/TP53 M), which corresponds with the approved indication for first-line CLL patients in Spain at the time. Methods. Adult patients diagnosed with CLL treated with single-agent ibrutinib in first-line, or at first or second relapse since its commercialization in Spain (between January 2016 to January 2019) were included in the IBRORS study. Clinical characteristics of patients, efficacy and tolerability of ibrutinib as first-line treatment were analyzed here. A Kaplan-Meier analysis was performed for overall survival (OS) and progression-free survival (PFS). Results. 84 patients, from a total of 269 included in IBRORS, received single-agent ibrutinib as first-line treatment. The median age was 71.3 years (range 63-77) at the time of ibrutinib initiation. 56.3% of patients presented with an intermediate/high-risk Rai-Binet stage, and the majority of patients (98.6%) had an ECOG PS of 0-1. 91.7% of patients had at least 1 high risk molecular cytogenetic factor (unmutated IGHV, TP53 abnormalities, 11q deletion or complex karyotype) described in table 1. Baseline comorbidities of patients are described in table 2. Concomitant medication included anticoagulants (9.5% patients; vitamin K antagonist [n=4], Apixaban [n=1] and LMWH [n=3] patients), antiplatelet agents (11.9% patients), and antihypertensives (50% patients). The overall response rate (ORR) was 79.5%; 14/84 (16.6%) achieved a complete response (CR), 14/84 (16.6%) achieved CR unconfirmed, 27/84 (32.14%) achieved a partial response (PR) and 12/84 (14.2%) a PR + lymphocytosis. The median PFS and OS were not reached, and the estimated PFS at 24 months was 84.5% (73.4-95.6%). OS and PFS curves are represented in figure 2. The PFS of each patient subgroup with high-risk cytogenetic characteristics was similar to that of all patients in the first-line cohort: del17p/TP53 mutation (HR = 0.963 [95% CI 0.188-4.928]; p = 0.964), del11q (HR = 0.042 [95% CI 0.000-682.736] ; p=0.521), unmutated IGHV (HR = 0.391 [95% CI 0.110-1.394]; p = 0.148). The median duration of exposure to ibrutinib was 17.3 (11.9-25.6) months. Dose reduction of ibrutinib occurred in 17/84 (20.2%) patients, 8/84 (9.52%) due to toxicity (4 hematologic toxicity and 4 non-hematologic toxicity). 27/84 (32.1%) patients had temporary interruption of treatment. 15/84 (17.8%) patients permanently discontinued ibrutinib including 6 (7.14%) patients due to progression, 4 (4.76%) due to toxicity and 5 for other reasons. Safety: 49/84 (58.3%) patients developed at least one adverse event (AE), while 12/84 (14.2%) patients developed at least one serious adverse event (SAE). Twelve (14.3%) patients reported at least one haematological toxicity while 53 patients (63.1%) recorded at least one non-haematological toxicity. Only 1 patient experienced grade 3 atrial fibrillation, which did not lead to discontinuation. The most common AEs are described in table 3. Conclusion. This population of previously-untreated CLL patients, enriched for high-risk genomic features, reflects the initial approval of ibrutinib for the treatment of first-line patients with del17p in Spain. Single-agent Ibrutinib as the first-line treatment in this real world population was effective regardless of risk factors and well tolerated, with a low rate of discontinuation due to toxicity. Findings are consistent with those reported in clinical trials. Disclosures Loscertales: AbbVie: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; AstraZeneca: Membership on an entity's Board of Directors or advisory committees; Roche: Honoraria. Arguiñano:AbbVie: Honoraria; Janssen: Honoraria; BMS-Celgene: Honoraria; Novartis: Honoraria. Hernandez-Rivas:Janssen: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Roche: Membership on an entity's Board of Directors or advisory committees; AstraZeneca: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Celgene/BMS: Membership on an entity's Board of Directors or advisory committees; Rovi: Membership on an entity's Board of Directors or advisory committees. Pérez Persona:Amgen: Consultancy; Celgene: Consultancy, Speakers Bureau; Roche: Consultancy, Speakers Bureau; Jannsen: Consultancy, Speakers Bureau; Abbvie: Consultancy, Speakers Bureau; Takeda: Consultancy. Loriente:Janssen Cilag: Current Employment. Villanueva:Janssen Cilag: Current Employment.

Kurzfassung: In acute myeloid leukemia (AML), FLT3 mutations are associated with a poor prognosis, particularly the internal tandem duplication (ITD/FLT3). In Latin America there are few  epidemiological data about these mutations.This study assessed the prevalence of FLT3 mutations in patients with AML at four reference hematology centers from Latin America. We included 138 samples of patients attending the Hematology Service of three Mexican University Hospitals (Monterrey,México D.F. and Puebla) and one Colombian center (Medellín) with a diagnosis of AML from different morphologic subgroups according to the French-American-British classification from 2006 to 2011. AML was diagnosed by morphology according to the FAB classification, by immunohistochemical staining and/or by immunophenotype according to each particular case. For sample processing DNA was extracted from peripheral blood or bone marrow with the automatic Maxwell®16 System (Promega Corporation, Madison, WI) using the principle of cellular lyses and binding nucleic acids to magnetized silice particles or the QIAAmp DNA Blood Kit (QIAGEN, Mexico City). The quality and DNA concentration was evaluated with the NanoDrop ND-1000 spectrophotometer (NanoDrop Technologies, Inc., Wilmington, DE). Then  internal tandem duplication and kinase domain mutations detection was performed with the GeneAmp PCR System 9700 (Applied Biosystems, Foster City, CA), through amplification of exons 14,15 with specific primers of FLT3 gen region, using the Seeplex® FLT3 Genotyping kit (Seegene, Rockville, MD, USA) or according to Kottaridis et al (1). Later, an electrophoretic analysis of the amplified products was made in a 2% agarose gel stained with ethidium bromide or in polyacrylamide gel electrophoresis (PAGE) and were observed by transilumination. In the Puebla samples the products were analyzed by capillary electrophoresis (ABI3130, Applied Biosystems, Foster City, CA). For detection of D835X mutations the exon 20 amplicom was subjected to digestion with EcoRV and analyzed by 4.5% PAGE. The patients were cytogenetically classified into three risk groups: favorable, intermediate, and adverse. Results We analyzed 138 samples of AML patients and found FLT3 mutations in 28 patients, for a prevalence of 20.3%. The median age was 47 years (5-96). Only four patients had the KD FLT3 mutation (3% of total population). The FLT3 mutation positive group was older than the negative (47 vs. 39 years), had higher WBC/mm3 (66.0 vs. 56.4 x 109/dl), higher hemoglobin values (9.3 vs. 8.6 g/dl), and lower platelet counts (72.6 vs. 92.5 x109/dl), although there were not statistically significant differences. Thirteen patients had AML M2, nine the monocytic variety, four had M3 and two M1. On cytogenetics  25% , 62.5% and 12.5% had favorable, intermediate and unfavorable risk karyotypes respectively. The rate response to standard Induction Chemotherapy was 78.3 % for the FLT3 positive group vs. 74.1 % for the non-mutated group. Nineteen of 28 patients (67.8%) with FLT3 mutations died, compared to 47 of 100 (42.72%) in those without the mutation. The median survival was 4.9 months for the FLT3 mutated group vs 20.4 months for the FLT3 negative group, P= 0.009.  The cytogenetic intermediate risk group (62.5%) was further analyzed and no statistically significant difference in overall survival between FLT3 non-mutated and FLT3 mutated patients was found (P= 0.22). Younger patients ( 〈 55 years)  had a higher mortality in the FLT3 positive group (P = 0.023).The presence or absence of the FLT3 mutation in patients who had the morphologic subtype M3 did not impact mortality (P = 0.28), but in non M3 subtypes, it did (P= 0.017). As conclusion, in this Latin American population the FLT3 mutation conferred a bad prognosis. References 1. Kottaridis PD, Gale RE,Frew ME et al. The presence of  a FLT3 internal tandem duplication in patients with acute myeloid leukemia (AML) adds importatnt prognostic information to cytogenetic risk group and response to the first cycle of chemotherapy: analysis of 854 patients from the United Kingdom Medical Research Council AML 10 and 12 trials. Blood.2001; 98: 1752-1759 2. Emerenciano M, Meneses J, Vasquez ML et al, Brazilian Collaborative Study Groupof Infant Acute L. Clinical relevance of FLT3 gene abnormalities in Brazilian patients with infant leukemia.Leuk Lymphoma. 2008; 49 (12):2291-2297. Disclosures: No relevant conflicts of interest to declare.

Kurzfassung: The tumoral clone of Waldenström’s macroglobulinemia (WM) shows a wide morphological heterogeneity which ranges from B lymphocytes (BL) to plasma cells (PC), including a lymphoplasmacytic population that defines the disease. The differences between these cell compartments and their cell-counterpart in other lymphoproliferative disorders have not yet been sufficiently explored. We compared the gene expression profiling (GEP) of BL and PC from patients diagnosed with WM, with clonal BL and PC from patients with chronic lymphocytic leukemia (CLL) and multiple myeloma (MM) respectively. Bone marrow samples from 10 WM and 10 MM patients and peripheral blood from 10 CLL patients were used for the analysis. The isolation of the different cell populations was carried out by multiparameter flow cytometry sorting with the following monoclonal antibodies combination: Kappa or Lambda-FITC, CD10-PE, CD38-PerCP-Cy5.5, CD19-PE-Cy7, CD34-APC and CD45-APC-Cy7. Total RNA (100–500 ng) was amplified and labeled using the “GeneChip Two-Cycle cDNA Synthesis Kit” and hybridized to “Human Genome U133A” microarray (Affymetrix). Processing of genechip data was carried out using the Robust Multi-chip Average (RMA) and the Affymetrix Microarray Suite v.5 (MAS5) gene expression algorithms. Two-way hierarchical cluster analysis showed that GEP was able to classify PC from WM and PC from MM in two different groups. In a similar way, BL from WM and CLL were grouped in different clusters. The Significance Analysis of Microarrays (SAM) algorithm identified gene expression changes in a total of 163 genes (103 up and 60 down-regulated) when MW PC were compared with MM PC. Some of these genes were related to regulation of PC development: PAX5 was overexpressed and IRF4 infraexpressed in WM PC. PC from WM displayed high expression levels of MYB and DEK oncogenes while MM PC showed an elevated expression level of v-MAF oncogene. Regarding gene functional categories, “immune response” and “signal transduction” were the biological process more active in WM PC. Comparison between BL from WM and BL from CLL revealed that 31 genes were differentially expressed: IL4 receptor, LEF1 (WNT/b-catenin pathway), fibromodulin (modulator of TGF-b activity) and FGR oncogene (tyrosine kinase protein) showed very low expression levels in WM BL compared to CLL BL. In contrast, the growth factor IL6 was over-expressed in WM BL. These results indicate that both PC and BL from WM are genetically different from the MM and CLL cell-counterpart. The differentially expressed genes have important functions in the B-cell differentiation and oncogenesis. Supported by Spanish Myeloma Network (G03/136) and “Ministerio de Ciencia y Tecnología” (SAF04/06587) and “Junta de Castilla y León” grants (SA032/04)

Kurzfassung: Background: Kinase domain (KD) mutations is a common resistance mechanism, secondary to the tyrosine-kinase inhibitors (ITKs) treatment in the case of chronic myeloid leukemia (CML) and Philadelphia (Ph)-positive acute lymphoblastic leukemia (ALL) patients. Sanger sequencing is the gold standard technique and already the currently recommended method for BCR-ABL1 KD mutation detection. However, Sanger sequencing has limited sensitivity and cannot firmly identify populations with variant allele frequencies (VAF) 〈 15-20%. Next-generation sequencing (NGS) allow us the screening of mutations in the whole KD with variants with a VAF greater than 1%. The aim of this study is to evaluate the clinical and prognostic implications of CML and Ph-positive ALL patients who have been studied for mutations in BCR-ABL1 by NGS. Methods: Seventy CML and Ph-pos ALL patients have been studied for BCR-ABL1 mutations between years 2015-2017. The study reason was warning or failure according to European Leukemia Net recommendations in the case of CML patients, and diagnostic or relapse in the case of ALL patients. Clinical characteristics of the patients are depicted on Table 1. Categorical variables are described as frequency, and quantitative variables as medians. Contingency tables were used to analyze associations between categorical variables (χ2). Median test was used to compare medians of continuous variables between groups. Overall survival (OS) was estimated using the Kaplan-Meier method and compared between patients using the log-rank test. Results: We have found 37 patients with mutations (51%), the most frequent being p.T315I, p.L248V and p.L387M. 28 out of 59 were found in CML (47%) vs 9 out of 13 (69%) in ALL. Of the 37 patients with mutations, double mutations have been found 10 times (27%). In the 72 analyses performed, 62 mutations were found in total, 41 of them were variants of uncertain significance (VUS) and 21 were well-known mutation. The median levels of BCR-ABL1 (IS) at the time of analysis were 3.00 (0.01-196.18) %. Regarding CML patients, we have found 12 and 16 cases with pathogenic mutations and VUS, respectively. The mean survival for CML and ALL were 75.2 months (CI 95%, 65.7-84.6) and 24.7 months (13.3-36.2), respectively. There are significant differences between the overall survival curves for patients with CML who have mutations in BCR-ABL1 compared to those who have VUS or do not (p-value = 0.024, n=59), suggesting a second role of the VUS variants in the resistance of the patients to the TKI. These two groups have no significant differences in ALL patients (p-value= 0.32, n=13). Overall survival at 10 years from the date of diagnosis is 74% for CML patients with mutations and 90% for CML patients without mutations. Data dropped significantly for ALL patients, but the number of cases is too low. Conclusions: - Mutations have been identified in 47% of CML patients studied in the case of failure or warning and 69% of the patients of ALL at diagnosis or relapse moments. - The identification of pathogenic variants has poor prognosis in patients with CML (p = 0.024), however no differences were observed in ALL. - The identification of VUS is not associated to poor prognosis and these variants could not confer resistance to ITK. Disclosures Sevilla: Rocket Pharmaceuticals, Inc.: Honoraria, Patents & Royalties: Inventor on patents on lentiviral vectors filled by CIEMAT, CIBERER and F.J.D and may be entitled to receive financial benefits from the licensing of such patents; NOVARTIS: Honoraria, Membership on an entity's Board of Directors or advisory committees; Rocket: Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Sobi: Membership on an entity's Board of Directors or advisory committees; Miltenyi Biotech: Honoraria. Steegmann:Incyte: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. García Gutiérrez:Novartis: Honoraria, Research Funding; BMS: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Incyte: Honoraria, Research Funding.

Kurzfassung: Background and aim: Current therapeutic protocols for adult ALL consider MRD together with the baseline risk factors (age, WBC count, immunophenotype, cytogenetics) and speed in response to therapy for treatment decisions. On the other hand, the systematic use of allogeneic SCT for all adult patients (pts) with Ph- HR-ALL is still a matter of debate. The aim of the prospective study ALL-AR-03 from the Spanish PETHEMA Group was to evaluate the response to a differentiated therapy (chemotherapy or allogeneic SCT) according to early bone marrow blast clearance and MRD levels (assessed by cytofluorometry at the end of induction and consolidation therapy) in HR Ph- adult ALL patients. Patients and methods: HR ALL included one or more of the following baseline parameters: age 30–60 yr, WBC count 〉 25x109/L and 11q23 or MLL rearrangements. Induction therapy included vincristine, prednisone and daunorubicin for 4 weeks. In pts with slow cytologic response to therapy (≥10% blasts in bone marrow assessed on d14) intensified induction with high dose ARA-C and mitoxantrone was administered. Early consolidation therapy included 3 cycles with rotating cytotoxic drugs including high-dose methotrexate, high-dose ARA-C and high-dose asparaginase. Pts. with slow cytologic response on d14 or MRD level 〉 0.05% after consolidation were assigned to allogeneic SCT (related or unrelated) and those with standard cytologic response on d14 and MRD level 〈 0.05% after consolidation received 3 additional cycles of delayed consolidation (identical to those of early consolidation) followed by maintenance therapy up to 2yr in CR. Results: On June 2008,192 patients were evaluable (mean (SD) age 37(10) yr, 105 males, 119 precursor B-ALL, 73 T-ALL, WBC count 65(99) x109/L). Induction death: 17(9%), resistance: 12 (6%), CR: 163 (85%). MRD 〈 0.1% was observed in 64% of CR patients. Early consolidation was completed in 126 patients. MRD 〈 0.05% was observed in 65% at the end of consolidation. On June 2008, allogeneic SCT was performed to 30 pts (15 from HLA-identical siblings and 15 MUD), TRM 11 pts, relapse 4, CCR 15. Delayed consolidation and maintenance was administered to 79 pts (toxic death 4 pts, relapse 21, CCR 54). Four-yr DFS for the whole series was 36±7% (37±19% for pts assigned to SCT and 56±12% for those assigned to chemotherapy). Slow cytologic response was associated with a lower CR probability and higher induction death. No baseline variable was associated with a higher probability of MRD negativity after induction or consolidation. Conclusions: These results suggest that in HR Ph- adult ALL pts with adequate response to induction and adequate clearance of MDR the results of therapy are not hampered by avoiding allogeneic SCT. Supported by grants P-EF/07 from FIJC and RD 06/0020/1014 from Instituto Carlos III

Kurzfassung: p15 and p14/p16 tumor suppressor genes, have been reported to be frequently inactivated by various mechanisms in haematological malignancies such us MM. Alterations of these cell cycle inhibitors in MM display a close correlation with the cell cycle and clinical outcome. We have evaluated by real time quantitative RT-PCR (RQ-PCR) the expression of the p14/p16 and p15 genes in purified bone marrow plasma cells (PBMPC) from MM patients in order to evaluate the possible clinical, biological and prognostic significance of these cell cycle regulators. RNA extracted from purified BMPC from 53 untreated symptomatic MM and a pool of buffy coat from healthy donors (reference value) was analyzed by RQ-PCR using Assays-on-Demand gene expression mixes specific for p14/p16 and p15 genes in an ABI PRISM 7700 SDS (Applied Biosystems, Foster City, CA, USA). Values were corrected with a control gene (ABL). The relative quantification of gene expression was performed through the cycle threshold (CT) increment method. Patients were classified into different groups depending on gene expression values. Thus, according to p15 expression, 29% of patients (n=14) showed higher levels than the control and this group was characterized by the presence of good prognostic markers such us low Lactato dehidrogenase levels (LDH), low b2-microglobulin (B2M) and C-Reactive Protein (CRP) serum levels and absence of monoclonal proteinuria. Similar results were found for p14/p16 expression. Fifteen patients (28%) displayed a high p14/p16 expression and the group was also characterized by good prognostic features: low CRP, B2M and LDH levels. When p14/p16 and p15 genes were simultaneously analyzed, clinical and biological parameters showed a statistically significant correlation with gene expression. Thus patients with low gene expression had a high B2M (≥3 mg/dl) and high C-reactive protein (≥3 mg/dl). As far as survival was concerned, patients with a high p15 expression had a longer overall survival of 100% vs. 58% at 30 months (p=0,01), with the additional value that no deaths have been observed in any such patients. Similar results were observed for the group of patients displaying a high p14/p16 expression since they displayed a much better OS (100% vs. 63% at 30 months, p=0,02). Again, we should note that no deaths have been presented in this group. All these findings were much more evident when the three genes were simultaneously considered. Thus, within the group of 21 patients with at least one of the two genes overexpressed there have been no deaths vs. 11 among the 27 patients with low levels. This resulted in quite different OS curves for the two groups of patients (Figure 1) of 100% vs. 49% at 30 months (p=0,00). In conclusion, these genes significantly determine patients’ outcome thanks to their ability to classify them into different groups with different clinical, biological and outcome characteristics.

Kurzfassung: Gene Expression Profiling through RNA arrays has provided new clues to Multiple Myeloma pathogenesis and prognostic pattern evaluation. Recently, ZHX2, CHC1L & RAN expression have been highlighted as key elements in MM. In the present paper, we have evaluated theses genes by real time quantitative RT-PCR (RQ-PCR) in purified plasma cells from 74 patients with plasma cell discrasias. MATERIAL AND METHODS: Purified Bone Marrow cells were obtained from the following patients: 6 MGUS, 7 smoldering MM, 59 newly diagnosed symptomatic MM patients and 2 Plasma cell leukemia (PCL). After RNA extraction, RQ-PCR of CHC1L(C), RAN(R) and ZHX2 (Z) genes was carried out using the standard protocol from TaqMan® gene expression Assays-on-Demand in an ABI-PRISM 7700 Sequence Detection System (Applied Biosystems, Foster City, CA, USA). Expression levels were normalized with ABL gene and expressed in n-fold times compared to the expression in a pool of RNA from mononuclear cells from healthy donors. The expression level of the different genes was evaluated for correlation with the diagnosis, clinical characteristics and prognosis of the patients. RESULT AND CONCLUSIONS: None of these genes displayed a clear relationship with the different stages of disease pathogenesis, although ZHX2 gene was slightly more expressed in the indolent forms of the proliferative disorders (MGUS and SMM). Within symptomatic MM patients, several interesting associations were observed. Thus, in hyperdiploid MM cases, CHC1L expressions observed were fewer than in those with a normal DNA index, confirming the participation of the gene product in chromosomal condensation during the mitosis. No other important associations were observed for this gene, although patients with the lowest expression displayed a very good prognosis, but without reaching statistically significant differences. As expected, RAN expression was related to S-Phase PC, since patients with high S phase values ( & gt;1.8 %) displayed higher levels of RAN transcripts. This, however, only resulted in a marginal impact on survival. ZHX2 provided the most interesting results, whereby decreased levels of ZHX2 were related to unfavorable prognostic indicators such as B2 microglobulin & gt;4 mg/L and Hemoglobin levels & lt;10.5 g/dl. This was significantly associated with a shorter survival. As such, patients with a level of ZHX2 expression lower than twice the expression of controls were associated with a very low survival (median of 6 months vs not reached, p=0.019). If we take into account the survival prediction value of these three genes, the following prognosis groups were defined: very good prognosis (Z+ C−), poor prognosis (Z− C+) and intermediate (the remaining patients). SUMMARY: From this study we can confirm that RAN, CHC1L and especially ZHX2 genes play a role in the pathogenesis and behavior of MM, this could be helpful in predicting survival of patients.

Kurzfassung: Introduction Multiple Myeloma (MM) is a heterogeneous disease with a complex clonal and subclonal architecture with few recurrent mutations. The arrival of next-generation sequencing (NGS) has allowed us to have a deeper understanding of the disease. Due to that complexity and the low recurrence of "driver" mutations, the study of general mutational profile, copy number variation (CNV) and translocations is crucial to make an accurate diagnosis and prognosis. For that reason, a clinically validated NGS capture panel has been designed to analyze in a single assay all interesting genetic aberrations simultaneously including SNVs, indels, CNVs and chromosomal translocations. Methods In addition to genomic DNA (gDNA) from 33 healthy donors to create a robust baseline forCNV detection,we studied 161 DNA samples from 149 newly diagnosed MM patients enrolled in the GEM2012MENOS65clinical trial and treated homogeneously: gDNA from 149 BM CD138+ plasma cells and 12 paired cfDNA from peripheral blood samples obtained at diagnosis. First, starting with 100 ng of gDNA and 200ng for cfDNA samples, a custom targeted NGS panel using SureSelect capture technology (Agilent) followed by NextSeq500 (Illumina) sequencing identified SNVs, indels, CNVs and the most relevant IGH translocations within 26genes involved with MM. The average sequencing depth was 609x across samples; and 98% of the targeted regions were sequenced with & gt;200x. Second, NGS custom panel results were compared with FISH (n=88) and whole exome sequencing (SureSelect XT V6) (n=48) results. Both NGS panel and exome raw data were analyzed by DREAMgenics applying a custom bioinformatic pipeline. Finally, 5 discordant cases were followed-up by SNP-arrays. Results We have identified 408 exonic and non-synonymous variants. At least 1 oncogenic mutation was detected in 86% (128/149) of patients. NRAS was mutated in25% of patients, followed by KRAS(23%), BRAF(12%) DIS3(11%) and TP53(9%). Other interesting pathogenic mutations were identified in FGFR3 andHIST1H1E genes in 9% and 5% of patients, respectively. In 92% (11/12) of cfDNA samples at least 1 oncogenic mutation was detected. For this 12 cases, paired samples (BM CD138+vscfDNA) were available. A total of 39 somatic mutations were identified in those cases. In cfDNA, 10 mutations were detected, and 5 were present in both samples. Furthermore, a mean decrease of 0.19VAF was observed in cfDNA (0.12; 0.01-0.48) vs plasma cells (0.31; 0.01-0.51). Regarding to CNV, 1q gain was detected in 32% of patients (28/88), and 1p and 17p deletions in 17% (15/88) and 13% (11/88), respectively. Additionally, ATR and CRBN gene amplifications were detected in 22% and 16%, respectively. When these data were compared to FISH, a 75% of sensitivity and 91% of specificity was achieved by our method, with a PPV of 68% and a NPG of 93%. Translocations were identified in 28% (25/88) of patients, including 7% (6/88)) t(11;14), 14% (12/88), t(4;14), and 1 patient t(14;16). We also detected t(6;14)(p21;q32) IGH/CCND3 in 3 patients that had also been described in MM.Translocations were detected with a 94% of sensitivity, 99% of specificity, with a predictive positive value of 94% and a predictive negative value of 99%. Importantly, NGS-based method revealed a t(10;14) in 3 patients that had not been identified by FISH, a new translocation implying the miRNA hsa-mir-4537. Finally, the impact on PFS from FISH and NGS results was analyzed separately. PFS was similar for translocations and 17p deletions. However, 1p-detected by NGS showed a higher negative prognostic impact (p=0,006 vs p=0,127 by FISH)(Fig.1). Furthermore, our panel showed that 7% of patients (6/88) had a bi-allelic TP53 inactivation. Survival analysis showed that these patients relapsed significantly earlier than the others (p=0.028). Amplifications (≥4 copies) in 1q+could not identified with this panel. Conclusions Our custom NGS-based test allows in a single assay a more comprehensive study of the genomic landscape of MM patients by (a) detecting with a high sensitivity the most important and recurrent mutations and cytogenetic alterations, (b) identifying translocations and CNVs not previously detected by FISH and (c) identifying a double-hit MM patient. Additionally, cfDNA could be analyzed with this NGS strategy identifying molecular alterations in most of the patients. Disclosures Oriol: Celgene: Consultancy, Speakers Bureau; Amgen: Consultancy, Speakers Bureau; Janssen: Consultancy. Sureda Balari:Takeda: Consultancy, Honoraria, Speakers Bureau; Celgene/Bristol-Myers Squibb: Consultancy, Honoraria; Roche: Honoraria; Sanofi: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Gilead/Kite: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Incyte: Consultancy; Celgene: Consultancy, Honoraria; BMS: Speakers Bureau; Merck Sharpe and Dohme: Consultancy, Honoraria, Speakers Bureau. de la Rubia:Janssen: Consultancy, Other: Expert Testimony; Celgene: Consultancy, Other: Expert Testimony; Amgen: Consultancy, Other: Expert Testimony; Ablynx/Sanofi: Consultancy, Other: Expert Testimony. Mateos:Oncopeptides: Honoraria, Membership on an entity's Board of Directors or advisory committees; Sanofi: Honoraria, Membership on an entity's Board of Directors or advisory committees; PharmaMar-Zeltia: Consultancy; Abbvie/Genentech: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Adaptive Biotechnologies: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen-Cilag: Consultancy, Honoraria; Regeneron: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Seattle Genetics: Honoraria, Membership on an entity's Board of Directors or advisory committees; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees; Pfizer: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; GlaxoSmithKline: Consultancy; Takeda: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Blade Creixenti:Amgen: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees; Oncopeptides: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees. San-Miguel:Amgen, BMS, Celgene, Janssen, MSD, Novartis, Takeda, Sanofi, Roche, Abbvie, GlaxoSmithKline and Karyopharm: Consultancy, Membership on an entity's Board of Directors or advisory committees. Garcia-Sanz:Novartis: Honoraria; Janssen: Honoraria, Research Funding; Incyte: Research Funding; Gilead: Honoraria, Research Funding; BMS: Honoraria; Amgen: Membership on an entity's Board of Directors or advisory committees; Pharmacyclics: Honoraria; Takeda: Consultancy, Research Funding. Martinez-Lopez:Novartis: Research Funding; BMS: Research Funding, Speakers Bureau; Incyte: Research Funding, Speakers Bureau; Janssen: Speakers Bureau; Roche: Speakers Bureau; Amgen: Speakers Bureau; Takeda: Speakers Bureau; Vivia Biotech: Honoraria; Altum: Membership on an entity's Board of Directors or advisory committees, Patents & Royalties; Hosea: Membership on an entity's Board of Directors or advisory committees, Patents & Royalties.